Langmuir Monolayer

CHEMICAL ENGINEERING METHODS AND TECHNOLOGY
LANGMUIR MONOLAYERS IN THIN FILM TECHNOLOGY
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LANGMUIR MONOLAYERS IN THIN FILM TECHNOLOGY
JENNIFER A. SHERWIN
EDITOR
Nova Science Publishers, Inc.

New York
Copyright 2011 by Nova Science Publishers, Inc.

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LIBRARY OF CONGRESS CATALOGING-IN-PUBLICATION DATA Langmuir monolayers in thin film technology / editors, Jennifer A. Sherwin. p. cm. Includes bibliographical references and index. ISBN 978-1-61209-188-4 (eBook) 1. Thin films, Multilayered. I. Sherwin, Jennifer A. QC176.9.M84L365 2010 621.3815'2--dc22 2010041312
Published by Nova Science Publishers, Inc. New York
CONTENTS
Preface Chapter 1 Carbons and Clays for Heavy Metals Removal A Review of Latest Literature John U. Kennedy Oubagaranadin and Z.V.P. Murthy Molecular Organization of Thermotropic Liquid Crystals and Their Mixtures with Azo Dyes in Langmuir and Langmuir-Blodgett Films Danuta Bauman, Anna Modliska, and Krzysztof Inglot Atomic Force Microscopy Characterization of Lipid/ Protein Nanostructures Formed in Langmuir-Blodgett Films Yih Horng Tan and Keith J. Stine Langmuir Monolayers in Biosensors Jadwiga Sooducho and Joanna Cabaj Adsorptive Characteristics of Bovine Serum Albumin onto Cationic Langmuir Monolayers of Sulfonated Poly (Glycidylmethacrylate)Grafted Cellulose: Mass Transfer Analysis, Isotherm Modeling and Thermodynamics T. S. Anirudhan and P. Senan Electrochemistry of Polymeric Thin Films Prepared by Langmuir-Blodgett Technique Paolo Bertoncello vii 1
Chapter 2
51
Chapter 3
101 131
Chapter 4 Chapter 5
151
Chapter 6
177 201
Index
PREFACE
The Langmuir-Blodgett (LB)technique for the preparation of ultrathin films of various organic, metallorganic, and polymeric compounds play an increasingly important role as a means or organizing molecular materials at the microscopic level. The LB technique has many potential applications in molecular electronics, nonlinear optics and conducting thin films. This book presents current research from across the globe in the study of Langmuir Monolayers, including the study of thermotropic liquid crystals and binary mixtures of dichroic azo dye/liquid crystal in Langmuir and Langmuir-Blodgett films; proteo-lipidic nanostructures generated via the Langmuir-Blodgett film method; Langmuir Monolayers in biosensors; as well as adsorptive characteristics of bovine serum albumin onto cationic Langmuir Monolayers of sulfonated poly-grafted cellulose. Chapter 1 - In this article, various adsorption isotherm models, kinetic models, adsorption thermodynamics and the technical viability of carbons made from different biomaterials and different types of naturally occurring clays for heavy metals removal by adsorption from contaminated water has been reviewed. Natural clays and carbons prepared from waste plant products can be employed for heavy metals removal from aqueous solutions and disposed of with little cost. Modification of these materials can also improve their adsorption capacity. In this review, an extensive literature survey from the past decade on the heavy metals removal characteristics of clays and carbons has been compiled to provide a summary. It is evident from this literature survey that carbons made from biomaterials and clays have demonstrated outstanding removal capabilities for certain metal ions. Some of the highest adsorption capacities reported are: for Cd(II), 180 mg/g by bean husk (Phaseolus vulgaris) carbon; for Cr(VI), 120.48 mg/g by date palm seed wastes carbon; for Pb(II), 279.92 mg/g by carbon made from Euphorbia rigida; for Hg(II), 151.5 mg/g by ZnCl2 activated walnut shells carbon; for Cr(III), 117.5 mg/g by Smectite clay with a small proportion of kaolinite; for Cd(II), 74.07 mg/g by petra clay; for Cu(II), 105.38 mg/g by modified Unye clay; for Pb(II), 104.28 mg/g by natural palygorskite clay; for Cu(II), 909 mg/g by Saudi bentonite. It is important to note that the adsorption capacities of the adsorbents presented in this paper vary, depending on the experimental conditions, characteristics of the individual adsorbent, the extent of chemical modifications, and the concentration of adsorbate. Chapter 2 - In this review articles the results of the study of thermotropic liquid crystals and of binary mixtures of dichroic azo dye/liquid crystal in Langmuir and Langmuir-Blodgett films are presented. The liquid crystals of rod-like shape from various homologous series and nine azo dyes with different molecular structure as well as different values and directions of
viii
Jennifer A. Sherwin
the dipole moment were chosen. It was found that the liquid crystals with the terminal isothiocyanato (NCS) group are not able to form a compressible monolayer at the water surface. Very short and very long alkyl or alkoxy chains attached to the rigid molecular core also hinder the creation of the stable film. Azo dyes cannot form the Langmuir film themselves; therefore, the liquid crystals 4-n-octyl-4'-cyanobiphenyl (8CB) and trans-4-noctyl(4-cyanophenyl)-hexane (8PCH) were used as supporting matrices. The Langmuir films were characterized by the surface pressure-area and surface potential-area isotherms and by Brewster angle microscopy (BAM). The analysis of the isotherms and BAM images of liquid crystals indicated that the organization of the mesogenic molecules at the air-water interface is dependent on their structure and to some extent reflects their ability to form an appropriate mesophase in the bulk. For the binary azo dye/8CB mixtures the miscibility of two components as well as the organization and the packing of molecules at the water surface were determined. The absorption spectra by using natural and linearly polarized light were recorded for both Langmuir and Langmuir-Blodgett films. Information about spectral properties of ultra-thin layers and ability of dye and liquid crystal molecules to form selfaggregates was obtained. The polarized absorption spectra allowed one to determine the alignment of molecules on the quartz surface. Chapter 3 - Recently, much of the emphasis in biotechnology has been on producing nanosystems and nanodevices for a vast range of medical applications, including nanoelectronic biosensors, drug delivery systems, and diagnostic and imaging techniques, to name a few. This chapter reviews the potentially useful creation of proteo-lipidic nanostructures generated via the Langmuir-Blodgett (LB) film method, and their direct characterization by atomic force microscopy (AFM). Following the introductory sections, a brief overview on LB film fabrication on surfaces suitable for AFM will be presented, LB films containing proteins, lipids, and biocompatible amphiphilic molecules will be described, and how they have been studied using various AFM imaging modes. We aim to highlight recent developments that illustrate the unique capability of AFM in elucidating nanometer scale organization and the physicochemical properties of artificially engineered biological membranes through the Langmuir-Blodgett method; as it could potentially open a new pathway toward the development of self-organized nanostructures of technological significance. Chapter 4 - In recent years, the Langmuir-Blodgett (LB) technique for the preparation of ultrathin films of various organic, metallorganic, and polymeric compounds plays an increasingly important role as a means of organizing molecular materials at the microscopic level. The LB technique has many potential applications in molecular electronics, nonlinear optics and conducting thin films. The most important advantage of this method is that the characteristic of the film can be varied by changing various LB parameters, namely, surface pressure of lifting, temperature, barrier speed, dipping speed, molar composition, etc. So it is important to study different molecules having various chromophores with interesting photophysical and electrical properties, confined in the restricted geometry of the LB films to fabricate various molecular electronic devices and also to realize the basic physicochemical processes involved at the mono and multilayer films [1]. Chapter 5 - Investigation on adsorption behaviour of Bovine Serum Albumin (BSA) on polymeric adsorbent materials is critical for many analytical and biomedical applications. In the present study a novel adsorbent poly(glycidylmethacrylate)-grafted-cellulose having sulfonate functional groups (PGMA-g-Cell-SO3H) was prepared by graft copolymerization of
Preface
ix
glycidylmethacrylate (GMA) onto cellulose in the presence of ethyleneglycoldimethacrylate as crosslinker using ,-azobisisobutryronitrile as initiator followed by the introduction of sulfonic acid groups through ring opening reaction of the epoxide groups of the grafted GMA with sodium sulfiteisopropanolwater mixture. The original and the modified materials were characterized by means of FTIR, SEM, XRD and BET analysis. Adsorption characteristics of BSA onto PGMA-g-Cell-SO3H were investigated under different optimized conditions of pH, contact time, initial BSA concentration, adsorbent dose and temperature. The maximum value of BSA adsorption was found to be 49.95 and 72.07 mg/g for an initial concentration of 100 and 150 mg/L, respectively at pH 4.5. Kinetic studies showed that the equilibrium conditions were achieved within 3 h. The kinetic data obtained at different concentrations and temperatures were analyzed using a pseudo-first-order and pseudo-second-order equation. The adsorption process followed pseudo-second-order kinetics. The experimental kinetic data were correlated by the external mass transfer and intraparticle mass transfer diffusion models. The intraparticle mass transfer diffusion model gave a better fit to the experimental data. Experimentally obtained isotherms were evaluated with reference to Langmuir, Freundlich and Sips equations. The isotherm data were best modelled by the Langmuir isotherm equation and the maximum monolayer adsorption capacity was found to be 124.85 mg/g at 30 C. Thermodynamic study revealed an endothermic adsorption process. The negative G values indicate feasible and spontaneous adsorption of BSA onto PGMA-g-Cell-SO3H. The positive and small value of enthalpy change Ho (9.50 kJ/mol) indicates the endothermic nature of adsorption primarily through weak physical forces between adsorbent and adsorbate. The positive and small value of entropy change, So (185.52 J/mol/K) indicates that the order less nature of adsorption system increases with adsorption of BSA onto adsorbent surface. Also at all temperatures Ho<T So, indicating that the BSA adsorption onto PGMA-g-Cell-SO3H is dominated by entropic rather than enthalpic changes. The values of isosteric heat of adsorption computed from linear isotherms remained almost constant (~ 26.45 kJ/mol) with different surface loadings suggest that the surface of PGMA-g-Cell-SO3H is energetically more or less homogeneous and the lateral interactions between adsorbed BSA ions do not exist. Adsorbed BSA was eluted completely by 0.2 M CH3COOH solution. Results obtained from repeated adsorption/desorption process showed that PGMA-g-Cell-SO3H can be used for the adsorption of BSA from various aqueous solutions. Chapter 6 - The utilization of the Langmuir-Blodgett (LB) technology for the fabrication of engineered supramolecular thin films has received an exceptional development in these last years due to possibility of different applications in materials science ranging from nanotechnology to biosensors. The materials fabricated by LB technology provide an accurate control of the order at the molecular level. The main objective of this chapter is to give an overview of the electrochemical properties of a particular class of polymeric thin films such as conducting polymers and ionomer polymers and describe the potentialities of some recent electrochemical technique for nanotechnological applications mainly scanning electrochemical technique (SECM) and SECM combined to Langmuir trough.
In: Langmuir Monolayers Editors: Jennifer A. Sherwin
ISBN: 978-1-61122-461-0 2011 Nova Science Publishers, Inc.
Chapter 1
CARBONS AND CLAYS FOR HEAVY METALS REMOVAL A REVIEW OF LATEST LITERATURE
John U. Kennedy Oubagaranadin 1,2 and Z.V.P. Murthy*2
Department of Ceramic and Cement Technology, PDA College of Engineering, Gulbarga, Karnataka, India 2 Department of Chemical Engineering, S.V. National Institute of Technology, Surat, Gujarat, India
1
ABSTRACT
In this article, various adsorption isotherm models, kinetic models, adsorption thermodynamics and the technical viability of carbons made from different biomaterials and different types of naturally occurring clays for heavy metals removal by adsorption from contaminated water has been reviewed. Natural clays and carbons prepared from waste plant products can be employed for heavy metals removal from aqueous solutions and disposed of with little cost. Modification of these materials can also improve their adsorption capacity. In this review, an extensive literature survey from the past decade on the heavy metals removal characteristics of clays and carbons has been compiled to provide a summary. It is evident from this literature survey that carbons made from biomaterials and clays have demonstrated outstanding removal capabilities for certain metal ions. Some of the highest adsorption capacities reported are: for Cd(II), 180 mg/g by bean husk (Phaseolus vulgaris) carbon; for Cr(VI), 120.48 mg/g by date palm seed wastes carbon; for Pb(II), 279.92 mg/g by carbon made from Euphorbia rigida; for Hg(II), 151.5 mg/g by ZnCl2 activated walnut shells carbon; for Cr(III), 117.5 mg/g by Smectite clay with a small proportion of kaolinite; for Cd(II), 74.07 mg/g by petra clay; for Cu(II), 105.38 mg/g by modified Unye clay; for Pb(II), 104.28 mg/g by natural palygorskite clay; for Cu(II), 909 mg/g by Saudi bentonite. It is important to note that the adsorption capacities of the adsorbents presented in this paper vary, depending on the experimental conditions, characteristics of the individual adsorbent, the extent of chemical modifications, and the concentration of adsorbate.
John U. Kennedy Oubagaranadin and Z.V.P. Murthy
1. INTRODUCTION
Heavy metals in the environment are not biodegradable and tend to accumulate in living organisms, causing various diseases and disorders. Eleven metals, viz., Pb, Cr, Hg, U, Se, Zn, As, Cd, Co, Cu and Ni, out of 20 classified metals as toxic, are emitted into environment in quantities that pose risks to human health (Kortenkamp et al., 1996). It is known that adsorption is cost effective and efficient in heavy metals removal from solutions. Application of carbons made from plant species and naturally occurring inorganic materials (clays) in heavy metals removal from various industrial effluents or water resources have been reported. Adsorption with clays has become an alternative to traditional methods of industrial wastewater treatment and it is relatively inexpensive, non-hazardous, and may permit recovery of the metals from the adsorbing mass.
Adsorbent carbons and clays are different in nature. A pure carbon surface is considered to be non-polar, but in actuality some carbon-oxygen complexes CxO, COx and CxO2 are usually present. When a metal ion is present in solution, surface complexes may be formed by ion exchange between H+ functional groups and metal ions in solution. Such mechanism is consistent with those proposed in earlier investigations (Namasivayam and Periasamy, 1993; Namasivayam and Kadirvelu, 1999). As metal ions are adsorbed onto the carbon, more hydrogen ions are released from the carbon into solution. The hydrogen ion sources are most likely the carboxylic, phenolic, sulfonic and lactonic groups in the carbon. These groups are generally considered to be responsible for cation exchange capacity of carbons (Kadirvelu et al., 2000).
Clays are basically aluminosilicate minerals containing sodium, potassium and calcium, in which magnesium and iron may be substituted for aluminium. The ion-exchange capacity of clays depends on the frame work of Si/Al ratio and decreases as the Si/Al ratio increases. The clays carry a net negative charge due to the broken bonds around the edges of the silicaalumina units that would give rise to unsatisfied charges, which would be balanced by the adsorbed cations. Substitutions within the lattice structure of trivalent aluminum for quadrivalent silicon in the tetrahedral sheet and of ions of lower valence, particularly magnesium for trivalent aluminum in the octahedral sheet, result in unbalanced charges in the structural units of clay minerals (Pehlivan et al., 2006). The process of adsorption is well explained by the well-known two-parameter Freundlich and Langmuir isotherms. However there are a number of other two-parameter, threeparameter and four-parameter models that can explain the adsorption equilibrium data better. Similarly kinetics of adsorption can be represented by a number of rate models. This article provides a collection of these models for the testing of adsorption equilibrium and kinetics data, and an overview of carbons made from bio-materials and clays for heavy metal removal from aqueous solutions based on recent publications (past decade).
2. ADSORPTION ISOTHERMS, KINETICS AND THERMODYNAMICS

2.1. Adsorption Isotherms
Adsorption is a surface process that leads to transfer of molecules from a fluid bulk to solid surface. This can occur because of physical forces or by chemical bonds. In most of the
Carbons and Clays for Heavy Metals Removal
cases, this process is described at the equilibrium by means of some equations that quantify the amount of substance attached on the surface given the concentration in the fluid. These equations are called isotherms because of the dependence of their parameters on the temperature, which is one of the most important factors affecting the adsorption. The uptake capacity of an adsorbent at equilibrium is given by:
qe
where, C0 and Ce are the initial and equilibrium concentration of the adsorbate (mg/L), m is mass of the adsorbent (g) and V is the volume of the solution (L). A brief description of various adsorption isotherm models used by researchers is given below.
(C0 Ce ) m /V
(2.1)
2.1.1. Henry Isotherm Henrys law is basically a one parameter model and has been applied in many cases (Ho, 2004a). It has the form of equation of a straight line with no intercept. This model normally does not represent the adsorption equilibrium data due to the unavailability of adsorption data in the lower range of metal concentration. In liquid-phase adsorption, the equilibrium adsorption data are normally obtained at higher equilibrium concentrations, where the adsorbent surface is almost at the edge of saturation. High concentrations may suggest the applicability of a model having a linear relationship between qe and Ce at the latter part of the equilibrium isotherm curve. This requisite may be partially fulfilled using Henrys law equation with an intercept. The addition of a constant term in Henrys law provides the model to integrate the basic characteristics of the equilibrium isotherm curve at the high concentration range. Henrys law is given as:
qe K H Ce b
(2.2)
where, KH and b are model constants. This model can be applied for high effluent concentration.
2.1.2. Freundlich Isotherm The Freundlich isotherm (Freundlich, 1907) habitually gives a better fit particularly for adsorption from liquids and can be expressed as:
qe K F Ce
(2.3)
In this model, the mechanism and the rate of adsorption are functions of the constants n and KF (L/g). For a good adsorbent, 1<n<10, and a higher value of n indicates better adsorption and formation of rather strong bond between the adsorbate and adsorbent (Treybal, 1981). It is a simple expression and shows no leveling.
2.1.3. Langmuir Isotherm The Langmuir (1916, 1917, 1918) model assumes uniform energies of sorption onto the surface and no transmigration of adsorbate in the plane of the surface. The Langmuir equation may be written as:
qe
qm K L Ce 1 K L Ce
(2.4)
In this model, qm (mg/g) is the amount of adsorption corresponding to complete monolayer coverage, i.e., the maximum adsorption capacity and KL (L/mg) is the Langmuir constant. It is also a simple expression with interpretable parameters and applicable for monolayer adsorption. For Langmuir type adsorption process, to determine if the adsorption is favorable or not, a dimensionless separation factor, RL, is defined (Hall et al., 1966) as:
RL
1 1 K L C0
(2.5)
If R L >1, the isotherm is unfavorable
RL =1, the isotherm is linear

0< R L <1, the isotherm is favorable
2.1.4. Dubinin-Radushkevich Isotherm The Dubinin-Radushkevich (D-R) adsorption isotherm (Dubinin and Radushkevich, 1966) is given as:
qe qm exp( D 2 )
here, (known as Polanyi potential) is given as (Rosene and Manes, 1977):
(2.6)
RT ln 1
1 Ce
(2.7)
where, R is the gas constant, T is temperature and the constant D (mol2/kJ2) given by the following Eq. (2.8), in terms of the mean free energy E (kJ/mol) of sorption per molecule of adsorbate. When the molecule is transferred to the surface of the solid from infinity in the solution, E is given by:
1 2D
(2.8)
Carbons and Clays for Heavy Metals Removal This isotherm is temperature independent and suitable for porous adsorbents.
2.1.5. Temkin Isotherm The derivation of the Temkin isotherm (Temkin and Pyzhev, 1940) assumes that the fall in the heat of sorption is linear rather than logarithmic, as implied in the Freundlich equation. The Temkin isotherm equation assumes that the heat of sorption of all the molecules in the layer decreases linearly with coverage due to sorbent-sorbate interactions, and that the adsorption is characterized by a uniform distribution of the binding energies, up to some maximum binding energy. The Temkin isotherm has generally been applied in the following form:
qe B ln( K T Ce )
where B
(2.9)
RT H o
(2.10)
Here, B is related to the heat of sorption (J/mol), R the gas constant (8.314 J/mol/K) and T the absolute temperature (K) and KT (L/mg) is the equilibrium binding constant corresponding to the maximum binding energy.
2.1.6. Harkins-Jura Isotherm The HarkinsJura adsorption isotherm can be expressed as (Baar, 2006):
qe
P1 P2 log Ce
(2.11)
where, P1 and P2 are isotherm constants. This model accounts to multi-layer sorption and can be explained with the existence of a heterogeneous pore distribution.
2.1.7. Halsey and Henderson Isotherms The Halsey isotherm (Halsey, 1948) and Henderson isotherm (Henderson, 1952) models explains multilayer sorption. They are generally applied in the following forms, respectively:
ln P3 ln C e qe exp P4
ln(1 C e ) qe P5
1 P6
(2.12)
2.13)
where P3, P4, P5 and P6 are isotherm constants.
2.1.8. Brunauer-Emmet-Teller Isotherm The Brunauer-Emmet-Teller (BET) isotherm (Brunauer et al., 1938) is an S-shaped isotherm, given as:
qe
q m kCe (C0 Ce )[1 (k 1)(Ce / C0 )]
(2.14)
The constant k is large as compared to unity and, therefore, the isotherm consists of two regions, i.e. low and high concentration regions. This isotherm is applicable for multilayer adsorption.
2.1.9. Redlich-Peterson Isotherm Redlich-Peterson sorption isotherm (Redlich and Peterson, 1959) is an empirical isotherm incorporating three parameters. It combines elements from both the Langmuir and Freundlich equations, and the mechanism of sorption is a hybrid and does not follow ideal monolayer sorption. This model is given as:
qe
K RP C e 1 a RP C e
(2.15)
Here KRP is Redlich-Peterson isotherm constant (L/g), aRP is also a constant (L/mg)1/ and is an exponent that lies between 0 and 1. For = 1, Eq. (2.15) reduces to Langmuir equation and for = 0, it reduces to Henrys equation. This three-parameter model approaches the Freundlich model at high concentrations and is in agreement with the low concentration limit of the Langmuir equation.
2.1.10. Sips Isotherm Recognizing the problem of the continuing increase in the adsorbed amount with an increase in concentration in the Freundlich equation, Sips (1948) proposed an equation similar in form to the Freundlich equation, but it has a nite limit when the concentration is sufficiently high. Sips isotherm has the following form:
qe
qm ( K S Ce ) 1 ( K S Ce )
(2.16)
where, qm is the maximum adsorption capacity (mg/g), KS is the Sips isotherm constant (L/g) and is the model exponent. At low sorbate concentrations, Sips equation reduces to the Freundlich isotherm. At high sorbate concentrations, Sips model predicts monolayer sorption capacity and distinctiveness of the Langmuir isotherm.
2.1.11. Radke-Prausnitz Isotherm The Radke-Prausnitz isotherm model (Radke and Prausnitz, 1972) can be expressed as:
qe
q m K rp Ce (1 K rp Ce )
(2.17)
where, Krp is Radke-Prausnitz model constant and the model exponent. It represents the Langmuir model for = 1 and Henrys model for = 0.
2.1.12. Tth Isotherm Tth (2000) has modied the Langmuir equation to reduce the error between experimental data and predicted values of equilibrium adsorption data. The application of this equation is best suited to multi-layer adsorption similar to BET isotherm which is a special type of Langmuir isotherm and has very restrictive validity. The Tth model can be represented as:
qe
q m K Th Ce [1 ( K Th Ce ) ]
1
(2.18)
where KTh is the Tth model constant and is the model exponent. For = 1, this isotherm reduces to the Langmuir adsorption isotherm.
2.1.13. FritzSchluender Isotherm (3-Parameter) The FritzSchluender equation (Fritz and Schluender, 1974) has the following form:
qe
q m K FS C e 1 qm Ce
(2.19)
where KFS the Fritz-Schluender equilibrium constant (L/mg), and is the FritzSchlunder model exponent.
2.1.14. FritzSchluender Isotherm (4-Parameter) FritzSchluender (Fritz and Schluender, 1974) proposed an empirical relation with four parameters to describe equilibrium data and it is Langmuir-Freundlich type of equation.
qe
JC e
K M
1 LCe
(2.20)
where J and L are the FritzSchluender parameters, and K and M are the FritzSchluender equation exponents. At high liquid-phase concentrations of the adsorbate, Eq.(2.20) reduces to the Freundlich equation and for K = M = 1, Eq. (2.20) reduces to the Langmuir equation.
2.1.15. Extended Langmuir Isotherm Originally, the Langmuir model was intended for monolayer adsorption with no interaction between adsorbed molecules. Thus, its ability to correlate experimental data is
limited. The extended Langmuir model (Yao, 2000) is a modified three-parameter version of the Langmuir model, which can provide better fit to the experimental data (Hall et al., 1966).
qe
q m K EL Ce (1 K EL Ce k EL K EL Ce )
(2.21)
where, KEL (L/mg) and kEL (dimensionless) are model parameters. The constant KEL of the model is a coefficient attributed to the affinity between the adsorbent and adsorbate.
2.1.16. Competitive Langmuir Isotherm The difficulties in describing the adsorption of metal ions from wastewater resulted from the presence of several different ions, causing interference and competition on adsorption sites. Therefore, for a wastewater containing two different metal ions, the competitive Langmuir model for two competing ions, can be used in order to express the relationships between the quantity of the first component adsorbed and the concentration of the second component (Ho and McKay, 2000). The model can be expressed as:
q e ,i
q m , i K i C e ,i 1 K i C e ,i K j C e , j
(2.22)
where, Ki and Kj are model parameters, and the subscripts i and j indicate two different metal ions.
2.2. Adsorption Kinetics

Two vital evaluation elements for an adsorption process operation unit are the mechanism and the reaction rate. Solute uptake rate determines the residence time required for completing the adsorption and can be enumerated from kinetic analysis. Numerous attempts were made in formulating a general expression to describe the kinetics of adsorption on solid surfaces for the liquid-solid adsorption system. In 1898, Lagergren presented the pseudo-firstorder rate equation for the adsorption of oxalic acid and malonic acid onto charcoal (Lagergren, 1898; Ho, 2004b). Lagergrens kinetics equation may have been the first one in describing the adsorption of liquid-solid systems based on solid capacity. The two kinetic models, namely pseudo-first-order (Lagergren, 1898; Ho, 2004b) and pseudo-second-order (Ho and McKay, 1999) equations have been widely used to describe adsorption data obtained under non-equilibrium conditions. In most of the adsorption kinetic studies, both pseudo-first-order and pseudo-second-order kinetic equations have been commonly employed parallely and one is often claimed to be better than another according to marginal difference in correlation coefficient. As noted by Rudzinski and Plazinski (2006), in the past decades no attempts were made to clearly explain the theoretical origins of these two equations, i.e., current understanding of adsorption kinetics is much less than theoretical description of adsorption equilibrium. A brief description of various kinetic models is given below.
2.2.1. Fractional Power Model The fractional power function model (Dalal, 1974) is a modified form of the Freundlich equation and may be expressed as:
qt k F t
(2.23)
where, qt is the amount of metal ion sorbed by sorbent at a time t, while kF and are constants with < 1. This expression is generally empirical, except for the case in which = 0.5, where it becomes indistinguishable from the parabolic diffusion equation.
2.2.2. Pseudo-First-Order Model Pseudo-first-order rate equation is expressed as follows (Lagergren, 1898; Ho, 2004b):
dqt k1 (qe qt ) dt
(2.24)
where, k1 is pseudo-first-order rate constant. After integration and applying boundary conditions t = 0 and qt = 0 to t = t and qt = qe at equilibrium, the Eq. (2.24) becomes
qt qe (1 e k1t )
(2.25)
where, value of k1 can be determined from the slope of the plot of log (qe qt) versus t.
2.2.3. Pseudo-Second-Order Model Pseudo-second-order rate equation is expressed as (Ho, 2006a,b):
dqt k 2 (q e qt ) 2 dt
(2.26)
where, k2 is pseudo-second-order rate constant. After integration and applying the same boundary conditions t = 0 and qt = 0 to t = t and qt = qe at equilibrium, Eq. (2.26) becomes
qt
qe k 2 t 1 k 2 qe t
(2.27)
In the linear form Eq. (2.27) becomes,
t 1 t = + 2 qt k 2 q e q e
(2.28)
10
John U. Kennedy Oubagaranadin and Z.V.P. Murthy If the initial adsorption rate is taken as h =
k2qe2. Now, Eq. (2.28) can be rearranged to obtain:
qt ; then when t approaches 0, h will become t
t 1 t qt h q e
(2.29)
2.2.4. Elovich Model The Elovich or RoginskyZeldovich equation (Low, 1960) is commonly used to describe the kinetics of chemisorption of gases onto heterogeneous solids. It is quite restricted as it only describes a limiting condition ultimately reached by the kinetic curve. The Elovich equation in its differential form is represented as,
dqt e exp( e qt ) dt
(2.30)
where, e is the initial adsorption rate (mg/g-min) and e is the desorption constant (g/mg) during any one experiment. Integration of Eq. (2.30), assuming the initial boundary condition qt = 0 at t = 0, gives
qt (1 / e ) ln(1 e e t )
(2.31)
To simplify the Elovich equation, Chien and Clayton (1980) assumed eet >> 1 and applied the boundary conditions qt = 0 at t = 0 and qt = qt at t = t, then the Eq. (2.31) becomes (Sparks, 1986):
qt (1 / e ) ln( e e ) (1 / e ) ln t
(2.32)
Thus, the constants can be obtained from the slope and intercept of the linearized plots of qt against ln(t).
2.2.5. Ritchie Model Ritchie kinetic model (Ritchie, 1977) was obtained assuming that the rate of adsorption depends only on the fraction of sites which are unoccupied at time t and is given as:
1 qt qe 1 r k r t
where, r and kr are constants.
(2.33)
11
2.2.6. Intra-Particle Diffusion The overall adsorption process may be controlled either by one or more steps, e.g., filmdiffusion, pore-diffusion, surface-diffusion and adsorption on the pore surface, or a combination of more than one step. Besides adsorption at the outer surface of the adsorbent, there is also a possibility of intra-particle diffusion of the metal ions from the bulk of the outer surface into the pores of the adsorbent material, which is usually a slow process. The possibility of intra-particle diffusion can be studied using the intra-particle diffusion model (McKay et al. 1987; Weber and Morris, 1963) given by:
qt k ip t I
(2.34)
where, kip is the intra-particle diffusion rate constant and I is a constant that gives idea about the thickness of the boundary layer, i.e., larger the value of I the greater is the boundary layer effect. When intra-particle diffusion is significant in controlling the kinetics of the adsorption process, the plot of qt vs. t yield straight lines passing through the origin and the slope gives the rate constant kip.
2.2.7. Film-Diffusion When the transport of the solute molecules from the liquid-phase to the solid-phase boundary plays a most significant role in adsorption, the liquid film-diffusion model (Boyd et al. 1947) can be applied, which is given as:
ln(1 F ) k fd t
(2.35)
where F is the fractional attainment of equilibrium (F = qt/qe), and kfd is the film diffusion rate constant. A linear plot of ln(1F) versus t with zero intercept would suggest that the kinetics of the adsorption process is primarily controlled by diffusion through the liquid film surrounding the solid adsorbent.
2.3. Thermodynamics
Based on the fundamental thermodynamics concept, it is assumed that in an isolated system, energy cannot be gained or lost and the entropy change is the only driving force. In environmental engineering practice, both energy and entropy factors must be considered in order to determine which process will occur spontaneously. The Gibbs free energy change, G, is the fundamental criterion of feasibility. Reactions occur spontaneously at a given temperature if G is a negative quantity. The Gibbs free energy of an adsorption process, considering the adsorption equilibrium constant KL (which is same as Langmuir constant) is given by the following equation:
G RT ln K L
(2.36)
12
where T is the solution temperature (K) and R is the Universal gas constant (8.314 J/mol/K). The thermodynamic parameters, such as changes in Gibbs free energy (G), enthalpy (H) and entropy (S) were determined by using the equations given below (Smith and Van Ness, 1987; Ho and Ofomaja, 2005, 2006). Considering the relationship between free energy and equilibrium constant, change in equilibrium constant with temperature can be obtained in the differential form as follows:
d H o ln K L dT RT 2
After integration and rearrangement, the Eq.(2.37) becomes:
(2.37)
RT ln K L H TRY
where, Y is integration constant. Let S = RY and substituting G will result into the following, G = H TS
(2.38)
(2.39)
(2.40)
The slope and the intercept of the plot of G versus T were used to determine the S and H values. Basically, the heat evolved during physical adsorption is of the same order of magnitude as the heat of condensation, i.e., 2.120.9 kJ/mol (Sag and Kutsal, 2000), while the heat of chemisorption generally falls in the range of 80200 kJ/mol (Hayward and Trapnell, 1964).
3. ADSORBENTS FOR HEAVY METALS

3.1. Carbons Made from Bio-Materials
Activated carbon has been the most popular and widely used adsorbent in wastewater treatment. In spite of its abundant use, activated carbon remains an expensive material since higher the quality of activated carbon, the greater the operation cost. Activated carbon also requires complexing agents to improve its removal performance for inorganic matters. Activated carbon is generally a good adsorbent for heavy metals in solutions. Nevertheless, it is essential to look for inexpensive sources from which activated carbon can be made. Wang et al. (2010) studied the adsorption of Cd(II) ions from aqueous solution by bamboo charcoal. The results showed that high pH (8.0) was favorable for the adsorption of Cd(II) ions. Higher initial Cd(II) concentrations led to lower removal percentages but higher adsorption capacity. As the adsorbent dose increased, the removal of Cd(II) increased, while
13
the adsorption capacity decreased. Adsorption kinetics of Cd(II) ions onto bamboo charcoal was best described by the pseudo-second-order model. The adsorption of Cd(II) ions followed Langmuir isotherm, with a maximum adsorption capacity of 12.08 mg/g. This study demonstrated that bamboo charcoal could be used for the removal of Cd(II) ions from water treatment. Removal of Cr(VI) ions from aqueous solution was investigated by Owlad et al. (2010) using modified palm shell activated carbon. Low Molecular Weight Polyethyleneimine (LMWPEI) was used for impregnation purpose. The adsorption experiments were carried out in a batch system using potassium dichromate K2Cr2O7 as the source of Cr(VI) in the synthetic waste water and modified palm shell activated carbon as the adsorbent. The effects of pH, concentration of Cr(VI) and PEI loaded on activated carbon were studied. The adsorption data were found to fit well with the Freundlich isotherm model. This modified Palm shell activated carbon showed high adsorption capacity for chromium ions. Monser and Adhoum (2009) modified activated carbon with tartrazine and applied it for the removal of Pb(II), Cd(II) and Cr(III) ions at different pH values. The presence of tartrazine as modifier enhanced attractive electrostatic interactions between metal ions and carbon surface. The adsorption capacity for Pb(II), Cd(II) and Cr(III) ions improved with respect to non-modified carbon reaching a maximum of 140%. The adsorption capacity was found to be pH dependent for both modified and non-modified carbons with a greater adsorption at higher pH values except for Cr(III). The enhancement percent of Pb(II), Cd(II) and Cr(III) at different pH values varied from 28% to 140% with respect to non-modified carbon. Oubagaranadin and Murthy (2009a) prepared physically and chemically activated carbons using sal wood (Shorea robusta) sawdust by thermal process and using sulfuric acid as the activation agent and tested them to remove Pb(II) from aqueous solutions. Adsorption equilibrium studies were done at a pH of 4 and at the room temperature of 30C. It was found that the adsorption isotherms were favorable and chemically activated carbons were better than physically activated carbon in terms of adsorption capacity ( 7 mg/g for chemical activated carbons and 4 mg/g for physically activated carbon). The Freundlich adsorption model provided best-fit. The first-order irreversible unimolecular reaction model and the pseudo-second-order kinetic model provided good fit. Kinetic and film diffusion studies showed that the adsorption of lead(II) on the activated carbons tested were both intra-particle and film diffusion controlled. Tajar et al. (2009) studied adsorption of Cd(II) onto low-cost activated carbons derived from nut shells. Four kinds of activated carbons: prepared activated carbon (PAC), commercial activated carbon (CAC), and sulfurized PAC (SPAC) and sulfurized CAC (SCAC) were tested for the adsorption of Cd(II). With increasing pH, the adsorption of Cd(II) increased and maximum removal was observed to be 92.4% for SPAC at pH values greater than 8.0 (C0 = 100 mg/L). The H-type adsorption isotherms, obtained for the adsorbents, indicated a favorable process. Adsorption data better indicated Frendlich isotherm. The maximum adsorption capacities were 90.09, 104.17, 126.58 and 142.86 mg/g for CAC, PAC, SCAC and SPAC, respectively. Removal of Pb(II), Zn(II), Cu(II), and Cd(II) from aqueous solutions using activated carbon prepared from Phaseolus aureus hulls was studied by Rao et al. (2009). The removal of metal ions by the adsorbent was found to be pH dependent and the optimum pH values were 7.0, 8.0, 7.0 and 6.0 for Cu(II), Cd(II), Zn(II), and Pb(II), respectively. The maximum
14
adsorption capacity values for metal ions were 21.8 mg/g for Pb(II), 21.2 mg/g for Zn(II), 19.5 mg/g for Cu(II) and 15.7 mg/g for Cd(II). It was observed that the kinetic data followed the pseudo-second-order kinetic model. Li and Wang (2009) studied the removal of Pb(II) by low-cost activated carbon prepared from Spartina alterniflora by phosphoric acid activation. It was found that pH played a major role in the adsorption process. The maximum adsorption capacity for Pb(II) on S. alterniflora activated carbon calculated from Langmuir isotherm was about 99 mg/g. The optimum pH range for the removal of Pb(II) was 4.8-5.6. The Freundlich isotherm model best described the experimental data. The kinetic data fitted best to the pseudo-second-order model. Thermodynamic study showed that the adsorption was a spontaneous and exothermic in nature. The adsorption of Cu(II) ions from aqueous solutions by hazelnut shell activated carbon was studied in a batch adsorption system by Demirbas et al. (2009). Maximum adsorption of Cu(II) ions occurred at around pH 6. The adsorption kinetics was best described by the pseudo-second-order rate equation. The adsorption equilibrium data fitted best with the Langmuir isotherm and the monolayer adsorption capacity of Cu(II) ions was determined to be 58.27 mg/g at 323 K. The positive value of the thermodynamic parameter H (18.77 kJ/mol) showed that the adsorption was endothermic. Acharya et al. (2009) studied the adsorption of Pb(II) on activated carbon prepared from Tamarind wood with zinc chloride activation. Adsorption studies were conducted in the range of 1050 mg/L initial Pb(II) concentration and at temperature in the range of 1050C. The maximum adsorption capacity was observed as 43.85 mg/g. The rates of adsorption were found to confirm to pseudo-second-order kinetics with good correlation and the overall rate of Pb(II) uptake was found to be controlled by pore-diffusion, film-diffusion and particlediffusion, in the entire adsorption period. Thermodynamic parameters indicated that the sorption was feasible, spontaneous and endothermic in nature. The use of olive-waste cake, a by-product of the manufacture of olive oil, as a potential feedstock for the preparation of activated carbon was studied by Baccar et al. (2009). Chemical activation of this precursor, using phosphoric acid as dehydrating agent, was adopted. To enhance the adsorption capacity of the optimal activated carbon for heavy metals, a modification of the chemical characteristics of the adsorbent surface was performed, using KMnO4 as oxidant. The efficiency of this treatment was evaluated considering the adsorption of Cu(II) ions as a model for metallic species. The results also indicated that copper uptake capacity was enhanced by a factor of up to 3 for the permanganate-treated activated carbon. The suitability and performance of cattle-manure-compost based activated carbons in removing heavy metal ions from aqueous solution was studied by Zaini et al. (2009). The influence of ZnCl2 activation ratios and solution pH on the removal of Cu(II) and Pb(II) were studied. Batch adsorption technique was used to determine the metal-binding ability of activated carbons. It was found that the uptake of aqueous metal ions by activated carbons well described by Langmuir equation. The increase of surface area and mesopore ratio as a result of increasing activation ratios favored the removal of Cu(II), while activated carbon rich in acidic groups showed selective adsorption towards Pb(II). Activated carbons prepared from chestnut shells and grape-seeds were used as adsorbents for the removal of Cu (II) ions from aqueous solutions by zimen and Ersoy-Meriboyu (2009). Adsorption experiments were performed by varying initial metal ion concentration, temperature and pH. Freundlich isotherm provided better fit to the equilibrium data. It was
15
observed that the adsorption capacities of adsorbents increased with increasing temperature, pH and surface area. The adsorption ability of a powdered activated carbons derived from walnut shells was investigated by Zabihi et al. (2009), in an attempt to produce more economic and effective sorbents for the control of Hg(II) from industrial liquid streams. Carbonaceous adsorbents were prepared by chemical activation methods using ZnCl2 as activating reagent. It was shown that Hg(II) uptake decreased with increasing pH of the solution. The proper choice of preparation conditions resulted in microporous activated carbons with different BET surface areas of 780 (carbon A, 1:0.5 ZnCl2) and 803 (carbon B, 1:1 ZnCl2) m2/g BET surface area. The monolayer adsorption capacities of these particular adsorbents were obtained as 151.5 and 100.9 mg/g for carbons A and B, respectively. It was determined that Hg(II) adsorption followed Langmuir isotherm and pseudo-second-order kinetics. A report was presented on the feasibility of using agricultural waste and timber industry waste carbons by Bansal et al. (2009), to remove Cr(VI) from synthetic wastewater under different experimental conditions. Rice husk and saw dust were used as adsorbents after sulphuric acid treatment. Maximum metal removal was observed at pH 2.0. The efficiencies of rice husk carbon (RHC) and saw dust carbon (SDC) for Cr(VI) removal were 91.75% and 94.33%, respectively, for aqueous solutions (250 mg/L) at 20 g/L adsorbent dose. It was found that Langmuir, Dubinin-Radushkevich and Temkin models fitted well. The results revealed that Cr(VI) was adsorbed considerably on RHC and SDC, and it could be an economical method for its removal from aqueous systems. The surface areas of RHC and SDC were 1.12 and 1.16 m2/g, respectively. Pomegranate husk was converted into activated carbon and tested for its ability to remove hexavalent chromium from wastewater by Nemr (2009). The activated carbon was obtained from pomegranate husk by dehydration process using concentrated sulfuric acid. Batch equilibrium experiments exhibited that a maximum chromium uptake was obtained at pH 1.0. The maximum adsorption capacity for pomegranate husk activated carbon was 35.2 mg/g as calculated by Langmuir model. The ability of activated carbon to remove chromium from synthetic sea water, natural sea water and wastewater was investigated as well. This study showed that the removal of toxic chromium by activated carbon developed from pomegranate husk was promising. Activated charcoal was prepared from Melocanna baccifera (Poaceae) charcoal by chemical pretreatment in order to make better use of this biomass (Lalhruaitluanga et al., 2009). Batch experiments were conducted under varying range of pH (2.06.0), contact time (15360 min) and metal ion concentrations (5090 mg/L). The optimum conditions for lead biosorption are almost same for M. baccifera raw charcoal (MBRC) and M. baccifera activated charcoal (MBAC), viz., pH 5.0, contact time 120 min, adsorption capacity 10.66 mg/g and 53.76 mg/g. The biomass of MBAC was found to be more suitable than MBRC for the development of an efficient adsorbent for the removal of Pb(II) from aqueous solutions. FTIR analysis revealed that -OH, C-H bending, C=O stretching vibration and carbonyl functional groups were mainly responsible for Pb(II) biosorption. A study by Hsu et al. (2009) evaluated the removal of Cr(VI) from water by carbon derived from the burning of rice straw. Rice straw was burned in the air to obtain rice carbon (RC), and then the removal of Cr(VI) by RC was investigated under various pH and ionic strengths. This study revealed that Cr bound to RC was predominately in the trivalent form. The results showed that upon reacting with RC, Cr(VI) was reduced to Cr(III), which was
16
either adsorbed on RC or released back into solution. The extent and rate of Cr(VI) removal increased with decreasing solution pH because the Cr(VI) adsorption and the subsequent reduction of adsorbed Cr(VI) to Cr(III) both occur preferentially at low pH. The minimal effect of ionic strength on the rates of Cr(VI) removal and Cr(III) adsorption indicated specific interactions between Cr(VI)/Cr(III) and their surface binding sites on RC. These results suggest that rice straw-based carbon may be effectively used at low pH as a substitute for activated carbon for the treatment of Cr(VI) contaminated water. In a study by Ghosh (2009), fresh activated carbon (AC) and waste activated carbon (WAC) were pretreated by heating with sulfuric acid and nitric acid at high temperature to prepare different grades of adsorbents and evaluated their performance for Cr(VI) removal from aqueous solution. Batch adsorption equilibrium data followed both, Freuindlich and Langmuir isotherms. Maximum adsorption capacity of the selected adsorbents treated with sulfuric acid (WAC1) and nitric acid (WAC2), calculated from Langmuir isotherm were found to be 7.485 and 10.929 mg/g, respectively. Adsorption capacities for Cd(II) on indigenously prepared, steam activated, untreated, surfactant-modified carbon powder, from husk and pods of Moringa oleifera were investigated by Nadeem et al. (2009). The optimized conditions for all the experimental runs were pH 8.0, temperature 30C, contact time 120 min, agitation speed 160 rpm, initial metal concentration 30 mg/L and adsorbent dosage 1.0 g/L. Maximum Cd(II) removal (98.0%) was observed when cetyltrimethyl ammonium bromide (CTAB), a cationic surfactant-treated carbon, was used as an adsorbent. The Cd(II) removal percentages for sodium dodecyl sulphate (SDS), anionic surfactant, Triton X-100 (non-ionic surfactant) treated and untreated powder activated carbons were found to be 95.60, 81.50 and 73.36%, respectively. SEM images and BET surface area, porosity and pore volume measurements revealed that surfactant-treated carbons had superior porosity and enhanced surface area than untreated carbons. The sorption data correlated better with the Langmuir adsorption isotherm. Oubagaranadin and Murthy (2009b) removed Cr(VI) from aqueous solutions of potassium dichromate by adsorption onto physically and chemically activated carbons derived from Zea mays (corn or maize cob), an agricultural waste product. Adsorption studies were done at the optimum pH of 2 and at the room temperature of 30C. The best fit to the equilibrium data was provided by the 3-parameter Fritz-Schluender model followed by the 2parameter Langmuir model. The adsorption isotherms were favorable and the adsorption on chemically activated carbons improved as the impregnation ratio was increased. Physically activated carbon showed better adsorption capacity (34 mg/g) than the chemically activated carbons (23.24, 27.1 and 33.4 mg/g). Kinetic modeling showed that the fractional power model provided the best fit. The adsorption of Cr(VI) on the activated carbons was filmdiffusion controlled. The use of an activated carbon developed from date palm seed wastes, generated in the jam industry, for removing toxic chromium from aqueous solution was investigated by Nemr et al. (2008). The activated carbon was obtained from date palm seed by dehydrating methods using concentrated sulfuric acid. A strong dependence of the adsorption capacity on pH was observed, the capacity increased as pH value decreased and the optimum pH value was 1.0. The adsorption process was fast and the equilibrium was reached within 180 min. The maximum removal was 100% for 75 mg/L of Cr(VI) concentration on 4 g/L carbon concentration and the maximum adsorption capacity was 120.48 mg/g. The Elovich equation
17
and pseudo-second-order equation provided the greatest accuracy for the kinetic data, and Koble-Corrigan and Langmuir models the closest fits for the equilibrium data. A low-cost activated carbon prepared from Tamarind wood material by chemical activation with sulphuric acid for the adsorption of Pb(II) from dilute aqueous solution was studied by Singh et al. (2008). The activated carbon developed showed substantial capacity to adsorb Pb(II) from dilute aqueous solutions. The kinetic data were best fitted to the Lagergren pseudo-first-order and pseudo-second-order models. The isotherm equilibrium data were well fitted by the Langmuir and Freundlich models. The maximum removal of Pb(II) obtained was 97.95% (experimental) and 134.22 mg/g (from Langmuir isotherm model) at an initial concentration 40 mg/L, adsorbent dose 3 g/L and pH 6.5. This high uptake showed Tamarind wood activated carbon as a promising adsorbent for Pb(II). The use of a date-pit activated carbon and a commercially available activated carbon for the removal of trivalent aluminum from aqueous solutions were examined at various conditions by Al-Muhtaseb et al. (2008). In the acidic range of aluminum solubility (up to pH value of 4), both adsorbents exhibited maximum (almost equivalent) capacities for adsorbing aluminum at the pH value of 4. At low initial concentrations of aluminum and low pH, the uptake of aluminum using date-pit activated carbon was 0.305 mg/g, while that using commercial activated carbon was only 0.021 mg/g. However, the commercial activated carbon was more effective in adsorbing aluminum with high concentrations and low pH. Furthermore, date-pit activated carbon exhibited higher initial adsorption rates as compared to commercial one, which showed higher rates at longer periods of time. Nickel removal efficiency of sulphuric acid-treated Parthenium carbon (SWC) from simulated wastewater was investigated by Lata et al. (2008). Ni(II) removal was pHdependent and found to be maximum at pH 5.0. Both Freundlich and Langmuir adsorption isotherm models represented the experimental data satisfactorily. The monolayer adsorption capacities of SWC as obtained from Langmuir isotherm was found to be 17.24 mg/g. The pseudo-first-order model was less applicable than pseudo-second-order reaction model. The FTIR study indicated the presence of O-H, C-H, C=O and C-O groups in the adsorbent. Production of a modified carbon by heat treating bean husk (Phaseolus vulgaris) at 270 C in Ar, followed by chemical activation using HNO3, was reported by ChvezGuerrero et al. (2008). Cd(II) sorption studies with this material were carried out at different concentrations and found that Cd(II) got effectively removed by the modified material obtained from bean husk (180 mg/g). An activated carbon prepared from olive stone was used for Cd(II) removal by Kula et al. (2008). Different activating agent (ZnCl2) amounts and adsorbent particle sizes were studied to optimize adsorbent surface area. The equilibrium time, optimum pH, adsorbent dosage were found to be 60 min, pH > 6 and 1.0 g/50 mL, respectively. The kinetic data supported pseudo-second-order model and intra-particle model. The activated carbons with 20% ZnCl2 solution was the best sample of the produced activated carbons from olive stone with the specific surface area of 790.25 m2/g. The results showed that the produced activated carbon from olive stone was an alternative low-cost adsorbent for removing Cd(II). Activated carbon prepared from hazelnut husks with zinc chloride activation at 973 K in nitrogen atmosphere was studied for adsorption of Cu(II) and Pb(II) by Imamoglu and Tekir (2008). BET surface area of the activated carbon was found as 1092 m2/g. The activated carbon exhibited good adsorption potential for copper and lead ions. The maximum
18
adsorption capacity of the adsorbent for Cu(II) and Pb(II) ions was calculated from the Langmuir isotherm and found to be 6.645 and 13.05 mg/g, respectively. The applicability of sulphurised activated carbon (SAC) as adsorbent for the effective removal of Co(II) from aqueous solutions was investigated by Krishnan and Anirudhan (2008). Bagasse pith, a sugarcane industry waste, was used for the synthesis of SAC. Maximum adsorption was observed in the pH range of 4.58.5. With an initial concentration of 50 and 100 mg/L of the adsorbate at pH 6.0, the percentage adsorptions were found to be 90.3 and 81.0%, respectively. SAC showed a high adsorption capacity for Co(II) removal when compared with laboratory and commercial grade activated carbons. The adsorption process obeyed Langmuir isotherm model. In a study by Demiral et al. (2008), activated carbon was prepared from olive bagasse by physical activation using steam. BET surface area of the activated carbon was determined as 718 m2/g. The maximum Cr(VI) adsorption yield was obtained at the initial pH of 2. The adsorption kinetics followed pseudo-second-order rate expression. Equilibrium results were analyzed by the Langmuir, Freundlich, Dubinin-Redushkevich, Temkin and Frumkin equations. Langmuir equation was found to fit the equilibrium data for Cr(VI) adsorption. Gerel and Gerel (2007) studied the adsorption of Pb(II) onto activated carbon prepared from renewable plant material, Euphorbia rigida. Adsorption data of Pb(II) onto activated carbon from E. rigida obeyed the Langmuir isotherm model. Maximum adsorption capacity of Pb(II) onto adsorbent was 279.72 mg/g at 40 C. It was indicated that the adsorption of Pb(II) onto activated carbon from E. rigida could be described by the pseudo-second-order kinetic model and also followed the simple external-diffusion model for the initial 10 min and then by intra-particle diffusion model up to 50 min. A carbon rich adsorbent prepared from the reaction of sugar beet pulp with sulphuric acid and gas formed during carbonization process have been studied for Cr(VI) removal from aqueous solutions by Altundogan et al. (2007). The SO2 rich gas was shown to be an excellent Cr(VI) reductant. The equilibrium and kinetic studies were conducted by using the carbonaceous adsorbent derived from sugar beet pulp. Lower pH favored Cr(VI) adsorption, hence substantial Cr(VI) reduction was observed. Langmuir model best fitted the equilibrium isotherm data. The maximum adsorption capacity of chromium calculated from Langmuir isotherm was about 24 mg/g at 25 C. The adsorption of Cr(VI) was endothermic and followed the pseudo-second-order rate kinetics. The sulphuric acid-carbonization was found to be economical particularly for chromium removal because the gas generated during carbonization exhibited good Cr(VI) reduction properties. In a study by Wilson et al. (2006) peanut shells were converted to activated carbons for use in adsorption of select metal ions; namely Cd(II), Cu(II), Pb(II), Ni(II) and Zn(II). Milled peanut shells were pyrolyzed in an inert atmosphere of nitrogen gas, and then activated with steam at different activation times. Following pyrolysis and activation, the carbons underwent air oxidation. The prepared carbons were evaluated for adsorption efficiency and these parameters were compared to the same parameters obtained from three commercial carbons, namely, DARCO 12 20, NORIT C GRAN and MINOTAUR. One of the peanut shell-based carbons had metal ion adsorption efficiency greater than two of the three commercial carbons but somewhat less than but close to MINOTAUR. This study demonstrated that the peanut shells can serve as a source for activated carbons with metal ion-removing potential and may serve as a replacement for coal-based commercial carbons in applications that warrant their use.
19
The performance of a commercially available palm shell based activated carbon to remove Pb(II) from aqueous solutions by adsorption was evaluated by Issabayeva et al. (2006). The adsorption experiments were carried out at pH 3.0 and 5.0. Palm shell activated carbon showed high adsorption capacity for Pb(II), especially at pH 5 with an ultimate uptake of 95.2 mg/g. This high uptake showed palm shell activated carbon as a promising adsorbent. Removal and recovery of chromium were carried out by using low-cost activated carbon made from sugar industry waste (Fahim et al., 2006). Three types of activated carbons, viz., C1 (the waste generated from sugar industry) and the others C2, C3 (commercial granular activated carbons) were used. The effect of pH, particle size and type of adsorbent on the adsorption isotherm of Cr(III) were studied in batch system. The sorption data fitted well with Langmuir adsorption model. The efficiencies of activated carbons for the removal of Cr(III) were found to be 98.86, 98.6 and 93 % for C1, C2 and C3, respectively. The order of selectivity was C1 > C2 > C3 for removal of Cr(III) from tannery wastewater. Carbon C1 of the highest surface area (520.66 m2/g) had the highest adsorptive capacity for the removal of Cr(III). The results revealed that the trivalent chromium was significantly adsorbed on activated carbon collected from sugar industry waste. Kannan and Rengasamy (2005) studied the removal of Cd(II) from aqueous solutions by adsorption on commercial activated carbon (CAC) and chemically prepared activated carbons (CPACs) from raw materials such as straw, saw dust and datesnut. The percentage removal increased with decrease in initial concentration and particle size of CPACs and an increase in contact time, dose and initial pH of the solution. The kinetics of adsorption was found to be first order with intra-particle diffusion as one of the rate determining steps. Results of the studies on adsorption of Cd(II) ions from simulated wastewater were compared with that of CAC and Tulsion CXO-9(H), a commercial ion exchange resin / cationic resin (CR). Straw carbon showed the maximum adsorption capacity towards Cd(II). Granular activated carbon (GAC) was treated chemically with potassium bromate for surface modification and its adsorption capacity was investigated with nickel ions by Satapathy and Natarajan (2006). There was an increase in the adsorption capacity of the modified carbon by 9095% in comparison with the raw granular activated carbon towards nickel ions. Potassium bromate oxidation treatment was employed for a period of about 30 min initially followed by 60 min and the oxidized carbons were adsorbed with nickel ions. Metal adsorption characteristics of as received and modified activated carbons were measured in batch experiments. Equilibrium data fitted well with Langmuir model which indicated monolayer adsorption. Effects of pH of initial solution, time of oxidation and mode of treatment on the adsorption process were studied. Experimental results showed that metal uptake increased with an increase in pH and oxidation time. An efficient adsorption process was reported for the decontamination of Cr(III) from tannery effluents by Mohan et al. (2006). A low cost activated carbon (ATFAC) was prepared from coconut shell fibers and utilized for Cr(III) removal from water/wastewater. A commercially available activated carbon fabric cloth (ACF) was also studied for comparative evaluation. The Langmuir model best fitted the equilibrium isotherm data. The maximum adsorption capacities of ATFAC and ACF at 25 C were 12.2 and 39.56 mg/g, respectively. Cr(III) adsorption increased with an increase in temperature (at 10 C, ATFAC: 10.97 mg/g, ACF: 36.05 mg/g; at 40 C, ATFAC: 16.10 mg/g, ACF: 40.29 mg/g). The adsorption of Cr(III) followed the pseudo-second-order rate kinetics. The sorption capacities of ATFAC
20
and ACF were comparable to many other adsorbents / carbons / biosorbents utilized for the removal of trivalent chromium from water/wastewater. Activated carbon prepared from Ceiba pentandra hulls, an agricultural solid waste byproduct, for the removal of copper and cadmium from aqueous solutions was studied by Rao et al. (2006). The adsorbent exhibited good sorption potential for copper and cadmium at pH 6.0. The C=O and S=O functional groups present on the carbon surface were the adsorption sites to remove metal ions from solution. The maximum adsorption capacity of Cu(II) and Cd(II) was calculated from Langmuir isotherm and found to be 20.8 and 19.5 mg/g, respectively. Desorption studies were carried out using dilute HCl and the effect of HCl concentration on desorption was also studied. Maximum desorption of 90% for Cu(II) and 88% for Cd(II) occurred with 0.2 M HCl. Apricot stones were carbonized and activated after treatment with sulphuric acid (1:1) at 200 C for 24 h by Kobya et al. (2005). The ability of the activated carbon to remove Ni(II), Co(II), Cd(II), Cu(II), Pb(II), Cr(III) and Cr(VI) ions from aqueous solutions by adsorption was investigated. Batch adsorption experiments were conducted to observe the effect of pH (16) on the activated carbon. The adsorption of these metals was found to be dependent on solution pH. Highest adsorption occurred at pH 1-2 for Cr(VI) and pH 3-6 for the rest of the metal ions, respectively. Adsorption capacities for the metal ions were obtained in the order of Cr(VI) > Cd(II) > Co(II) > Cr(III) > Ni(II) > Cu(II) > Pb(II) for the activated carbon prepared from apricot stone. Adsorption capacity of Cr(VI) onto Hevea brasilinesis (rubber wood) sawdust activated carbon was investigated in a batch system by considering the effects of various parameters like contact time, initial concentration, pH and temperature by Karthikeyan et al., 2005. Cr(VI) removal was found to be maximum at pH 2.0. Increase in adsorption capacity with increase in temperature indicated that the adsorption reaction was endothermic. Pseudosecond-order model was found to explain the kinetics of Cr(VI) adsorption most effectively. Intraparticle diffusion studies at different temperatures show that the mechanism of adsorption is mainly dependent on diffusion. Langmuir isotherm showed better fit than Freundlich and Temkin isotherms in the temperature range studied. The results showed that the rubber wood sawdust activated carbon can be efficiently used for the treatment of wastewaters containing chromium as a low cost alternative. Removal of Pb(II) from aqueous solutions by adsorption onto coconut-shell carbon was investigated by Sekar et al. (2004). Adsorption of Pb(II) was strongly affected by pH. The coconut-shell carbon (CC) exhibited the highest lead adsorption capacity at pH 4.5. The equilibrium data fitted well to the Langmuir model. At pH 4.5, the maximum lead adsorption capacity of CC estimated with the Langmuir model was 26.50 mg/g. The thermodynamics of Pb(II) on CC indicated spontaneous and endothermic nature of the adsorption. Quantitative desorption of Pb(II) from CC was found to be 75% which facilitated the adsorption of metal by ion exchange. In a study, the technical feasibility of coconut shell charcoal (CSC) and commercial activated carbon (CAC) for Cr(VI) removal was investigated in batch studies using synthetic electroplating wastewater (Babel and Kurniawan, 2004). Both the granular adsorbents were made up of coconut shells (Cocos nucifera L.). Surface modifications of CSC and CAC with chitosan and/or oxidizing agents, such as sulfuric acid and nitric acid, respectively, were also conducted to improve removal performance. The adsorbents chemically modified with an oxidizing agent demonstrated better Cr(VI) removal capabilities than as-received adsorbents
21
in terms of adsorption rate. Both CSC and CAC, which were oxidized with nitric acid, had higher Cr(VI) adsorption capacities (CSC: 10.88, CAC: 15.47 mg/g) than those oxidized with sulfuric acid (CSC: 4.05, CAC: 8.94 mg/g) and non-treated CSC coated with chitosan (CSCCC: 3.65 mg/g), respectively, suggesting that surface modification of a carbon adsorbent with a strong oxidizing agent generated more adsorption sites on their solid surface for metal adsorption. Hasar (2003) prepared activated carbon from almond husk by activating without (MACI) and with (MAC-II) H2SO4 at different temperatures. The ability of the activated carbon to remove Ni(II) from aqueous solutions by adsorption has been investigated under several conditions such as pH, carbonization temperature of husk, initial concentration of metal ions, contact time, and adsorbent concentration. Optimal conditions were pH 5.0, the carbonization temperature of 700 C, 50 min of contact time and adsorbent concentration of 5 g/L. The results indicated that the effective uptake of Ni(II) ions was obtained by activating the carbon, prepared from almond husk at 700 C, through the addition of H2SO4. The removal of Ni(II) was found to be 97.8% at initial concentration of 25 mg/L and the adsorbent concentration of 5 g/L. When the adsorbent concentration was increased up to 40 g/L, the adsorption capacity decreased from 4.89 to 0.616 mg/g for MAC-II. In the isotherm studies, the experimental adsorption data fitted reasonably well to Langmuir isotherm for both MAC-I and MAC-II. The use of low-cost activated carbon derived from bagasse was investigated by Mohan and Singh, (2002) for the removal of Cd(II) and Zn(II). The uptake of Cd(II) was found to be slightly greater than that of Zn(II) and the adsorption capacity increased with increase in temperature. The adsorption equilibrium data were better fitted by the Freundlich isotherm as compared to Langmuir in both the single- and multi-component systems. The adsorption occurred through a film diffusion mechanism at low as well as at higher concentrations. Activated carbon (AC) prepared from waste Parthenium was used to eliminate Ni(II) from aqueous solution by adsorption (Kadirvelu et al., 2002). The adsorption capacity calculated from the Langmuir isotherm was 54.35 mg Ni(II)/g of AC at initial pH of 5.0 and 20C, for the particle size 250-500 m. Increase in pH from 2 to 10 increased percent removal of the metal ions. The regeneration of Ni(II)-saturated carbon by HCl indicated an adsorption mechanism by ion-exchange between metal ions and H+ ions on the AC surfaces. Quantitative recovery of Ni(II) was possible with HCl. Activated carbon prepared from coconut tree sawdust was used as an adsorbent for the removal of Cr(VI) from aqueous solution by Selvi et al. (2001). Adsorption capacity was calculated from the Langmuir isotherm and was 3.46 mg/g at an initial pH of 3.0 for the particle size 125-250 m. The adsorption of Cr(VI) was pH dependent and maximum removal was observed in the acidic pH range. A summary of carbons made from different starting materials and their adsorption capacities for different heavy metal ions is given in Table 3.1.
3.2. Clays
Clays are one of the prospective low cost alternatives to activated carbons as adsorbents for the heavy metals removal. The clays sorption capabilities come from their high surface area (up to 800 m2/g) and exchange capacities (Cadena et al., 1990). The negative charge on the structure of clay minerals gives clay the capability to attract metal ions. There are three
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basic species of clay, viz., smectites (such as montmorillonite), kaolinite, and micas; out of which montmorillonite has the highest cation exchange capacity. Therefore, a number of studies have been conducted using clays to show their effectiveness for removing metal ions. Table 3.1. Activated carbon adsorbents for heavy metals from solutions
Source material for activated carbon Bamboo Shorea robusta Nut shells Adsorbate metal ion Cd(II) Pb(II) Adsorption capacity,Reference qm (mg/g) 12.08 Wang et al., 2010 PAC: 4.0 Oubagaranadin and CACs : 7.0 Murthy, 2009a Cd(II) CAC: 90.09 Tajar et al., 2009 PAC: 104.17 SCAC: 126.58 SPAC: 142.86 Pb(II), Zn(II), 21.8, 21.2, 19.5, 15.7 Rao et al., 2009 Cu(II), Cd(II) Pb(II) 99 Li and Wang, 2009 Cu(II) 58.27 Demirbas et al., 2009 Pb(II) 43.85 Acharya et al., 2009 Hg(II) A: 151.5; B:100.9 Zabihi et al., 2009 Cr(VI) 35.2 Nemr, 2009 Pb(II) MBRC: 10.66 Lalhruaitluanga et al., MBAC: 53.76 2009 Cr(VI) WAC1: 7.48 Ghosh, 2009 WAC2: 10.93 Cr(VI) PAC: 34 CAC1: 23.24 CAC2: 27.1 CAC3: 33.4 120.48 0.305 17.24 180 279.72 24 95.2 12.2 20.8, 19.5 26.5 10.88 54.35 3.46 Oubagaranadin and Murthy, 2009b
Phaseolus aureus hulls Spartina alterniflora Hazelnut shell Tamarind wood Walnut shells Pomegranate husk Melocanna baccifera Waste activated carbon (WAC) Zea Mays
Date palm seed wastes Date-pit Parthenium Bean husk (Phaseolus vulgaris) Euphorbia rigida Sugar beet pulp Palm shell Coconut shell fibers Ceiba pentandra hulls Coconut-shell Coconut shell Parthenium Coconut tree sawdust
Cr(VI) Al(III) Ni(II) Cd(II) Pb(II) Cr(VI) Pb(II) Cr(III) Cu(II), Cd(II) Pb(II) Cr(VI) Ni(II) Cr(VI)
Nemr et al., 2008 Al-Muhtaseb et al., 2008 Lata et al., 2008 Chvez-Guerrero et al., 2008 Gerel and Gerel, 2007 Altundogan et al., 2007 Issabayeva et al., 2006 Mohan et al., 2006 Rao et al., 2006 Sekar et al., 2004 Babel and Kurniawan, 2004 Kadirvelu et al., 2002 Selvi et al., 2001
Vieira et al. (2010) studied the adsorption of nickel on calcined Bofe bentonite clay. The influence of parameters such as pH, amount of adsorbent, adsorbate concentration and temperature was investigated. The kinetics data were better represented by the second-order model. The Bofe clay removed nickel with a maximum adsorption capacity of 1.91 mg/g of clay (20C, pH 5.3) and that the thermodynamic data indicated that the adsorption reaction was spontaneous and exothermic. The Langmuir model provided the best fit for sorption isotherms.
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Ghorbel-Abid et al. (2009) characterized local clay from Jebel Chakir (Tunisia, North Africa). The clay was basically a smectite with a small proportion of kaolinite. The adsorption properties of the natural clay and the Na-purified clay in a chromium rich aqueous solution was studied by batch technique. The amount of adsorbed chromium ions was determined for the adsorption systems as a function of contact time, pH, adsorbent, and metal ion concentration. The results showed that the uptake of Cr (III) at pH 4, by the purified clay was very rapid. The quantity removed from the solution reached a maximum value at 15 min after mixing, and 1 h for the natural clay, although the latter removes greater quantities of Cr(III) ions compared to the Na-purified clay. The amounts adsorbed by the natural clay were about 117.5 mg of Cr(III)/g of clay and 61.4 mg/g with the Na-purified clay. Moreover, the results showed that the adsorption behavior of both the clays depended highly on the pH. Adsorption increased with the pH of the suspension in the range of 3-5.3. The pH was limited to values equal or less than 5.3 because of the precipitation of the hydroxide chromium at higher pH. The equilibrium data well fitted to the linearized Freundlich isotherm for the natural clay and Langmuir model for the Na-purified clay. Three types of Saudi Arabian clays, viz., Tabuk, Baha, and Khaiber; were tested for their abilities to adsorb Pb(II) from wastewater by Al-Jlil and Alsewailem (2009). The clays were treated with hydrochloric acid to activate adsorption sites within clay particles. Untreated Tabuk clay had the largest adsorption capacity for Pb(II), about 30 mg/g, in comparison with those of Baha and Khaiber clays. Least adsorption was observed with Khaiber clay, about 10 mg/g, which was attributed to the prior existence of lead within the clay. Adsorption of the acid-activated clays was not enhanced compared to those of untreated clays. The Langmuir model described the experimental data for all untreated clays, while the Freundlich model described the experimental data of untreated Khaiber clay and treated Baha clay. The results showed that the Tabuk clay could be utilized as a cost-effective and efficient adsorbent for removing heavy metals from wastewater in Saudi Arabia. Modified kaolinite clay with 25% (w/w) aluminium sulphate and unmodified kaolin were investigated as adsorbents to remove Pb(II) from aqueous solution by Jiang et al., 2009. The results showed that the amount of Pb(II) adsorbed onto modified kaolin (20 mg/g) was more than 4.5 times than that adsorbed onto unmodified kaolin (4.2 mg/g) under optimized conditions. It was observed that the data from both the adsorbents fitted well to the Langmuir isotherm. The kinetic adsorption of modified and unmodified kaolinite clay fitted well to the pseudo-second-order kinetic model. Experiments with real wastewater showed that higher amount of Pb(II) (the concentration reduced from 178 to 27.5 mg/L) and other metal ions were removed by modified kaolinite clay as compared with unmodified adsorbent (the concentration reduced from 178 to 168 mg/L). In a work by Wang et al. (2009), removal of Pb(II) from aqueous solution by adsorption onto Na-bentonite was reported under ambient conditions as a function of shaking time, pH, ionic strength, Na-bentonite content and temperature using batch technique. The kinetic adsorption was well described by the pseudo-second-order rate equation. The adsorption of Pb(II) on Na-bentonite was strongly dependent on pH. The Langmuir model fitted the adsorption isotherm very well. The thermodynamic parameters suggested that the adsorption of Pb(II) was endothermic and spontaneous. At low pH, the adsorption of Pb(II) was dominated by outer-sphere surface complexation and ion exchange with Na+/H+ on Nabentonite surfaces, whereas inner-sphere surface complexation was the main adsorption mechanism at high pH.
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A hectorite (H) clay sample was modified with 2-mercaptobenzimidazole using homogeneous and heterogeneous routes by Guerra et al. (2009). Both modification methodologies resulted in similar products, named HHOM and HHET, respectively. The effect of two variables (contact time and metal concentration) was studied using batch technique at room temperature and pH 2.0. After achieving the best conditions for Cr(VI) adsorption, isotherms of this adsorbate on using the chosen adsorbents were obtained, which were fitted to non-linear Sips isotherm model. The maximum number of moles adsorbed was determined to be 11.63, 12.85 and 14.01 mmol/g for H, HHOM and HHET, respectively, reflecting the maximum adsorption order of HHET > HHOM > H. Batch sorption experiments were conducted with cadmium and lead ions at low equilibrium concentrations in 0.01 M of NaNO3 onto Petra clay in single component systems by Baker (2009). The equilibrium isotherms were determined at pH 6 under constant ionic strength and at different temperature. From the Langmuir isotherm, the equilibrium adsorption capacity for Cd(II) was found to be 74.074 to 144.927 mg/g and that for Pb(II) was 83.333 to 263.158 mg/g. The results showed that Petra clay, which was mainly composed of 20% of kaolinite and 55% of calcium montmorillomite, exhibited higher selectivity for Pb(II), whereas its selectivity for Cd(II) was often lower at all concentrations studied. From the R2 values for different isotherm models it was found that the sorption was good for the two metal ions and good correlation confirmed the formation of a monolayer of Cd(II) and Pb(II) on the surface of the clay. Isotherms analysis showed that the binding for these metal ions with Petra clay minerals was physisorption and the process was endothermic. Oubagaranadin and Murthy (2009c) studied the removal of Pb(II) from aqueous solution by adsorption on a montmorillonite-illite type of clay (MIC) collected from the Gulbarga district of Karnataka, India. Batch adsorption equilibrium data were determined with different initial Pb(II) concentrations (100, 150, and 200 ppm) at pH 4 and 37 C, and the data were tested with isotherm models. The three-parameter Freundlich-Langmuir model gave the best fit to the equilibrium data. However, as the initial Pb(II) concentration was increased (150 and 200 ppm), then multi-layer adsorption was observed. The maximum monolayer adsorption capacity of the clay was determined to be ~ 52 mg/g. Kinetic studies indicated that the rate of adsorption of Pb(II) on the clay followed a second-order rate mechanism, with decreasing rate constant values of 0.1097, 0.0571, and 0.0022 g/(mg min) as the initial Pb(II) concentration was increased in the order of 100, 150, and 200 ppm, respectively. The value of the Freundlich constant (n) in the range of 2.5-4.6 indicated that MIC was a good adsorbent of divalent lead. At a higher initial Pb(II) concentration (200 ppm), the adsorption process was determined to be film-diffusion controlled, with a rate of 0.051 min-1. The mean values of the thermodynamic parameters showed that the adsorption process was endothermic, thermodynamically favorable, and spontaneous. The proposed two-stage adsorber system has reduced the clay dose by 8.5%, as compared to that of a single-stage adsorption system. In another work by Oubagaranadin and Murthy (2010), the natural montmorillonite-illite clay was activated with sulfuric acid and used for lead(II) removal. Raw clay disintegrated on acid activation and showed a particle size distribution. The montmorillonite and illite phases in the raw clay disappeared on acid activation and the activated clay, showed with sodiumaluminum-silicate and beidellite phases apart from quartz (low) phase. When tested for adsorption of Pb(II) in aqueous solutions, the acid-activated clay showed about 50% increased adsorption than raw clay. Sips adsorption isotherm and pseudo-second-order kinetic models were found to be best for the batch adsorption data. Kinetic studies showed the
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existence of film diffusion and intraparticle diffusion. A two-stage batch adsorber was designed for the removal of Pb(II) from aqueous solutions. There was about 63% increase in the monolayer adsorption capacity of the clay adsorbent due to acid activation. From the twostage batch adsorption system proposed using the experimental data, it was observed that the two-stage system reduced the adsorbent dose by about 14 and 30% of raw and activated clay, respectively, when compared with that of single-stage adsorption system. Adsorption of Cu(II) ions on a zeolite, clay and diatomite from Serbia was studied by ljivi et al. (2009), at different pH. The amounts of Cu(II) removed from the solution increased with increasing initial pH, reaching nearly 100% at pH > 7, due to precipitation of Cu(OH)2. Relatively constant final pH values and less significant increase of Cu(II) uptake was observed in the initial pH range 46, which have pointed out the role of buffering properties of investigated adsorbent materials. The maximum adsorption capacities decreased in the order of zeolite (0.128 mmol/g) > clay (0.096 mmol/g) > diatomite (0.047 mmol/g). The Langmuir isotherm gave the best fit. Ion exchange of exchangeable cations and protons were identified as main adsorption mechanisms. Removal of Cr(III) and Cr(VI) from aqueous solutions by white, yellow and red sands from the United Arab Emirates, as low cost abundant adsorbents, was investigated by Khamis et al. (2009). The effect of contact time, pH, temperature, metal concentration and sand dosage were studied. The optimal pH for adsorption was 5.0 for Cr(III) and 2.0 for Cr(VI). The optimal adsorption time for both ions was 3 h. Even at the optimal pH, adsorption of Cr(VI) on all sand forms was very low (removal 10%) and could not be fitted to any of the common isotherms. While at pH 5.0, Cr(VI) was not at all adsorbed and Cr(III) was totally removed. Adsorption of Cr(III) by the three sand forms obeyed Lagergren first-order kinetics. For Cr(III), the Langmuir isotherm gave best fit for adsorption. At 25 C, the maximum mass of Cr(III) removed per gram of sand was 62.5, 9.80 and 2.38 (mg/g) for white, yellow and red sands, respectively. H was 14.5, 51.2 and 45.8 kJ/mol and S was 24.0, 136 and 111 J /K/mol for adsorption on white, yellow and red sands, respectively. Unuabonah et al. (2008a) studied the adsorption behavior of a polyvinyl alcohol modified (PVA-modified) kaolinite clay. Modification of kaolinite clay with PVA increased its adsorption capacity for 300 mg/L Pb(II) and Cd(II) by a factor of about 6, i.e., from 4.5 mg/g to 36.23 mg/g and from 4.38 mg/g to 29.85 mg/g, respectively, at 298 K. Binary mixtures of Pb(II) and Cd(II) decreased the adsorption capacity of unmodified kaolinite clay for Pb(II) by 26.3% and for Cd(II) by 0.07%, respectively. For PVA-modified kaolinite clay, the reductions were up to 50.9% and 58.5% for Pb(II) and Cd(II), respectively. The adsorption data of Pb(II) and Cd(II) onto both unmodified and PVA-modified kaolinite clay adsorbents were found to fit the pseudo-second-order kinetic model, indicating that adsorption on both the surfaces was mainly by chemisorption. Kaolinite clay obtained from Ubulu-Ukwu in Nigeria was modified with sodium tetraborate (NTB) to obtain NTB-modified kaolinite clay by Unuabonah et al. (2008b). Modification with sodium tetraborate reagent increased the adsorption capacity of kaolinite clay from 16.16 mg/g to 42.92 mg/g for Pb(II) and 10.75 mg/g to 44.05 mg/g for Cd(II) at 298 K. Increasing temperature was found to increase the adsorption of both the metals onto both the adsorbents suggesting endothermic adsorption. The simultaneous presence of electrolyte in aqueous solution with Pb(II) and Cd(II) was found to decrease the adsorption capacity of NTB-modified adsorbent for Pb(II) and Cd(II). The thermodynamic calculations for the modified kaolinite clay sample indicated an endothermic nature of adsorption and an
26
increase in entropy as a result of adsorption of Pb(II) and Cd(II). The small positive values of free energy change indicated that the adsorption of Pb(II) and Cd(II) onto the modified adsorbent may require some small amount of energy to make it more feasible. Modeling equilibrium adsorption data suggested that NTB-modified adsorbent sample had homogeneous adsorption sites and fitted well with Langmuir model. NTB-modified kaolinite clay sample showed good potentials as a low-cost adsorbent for the adsorption of Pb(II) and Cd(II) from aqueous solutions. Adsorption of Cu(II) by raw bentonite (RB) and acid-activated bentonite (AAB) samples was investigated by Eren and Afsin (2008) as a function of initial Cu(II) concentration, solution pH, ionic strength, temperature, the competitive and complexation effects of ligands (Cl, SO42, PO43). Langmuir monolayer adsorption capacity of the RB (42.41 mg/g) was found to be greater than that of the AAB (32.17 mg/g). The spontaneity of the adsorption process was established by decrease in G which varied from 0.34 to 0.71 kJ/mol (RB), 1.13 to 1.49 kJ/mol (AAB) in the temperature range of 303-313 K. Infrared (IR) spectra of the bentonite samples showed that the positions and shapes of the fundamental vibrations of the OH and Si-O groups were influenced by the adsorbed Cu(II) cations. Differential thermal analysis (DTA) results showed that adsorbed Cu(II) cations have a great effect on the thermal behavior of the bentonite samples. The X-ray diffraction (XRD) spectra indicated that the Cu(II) adsorption onto the bentonite samples led to changes in unit cell dimensions and symmetry of the parent bentonites. Eren (2008) reported the adsorption of Cu(II) from aqueous solution on modified Unye bentonite. Adsorption of Cu(II) by manganese oxide modified bentonite (MMB) sample was investigated as a function of the initial Cu(II) concentration, solution pH, ionic strength, temperature and inorganic ligands (Cl, SO42, HPO42). The adsorption properties of raw bentonite were further improved by modification with manganese oxide. Langmuir monolayer adsorption capacity of the MMB (105.38 mg/g) was found to be greater than that of the raw bentonite (42.41 mg/g). The spontaneity of the adsorption process was established by decrease in G which varied from 4.68 to 5.10 kJ/mol in temperature range of 303 313 K. The high performance exhibited by MMB was attributed to the increased surface area and higher negative surface charge acquired after modification. The adsorption of Pb(II) onto Tunisian smectite-rich clay in aqueous solution was studied in a batch system by Chaari et al. (2008). In this study, four samples of clay (AYD, AYDh, AYDs, AYDc) were used. The AYD was raw clay. AYDh and AYDs correspond to AYD activated by 2.5 mol/L hydrochloric acid and 2.5 mol/L sulphuric acid, respectively. AYDc corresponds to AYD calcined at different temperatures (100, 200, 300, 400, 500 and 600 C). Preliminary adsorption tests showed that sulphuric acid and hydrochloric acid activation of raw AYD clay enhanced its adsorption capacity for Pb(II). However, the uptake of Pb(II) by AYDs was very high compared to that of AYDh. This fact was attributed to the greater solubility of clay minerals in sulphuric acid compared to hydrochloric acid. Thermo activation of AYD clay reduced the Pb(II) uptake as soon as calcination temperature reaches 200 C. Kinetic experiments showed that the sorption of lead ions on AYDs was very fast and the equilibrium was practically reached after only 20 min. The results also revealed that the adsorption of lead increased with increase in the solution pH from 1 to 4.5 and then decreased slightly between pH 4.5 and 6, and rapidly at pH 6.5 due to the precipitation of Pb(II) ions. The equilibrium data were analyzed using Langmuir isotherm model. The maximum adsorption capacity increased from 25 to 25.44 mg/g with increasing temperature from 25 to
27
40 C. It was observed that sulphuric acid activated clay was more efficient than physically activated clay. Adsorption of Cr(VI) onto spent activated clay (SAC), a waste produced from an edible oil refinery company, was investigated for its beneficial use in wastewater treatment by Weng et al. (2008). After pressure steam treatment, SAC was used as an adsorbent. The adsorption kinetic data were analyzed and fitted well with pseudo-first-order equation and the rate of removal was found to speed up with decreasing pH and increasing temperature. The maximum adsorption capacities for Cr(VI) were ranged from 0.743 to 1.422 mg/g for temperature between 4 and 40 C under a condition of pH 2.0. The studies conducted show the process of Cr(VI) removal to be spontaneous at high temperature and endothermic in nature. Kaolinite and montmorillonite were used as adsorbents for Fe(III), Co(II) and Ni(II) in aqueous medium by Bhattacharyya and Gupta (2008). The effect of different variables, namely, concentration of metal ions, amount of clay adsorbents, pH, time and temperature of interaction was investigated. Adsorption increased with pH till precipitation of insoluble hydroxides became dominant. The processes conformed to second-order kinetics. Montmorillonite had a much higher adsorption capacity for the metal ions with the Langmuir monolayer capacity of 28.4 to 28.9 mg/g compared to that of 10.4 to 11.2 mg/g for kaolinite. All the interactions were exothermic except those between Co(II) and kaolinite. The adsorption processes were accompanied by an appreciable decrease in Gibbs energy. Both kaolinite and montmorillonite were observed to be suitable for treating water contaminated with Fe(III), Co(II) and Ni(II). Argun (2008) described the removal of Ni(II) ions from aqueous solutions using clinoptilolite. The effect of clinoptilolite level, contact time, and pH were determined. Different isotherms were also obtained using concentrations of Ni(II) ions ranging from 0.1 to 100 mg/L. The ion-exchange process followed second-order kinetics and the Langmuir isotherm. The work revealed that the ion-exchange process was spontaneous and exothermic under natural conditions. The maximum removal efficiency obtained was 93.6% at pH 7 and with a 45 min contact time (for 25 mg/L initial concentration and a 15 g/L solid-to-liquid ratio). The use of bentonite for the removal of Pb(II) from aqueous solutions for different contact times, pH of suspension, and initial concentration of lead and particle sizes of absorbent was investigated by Zhu et al. (2008). Batch adsorption kinetic experiments revealed that the adsorption of Pb(II) onto bentonite involved fast and slow processes. The adsorption mechanisms in the lead/bentonite system followed pseudo-second-order kinetics with a significant contribution of film-diffusion. The Langmuir model represented the adsorption process. The maximum adsorption capacity of Pb(II) onto natural bentonite was 78.82 mg/g. oruh (2008) investigated the effects of conditioning with NaCl and HCl solutions on the removal of Zn(II) from aqueous solutions using natural clinoptilolite collected from ManisaGrdes region of Turkey. The clinoptilolite sample was used in four different forms, which consisted of one unconditioned (NC) and three conditioned (CC1, CC2 and CC3). The results clearly showed that the conditioning improved both the exchange capacity and the removal efficiency. It was found that the highest removal efficiency was obtained with CC2 sample. Adsorption isotherm of Zn(II) was best modeled by the Langmuir equation. The maximum adsorption capacities for Zn(II) shown by NC, CC1, CC2 and CC3 samples were 21.2, 20.8,
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22.2 and 17.9 mg/g, respectively. Results indicate a significant potential for the natural and conditioned clinoptilolites as an adsorbent/ion-exchange material for heavy metal removal. Trivalent chromium was removed from synthetic wastewater using low-cost diatomite in batch and continuous systems by Gr et al. (2008). In batch system, four different sizes and five different amounts of adsorbent were used. Langmuir adsorption capacities were found to be 28.1, 26.5 and 21.8 mg Cr(III)/g diatomite at 15, 30 and 45 C, respectively. Adsorption process was exothermic in nature. The kinetic data showed that the pseudo-second-order equation was more appropriate, which indicated that the intra-particle diffusion is the ratelimiting factor. Laboratory grade Fullers earth (FE) was used as an adsorbent in a work by Oubagaranadin et al. (2007) to remove mercury from aqueous solutions. For the purpose of comparison, simultaneous experiments using laboratory grade activated carbon (AC) was also done. Isotherms such as Freundlich, Langmuir, DubininRadushkevich, Temkin, HarkinsJura, Halsey and Henderson were tested to the equilibrium data. Kinetic studies based on Lagergren first-order, pseudo-second-order rate expressions and intra-particle diffusion studies were done. The batch experiments were conducted at room temperature (30C) and at the normal pH (6.7) of the solution. It was observed that Hg(II) removal rate was better for FE than AC, and the adsorption capacity of AC (69.44 mg/g) was found to be much better than FE (1.15 mg/g). Hybrid fractional error function analysis showed that the best-fit for the adsorption equilibrium data were represented by Freundlich isotherm. Kinetic and filmdiffusion studies showed that the adsorption of mercury on FE and AC was both intra-particle diffusion and film-diffusion controlled. In a study by Abu-El-Shar and Haddad (2007), lead adsorption onto soil samples from Irbid, which were subjected to high temperatures, was investigated. These samples were collected from ground surface and heated to temperatures of 25, 70, 100, 200, 225, 250, 275, 300, 400, and 550C. Based on these temperatures the soil was divided into ten different groups. Each group was first characterized by conducting a set of experiments to estimate the liquid limit (LL), plastic limit (PL), and plasticity index, the organic carbon content, and a set of batch experiments to study lead adsorption. Results indicated that the LL, PL, total organic carbon were slightly affected by high temperatures less than 200C, showed an abrupt change between temperature from 200 and 300C, and then slight change above 300C. Adsorption of lead onto heated samples, however, was not significantly changed. This may be explained by the fact that adsorption of heavy metals mainly occurs onto the soil mineral parts which are slightly affected by the temperature range used in this study. The adsorption of Pb(II) onto Turkish (Bandirma region) kaolinite clay was examined in aqueous solution with respect to the pH, adsorbent dosage, contact time, and temperature by Sari et al. (2007). The linear Langmuir and Freundlich models were applied to describe equilibrium isotherms and both models fitted well. The monolayer adsorption capacity was found as 31.75 mg/g at pH 5 and 20 C. DubininRadushkevich isotherm model was also applied to the equilibrium data. The mean free energy of adsorption (13.78 kJ/mol) indicated that the adsorption of Pb(II) onto kaolinite clay may be carried out via chemical ion-exchange mechanism. The adsorption of Pb(II) onto kaolinite clay was feasible, spontaneous and exothermic in nature. Furthermore, the experimental data fitted well with the pseudo-secondorder kinetics. Removal of copper and zinc from aqueous solutions was investigated by Veli and Alyz, (2007) using Cankiri bentonite natural clay. The effects of pH, clay amount, heavy metal
29
concentration and agitation time on adsorption efficiency were studied. Langmuir, Freundlich and DubininRadushkevich isotherms were applied in order to determine the efficiency of natural clay used as an adsorbent. Results showed that all isotherms were linear. It was observed that adsorption data of Cu(II) and Zn(II) were well-fitted by the second-order reaction kinetics. In addition, calculated and experimental heavy metal amounts adsorbed by the unit clay mass were too close to each other. It was concluded that natural clay could be used as an effective adsorbent for removing Cu(II) and Zn(II) from aqueous solutions. A work by Santos and Masini (2007) presented an evaluation of vermiculite as a low cost adsorbent for treatment of wastewater from a coatings industry containing Cd(II), Pb(II) and Cu(II). Adsorption data were fitted by Freundlich isotherms, as well as by partition constants at pH 4, 5 and 6. It was observed that the non-expanded vermiculite presented the following affinity orders by the studied ions: pH 4: Cu(II) < Cd(II) < Pb(II); pH 5: Cu(II) Cd(II) < Pb(II); pH 6: Cd(II) < Cu(II) < Pb(II). For Pb(II) and Cd(II), the adsorption percentages determined in real wastewaters were around 20% lower than the removal percentage previewed by the Freundlich parameters, a fact that may be explained by competition of the studied cations among themselves, and with Fe(III) species, which were present in the water at concentration levels similar to Cd(II), Cu(II) and Pb(II). Additionally, the high content of organic compounds in the wastewater might have decreased the adsorption of Cu(II) and Pb(II) because of possible formation of soluble complexes between the heavy metal cations and the organic compounds. On the other hand, adsorption of Cd(II) from the real wastewater was about 20% higher than that previewed by the Freundlich parameters, denoting the complexity of interactions that ions are liable in matrices of wastewaters. A series of activated palygorskite clays by HCl with different concentrations were prepared and applied as adsorbents for removal of Cu(II) from aqueous solutions by Chen et al. (2007). The results showed that adsorption capacity of activated palygorskites increased with increasing the HCl concentration and the maximum adsorption capacity with 32.24 mg/g for Cu(II) was obtained at 12 mol/L of HCl concentration. Kinetic studies indicated that the adsorption mechanisms in the Cu(II)/acid-activated palygorskite system followed the pseudosecond-order kinetic model with a relatively small contribution of film-diffusion. Equilibrium data fitted well with the Freundlich isotherm model compared to the Langmuir isotherm model, indicating that adsorption taking place on heterogeneous surfaces of the acid-activated palygorskite. The use of natural palygorskite clay for the removal of Pb(II) from aqueous solutions for different contact times, pH of suspension, adsorbent amounts and particle sizes of palygorskite clay were investigated by Chen and Wang (2007). Batch adsorption kinetic experiments revealed that the adsorption of Pb(II) onto palygorskite clay involved fast and slow processes. It was found that the adsorption mechanisms in the lead/palygorskite system followed pseudo-second-order kinetics with a significant contribution from film-diffusion. The Langmuir model represented the adsorption process better than the Freundlich model. The maximum adsorption capacity of Pb(II) onto natural palygorskite was found to be 104.28 mg/g. Adsorption of Pb(II) ions from aqueous solution onto clinoptilolite was investigated by Gnay et al. (2007) to evaluate the effects of contact time, initial concentration and pretreatment of clinoptilolite on the removal of Pb(II). Maximum experimental adsorption capacity was found to be 80.933 and 122.400 mg/g for raw and pretreated clinoptilolite,
30
respectively, for the initial concentration of 400 mg/L. Results of the kinetic studies showed that the best fitted kinetic models were obtained to be in the order: the pseudo-first-order, the pseudo-second-order and Elovich equations. The negative value of change in Gibbs free energy (G) indicates that the adsorption of Pb(II) on clinoptilolite was spontaneous. Bhattacharyya and Gupta (2007) investigated the influence of acid activation of montmorillonite on adsorption of Cd(II), Co(II), Cu(II), Ni(II), and Pb(II) from aqueous medium and comparison of the adsorption capacities with those on parent montmorillonite. The clay-metal interactions were studied under different conditions of pH, concentration of metal ions, amount of clay, interaction time, and temperature. The interactions were dependent on pH and the uptake was controlled by the amount of clay and the initial concentration of the metal ions. The adsorption capacity of acid-activated montmorillonite increased for all the metal ions. The interactions were adsorptive in nature and relatively fast and the rate processes were more akin to the second-order kinetics. The Langmuir monolayer capacity of the acid-activated montmorillonite was more than that of the parent montmorillonite (Cd(II): 32.7 and 33.2 mg/g; Co(II): 28.6 and 29.7 mg/g; Cu(II): 31.8 and 32.3 mg/g; Pb(II): 33.0 and 34.0 mg/g; and Ni(II): 28.4 and 29.5 mg/g for montmorillonite and acid-activated montmorillonite, respectively). The thermodynamics of the rate processes showed that the adsorption of Co(II), Pb(II), and Ni(II) to be exothermic, accompanied by decreases in entropy and Gibbs free energy, while the adsorption of Cd(II) and Cu(II) was endothermic, with an increase in entropy and an appreciable decrease in Gibbs free energy. The results established the potential use of montmorillonite and its acid-activated form as adsorbents for Cd(II), Co(II), Cu(II), Ni(II), and Pb(II) ions from aqueous media. Application of riverbed sand for the adsorptive separation of Cd(II) from aqueous solutions has been investigated by Sharma et al. (2007). Metal removal increased from 26.8 to 56.4% by decreasing the initial concentration of cadmium from 7.5 105 to 1.0 105 M at pH 6.5, 25 C temperature, agitation speed of 100 rpm, 100 m particle size and 1.0 102 NaClO4 ionic strength. Process of separation is governed by first-order rate kinetics. Values of thermodynamic parameters Go, Ho and So were also calculated and were recorded as 0.81 kcal/mol, 9.31 kcal/mol and 28.10 kcal/mol, respectively, at 25C. The solution pH has been found to affect the removal of cadmium significantly and maximum removal (58.4%) has been found at pH 8.5. In a work by Shawabkeh et al. (2007), natural bentonite was treated by hydrochloric, nitric, and phosphoric acids followed by washing with sodium hydroxide in order to enhance its adsorption capacity. The sample which was treated with hydrochloric acid, followed by further treatment with NaOH, showed the highest cation exchange capacity with a value of 51.20 meq/100 g. Adsorption isotherms for both cobalt and zinc were fitted using Langmuir, Freundlich, and Redlich-Peterson and showed an adsorption capacity of 138.1 mg Co(II) and 202.6 mg Zn(II) per gram of treated adsorbent sample. The adsorption characteristics of palygorskite with respect to cadmium were studied with the aim of assessing its use in water purification systems by lvarez-Ayuso and GarcaSnchez (2007). The adsorption of Cd on palygorskite appeared as a fast process, with equilibrium being attained within the first half-an-hour of interaction. This process was described by the Langmuir model and gave a maximum Cd sorption of 4.54 mg/g. This sorption capacity value was greatly affected by both pH and ionic strength. High competing electrolyte concentrations have decreased the amount of Cd sorbed (close to 60%), suggesting a great contribution of replacement of exchange cations in this metal removal by palygorskite.
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The increase of mineral dose provoked a Cd removal raise; removal values in the range of 8545% were attained for Cd initial concentrations of 10150 mg/L (0.0891.34 mmol/L) when the palygorskite dose of 20 g/L was employed. Column studies were also performed in order to estimate the potential of palygorskite to be used in continuous flow purification systems, showing the effectiveness of this mineral to purify, down to the legal limit of waste, moderate volumes of Cd-containing solutions with a similar concentration (50 mg/L or 0.445 mmol/L) to those mostly found in the upper range of concentrations usually present in industrial wastewaters. In a study by Kubilay et al. (2007), the removal of Cu(II), Zn(II) and Co(II) ions from aqueous solutions using adsorption onto natural bentonite was investigated as a function of initial metal concentration, pH and temperature. For all the metal cations studied, the maximum adsorption was observed at 20C. The batch method was employed using initial metal concentrations ranging from 15 to 70 mg/L at pH 3.0, 5.0, 7.0 and 9.0. It was found that in every concentration range, adsorption data of bentonitic clay - heavy metal cations, matched to Langmuir, Freundlich and Dubinin-Kaganer-Radushkevich (DKR) isotherms. Bentonite used was sensitive to pH changes and the amounts of heavy metal cations adsorbed increased as pH was increased in adsorbent-adsorbate system. According to the adsorption equilibrium studies, the selectivity order was found to be: Zn(II)>Cu(II)>Co(II). These results showed that bentonitic clay holds great potential to remove the relevant heavy metal cations from wastewater. The sorption of Cr(III) from aqueous solutions on kaolinite was studied by a batch technique by Turan et al. (2007). The adsorbed amount of chromium ions on kaolinite increased with increasing pH and temperature and decreased with increasing ionic strength. The sorption of Cr(III) on kaolinite was found to be endothermic. Sorption data have been interpreted in terms of Freundlich and Langmuir equations. The experimental data were correlated reasonably well by the Langmuir adsorption isotherm. The enthalpy change for chromium adsorption was estimated to be 7.0 kJ/mol. In a work by Manohar et al. (2006), a natural bentonite clay collected from Ashapura Clay Mines, Gujarat State, India, was utilized as a precursor to produce aluminum-pillared bentonite clay (Al-PILC) for the removal of Co(II) ions from aqueous solutions. Adsorption experiments were conducted under various conditions, i.e., pH, contact time, initial concentration, ionic strength, adsorbent dose and temperature. The most effective pH range for the removal of Co(II) ions was found to be 6.08.0. The maximum adsorption of 99.8% and 87.0% took place at pH 6.0 from an initial concentration of 10.0 and 25.0 mg/L, respectively. Kinetic studies showed that an equilibrium time of 24 h was needed for the adsorption of Co(II) ions on Al-PILC and the experimental data were correlated by the external mass transfer diffusion model for the first-stage of adsorption and the intraparticle mass transfer diffusion model for the second-stage of adsorption. The intraparticle mass transfer diffusion model gave a better fit to the experimental data. The equilibrium isotherm data were analyzed using the Langmuir, Freundlich and Scatchard isotherm equations and the adsorption process was expressed by Freundlich isotherm. The adsorption behavior of vermiculite was studied with respect to cadmium, copper, lead, manganese, nickel, and zinc as a function of pH and in the presence of different ligands by Malandrino et al. (2006). The continuous column method was used in order to evaluate the feasibility to use the clay in wastewater purification systems. The total capacity of vermiculite was found to increase in the following order: Mn > Ni > Zn > Cd > Cu > Pb. The adsorption
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of metal ions on vermiculite decreased with decreasing pH and increasing ionic strength. The metal uptake on the clay was hindered by the presence of strong complexing agents in solution and it decreased with increasing of the complexation of the ligands with exception of cysteine and tiron. It was concluded that the vermiculite has good potentialities for costeffective treatments of metal-contaminated wastewaters. Natural Jordanian adsorbent (consisting of quartz and aluminosilicates and secondary minerals, i.e., calcite and dolomite) was shown to be effective for removing Zn(II), Pb(II) and Co(II) from aqueous solution as reported by Al-Degs et al. (2006). The major mineral constituents of the sorbent were calcite and quartz. Dolomite was present as minor mineral and palygorskite was present as trace mineral. The sorbent had microporous structure with a modest surface area of 14.4 m2/g. The adsorption capacities of the metals were: 2.86, 0.32, 0.076 mmol cation/g for Zn(II), Pb(II) and Co(II) at pH 6.5, 4.5 and 7.0, respectively. Adsorption data of metals were described by Langmuir and Freundlich models over the entire concentration range. It was found that the mechanism of metal adsorption was mainly due to the precipitation of metal carbonate complexes. The overall adsorption capacity has decreased after acid treatment, as this decreases the extent of precipitation on calcite and dolomite. Kinetic data were accurately fitted to pseudo-first-order and external diffusion models, which indicated that adsorption of Zn(II) occurred on the exterior surface of the adsorbent and the contribution of internal diffusion mechanism was insignificant. Gupta and Bhattacharyya (2006) investigated the adsorptive interactions of Ni(II) ions with kaolinite, montmorillonite, and their poly(oxo-zirconium) and tetrabutylammonium (TBM) derivatives in aqueous medium. The adsorption strongly depended on pH of the medium with enhanced adsorption as the pH turns from acidic to alkaline side till precipitation started. The process was fast initially and maximum adsorption was observed within 180 min of agitation. The kinetics of the interactions showed better agreement with second-order kinetics. The adsorption data gave Langmuir monolayer capacity of 2.75 to 21.14 mg/g for the clay adsorbents. The adsorption process was exothermic accompanied by decrease in entropy and Gibbs free energy. The results showed that montmorillonite had the largest adsorption capacity followed by ZrO-montmorillonite, TBA-montmorillonite, kaolinite, ZrO-kaolinite and TBA-kaolinite. Introduction of ZrO- and TBA- groups into the clays reduced their adsorption capacity by blocking the available adsorption sites. Bhattacharyya and Gupta (2006) investigated the removal of Fe(III) ions from an aqueous solution using kaolinite, montmorillonite and their acid activated forms. The specific surface areas of kaolinite, acid activated kaolinite, montmorillonite and acid activated montmorillonite were 3.8, 15.6, 19.8 and 52.3 m2/g, respectively, whereas the cation exchange capacity (CEC) was measured as 11.3, 12.2, 153.0, and 341.0 meq/100g for the four clay adsorbents, respectively. Adsorption increased with pH till Fe(III) became insoluble at pH > 4.0. The second-order kinetics (k2 = 4.710 2 to 7.410 2 g/mg/min) gave a better description of the kinetic data. The Langmuir monolayer capacity of the clay adsorbents was from 11.2 to 30.0 mg/g. The adsorption was exothermic with Ho in the range of 27.6 to 42.2 kJ/mol accompanied by decrease in entropy (So = 86.6 to 131.8 J/mol/K) and decrease in Gibbs free energy. The results showed that the kaolinite, montmorillonite and their acid activated forms could be used as adsorbents for separation of Fe(III) from aqueous solution. Acid activation enhanced the adsorption capacity as compared to the untreated clay minerals.
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A study was carried out by Sprynskyy et al. (2006) on the adsorption of heavy metals (Ni(II), Cu(II), Pb(II), and Cd(II)) from single- and multi-component aqueous solutions by raw and pretreated clinoptilolite. The adsorption had ion-exchange nature and consisted of three stages, i.e., the adsorption on the surface of microcrystals, the inversion stage, and the moderate adsorption in the interior of the microcrystals. The finer clinoptilolite fractions adsorbed higher amounts of the metals due to relative enrichment by the zeolite proper and higher cleavage. The slight difference between adsorption capacity of the clinoptilolite toward lead, copper, and cadmium from single- and multi-component solutions testified to individual adsorption centers of the zeolite for each metal. The decrease of nickel adsorption from multicomponent solutions was probably caused by the closeness of its adsorption forms to the other metals and by competition. The maximum sorption capacity towards Cd(II) was determined as 4.22 mg/g at an initial concentration of 80 mg/L and towards Pb(II), Cu(II), and Ni(II) as 27.7, 25.76, and 13.03 mg/g, respectively, at 800 mg/L of initial concentration. The adsorption results fitted well to the Langmuir model. Vermiculite, a 2:1 clay mineral, was applied as adsorbent for removal of cadmium, zinc, manganese, and chromium from aqueous solutions by Fonseca et al. (2006). All isotherms observed were L-type of the Gilles classification, except zinc (S-type). The adsorbent showed good adsorption potential for these cations. The experimental data were analyzed by the Langmuir isotherm model showing reasonable adjustment. The quantity of adsorbed cations was 0.50, 0.52, 0.60, and 0.48 mmol/ g of Cd(II), Mn(II), Zn(II), and Cr(II), respectively. Manohar et al. (2005) investigated the possibility of using a natural bentonite clay as a precursor to produce aluminum-pillared clay (Al-PILC) for the removal of vanadium(IV) from aqueous solutions. Batch experiments were carried out as a function of solution pH, contact time, vanadium(IV) concentration, adsorbent dose, ionic strength, and temperature. The maximum adsorption capacity was observed in the pH range of 4.56.0. The maximum adsorption of 99.8 and 88.5% took place at pH 5.0 from an initial concentration of 5 and 10 mg/L, respectively. It was shown that the adsorption of vanadium(IV) could be fitted to the intra-particle mass-transfer model. The temperature dependence indicated the endothermic nature of adsorption. The percentage removal of vanadium(IV) decreased with increasing ionic strength. The Freundlich isotherm was found to well represent the measured adsorption data. Kaolin (bright white lumps) from Ubulu-Ukwu in Nigeria was modified with 200 g/mL of phosphate and sulphate anions to give phosphate- and sulfate-modified adsorbents, respectively and the adsorption of four metal ions (Pb(II), Cd(II), Zn(II), and Cu(II)) was studied as a function of metal ions concentration by Adebowale et al. (2005). The metal ions showed stronger affinity for the phosphate-modified adsorbent with Pb(II), Cu(II), Zn(II), and Cd(II) giving an average of 93.28%, 80.94%, 68.99%, and 61.44% uptake capacity, respectively. The order of preference for the various adsorbents shown by the metal ions was as follows: Pb(II) > Cu(II) > Zn(II) > Cd(II). Desorption studies showed that the phosphatemodified adsorbent had the highest affinity for the metal ions, followed by the sulfatemodified clay, while the unmodified clay had the least affinity. The experimental data were fitted by both the Langmuir and Freundlich models. In a study, Srivastava et al. (2005) investigated the adsorption of Cd(II), Cu(II), Pb(II), and Zn(II) onto kaolinite in single- and multi-element systems as a function of pH and concentration, in a background solution of 0.01 M NaNO3. The pH was varied from 3.5 to 10.0 with total metal concentration of 133.3 M in the single-element system and 33.3 M
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each of Cd(II), Cu(II), Pb(II), and Zn(II) in the multi-element system. The value of pH50 (the pH at which 50% adsorption occurs) was found to follow the sequence Cu < Zn < Pb < Cd in single-element systems, but Pb < Cu < Zn < Cd in the multi-element system. Adsorption isotherms at pH 6.0 in the multi-element systems showed that there was competition among various metals for adsorption sites on kaolinite. The adsorption and potentiometric titrations data for various kaolinite-metal systems were modeled using an extended constantcapacitance surface complexation model that assumed an ion-exchange process below pH 7.0 and the formation of inner-sphere surface complexes at higher pH. Inner-sphere complexation was more dominant for the Cu(II) and Pb(II) systems. The adsorption of the heavy metals (Cd(II), Cu(II), Mn(II), Pb(II), and Zn(II)) from aqueous solutions by a natural Moroccan stevensite was studied by Benhammou et al. (2005a). The surface area of stevensite was 134 m2/g and the cation exchange capacity (CEC) was 76.5 meq/100 g. Adsorption tests of Cd(II), Cu(II), Mn(II), Pb(II), and Zn(II) in batch reactors were carried out at ambient temperature and at constant pH. The increasing order of the adsorption rates followed the sequence: Mn(II) > Pb(II) > Zn(II) > Cu(II) > Cd(II). The maximal adsorption capacities at pH 4.0 determined from the DR and Langmuir models vary in the following order: Cu(II) > Mn(II) > Cd(II) > Zn(II) > Pb(II). The equilibrium data fitted well with the three-parameter RedlichPeterson model. The values of mean energy of adsorption show mainly an ion-exchange mechanism. The objective of a study by Benhammou et al. (2005b) was to investigate the adsorption of the heavy metals Hg(II) and Cr(VI), from aqueous solutions, onto Moroccan stevensite. In order to improve the adsorption capacity of stevensite for Cr(VI), a preparation of stevensite was carried out. It consisted of saturating the stevensite in ferrous iron Fe(II) and reducing the total Fe by Na2S2O4. Then, the adsorption experiments were studied in batch reactors at 25 C. The influence of the pH solution on the Cr(VI) and Hg(II) adsorption was studied in the pH range of 1.5-7.0. The optimum pH for the Cr(VI) adsorption was in the pH range of 2.0-5.0, and for Hg(II) above 4.0. The adsorption kinetics was tested by a pseudo-secondorder model. The adsorption rate of Hg(II) was 54.35 mmol/kg-min and that of Cr(VI) was 7.21 mmol/kg-min. The adsorption isotherms were described by the Dubinin-Radushkevich model. The maximal adsorption capacity for Cr(VI) increased from 13.7 (raw stevensite) to 48.86 mmol/kg (modified stevensite) and for Hg(II) it decreased from 205.8 to 166.9 mmol/kg. A study was carried out by Sarioglu et al. (2005) to examine the removal of copper from an aqueous solution by phosphate rock (PR). The optimum conditions for adsorption were evaluated by changing various parameters, viz., effects of pH, adsorbent concentration, initial metal concentration and contact time. The study showed that copper removal from an aqueous solution increased with increasing pH and adsorbent concentration (up to 5 g/L) and decreased with increasing initial copper concentration, and the equilibrium (contact) time was 40 min. The adsorption capacity of PR was determined as 0.17 mmol/g by fitting the experimental results to Langmuir isotherm. The potential of using activated phosphate as a new adsorbent for the removal of Pb from aqueous solutions was investigated by Mouflih et al. (2005). The kinetics of lead adsorption and the adsorption process were compared for natural phosphate (NP) and activated phosphate (AP). The results indicate that equilibrium was established in about 1 h for NP and 3 h for AP. The effect of the pH was examined in the range of 26 and the maximum removal obtained was between 2 and 3 for NP and between 3 and 4 for AP. The maximum adsorption
35
capacities at 25 C were 155.04 and 115.34 mg/g for AP and NP, respectively. The thermodynamic parameter showed that adsorption of lead on NP and AP was an endothermic process. These results showed that AP was a good adsorbent for heavy metals from aqueous solutions and could be used as a purifier for water and wastewater. Kinetic and equilibrium adsorption experiments on removal of Zn(II) from aqueous solutions by scoria (a vesicular pyroclastic rock with basaltic composition) from Jeju Island, Korea, in order to examine its potential use as an efficient sorbent, was conducted by Kwon et al. (2005). The batch-type kinetic sorption tests under variable conditions indicated that the percentage of Zn(II) removal by scoria increased with decreasing initial Zn(II) concentration, particle size, and sorbate/sorbent ratio. However, the adsorption capacity decreased with the decrease of initial Zn(II) concentration and sorbate/sorbent ratio. Equilibrium adsorption tests showed that Jeju scoria had a larger capacity and affinity for Zn(II) sorption than commercial powdered activated carbon (PAC), at initial Zn(II) concentrations of more than 10 mM, the adsorption capacity of Jeju scoria was about 1.5 times higher than that of PAC. The acquired adsorption data better fitted to the Langmuir isotherm. The adsorption behavior was mainly controlled by cation exchange and typically displayed characteristics of cation sorption. The Zn(II) removal capacity decreased when solution pH decreased because of the competition with hydrogen ions for adsorption sites, while the Zn(II) removal capacity increased under higher pH conditions, which was likely due to hydroxide precipitation. At an initial Zn(II) concentration of 5.0 mM, the removal increased from 70% to 96% with the increase of initial pH from 3.0 to 7.0. The kinetics of sorption of Cu(II) on a Saudi clay mineral (bentonite) was investigated by Al-Qunaibit et al. (2005) at 20C using different weights of the clay (0.5, 1.0, 1.5, and 2 g). Each weight represented a certain sample size. The order of the process appeared to be 1 with respect to Cu(II), and 1 with respect to the clay surface area. The adsorption rate was found to depend on internal diffusion, which produced certain decrease in the specific rate of sorption as a function of time. Adsorption characteristics were described using two-site Langmuir isotherm. The desorption experiments proved that Cu(II) ions were chemisorbed on the bentonite surface. The maximum adsorption obtained was 909 mg Cu(II)/g clay. Calcined phosphate (CP) was evaluated as a new adsorbent for removal of heavy metals from aqueous solution by Aklil et al. (2004). Removal of Pb(II), Cu(II), and Zn(II) on the CP was investigated in batch experiments. The influence of pH was studied and the adsorption capacities obtained at pH 5 were 85.6, 29.8, and 20.6 mg/g for Pb(II), Cu(II) and Zn(II), respectively. The adsorption between phosphate rock (PR) and metals (Pb, Cu, and Zn) was studied by Cao et al. (2004). Phosphate rock had the highest affinity for Pb, followed by Cu and Zn, with sorption capacities of 138, 114, and 83.2 mmol/kg PR, respectively. In the PbCuZn ternary system, competitive metal sorption occurred with adsorption capacity reduction of 15.2%, 48.3%, and 75.6% for Pb, Cu, and Zn, respectively, compared to the mono-metal systems. A fractional factorial design showed the interfering effects in the order of Pb > Cu > Zn. The capacity of sepiolite for the removal of lead ions from aqueous solution was investigated under different experimental conditions by Bekta et al. (2004). The Langmuir and Freundlich equations were applied to fit the data. The constants and correlation coefficients of these isotherm models at different conditions, such as pH, temperature and particle size were calculated and compared. The equilibrium process was well described by the Langmuir isotherm model and the maximum sorption capacity was found to be 93.4 mg/g
36
for the optimal experimental conditions. The best correlation coefficients were obtained using the pseudo-second-order kinetic model. Removal of heavy metals Mn(II), Co(II), Ni(II), and Cu(II) from aqueous solutions was studied using a raw kaolinite by Yavuz et al. (2003). The adsorption of these metals on kaolinite conformed to linear form of Langmuir adsorption equation. Langmuir adsorption capacity for each metal was found as 0.446 mg/g (Mn), 0.919 mg/g (Co), 1.669 mg/g (Ni), 10.787 mg/g (Cu) at 25C. The thermodynamic parameters showed that the adsorption of these heavy metals on kaolinite were an endothermic process. Dho and Lee (2003) conducted a combined adsorption-sequential extraction analysis by which five phases (i.e., exchangeable, carbonate, Mn-Oxide, organic, and Fe-Oxide phases) of adsorbed heavy metals were analyzed, to investigate temperature effects on single and competitive adsorptions of Zn(II) and Cu(II) onto natural clays. In the case of single adsorption of Zn, the exchangeable phase adsorption decreased from 65 to 40%, but the carbonate phase adsorption increased from 30 to 40%, with an increase in temperature from 15 to 55C. However, in its competitive adsorption with Cu, Zn was mostly present in the exchangeable phase (over 90%), and with an increase in temperature, the exchangeable phase adsorption decreased only 10%. In the case of Cu, over 50% among the total amount of adsorption was present in the carbonate phase in both cases of single and competitive adsorptions. The carbonate phase adsorption of Cu increased from 56 to 61% and from 60 to 66% in single and competitive adsorptions, respectively, with a temperature increase. These results showed that in the case of Zn, the major mechanism of retention in natural clay soils could be exchangeable phase adsorption, especially in the case of competitive adsorption with Cu. However, in the case of Cu, the major mechanism could be carbonate phase adsorption. It was observed that the adsorption of Zn and Cu onto natural clays was endothermic, which indicated that the adsorption equilibrium constants and capacities increase with a temperature increase, with the exception of exchangeable phase adsorption. The adsorption behavior of sepiolite was studied with respect to cadmium and zinc in order to consider its application to remediate soils polluted with these metals by lvarezAyuso and Garca-Snchez (2003a). The Langmuir model was found to describe the sorption processes well, offering maximum sorption capacities of 17.1 and 8.13 mg/g for cadmium and zinc, respectively, at pH 6. The sorption capacities were pH dependent, undergoing a decrease with H+ concentration increase. The column studies also showed a high reduction in the leaching of cadmium and zinc (69 and 52%, respectively) when a sepiolite dose of 4% was applied. The adsorption behavior of palygorskite was studied with respect to lead, copper, zinc and cadmium in order to consider its application to remediate soils polluted with these metals by lvarez-Ayuso and Garca-Snchez (2003b). The Langmuir model was found to describe the adsorption processes well offering maximum sorption values of 37.2 mg/g for lead, 17.4 mg/g for copper, 7.11 mg/g for zinc and 5.83 mg/g for cadmium at pH 56. The column studies also showed a high reduction in the metal leaching (50% for lead, 59% for copper, 52% for zinc and 66% for cadmium) when a palygorskite dose of 4% was applied. The adsorption characteristics of heavy metals such as Cd(II), Cr(III), Cu(II), Ni(II), Pb(II), and Zn(II) ions by kaolin (kaolinite) and ballclay (illite) from Thailand were studied by Chantawong et al. (2003). It was found that, except Ni, metal adsorption increased with increased pH of the solutions and their adsorption followed both Langmuir and Freundlich isotherms. Adsorption of metals in the mixture solutions by kaolin was: Cr > Zn > Cu Cd
37
Ni > Pb, and for ballclay was: Cr > Zn > Cu > Cd Pb > Ni. The adsorption of metals was endothermic, with the exception of Cd, Pb and Zn for kaolin, Cu and Zn for ballclay. The presence of Cr(III) induced the greatest reduction of metal adsorption onto kaolin, as did the presence of Cu(II) for ballclay. A bentonite and an expanded perlite (Morocco) were used for the removal of trivalent chromium from aqueous solutions by Chakir et al. (2002). The kinetic study showed that the uptake of Cr(III) by bentonite was very rapid as compared to expanded perlite. For both the adsorbents the adsorption capacity increased with increasing pH of the suspensions. Results showed that bentonite was more effective in removing trivalent chromium (96%) from aqueous solution than expanded perlite (40%). Surface complexation played an important role in the sorption of Cr(III) species on expanded perlite. In the case of bentonite, cationexchange was the predominate mechanism for adsorption of trivalent chromium ions. A thiol-functionalized layered magnesium phyllosilicate material (Mg-MTMS) was investigated as a high-capacity adsorbent for heavy metal ions by Lagadic et al. (2001). Structural characterization by powder X-ray diffraction, infrared spectroscopy, NMR spectroscopy and elemental analyses confirmed the smectite-type structure. Mg-MTMS was found to be highly effective for the adsorption of Hg(II), Pb(II), and Cd(II) ions, exhibiting extraordinary metal ion uptake capacities of 603, 365, and 210 mg of metal/g of adsorbent, respectively. The high effectiveness of Mg-MTMS for the capture of metal ions is attributed to both high concentration of complexing thiol groups (6.4 mmol of SH/g of Mg-MTMS) and expansion capability of the framework, which facilitates the accessibility of the binding sites. De-oiled spent bleaching clay was activated either by acid treatment followed by heat activation or by heat activation alone at temperatures between 200 and 800C by Seng et al. (2001). The surface area of the heat-activated clay attained a maximal value of 120 m2/g at temperatures between 400 and 500C, while the acid-heat-treated clay attained maximal surface area of 140 m2/g. The adsorption capacities of Cr(VI) for both series studied increased as the activation temperature increased until 300C and decreased again at higher temperatures. At lower pH, more than 95% of the Cr(VI) was absorbed in a solution with initial concentration of 1 mg/L per gram of adsorbent activated at 300C. The adsorption patterns followed Freudlich isotherm. The amount of Ni(II) adsorbed increased with the pH of the solution for all samples studied. The maximal adsorption capacities of the adsorbents in solution containing initial Ni(II) concentration of 5 mg/L per 0.5 g of adsorbent and at pH 6 were found to be 44 and 42%, respectively, for the acid-treated sample activated at 500C and for the nonacid-treated sample activated at 700C. The use of an adsorbent produced by the chemical treatment of locally available clay for the removal of some metals from waste water was investigated by Vengris et al. (2001). The modification of the natural clay was performed by treatment with hydrochloric acid and subsequent neutralization of the resultant solution by sodium hydroxide. The adsorption amounts of iron, aluminium and magnesium compounds were increased with the modified sorbent. Acidic treatment led to the decomposition of the montmorillonite structure. Adsorption studies were carried out by both batch and column methods. The uptake capacity of the modified clay for nickel, copper and zinc significantly increased. Batch and column sorption methods enabled the removal of nickel, copper and zinc ions till the permissible sewerage discharge concentration. The sorption process was reflected by Langmuir-type isotherm.
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John U. Kennedy Oubagaranadin and Z.V.P. Murthy Table 3.2. Clay adsorbents for heavy metals from solutions
Type of clay Adsorbate metal ion Ni(II) Adsorption capacity, Reference qm (mg/g) 1.91 Vieira et al., 2010 117.5 Ghorbel-Abid et al., 2009
Calcined Bofe bentonite clay Smectite with a small Cr(III) proportion of kaolinite (From Jebel Chakir, Tunisia, North Africa). Tabuk and Khaiber (From Pb(II) Saudi Arabia) Kaolin Pb(II) Petra clay Cd(II), Pb(II) Montmorillonite Illite clay Pb(II) White, yellow and red sands Cr(III) from the United Arab Emirates Raw and acid-activated Cu(II) bentonite Modified Unye bentonite Cu(II) Tunisian smectite-rich clay Pb(II) Spent activated clay Cr(VI) Kaolinite and Fe(III), Co(II), montmorillonite Ni(II) Bentonite Pb(II) Natural clinoptilolite Zn(II) Diatomite Cr(III) Laboratory grade Fullers Hg(II) earth Turkish kaolinite Pb(II) Activated palygorskite Cu(II) Natural palygorskite Pb(II) Clinoptilolite Pb(II) Montmorillonite Cd(II), Co(II), Cu(II), Ni(II), Pb(II) Palygorskite Cd(II) Natural and activated phosphate Saudi bentonite Calcined phosphate Sepiolite Raw kaolinite Sepiolite Palygorskite Pb(II) Cu(II) Pb(II), Cu(II), Zn(II) Pb(II) Mn(II), Co(II), Ni(II), Cu(II) Cd(II), Zn(II) Pb(II), Cu(II), Zn(II), Cd(II)
30, 10 4.2 74.07, 83.33 Raw: 52 Acid activated: 78 62.5, 9.8, 2.38
Al-Jlil and Alsewailem, 2009 Jiang et al., 2009 Baker, 2009 Oubagaranadin and Murthy, 2009c and 2010 Khamis et al., 2009
42.41, 32.17 105.38 25 1.422 10.4 to 11.2 28.4 to 28.9 78.82 21.2 26.5 1.15 31.75 32.24 104.28 80.93 32.7, 28.6, 31.8, 28.4, 33.0 4.54 115.34, 155.04 909 85.6, 29.8, 20.6 93.4 0.446, 0.919, 1.669, 10.787 17.1, 8.13
Eren and Afsin, 2008 Eren, 2008 Chaari et al., 2008 Weng et al., 2008 Bhattacharyya and Gupta, 2008 Zhu et al., 2008 oruh, 2008 Gr et al., 2008 Oubagaranadin et al., 2007 Sari et al., 2007 Chen et al., 2007 Chen and Wang, 2007 Gnay et al., 2007 Bhattacharyya and Gupta, 2007 lvarez-Ayuso and GarcaSnchez, 2007 Mouflih et al., 2005 Al-Qunaibit et al., 2005 Aklil et al., 2004 Bekta et al., 2004 Yavuz et al., 2003
lvarez-Ayuso and GarcaSnchez, 2003 37.2, 17.4, 7.11, 5.83lvarez-Ayuso and GarcaSnchez, 2003b
Rashed (2001) reported suitable conditions for the use of naturally occurring minerals (talc, chalcopyrite and barite) as adsorbents for the removal of lead ions. The adsorption of lead ions from solutions containing different initial lead concentrations (50, 100, 200, 400, 600, 800 and 1000 mg/L Pb as lead nitrate) using different size fractions (<63m, 63150m) of talc, chalcopyrite and barite at different pH (3, 5, 7 and 9) and different adsorption times (24, 48, 72 and 96 h) was examined. The results revealed that the chalcopyrite fraction
39
between 63150m showed the highest adsorption capacity. It was concluded that the equilibrium time of adsorption was 72 h at optimum pH between 79. Amorphous derivatives of kaolinites prepared by thermal modification followed by acid activation which improved the exchangeability of kaolinites were studied by Suraj et al. (1998). Adsorption of cadmium and copper on these modified kaolinites was studied as a function of equilibration time and temperature and it was found that the initial 1 h was sufficient to exchange most of the metal ions. Two kaolinitic clay samples obtained from Thonnakkal, south Kerala (TK) and Madai, north Kerala (MK) were modified to study their exchange behaviour. The exchangeability was found to decrease with an increase in the calcination temperature (to 600C) of kaolinite samples. An improved exchange kinetics applies for their modified (calcined and acid activated) counterparts. The amount of metal ions adsorbed showed a direct correlation with the surface area and cation exchange capacity (CEC) values. The uptake of Cd and Cu at 30, 40, 50 and 60C showed similar kinetics with maximum uptake at 40C for both Cd and Cu. Chen et al. (1997) investigated the adsorption and desorption of dissolved lead, cadmium and zinc from aqueous solutions using a North Carolina mineral apatite (contaminated soil). Aqueous solutions of Pb, Cd, and Zn were treated with the apatite, followed by desorption experiments under a wide pH conditions ranging from 3 to 12. The adsorption results showed that the apatite was very effective in retaining Pb and was moderately effective for Cd and Zn at pH 45. Approximately 100% of the Pb was removed from solutions, representing a capacity of 151 mg of Pb/g of apatite, while 49% of Cd and 29% of Zn added were attenuated, with removal capacities of 73 and 41 mg/g, respectively. The apatite was also effective in removing dissolved Pb, Cd, and Zn leached from the contaminated soil using pH 312 solutions by 62.399.9, 2097.9, and 28.698.7%, respectively. A summary of different clays and their adsorption capacities for different heavy metal ions is summarized in Table 3.2.
CONCLUSIONS AND RECOMMENDATIONS

A wide range of low-cost adsorbents has been studied worldwide for heavy metal removal. It is evident from our literature survey on adsorbents for heavy metals that inexpensive and locally available materials could be used instead of commercial activated carbon. In spite of the scarcity of consistent cost information, the widespread uses of low-cost adsorbents in industries for wastewater treatment applications today are strongly recommended due to their local availability, technical feasibility, engineering applicability, and cost effectiveness. Some low-cost adsorbents such as carbons made from bio-materials and clays have demonstrated outstanding removal capabilities for heavy metals. These adsorbents are efficient and can be effectively used for inorganic effluent treatment containing metal ions. If the alternative adsorbents are found highly efficient for heavy metals removal, not only the industries, but the living organisms and the surrounding environment will also be benefited from the potential toxicity due to heavy metals. Thus, the use of lowcost adsorbents may contribute to the sustainability of the surrounding environment. Undoubtedly low-cost adsorbents offer a lot of promising benefits for commercial purpose in the future.
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NOMENCLATURE
aRP: B: b: C0: Ce: Ct: D: E: F: h: I: J: k K: k1: k2: Kc kEL: KEL: KF: kF: kfd: KFS: KH : kip: KL: kr: KRP: Krp: KS: KT : KTh: L: M: m: n: N: P1: P2: P3: P4: P5: P6: qe: qm: qt: R: RL: t: T: Constant in Redlich-Peterson isotherm model (L/mg1/) Temkin adsorption model contant (RT/-H) (mg/g) Henry isotherm model (with intercept) constant (mg/g) Initial adsorbate concentration in solution (mg/L) Equilibrium (residual) concentration of adsorbate in solution (mg/L) Adsorbate concentration in solution at time t (mg/L) Dubinin-Radushkevich adsorption model constant (mol2/J2) Mean free energy (J/mol) Fractional attainment of equilibrium (qt/qe) Initial adsorption rate (mg/g-min) Intercept (mg/g) 4-p Fritz Schluender isotherm model constant (L/g) B-E-T isotherm model constant 4-p Fritz-Schluender isotherm model exponent Pseudo-first-order (Lagergren) adsorption rate constant (min-1) Pseudo-second-order (Ho) adsorption rate constant (g/mg-min) Competitive Langmuir isotherm model constant Extended Langmuir isotherm model constant Extended Langmuir isotherm model constant (L/mg) Freundlich isotherm model constant (L/g) Fractional power kinetic model constant (mg/g) Film diffusion rate constant (min-1) 3-p Fritz Schluender isotherm model constant (L/mg) Henry isotherm model constant (L/g) Intraparticle diffusion constant (mg/g-min0.5) Langmuir adsorption isotherm model constant (L/mg) Rate constant in Ritchies equation (min-1) Redlich-Peterson isotherm model constant (L/g) Radke-Prausnitz isotherm model constant (L/mg) Sips isotherm model constant (L/g) Temkin isotherm model constant (L/mg) Toth isotherm model constant (L/mg) 4-p Fritz Schluender isotherm model constant (L/mg) 4-p Fritz Schluender isotherm model exponent Mass of adsorbent (g) Freundlich adsorption model exponent Sips adsorption model exponent Harkins-Jura model constant (mg/g)2 Harkins-Jura model constant Halsey model constant Halsey model constant Henderson model constant Henderson model exponent Amount of adsorbate adsorbed at equilibrium (mg/g) Maximum adsorption capacity (mg/g) Amount of adsorbate adsorbed at time t (mg/g) Universal gas constant (8.314 J/mol-K) Langmuir separation factor Time (min) Absolute temperature (K)

V: Volume of solution (L) Y: Integration constant Greek symbols : Radke-Prausnitz isotherm model exponent e: Elovich kinetic model constant (mg/g-min) : Redlich-Peterson isotherm model exponent e: Elovich kinetic model constant (g/mg) r: Ritchie kinetic model constant : Sips isotherm model exponent : Toth isotherm model exponent H: Heat of adsorption (J) H: Standard enthalpy change (J) G: Standard Gibbs energy change (J) S: Standard entropy change (J/K) : Polanyi potential (J/mol) : 3-p Fritz Schluender isotherm model exponent : Fractional power model kinetic model exponent (min-1)
41
LITERATURE CITED
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In: Langmuir Monolayers Editor: Jennifer A. Sherwin
Chapter 2
MOLECULAR ORGANIZATION OF THERMOTROPIC LIQUID CRYSTALS AND THEIR MIXTURES WITH AZO DYES IN LANGMUIR AND LANGMUIR-BLODGETT FILMS
Danuta Bauman, Anna Modliska and Krzysztof Inglot
Faculty of Technical Physics, Pozna University of Technology, Pozna, Poland
ABSTRACT
In this review article the results of the study of thermotropic liquid crystals and of binary mixtures of dichroic azo dye/liquid crystal in Langmuir and Langmuir-Blodgett films are presented. The liquid crystals of rod-like shape from various homologous series and nine azo dyes with different molecular structure as well as different values and directions of the dipole moment were chosen. It was found that the liquid crystals with the terminal isothiocyanato (NCS) group are not able to form a compressible monolayer at the water surface. Very short and very long alkyl or alkoxy chains attached to the rigid molecular core also hinder the creation of the stable film. Azo dyes cannot form the Langmuir film themselves; therefore, the liquid crystals 4-n-octyl-4'-cyanobiphenyl (8CB) and trans-4-n-octyl(4-cyanophenyl)-hexane (8PCH) were used as supporting matrices. The Langmuir films were characterized by the surface pressure-area and surface potential-area isotherms and by Brewster angle microscopy (BAM). The analysis of the isotherms and BAM images of liquid crystals indicated that the organization of the mesogenic molecules at the air-water interface is dependent on their structure and to some extent reflects their ability to form an appropriate mesophase in the bulk. For the binary azo dye/8CB mixtures the miscibility of two components as well as the organization and the packing of molecules at the water surface were determined. The absorption spectra by using natural and linearly polarized light were recorded for both Langmuir and Langmuir-Blodgett films. Information about spectral properties of ultrathin layers and ability of dye and liquid crystal molecules to form self-aggregates was obtained. The polarized absorption spectra allowed one to determine the alignment of molecules on the quartz surface.
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Danuta Bauman, Anna Modliska, and Krzysztof Inglot
1. INTRODUCTION
In the XXI century, modern electronic devices tend to increasing miniaturization. The smaller the system, the faster and more economically it can work. Unfortunately, it is more and more difficult and expensive to produce it. Difficulties arise, among other things, because of the non-applicability of classical physics laws at the atomic scale. Indeed it is expected that some progress towards miniaturization could yet be made using three-dimensional (3D) systems, but it is necessary to work more diligently on the use of single molecules or small molecular aggregates as functionalizing materials in electronic devices. Various nanotechnologies come here with the help it is searching for quick, reliable and economically efficient methods which will produce systems on the nanoscale. The LangmuirBlodgett (LB) method is considered as one of the most versatile techniques currently used worldwide to produce nanostructures. As this is a unique technique that allows us to create two-dimensional (2D) ordered layers of certain molecules or colloidal particles, the architecture of which can be manipulated with ease, there exists good opportunity to optimize the physical parameters (mainly electro-optical) of the material used. Thus, LB films are now an integral part of a knowledge area related to the molecular electronics [1]. LB film properties depend strongly on the properties of a Langmuir film; therefore, it is necessary to study comprehensively both types of films. Stable monolayers at gas-liquid (Langmuir films) and gas-solid (LB films) interfaces are generally formed by amphiphilic molecules; thus, it is not astonishing that they can be formed of lyotropic liquid crystals. Thermotropic liquid crystal molecules cannot be directly treated as amphiphilic, nevertheless many of them are able to create stable and compressible monolayers at the liquid surface, which can be next transferred at the solid substrates [2]. Already in the eighties and nineties of the last century, it was found that some thermotropic liquid crystals with rod-like shaped molecules possessing strongly polar hydrophilic head group and hydrophobic alkyl chain can form stable 2D monolayers at the air-water interface, which can be transferred onto the solid substrates [3-7]. Such monolayers are very simple systems, in which the intermolecular interactions can be investigated [1,8]. Moreover, studying Langmuir and LB films formed from liquid crystals, it is possible to obtain information about the way in which the molecules are anchored to the interface, and how these interfacial interactions affect the macroscopic molecular alignment in the bulk. Thermotropic liquid crystals can also be used, likewise fatty acids, as supporting matrices for compounds which are not able to form compressible and stable monolayers at the liquid surface themselves. They were used, e.g., to support molecules of various azo dyes, and stable Langmuir and LB films of azo dye/liquid crystal mixtures were obtained and investigated [917]. The opportunity to study the properties of azo dyes in Langmuir and LB films is very essential due to the strong tendency to formation of self-aggregates by molecules of these compounds. In films fabricated by using LB technique, molecules are in a highly ordered environment, similarly as in solid matrices, where aggregation of aromatic molecules is frequently observed [17]. Interest in molecular aggregates has grown in recent years because of intriguing optical properties of these systems, giving the possibility to use them as modern functional materials, both in the field of molecular electronics and photonics. Moreover, azo dyes are characterized by the large change of the dipole moment at * transition, and thus
Molecular Organization of Thermotropic Liquid Crystals
53
by a high value of hyperpolarizability. Therefore, incorporated into highly oriented matrices, they can find application in nonlinear optics [19]. In this review article, the results of the study of 2D layers formed of thermotropic liquid crystals and of binary mixtures of dichroic azo dye/liquid crystal at the air-water and airquartz interfaces are presented. The liquid crystals of rod-like shaped molecules from various homologous series were investigated. The aim of our study was to recognize the relationship between the structure of mesogenic molecules and stability of monolayers formed by them at interfaces. We also tried to explore to what extent only the orientational order in the nematic phase, and both the orientational and positional orders leading to the layered structure in the smectic phase, are preserved if we consider a liquid crystal layer with a thickness of the order of the molecular dimension. Next, two liquid crystals, 4-n-octyl-4-cyanobiphenyl (8CB) and trans-4-n-octyl(4-cyanophenyl)-hexane (8PCH) were chosen as supporting matrices for nine azo dyes with different molecular structure. We would like to know the effect of the structure as well as the value and the direction of the dipole moment of dye molecules on their polar ordering and the tendency to creation of self-aggregates in Langmuir and LB films. Table 1. Molecular structure of liquid crystals investigated.
Abbreviation nCB Molecular structure n for compounds investigated 49 4 10 2 14 2 12 39 58
nPCH
nOCB
nCPB
nCHBT
nBOBT
8OCFPB
8OCPFB
54
2. EXPERIMENTAL
The thermotropic rod-shaped (calamitic) liquid crystals often used as components for preparation of mixtures for liquid crystal displays (LCDs) were studied. The molecular structure of these liquid crystals is seen in Table 1. All compounds were synthesized and chromatographically purified at the Institute of Chemistry, Military University of Technology, Warsaw (Poland). The most of the liquid crystals under investigation has only nematic (N) phase between solid and isotropic phases. Some of them, as for example 8CB, 9CB, 8OCB, 9OCB, have both nematic and smectic A (SmA) phases, while some as 10CB or 12PCH show only SmA phase. The molecular structure of the dyes used is presented in Table 2. All the dyes were synthesized and chromatographically purified in Institute of Polymer Technology and Dyes at d University of Technology, Poland. A commercially available Minitrough 2 Langmuir-Blodgett system (KSV Instruments Ltd., Finland) was used for fabrication of spread monolayers on the water surface and for LB films deposition. Ultrapure water characterized by resistivity of 18.2 Mcm, used as a subphase in the trough, was destilled and passed through a Milli-Q water purification system. The liquid crystals and azo dyes were first dissolved in chloroform (Uvasol, for spectroscopy, E. Merck) at a concentration of 0.1 mM in order to obtain stock solutions and kept refrigerated. The concentration of solutions was confirmed spectroscopically. The dye/liquid crystal mixture solutions were made at a constant concentration of the liquid crystal (0.3 g/l) and appropriate amounts of the dye to obtain the molar fraction (XM) of the dye in the whole range of possible concentrations (depending on the dye solubility). The solution of the appropriate liquid crystal or dye/liquid crystal mixture was then spread drop by drop from a microlitre syringe (Hamilton, England) onto the subphase to form a monolayer and was equilibrated for about 15 minutes to allow the chloroform to evaporate. Then the floating monolayer was slowly compressed, symmetrically from both sides at a barrier motion speed of 5 mm min-1. All measurements were repeated on fresh subphases three to five times to confirm reproducibility. Standard trough cleaning procedure was adopted between measurements. A constant subphase temperature was maintained by a cooling circulator and kept constant at (210.5)oC. Additionally, some experiments for liquid crystal 8CB were made at different temperatures, up to 45oC. The most basic characterization of Langmuir films is the measurement of the surface pressure versus the average area available for one molecule isotherm (-A isotherm) [1,8]. For monolayers, is defined as the surface tension of the pure subphase minus the surface tension of the subphase-monolayer system. In our experiment the surface pressure was monitored by a platinium Wilhelmy plate balance with an accuracy of 0.1 mN/m. Additionally, the surface potential (V) of the monolayer as a function of the mean molecular area (V-A isotherm) was measured using the vibrating plate method by means of a SPOT 1 head from KSV. An accuracy of this method was 1.0 mV. V is defined as the difference in the potential between a monolayer-covered surface and a clean subphase surface [8] and can be related to an average effective dipole moment for monolayer-forming molecules by Helmholtz equation [20,21]:
55
, A r 0
Table 2. Molecular structure of dyes investigated.
(1)
Compound 1
Molecular structure
where = cos is the average vertical component of the molecular dipole moment, is the angle between the surface normal and the dipole axis, and r and 0 are the dielectric constant of the monolayer and the electric permittivity of the free space, respectively.
56
The morphology of the films at the air-water interface was monitored by means of a Brewster angle microscope (BAM). The instrument we used is based on Hoenig and Moebius setup [22] and was built in our laboratory [14]. The green (532 nm) laser beam was directed at the Brewster angle (53.1o) onto the pure water. The light reflected from the monolayer was imaged by means of a CCD camera. The BAM images obtained were recorded directly on the hard disc of PC computer both on the continuous slow film compression and expansion, simultaneously with the isotherm record. Typical reproducible images for the films at various surface pressures were saved in files using a framegraber and printed. The image features were observed with a lateral resolution of 5 m. Absorption spectra of spread monolayers on the water surface were recorded in situ by means of a spectrophotometer Varian CARY 400 equipped with fiber optic accessory supplied by Varian. The quartz fiber bundle delivered UV and Vis radiation from appropriate lamps in the spectrophotometer to the Langmuir film on a trough with a quartz window at the center. Another bundle collected the light transmitted trough the film and led it back to the spectrophotometer (detector) sample compartment. Both fiber bundles were fixed in the holder, which enabled us to position the end of the bundle vertically to the water surface on both sides of the trough. The fiber bundles were precisely focused on the film to maximalize the transmitted light level. The reference light beam of the spectrophotometer was properly attenuated to reduce a level of noise. Stability tests were done for the Langmuir films to keep the pressure constant and LB films were fabricated using electronically controlled dipping device. The substrates for LB films were polished quartz plates (35x10x1mm3) with hydrophilic surfaces. The dipper speed was ca. 5 mm/min, and the dipping stroke was 25 mm. Langmuir films were transferred onto both sides of quartz slides at the surface pressure below the collapse point, which corresponds to the stage of the formation of the compressed monolayer. The transfer ratio (TR), defined as the ratio of the actual decrease in the subphase area to the actual area on the substrate coated by the floating layer, was estimated to be between 1.0 and 1.2. The deposition of the Langmuir film of the compounds under investigation onto the quartz was successful only at raising the substrate. Repeated attempts to transfer a floating layer onto quartz slides failed. Therefore, only one dipping and one raising were made.
Figure 1. Geometry for the polarized absorption measurements.
the incident light polarized, respectively, parallel and perpendicularly to the plane of incidence. is the incidence angle. AP and AS are components of absorbance.
EP and ES
are the electric vectors of
57
The absorption spectra of LB films were recorded in the UV-Vis region by means of a spectrophotometer CARY 400, equipped with an angular sample holder. The measurements were performed in the geometry presented in Fig. 1. Both natural and linearly polarized light were used. The incident light beam was normal to the substrate surface by using natural light, whereas polarized spectra were recorded at the incidence angle = 0o, 30o and 60o. In order to obtain the polarized light, the Glan-Thomson polarizers were used.
3. RESULTS AND DISCUSSION

3.1. Langmuir and Langmuir-Blodgett Films of Liquid Crystals
3.1.1. Langmuir Film Characterization The amphiphilic character of rod-shaped liquid crystal molecules is not sufficient condition for the formation of stable and compressible monolayers at the air-water interface. From our study follows that all the members of the homologous series of 4-(trans-4-nalkylcyclohexyl)-isothiocyanato-benzene (nCHBT), and n-alkyl-4-(4-isothiocyanatophenyl)bicyclo[2,2,2]-octanes (nBOBT) give monolayers which are unable to offer any significant resistance to barrier compression. All these compounds have the terminal NCS group and it seems that already this group causes the formation of the Langmuir film impossible. The other liquid crystals under investigation have the terminal CN group. Among them only those with very short and very long alkyl or alkoxy chain could not form a stable monolayer [14,23,24]. The most comprehensive investigations were carried out for the liquid crystal 4-n-octyl4-cyanobiphenyl (8CB) in the Langmuir film and the results can be found in references [3,57,14,23-26]. Figure 2 illustrates the dependence of the surface pressure, , on the average area available for one molecule, A, while compressing and expanding the monolayer of 8CB on the water surface. The expansion isotherm shows only slight hysteresis, which means that the equilibrium conditions were obtained. Following Xue et al. [5] we can distinguish in the measured area range five regions. In the region where the mean molecular area is greater than 0.50 nm2 (region I), is near to zero and constant, which indicates the coexistence of gas and liquid phases. Between 0.48 and 0.41 nm2 (region II), the first significant increase in up to the collapse point occurs (the collapse point is recognized as the point in the -A isotherm where the ratio /A begins to decrease due to the next phase transition), indicating the formation of the completely homogeneous monolayer. However, the collapse point occurs at the mean molecular area larger by a factor of two than the theoretical molecular cross-section. This effect was explained in terms of strong repulsive interactions between the electric dipoles of the cyano groups. As a result of such interactions, the 8CB monolayer is fragile, molecules are not densely packed and are tilted to the water surface. By further compression of the 8CB film (region III), the plateau region is observed: remains constant with decreasing the A value. When A reaches about 0.11 nm2 (region IV), rises sharply. This value is too small for all the molecules to remain on the water surface and therefore it was postulated that in this A region an interdigitated bilayer on the top of the monolayer (trilayer) is created [5,6]. After further compression (region V) a second plateau appears suggesting the creation of the uniform multilayer.
58
Figure 2. Surface pressure-area isotherms of 8CB Langmuir film recorded during compression and expansion processes [23] with schematic representation of molecular alignment in various stages of the film formation.
Figure 3. Surface pressure-area isotherms of Langmuir films formed of representative liquid crystals with n = 8.
Figure 3 presents -A isotherms obtained during compression process for members with n = 8 for series investigated. With a decrease of the available area, the surface pressure rises up to the collapse point, indicating the formation of the compressed monolayer. In the case of the liquid crystals 8CB, 8PCH (trans-4-n-octyl(4-cyanophenyl)-hexane) and 8OCFPB (4-
59
cyano-3-fluorophenyl 4-n-octyloxybenzoate), behind the collapse point in the -A isotherms a broad plateau is observed, and it was found that the expansion isotherms show only a slight hysteresis with respect to the compression ones. This implies that the films created are very stable. In the case of the liquid crystals 8OCB (4-n-octyloxy-4-cyanobiphenyl), 8CPB (4-nalkyl(4-cyanophenyl)benzoate) and 8OCPFB (4-cyanophenyl 4-n-octyloxy-2-fluorobenzoate), just behind the collapse point in the -A isotherm a 'spike', i.e. a rapid fall of the surface pressure appears and a large hysteresis of expansion and compression isotherms is observed, indicating on the instability of the monolayers. This leads to the conclusion that the molecular structure of the rigid core of calamitic liquid crystals strongly influences the stability of the monolayer at the air-water interface. The difference in the -A isotherm shapes for two fluoro-compounds points out that the monolayer stability can be also affected by the position of the lateral group. The steepness of the isotherm reflects the monolayer rigidity. It is seen that it is dependent on the molecular structure of the liquid crystal too. Figures 4 and 5 show isotherms for some members of 4-n-alkyloxy-4-cyanobiphenyl (nOCB) and trans-4-n-alkyl(4-cyanophenyl)-hexane (nPCH) series. It is seen that the values of the area and the pressure at the collapse point as well as the isotherm steepness are dependent also on the hydrocarbon chain length in the molecule. The values of the collapse areas found from the isotherms suggest that in the monolayer formed of all the compounds investigated the rigid cores of the molecules must be tilted with respect to the water surface (Fig. 6).
Figure 4. Surface pressure-area isotherms of Langmuir films formed of members of nOCB series [24].
Further information about a polar molecules organization, particularly at early stages of the film compression when the surface pressure is still zero, gives the surface potential measurement. For all the liquid crystals under investigation the dependence of the surface potential, V, on the average area, A, were recorded and Figs. 7-10 present representative
60
examples of V-A isotherm (solid curve), recorded simultaneously with the -A isotherm (dotted curve curve), as well as the dipole moment-area (-A) isotherm (dashed curve), calculated on the basis of Eq. (1), for the liquid crystals 7PCH, 7OCB, 6CPB, and 8OCFPB. In Tables 1-5 the features of -A and V-A isotherms of the monolayers of all the liquid
crystals which were able to form the compressible Langmuir film are gathered. A 0 and A V 0
are the values of the area at which, respectively, and V start to increase, whereas AC, C, and VC are the values of, respectively, the area, the surface pressure, and the surface potential at the collapse point. On the basis of AC the average tilt angle between the normal to the water surface and the long molecular axis (Fig. 6) were calculated by assuming that the mean molecular area is determined only by the rigid core of the liquid crystal molecule. The values of the angle as well as of , calculated at the collapse pressure, are also given in Tables 3-7.
Figure 5. Surface-pressure-area isotherms of Langmuir films formed of members of nPCH series [23].
Figure 6. Definition of the angle between the normal to the water surface and the long molecular axis (when instead of water the surface is quartz slide, this angle is marked as ) [24].
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Table 3. Features of -A and V-A isotherms, the average angle between the long molecular axis and the normal to the water surface, , and the effective dipole moment, , of members of homologous series of nCB in Langmuir films.
Compound A 0 /nm2 AC/nm2 C/mNm-1 /deg 4CB 5CB 6CB 7CB 8CB 9CB 10CB 11CB 12CB 0.29 0.39 0.45 0.46 0.47 0.46 0.51 0.47 0.49 0.20 0.30 0.36 0.37 0.41 0.40 0.44 0.40 0.43 4.3 4.8 5.3 5.0 4.8 4.6 4.4 4.6 4.6 25 39 49 51 60 57 68 57 65
A V /nm2 VC/V 0
0.41 0.60 0.57 0.56 0.65 0.54 0.58 0.64 0.72 0.590 0.610 0.618 0.663 0.630 0.619 0.683 0.744 0.787
/D 0.31 0.48 0.58 0.65 0.67 0.65 0.79 0.94 0.90
Table 4. Features of -A and V-A isotherms, the average angle between the long molecular axis and the normal to the water surface, , and the effective dipole moment, , of members of homologous series of nPCH in Langmuir films.
Compound 4PCH 5PCH 6PCH 7PCH 8PCH 9PCH 10PCH
A 0 /nm2
AC/nm2 0.42 0.45 0.45 0.47 0.48 0.47 0.47
C/mNm-1 6.0 5.4 5.3 4.8 5.3 4.6 3.5
/deg 45 50 50 53 54 53 53
A V /nm2 0
0.95 0.90 0.65 0.86 0.71 0.73 0.72
VC/V 0.502 0.534 0.551 0.544 0.520 0.589 0.649
/D 0.56 0.64 0.67 0.68 0.73 0.73 0.81
0.61 0.59 0.58 0.61 0.57 0.54 0.53
The shapes of V-A and -A isotherms for all the liquid crystals investigated are quite similar, although the values of V and at the characteristic points of the isotherms are different for various liquid crystals. In all the cases the rise of V is observed earlier than the rise of . V first increases rather sharply up to the area which coincides with the onset area for . Next, V keeps on growing but at a slower rate and reaches the maximum at the collapse point. Behind this point V remains constant. As the measured value of V is contributed only by the first monolayer coming into contact with the water [27], this suggests that the dipole density in this layer does not change during reduction of the area. Thus, during the compression, the molecules do not assume more and more vertical alignment, but are pushed out above the first monolayer being in the contact with the water and align on its top in more or less ordered way. Therefore, the dipole moment, , calculated directly from Eq. (1) has a physical meaning only up to the collapse point and is not shown behind this point in Figs. 7-10.
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Table 5. Features of -A and V-A isotherms, the average angle between the long molecular axis and the normal to the water surface, , and the effective dipole moment, , of members of homologous series of nOCB for Langmuir films.
Compound A 0 /nm2 AC/nm2 C/mNm-1 /deg 5OCB 6OCB 7OCB 8OCB 9OCB
a
A V /nm2 VC/V 0
0.44 0.61 0.62 0.57 -a 0.611 0.643 0.683 0.675 -a
/D 0.40 0.54 0.69 0.70 -a
0.34 0.45 0.45 0.44 0.38
0.22 0.32 0.36 0.40 0.33
5.8 5.8 5.8 3.6 1.9
24 36 41 47 37
- V-A isotherm impossible to record
Table 6. Features of -A and V-A isotherms, the average angle between the long molecular axis and the normal to the water surface, , and the effective dipole moment, , of members of homologous series of nCPB in Langmuir films.
Compound A 0 /nm2 AC/nm2 C/mNm-1 /deg 6CPB 7CPB 8CPB 9CPB 0.67 0.62 0.62 0.57 0.42 0.42 0.40 0.41 7.0 7.2 6.2 7.8 46 46 43 45
A V /nm2 VC/V 0
0.88 0.77 0.85 0.83 0.624 0.624 0.710 0.740
/D 0.75 0.75 0.87 0.81
Table 7. Features of -A and V-A isotherms, the average angle between the long molecular axis and the normal to the water surface, , and the effective dipole moment, , of 8OCFPB and 8OCPFB in Langmuir films.
Compound A 0 /nm2 AC/nm2 C/mNm-1 /deg 8OCFPB 8OCPFB 0.72 0.76 0.45 0.45 7.4 8.3 40 40
A V /nm2 VC/V 0
0.91 1.02 0.551 0.577
/D 0.67 0.73
The quantity is the so-called apparent or effective dipole moment and cannot be mistaken for the dipole moment of an isolated molecule, because it is influenced by several factors, first of all by various dipole fields. According, e.g. to the Demchak-Fort model [28], is composed of three independent contributions arising from (i) orientation of water dipoles in the vicinity of the headgroup, (ii) the headgroup dipoles, and (iii) the hydrophobic part dipoles. Further, in our calculations we assumed, as it is made customary [27,29], that r = 1, although it is known that dipoles in a monolayer may be embedded in media with distinct dielectric constant values (r > 1), which depends on whether these dipoles are at the monolayer-water or monolayer-air interface [30,31]. For the liquid crystals under investigation rises together with the rise of V up to the highest value being reached at the area corresponding to the onset for the surface pressure. Such a value of remains almost constant up to the collapse point (the product VA is constant in the region between vertical lines in Figs. 7-10). The runs of -A isotherms
63
indicate that at a large area available for the liquid crystal molecules, they lie almost horizontally at the water surface. Upon compression, the molecules start to interact and at some critical area the hydrophobic parts of them lift up from the water surface causing their tilted alignment with respect to the horizontal direction. Such changes of the orientation are confirmed by the significant increase of V and at this stage. Since reaches the maximal value at the area corresponding to the surface pressure onset and then remains constant (or almost constant), we suppose that the tilt angle of the mesogenic molecules at the air-water interface is settled already at the beginning of the formation of the condensed monolayer.
Figure 7. Surface potential, V (solid curve), surface pressure, (dotted curve) and effective dipole moment, (dashed curve) as a function of the mean molecular area, A for Langmuir film of 7PCH [24]. The scale in ordinate axis for V and is the same.
Figure 8. Surface potential, V (solid curve), surface pressure, (dotted curve) and effective dipole moment, (dashed curve) as a function of the mean molecular area, A for Langmuir film of 7OCB [24]. The scale in ordinate axis for V and is the same.
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Figure 9. Surface potential, V (solid curve), surface pressure, (dotted curve) and effective dipole moment, (dashed curve) as a function of the mean molecular area, A for Langmuir film of 6CPB [24]. The scale in ordinate axis for V and is the same.
Figure 10. Surface potential, V (solid curve), surface pressure, (dotted curve) and effective dipole moment, (dashed curve) as a function of the mean molecular area, A for Langmuir film of 8OCFPB [24]. The scale in ordinate axis for V and is the same.
65
(a)
(b)
(c)
(d)
Figure 11. BAM images of 8CB Langmuir film at mean molecular areas A = 0.55 nm2 (a), 0.44 (b), 0.26 (c) and 0.09 nm2 (b). The area of images is 0.35 x 0.30 mm2.
The direct observation of the morphology of Langmuir films enables Brewster angle microscopy. BAM images of the liquid crystal 8CB recorded by us were similar to those known from the literature [6,7]. In the region of coexistence of the gas and liquid phases (region I Fig. 2) we observed condensed monolayer islands in equilibrium with a foam-like structure (Fig. 11a). As the surface pressure raised (region II), the islands joined together into a completely packed monolayer, giving a homogeneous picture (Fig. 11b). Just after the collapse point in the first plateau region (III), small brighter circular or oval domains were observed, and these grew with reduction of the film area (Fig. 11c). By further reduction of the available area, when the domains observed became sufficiently large, they deformed and coalesced. The domains of 8CB in the plateau region appeared to be of homogeneous reflectivity, meaning that they have equal thickness. At the second rise of (region IV), a homogeneous multilayer structure (trilayer: monolayer covered by bilayer) was created, and in the second plateau region (V), new domains were formed (Fig. 11d). These domains are significantly brighter than those in region III, thus they indicate the creation of the next uniform layer.
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(a)
(b)
Figure 12. BAM images of 5CB in Langmuir film at A = 0.25 (a) and 0.10 (b). The area of images is 0.35x0.30 mm2.
Similar BAM images as for 8CB were obtained for 9CB and 10CB, i.e. for these liquid crystals which have the smectic A (SmA) phase in the bulk and are able to create a stable Langmuir film. However, BAM images of the liquid crystals having only the nematic (N) phase were similar to those of 8CB only up to the collapse point. By the compression beyond this point we observed the appearance of the domains of much higher brightness surrounded by interference rings. Fig. 12 presents, as an example, BAM images of the Langmuir film formed from 5CB at two different areas in the plateau region of the surface pressure. It is seen that with the decrease of the A value the number of the domains rises. However, they are not in collision and do not join together. The higher brightness of the domains in comparison with those observed for 8CB-10CB in this region indicates that they have higher thickness. Similar BAM images behind the collapse point as for 5CB were obtained for all the nematogens giving stable monolayers (4CB, 5CB, 5OCB, 6OCB, 4PCH-8PCH, 6CPB, 7CPB, 8OCFPB). For 7CB and 7OCB, which are precursors of the smectogenic compounds in the homologous series, the domains with the interference rings appeared at the end of the plateau region [14,23,24]. The detailed analysis of the rings structure suggests that the domains grow in the third dimension when A decreases. This leads to the conclusion that we have to deal here with 3D objects. Our observations seem to suggest that the shape and character of the domains observed beyond the collapse point can be related to the mesophase which the compounds possess in the bulk. This suggestion could be confirmed by BAM images of 8CB in the Langmuir film recorded at different temperatures in the whole mesophase region, i.e. both in SmA and N phases. Previously, Suresh and Bhattacharyya [32] investigated the 8CB monolayer on the water surface at different temperatures by means of a fluorescence microscope. At temperatures corresponding to the existence of SmA phase in the bulk they observed optically flat domains representing trilayer phase (first plateau region of -A isotherm) or regular multilayer phase (second plateau region). With an increase of temperature above the SmA-N transition, instead of the multilayer domains, the lens-shaped 3D domains came into existence. Similar domains were observed also at temperatures corresponding to the isotropic phase in the bulk.
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Figure 13. Surface pressure-area isotherms of 8CB in Langmuir film at T = 26oC (SmA phase), 40oC (N phase), and 45oC (isotropic phase). Marks indicate the area values at which BAM images presented in Fig. 14 were recorded.
Our BAM studies revealed similar results. Figs. 13 and 14 illustrates how -A isotherms and BAM images obtained at A 0.2 nm2 change when the temperature rises (when the available area is greater than 0.2 nm2, the run of the isotherm and BAM images recorded at temperatures corresponding to SmA and N phases do not change and resemble those obtained at 21oC and presented in Fig. 11). At T = 26oC (SmA phase) the second plateau on the isotherm is still observed (Fig. 13a) and flat domains are seen to the end of the compression
68
process (Fig. 14b), although they have now smaller sizes than at lower T. The second plateau disappears in the N phase (Fig. 13b) and the domains with the interference rings appear, but not before A < 0.1 nm2 (Fig. 14d). At the temperatures corresponding to the isotropic phase (Fig. 13c) the 3D objects are seen earlier, already at A 0.25 nm2 (Fig. 14e). Thus, following Suresh and Bhattacharyya [32] we can postulate that the domains found for 8CB monolayer for A less than 0.2 nm2 exhibit different phases depending on temperature. At temperatures below the SmA-N transition in the bulk they are flat and contain molecules aligned in an interdigitated bilayer, like in SmA phase. Next they become lens-shaped with a nematic order, and at higher temperatures the domains may contain molecules with random (isotropic) arrangement.
(a) T = 26C (SmA)
b)
(c) T = 40C (N)
d)
(e) T = 45C (Iso)
(f)
Figure 14. BAM images of 8CB in Langmuir film at A = 0.16 (a,c), 0.09 (b,d,f), and 0.26 (e) nm 2. The area of images is 0.35x0.30 mm2.
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On the basis of our BAM studies we can conclude that there exists some correlation between the molecular organization of liquid crystals at the air-water interface and their ability to create the appropriate phase in the bulk. Whereas for smectogenic molecules only optically flat domains have been found, the Langmuir films formed of nematogenic molecules have been characterized by existence of 3D lens-shaped domains just beyond the collapse point or at the end of the plateau region. When the liquid crystal has both SmA and N phases the type of the domains depends strongly on the temperature. The proposed organization of liquid crystal molecules in the flat and droplet-like domains is illustrated in Fig. 15. As for nematogenic liquid crystals the constant value of V beyond the collapse point is also observed this leads to the conclusion that in 3D domains created on the top of the homogeneous monolayer mostly antiparallel alignment of molecules occur, similarly as it takes place in the flat domains.
Figure 15. Schematic representation of molecular organization in flat (a) and lens-like (b) domains [23].
3.1.2. Electronic Absorption Spectra of Langmuir-Blodgett Films

Figure 16 shows the absorption spectra of the LB films of the liquid crystals recorded in UV region from 225 nm to 350 nm, whereas in Table 8 the position of the absorption maximum, max, and the half-bandwidth of the absorption band, , for the representative liquid crystals with n = 8 as the LB films are presented. The absorption maximum of the liquid crystals from nPCH series appears outside the spectral region used in the experiment (< 225 nm). It follows from the data presented here that both the absorption band position as well as the band width strongly depend on the molecular structure of the liquid crystals. For comparison, Table 8 gathers the values of max and for the liquid crystals dissolved in chloroform. The data given in Table 8 indicate that the wavelengths of the absorption band maximum of liquid crystals in the LB film are almost the same as in the diluted solution. However, the half-bandwidths in the LB film are significantly larger than those in chloroform. As in the chloroform the measurements were made at very low liquid crystal concentration, it is clear that the absorption spectra obtained are characteristic for monomers. Thus, the significant broadening of the spectra of the LB films could be attributed to the creation of some kinds of
70
aggregates between liquid crystal molecules, although the influence of the surface interaction cannot be excluded.
Figure 16. Absorption spectra of Langmuir film of representative liquid crystals with n = 8 [24].
Table 8. The position of the absorption maximum, max, and the half-bandwidth of the absorption band, , for liquid crystals with n = 8 dissolved in LB film and in chloroform at XM = 1.710-7.
LB film Chloroform max/nm /cm-1 max/nm /cm-1 -1 max = 1nm = 50 cm max = 1nm = 10 cm-1 281 5700 283 5180 295 6250 298 5650 249 7250 251 5000 276 5450 277 5010 264 6600 263 5900
Compound 8CB 8OCB 8CPB 8OCFPB 8OCPFB
In order to evaluate quantitatively the orientation of the liquid crystal molecules in the LB films, polarized absorption spectra were recorded and the linear dichroism (LD) was determined. We defined LD as follows [33]:
LD
AP AS , AP AS
(2)
where AP and AS are the absorbance values at the band maximum for the light polarized parallel and perpendicularly to the plane of incidence, respectively.
Molecular Organization of Thermotropic Liquid Crystals LD can be related to the angle of incidence, (Fig. 1) in the following way:
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LD
2 tan 2 , 1 cos 2 2 2 tan sin 2
(3)
where is the angle between the absorption transition dipole moment vector and the normal to the plane of the LB film. Figure 17a shows the polarized absorption spectra recorded at = 0 for 7OCB in the LB film, as an example. It can be seen that, for this liquid crystal, the value of the linear dichroism for the light incident perpendicularly to the quartz surface, LD=0, is zero. This means that the movement of the quartz slide during deposition did not disturb the homogeneity of the molecular alignment. Thus, the angle can be calculated from Eq. (3) on the basis of the spectra measured at the angle 0o. The spectra of polarized absorption components recorded at = 60o for 7OCB are presented in Fig. 14b.
Figure 17. Absorption spectra of the light polarized parallel (Ap) and perpendicularly (As) to the incidence plane for 7OCB as LB film. The incidence angle was 0o (a) and 60o (b) [24].
On the basis of the angle it is possible to obtain information about the orientation of the liquid crystals molecules with respect to the normal to the solid substrate surface. It is, however, necessary to know the angle between the absorption transition moment and the long axis of the liquid crystal molecule. To a first approximation, we can assume that for the liquid crystals under investigation this angle is equal to zero. Thus, the angles reflect directly the orientation of the mesogenic molecules in the LB films and for the liquid crystals with n = 8 they are collected in Table 9. It is seen that for all the liquid crystals the angle has similar value, equal to about 60o. This means that the liquid crystal molecules are significantly tilted towards the quartz slide surface, like to the water surface in the Langmuir films. Comparing
72
the values of and (Tables 3-7) it is seen that the rigid cores of the rod-shaped liquid crystal molecules are significantly more tilted toward the solid surface in the LB films than toward the water in the Langmuir films. This indicates that during the transfer of the monolayer from the air-water interface onto the quartz slide a molecular rearrangement can take place. However, one should keep in the mind that the different tilt of molecules with respect to the surface can be also, at least partially, due to the surface interactions of the liquid crystal molecules with the solid substrate which are different than with the water. Table 9. Average angle between the rigid molecular core and the normal to the quartz surface, , for liquid crystals with n = 8.
Compound 8CB 8OCB 8CPB 8OCFPB 8OCPFB /deg 65 60 59 62 60
3.2. Langmuir and Langmuir-Blodgett Films of Azo Dye/Liquid Crystal Mixtures

3.2.1. Langmuir Film Characterization The azo dyes dissolved in chloroform were spread on the water surface in the Langmuir through and the monolayer compression test was performed. We found, however, that the monolayer cannot be compressed: the surface pressure remained continuously at zero and the bulk phase was observed to form. This means that none of the azo dyes under investigation can produce stable and compressible monolayer, although all they possess terminal and/or lateral polar group in the molecular structure. Therefore, in order to study the dye properties in Langmuir and LB films it was necessary to use the supporting matrix. The azo dyes were mixed with two chosen liquid crystals, namely 8CB and 8PCH, at various concentrations. It was ascertained that up to the dye molar fraction XM = 0.5 (sometimes up to 0.6), the compression of the monolayer formed of dye/liquid crystal mixture was possible and stable Langmuir films were obtained. Figures 18 and 19 present -A isotherms of the Langmuir films of the liquid crystals and of 1/8CB and 3/8PCH mixtures, as examples, whereas the characteristic values of -A isotherms for monolayers of all azo dyes mixed with the liquid crystal (dyes 5-9 were mixed only with 8CB) at XM = 0.1, 0.3 and 0.5 are resumed in Tables 10 and 11. The isotherms were recorded both during compression and expansion processes and only small differences in the isotherm runs were found. From the course of the -A isotherms for all the dye/liquid crystal mixtures it follows that the rise of begins at smaller and smaller values as XM of the dye increases. This indicates that the presence of the dye improves the packing of molecules in the Langmuir film. As a result the tilt angle decreases as compared with the tilt angle of 8CB and 8PCH molecules (see: Tables 3 and 4). Knowing the area of the dye molecule and taking into account the mixture composition we were able to estimate the average angles, av, for mixed monolayers. They are gathered in Tables 10 and 11, too. One should keep, however, in
73
the mind that such estimation is very rough, because in general the splay deformation of the Langmuir film can occur.
Figure 18. Surface pressure-mean molecular area isotherms of Langmuir films of pure 8CB (1) and of binary mixtures of dye 1 in 8CB with XM of 0.1 (2), 0.2 (3), 0.3 (4), 0.4 (5), and 0.5 (6).
Figure 19. Surface pressure-mean molecular area isotherms of Langmuir films of pure 8PCH (1) and of binary mixtures of dye 3 in 8PCH with XM of 0.1 (2), 0.2 (3), 0.3 (4), 0.4 (5), 0.5 (6), and 0.6 (7) [13].
74
The increase of the dye content in the mixture causes the better rigidity and stability of the Langmuir film, as is confirmed by the rise of the isotherm steepness and the value of C. For the most mixtures the characteristic dye concentration is XM = 0.3 or 0.4. At such content of the dye, both the tilt angle of the -A plot (with respect to the horizontal direction) and C have the highest values. Table 10. Features of -A and V-A isotherms, the average angle between the long molecular axis and the normal to the water surface, av, and the effective dipole moment, , of azo dye/8CB mixtures in Langmuir films. Nup/Ndown is the ratio of molecules with dipole moment pointing in opposite directions in the monolayer.
A0
Compound XM 8CB 1/8CB 2/8CB 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5
/nm 0.48 0.38 0.24 0.18 0.36 0.31 0.30 0.38 0.25 0.24 0.41 0.34 0.24 0.42 0.32 0.26 0.46 0.30 0.25 0.43 0.27 0.20 0.47 0.39 0.27 0.44 0.37 0.27
AC C av /nm2 /mNm-1 /deg 0.41 0.29 0.20 0.14 0.30 0.27 0.28 0.31 0.17 0.20 0.35 0.27 0.20 0.32 0.24 0.20 0.37 0.24 0.20 0.29 0.20 0.14 0.37 0.30 0.20 0.37 0.31 0.23 4.8 6.1 19.0 15.7 7.2 13.0 12.4 6.0 21.0 19.0 6.5 6.5 5.9 5.4 7.0 4.0 5.2 7.0 9.0 6.7 11.6 6.7 4.9 5.1 4.2 4.6 4.3 4.1 61 37 23 15 38 31 30 39 18 20 39 29 18 31 15 11 35 15 10 33 19 11 45 30 17 45 31 19
A0 V
/nm 0.65 0.54 0.46 0.35 0.52 0.55 0.50 0.50 0.51 0.46 0.60 0.65 0.43 0.65 0.70 0.67 0.60 0.45 0.50 0.52 0.50 0.37 0.60 0.47 0.44 0.53 0.45 0.38
VC /V 0.630 0.626 0.415 0.395 0.611 0.422 0.319
Nup/Ndown /D 0.67 0.48 0.22 0.15 0.49 0.30 0.24 0.15 0.34 0.39 0.14 0.28 0.32 0.15 0.39 0.40 0.10 0.19 0.28 0.07 0.22 0.34 0.07 0.30 0.34 0.13 0.31 0.34 0.01 0.10 0.22 0.05 0.12 0.24
3/8CB
0.584 0.48 0.341 0.15 0.251 0.13 0.595 0.580 0.568 0.618 0.524 0.352 0.624 0.461 0.388 0.607 0.411 0.465 0.640 0.671 0.627 0.636 0.638 0.600 0.55 0.42 0.30 0.61 0.40 0.23 0.61 0.29 0.23 0.52 0.27 0.23 0.70 0.57 0.40 0.64 0.54 0.37
4/8CB 5/8CB 6/8CB 7/8CB 8/8CB 9/8CB
This observation points out that the azo dye molecules are able to stiffen the molecules of liquid crystals. As the XM of the dye increases further, C value decreases indicating that the monolayer stability grows smaller. This means that for XM 0.4, the liquid crystal cannot play further the role of a host. Although the compression is still possible, in the most cases the average area per molecule is too small for even the most dense packing of molecules in a
75
monolayer, which suggests the creation of 3D domains. Comparing the results gathered in Table 10 with those in Table 11 it is seen that all the effects mentioned above depend in some extent on the kind of the liquid crystal. The influence of the molecular structure of the dyes on the behavior of the azo dye/liquid crystal mixtures in the Langmuir film is also distinctly seen. Table 11. Features of -A and V-A isotherms, the average angle between the long molecular axis and the normal to the water surface, av, and the effective dipole moment, , of azo dye/8PCH mixtures in Langmuir films. Nup/Ndown is the ratio of molecules with dipole moment pointing in opposite directions in the monolayer.
A0
Compound XM 8PCH 1/8PCH 2/8PCH 3/8PCH 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5
4/8PCH
/nm 0.58 0.54 0.35 0.23 0.58 0.41 0.30 0.54 0.46 0.36 0.51 0.47 0.32
AC /nm2 48 0.45 0.23 0.17 0.49 0.32 0.25 0.45 0.34 0.27 0.43 0.40 0.26
C av /mNm-1 /deg 5.3 6.3 16.0 15.7 6.4 7.9 11.5 6.6 6.0 5.2 5.8 5.9 5.9 54 51 23 17 42 32 24 49 33 27 46 39 24
A0 V
/nm 0.72 0.70 0.63 0.44 0.73 0.61 0.49 0.66 0.60 0.52 0.64 0.65 0.48
VC /V 0.520 0.491 0.379 0.298 0.470 0.329 0.270 0.492 0.431 0.364 0.528 0.513 0.511
Nup/Ndown /D 0.64 0.57 0.23 0.13 0.59 0.28 0.18 0.59 0.39 0.26 0.60 0.55 0.35 0.05 0.33 0.40 0.04 0.28 0.36 0.04 0.20 0.30 0.03 0.07 0.23
The changes of the isotherm run with the Langmuir film composition for the azo dye/liquid crystal mixtures indicate on the interactions between two components molecules. The kind of these interactions can be followed in Figs. 20-22, in which the excess of the average area per molecule, AE as a function of XM for the dye/liquid crystal mixtures in the Langmuir films is presented. The excess area is defined as follows:
AE A12 ( X M 1 A1 X M 2 A2 ).
(4)
A12 is here the average molecular area in the two-component film, XM1 and XM2 are the molar fractions of the components, and A1 and A2 are the single component areas taken at the same as A12. In our experiment the values of A were taken at = 3 mN/m for dyes 1-4 and 4 mN/m for dyes 5-9. If AE is equal to zero, the average area per molecule follows the additivity rule, which means that in the mixture ideal mixing or complete immiscibility occurs. Deviation from zero, either positive or negative, indicates miscibility and non-ideal behavior. In Figs. 20-22 the noticeable deviation from the additivity rule is observed. For dyes 1 mixed with both liquid crystals and dyes 5, 6 and 7 mixed with 8CB this deviation is predominantly negative meaning a contraction of the two-component films due to attractive interactions among dye and liquid crystal molecules [8,34]. In the case of dyes 2, 3 and 4 the deviation is mostly
76
negative for their mixtures with 8CB, and positive when they are mixed with 8PCH. This indicates that the cyclohexane ring present in the 8PCH molecule can cause repulsive interactions. The positive values of AE occur here also for 8/8CB and 9/8CB mixtures, indicating on repulsive intermolecular interactions [8,34]. The values of AE 0 for all the binary mixtures investigated would suggest that two components in the Langmuir films are always good miscible. However, the information from the surface phase rule [8] should be taken into account additionally. This rule states that if the components are miscible, the C value should change with the mixture composition. Meanwhile, from the dependence of the C value on the composition of the Langmuir films of the dye/liquid crystal mixtures (Table 10 and 11) follows that in the case of dyes 4, 8 and 9 mixed with 8CB and 3 and 4 mixed with 8PCH the immiscibility or at least only partial miscibility of two components can be postulated. This results from the fact that although the additivity rule is not fulfilled, C remains constant.
Figure 20. Plot of the excess of the mean molecular area per molecule, AE of dyes 1-4 mixed with 8CB versus XM of dyes at = 3 mN/m: 1 (1), 2 (2), 3 (3), and 4 (4).
Figure 21. Plot of the excess of the mean molecular area per molecule, A E of dyes 5-9 mixed with 8CB versus XM of a dye at = 4 mN/m: 5 (1), 6 (2), 7 (3), 8 (4), and 9 (5) [17].
77
Figure 22. Plot of the excess of the mean molecular area per molecule, AE of dyes 1-4 mixed with 8PCH versus XM of dyes at = 3 mN/m: 1 (1), 2 (2), 3 (3), and 4 (4).
For the Langmuir films of all the azo dye/liquid crystals mixtures under investigation the dependence of the surface potential, V, on the average area, A, were recorded and Fig. 23 illustrates the V-A isotherm (solid curve), recorded with the -A isotherm (dotted curve), and the dipole moment-area (-A) isotherm (dashed curve), calculated from Eq. (1), for 8CB and 2/8CB mixtures at various XM, as examples. In Tables 10 and 11 the features of V-A isotherms of the monolayers of all the mixtures are listed.
20 18 16 14
-1
a)
0.7 0.6 0.5
20 18 16 14
b)
0.7 0.6 0.5 0.4 0.3 0.2 0.1 0.0 0.2 0.3 0.4 0.5
2
V/V; /D
-1
/mNm
0.4 0.3 0.2 0.1 0.0 0.2 0.3 0.4
10 8 6 4 2 0 0.1
20 18 16 14
/mNm
12
12 10 8 6 4 2 0 0.1 0.6
A/nm
c)
0.5
0.6
0.7
0.7 0.6 0.5
0.7
A/nm
20 18 16 14
d)
0.7 0.6 0.5 0.4 0.3 0.2 0.1 0.0 0.2 0.3 0.4 0.5
2
/mNm
10 8 6 4 2 0 0.1 0.2 0.3 0.4 0.5

2
/mNm
12
V/V; /D
0.4 0.3 0.2 0.1 0.0 0.6 0.7
12 10 8 6 4 2 0 0.1 0.6
0.7
A/nm
A/nm
Figure 23. Surface potential, V (solid curve), surface pressure, (dotted curve), and effective dipole moment, (dashed curve) as a function of the mean molecular area, A for Langmuir film of 8CB (a) and 2/8CB mixtures at XM equal to 0.1 (b), 0.3 (c) and 0.5 (d) [15].
V/V; /D
-1
-1
V/V; /D
78
From the data presented in Fig. 23 and in Tables 10 and 11 follows that with the increase of the dye content in the mixture, although C values rises or remains constant, VC decreases. As a result, for the dye/liquid crystal mixtures is always smaller than that for the liquid crystal itself and decreases as the XM of the dye rises. This is rather unexpected result taking into account the fact that the molecules in the Langmuir films formed of the mixtures align more and more perpendicularly to the water surface with the increase of the dye content (see values in Table 8 and 9) in this case should be higher and higher. Thus, we need to assume that the dye dipoles are aligned mostly opposite to the dipoles of the liquid crystal molecules leading to the diminishing of the total effective dipole moment. The dipole moments of the liquid crystal molecules in the first monolayer at the air-water interface are directed towards the water (down) giving the effective dipole moment . Let us now estimate the number of the molecules in the dye/liquid crystal mixtures with the dipole moments aligned in the opposite direction (up). Assuming that in the monolayer at the water surface there exists Nup and Ndown molecules, the effective dipole moment ' is obtained, which is smaller than . This decrease of is due to the compensation of the resultant dipole moment by pairs of molecules, in which the dipole moments are antiparallel arranged. As the dipole moments of the polar groups CN and NO2 are similar ( 4 D), we assume for simplicity that values of the dye and liquid crystal molecules are the same. Then the ratio of Nup/Ndown can be calculated from the relation:
' ( N down 2 N up ) / N down / .
(5)
The results for various XM of the dyes mixed with 8CB or 8PCH are presented in Tables 10 and 11. Comparison of the data for given dye in two liquid crystals indicate on the different arrangement of the dye molecules in the mixtures with 8CB and 8PCH at low concentration. In the mixtures with 8PCH only some part of the dye molecules align with the dipole moment directed from the water to the air. For the mixtures of some dyes (1, 2, 3, 7) with 8CB it is necessary to assume that not only the dye molecules but also some liquid crystal molecules are directed "up". When XM rises, the fraction of Nup molecules in the mixtures with both liquid crystals becomes similar and at MF > 0.3 it is smaller than the whole number of the dye molecules. However, the alignment of the dye molecules with the dipole moment directed towards the air is further strongly preferred. We looked for the correlation between the value and the direction of the dipole moment of the given dye molecule and the diminishing of values of the monolayer after the dye addition. Dyes 1, 2, 4, 5, 7 and 8 have similar value of the dipole moment, with different directions of it. The value of the dipole moment of dyes 3, 6 and 9 is smaller because here the NO2 group ( 4 D) is replaced by Cl group ( 1.5 D). However, the values of calculated from Eq. (1) are not associated with these facts. Meanwhile, the smallest changes of V and appear for the Langmuir films of 4/8CB, 8/8CB, 9/8CB, 3/8PCH and 4/8PCH mixtures. As in these mixtures the immiscibility or at least partial miscibility of the dye and liquid crystal molecules is observed, it is reasonable to assume that in such a case, the dye molecules form domains in which the dipolar groups of molecules are directed mostly down as a result of attractive interactions with the water (the dye molecules have no distinct hydrophobic part). Thus, the dipole-dipole interaction between both kinds of molecules is
79
hindered making the creation of the antiparallel pairs consisted of the dye and liquid crystal molecules more difficult, which is confirmed by the small value of the ratio of Nup/Ndown for these mixtures. Moreover, V does not change significantly and the presence of the dye does not improve the stability of the Langmuir film (C remains constant). This is illustrated schematically in Fig. 24a. However, in the case of good miscibility, individual dye molecules can point the dipole moment towards air and form antiparallel aligned dimers with liquid crystal molecules (Fig. 24b).
Figure 24. Schematic representation of microscopic polar ordering in Langmuir film of dye/liquid crystal mixture before the collapse point in the case of immiscibility or partial miscibility (a) and good miscibility (b) of both components. Filled and open arrows represent dipole moments of molecules of liquid crystal and dye, respectively [15].
The runs of V behind the collapse point indicate that this value is either constant or drops slightly upon reduction of the available area. The decreasing of V is not bigger than 20% and is observed only to XM = 0.3 (Figs. 23b and 23c). At XM > 0.3, V stays constant, which is seen in Fig. 23d. This means that in the interdigitated bilayers (dye/8CB mixtures) or 3D domains (dye/8PCH mixtures) created on the top of the first monolayer coming into the contact with the water, the antiparallel arrangement of the dye and liquid crystal molecules dominates (Fig. 25) causing that V does not change significantly. The diminishing of V, observed for some mixtures, could solely suggest the very small excess of the molecules directed with the dipole moment towards the air. In order to obtain further information about organization of azo dye and liquid crystal molecules in the two-component monolayer at the air-water interface, the BAM images were recorded at various stages of the Langmuir film creation. In Figs. 26a and 27a are shown -A isotherms for Langmuir films of 8CB mixed with dyes 1 and 4, respectively, at XM = 0.1 with marks of the compression stages at which BAM images, presented in Figs. 26b and 27b, were recorded. There were almost no differences between the structures of pure 8CB film and those of 1/8CB film. Before the surface potential and the surface pressure start to rise (1), we observed brighter islands of molecules on the dark water surface. These islands come together to form a homogeneous film when the surface pressure is still zero but the surface potential starts to rise. Further compression makes the film completely homogeneous (2). After the collapse point small domains appear (3),
80
which grow continuously as the monolayer is further compressed. The surface pressure increases in this region (unlike to the case of the Langmuir film of 8CB), indicating the presence of the dye molecules in the monolayer, which, however, do not perturb the alignment of 8CB molecules and the liquid crystal domains can join together as the available area is limited (4). The images of the mixtures of dyes 2, 3, 5, 6 and 7 with 8CB are similar to those of 1/8CB. The behaviour of 4/8CB film is different. Already at = 0 (1) in the foamlike structure bright shimmer areas with irregular boundaries are seen, which exist also beyond the collapse point (2). The creation of the oval domains is also seen (3,4), but they do not pack together, solely their brightness increases. As the similar BAM images for dyes 8 and 9 mixed with 8CB were obtained, we suppose that this effect is connected with the immiscibility of two components in the Langmuir film. For dyes 3 and 4 mixed with 8PCH also the phase separation was observed and it was reflected in BAM images too (data not shown). However, for the Langmuir film of 1/8PCH and 2/8PCH at XM = 0.1 we obtained very similar images as for pure 8PCH [14,23]. They are shown for 2/8PCH in Fig. 28, together with -A isotherm. The objects with interference rings are seen very distinctly. The number of these objects rises when the available area on the water surface decreases.
Figure 25. Schematic representation of microscopic polar ordering in Langmuir film of dye/8CB (a) and dye/8PCH (b) mixtures beyond the collapse point. Filled and open arrows represent dipole moments of molecules of liquid crystal and dye, respectively [15].
81
(1)
(2)
(3)
(4)
Figure 26. Surface pressure-mean molecular area isotherms of 1/8CB mixture (XM = 0.1) in monolayer at the air-water interface (a) and BAM images obtained during the compression (b) at A = 0.56 nm2 (1), 0.34 nm2 (2), 0.26 nm2 (3), and 0.16 nm2 (4). The scale of the images is 0.35 x 0.30 mm2 [16].
82
(1)
(2)
(3)
(4)
Figure 27. Surface pressure-mean molecular area isotherms of 4/8CB mixture (XM = 0.1) in monolayer at the air-water interface (a) and BAM images obtained during the compression (b) at A = 0.46 nm2 (1), 0.30 nm2 (2), 0.21 nm2 (3), and 0.15 nm2 (4). The scale of the images is 0.35 x 0.30 mm2 [16].

a)
10 8 6 4 2 0
83
3 2
/mNm
-1
0.2
0.3
0.4
0.5
2
0.6
b)
A/nm
(1)
(2)
(3)
(4)
Figure 28. Surface pressure-mean molecular area isotherms of 2/8PCH mixture (XM = 0.1) in monolayer at the air-water interface (a) and BAM images obtained during the compression (b) at A = 0.64 nm2 (1), 0.43 nm2 (2), 0.24 nm2 (3), and 0.15 nm2. The scale of the images is 0.35 x 0.30 mm2 [16].
84
(1)
(2)
Figure 29. Surface pressure-mean molecular area isotherms of 2/8PCH at XM = 0.3 in monolayer at the air-water interface (a) and BAM images obtained during the compression (b) at A = 0.26 nm2 (1) and 0.15 nm2 (2). The scale of the images is 0.35 x 0.30 mm2 [15].
The contrast between BAM images recorded behind the collapse point for the dye/liquid crystal mixtures and those for the liquid crystals grows with increasing XM of the dye. Up to XM = 0.5 it is possible to observe in the Langmuir film the creation of small domains, characteristic for given liquid crystal. Upon continuous compression, the number of domains increases, as it is seen in Fig. 29 for 2/8PCH at XM = 0.3. For mixtures of dyes with 8CB the size of domains does not grow, even when two components are miscible. The background monolayer at which the domains of interdigitated bilayer are formed, starts to be rough. As the available area is further reduced, the domains are very close to one another but do not coalesce. It seems that the rough bottom monolayer prevents the formation of structures bigger in the diameter. The mobility of the bilayer created on the rough monolayer is much smaller than in the Langmuir film of 8CB, where the domains of the interdigitated bilayer coalesce easily. It means that the dye molecules are present both in the monolayer and in the upper part of the Langmuir film. This situation remains almost unchanged to the end of the compression process. When for the dye/8CB mixture the phase separation occurs, as in the case presented in Fig. 30 for 4/8CB mixture at XM = 0.3, one notes in BAM images of the mixtures at XM 0.3 small non-regular patches just after spreading the chloroform solution on the water surface. Upon reduction of the available area after the collapse point, the patches grow in size but to the end of the compression process the circular domains are further seen.
85
(1)
(2)
Figure 30. Surface pressure-mean molecular area isotherms of 4/8CB at XM = 0.3 in monolayer at the air-water interface (a) and BAM images obtained during the compression (b) at A = 0.26 nm2 (1) and 0.15 nm2 (2). The scale of the images is 0.35 x 0.30 mm2 [15].
3.2.2. Electronic Absorption Spectra of Langmuir and Langmuir-Blodgett Film
The azo dyes under investigation are characterized by high extinction coefficient (over 20 000 dm3/molcm), thus it was possible to measure the absorption of
their mixtures with the liquid crystal in the monolayers floating on the water surface (in situ). Figure 31 presents the representative long-wavelength absorption spectra of the Langmuir film of dye 1 mixed with 8CB at XM = 0.3. The spectra were recorded at various surface pressure: before compression (1), at the moment when the surface pressure start to increase (2), below the collapse point (3) and behind this point, when the surface pressure remains constant (4). It can be noticed that there is no significant change in the shape and peak position as the surface pressure is increased. In the case of low XM, the absorbance values were very small and the spectra were strongly noised. Therefore, it was difficult to visualize the maximum position. It succeeded only upon processing the spectra, thus the maxima were identified and they are given in Tables 12-14, together with the half-bandwidth values.
86
Figure 31. Long-wavelength absorption spectra in situ of Langmuir film formed of 1/8CB mixture (XM = 0.3) at = 0 (1), 2.5 (2), 6.3 (3), and 8.7 (4) mN/m [16].
Table 12. The position of absorption maximum, max, and the half-bandwidth, , of the long-wavelength absorption band of dyes 1-4 dissolved in ethanol and mixed with 8CB in Langmuir and LB films.
Ethanol XM = 1.510-7 Dye max/nm /cm-1 =1nm =10cm-1 1 2 3 4 a 580 508 3210 4530 8CB Langmuir film LB film max/nm /cm-1 max/nm /cm-1 -1 =1nm =50cm =1nm =50cm-1 612 2850 593 3350 585 3450 598 4150 588 3750 583 4550 568 4100 570 5300 562 4300 560 5550 538 4700 543 5850 -a -a 537 5400 -a -a 518 5850 -a -a -b -b 582 4150 579 5550 546 4600 598 5700 530 5900 575 6950
XM 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5
486
4720
512 b
4470
- impossible to record,
- transfer impossible
87
Table 13. The position of absorption maximum, max, and the half-bandwidth, , of the long-wavelength absorption band of dyes 1-4 dissolved in ethanol and mixed with 8PCH in Langmuir and LB films.
Ethanol 8PCH XM = 1.510-7 Langmuir film LB film Dye max/nm /cm-1 max/nm /cm-1 max/nm /cm-1 XM =1nm =10cm-1 =1nm =50cm-1 =1nm =50cm-1 0.1 602 3550 587 3350 499 4700 0.3 592 3750 590 4150 1 0.5 590 4300 583 4550 0.1 535 4050 551 4200 481 4800 2 0.3 531 4650 544 5100 0.5 530 5150 533 5900 3 536 3900 0.1 -a -a 522 4750 0.3 -a -a 524 4950 0.5 -a -a 524 5150 4 527 3650 0.1 0.3 0.5 550 550 537 5450 5850 5850 531 531 556 -a 7250 8250
- impossible to record
Table 14. The position of absorption maximum, max, and the half-bandwidth, , of the long-wavelength absorption band of dyes 5-9 dissolved in chloroform and mixed with 8CB in Langmuir and LB films.
Chloroform XM = 1.510-7 Dye max/nm /cm-1 XM =1nm =10cm-1 499 481 4700 4800 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 0.1 0.3 0.5 8CB Langmuir film LB film max/nm /cm-1 max/nm /cm-1 =1n -1 =50cm =1nm =50cm-1 m 534 5050 537 5100 524 5250 529 5150 525 5450 524 5550 520 5000 518 4950 510 5100 504 5400 505 5350 508 5450 560 4500 520 6800 571 4650 522 7250 552 5900 524 7350 535 3800 531 3900 530 4100 531 4350 546 4500 556 4650 520 3900 521 3950 530 4000 534 4100 539 4250 541 4100
5 6 7
536
3900
527
3650
512
3600
The LB films of the dye/liquid crystal mixtures were obtained after deposition of the floating monolayer onto quartz slides at the surface pressure corresponding to the value of
88
before the collapse point at the -A isotherm. The absorption spectra were recorded and the positions of the maximum and the half-bandwidths of the long-wavelength absorption band for dyes 1-9 mixed with liquid crystals are gathered in Tables 12-14. For comparison in these tables the data for dyes dissolved in ethanol (dyes 1-4) and chloroform (dyes 5-9) at XM = 1.510-7 are also given. Figure 32 shows normalized absorption spectra of the Langmuir and LB films of 1/8CB (a), 2/8PCH (b), 4/8CB (c) and 4/8PCH (d) mixtures at XM = 0.5, as examples, compared with the spectra of the appropriate dye in ethanol. The solution spectra are characteristic for azo dyes in the monomeric form. In the Langmuir and LB films, the absorption band of dyes 1-9 mixed with the liquid crystal is distinctly broadened in comparison with that in solution (for dye 1 at each concentrations two maxima or maximum with distinct shoulder in 8PCH at higher XM can be distinguished). This observation could suggest that in the monolayers at the air-water and air-solid substrate interfaces some fraction of self-aggregates by dye molecules is created. Indeed neither an additional peak, nor a shoulder, which would indicate the dimer creation in the ground state, is observed, but the dye absorbance in the Langmuir and LB films does not vary proportional to its concentration. This effect can be seen in Figs. 33 and 34. Fig. 33 displays the long-wavelength absorption spectra of 1/8PCH mixture in Langmuir (a) and LB (b) films, whereas in Fig. 34 the UV-Vis absorption spectra of the LB films of 2/8CB and 2/8PCH mixtures at various XM are presented. The band in UV region with the maximum at 280 nm is related to the absorption of 8CB with very small contribution of the dye absorption. For other dyes similar behaviour was found and it can confirm the suggestion about the occurrence of some fraction of 1-9 aggregates in monolayers already in the ground state.
Figure 32. Long-wavelength absorption spectra of Langmuir (1) and LB (2) films of 1/8CB (a), 2/8PCH (b), 4/8CB (c), and 4/8PCH (d) at XM = 0.5. Dashed curve (3) presents a spectrum of a dye dissolved in ethanol at XM = 1.510-7 [16].
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Figure 33. Long-wavelength absorption spectra of Langmuir (a) and LB (b) films formed of 1/8PCH at XM of a dye of 0.1 (1), 0.3 (2) and 0.5 (3) [16].
The creation of self-aggregates by the azo dye molecules was considered in term of the molecular exciton model, proposed by Kasha et al. [35]. From this model follows that strong interactions among molecules in the aggregate can cause the splitting of the electronic excited state, which appears as a spectral shift of the absorption band and/or its splitting, known in literature as the Davydov splitting [36]. The physical basis of this model is a state interaction theory. In the simplest case, i.e. a binary system composed of two identical molecules or chromophores (dimer), the ground state wave function is expressed by the product of wave functions of molecules u and v:
0 u 0 0 .
The Hamiltonian operator for the dimer is as follows:
(6)
H H u H H u ,
(7)
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Figure 34. Absorption spectra of LB films formed of 2/liquid crystal mixtures at XM of a dye of 0.1 (1), 0.3 (2), and 0.5 (3) for: a) 2/8CB and b) 2/8PCH mixtures [13].
where Hu i Hv are the Hamiltonian operators for individual molecules, and Huv is the term connected with the energy interaction between molecules. Solving the Schrdinger equation, one obtains the following expression for the energy of the ground state of the dimer:
E0 Eu Ev u 0 v 0 H uv u 0 v 0 d u d v .
(8)
The last term represents the van der Waals interaction energy (an energy lowering) between the ground states of molecules u and v, and Eu and Ev are the ground state energies of the isolated molecules (monomers). When each molecule in the dimer undergoes the excitation from the state 0 to the state a, their wave functions of the excited state (exciton wave functions) are represented by the products of two wave functions: of a molecule u in the excited state and a molecule v in the ground state, uav0, and of molecule v in the excited state and molecule u in the ground state, u0va. The wave function of the total system is the linear combination of these two products:
a Cua 0 ua 0 Cu 0a u 0 a .
(9)
Molecular Organization of Thermotropic Liquid Crystals The Schrdinger equation for the excited state has the following form:
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H (Cua 0 ua 0 Cu 0a u 0 a ) E (Cua 0 ua 0 Cu 0a u 0 a ).
Solving Eq. (10), the following eigen functions and eigen values are obtained:
(10)
a 21/ 2 ( ua 0 u 0 0 ),
E Eua Ev 0 ua v 0 H uv ua v 0 d u d v ua v 0 H uv u 0 va d u d v Ea .
(11)
(12)
The last term in Eq. (12) is exciton splitting term, which in the point-dipole point-dipole approximation becomes [31]:
u 0a v0a
3 Ruv
3( u 0 a R )( v 0 a R) 5 Ruv
(13)
and represents an interaction energy due to exchange of the excitation energy between molecule u and molecule v. The third term in Eq. (12) is analogous to the corresponding term in Eq. (8), and represents the van der Waals interaction between an excited molecule u and the ground state molecule v.
u 0 a and v0 a are the electric transition moment of molecules u
and v, respectively, and Ruv is the distance between centers of two molecules. If one takes the difference of the van der Waals interaction energy for the excited and ground states as D, the transition energy for dimer will be the difference between Eq. (12) and Eq. (8):
Edimer E E0 Emonomer D
(14)
This is the characteristic form of the transition energy between ground and excited states of an aggregate, resulting from the exciton model. From Eq. (14) it follows that the interaction between dipole moments of two molecules (chromophores) splits the singly excited state of the dimer into two states, called and states, according to [37], with the following eigen values and eigen functions:
E Ea , a 2 1 / 2 ( ua 0 u 0 0 ),
E Ea , a 21/ 2 ( ua 0 u 0 0 ).
(15) (16)
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The energy gap between and states, expressed by = 2, is called exciton splitting or Davydov splitting [36]. Figure 35 presents three possibilities of the mutual arrangement of transition dipole moments of two molecules: parallel, in-line, and oblique. In the case (a) the transition to the state is forbidden due to the zero value of the resultant transition moment for antiparallel dipoles arrangement. As a result the maximum of the absorption band is shifted toward shorter wavelengths with respect to the maximum of the appropriate absorption band of the monomer. In the case of in-line alignment (c), the transition to the state is forbidden, which leads to the observation of a spectral red shift for the transition in the dimer compared with that of the monomer. For the oblique alignment (b), the vector sum and difference of two dipole transition moments are always different from zero, giving in the absorption spectrum the band with two maxima.
Figure 35. Exciton band energy diagram for a molecular dimer with parallel (a), oblique (b), and in-line (c) arrangement of the transition dipole moments. The ovals correspond to the molecular profile, and the double arrows indicate the transition dipole moment direction (the transition moment direction need not be parallel to the long molecular axis).
Coplanar arrangement of the transition dipole moments in dimer leads to the exciton energy diagram shown in Fig. 36. This case corresponds to the variation of the angle between the direction of the transition moment of molecules and the line of their centers from 0o to 90o. Thus, = 0o corresponds to the case in Fig. 34c, and = 90o corresponds to the case in Fig. 34a. The energy splitting of the exciton band is now given by the formula [35]:
2 u 0a v0a
3 Ruv
(1 3 cos 2 ).
(17)
From Eq. (17) and Fig. 36 it follows that for 0o<<54.7o, the exciton band is energetically located below the monomer band causing a red shift, and created aggregates are called Jaggregates [38]. For 54.7o<<90o, the exciton band is located energetically above the monomer band causing a blue shift, and corresponding aggregates are called H-aggregates [38]. When = 54.7o, no shift in the absorption spectrum is observed, and the aggregates are then called I-aggregates [39].
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Figure 36. Exciton band energy diagram for dimer with coplanar arrangement of the transition dipole moments.
Looking at the data presented in Tables 12-14 it is seen that the absorption band of all the dyes in the monolayers becomes broader and broader as the dye content increases. However, the shift of the maximum position is different for various dyes. For dyes 1-7 studied at low XM in the Langmuir and LB films the absorption maximum position shifts towards longer wavelengths with respect to the position of the monomeric absorption band. Upon increasing the dye content, the wavelength of the maximum band shortens. Indeed significant differences in the change of the absorption band shape ( and ) at the same dye content in the liquid crystal for various dyes occur, indicating the influence of the molecular structure of the substituents on the spectral properties of the dyes under investigation, but above mentioned behavior of the maximum position for all the mixtures can be noticed. It seems, therefore, that at the change of the dye content in the monolayers at interfaces various kinds of aggregates are formed. One of the possibilities is assumption that at low XM some fraction of Jaggregates is created, whereas at higher concentrations the H-aggregates appear too. We tried to separate the long-wavelength absorption spectra for the azo dye/liquid crystal mixtures under investigation into two or three bands, corresponding to absorption of monomers, Jand/or H-dimers. We supposed that up to the dye molar fraction equal to 10-2, the dye molecules occur only in their monomeric form. Thus, the values of and for the monomer (M) absorption band were taken from the measurements of the absorption of the dyes dissolved in the appropriate liquid crystal at XM = 10-2 in the sandwich cell of 10 m in thickness [13,14,17]. Next, one or two additional maxima were assumed, which should appear at the longer (J-type dimer) and shorter (H-type dimer) wavelengths with respect to the band mximum position of the monomer. A sum of normalized Gaussians as a model function for experimentally obtained absorption band of dye/liquid crystal mixtures in the Langmuir and LB films were used.
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Figure 37 illustrates the exemplary decomposition of the absorption band into Gaussiantype components for 4/8CB mixture at XM = 0.1 (a) and 0.5 (b). The detailed analysis of the data obtained from the decomposition of the absorption spectra of the azo dye/liquid crystal mixtures in LB films is presented in refs. [13,14,17]. For all the mixtures investigated the general observation was made that at low dye content apart from monomers, only J-dimers appear, as in Fig. 37a. When the dye concentration rises, the H-dimers can be also created. In the mixtures of the most dyes with 8CB and 8PCH at XM = 0.5 the monomers, J-dimers and H-dimers in various fractions exist both in the Langmuir and LB films (Fig. 37b). It was found, however, that the mutual orientation of molecules in the aggregate as well as the amount of the appropriate kind of aggregates at given XM strongly depend on the molecular structure of the dye, and first of all on the value and the direction of the resultant dipole moment. For example, for dye 3 the best fitting for the decomposition of the absorption band of its mixture with 8PCH was obtained by assumption that at XM 0.3 only J-type and H-type dimers occur, without presence of the monomers. In the case of dyes 8 and 9 the creation of Jdimers was observed only [17]. With the rise of the dye content more and more aggregates appeared and the amount of the monomers decreased, however H-type dimers were not created. It seems that the direction of the dipole moment in the molecules of dyes 8 and 9 is not conducive to such alignment of the molecules that enables H-dimers creation.
Figure 37. Decomposition of the absorption spectrum of 4/8CB mixture at XM = 0.1 (a) and 0.5 (b) into two (a) and three (b) Gaussian-type absorption bands [13].
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In order to evaluate quantitatively the orientation of molecules of the azo dyes and the liquid crystals in the monolayer at the solid substrate, the polarized absorption spectra of the LB films were recorded. This kind of measurement was made only for dyes 1-4 mixed with two liquid crystals. Figs. 38 and 39 show examples of the polarized components of the absorption spectra of the LB films of 1/8CB and 4/8PCH mixtures, respectively. For all the mixtures investigated, at the angle of incidence = 0o (see: Fig. 1), both components were equal to each other. This means that the movement of the quartz slide during transfer did not disturb the homogeneity of the molecules arrangement. The shape of the band at 280 nm, related to the absorption of 8CB (Fig. 38), was independent of XM and . However, the AP and AS for the band attributed to the dye absorption had different shapes at various XM and for the mixtures, both with 8CB and 8PCH. As the orientation of monomers and dimers can be, generally, different, the results obtained confirm previous suggestion that in the monolayers at interfaces various kinds of aggregates can be created.
Figure 38. Absorption spectra for light polarized parallel (AP) and perpendicularly (AS) to the incidence plane for 5/8CB mixture as LB film at XM = 0.1 (a) and 0.3 (b). The incidence angle was 60o [13].
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Figure 39. Absorption spectra for light polarized parallel (AP) and perpendicularly (AS) to the incidence plane for 4/8PCH mixture as LB film at XM = 0.5. The incidence angle was 0o (a) and 60o (b) [13].
On the basis of the polarized absorption spectra the angle between the absorption transition moment and the normal to the quartz surface was calculated from Eqs. (2) and (3). Similarly as in the case of the liquid crystals (see: section 3.1.2), it was assumed that the angle between the transition moment direction and the long axis of dyes 1-4, for the longwavelength absorption band, resulting from * transition, is equal to zero. Thus, the angle gives direct information about the orientation of the dye molecules at the quartz surface. Table 15 contains the values of the angle for dyes 1-4 mixed with 8CB or 8PCH in the LB films at XM = 0.1, 0.3 and 0.5. These are the mean values of the results obtained at the angles of incidence, = 30o and 60o, for at least three independently prepared samples. The values of AP and AS were taken at the wavelengths corresponding to the monomer, J-dimer and H-dimer maximum positions, obtained from the best fitting of the Gaussian functions to the experimental spectrum [13,14]. The results obtained indicate that, in general, the angle of all the absorbing species decreases with the rise of the dye content as does the angle between the long molecular axis and the normal to the water surface in the Langmuir films
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(see data in Tables 10 and 11). However, the value of for a given dye/liquid crystal mixture is greater than that of . This difference can be due to three reasons: (i) the molecular packing can be deteriorated by the stress generated by the transfer of the monolayer from the water surface to the quartz slide, (ii) the methods used to calculate the angles and are different, and (iii) the surface interactions with the water differ from those with the solid substrate. The various angles between the long molecular axis and the normal to the surface in the Langmuir and LB films were found also previously for other dyes [40-45]. The molecular orientation in the LB films strongly depends on the structure of both dye and liquid crystal molecules. For dyes 1 and 4, having a lateral NO2 group, the greatest values for all absorbing species were observed, indicating that the presence of such group can loosen the molecular packing during the monolayer transfer. Moreover it is seen that, in general, in the mixtures with 8PCH, the molecules are more tilted with respect to the quartz surface than in the corresponding mixtures with 8CB. It is possible that repulsive interactions, observed also in the Langmuir films (see Fig. 22) play here some role. From the data in Table 15 follows that, when the dye molecules form H-type dimers, their long axes show a more vertical orientation with respect to the film surface than in the case of J-type dimers and monomers. Such an observation was made also for other compounds forming self-aggregates in films at interfaces [40]. The greatest angle was obtained for J-type dimers. This seems to be reasonable as these dimers are long and thus not very rigid. One should take also into account that the transition moment direction does not need to be directed along the J-dimer long axis. Conclusions drawn from analysis of the absorption spectra of Langmuir films formed from mixtures of azo dye/liquid crystal allow to propose a model of organization of the molecules in the monolayers at the air-solid substrate interface at low and high contents of a dye. This model is presented in Fig. 40.
Figure 40. Schematic representation of alignment of azo dye and liquid crystal molecules in LB films at low and high concentrations.
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Table 15. Values of the angle, av, between the molecular long axis and the normal to the quartz slide, in LB films of dyes 1-4 mixed with liquid crystals.
av/deg Dye/MF 1 XM = 0.1 0.3 0.5 2 XM = 0.1 0.3 0.5 3 XM = 0.1 0.3 0.5 4 XM = 0.1 0.3 0.5 M 61 62 43 43 43 41 60 43 38 40 8CB H 45 36 37 38 45 37 J 62 70 52 47 53 49 62 48 50 51 M 62 60 62 54 49 50 56 69 62 8PCH H 66 59 34 37 25 26 60 55 J 65 66 69 55 61 56 60 52 43 70 78
CONCLUSION
The thermotropic liquid crystals with rod-shaped molecules are able to create monolayers at the air-water and air-solid substrate interfaces, but this ability strongly depends on the molecular structure of the terminal groups attached to the rigid core of the molecule. The measurements of the surface pressure during compression process of the Langmuir films showed that the liquid crystals with the terminal NCS group are not able to form a compressible monolayer on the water surface. The mesogenic molecules with very short or very long alkyl or alkoxy chain also do not create Langmuir films. The shapes of the surface pressure-area and surface potential-area isotherms for the Langmuir films of the liquid crystals that make the compressible monolayer at the air-water interface are dependent on the molecular structure of the mesogen rigid core and on the chain length. Brewster angle microscope images revealed that in the case of nematogens giving stable monoayers (4CB6CB, 5OCB, 6OCB, 4PCH-8PCH, 6CPB, 7CPB), behind the collapse point 3D lens-like domains are formed. In the case of smectogens (8CB-10CB) or their precursors (7CB, 7OCB) interdigitated bilayer on the top of the monolayer is formed. Thus, the organization of the liquid crystal molecules in Langmuir films can to some extent be correlated with the type of the mesophase that they form in the bulk. The electronic absorption spectra recorded for the liquid crystals as the LB films indicated that the mesogenic molecules have a tendency to formation of self-aggregates. The polarized absorption spectra of the LB films enabled us to calculate the angle between the long molecular axis and the normal to the quartz surface. Some differences between the molecules tilt angle in the Langmuir and LB films were observed, which can be due to the rearrangement of molecules by the transfer of the floating monolayer on the quartz surface. The different tilt of molecules can be, at least partially, due to the surface interactions of the liquid crystal molecules with the solid surface that are different than with the water.
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Azo dyes cannot form the Langmuir films themselves, but when one of the thermotropic liquid crystals (here 8CB and 8PCH) was used as supporting matrix, the compression was possible and stable films of azo dye/liquid crystal mixture up to the dye content of at least XM = 0.5 were obtained. The analysis of -A isotherms of the Langmuir films revealed that properties of the monolayer on the water surface (packing density, stability and rigidity) depend strongly on the structure of both the dye and liquid crystal molecules as well as on the mixture composition. The surface potential measurements pointed out that in the spread monolayer, the molecules of the dye are aligned with the dipole moment directed mostly opposite to the dipole moment of the liquid crystal molecules, i.e., from the water to the air. The influence of the miscibility or the immiscibility of both components in the Langmuir film on the change of the surface potential and on the value of the resultant effective dipole moment was found. The electronic absorption spectra recorded for dye/liquid crystal mixtures in both the Langmuir and LB films showed very modest changes with respect to the spectra of the dyes in the diluted solution, primarily in the form of a broadening of the spectrum. The shift of the maximum position of the absorption band, different for various dyes and dependent on the dye molar fraction, was also observed. It was found that in the monolayers at interfaces J-type and H-type aggregates are created by azo dye molecules. The fraction and the type of the aggregates strongly depend on the molecular structure of the dye and can be quite different in both kinds of monolayers. Similarly, as in the case of the liquid crystals, the angle between the long molecular axis of the azo dye and the normal to the solid surface is different from the angle formed with the normal to the water surface. This implies that the molecular packing is loosened during the monolayer transfer. However, some tendency to the tilt angle decrease in H-type dimers and increase in J-type dimers with respect to this angle for monomers can be seen. Such observation indicates that only aggregates in which the transition moment is aligned at high angles (>54.7o-90o) to their line of the centre are able to improve the molecular packing in the LB film.
REFERENCES
[1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] Roberts, G. Langmuir-Blodgett Films; Plenum Press: New York, 1990. Janietz, D. In: Handbook of Surfaces and Interfaces of Materials; Nalwa, H.S.; Ed.; Academic Press: New York, 2001; Vol.1, pp. 423-446. Daniel, M.F.; Lettington, O.C.; Small, S.M. Thin Solid Films 1983, 99, 61. Sakuhara, T.; Nakahara, H.; Fukuda, K.; Thin Solid Films 1980, 159, 345. Xue, J.; Jung, C.S.; Kim, M.W. Phys.Rev.Lett. 1992, 69, 474. Friedenberg, M.C.; Fuller, G.G.; Frank, C.; Robertson, C.R. Langmuir 1994, 10, 1251. de Mul, M.N.G.; Mann, J.A. Jr. Langmuir 1994, 10, 2311. Gaines, G.L. Insoluble Monolayers at Liquid-Gas Interface; Interscience: New York, 1996. Martyski, T.; Biadasz, A.; Bauman, D. Liq.Cryst. 2002, 29, 281. Martyski, T.; Miyake, J. Z.Naturforsch. 2003, 58a, 23. Martyski, T.; Biadasz, A.; Bauman, D. Z. Naturforsch.2003, 58a, 97. Bauman, D.; Inglot, K.; Martyski, T. Mol.Cryst.Liq.Cryst. 2007, 479, 49. Inglot, K.; Kaleta, A.; Martyski, T.; Bauman, D. Dyes and Pigments 2008, 77, 303.
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Danuta Bauman, Anna Modliska, and Krzysztof Inglot Inglot, K. PhD Thesis, Pozna 2008 (in Polish). Inglot, K.; Martyski, T.; Bauman, D. Dyes and Pigments 2009, 80, 106. Inglot, K.; Martyski, T.; Bauman, D. Opto-Electronics Review 2009, 17, 120. Bauman, D.; Pciennik, A.; Inglot, K. Acta Phys.Polon.A 2009, 115, 203. Organic molecular aggregates: Electronic Excitation and Interaction Processes; Reineker, P.; Haken, H.; Wold, H.C.; Eds.; Springer: Berlin, 1983. Side-chain Liquid Crystal Polymers; Mc Ardle, C.B.; Ed.; Blackie: Glasgow, 1984. Davies, J.T.; Rideal, E.K. Interfacial Phenomena; Academic Press: New York, 1963. Myers, D. Surfaces, Interfaces and Colloids; Willey-VCH: New York, 1999. Hoenig, D.; Moebius, D. J.Phys.Chem. 1991, 95, 4590. Inglot, K.; Martyski, T.; Bauman, D. Liq.Cryst. 2006, 33, 855. Modliska, A.; Inglot, K.; Martyski, T.; Dbrowski, R.; Jadyn, J.; Bauman, D. Liq.Cryst. 2009, 36, 197. Martyski, T.; Hertmanowski, R.; Bauman, D. Liq.Cryst. 2001, 28, 437. Martyski, T.; Hertmanowski, R.; Bauman, D. Liq.Cryst. 2002, 29, 99. Schmitz, P.; Gruler, H. Europhys Lett. 1995, 29, 451. Demchak, R.J.; Fort, T.J., Jr. J.Colloid Interface Sci. 1974, 46, 191. Dynarowicz-tka, P. Adv.Colloid Interface Sci. 1993, 45, 215. Oliveira, O.N., Jr.; Taylor, D.M.; Lewis, T.J.; Salvagno, S.; Stirling, C.J.M. J.Chem.Soc. Faraday Trans. 1989, 85, 85. Taylor, D.M.; Baynes, G.F. Mat.Sci.Engng.C 1999, 8-9, 65. Suresh, K.A.; Bhattacharyya, A. Langmuir 1997, 13, 1377. NSoukpo-Kossi, Ch.N.; Sielewiesiuk, J.; Leblanc, R.M.; Bone, R.A.;. Landrum, J.T. Biochim.Biophys. 1988, 940, 255. Angelova, A.; van der Auweraer, M.; Ionov, R.; Vollhardt, D.; de Schryver, F.C., Langmuir 1995, 11, 3167. Kasha, M.; Rawls, H.R.; Ashraf El-Bayoumi, M. Pure Appl.Chem. 1965, 11, 371. Davydov, A.S. Theory of Molecular Excitons; Mc Graw-Hill Book Company, Inc.: New York, 1962. Harada, N.; Nakanishi, K. In: Circular Dichroism Spectroscopy Exciton Coupling in Organic Chemistry; University Science Books: Mill Valey, CA, 1983. Moebius, D. Adv.Mat. 1995, 7, 437. Miyata, A.; Heard, D.; Unuma, Y.; Higashigaki, Y. Thin Solid Films 1992, 210/211, 175. Kawai, T.; Umemura, J.; Takenaka, T. Langmuir 1989, 5, 1378. Johnson, E.; Aroca, R.; Nagao, Y. J.Phys.Chem. 1991, 95, 8840. Hertmanowski, R.; Chudziski, .; Martyski, T.; Stempniewicz, P.; Wolarz, E.; Bauman, D. Liq.Cryst. 2004, 31, 791. Biadasz, A.; Martyski, T.; Stolarski, R.; Bauman, D. Liq.Cryst. 2004, 31, 1639. Biadasz, A.; Martyski, T.; Stolarski, R.; Bauman, D. Liq.Cryst. 2006, 33, 307. Bielejewska, N.; Chrzumnicka, E.; Stolarski, R.; Bauman, D. Opto-Electronics Review 2010, 18,197. Reviewed by Prof. Dr. Piotr Bojarski, Institute of Experimental Physics, University of
Gdask, Poland.
Chapter 3
ATOMIC FORCE MICROSCOPY CHARACTERIZATION OF LIPID/PROTEIN NANOSTRUCTURES FORMED IN LANGMUIR-BLODGETT FILMS

Yih Horng Tan and Keith J. Stine
Department of Chemistry and Biochemistry and Center for Nanoscience, University of Missouri Saint Louis, Saint Louis, MO
ABSTRACT
Recently, much of the emphasis in biotechnology has been on producing nanosystems and nanodevices for a vast range of medical applications, including nanoelectronic biosensors, drug delivery systems, and diagnostic and imaging techniques, to name a few. This chapter reviews the potentially useful creation of proteo-lipidic nanostructures generated via the Langmuir-Blodgett (LB) film method, and their direct characterization by atomic force microscopy (AFM). Following the introductory sections, a brief overview on LB film fabrication on surfaces suitable for AFM will be presented, LB films containing proteins, lipids, and biocompatible amphiphilic molecules will be described, and how they have been studied using various AFM imaging modes. We aim to highlight recent developments that illustrate the unique capability of AFM in elucidating nanometer scale organization and the physicochemical properties of artificially engineered biological membranes through the Langmuir-Blodgett method; as it could potentially open a new pathway toward the development of self-organized nanostructures of technological significance.
INTRODUCTION
The utilization of biological molecules such as enzymes, peptides or antibodies concurrently with artificially engineered amphiphilic lipid monolayer films to elucidate the origin, composition, size and lifetime of microdomains in biological membranes has been the subject of scientific curiosity for most of the past three decades. In this respect, LangmuirBlodgett (LB) monolayers at the air/water (A/W) interface incorporating proteins are
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promising model systems that can be used to correlate the relationship between artificial structures and natural biological membranes. The imitation of natural biological membranes using proteo-lipidic LB model systems is made possible by introduction of biomolecules into the amphiphilic LB films. This approach to understanding biological activities, including biospecific recognition properties, may yield improved insight into these processes and provide the basic understanding needed for many applications in nanobiotechnology that are poised to play a major role in next-generation drug delivery systems, signal detection, biosensors and molecular labeling technology.[1-12] There have been some excellent detailed reviews of the development of biomimetic and functional proteo-lipidic nanostructures based on Langmuir-Blodgett technology over the past few years.[6-8, 11-16] For example, Godoy et al. demonstrated that the ability to control the molecular architecture of LB films was essential for preparing a biosensor with a highly organized proteo-lipidic structure able to partition a non-inhibitory monoclonal antibody in a functional orientation in order to bind an enzyme in an active and stable orientation.[15] In another paper from the same group (Girard-Egrot et al. in 2003), it was shown that the parameters which influence the kinetics of the surface film formation can affect the orientation of the immunoglobulin G, when it is inserted into a mixed glycolipid monolayer formed by spreading of proteo-glycolipid vesicles.[16] Along with the rapid progress in understanding the molecular organization of amphiphiles in LB films, and their association with proteins/enzymes, many optical microscopy techniques have been used to gain insight into the morphological texture, lattice structure, phase behavior and the molecular structure of amphiphilic monolayers.[17-18] Similarly, Brewster angle microscopy (BAM), has been used to reveal that amphiphilic monolayers can self-assemble into mesoscopic domain structures, and the introduction of synchrotron X-ray diffraction at grazing incidence (GIXD) has made possible the determination of two-dimensional lattice structures.[14, 19-20] Furthermore, examples of membrane microdomains resulting from lipid-protein, lipidlipid and protein-protein interactions were reported by using fluorescence photobleaching recovery, fluorescence digital imaging microscopy and single molecule fluorescence microscopy.[12, 21-23] Among all the available surface analysis tools and above mentioned techniques, atomic force microscopy (AFM) was also applied to investigate the amphiphilic monolayer films generated via the LB technique.[24-25] Many high molecular resolution images of single component LB monolayers and multilayers were reported recently.[26-27] For example, by preparing a LB film of a 1:1 molar mixture of arachidic acid and a partially fluorinated carboxylic acid, Gotoh et al., showed that phase separation in multicomponent LB films can be revealed using both AFM normal and lateral force imaging. In their report, the vertical resolution of the domain topography identified reached a resolution of approximately 5 and the typical lateral dimension of the round shaped domains ranged from 100 nm to 1 m in diameter. Further, the chemical makeup of each phase was able to be assigned by making reference to the frictional information acquired on the surfaces of LB films of the pure components.[28] Also in connection with biological applications, aside from visualizing objects on a surface at nanometer scale, AFM is capable of in-situ surface manipulation followed by insitu measurements, and real time data acquisition.[29-38] Niehus et al. reported the morphology and the stability of supported LB films of phospholipids such as 1,2-dipalmitoylsn-glycero-3-phosphocholine, 1,2-dimyristoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-
Atomic Force Microscopy Characterization
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sn-glycero-3-phosphocholine/1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC, DMPC, and DOPC/DPPC, respectively) layers, and demonstrated that using AFM is suitable for studying time-dependent biological and physical phenomena.[38] Other applications of AFM also extended to examining phospholipid layers supported on various solid substrates, such as mica or glass, characterizing vesicle fusion and membrane lithography via lipid backfilling methods, and AFM anodization lithography on specialized resists using amphiphilic molecules of hexadecylamine (HDA) and palmitic acid (PA).[38-44] In this review chapter, we aim to highlight the nanometer scale organization and the physicochemical properties of artificially engineered biological membranes elucidated by using AFM; in particular, high resolution imaging of biologically relevant thin films that extends from the molecular to the microscopic level. Studies using real-time AFM imaging techniques illustrating (i) the effect of lipidic LB films upon protein adsorption, (ii) the stability of proteo-lipidic LB films, and (iii) the kinetic behavior of immobilized proteins/drug complexes in a biomimetic environment via real-time AFM imaging techniques will be discussed to emphasize the information that AFM can provide. A brief discussion of a variety of recent AFM imaging techniques or developments will be presented, that might open new avenues in model and biomembrane AFM applications.
LANGMUIR-BLODGETT TECHNOLOGY
Langmuir-Blodgett films with proteo-lipidic nanostructures have been extensively used as model membrane systems in biophysics to study/investigate biocatalytic or biomimetic phenomena.[7-8, 45] The use of Langmuir-Blodgett films is possible due to the natural properties of organic molecules such as fatty acids, phospholipids, glycolipids, sterols, etc, that can orient themselves at the air/water interface. Traditionally, the molecules forming Langmuir monolayers are amphiphilic molecules that contain both hydrophilic and hydrophobic structural regions. The hydrophilic (water loving) end typically is either charged or contains a polar head group. In contrast, the hydrophobic (water hating) end typically is nonpolar and not easily dissolved in water.[46] In general, all amphiphiles are potentially ideal materials for monolayer formation. Regardless of whether they are naturally occurring or synthetic, these molecules will spread/accumulate at the air/water interface. The strength of their affinity to the air/water interface, is determined by the balance of hydrophilic and hydrophobic components in their structure, directly determines the physicochemical properties of the monolayers formed. Figure 1 illustrates Langmuir-Blodgett monolayer formation by typical lipid amphiphiles having two hydrocarbon chains and a head-group. At a controlled initial condition just after spreading (from a dilute amphiphile solution in a spreading solvent such as chloroform), the average separation distance between molecules is large and the lateral molecular attractions are usually at a minimum, as depicted in Figure 1A. The available surface area in the Langmuir-Blodgett trough is then decreased by applying compression using the moving barrier, and the average distance between molecules decreases, as depicted in Figure 1B. Continuing to increase the compression can lead to a highly organized monomolecular monolayer at the air/water interface, as depicted in Figure 1C. The formation of a highly organized monolayer upon compression depends upon the amphiphile structure, including
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variables such as the chain length. For example, the phospholipid DPPC (two C16 chains) forms an ordered liquid-condensed phase (LC) upon compression at room temperature, while the phospholipid DMPC (two C14 chains), remains in a liquid-like liquid-expanded phase (LE) even at the full extent of compression.[47] Further compression will result in monolayer collapse into the third dimension, by a variety of mechanisms, as has been recently reviewed.[48]
Wilhelmy's surface balance
A
0 mN/m
Amphiphiles Floatation device
Langmuir Blodgett Trough
Water
Compression
Compression
40 mN/m
Figure 1. Langmuir Monolayer Formation. (A) Amphiphiles not yet under any external compression by floating barrier device. (B) Some molecular interaction and initial organization of amphiphiles occurs when film compression is initiated by the floating device. (C) Well-ordered Langmuir monolayer formed, at elevated surface pressure.
SURFACE PRESSURE AND ITS RELATION TO SURFACE TENSION

As discussed in greater detail elsewhere, water molecules at the air/water interface possess an excess free energy originating from the difference in environment between the surface molecules and those in the bulk.[6, 12, 49] This interfacial free energy is accessible by measurements of the surface tension, denoted as (). When amphiphilic molecules are spread on the water surface, the formation of a Langmuir monolayer will affect, and in particular decrease, the surface tension.[6, 46] This change in surface tension occurs because when amphiphilic molecules (primarily the hydrophobic ends) are in close proximity, interaction and exertion of repulsive forces among the molecules occurs. The force exerted by the film is referred to as the surface pressure () and is equal to the reduction of the surface tension of the pure liquid subphase by the film, and is given by the following relationship
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o
where o represents the surface tension of the pure liquid subphase and represents the surface tension in the present of amphiphiles. The surface tension of pure water is 72.8 mN m1 at 20 oC and depends slightly on the addition of electrolytes.
SURFACE PRESSURE VERSUS AREA ISOTHERMS

Traditionally, the primary method used to evaluate and monitor the Langmuir film properties of a given amphiphilic molecule is by measuring surface pressure versus mean molecular area isotherms. In the isotherm measurement, essential information such as the total number of molecules spread on the water surface at a given area is known, the experiment is carried out at a constant controlled temperature, and the subphase conditions are specified (pH, electrolyte concentration, concentrations of other dissolved species). Classically, the area per molecule, which represents the mean area available to each molecule, is calculated by dividing the film area (determined by the barrier position during the compression) by the total number of molecules spread on the water surface.[8] The area per molecule, A, is typically expressed in 2/molecule or nm2/molecule. A characteristic surface pressure versus area (-A) isotherm is shown in Figure 2.
Maximum surface pressure reached before collapse mechanisms become dominant
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Surface Pressure, (mN/m)
70
Water
Collapse phase
60 50 40 30
Solid Phase (S)
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
LC/ S
Water
Liquid Condensed, (LC)

Water
LE /LC
-+ -+ -+ -+ -+ -+ -+
LE - LC
20 10
Liquid Expanded, (LE) Gas Phase, (G)
-+
-+
Water
LE
-+
-+
- + +-
Nonpolar tail Polar head
-+
Water
-+
50 100 Area per molecule, (2/molecule)
Figure 2. Surface pressure (-A) isotherm for a lipid with distinct phase regions of differing compressibility. The isotherm curve represents a typical surface pressure versus area per molecule dependence at a fixed temperature, and often shows sharp bends or kinks to indicate the phase transitions occurring in the two dimensional layer at the air/water interface. The isotherm shown is typical for a phospholipid that forms a condensed phase.
The -A isotherm in Figure 2 indicates a number of distinct segments; these segments correspond to the presence of different two-dimensional phases. When a monolayer is compressed it can go through several diverse phases. The phase behavior of the monolayer is
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based on the physicochemical properties of the amphiphilic molecules, at constant temperature, subphase pH, surface pressure, and other subphase variables. It is worth noting that the hydrophobic and hydrophilic regions of amphiphiles each play critical roles. An increase in the chain length of the hydrophobic group will result in increased attraction between molecules and favor the formation of more ordered condensed phases. The cohesive and repulsive forces will change if an ionizable or charged amphiphile are used, and the presence of charged head groups generally makes the formation of condensed phases less favorable. Conventional terminology used to classify different phases has been adopted.[13] The gaseous phase (G) typically represents the largest areas per molecule, with small molecular interactions and minimum to no lateral cohesion in the monolayer. When compressed, the hydrophobic ends of the amphiphiles will start to experience some lateral cohesion; however, the hydrophilic ends of the amphiphiles will remain randomly oriented. This phase is known as the liquid-expanded phase (LE), as show in Figure 2. Continued compression of the LE phase will, for certain chain lengths and depending on temperature, result in a first-order phase transition during which the liquid-condensed (LC) phaseemerges. This first-order phase transition has a broad coexistence region signified by a quasi-plateau in the surface pressure. The LE-LC coexistence region is analogous to the coexistence of liquid and vapor phases in three-dimensions and the proportion of the two phases at points along the isotherm within this segment is governed by the thermodynamic lever rule. The LC phase is characterized by a significant degree of chain ordering together with only short-range positional ordering of the head-groups. LC phases can have either vertically oriented or tilted chains, and transitions between tilted and vertical phases are possible. Upon compression out of the plateau-like segment, the LE phase is completely converted to the LC, and at even higher densities the monolayer finally reaches the solid phase (S), in which the molecules are closely packed and highly ordered both in terms of chain orientation and head-group positional ordering. If the monolayer is further compressed after reaching the S phase, the monolayer will collapse into three-dimensional structures (as illustrated on the right side of Figure 2 at the top).
LANGMUIR-BLODGETT FILM DEPOSITION

Langmuir-Blodgett films are generated by a successive deposition process in which a monolayer on the water surface is transferred onto a solid substrate. This process was demonstrated by Dr. Katherine Blodgett, in 1935, after the introduction of the Langmuir film by Dr. Irving Langmuir in 1917. The deposition process require surface pressure, temperature, pH, and other parameters be controlled at constant values while the transfer process is accomplished by successively dipping a solid substrate up or down at a controlled rate through the surface monolayer, as depicted in Figure 3. The solid substrate may start the process either held above the air/water interface and ready for the immersion (down stroke), or immersed and ready to be pulled out at a fixed rate (upstroke). If the substrate is hydrophobic, as is the case for chemically modified gold or silicon, the substrate will be positioned above the surface and ready for a down stroke. If the substrate is hydrophilic, such as a clean glass slide, the substrate will be position immersed into the subphase and ready for the first upstroke. When the substrate is hydrophilic and starts out immersed, the subphase
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will wet the substrate and the contact angle will be near zero at the three-phase contact line, which represents the border where monolayer transfer occurs during substrate motion. The monolayer will naturally flow onto the substrate as it is raised in this case and transfer in a head-groups down orientation. For hydrophobic substrates, the aqueous subphase does not wet the substrate and the contact angle will be near 180o upon immersion, facilitating transfer in a tails-down orientation. The degree of success in transfer is expressed in the form of the transfer ratio, defined as the ratio of the decrease in area of the monolayer on the water surface divided by the substrate area. Perfect transfer of a stable monolayer at constant surface pressure will result in a transfer ratio of unity.
A
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
Hydrophilic Substrate
B
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
-+ -+ -+ -+ -+ -+ -+
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
Hydrophobic Substrate
D
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
-+ -+ -+ -+ -+ -+ -+
-+ -+ -+ -+
-+ -+ -+ -+
-+ -+
-+ -+ -+ -+ -+ -+ -+
Figure 3. Langmuir-Blodgett Deposition (see text for detailed discussion). (A) Deposition onto a hydrophilic substrate during emersion, (B) deposition of the second layer during immersion, (C) deposition onto hydrophobic substrate during immersion, and (D) deposition of the second layer during emersion.
When the substrate is hydrophilic, during the first upstroke the very first monolayer transferred onto the substrate will orient with its hydrophilic head-groups interacting with the substrate. When the first transfer step is completed, the initial hydrophilic substrate becomes hydrophobic as it is now covered by a monolayer with hydrocarbon chains oriented towards the air. If the motion of the dipper is reversed and a subsequent immersion step is now carried out, the newly formed hydrophobic outermost layer will attract the hydrophobic chains of the surface-monolayer, as seen in Figure 3B. The result will be a bilayer structure. Continuing the deposition process (emersion and immersion of substrate) will lead to multiple layers of monomolecular amphiphiles of alternate orientation. This process is known as Y-type deposition, as shown in Figure 4 (for the case of a hydrophilic substrate). If the substrate is hydrophobic, the first monolayer will transfer tails-down on the first immersion and as the substrate is pulled out, the second layer will transfer with the headgroups interacting with those of the first monolayer. The result after multiple cycles of
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immersion and emersion resulting in multilayer formation will still be Y-type deposition, but with the first monolayer being tails-down instead of heads-down.
A
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
B
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
C
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
-+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+ -+
X-type deposition
Y-type deposition
Z-type deposition
Figure 4. X-, Y- and Z-type multilayers.
Besides the Y-type deposition, two other monolayer deposition orientations on solid substrates have been reported.[6, 46] X-type results from transfer only upon immersion steps and the resulting layers are all oriented in the same manner with the tails pointing towards the substrate. Z-type deposition occurs only on emersion steps and yields multilayers whose individual monolayers are all oriented with the head-groups pointing towards the substrates. It has been reported that the X- or Z-type deposited multilayers, as indicated by the transfer ratios, sometime rearrange into a Y-type repeating bilayer organization.[50] The above mentioned deposition types (or modes), can be influenced by many factors, such as the nature of the amphiphiles, transfer surface pressure, deposition speed, pH, temperature, packing of the monomolecular layer at the air/water interface, and the presence of dissolved subphase cations that interact with the head-groups.[51-55] Langmuir-Blodgett films can be formed by transfer of phospholipid monolayers, and also by transfer of phospholipid mixed monolayers or phospholipid-cholesterol mixtures. It is possible to transfer single monolayers, bilayers, or multilayers.[56] The transfer of multilayers of DPPC is not easy but has been enhanced by addition of DPPA (dipalmitoylphosphatidic acid) as a transfer promoter. It has been reported that for transfer onto hydrophilic substrates, transfer of the first layer is generally possible with transfer ratio of nearly 1.0; however, successful transfer of the second layer was noted as being successful only in a narrow range of surface pressure.[57] It is also possible to transfer a different phospholipid in the second layer than in the first. In a study making use of AFM, Kruijff et al. revealed that when the first layer was DPPC, AFM topographic image indicated that DPPC in general looked smooth with a few small defects, as seen in Figure 5A.[58] Furthermore, when the second layer was DPPC, the films appeared relatively smooth with a small number of round defects covering less than 6% of the surface, as seen in Figure 5B. These features were well below the resolution limit of optical microscopy methods such as fluorescence microscopy. Futhermore, Kruijff et al. also indicated the shape of the defects in asymmetric LB bilayers was influenced by the nature of the first leaflet; for example, polygonal lineshaped defects were observed in 1,2-dipalmitoyl-sn-glycero-3-phosphocholine/1,2dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPC/DPPG) bilayers. Again, the irregularly shaped features continued to be present and display elevations from the rest of 1,2-
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dipalmitoyl-sn-glycero-3-phosphocholine/1,2-dimyristoyl-sn-glycero-3-[phospho-rac-(1glycerol)] (DPPC/DMPG) bilayers as illustrated in Figure 5C and 5D (second layer leaflets of the bilayer are in the condensed phase). Whereas mainly line-shaped, polygonal features and some small irregularly shaped defects were seen when 1,2-dimyristoyl-sn-glycero-3[phospho-rac-(1-glycerol)] (DMPG) or 1,2-dioleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DOPG) were used as a second layer in the bilayer system, as seen in Figure 5E and 5F (second layer leaflets of the bilayer are in the liquid phase).[58]
Figure 5. Morphology of LB Phospholipid Layers Revealed by AFM. (A) Monolayer of DPPC (condensed phase), transferred at a surface pressure of 35 mN/m. Such a monolayer was used as the first leaflet of the following bilayers, of which the second leaflet was transferred from 10 mM Tris buffer, with 100 mM NaCl, pH 7.4, and consisted of (B) DPPC deposited at a surface pressure of 26 mN/m (condensed phase); (C) DPPG deposited at 26 mN/m (condensed phase); (D) DMPG deposited at 35 mN/m (condensed phase); (E) DMPG deposited at 22 mN/m (liquid-expanded phase); and (F) DOPG deposited at 26 mN/m (liquid-expanded phase). All images are 10 m x 10 m; scale bar = 2 m; z-scale is 10 nm. (see original article, de Kruijff, B. 1999). Reprinted with permission from reference 58.
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MOLECULES THAT FORM BIOMIMETIC MONOLAYERS

In theory, any compound or molecule possessing both hydrophilic and hydrophobic properties is a good candidate for monolayer formation. Many of the common amphiphiles, such as fatty acids, cholesterol, glycolipids, phospholipids, and others, are commonly used to prepare Langmuir-Blodgett films, because they have the potential to serve as biomimetic membranes. The structures of some of the molecules most commonly used to form biomimetic membranes are shown in Figure 6.[54-55, 59-68]
Phospholipids
Phosphatidylethanolamine
Phosphotidylglycerol
Phosphatidylcholine
Phosphotidylinositol
Glycolipids
Monogalactosyl diglyceride Digalactosyl diglyceride
Sphingolipids
Figure 6. Representative structures of some of the main membrane lipids that are potentially of interest for formation of Langmuir-Blodgett films acting as biomimetic membranes.
The hydrophobic portions of the amphiphilic molecules prevent water solubility, while the hydrophilic ends interact favorably with water. It is crucial that there be an appropriate balance between the head-group hydrophilicity and the hydrophobicity of the hydrocarbon chains for successful monolayer formation. If the hydrocarbon region is too short, the amphiphilic molecules will simply dissolved in the subphase shortly after spreading. A general rule of thumb, for the spreading of single saturated chain derivatives, is that a minimum hydrocarbon chain length of 12 ((-CH2-)n, n 12) is necessary to ensure that the hydrocarbon chain is long enough for the compound to form a spread monolayer that remains at the air/water interface long enough for most experiments.[6, 14] Monolayers are thermodynamically metastable systems, and should ultimately dissolve into the subphase; however, they do so extremely slowly if the chain length is sufficient long. It is also worth noting, introduction of a double bond (either cis or trans conformation, Figure 7A and B, respectively) into the middle of the hydrocarbon chain will significantly disrupt the molecular packing of the Langmuir firm and change the surface area per molecule and ability to undergo
Galactosylceramide
Sphingomyelin
Diphosphotidyl glycerol
Phosphotidylserine
Cholesterol
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deposition onto substrate surfaces, especially for cis double bonds in the middle of the chain. On the other hand, disruption of the molecular packing is minimal if the double bond is at the end of hydrocarbon chain, as seen in Figure 8.[46]
Oleic acid
Figure 7. Structures of Oleic Acid and Elaidic Acid.
Elaidic acid
Tricosenoic acid
Figure 8. Structure of Tricosenoic acid.
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For the case of phospholipids and related two-chain amphiphiles, monolayer formation can sometimes be successful for chain lengths shorter than 12 CH2 units.[69] For example, the surface pressure-area isotherms of 1,2-didecanoyl phosphatidylcholine and 1,2-didecanoyl phosphatidylethanolamine were reported in early studies. Such shorter chain length phospholipid derivatives form highly fluid monolayers which have featureless isotherms showing smoothly increasing surface pressure and high compressibility.[70] LangmuirBlodgett films of these very short phospholipids have not been reported. Under ambient conditions, the phospholipid DMPC is in the liquid-expanded phase, and increasing the chain length by two CH2 units to C16 in DMPC results in a monolayer that is in the liquidcondensed phase at ambient conditions and surface pressures of most interest for LB transfer.
LIPID/PROTEIN LANGMUIR-BLODGETT FILMS

The LB technique offers one of the best possibilities to perfectly control each step of the film preparation, largely due to the molecular arrangement and perfectly organization at the air/water interface. The molecular packing itself is robust and can be maintained during the transfer process onto the desired solid support, when all the parameters are optimized. In the context of biomimetic studies, LB films composed of phospholipids, such as 1palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) [71] and 1,2-dipalmitoyl-snglycero-3-phosphocholine (DPPC),[71] other phosphatidylcholines,[72-75] phosphatidylethanolamines, [76] etc, are a few of the many essential components of biological membranes of interest. The functionalization of LB films with biological molecules can be achieved by association with proteins presenting specific recognition properties, for example, enzymes, antibodies or bio-specific ligands. In the past decades, utilization of biomolecules such as enzymes or antibodies in conjunction with amphiphilic molecules via LB thin film approaches have been reported and several bioactive protein-lipid LB films have been studied with regard to their potential application in biosensor development. [4, 6, 77-83] The popularity of this approach is mainly due to the ease of preparation and incorporation of biological molecules that act as models for biological membrane structures or functions, and may be used for studying and testing isolated functions by systematic control of composition. Some crucial and lingering questions (to name a few) include: (i) determination of the minimum amount of enzyme needed for membrane preparation for biosensor function; (ii) whether one-step preparation of a bioactive sensing layer and its association with the transducer is possible; (iii) the possibility to work at ambient environmental conditions (atmospheric pressure and temperature) to avoid denaturing the biological compounds; (iv) the ability to modulate the sensor performance (detection limit, sensitivity, dynamic range) by varying the number of the deposited proteolipidic layers.[77, 84-89] The resolution of these issues remain vital for further development or design of microscale or nanoscale sensors capable of operating in controlled environments and with ultra-rapid response times. Several methodologies for incorporation of proteins, enzymes, or biological compounds into LB films have been reported including: (i) adsorption of protein from the subphase onto the interfacial film by means of spreading the monolayer onto a subphase containing the dissolved protein or by injecting the protein into the subphase after lipid monolayer
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formation,[77, 80-82, 84-86, 89-108] (ii) adsorption of the protein onto the LB film after transfer, and (iii) co-spreading of protein included in the spreading solution just prior to LB monolayer formation has also been reported.[4, 109-110] Although many successful incorporations of protein in LB monolayers have been reported, the presence of protein in LB monolayers in general can modify the packing of the amphiphile at the air/water interface and inevitably complicate the transfer procedure onto a solid support.[89, 104, 111] These complications could be attributed to the fact that protein incorporation changes the surface pressure behavior of the monolayer and its stability, can induce poor monolayer adhesion to the solid support which can lead to peeling off during subsequent immersion/deposition steps, and release of protein molecules into the subphase during deposition due to their weak association with the amphiphilic molecules, etc.[112-113] The reported strategies to overcome the above mention problems, include but are not limited to: (i) usage of a covalent binding approach, to directly immobilize protein onto the cross-linking agents in the LB film,[114-115] (ii) stabilization the proteo-lipidic monolayer by using glutaraldehyde vapour as a protein cross-linker,[81-82, 104] and, (iii) stabilizing the choline oxidase enzyme in the proteo-lipidic LB film by using the heads of the two opposing lipidic layers (referred to as inclusion by sandwiching).[7, 116-117]
PROTEIN ORIENTATION OR DIRECTED PROTEIN PRESENTATION

With the ultimate goal of biomimetic membrane applications in nanobiotechnology, understanding and controlling macromolecule presentation and orientation in proteo-lipidic nanostructures intended for use in biocatalysis or based on biomolecular recognition by the immobilized biological macromolecules, e.g. enzymes, antibodies, etc., is crucial for the success of innovative strategies in the future development of integrated nanobioelectronic devices. Many strategies have so far been reported to incorporate proteins in LB layer(s) and to build stable proteo-lipidic LB films onto solid supports. The control of the protein incorporation and presentation in a defined orientation that mimics that in a natural biological membrane to achieve maximum functionality is important and remains one of the many challenges in designing proteo-lipidic LB layer(s) for biomimetic sensing applications. To meet these challenges, numerous systematic tactics have been reported in the past decades, towards the goal of generating oriented proteins in lipidic LB films. One of the methods involved covalently associating antibody fragments to lipids bearing thiol or lipoyl head-groups via disulfide bonds to the free thiols formed after the hinge regions of the immunoglobulins had been cleaved by the enzyme pepsin. A different approach used histidine tag containing proteins associated onto monolayers of lipids bearing Ni2+ chelating head-groups.[118-121] An alternate approach reported by Girard-Egrot et al., involved inserting a non-inhibitory monoclonal antibody in LB films as a form of anchor, thus the hydrophilic protein can be associated in an oriented position at the surface of lipidic matrix.[15] In fact, the reported strategy by Girard-Egrot et. al., was also capable of increasing the shelf life of the proteo-lipidic LB films and retain the biological activity of the immobilized proteins, for up to few months.[45] In this study, activity of the
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acetylcholinesterase monomers (AChE), in an orientated position, at the surface of the IgGglycolipid LB film after immunoassociation was demonstrated. Furthermore, the approach of incorporating biological molecules, such as proteins, in liposomes and the applying Langmuir-Blodgett techniques was successful. The proteo-lipidic vesicle can be spread onto the air/water or air/buffer interface in the Langmuir Blodgett trough. Upon the disintegration of the proteo-lipidic liposomes, the proteo-lipidic LB films can be generated by compressing the film up to the transfer surface pressure (eg: 30 mN/m) and then transferring onto the solid support.[16, 122-128]
AFM CHARACTERIZATION OF SUPPORTED BILAYERS

For the characterization of the monolayers at the air/water interface that are the precursors to LB films, many advanced and promising techniques such as fluorescence microscopy and fluorescence recovery after photobleaching (FRAP) methods[129-130], Brewster-angle microscopy (BAM), x-ray and neutron scattering techniques, external reflection Fourier transform infrared (FTIR) spectroscopy, and polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) have been applied.[131-135] The above mentioned techniques have collectively provided much valuable information on the LB monolayers, such as how amphiphilic molecules pack in the liquid-expanded phase, and in the variety of possible liquid-condensed and solid-like phases. Many of the above techniques, and additional ones such as total internal reflection fluorescence (TIRF), have been applied to characterize LB films after transfer onto a solid substrate.[136] However, due to the lateral resolution limitations of the above mentioned techniques, detailed information on the arrangement of the amphiphiles at the nanometer scale level remains uncertain or ambiguous, such as the coexistence of LC and LE phases in films on solid supports, segregation of components in transferred films of lipid mixtures, lateral distribution of incorporated proteins, and the structure of the boundary region between the phases in phase-segregated supported films.[12, 137] Since the invention and introduction of scanning probe microscopy in 1986, atomic force microscopy has become a unique tool presenting new possibilities and limits for characterization of simple model systems.[29-34, 138-140] AFM provides the capability of acquiring data (such as, friction, adhesion forces measurements, viscoelastic properties, determination of Young modulus, acquiring magnetic or electrostatic properties) or topography both in air and in liquid medium down to molecular scale resolution. The Hansma laboratory was one of the first to use AFM at the high resolution of approximately 1.0 nm laterally and approximately 0.1 nm in the vertical direction, to capture images of phospholipids in LB and supported lipid bilayer (SLB) model membranes, as reproduced in Figure 9.[141-142] As illustrated in Figure 9, AFM images of the polar or headgroup regions of bilayers of dimyristoylphosphatidylethanolamine (DMPE), deposited by Langmuir-Blodgett deposition onto mica substrates were obtained.[141] The DMPE was transferred at specific molecular area of 0.41 nm2, at a surface pressure of 40 mN/m and the images captured under water at ambient temperature and pressure. The distinct features of the images are parallel lines with uniformly spaced rows roughly 0.7 - 0.9 nm in spacing. A modulation also can be seen along the rows, with rounded bright spots roughly every 0.5 nm, that correspond to the individual
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head groups of the DMPE molecule, which only the incredible resolution of the AFM can reveal.[141] Figure 9B is a subsequent trace of approximately the same area as in Figure 9A, on the LB film. The arrows point to a small patch of molecules believed to be the same in both images. Although the pattern of modulated rows is common to both images, the images are not identical and the difference could arise due to several reasons. A small amount of thermal drift (tenths of nanometer per minute) is difficult to avoid between images, which leads to small offsets between images. In addition to the offsets, depending on if the raster scan was performed top to bottom (Figure. 9A) or bottom to top (Figure. 9B), the lattice spacing and symmetry of the image are slightly different due to the distortion caused by the thermal drift during the 10 s of image acquisition. The images are also not expected to be identical because of the thermal motion of the DMPE molecules; bilayers are fairly fluid materials, and molecules likely move both horizontally and vertically between scans.
Figure 9. (A, B) Sequential grey scale AFM images of the polar region of a bilayer of dimyristoyl phosphatidylethanolamine (DMPE) deposited by the Langmuir-Blodgett technique at a specific molecular area of 0.41 nm2 and a surface pressure of 40 mN/m on a freshly cleaved mica substrate. The images were taken under water at ambient temperature and pressure. (see original article, Zasadzinski, J. A, 1991). Reprinted with permission from reference 141.
Another common short coming of contact mode AFM imaging is that the AFM probe remains continuously in contact with the sample during the raster scan of the surface. As a result, features could be affected or distorted due to AFM tip induced deformation of the relative height of surface structures by the scanning force applied during scanning. It is worth noting that acquiring AFM topography is ideal under a liquid medium when compare to imaging LB film in air under ambient conditions, because significantly less capillary or adhesion forces are experienced between the tip and the film in liquid medium.
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With the advancement and achievement of organized proteo-lipidic nanostructures based on Langmuir-Blodgett technology and their potential applications in the nanobioscience area, many available amphiphiles were used to form LB films and then examined using AFM. Studies using AFM can be found in the literature for LB films made of 1-palmitoyl-2oleoylsn-glycero-3- phosphoethanolamine (POPE)/1-palmitoyl-2oleoyl-sn-glycero-3-phospho-Lserine (POPS) and POPE/POPS/sphingomyelin (SM),[143] DPPC/1,2-dipalmitoyl-snglycero-3-phospho-(1-rac-glycerol) (DPPG),[144] 1,2-dioleoyl-sn-glycero-3phosphocholine/1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DOPC/DPPC),[145-146] and others.[137, 147-148] Figure 10 shows an example of (liquid-expanded)-(liquid-condensed) (LE-LC) phase separation in a binary mixture of 1,2-dioleoyl-sn-glycero-3-phosphocholine/1,2-dipalmitoylsn-glycero-3-phosphocholine (DOPC/DPPC, 1:1) in a LB film. Figure 10A shows the phase separated domains of DOPC/DPPC that were transferred at 32.0 mN/m onto mica. The typical height profile of the separated domains of DOPC/DPPC is approximately 6.0 , between the dark and protruding light domains, with lateral sizes ranging from 1.0 m to 2.2 m for larger features. The highly abundant small domains, approximately 100.0 nm to 400.0 nm in lateral size, would not be detected by typical optical microscopy methods, but are vividly revealed by AFM.[12]
Figure 10. Example of LE-LC phase separated binary mixture in LB film. DOPC/DPPC (1:1) films were transferred at 32 mN/m onto mica and examined in air with an AFM working in contact mode. A: low magnification height image: bar 5 m, z scale 20 nm; B and C: samples from two other preparations, bar: 2 m, z scale 15 nm; D: height image at a higher magnification of A: bar 400 nm, z scale 7 nm. E and F, corresponding lateral force (friction) images in the forward and backward direction of the tip scanning, z: 0.2 V. Reprinted with permission from reference 12.
The taller larger domains exhibit irregular shapes, as seen in Figure 10A, often with linear and angular boundaries (white arrows), indicating they correspond to LC phase domains. Figure 10B and 10C, show general characteristics of the separated domains in mixtures of DOPC/DPPC, showing the coexistence of large and much smaller domains, but also indicates that their form and relative size can vary (see original article for details).[12] More interestingly, using higher resolution or smaller scan size, Giocondi et al. revealed that
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the boundaries of the light domains also adopt angular shapes, as seen in Figure 10D. Furthermore, the frictional force images (black arrows) in Figure 10E and 10F, indicated the existence of heterogeneity of the mixture DOPC/DPPC and its molecular packing density or quality was revealed.[12]
LIPID INTERACTION WITH PEPTIDES

Due to its broad applicability, AFM rapidly evolved from a mere physical microscopy technique to one of the standard methods in life sciences. In drug delivery research, AFM has already been used in pharmacological settings.[149-150] The advancement of AFM imaging techniques extended to real-time imaging of bilayerpeptide interactions and bilayerdrug interactions.[7, 35, 37, 151-155] This unique feature of AFM provides the ability to monitor dynamic processes, such as the interaction of bilayers with proteins or drugs at nanometer scale and at picomolar concentration.[151] Dufrne et al. used phase-separated 1,2-dioleoylsn-glycero-3-phosphocholine/1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DOPC/DPPC) bilayers supported on mica, and imaged their nanoscale interaction with the antibiotic azithromycin using AFM (Figure. 11).[151] Based on the findings of Dufrne et al., increasing the incubation time progressively led to the decrease of the DPPC domains size and to finally to their disappearance. As shown in Figure 11, after 12 min the smallest DPPC domains disappeared; after 63 min, the DPPC domains were no longer visible. He attributed these observations to the incubation of DPPC/DOPC bilayers with azithromycin causing a time-dependent erosion and disappearance of the DPPC domains attributed to the disruption of the molecular packing by the drug.[151] Similarly, there is a strong interest in using simple membrane model systems, such as those of the widely studied supported lipid bilayers, to understand the interactions between proteins and lipids.[35] These kinds of model membrane systems truly allow researchers to gain insight into protein-lipid interactions in two ways. First, the presentations of the local structure of amphiphiles or lipidic LB films influence the protein adsorption behavior and the dynamics of proteins upon adsorption or interaction with amphiphiles. Also of great interest, the adsorption of protein or peptides molecules affects the lipidic bilayer. Shao's group has lead the way, using AFM to study lipidpeptide and peptidepeptide interactions, revealing the exquisite supramolecular organization adopted by gramicidin A in DPPC bilayers. Gramicidin incorporated into DPPC bilayers was observed to form linear and point-like aggregates at 2 mol%, while above 5 mol%, a percolation transition occurred yielding a network of interconnected linear aggregates. The dimensions of the aggregates were consistent with a basic hexamer unit. For phospholipids with longer chains than C16, gramicidin induced formation of multilamellar structures. The fundamental information gain from the study, could not have been obtained by other techniques.[36] In 2006, Brasseur et al. expanded AFM real-time imaging to investigate bilayerpeptide interactions in LB films. He successfully utilized and captured images of mixed DOPC/DPPC/DOPA (495:500:5 molar ratio) bilayers using AFM in buffer medium in the absence and presence of the simian immunodeficiency virus (SIV) fusion peptide, as seen in Figure 12.[156]
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Figure 11. Real-time imaging of bilayerdrug interactions. Atomic force microscopy height images (zscale: 10 nm) of a mixed dioleoylphosphatidylcholine/ dipalmitoylphosphatidylcholine (1/1, mol/mol) bilayer supported on mica, recorded following incubation with the antibiotic azithromycin (1 mM) at increasing incubation times. Reprinted with permission from reference 151.
As illustrated in Figure 12B, the addition of the SIV peptide to the DOPC/DPPC/DOPA bilayer induced profound, time-dependent modification of the bilayer morphology. With time, incubation of DOPC/DPPC/DOPA bilayers with the SIV peptide induces two consecutive, marked morphological changes, from Figure 12 B-H, i.e., depression of whole DPPC domains followed by the formation of a third elevated phase slowly covering the initial DPPC domains.[156] Again, the DPPC phase was depressed and exhibited a thickness reduction of 1.9 0.1 nm, as seen in Figure 12B, and the elevated bright features protruded 3.2 0.2 nm above the DOPC surface, after 30 min incubation of SIV peptide (Figure 12C). Higher magnification scanning of the box highlighted in Figure 12F revealed the formation of nanotubular structures suggested to be of an inverse micellar form. The depressed regions were interpreted as due to interdigitation of the peptide into the bilayer resulting in bilayer contraction. The characterization of this interaction at this resolution could not have been obtained by any technique other than AFM.
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Figure 12. Influence of the SIV peptide on DOPC/DPPC/DOPA bilayers. AFM topographic images (15 m 15 m, z-range of 10 nm) of a DOPC/DPPC/DOPA (495:500:5 molar ratio) bilayer recorded in Tris and EDTA (pH 7.4) (A, 0 min) and in a 10 M SIV peptide solution (Tris/EDTA) after (B) 15, (C) 30, (D) 40, (E) 50, (F) 60, (G) 80, (H) 110, and (I) 120 min. These results are representative of four independent experiments with at least two different regions explored. Reprinted with permission from reference 156.
FUTURE PERSPECTIVE AND CONCLUSION

For many years, researchers all over the world have been actively utilizing the Langmuir Blodgett (LB) technique to generate well-ordered ultra-thin films using natural or synthetic biomolecules (amphiphiles) that are capable of self-organization. The versatility of such LB films, with careful design or selection of organic amphiphiles, alteration of molecular orientation and packing can be controlled. These films show promise for future applications, e.g. as solar energy converting devices and as model systems of biological membranes.[5455, 157-158] In addition, relatively recently, AFM has become one of the promising and popular tools in surface science due to its unique capability and capacity in providing topographic images of model and biological membrane at a molecular scale. The distinct advantages of AFM as an imaging tool in biological science attested to its ability in capturing images under physiological conditions in real time with nanometer resolution and its ability to provide
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detailed chemical and mechanical information. With the continued demand and development, a significant amount of work has been carried out recently, where the temperature dependence of membrane domains (DMPC and DMPC/1,2-distearoyl-sn-glycero-3phosphocholine (DSPC) samples) using AFM imaging was reported.[159-160] Further new imaging modes such as AC modes, phase modulation AFM, higher harmonic AFM, bimodal AFM modes and high-speed AFM are expected to lead the way in improving special resolution of topographic features, widen the nature of the information collected, and improve the AFM sampling rate to better than 1 image per second for nano-visualization of biomolecular processes.[139, 161-169] By using the adaptive combination of both LB and AFM characterization techniques, the understanding of protein/enzyme association with LB monolayers, conformational change, structural alteration and denaturation of protein/enzyme in LB films can be realized. Undoubtedly there are challenges to be faced in extending the use of AFM to study real cell membranes, but the ability of the instrument to characterize membrane morphology and membrane bound protein structures under native conditions with high spatial resolution makes this effort highly worthwhile for providing basis support for many application in nanobiotechnology or next generation bioelectronics design or development of novel nanobiosensors.
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Chapter 4
LANGMUIR MONOLAYERS IN BIOSENSORS

Wrocaw University of Technology, Wrocaw, Poland
Jadwiga Sooducho* and Joanna Cabaj
INTRODUCTION
In recent years, the Langmuir-Blodgett (LB) technique for the preparation of ultrathin films of various organic, metallorganic, and polymeric compounds plays an increasingly important role as a means of organizing molecular materials at the microscopic level. The LB technique has many potential applications in molecular electronics, nonlinear optics and conducting thin films. The most important advantage of this method is that the characteristic of the film can be varied by changing various LB parameters, namely, surface pressure of lifting, temperature, barrier speed, dipping speed, molar composition, etc. So it is important to study different molecules having various chromophores with interesting photophysical and electrical properties, confined in the restricted geometry of the LB films to fabricate various molecular electronic devices and also to realize the basic physicochemical processes involved at the mono and multilayer films [1]. In particular LB films of conjugated polymers have been of interest, offering a possibility of obtaining extended twodimensional -electron systems [2]. Although, majority of conventional conducting polymers are not soluble in common solvents, making any LB deposition impossible, one can increase their solubility by attachment of side groups (usually nalkyl ones) to the main chains. According to this Langmuir-Blodgett, horizontal lifting or other selfassembled method is employed for obtaining molecular films of conducting structures. This type of material is popular in designing of sensor devices. Biosensors generally offer simplified reagentless analyses for a range of biomedical and industrial application. For any sensor, speed of response and reversibility are often paramount. In any solid-state sensor, analyte molecules have to diffuse into and react with the acting sensing component and any product of the reaction must diffuse out. It therefore
* Corresponding author: Department of Medicinal Chemistry and Microbiology, Faculty of Chemistry, Wrocaw University of Technology, Wybrzee Wyspiaskiego 27, 50-370 Wrocaw, Poland
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Jadwiga Sooducho and Joanna Cabaj
follows that the thinner the sensing layer is, the less time this will take and thereby speed and reversibility being improved. Such model of molecular assemblies can be prepared also by Langmuir-Blodgett, Langmuir-Schaefer (LS) techniques or by using self-assembly monolayer. The Langmuir-Blodgett layers of amphiphile proteins complexes with embedded immobilized enzymes could be also deposited directly on transducers (such as amperometric or potentiometric electrodes or field effect transistors) and be used as recognition elements (Scheme 1) [3]. In fact, proteins differ strongly from usual amphiphilic molecules. Nevertheless, the first approach to making LB films of proteins was performed by Langmuir and Schaefer in 1938. They worked with pepsin and urease and were able to demonstrate the preservation of protein activity in the deposited layer.
BIOSENSORS
O2
Substrate
En En En
En
En En
Au
SiO 2
Si Product
H2 O
Measured signal (i.e. amperometric)
Scheme 1. Simplified diagram of fabrication layered biosensor
MONOLAYERS OF BIOLOGICAL MOLECULES ON WATER

When a monolayer at the airwater interface is compressed, the surface tension of the water is reduced. This surface pressure can be measured by various means, such as a piece of filter paper dipped into the subphase and attached to a balance. Since the concentration and volume of solution spread and the area of the monolayer are known, we can plot surface area per unit against surface pressure. This plot is known as the
Langmuir Monolayers in Biosensors
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isotherm; a steep portion of the curve obtained is indicative of the formation of a well-packed structure. When deposition of a monolayer onto a solid surface occurs, a feedback circuit may be used to keep the surface pressure content. Since the area of the substrate is known, we can obtain a deposition ratio, i.e. area of monolayer deposited/area substrate. A ratio of 1 is indicative of good transfer of the monolayer. Much of the early work on monolayers at the airwater interface was performed using relatively simple chemical structures, e.g. fatty acids and amines, steroids, phospholipids [4]. However, attention was turned to formation of thin films of biological molecules, either pure or as a mixture with the more classical amphiphiles. Much of the early studies on monolayers of biological molecules have been extensively reviewed [5]. Whereas for simple molecules such as fatty acids, the behaviour of the monolayer is dependent on the nature of the hydrophilic head-group and the alkyl chain, for biomolecules a different situation occurs. The monolayer properties of these species are dependent on the balance between hydrophilic and lipophilic groups within the macromolecule. Proteins tend to form stable monolayers at the airwater interface because of their mixture of hydrophilic and lipophilic groups. Often spreading species such as proteins at the airwater interface can effect the conformation of the molecule such as causing unfolding. Many experimental conditions directly affect the structure and activity of protein monolayers. An enzyme monolayer having the desired structure and activity is a requisite for biosensor development. In contrast to Langmuir monolayers studies of small amphiphilic lipids, enzymes are macromolecules readily dissolved in the water. Buffered water is the most widely used and also most appropriate solvent for protein engineering. Polar organic solvents such as methanol, ethanol, propanol, can be used in pure form or mixed with small amount of water for protein monolayer preparation. Other relatively nonpolar organic solvent rather cannot be used for preparation of protein monolayer due to the insolubility or denaturation of macromolecules. Since water is the best solvent for proteins, the problem of preventing the enzymes from leaving the interface and sinking into the bulk aqueous suphase in the main concern involved in the Langmuir monolayer fabrication of i.e. enzymes. Earlier was reported that the addition of small amounts of salt to the suphase could effectively prevent the dissolution of proteins to the bulk solution, then to form more stable monolayers at the air-water interface. This situation was found for i.e. acetylcholinesterase [6], bovine serum albumine [7]. The solubility of proteins and enzymes in bulk form decreases in the presence of electrolytes solution, leading to growth adsorption to the interface. Other parameters such as pH and temperature also have significant effects on protein monolayers and may directly alter the activities of the enzymes, i.e. an enzyme can be charged or in its isoelectric point. With different charges the molecule can adopt a completely different conformation during adsorption process on the air-water interface.
PROTEIN-MONOLAYER ENGINEERING
Monolayers engineering aims to create complex molecular assemblies with a specific layered structure. The techniques applied are based mainly on the original Langmuir-Blodgett (LB) [9] or Langmuir-Schaefer (LS) [9] methods, very often combined with self-assembly
134
[10] and adsorption processes. Generally, monolayers of amphiphilic organic molecules are formed at the liquid-air interface in the LB through by first spreading and then compressing the organic surface layer to a defined surface pressure (Fig. 1).
substrate solvent solution barrier
subphase
Figure 1. Formation of Langmuir layer on air/subphase interface
subphase removing by pump

Figure 2. Fabrication of LB (left) and LS (right) films
The monolayer thus formed is transferred onto solid substrate that is arranged vertically and dipped through monolayer. Successive layers are built up by repeating this process. However, protein monolayers are usually prepared by adopting the horizontal-lift LS technique for transfer onto the solid substrate (Fig. 2). For enzymes, an adsorption through has proved to be even more effective in preserving the native protein function in the engineered monolayers [11].
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The latter technique of monomolecular layers formed at the air-water interface has an advantage over the other methods since it allow for a continuous control of both quality of the surface and such a parameters as molecular packing, physical state, lateral pressure and composition. In particular, mixed Langmuir monolayers composed of constituents of biological membranes, such as phospholipids, sterols, sphingo- and glycolipids, provide a highly informative approach for studying e.g. intermolecular interactions between membrane components and biomolecule [12]. As proteins are not ideal amphiphilic molecules, the techniques need to be adapted, either by chemical methods (e.g., derivatization methods [13] or varying the subphase composition [14]) or by applying some mimetic systems of biological membranes, due to preserve their native structure and function in monolayer. The quality of protein-monolayer formation at the air-water interface is related to the degree of preservation of the native properties of all proteins. The magnitude of the electrostatic forces maintaining the protein structure is comparable with that of the surface tension. Proteins tend to form stable monolayers at the air-water interface because of their mixture of hydrophilic and lipophilic groups. Often spreading species such as proteins at the air-water interface can effect the conformation of the molecule such as causing unfolding. For example insulin or ovalbumin unfold completely whereas myoglobin and cytochrome C are only partially unfolded [5]. This again is thought to be a function of the ratio of polar to nonpolar amino acids residues. Highly polar proteins such as xanthine oxidase do not form stable monolayers [14]. In all these circumstances the convenient matrix may be required. For instance, according to Girart-Ergot et al. [15] enzyme bioactivity in mixed lipid LB films is preserved due to the lipid molecular assembly protects the enzyme, positioning the polypeptide moiety in such a way as to allow the recognition and signal events. In fact, phospholipids have been used as protecting agents for several types of material, not only for membrane cell proteins [16] but also for polysaccharides [17] and synthetic polymers [18]. The lipid fraction of biological membranes is mainly composed of phospholipids with different chain lengths and ionic character. Therefore, phospholipids are widely used as mimetic systems in studies involving the cross resistance to drugs. Usually, these studies are mainly based on the interactions between immobilized biomolecule and for example drug, which could be incorporated in phospholipids structures as ordered Langmuir or LB/LS films. Consequently, the development or improvement of techniques that allow immobilization of phospholipids onto solid substrates is desirable. In this context, not only LB method but also the layer-by-layer (LbL) technique or combination of both LB and LbL procedures have been explored in the fabrication of supramolecular architectures with molecular level control. Dipalmitoylphosphatidylglycerol (DPPG) as well as dimirystylphosphatidylglycerol (DMPG) are phospholipids extensively applied in studies involving mimetic systems in the form of Langmuir monolayers [19,20], LB films [20,21] and other lamellar vesicles [22]. Phospholipids are insoluble molecules, so their evaporation and dissolution is neglible. The major cause for the poor respreading ability of phospholipids monolayers is film collapse. This collapse phenomenon was first described by Langmuir to explain the compressionextension hysteresis in surface pressure-area curves [23]. Since then, the collapse mechanism of an insoluble monolayer, such as long-chain fatty acids with chain lengths of more than eighteen carbons, has been studied [24]. The possibility of preparing multilayer films opens the perspectives of characterizing these mimetic systems using a wider range of techniques in opposition to LB method, which
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usually restricts the phospholipids films to one or two layers [21]. However, despite this disadvantage, the LB technique is still a distinctive way to produce phospholipids structured as mono- or bilayers like they are found in the cell membrane models. The monolayer of DPPG exhibits a liquid-expanded to liquid condensed (LE-LC) phase transition evidenced as a plateau in the -A isotherm. This transition is typical of phospholipids films at temperatures below that of the gel-liquid crystal transition temperature (41oC) [25]. The surface pressures at the beginning and the end of the transition are 4.8 and 6.3 mN/m, respectively (Fig. 3). Consequently, this is not a true first-order phase transition due to the fact the surface pressure does not remain constant. The increase in surface pressure during the transition may be due to the fact that film was not compressed sufficiently slowly. However, the use of faster rates of compression leads to an increase in -transitions, as described [26]. The same phenomenon occurs, even more significantly, upon increasing the temperature or pH [27,28]. A technique suitable for the deposition of enzyme LS films onto glass substrate processes that the surface of the solid is sometime activated (e.g. siloxane polymer in case of urease [29]).
60
Surface pressure [mN/m]
40
20
0 30 40 50 60 70
2
80
90
Area per molecule [ ]
120
Compression modulus [mN/m]
100 80 60 40 20 0 0 10 20 30 40 50 60 Surface pressure [mN/m]
Figure 3. Surface pressure ()-mean molecular area (A) isotherm of dipalmitoylphosphatidylglycerol (DPPG) - a. Subphase: phosphoric buffer, pH 7.0, T = 295 K. B the elasticity (compression modulus, Cs-1) values versus surface pressure plots for investigated monolayer
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The collapse behavior of DPPG monolayer is different from that of fatty acid monolayer. The collapse pressure (c) and the equilibrium spreading pressure (e) are two properties commonly used to describe the collapse behavior of a surface film. The collapse pressure is generally taken as the maximum of a spike or a plateau appearing in the dynamic -A curves [24], while the equilibrium spreading pressure (c) is the maximum equilibrium surface pressure attained by gradually dropping the finely dispersed sample on the substrate surface [30]. From a thermodynamic point of view, the monolayers essentially collapse instantaneously at their respective equilibrium spreading pressures (e). However, solid films of surfactants, both phospholipids and fatty acids, can be over-compressed to surface pressure much greater than their e. For a mononlayer with e< < c, the monolayer is suggested to be a metastable (supersaturated) state [31]. Usually, the monolayers of fatty acids exhibit collapse behavior at the surface pressure slightly greater than their e, and a nucleation and growth model was proposed to be the collapse mechanism of these supersaturated monolayers. On the other hand, the relaxation of supersaturated DPPG monolayer showed that it does not collapse until surface pressure nears its c (Fig. 3a). Smith and Berg [24] have therefore suggested that these films collapse by a fracture process. Additionally, the DPPG monolayer and its c is irreversible. From the result of the static saturated spreading surface pressure and dynamic -A curves measurements, there was obtained the values of 35 and 55 mN/m for e and c of DPPG monolayer at 22oC, respectively. The value of c is dependent on experimental conditions. For monolayers of fatty acids, such as stearic acid, their c will increase as the compression rate increases. Nevertheless, for DPPG, the compression rate had no significant influence on the value of its c. Obviously, the relaxation behavior of DPPG at e< < c suggests that the monolayer does not collapse in this region, it is believed that with the DPPG monolayer at e< < c, the monolayer should be at a metastable state. In this state DPPG behave more like an elastic solid film instead of like supersaturated liquid monolayer. DPPG monolayers collapse at > c, which is like the yield point of a solid film.
INTERROGATION TECHNIQUES FOR THE STUDY LAYERS

Protein monolayer films can be characterized by almost all known biophysical techniques circular dichroism, Brewster angle microscopy, atomic force microscopy, scanningtunneling microscopy, infrared and electron-paramagnetic-resonance spectroscopy, X-ray diffraction, absorption spectra, nanogravimetry and other. It is in no way intended here to provide a full description of the methods of characterizing thin films, since that would comprise an entire book, not just a short review. Many of the techniques described briefly below are reviewed moreover in more detail elsewhere [32-34].
Monolayers at Air-Water Interface

During compression a monolayer at the air-water interface, the surface tension of water is reduced. This pressure can be measured by various means, such as piece of filter paper and mostly by platinum Wilhelmy plate dipped into the subphase and attached to a balance. Since
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the concentration and volume of solution spread and the area of the monolayer are known, it is possible to plot surface area per unit against surface pressure. This plot is known as isotherm. When deposition of monolayer onto solid surface occurs, a feedback circuit may be used to keep the surface pressure content. Since area of the substrate is known, it is possible to obtain a deposition ratio, i.e. area of monolayer deposited per area of substrate. A ratio of 1 is indicative of good transfer of the monolayer.
Brewster Angle Microscopy (BAM)

Brewster angle microscopy (BAM) is a well established method for visualization the morphology of ultra-thin surface films over aqueous subphase. BAM images show contrast within a film from differences in reflectivity to polarized light incident at Brewsters angle for the clean air-water interface. Without a surface film the reflection is zero (black field), but when the thin film forms the reflective index is different from that of the subphase (white field). This technique permits to observe separation of the monolayer of domains and surrounding film after interaction between different surface active molecules. The value of the Brewster angle depends upon the material at the surface, and so it is possible to visualize the domain structure of the monolayer, as the presence of the monolayer changes the Brewster angle [35]. The morphology of phospholipids monolayers over water subphase are well known, but one interesting morphologic feature regarding these lipid layer is the formation of well defined, bean-shaped domains that appear when the film undergoes the liquid-expanded-toliquid-condensed transition (Fig. 4). At the beginning of the compression, the film is completely homogenous, and on decreasing of surface area, small spots appear. Such spots change to bean shaped domains. After the end of the phase transition, these domains coalesce and the film becomes homogenous on the liquid-condensed phase until they collapse [36].
Figure 4. BAM images of L--dipalmitoylphosphatidylcholine at the air-water interface [37]
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Addition of hydrophilic unit (i.e. cytochrome b6f complex) in the subphase to spread phospholipid monolayer leads to considerable changes of surface morphology and homogeneity of mixed film. As a result of surface interaction between the protein and the lipid molecules are formed aggregates and large surrounding areas. The obtained result suggests stronger surface interactions between the protein complex b6f and lipid molecules in mixed monolayer and formation of more packed structure at the surface. In case of phospholipids frutalin (-D-galactose lectin) film formation it was observed that galactocerebroside stabilizes the formation of liquid-condensed domains in the phospholipids film, which could be formed at lower values of surface pressure [37]. Frutalin showed two main effects over the morphology of the mixed films: first, it favors the appearing of clusters even for the noncompressed film, that is also a reason for the more expanded isotherm at high areas (Fig. 5). Second, the lectin promoted the blending of the lipids, since the duality of domains existent in the absence of the protein vanished in presence of it. Probably, it is due to the expulsionm of frutalin at high states of packing, predicted by the surface pressure-area isotherm.
Figure 5. Surface pressure vs area isotherms phospholipids, Langmuir monolayers formed over subphases of pure PBS buffer (, ), and PBS buffer with 0.1g/mL frutalin dissolved (, ) [36]
Mass-Sensitive Techniques
The mass of a thin film can be monitored using a quartz crystal microbalance (QCM). It is useful for monitoring the rate of deposition in thin film deposition systems under vacuum. In liquid, it is highly effective at determining the affinity of molecules (i.e. proteins, in particular) to surfaces functionalized with recognition sites. Larger entities such as viruses or polymers are investigated, as well. QCM has also been used to investigate interactions
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between biomolecules. This if often especially suitable for sensing applications since it can monitor mass changes taking place whilst the sample is immersed in a solution. Binding or desorption of species will cause the mass of the film to be changed, and this can be monitored in real time. Reviews on this subject include [38] and [39]. The change in resonance frequency is recorded after each deposition step and correlated to the deposited mass (m, ng) and layer thickness (t, nm) by the Sauerbrey equation [40]. -f = [2f02 /Aq q]m (1)
Where f0 (Hz) is the resonance frequency, A is the area of the electrode, q is the quartz density, and q is its shear modulus. The Sauerbrey equation was developed by G. Sauerbrey in 1959 as a method for correlating changes in the oscillation frequency of a piezoelectric crystal with the mass deposited on it. He simultaneously developed a method for measuring the characteristic frequency and its changes by using the crystal as the frequency determining component of an oscillator circuit. His method continues to be used as the primary tool in quartz crystal microbalance experiments for conversion of frequency to mass and is valid in nearly all applications. Because the film is treated as an extension of thickness, Sauerbreys equation only applies to systems in which the following three conditions are met: the deposited mass must be rigid, the deposited mass must be distributed evenly and the frequency change f / f < 0.02.
Spectroscopy and Microscopy

Many of the spectroscopic techniques used to characterize bulk samples can also be used to characterize deposited thin films. UVvis spectroscopy can be performed on samples deposited on suitable transparent or reflective substrates and can give quantitative measurements of the amount of material present. FTIR spectroscopy can also be used and can give information about any reactions that may have taken place within the film. The problems of sensitivity can be overcome by depositing the thin film on an ATR crystal or on a gold-coated substrate and using reflection/adsorption FTIR spectroscopy. Xray photoelectron spectroscopy (XPS) can give the elemental composition of the film and also some depth profiling-related information, giving an idea of the physical location of various moieties within the film. Low-angle X-ray diffraction can give Bragg peaks which give a measurement of order and repeat spacing within the film [23]. Ellipsometry can be used to measure film thickness and refractive index [41]. Atomic force microscopy (AFM), scanning tunneling microscopy (STM) and scanning electron microscopy (SEM) can be used to visualize the thin film and can give information about the regularity of the film, any phase separation or aggregation and may be able to visualize any binding events that have occurred [33]. To further explore the nature of chain organization within each thin film is utilized by UV/vis absorption and PL spectroscopy. It has been known that a specific interchain stacking
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of conjugated polymers results in the formation of new electronic species called aggregates [42, 43-48].
Atomic Force Microscopy (AFM)

Atomic force microscopy (AFM) or scanning force microscopy (SFM) is a very highresolution type of scanning probe microscopy, with demonstrated resolution on the order of fractions of a nanometer, more than 1000 times better than the optical diffraction limit (Fig. 6) Because the atomic force microscope relies on the forces between the tip and sample, knowing these forces is important for proper imaging. The force is not measured directly, but calculated by measuring the deflection of the lever, and knowing the stiffness of the cantilever. Hooks law gives F = -kz (2)
ent.
where F is the force, k is the stiffness of the lever, and z is the distance the le Laser Photodiode
where F is the force, k is the stiffness of the lever, and z is the distance the lever is bent. Because of AFMs versatility, it has been applied to a large number of research topics.
Cantilever Sample
Scanner
Figure 6. General principle of AFM
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Direct interaction between the scanning stylus and the biological sample is required, there is a potential risk of sample deformation. Therefore, samples well known from electron microscopic and x-ray analyses have been studied with the AFM to demonstrate the precision of topography recorded with the AFM. The structure of the heptametrical GroES complex was seen at a resolution of 1 nm with the AFM before the x-ray structure was available to confirm the topography data [49]. Furthermore, comparative electron microscopy and AFM analyses of DNAprotein complexes and DNA triplexes have been reported as well [50]. Although reliable instruments allowing routine operation have been available for several years, progress has been made recently by improving the sample-preparation methods and by finding buffer conditions that optimize the tipsample interaction. The AFM is now a powerful tool and can reveal the surface structure of protein assemblies in their native environment at submolecular resolution. As demonstrated with the few examples here, it is not just the resolution that makes these topography attractive but also the ability to monitor conformational changes of biological assemblies directly and under native conditions.
EFFECTS OF SOLVENT
Protein-based catalysts are less stable than many traditional chemical catalysts, particularly in water-miscible organic solvents. These commonly used solvents disrupt the tertiary structure of the enzyme, so inactivated it. Nevertheless, it is well known that proteins are stable in some organic solvents and even multicomponent enzymes have been shown to retain catalytic activity, albeit at significantly lower levels than in water [51]. Proteins in hydrophobic solvents are thought to retain their native structure as a result of kinetic trapping, which results from stronger hydrogen bonding between the protein atoms and a more rigid structure in the absence of water. In hydrophobic water-immiscible solvents, any water that might be present will tend to stay at the protein surface because of the solvophobic and hydrophilic nature of the protein surface [52]. In fact, the addition of even a minute amount of water (1% v/v) is sufficient to drastically increase catalytic activity in these unnatural solvents; this observation is linked to the role that water plays in the structure and dynamics of the protein [53]. Conversely, polar solvents that can easily strip water from the surface of the protein and compete strongly for hydrogen bonds between protein atoms (e.g. dimethyl sulfoxide [DMSO], dimethylformamide [DMF], formamide) usually denature the structure to a largely unfolded state [54]. Alcohols have some hydrophilic component, but are only moderate competitors for amide hydrogen bonds. They tend to disrupt tertiary structure and leave secondary structure interactions largely undisturbed. Indeed, methanol has gained some attention as a denaturant that increases the concentration of possible folding intermediates and has therefore been used in protein folding studies [55]. In this light, it is noteworthy that, although there is no doubt that there exists a minimum structural requirement for catalytic activity, the idea that all proteins must be intact relative to the native state for catalysis to occur is not completely general, as partially unfolded subtilisin Carlsburg was recently found to be catalytically active in organic solvents [56].
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Most of organic solvent-tolerant enzymes are lipolytic and proteolytic enzymes. Although impurities often influence the stability of enzymes in the presence of organic solvents, some were investigated without enzyme purification. High thermal stability of enzymes is considered to be positively correlated with stability in the presence of organic solvents [57]. Thermophiles and hyperthermophiles are host for many useful thermophilic enzymes, i.e. an esterase from Pyrobaculum calidifontis was stable in water-miscible solvents including methanol, ethanol, 2-propanol, acetonitrile and DMF [58]. However, the enzyme activity was markedly reduced in these solvents.
IMAGING SURFACE STRUCTURE

During the last decade, the unique ability of AFM to image specimens at subnanometer resolution and in aqueous solution has been intensively exploited to investigate the structure of cell surface layers made of two-dimensional protein crystals. Traditionally, X-ray crystallography has been the premier technique for atomic resolution of protein crystals, while electron crystallography examines the specimen in a more native-like environment at nearatomic resolution. More recently, the oblique lattice of the S-layer of Bacillus stearothermophilus was visualized to a lateral resolution of about 1.5 nm [59], and nanometer lateral resolution was achieved for the S-layers of B. coagulans and B. sphaericus strains recrystallized on silanized silicon substrates [60]. Interestingly, an elegant preparation method has been developed to investigate S-layers from Bacillus species in conditions relevant to their native state [61]. The proteins are recrystallized on a lipid monolayer in a Langmuir-Blodgett trough, and the composite lipid/S-layer structure is then deposited on a flat substrate. Under these conditions, S-layers attach to the lipid film with their inner face, which corresponds to the orientation found in the living organism (Table 1). The morphology of the deposited multilayered structures is characterized at nanometer level by AFM that is a tool with different possibilities and limits. Multilayered structures are possible to form because the technique gives the possibility to form multilayered architectures controlled at molecular level. Atomic force microscopy (AFM) can image biological samples under aqueous conditions with high resolution in three dimensions without the use of any probes. AFM has been successfully used to image isolated phase separated bilayers and peptidelipid domains in supported bilayers. Also monolayers containing glycosphingolipids and cholesterol have been imaged [66] as well as phenoloxidases or glucose oxidase mixed (with linoleic acid, phospholipids) LB/LS films [3]. The phenoloxidases (laccase, tyrosinase) and glucose oxidase hetero layers were visualized by contact mode AFM (Figs. 7ab and 8ab). The enzyme molecules were fairly well deposited onto solid substrate. Immobilized phenoloxidases as well as glucose oxidase were observed as an aggregated pattern in solid-like state with keeping their characteristic random cloud-like or island structure. The heterogeneous films roughness was found relatively high (especially in case of tyrosinase film) for an LB film, which indeed shows that the enzymes were transferred. The roughness of linoleic acid - laccase film was measured as 7.17 nm (similar results was found for film of lipase [67]), when the roughness value of tyrosinase film was found as 19 nm. To compare, the roughness of glucose oxidase LB film has been
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measured as 0.38 nm [68] or 1.9 for LS films. These obtained values were attributed to the immobilization process of comparatively large molecule aggregates of enzymes (laccase, tyrosinase, glucose oxidase) incorporated to LB/LS films. This leads to conclude that there is sometime formation of an agglomerate of enzymes rather than an organized monolayer at the air/water interface. The AFM results showed that the effect could be also associated with changes in the enzyme conformations A monolayer rearrangement, such as two- dimensional formation or hindered molecular orientation, might take place during the phase transition behaviour resulted in the molecular aggregates on the protein layer. Table 1. Imaging the ultrastructure of different surface layers
Organism Sample Immobilization Observationsa Reference procedure Covalent linkage Square lattice; r 12 nm 62 to glass/mica (agreement with electron microscopy) Recrystallization Square lattice; r = 1-2 nm 61 on supported (novel S-layer/lipid lipid bilayers bilayer structures) Recrystallization Oblique lattice; r 1.5 59 on various siliconnm (study of surfaces recrystallization process) Adsorption on Conformational change 63 mica of central pores
Bacillus sphaericus S-layer
Bacillus sphaericus S-layer
Bacillus stearothermophilus Deinococcus radiodurans Halobacterium halobium
S-layer
S-layer Purple membrane
Escherichia coli
Adsorption on Hexagonal symmetry; r = 64 mica, silanized 1.1 nm glass, and supported lipid bilayers Aquaporin Z Assembly on Tetramers; p42(1)2 and 65 mica in the p4 symmetry, r < 1 nm presence of lipids (proteolytic cleavage force-induced conformational changes)
r values are lateral resolution determined directly from the images or after image processing
Figure 7. AFM topography images of a) linoleic acid laccase LB film, b) linoleic acid tyrosinase LB film [3]
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Figure 8. AFM images of a) phospholipids tyrosinase LS film, b) phospholipids glucose oxidase LS film
LB FILMS, INCORPORATION WITHIN BIOSENSORS

After the initial work on bioactive molecules in monolayers, it was inevitable that workers would attempt to transfer these films onto solid substrates since a major problem with the use of LB films is their extreme fragility, requiring deposition onto a suitable substrate for support and to allow measurements to be made on the film. One of the earliest papers [69] reports deposition of phospholipids or cholesterol onto an ionically conductive polyacrylamide hydrogel, giving a structure capable of an electrochemical response. The sensing of i.e. glucose is of paramount interest and much work has been done on developing biosensors based on glucose oxidase which can be adsorbed into a polymeric matrix or cast as a thick film and cross-linked with glutaraldehyde. Problems with these sensors include stability and slow response times. Table 2 shows the behaviour of some of these conventional sensors and compares them with some of the sensors manufactured using the techniques described. Often species of biological interest are water-soluble which means they cannot be directly deposited by the LB method, however, should they be charged molecules, they can be dissolved in the subphase. If a layer of an oppositely charged amphiphile is then spread on the water surface and formed into an LB film, the biomolecule will then be incorporated into the LB film. Penicillinase could be co-deposited [70] with stearic acid onto an ISFET to give a penicillin sensor. Glucose oxidase was also studied [71,72] by using different lipids to adsorb the enzyme from solution and showing that a glucose sensor could best be made by codepositing glucose oxidase with octadecyltrimethylammonium chloride. Okahata et al. [73] mixed glucose oxidase with a cationic lipid and deposited a bilayer on a platinum electrode and showed it to respond to glucose with a response time of 5 s, much faster than other techniques (Table 2). FTIR studies [74] were performed on fatty acid/glucose oxidase and phospholipids/glucose oxidase monolayer deposited onto ATR crystals, the fatty acid was shown to incorporate more of the enzyme. Chymotrypsin and urease as well as fenoloxidases were also studied [75, 3]. Glutathione-S-transferase is another enzyme which can be spread at the interface and compressed and deposited as an LB film [76]. The resultant film was shown to maintain its biological activity towards pesticides such as atrazine, even after being heated to 423 K,
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whereas in solution all activity is lost at 353 K. The same group also successfully deposited enzymes such as urease onto silanised glass surfaces [77] and studied the films using QCM and potentiometric measurements. Phenolooxidase (laccase, tyrosinase) enzymes could be immobilized via glutaraldehyde coupling onto amine-terminated thiol monolayers [78] and then used to detect catechol. A combination of fungal laccase and a cystamine monolayer gave optimal results, giving a linear amperometric response between concentrations of 0.001 and 0.4 mM. An interesting variant on the use of goldthiol monolayers has been rather than attaching the thiol to gold electrode, instead to use small colloidal gold particles as the substrate. Table 2. Comparison of selected protein sensors immobilized in thin films
Immobilization method Sample Response thickness time 2 layers 0.12 min 2 min Stability Reference 3 month 73 40 days 79 80 81 82
Glucose oxidase LB deposition with lipids Glucose oxidase LB deposition with 1 layer polythiophene Catalase immobilized in LB film of 1 layer phospholipids Alcohol dehydrogenase in LB film of 1 layer phospholipids Laccase LB deposition with N-heptyl- 5 layers bis(thiophene)carbazole and tricosenoic acid cross-linked with glutaraldehyde Horseradish peroxidase LB deposition 1 layer with phsospholipids Laccase LB deposition with N-nonyl- 5 layers bis(thiophene)diphenylamine and stearic acid cross-linked with glutaraldehyde Tyrosinase LB deposition with N5 layers nonyl-bis(thiophene)diphenylamine and stearic acid cross-linked with glutaraldehyde Laccase LB deposition with 5 layers benzothiadiazole-based copolymer
not reported >3 month not reported >3 month 1.5 min >3 month not reported > 2 weeks 1.5 min >3 month
83 84
2 min
>3 month
84
1 min
>3 month
CONSLUSION
The LB technique has a myriad of uses, but generally takes on one of two roles. First the LB trough can be used to deposit one or more monolayers of specific amphiphiles onto solid substrates. They are in turn used for different areas of science ranging from optics to rheology. Secondly, the LB technique can be used itself in an experimental device to test interfacial properties such as the surface tension of various fluids, as well as the surface pressure of a given system. The system can also be used as an observation mechanism to
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watch how drugs interact with lipids, or to see how lipids arrange themselves as the number to area ratios are varied. The use of thin films, especially self-assembly ones provide a simple method for the fictionalization of electrode surfaces using nanogram amounts of material. Langmuir-Blodgett technique can give either highly ordered films, giving a high level of control of the environment and often resemble the environment found inside biomembranes, thereby helping to stabilize proteins. Biosensors produced using these various types of films can display high sensitivities, be easily interrogated using electronic, optical or mass-sensitive techniques, can often be regenerated and display good stability. The potential for usable devices based on these types of films devices in the medical diagnostic and environmental monitoring applications is immense. Accordingly, for nearly 50 years we have witnessed tremendous progress in the development of electrochemical biosensors. Elegant research on new sensing concepts, coupled with numerous technological innovations, has thus opened the door to widespread applications of electrochemical biosensors. Major fundamental and technological advances have been made for enhancing the capabilities and improving the reliability of chemical measuring devices. As this field enters its fifth decade of intense research, we expect significant efforts that couple the fundamental sciences with technological advances.
LITERATURE
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Chapter 5
ADSORPTIVE CHARACTERISTICS OF BOVINE SERUM ALBUMIN ONTO CATIONIC LANGMUIR MONOLAYERS OF SULFONATED POLY (GLYCIDYLMETHACRYLATE)GRAFTED CELLULOSE: MASS TRANSFER ANALYSIS, ISOTHERM MODELING AND THERMODYNAMICS
T. S. Anirudhan and P. Senan
Department of Chemistry, University of Kerala, Trivandrum, India
ABSTRACT
Investigation on adsorption behaviour of Bovine Serum Albumin (BSA) on polymeric adsorbent materials is critical for many analytical and biomedical applications. In the present study a novel adsorbent poly(glycidylmethacrylate)-grafted-cellulose having sulfonate functional groups (PGMA-g-Cell-SO3H) was prepared by graft copolymerization of glycidylmethacrylate (GMA) onto cellulose in the presence of ethyleneglycoldimethacrylate as crosslinker using ,-azobisisobutryronitrile as initiator followed by the introduction of sulfonic acid groups through ring opening reaction of the epoxide groups of the grafted GMA with sodium sulfiteisopropanolwater mixture. The original and the modified materials were characterized by means of FTIR, SEM, XRD and BET analysis. Adsorption characteristics of BSA onto PGMA-g-Cell-SO3H were investigated under different optimized conditions of pH, contact time, initial BSA concentration, adsorbent dose and temperature. The maximum value of BSA adsorption was found to be 49.95 and 72.07 mg/g for an initial concentration of 100 and 150 mg/L, respectively at pH 4.5. Kinetic studies showed that the equilibrium conditions were achieved within 3 h. The kinetic data obtained at different concentrations and temperatures were analyzed using a pseudo-first-order and pseudo-second-order
Corresponding author, Tel : +914712308682, E-mail address : tsani@rediffmail.com (T. S. Anirudhan)
152

equation. The adsorption process followed pseudo-second-order kinetics. The experimental kinetic data were correlated by the external mass transfer and intraparticle mass transfer diffusion models. The intraparticle mass transfer diffusion model gave a better fit to the experimental data. Experimentally obtained isotherms were evaluated with reference to Langmuir, Freundlich and Sips equations. The isotherm data were best modelled by the Langmuir isotherm equation and the maximum monolayer adsorption capacity was found to be 124.85 mg/g at 30 C. Thermodynamic study revealed an endothermic adsorption process. The negative G values indicate feasible and spontaneous adsorption of BSA onto PGMA-g-Cell-SO3H. The positive and small value of enthalpy change Ho (9.50 kJ/mol) indicates the endothermic nature of adsorption primarily through weak physical forces between adsorbent and adsorbate. The positive and small value of entropy change, So (185.52 J/mol/K) indicates that the order less nature of adsorption system increases with adsorption of BSA onto adsorbent surface. Also at all temperatures Ho<T So, indicating that the BSA adsorption onto PGMA-gCell-SO3H is dominated by entropic rather than enthalpic changes. The values of isosteric heat of adsorption computed from linear isotherms remained almost constant (~ 26.45 kJ/mol) with different surface loadings suggest that the surface of PGMA-g-Cell-SO3H is energetically more or less homogeneous and the lateral interactions between adsorbed BSA ions do not exist. Adsorbed BSA was eluted completely by 0.2 M CH3COOH solution. Results obtained from repeated adsorption/desorption process showed that PGMA-g-Cell-SO3H can be used for the adsorption of BSA from various aqueous solutions.
Keywords: Cellulose; Graft copolymerization; BSA adsorption; Kinetics; Desorption
1. INTRODUCTION
Conventional systems for the separation of proteins from aqueous solution are usually very complex, and involve different techniques in many process steps, which led to very low overall process yield and high product cost. This was solved by selective adsorption technique by the specific interaction of proteins with adsorbent surface so that the protein can directly extracted through a simple adsorption process. The overall protein adsorption process comprises various steps or stages. Transport of the protein from the bulk solution into the interfacial region, attachment of the protein at the adsorbent surface and the relaxation of the protein on the surface. Depending on the extent of relaxation the molecule may detach more or less readily [1]. Several interactions involved in the protein adsorption are hydrophobic interaction, hydrogen bonding, and electrostatic interaction; simultaneously take part in controlling its behaviour [2]. The adsorption of proteins onto polymer grafted cellulose has been studied extensively; however the results were not consistent owing to the complexity of the adsorption phenomenon [3-5]. In particular, graft polymerization technique allows introducing the chains of a precursor-monomer, which can be subsequently modified to desirable functional groups. Bovine Serum Albumin (BSA) was adopted as a model protein since it is easily available and because it displays high structural homology with its human counterpart (HSA). The most important physiological function of serum albumin is to maintain the osmotic pressure of blood and to transport a wide variety of endogenous and exogenous compounds including fatty acids, metals, amino acids, steroids and drugs. Selective adsorption of BSA on various synthetic adsorbents has been examined under
Adsorptive Characteristics of Bovine Serum Albumin
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different conditions (such as solution pH and protein concentration) and the mechanisms of selective adsorption have been reported [6-10]. In the present study, an epoxy group containing monomer, glycidylmethacrylate (GMA), is successfully used as a precursor monomer and subsequent ring opening of the epoxy groups with sodium sulfite/H2SO4. [11]. The discussion will cover the synthesis of sulfonated GMA grafted cellulose adsorbent (PGMA-g-Cell-SO3H), characterization of PGMA-g-Cell-SO3H, adsorption parameters such as pH, temperature and initial BSA concentration were investigated in batch systems.
2. MATERIALS AND METHODS

2.1. Reagents
BSA (66 kDa) purchased from Sigma (A-7638 globulin free) and reagent grade cellulose (Cell) powder obtained from S. D. Fine-Chem. Ltd. (Mumbai) were used as received. Analytical grade GMA, ethyleneglycoldimethacrylate (EGDMA), ,-azobisisobutyronitrile (AIBN), isopropyl alcohol, cyclohexane, polyvinyl alcohol (PVA) and sodium sulfite were Aldrich products. Sulfuric acid was purchased from E, Merk, India, Ltd. All the reagents were used without further purification. All aqueous solutions were prepared in double distilled water. The particle size of the adsorbent was between 80 and 230 US standard mesh (average particle size 0.096 mm).
2.2. Synthesis of PGMA-g-Cell-SO3H

The preparation scheme for PGMA-g-Cell-SO3H is shown in Scheme 1. This scheme consists of the following three steps. (1) Graft copolymerization of GMA onto Cell (GMACell) (2) Introduction of the -SO3H group onto GMA-Cell using sodium sulfiteisopropanol water mixture. (3) Blocking of the remaining epoxy group with H2SO4 .
2.2.1. Graft Copolymerization of GMA Onto Cell (GMA-Cell)

GMA-Cell is prepared via suspension polymerization. About 20 g of Cell was suspended in 100 mL distilled water in a beaker with constant stirring. The organic phase containing GMA (19.5 mL), EGDMA (1.0 mL; cross-linker), cyclohexane (15 mL) and AIBN (0.2 g; initiator) were added to the flask containing 3% PVA (100 mL; stabilizer). The contents were added to the beaker containing Cell and the polymerization reaction was carried with continuous stirring in a water bath at 65 C for 6 h. Finally the temperature of the reaction system was raised to 80 C for 30 min to evaporate cyclohexane. After the reaction, the product was filtered under suction and washed repeatedly with hot water (65 C) and ethanol to remove unreacted chemicals and then dried in vacuum at 60 C.
154
CN CH3 CH3 C N=NC CH3 CH3CN AIBN O CN CH3 C. + CN OH-H2 H C H O H H N N

CN CH 3 C CN 2-cyanoprop-2-yl radical 2
OH
OH O
O H NC CH3 O C-O-H2C H CN n
OH H
H O
O.
CH3 + CH2=C-C-O-CH2-CH-CH2 O O GMA
Cell
n 2-cyanoprop-2-yl initiated cell radical H2C O O CH3 O CH2
C H3C
EGDMA
O H CN CH3 C-O-H2C CN O H
Na 2 SO3 /H2 SO4
O C-O-CH2-CH CH2 H O (CH2-C)n OH H2C O CH3 C H O HO O H O n O H CH OH CN H H2 3 O CH2-O-C H CH3CN C-(C-CH2)n O O O H H2C HC-H2C-O-C H H O O n
O C-O-CH2-CH CH2 SO3 H H (CH2-C)n OH O H OH H2C O O CN CH3 CH3 C-O-H2C C H O HO H O CN H O n O H CH OH CN H H2 3 O CH2-O-C H CH3CN C-(C-CH2)n O O O H H3OS H2C HC-H2C-O-C H H O OH n
GMA-Cell
PGMA-g-Cell-SO H
3
Scheme 1. Preparation of PGMA-g-Cell-SO3H
2.2.2. Functionalization of GMA-Cell

The adsorbent, PGMA-g-Cell-SO3H was prepared by the ring opening reaction of the epoxide group of GMA-Cell [12]. Briefly, 10 g of GMA-Cell was allowed to react with 30 g sodium sulfite (Na2SO3), dissolved in 57.5 mL isopropyl alcohol (IPA) and 225 mL water. The composition was Na2SO3:(IPA):water equal to 10:15:75 wt%. The mixture was refluxed at 80 C for 4 h. The product was filtered, washed several times with distilled water. The remaining epoxy groups were converted into diol groups by reaction with 0.5 M H2SO4 for 2 h at 80 C [13]. This was done to reduce non selective adsorption. The resulted product,
155
PGMA-g-Cell-SO3H was filtered, washed with distilled water and dried at 65 C. It was sieved and particles having the average diameter 0.096 mm were used through out the study.
2.3. Characterization of PGMA-g-Cell-SO3H

2.3.1. Determination of % Graft Properties
The percentage graft yield (%G) and grafting efficiency (%GE) were calculated on dry weight of Cell from the increased weight of Cell after grafting by using the following relationships [14].
%G
( w2 w1 )100 w1 ( w2 w1 )100 w3
(1)
%GE
(2)
where w1, w2 and w3 represent the weights of Cell, graft copolymer and monomer, respectively.
2.3.2. Determination of the Epoxy Groups Content in GMA-Cell

The amount of surface functional epoxy groups content in GMA-Cell was determined by pyridineHCl method [15]. About 1 g of GMA-Cell was refluxed with 50 mL of pyridine HCl mixture (2 mL HCl and 123 mL pyridine) for 20 mins. After cooling down the solution, the amount of available epoxy groups was determined by titration of pyridineHCl solution with 0.1M NaOH.
2.3.2. Determination of the Sulfonic Acid Groups Content in PGMA-g-Cell-SO3H

The sulfonic acid group in PGMA-g-Cell-SO3H was determined by titrating the H-form of PGMA-g-Cell-SO3H with 0.05 M NaOH. The -SO3H group density and its conversion by sulfonation were calculated as follows:
SO3 H group density (mmol / g )
Amount of SO3 H groups w
(3)
Conversion%
100 Amount of SO3 H groups Amount of epoxy groups before sulfonatio n
(4)
where w is the weight of PGMA-g-Cell-SO3H in the H-form.
156
2.3. Characterization Techniques

The adsorbent was examined with FTIR spectroscopy (Schimadzu FTIR model 1801, range 4000-400 cm-1), X-ray diffraction patterns (Rigaku Dmax IC model, Japan), Scanning electron microscope (Phillips XL-3CP microscope unit), N2 adsorption-desorption technique (Quantasorb Surface area analyzer, QS/T, zero point charge (potentiometric titration) [16] and anion exchange capacity, AEC (Column operation using 0.1 M NaNO3) measurements [17].
2.3. Protein Adsorption-Desorption Experiments

Batch experiments were performed by shaking 0.1 g of adsorbent with 50 mL of BSA solution in 100 mL stoppered conical flasks at constant temperature (30 C) in a water bath shaker (200 rpm) for 3 h. The concentration of BSA was measured at 280 nm by using a double beam UV/Vis spectrophotometer (JASCO UV/VIS-V-530). The adsorption capacities were calculated from the difference between the final concentration and BSA solution of the same concentration without any adsorbent left in the water bath after the same period of time (blank experiments). The pH dependent adsorption of BSA was studied at various pHs ranging from 2.0 to 8.0, in either acetate (50 mM pH 4.05.5), in phosphate buffer (50 mM, pH 6.06.5) or in TrisHCl buffer (50 mM, pH 7.08.0). The sorption kinetics was performed at the optimum pH with four different BSA concentrations ranging from 100 to 200 mg/L at 30 C. Samples were taken at appropriate time intervals, centrifuged, and analyzed to obtain the corresponding BSA concentration in supernatant solution. The effects of adsorbent dose, and temperature on the sorption process were also studied. The contact time was varied from 3 to 240 min, the temperature from 10 to 40 C and the amount of adsorbent from 1 to 10 g/L. To investigate the possibility of recycle of the PGMA-g-Cell-SO3H, desorption and regeneration experiments were conducted with different types of desorbing agents through batch technique. The BSA adsorbed onto PGMA-g-Cell-SO3H was placed in the desorption medium while stirring at 200 rpm at 30 C for 3 h. The elution ratio was calculated from the amount of BSA adsorbed on the PGMA-g-Cell-SO3H and the amount of BSA desorbed. The adsorptiondesorption cycle was repeated four times to determine the reusability of the adsorbent.
3. RESULTS AND DISCUSSION

3.1. Preparation and Adsorbent Characterization
Scheme 1 represents the functional reaction involved in the adsorbent preparation. The %G and %GE were found to be 29.8 and 112.3 %, respectively. There was remarkable weight gain of cellulose, after graft copolymerization of GMA/ EGDMA. The amount of surface functional epoxy group content in GMA-Cell was determined to be 1.44 mmol/g, which indicates that epoxy groups were introduced into Cell successfully. The resultant density of sulfonic acid group was found to b 1.23 mmol/g.
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The BET surface area of PGMA-g-Cell-SO3H and Cell was determined from the N2 adsorption data and the values were found to 47.3 and 18.2 m2/g, respectively. It was found that PGMA-g-Cell-SO3H has surface area 2.6 times than Cell after chemical modification. The cation exchange capacity and apparent density were found to be 1.32 meq/g and 1.15 g/L for PGMA-g-Cell-SO3H and 0.71 meq/g and 0.83 g/L for Cell, respectively. The values indicate that PGMA-g-Cell-SO3H is more capable of adsorbing protein than Cell. The values of pHpzc determined using potentiometric titration of PGMA-g-Cell-SO3H and Cell were found to be 3.5 and 5.0 respectively. The low pHpzc of PGMA-g-Cell-SO3H indicates that the surface became more negative due to chemical treatment and this increases the extent of BSA adsorption onto PGMA-g-Cell-SO3H.
3245 1658 D
1055 650
1539
3329 C
Transmittance, %
1539 1393 1658
3348
1246
650 1055
B 3329
2997 758 846 1270 906
A 2900 3211 1028 670
1790
4000 3400 2800 2200 1600 1000

Wave number, cm
-1
400
Figure 1. FTIR spectra of Cell (A), GMA-Cell (B) PGMA-g-Cell-SO3H. (C), BSA (D) and BSAPGMA-g-Cell-SO3H. (E)
158
The FTIR spectra of Cell, GMA-Cell, PGMA-g-Cell-SO3H, BSA and BSA-PGMA-gCell-SO3H are presented in Fig. 1. In the spectrum of Cell the broad peak at 3211 cm-1 could be related to hydrogen bonded O-H stretching vibration and the peaks at 2900 and 1028 cm-1 could be attributed to the C-H stretching and the C-H bending from the CH2 group. The peaks at 1790 and 670 cm-1 are attributed due to the C=O stretching of hemicellulose and glycosidic linkages in Cell, respectively. The FTIR spectrum of GMA-Cell maintained the Cell profile having the ester and epoxide group signals around 2997 cm-1 (C-H stretching typical of the epoxide ring) and 1270 and 846 cm-1(COC stretching typical of the epoxide ring). Absorption bands of epoxy groups at 758 and 906 cm-1 corresponds to terminal oxiranes groups derived from GMA [18]. In addition involvement of hydroxyl group for GMA grafting was conformed by the observed shifting of stretching frequency corresponding to the 3211 cm-1 to 3329 cm-1 in GMA-Cell (Fig. 1B). The appearances of new peaks in the spectrum of GMA-Cell gave supporting evidence for the grafting reaction and suggest that GMA had been successfully grafted onto Cell. The characteristic absorption bands of epoxy rings at 1270, 846, 906 and 758 cm-1 were not shown in the spectrum of PGMA-g-Cell-SO3H. In addition, the band observed at 3329 cm-1 in the spectrum of GMA-Cell shifted to 3348 cm1 and has become broad with the IR spectrum of PGMA-g-Cell-SO3H, which correspond to OH stretching vibration.
10
20
30
40
50
60
70
80
Figure 2. XRD patterns of Cell (A), PGMA-g-Cell-SO3H. (B) BSA (C) and BSA-PGMA-g-Cell-SO3H. (D)
The characteristic absorption bands of the sulfonate group were also observed at 1246, 1055 and 650 cm-1, respectively (Fig. 1C). The presence of these characteristic bands
159
confirms the grafting with GMA and chemical modification of the GMA-Cell with sulfonic acid [19]. The main spectral features of the BSA solid with KBr discs that are related to this discussion are shown in Fig. 1(C). The protein amide 1 (C=Ostretching) was observed as a strong band at 1658 cm-1, while amide II (N-H bending and C-N stretching) appeared at 1537 cm-1. Bands at 1451, 1393 cm-1corresponds to (CH2) mode and stretching of COO- groups. The presence of the amide I absorption at 1658 cm-1 as strong feature of the spectrum is indicative of the BSA molecule being mainly in the -helix conformation [20]. In the FTIR spectra of BSA-PGMA-g-Cell-SO3H, the characteristic peak corresponding to -helix conformation (1651 cm-1) and amide II frequency (1539 cm-1) are present which suggests that there is no conformational change in BSA even after adsorption.
A
Figure 3. SEM micrographs of Cell (A), PGMA-g-Cell-SO3H. (B) and BSA-PGMA-g-Cell-SO3H. (C)
160
Fig. 2 presents the XRD patterns of Cell, PGMA-g-Cell-SO3H, BSA and BSA-PGMA-gCell-SO3H. The X-ray analysis of Cell reveals that the diffraction maxima at 2 = 21.6 , 25.1 and 36.5 are narrow and distinct, corresponds to crystalline domain of cellulose structure. After graft copolymerization there is considerable decrease in scattering angle from 21.6 to 20.16 , 25.1 to 22.6 and 36.5 to 34.6 [21]. Also it was noticed that there is decrease in reflexion sharpness. This indicates that some rearrangement in the morphology of Cell chain occurs as a result of graft copolymerization and also a significant decrease in crystallinity upon incorporation of amorphous copolymer. BSA-PGMA-g-Cell-SO3H has XRD output signal at 34.2 in addition to the BSA phase. Also the scattering angle in PGMA-g-CellSO3H at 20.16 disappeared. This suggests that on adsorption of amorphous BSA, the crystallinity of BSA-PGMA-g-Cell-SO3H decreased. Fig. 3 shows the electron micrographs of Cell, PGMA-g-Cell-SO3H and BSA-PGMA-gCell-SO3H. The surface morphology of Cell was different from PGMA-g-Cell-SO3H. SEM micrograph of Cell appears as corn flake like appearance revealing its extremely fine platy structure. After graft copolymerization, PGMA-g-Cell-SO3H has become more porous and fluffy. The porous surface property of the PGMA-g-Cell-SO3H would favor higher adsorption capacity for the protein due to increase in the surface area. The BET surface area of PGMAg-Cell-SO3H is larger than that of Cell, which implies that PGMA-g-Cell-SO3H has higher adsorption capacity than Cell, which agrees with the observation of SEM images. The SEM image obtained after the sorption of BSA has smooth appearance which may be due to the protrusion of BSA molecules to the surface of PGMA-g-Cell-SO3H indicating that the surface of PGMA-g-Cell-SO3H was covered with BSA.
120 100
Adsorption (%)
80 60 40 20 0 0 2 4 6
PGMA-g-Cell-SO H Cell
Initial concentration : 100 mg/L pH : 4.5 Equilibrium time :3h

8 10 12
Adsorbent dose (g L-1 ) Figure 4. Effect of adsorbent dose on the adsorption of BSA from aqueous solution by PGMA-g-CellSO3H.and Cell.
3.2. Effect of Surface Modification

The effect of surface modification on BSA adsorption was studied by conducting batch experiments using an initial concentration of 100 mg/L with varying adsorbent doses of
161
PGMA-g-Cell-SO3H and Cell. Fig. 4 shows that the percentage of adsorption increased with increase in adsorbent doses and for the quantitative removal of 100 mg BSA from 1000 ml aqueous solution, a minimum adsorbent dosage of 2 g PGMA-g-Cell-SO3H or 9 g Cell was required. The increase in removal percentage with doses may be due to the availability of more adsorption sites at high doses. The results clearly show that PGMA-g-Cell-SO3H is 4.5 times more effective than Cell for BSA adsorption from aqueous solutions. The high percentage removal obtained for PGMA-g-Cell-SO3H may be due to the introduction of SO3H group on the PGMA-g-Cell-SO3H surface through chemical treatment. Since PGMA-gCell-SO3H possess high adsorption capacity relative to Cell, further batch studies were done only with PGMA-g-Cell-SO3H.
3.3. Effect of pH on Adsorption Capacity

The adsorption capacity of BSA on PGMA-g-Cell-SO3H is significantly influenced by pH, as shown in Fig. 5. It is evident that the adsorption capacity increases with the increase of pH and it reaches a maximum value around pH 4.5. The isoelectric pH of BSA is 4.7. So at pH 4.5, BSA has little net electrical charge. It has been observed that proteins have no net charge at their isoelectric points, and therefore the maximum adsorption from aqueous solutions is usually observed at their isoelectric points. This may be due to the minimum electrostatic repulsion between the adsorbed molecules and the molecules to be adsorbed [22]. Moreover, proteins at pH values near their isoelectric point also tend to have a higher structural stability. This will result in minimum intermolecular repulsion and therefore occupy smaller space on the surface of the adsorbent [23]. The maximum removal percentage of BSA was found to be 99.9 and 96.7% at an initial concentration of 100 and 150 mg/L, respectively, the results confirm the previous suggestions. An attractive electrostatic interaction between PGMA-g-Cell-SO3H and BSA would favor the initial attachment of proteins on the surfaces of the adsorbent, however once the adsorbent surface were covered with some proteins, there would be interaction between the already adsorbed proteins and the arriving proteins to be adsorbed from the solution. Since the same type of proteins carry the same type of electric charges, the repulsive electrostatic interaction between the absorbed proteins and the proteins to be adsorbed in the solution would occur, this would hinder the further adsorption of the proteins from the solution on to the adsorbent [24]. At pH values lower and higher than pH 4.5, the adsorbed amount of BSA drastically decreased. The decrease in the BSA adsorption capacity below and above pH 4.5 can be attributed to increase in the conformational size and also due to the electrostatic repulsion effects between the oppositively charged groups. Moreover, the net charge would induce expanded conformation of the BSA molecule, which resulted in the more occupancy areas of the single BSA molecule on PGMA-g-Cell-SO3H [25]. The electrostatic interaction between BSA and PGMA-g-Cell-SO3H above pH 4.5 is highly repulsive due to the same type and high values of surface charges (negative) on them. The experimental results indicate that the BSA molecules which exist as P-NH3+ species in aqueous medium may be exchanged with H+ ion from the SO3H groups from the adsorbent surface.
PGMA- g - Cell SO3 H P NH 3 PGMA- g - Cell SO3 NH 3 P H
(5)
162
In view of ion exchange between sorbent and sorbate system, it was decided to maintain the pH at 4.5 for further experiments.
120 100
Removal (%)
80 60 40 20 0 0 2 4 pH 6
50 mg/L 100 mg/L
Sorbent dose : 2g/L Temperature :30 C

8
Figure 5. Effect of pH on the adsorption of BSA onto PGMA-g-Cell-SO3H.
3.4. Kinetic Model

The effect of contact time on the adsorption of BSA on PGMA-g-Cell-SO3H was investigated by batch adsorption experiments for different initial concentrations. The results are shown in Fig. 6. BSA adsorption increases sharply at a short contact time and slow down gradually with approaching equilibrium. Equilibrium was attained at 3 h. It can be shown that equilibrium time is quite independent of initial BSA concentration. An increase of BSA initial concentration accelerates the diffusion of BSA molecule from solution to adsorbent due to the increase in the driving force of the concentration gradient. Hence, the amount of adsorbed BSA at equilibrium increased with the increase of initial BSA concentration. Inorder to investigate the mechanism of BSA adsorption onto PGMA-g-Cell-SO3H, the rate constant of adsorption is determined from the Lagergren pseudo-first-order kinetic expression [26]:
qt q e [1 e ( k1t ) ]
(6)
where qe and qt are the amounts of BSA adsorbed (mg/g) at equilibrium and at time t (min), respectively, and k1 the rate constant of adsorption (min1). Values of k1 were calculated by non-linear regression analysis for different concentrations and temperatures of BSA. Model parameters including kinetic constants, maximum uptake capacities and the correlation coefficients are presented in Table 1. The values of correlation coefficients were lower than 0.99 and also the experimental qe values do not agree with the calculated ones, obtained from
163
the linear plots (Table 1). This shows that the adsorption of BSA onto PGMA-g-Cell-SO3H is not a first-order reaction.
180 160 140
Pseudo-second- order Adsorbent dose: 2 g/L pH : 4.5 Temperature : 30C
Amount adsobed (mg/g)
120 100 80 60 40 20 0 0
Experimental 100 mg/L 150 mg/L 200 mg/L 250 mg/L
50
100
150 Time (min)
200
250
300
Figure 6. Amount adsorbed versus time plots for the adsorption of BSA onto PGMA-g-Cell-SO3H
Table 1. Kinetic parameters for adsorption onto PGMA-g-Cell-SO3H at different temperatures and initial BSA concentrations
Con: (mg/L) Pseudo-first- order Pseudo-second-order External mass transfer diffusion Coefficient BL (m2/ s) 8.7x10-4 4.9x10-4 3.1x10-4 3.0x10-4 R2 0.991 0.992 0.991 0.993 Intraparticle mass transfer diffusion coefficient Di (cm/s) 3.8x10-15 3.0x10-15 2.3x10-15 2.2x10-15 R2 0.998 0.999 0.997 0.998
100 150 200 250
qe (exp) k1 qe (mg/g) (min-1) (mg/g) 49.5 2.8x10-1 46.8 72.0 2.0x10-1 69.0 96.2 1.5x10-1 67.0 116.0 1.3x10-1 64.2
R2 0.901 0.929 0.935 0.956
2 63.1 41.2 26.4 14.9
qe k2 (mg/g) -1 (min ) 4.7x10-3 49.7 3.2x10-3 73.8 2.4x10-3 96.8 2.2x10-3 116.9
R2 0.998 0.999 0.997 0.998
2 0.4 0.4 0.4 0.9
Temperature (C) Initial concentration: 100 mg/L 10 20 30 40 35.1 40.3 49.5 55.1 0.9x10-1 1.3x10-1 2.8x10-1 3.9x10-1 33.8 38.0 46.8 52.3 0.921 0.923 0.935 0.965 51.1 44.2 26.4 18.9 3.5x10-3 3.0x10-3 4.7x10-3 7.2x10-3 36.1 40.9 49.7 54.9 0.999 0.998 0.997 0.998 0.5 0.4 0.3 0.5 4.5x10-4 6.6x10-4 8.7x10-4 9.9x10-4 0.992 0.991 0.995 0.992 1.8x10-15 3.0x10-15 3.8x10-15 4.2x10-15 0.997 0.998 0.999 0.998
The pseudo second-order kinetic model is expressed as [27]:
k q t qt 2 e 1 k 2 qe t
(7)
164
where k2 is the pseudo-second-order rate constant of adsorption (g/mg/min). The theoretical qeq values estimated from the pseudo-second-order kinetic model gave almost similar values compared to experimental values. The values of correlation coefficients are greater than 0.998 indicating the applicability of the pseudo-second-order equation for the adsorption process of BSA onto PGMA-g-Cell-SO3H.
3.5. Mass Transfer Analysis

It may be considered that usually there are three steps involved in protein adsorption from a bulk solution onto a solid adsorbent, all of which can be considered to offer resistance to protein uptake. These steps include (1) mass transfer from the bulk liquid to the outer surface of the particles (external mass transfer resistance) (2) movement of the adsorbate into the pores of adsorbent by diffusion (internal mass transfer resistance), and (3) protein binding to n internal site. Step (3) is assumed to be rapid with respect to (1) and (2) and is neglected in kinetic analysis. To predict the kinetics of the adsorption process, researchers have assumed that either of the mass-transfer resistances outlined in steps (1) and (2) may be ratecontrolling, acting either individually or in combination [28]. In order to quantify the changes in adsorption with time and also to evaluate kinetic parameters, we use two mass transfer diffusion models, namely, external mass transfer diffusion and intraparticle mass transfer diffusion models. The external mass transfer analysis of BSA during the adsorption process onto PGMA-g-Cell-SO3H was studied using the kinetic model developed by McKay et al [29]. ln
Ct 1 = ln C0 (1 mK L )
mK L (1 mK L ) BL S S t (1 mK L ) mK L
(8)
where Ct is the concentration of adsorbate at time t, Co is the initial concentration of the adsorbate, m is the mass of the adsorbent per unit volume of particle free adsorbate solution, KL is the Langmuir constant (Obtained by multiplying the Langmuir constants, Qo and bL), BL is the mass transfer coefficient (cm/s). Ss is the specific surface per unit volume of particle free slurry and is calculated as:
SS
6m d P P (1 P )
(9)
where dp is the particle diameter (cm), p is the density of the adsorbent (g/mL), and p is the porosity of adsorbent particles. The values of the mass-transfer coefficient (BL) were determined for different concentrations and temperatures from the slopes and intercepts of the plots of ln
Ct (1 mK L ) versus t. C0
Due to the porous nature of the adsorbent, intraparticle mass-transfer diffusion model developed by Urano and Tachikawa has been utilized [30]. In this model the sorption rate is
165
considered to be independent of the stirring speed and external diffusion to be negligible relative to the low overall sorption rate. The Urano Tachikawa model is given by the following equation:
q q f t log 1 t qe qe
4 2 D t i 2.303d 2
(10)
where qt and qe are the BSA concentrations in the solid at time t and at equilibrium, respectively, d is the particle diameter and Di is the diffusion coefficient in the solid (cm2/s). The validity of this model was checked using the linear plots of log[1-( qt/ qe)2] versus t (Fig 7). Linearization was carried out using the initial time of contact between 0 and 90 min. According to the earlier workers, for pore diffusion to be rate limiting, the diffusion coefficient should be in the range 10-15-10-17 m2/s [31].
0 -0.4 -0.8 -1.2 -1.6 -2 -2.4 -2.8 -3.2 -3.6 -4

0 -4.4 -0.4 -0.8
100 mg/L 150 mg/L 200 mg/L 250 mg/L Linear (250 mg/L)
log[1-(qt/qe)2 ]
pH : 4.5 Adsorbent dose : 2 g/L Temperature : 30 0C
log[1-(qt/qe)2 ]
-1.2 -1.6 -2 -2.4 -2.8 -3.2 0
10 20 30 40
C C C C
Initial Concentration : 100 mg/L Adsorbent dose : 2 g/L pH : 4.5

20 40 60 80
Time (min)
Figure 7. UranoTachikawa intraparticle mass transfer plots for the adsorption of BSA onto PGMA-gCell-SO3H at different initial concentrations (A) and temperatures (B)
166
The kinetic parameters obtained from McKay and Urano Tachikawa model are shown in Table 1. It was observed that the BL values are affected by the initial BSA concentration. They gradually decreased, which has been the reason for the decreasing trend of percentage of BSA adsorption by PGMA-g-Cell-SO3H with a gradual increase of BSA initial concentration. Also it can be seen that the BL values increase with increase in temperature, suggesting the process is endothermic. The values of correlation coefficient were lower than 0.997. This can be explained due to the fact that the film diffusion did not dominantly control the overall adsorption process. The values of Di presented in Table 1 showed that the values of Di increased with increasing initial BSA concentrations. With the increase in BSA concentration in the solution, the diffusion of BSA molecules into the boundary layer increases and enhances the diffusion into the solid. The increase in the Di values with temperature may be due to enhanced rate of intraparticle diffusion of BSA molecules. This suggests that a large number of BSA molecules acquire sufficient energy to undergo an interaction with active sites at the surface. The diffusion coefficients are in the order of 10-15 m2/s, which suggests that rate limiting step appears to be pore diffusion.
3.6. Isotherm Analysis

The equilibrium adsorption isotherm is of immense importance in the design of adsorption systems. The isotherm shape can also provide qualitative information on the nature of the solutesurface interaction. In addition, adsorption isotherms are developed to evaluate the capacity of the adsorbent for the adsorption of a particular protein molecule. There are several isotherm equations available for analyzing experimental adsorption isotherm data. Equilibrium data for BSA adsorption was modeled using Langmuir, Freundlich and Sips isotherms. The Langmuir isotherm assumes monolayer coverage of adsorbate over a homogeneous adsorbent surface, and the adsorption of each molecule on the surface has equal adsorption activation energy. The well known Freundlich isotherm used for isothermal adsorption is a special case for heterogeneous surface energy in which the energy term in the Langmuir equation varies as a function of surface coverage strictly due to variation of the sorption. The Freundlich isotherm (Eq. (12)) is more flexible and assumes that the energy of adsorption decreases logarithmically as the fractional coverage increases. However, the Freundlich isotherm does not tend to a limiting coverage as the concentration tends to infinity. Additionally, Sips proposed a model that eludes this limitation of Langmuir and Freundlich models. The Sips or LangmuirFreundlich (LF) isotherm is derived from the Langmuir and Freundlich models. At low sorbate concentrations it effectively reduces to a Freundlich isotherm, while at high sorbate concentrations it predicts a monolayer adsorption capacity characteristic of the Langmuir isotherm. The Langmuir, Freundlich and Sips models can be expressed as follows [32-34]: Langmuir model: q e
q m bC e 1 bC e
1 / nF
(11)
Freundlich model: qe K F Ce
(12)
Adsorptive Characteristics of Bovine Serum Albumin Sips model: qe
167
bs q s C e
1 / nS 1 / nS
1 bs C e
(13)
where qm and b are Langmuir adsorption constants related to adsorption capacity and binding energy of adsorption of BSA onto PGMA-g-Cell-SO3H, respectively. KF and nF are Freundlichs constants related to adsorption capacity and the heterogeneity factor, respectively. qs, bs and 1/nS are Sips maximum adsorption capacity, equilibrium constant related to adsorption capacity and surface heterogeneity, respectively. For a highly heterogeneous system, the deviation of 1/n value from unity will be higher.
140 120 100 80 60 40 20 10 C Langmuir Freundlich Sips
qe, mg/g
Sorbent dose : 2 g/L Initial pH :4.5 Agitation time :2 h
140 0 120 100
qe, mg/g
80 60 40 20 0 140 120 100 80 60 40 20

160 0 140 120
20 C Langmuir Freundlich Sips
qe, mg/g
30 C Langmuir Freundlich Sips
q e,mg/g
100 80 60 40 20 0 0 100 200 300 400 40 C Langmuir Freundlich sips
Ce, mg/L
Figure 8. Comparison of isotherm data for the adsorption of BSA onto PGMA-g-Cell-SO3H
168
Fig. 8 shows the data fittings to different isotherm models of BSA adsorption onto PGMA-g-Cell-SO3H together with the experimental points. It was observed that the best results were obtained when the Langmuir isothem model was used. This is also corroborated by the values of 2 and R2, because the lower the value of 2 and the closer to unity the value of R2, the better the fit (Table 2). The applicability of the isotherm models for the present experimental data approximately follows the order: Langmuir> Sips > Freundlich. The results obtained with the Langmuir isotherm show that an increase in temperature within the range studied increased the maximum adsorption capacity and increased the value of the Langmuir constant. This indicates that the adsorption capacity and the affinity between the active sites and BSA increase with increasing temperature. The value of b, relating to the binding energy, also increases as the temperature increases, suggesting the contribution of stronger binding sites at higher temperature conditions. As temperature increases, the weaker binding sites are occupied first and that the binding strength increases with increasing degree of site occupation [35]. The Langmuir adsorption model assumes that the molecules are adsorbed at a fixed number of well-defined sites, each of which can only hold a single molecule. All adsorption sites have the same affinity for adsorbate, and there are no lateral interactions between molecules adsorbed to adjacent sites [24]. The Langmuir isotherm model has most widely been used as the simple adsorption model for various adsorbentprotein systems [36], even though the adsorption mechanism of proteins may not strictly obey the assumptions of the Langmuir model [37]. As seen from the figure an increase in the BSA concentration in the adsorption medium led to an increase in the amount of adsorbed BSA on the PGMA-g-CellSO3H but this relation leveled off at around 350 mg/L BSA in the adsorption medium. This could be explained by saturation of interacting group of PGMA-g-Cell-SO3H with the adsorbed BSA molecules, as a result of which maximum adsorption capacity is reached. The adsorption capacity (qmax) corresponding to all the three models exhibited significant increase with rise in temperature, which is an indication of the endothermic nature of the process. As deduced from data in Table 2, the maximum adsorption capacity of BSA at 30 C on PGMAg-Cell-SO3H is 124.85 mg/g. Table 2. Isotherm parameters for the adsorption of BSA onto PGMA-g-Cell-SO3H
Langmuir
Temp Q ( oC ) (mg/g)
o
b R2 (L/mg) 0.23 0.998 0.32 0.999 0.50 0.998 0.71 0.997
Freundlich 2 KF 1/n R2
2 qmax bs (mg/g) 127.6 130.5 139.7 140.5 0.219 0.269 0.356 0.654
Sips 1/ns
R2
10 20 30 40
126.01 132.57 141.67 155.37
0.2 0.3 0.4 0.3
72.7 78.6 83.4 92.6
0.097 0.095 0.099 0.105
0.776 8.5 0.832 7.2 0.899 6.9 0.905 5.5
0.789 0.882 0.901 0.978
0.921 0.934 0.944 0.951
1.5 2.5 3.5 4.7
The Langmuir parameters given in Table 2 can be used to predict the affinity between the sorbate and sorbent using dimensionless separation factor (RL):
RL
1 1 b C0
(14)
169
where b is the Langmuir isotherm constant relating to binding energy and Co is the initial solute concentration. The values of RL at 30 C were determined and were formed to be 0.0057, 0.0038, 0.0028, 0.0023, 0.0019, 0.0016, 0.0014, 0.0012 and 0.0016 at an initial BSA concentration of 100, 150, 200, 250, 300, 350, 400, 450, 500 and 550 mg/L, respectively. The RL values for the present experimental data fell between 0 and 1, which is indicative of favorable adsorption of BSA onto PGMA-g-Cell-SO3H. These results indicate that the adsorption of BSA is more favourable at higher initial BSA concentration than lower ones. Earlier workers have already been demonstrated that the Langmuir isotherm model gives adequate results for the adsoption of BSA onto different adsorbents. According to Langmuir model, the maximum adsorption capacity obtained for BSA was reported to be 39.49, 48.9 and 68.7 mg/g for adsorption onto modified chitosan [38], polypyrrole-based adsorbents doped with chloride [24] and macroporous poly(glycidylmethacrylatetriallyl-isocyanurate divinylbenzene) matrix [39], respectively. Inorder to determine the theoretical number of stages for the adsorption of BSA from aqueous solution, operational lines are drawn with a slope of V/m (where V is the volume of BSA solution and m is the mass of PGMA-g-Cell-SO3H). The operational lines join Co,qo to Ce,qe at equilibrium and effect of changing of Co with constant mass generates a series of operating lines as shown in Fig. 9. The operating lines having a slope V/m=0.5 were drawn through two BSA initial concentrations, 200 and 300 mg/L. The values of qe obtained from the operating lines (96.2 and 118.1 mg/g) and from the Langmuir isotherm equation (95.4 and 117.3 mg/g) exhibit reasonable correlation for initial concentrations, 200 and 300 mg/L. Also it was found that in both cases, ~ 100% BSA adsorption can be achieved in two stages.
140 120 100 30 C
qe(mg/g)
80 60 40 20 0 0 50 100 150
Sorbent dose : 2 g/ L Initial pH :4.5 Agitation time :3 h
200
250
300
350
Ce (mg/L)
Figure 9. The operational lines along with the theoretical number of stages for the adsorption of BSA onto PGMA-g-Cell-SO3H
3.7. Thermodynamic Studies

In order to explain the effect of temperature on the adsorption thermodynamic parameters, standard free energy (G), standard enthalpy (H), and standard entropy (S) were determined. To calculate the value of the parameters, the following equations were used:
170 G0=-RTln b
T. S. Anirudhan and P. Senan (15)

0
ln b
S H R RT
(16)
where R is the ideal gas constant 8.314 (J/mol/K) and T is temperature (K). The values of ln b were found to be 18.25, 18.43, 18.55 and 18.64 at 10, 20, 30 and 40 C, respectively. Plotting of ln b vs 1/T (vant Hoff plot) gives a straight line with a slope and intercept equal to H/RT and S/R, respectively. The G values for BSA adsorbed on PGMA-g-Cell-SO3H were calculated for each temperature and were found to be -42.94, -44.91, -46.73, -48.51 kcal/mol for 10, 20, 30 and 40 C, respectively. The Gibbs free energy indicates the spontaneity of the adsorption process, where higher negative values reflect a more energetically favorable adsorption process. The negative G values obtained for each temperature in this study confirm the feasibility of the adsorbent and spontaneity of adsorption of BSA onto PGMA-g-Cell-SO3H. Also the increase in the negative Go with temperature suggests that the adsorption is more favorable at high temperatures. Therefore, high temperatures favor the adsorption of BSA onto PGMA-g-Cell-SO3H. The So value for the adsorption of BSA to PGMA-g-Cell-SO3H was found to be 185.52 J/mol/K. Positive value of So indicates an increase in the total disorder of the system during adsorption [40]. The calculated Ho value of the system for the interaction for BSA with PGMA-g-Cell-SO3H was 9.50 kJ/mol indicates that BSA adsorption is endothermic in nature, which is supported by the increase in the adsorption onto PGMA-g-Cell-SO3H with a rise in temperature, as shown in Fig. 7. Isothermal data at four different temperatures were used to estimate the isosteric heat of adsorption process (Hx) and is calculated using ClausiusClapeyron equation [41]
d ln Ce H x dt RT 2
(17)
The plots of ln Ce versus 1/T (Figure not shown) for different amounts of BSA adsorption were found to be linear and the values of Hx were measured from the slopes of the plots. The values of Hx were found to remain almost constant (~ 26.45 kJ/mol) with increase in surface loading from 60.00 to 105.00 mg/g. This indicates that the surface of PGMA-g-Cell-SO3H is energetically more or less homogeneous and the lateral interactions between adsorbed BSA ions do not exist.
3.3. Design of Single Stage Batch Reactor

Scheme 2 represents the schematic diagram of the single stage batch adsorption system designed from the adsorption isotherm data. The solution containing Co (mg/L) of BSA in V(L) of water is reduced to C1 (mg/L) after the adsorption process. During the adsorption process W (g) of PGMA-g-Cell-SO3H is added, and the BSA concentration on PGMA-gCell-SO3H increases from qo (initially) to qe. The mass balance equation of BSA adsorption from the aqueous solution to that loaded on the adsorbent is:
171 (18)
C1 C0 V qt qo W
When fresh adsorbent is used, q0 = 0 and if the system is allowed to reach equilibrium, then Eq. (18) can be expressed as:
C1 C o W V qe
Substituting for qe from Eq. (11) and rearranging gives:
(19)
C Co [1 bCe ] W 1 V Qo bC e
(20)
C Co [1 1.73Ce ] W 1 V 212.24 C e
(21)
This equation can be used to calculate the mass of PGMA-g-Cell-SO3H required to achieve certain percentage removal by treating a definite volume of BSA initial concentration Co. Fig. 10 represents the experimental and theoretical masses of PGMA-g-Cell-SO3H against different volumes for different percentages of removal of BSA and mass of PGMA-g-CellSO3H against different concentrations and different volumes for the removal of BSA (>99.0%) from aqueous solutions. The amounts of adsorbent calculated using the model Eq. (11) match those observed experimentally for different volumes of effluent and concentrations.
W g PGMA-g-Cell-S O 3H
qo mg of BS A in W g of PGMA-g-Cell-S O 3H
V L of aqueous solution Co mg of BS A in 1 L of aqueous solution
V L of aqueous solution C t mg of BS A in 1 L of aqueous solution
qe mg of BS A in W g of PGMA-g-Cell-S O 3H
W g PGMA-g-Cell-S O 3H
Scheme 2. Diagram of a single-stage batch reactor
172
20

Experimental
99.6% 95.60% 88.80% 10 82.70%
Calculated
Mass of adsorbent (g)
Mass of adsorbent (g)
2.5 2 1.5 1 0.5 0 0
Experimental
125 mL 100 mL 75 mL 50 mL
Calculated
A
0 0 5 10 15
B
50 100 150 200 250
Volume of effluent (L)
Co (mg/L)
Figure 10. (A) Mass of PGMA-g-Cell-SO3H against different volumes for different percentages of adsorption of BSA and (B) mass of PGMA-g-Cell-SO3H against different concentrations and different volumes for adsorption of BSA from aqueous solutions.
150
Adsorption/Desorption, %
Initial concentration :100 mg/L pH : 4.5 Equilibrium time : 3 h
Adsorption Desorption
100
50
0 1 2 3 4
No of cycles
Figure 11. Adsorption-desorption cycles of BSA
3.9. Desorption Experiments

The use of an adsorbent in the adsorption of proteins depends not only on the adsorption capacity, but also on how well the exhausted adsorbent can be regenerated and used again. To test repeatedly using the adsorbent and to recover the adsorbed BSA, the recovery and regeneration tests were conducted with different types of desorbing agents (NaCl, CH3COOH, KSCN, sodium acetate buffer and sodium lauryl sulfate) through batch adsorption technique. The percentage desorption of BSA from spent adsorbent was found to be 60.1, 93.1, 79.5, 80.1, and 81.2% for 0.1 M concentration of NaCl, CH3COOH, KSCN, sodium acetate buffer and sodium lauryl sulfate, respectively. CH3COOH was found to be more effective desorbing agent and hence desorption of BSA from PGMA-g-Cell-SO3H was carried out with different concentrations of CH3COOH. The % desorpion was found to be
173
55.4, 69.9, 77.5, 93.1 and 98.2% for 0.025, 0.05, .075, 0.1 and 0.2 M CH3COOH, respectively. The adsorption/desorption cycles were repeated for 4 cycles using the same amount of adsorbent. The results of regeneration study of BSA by CH3COOH solution are shown in Fig. 11. The results indicate that the adsorbent can be effectively reused upon treatment with 0.2 M CH3COOH solution, which may be attributed to the displacement of BSA bound to the adsorbent with H+ ions. After four cycles the adsorption capacity of PGMA-g-Cell-SO3H decreased from 49.1 mg/g (99.9%) to 44.7 mg/g (95.1%), while the recovery of BSA decreased from 98.2 % in the first cycle to 94.1 % in the fourth cycle. These results show that PGMA-g-Cell-SO3H can be repeatedly used in protein adsorption/desorption without much noticeable losses in its initial adsorption capacity.
CONCLUSION
The present investigation showed that the PGMA-g-Cell-SO3H prepared from cellulose was an effective adsorbent for the adsorption of BSA from aqueous solutions. The adsorption behaviour of BSA onto PGMA-g-Cell-SO3H was investigated under various reaction conditions. The pH of the medium has an important effect on the adsorption equilibrium of BSA, and maximum adsorption occurs at pH 4.5. Kinetic studies showed that the equilibrium conditions were achieved within 3 h. The kinetic data were described using pseudo first-order and pseudo second-order equations and pseudo second-order equation was found to explain the kinetics most effectively. It was also found that the intraparticle mass transfer diffusion played an important role in the adsorption. The isotherm data were best modeled by the Langmuir isotherm equation and the maximum monolayer adsorption capacity was found to be 124.85 mg/g at 30 C. Thermodynamic parameters were calculated and the results show that the adsorption process is spontaneous and endothermic in nature. The isosteric heat of adsorption process (Hx) was investigatd using ClausiusClapeyron equation and the values remained almost constant, suggesting that the surface of PGMA-g-Cell-SO3H is energetically more or less homogeneous. More than 98.0% of the adsorbed BSA was desorbed using 0.2 M CH3COOH as the elution agent. Repeated BSA adsorption/elution processes showed that PGMA-g-Cell-SO3H can be used for the separation of BSA in aqueous solutions. Thus PGMA-g-Cell-SO3H seems to provide an adequate approach to adsorb BSA from aqueous solutions.
ACKNOWLEDGMENTS
The authors are thankful to the Department of Chemistry, University of Kerala, Thiruvananthapuram, India, for providing the laboratory facilities.
REFERENCES
[1] Ball, V.; Schaaf, P.; Voegel, J.C. Malmsten, M. (Ed.), Biopolymers at interfaces. In: Surfactant Science Series, Marcel Dekker, New York, 1998; Vol. 75. pp. 453484
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T. S. Anirudhan and P. Senan Andrade, J. D. Protein Adsorption, Plenum, New York, 1985; Vol. 2. pp. 180. Hu, J.; Songjun, L.; Bailing, L. Biochem. Eng. J. 2005, 23, pp. 259263 Suen, S. Y.; Lin, S. Y.; Chiu, H. C. Ind. Eng. Chem. Res. J. 2000, 39, 478-487. Hao, W.; Wang, J.; Li, J. J. Chromatographia. 2004, 60, pp. 449454. Zhou, D.; Zhang, L.; Zhou, J.; Guo, S. Water Res. 2004, 38, pp. 2643-2650. Tanyola, D.; Snmezk, H.; zdural, A.R. Biochem. Eng. J. 2005, 22, pp. 221-228 Liu, C.; Bai, R. J.Memb. Sci. 2005, 267, pp. 68-77 Purohit, G.; Sakthivel, T.; Florence, A.T. J. Pharmaceutics. 2003, 254, pp. 37-41. Zhao, Z.P.; Wang, Z.; Wang, S.C. J. Memb. Sci. 2003, 217, pp. 151-158. Kim, M.; Saito, K.; Furusaki, S.; Sugo, T. J. Membr. Sci. 1993, 85, pp. 2128. Yu, Y.H.; Sun, Y. J. Chromatrogr. A 1999, 855, pp. 129136. Kim, M.; Saito, K.; Sugo, T.; Okamoto, J. J. Membrane Sci. 1991, 56, pp. 289302. Hebeish, A. ; Guthrie, J.T. Polymers - Properties and Applications. Springer-Verlag, Berlin Heidelberg, New York. 1981, 4 Sidney, S. Quantitative Organic Analysis, 3rd ed., John Wiley and Sons, New York, 1967; pp. 124-127 Schwarz, A.; Driscoll, C. T.; Bhanet, A. K. J. Colloid. Interface Sci. 1984, 97(1), pp. 55-61. Shubha, K. P.; Raji, C.; Anirudhan, T. S. Water Res. 2001, 35(1), pp. 300-310. Alberti, A.; Bertini, S.; Gastaldi, G.; Iannaccone, N.; Macciantelli, D.; Torri, G.; Vismara, E. Europ. Polym. J. 2005, 41, pp. 17871797 Silverstein, R. M. ; Bassler, G. C.; Terrence, C. M. Spectroscopic identification of organic compounds. John Wiley and Sons, Inc. New York. 1991. Wang, F.; Yang, Z.; Zhou, Y.; Weng, S.; Zhang, L.; Wu, J. J. Molec. Struc. 2006, 794, pp. 111 Anirudhan, T. S. ; Jalajamony, S. ; Suchithra, P. S. Colloids and Surf. A: Physicochem. Eng. Aspects. 2009, 335, pp. 107113 Joon, T. O.; Kim, J. H. Enzyme Microb. Technol. 2000, 27, pp. 356361. Haynes, C. A.; Norde, W. Colloids Surf. B. 1994, 2, pp. 517-566. Zhang, X.; Bai, R.; Tong, Y. W. Sep. Purif. Tech. 2006, 52, pp. 161169 Hu, J.; Li, S.; Liu, B. Biochem. Eng. J. 2005, 23, pp. 259263 Lagergren, S.; Svenska, B. K. Zur theorie der sogenannten adsorption geloester stoffe, Veternskapsakad Handlingar 1898, 24 (4), pp. 139. Ho, Y. S.; Mckay, G. Process Biochem. 1999, 34, pp. 451465. Ruthven, D. M. Principles of Adsorption and Adsorption Processes, Wiley, New York, 1984. McKay, G.; Otterburn, M. S.; Sweeny, A. G. Water Res. 1981, 15, pp. 327-331. Urano, K.; Tachikawa, H. Ind. Eng. Chem. Res. 1991, 30, pp. 1897-1899. Michelson, L. D.; Gideon, P. G.; Pace, E. G.; Kutal, L. H. Bulletin No.74 Office of Water Research and Technology. U.S. Department of Industry. 1975. Langmuir, I. J. Am. Chem. Soc. 1918, 40, pp. 1361-1403. Freundlich, H. M. F. J. Phys. Chem. 1906, 57, pp. 385-470. Sips, R. J. Chem. Phys. 1948, 16, pp. 490-495. Ruzic, I. Marine Chem. 1996, 53, pp. 1-15. Fernandez, M. A.; Carta, G. J. Chromatogr. A. 1996, 746, pp. 169-183. Haynes, C. A.; Norde, W. Colloids Surf. B. 1994, 2, pp. 517-566.
Adsorptive Characteristics of Bovine Serum Albumin [38] [39] [40] [41]
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Fu, G.; Zhao, J.; Yu, H.; Liu, L.; He, B. React. Func. Polym. 2007, 67, pp. 442450 Yu, Y.; Sun, Y. J. Chromatogr. A. 1999, 855, pp. 129136 Finette, G. M. S.; Mao, Q.M.; Hearn, M. T. W. J. Chromatogr. A. 1997, 763, pp. 71-90. Young, D. M.; Crowell, A. D. Physical Adsorption of Gases. Butterworth, London: 1962
Chapter 6
ELECTROCHEMISTRY OF POLYMERIC THIN FILMS PREPARED BY LANGMUIR-BLODGETT TECHNIQUE

*
Paolo Bertoncello
Department of Chemistry, The University of Warwick, United Kingdom
ABSTRACT
The utilization of the Langmuir-Blodgett (LB) technology for the fabrication of engineered supramolecular thin films has received an exceptional development in these last years due to possibility of different applications in materials science ranging from nanotechnology to biosensors. The materials fabricated by LB technology provide an accurate control of the order at the molecular level. The main objective of this chapter is to give an overview of the electrochemical properties of a particular class of polymeric thin films such as conducting polymers and ionomer polymers and describe the potentialities of some recent electrochemical technique for nanotechnological applications mainly scanning electrochemical technique (SECM) and SECM combined to Langmuir trough.
LANGMUIR-BLODGETT TECHNIQUE
This technique originates his name from the pioneeristic works of I. Langmuir and K. Blodgett that, in 1917 and in 1935, respectively, studied the properties of organic surfactants (fatty acids) spread from solution onto the air-water interface of a trough [1, 2]. In particular,
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A version of this chapter also appears in Progress in Electrochemistry Research, edited by Magdalena Nuez, published by Nova Science Publishers, Inc. It was submitted for appropriate modifications in an effort to encourage wider dissemination of research.
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they found that fatty acids having long chains can form a molecular monolayer at the airwater interface, resulting in an overall orientation of the molecules perpendicular to the water surface. By using a barriers system, it is possible to control the area available per molecule. The 2-dimensional pressure exerted on the barriers by the molecules can be measured allowing the pressure-area relationship. By monolayer compression, it is possible to pack the film and transfer it onto a solid substrate. There are different techniques to transfer monolayer: for films having low viscosity, the most used is the Langmuir-Blodgett technique, in which the substrate is simply dipped and raised vertically while maintaining constant the surface pressure (figure 1 A-D). This procedure can be repeated several times resulting in the formation of a multilayers structure. In the standard Langmuir-Blodgett transfer process, it will be built up centrosymmetrical or Y-type films. Other two kind of multilayers structure can be possible: the first one called Z-type, if the film is transferred at the upstroke and X-type if the monolayer is transferred at the downstroke [3, 4].
Figure 1. Schematic of the Langmuir-Blodgett technique: deposition of amphiphilic molecule on the water subphase with a solid substrate (A), monolayer compression (B), monolayer transfer to both side of substrate (C), a multilayers structure is built up by repeating monolayer transfer (D), LangmuirSchaefer method (horizontal lifting) (E); different types of multilayers structures (X,Y and Z). From Ref. [7], Reproduced by permission of the PCCP Owner Societies, Copyright (1999)
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Usually, the vertical dipping technique is the most successful for liquid-like monolayers such as phospholipids or fatty acids having long chains. Instead, if the film is more viscous and there is formation of aggregates or crystallites, the vertical transfer can be difficult: in this case is more preferable a horizontal dipping of the substrate thus leading to a monolayer transfer. This approach is called Langmuir-Schaefer method [5]. This approach were then transferred in the 60 by Kuhn et al to exploit the LB formation of dye containing surfactants and in these last decades in the field of molecular electronics [6]. Currently, in the organic electronics, the focus of the research is to emphasize the need to control the micro and nanoscale structure of molecular architectures [7]. Recently, it has been demonstrate the possibility to fabricate LB films of ionomeric polymers [8-11]. Among perfluorinated ionomers, Nafion (cations exchanger) is still object of study due to the different possible application, ranging from fuel cells to chemically modified electrodes [9-21]. Nafion (Trademark of DuPont de Nemours Co.) (Figure 2) is constituted by a long fluorocarbon chains intercalated with oxygen groups and terminating with a sulfonated group. The sulfonated group is responsible of the partial solubility in water and confers to the polymer good cations exchange properties. Nafion is expected to attain a micellar conformation with the polar sulfonated groups located on the surface of the micelles and the hydrophobic fluorocarbon chains in the inner part [22]. These properties of cations exchange has been utilized to fabricate chemically modified electrodes by incorporation of different molecules having cationic character for the determination of heavy metals in the water and also to preconcentrate biological molecules such as proteins and cytochromes [9-11]. Figure 3 shows the pressure-area isotherms of Nafion obtained in subphases containing different electrolytes.
Figure 2. formula of Nafion
The addition of strong electrolytes in the subphase is a key factor for the obtaining of a stable and reproducible isotherm. In particular, the degree of condensation of the LB film depends on the specific interactions between Nafion and the cations dissolved in the subphase. The degree of condensation follows the sequence Na+>H+>Li+, in agreement to the ion exchange selectivity coefficients [23]. This fact suggests that the factor, which favours the ion exchange incorporation, also helps the aggregation of the interfacial films at the air water interface. Another important factor influencing the interfacial film at the air-water interface is the cationic charge utilized in the subphase as demonstrated recently in the case of the incorporation of TiO2 into Nafion LB films [24]. The electrochemical properties of Nafion LS
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fims have been investigation employing methylviologen as cationic electroactive specie: Figure 4 reports the voltammograms of 40 Nafion LS films after loading in 10-5 M MV2+;
Figure 3. Pressurearea (A) isotherm curves of Nafion at different subphases: (1) H2O, (2) 0.1 M NaCl, (3) 0.1 M NaCl + HCl (pH 2), (4) HCl (pH 2), (5) 0.1 M LiClO4, (6) 0.1 M Ca(NO3)2 ; barrier speed: 1.67 mm s-1. From Ref. [8], Reproduced by permission of the PCCP Owner Societies, Copyright (2002)
Figure 4. CVs of 40 Nafion LS films at different scan rates after loading in 10-5 M MV2+, supporting electrolyte 10-2 M NaNO3. From Ref. [8], Reproduced by permission of the PCCP Owner Societies, Copyright (2002)
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The analysis of the voltammograms indicate a linear dependence of the peak current on the square root of the scan rate, v, between 40 mV s-1 and 200 mV s-1. At scan rate lower than 40 mV -1 and in particular between 5 and 20 mV -1 the peak current depends directly on the scan rate. In this last case the peak separation, Ep is 30 mV, meanwhile in the first case the peak separation is about 60 mV. These data evidence the occurrence of a one-electron reversible reduction and a process diffusion controlled at v>40 mV s-1. The following equation correlates the thickness of the diffusion layer to the scan rate: = (RTDapp/Fv)1/2
4,0
2,0
Current / A
0,0 v (mV s ) 2 5 10 20 50 100

-1
-2,0
-4,0
-6,0 -0,4
-0,3
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Potential / V
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ip (A)
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20
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Figure 5. CVs recorded at 10 layers LB Nafion-cyt c in 0.01 M phosphate buffer (pH 7.0), at different scan rates: 2, 5, 10, 20, 50, 100 mV s-1 (a); Dependence of the anodic peak current on the scan rate (b). Reprinted from Ref. [11], Copyright (2004) with permission from Elsevier.
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Where is the diffusion layer thickness, Dapp is the apparent diffusion coefficient, v the scan rate, R and T the gas constant and the temperature, respectively [18]. When the diffusion layer is smaller than the film thickness (at high scan rate), a semiinfinite diffusion condition takes place. In contrast, a surface (or thin-layer like) process is observed at lower scan rate. It is interesting to note that, in the case of very thin films (10 Nafion LS films) a surface process is observed even at higher scan rate as high as 200 mV s-1. Other results point to a proportionality between the amount of MV2+ (calculated by integrating the peak area of the first reduction peak (figure not shown) with the number of layers deposited and then with the amount of Nafion deposited employing LB procedure. In addition, Nafion LS films evidence permselectivity properties, repelling anions (such as [Fe(CN)6]3-, as expected. LB films of perfluorinated ionomer can be used also for the incorporation of biological molecules such as cytochrome c [11]. Figure 5 shows the voltammograms at different scan rate related to 10 Nafion/cytochrome c LB films. It observes a reduction peak between 14 and 19 mV (at scan rate of 5 and 100 mV, respectively) and an oxidation peak at about -10 mV vs Ag/AgCl. The quasi-reversible process is due to the oxidation of the heme group of the cytochrome c accordingly to the following reaction: [cyt c- Fe(III)] + e- [cyt c-Fe(II)] (2)
The calculated E1/2 (-0.12 mV) is practically constant respect to the scan rate. Interestingly, this value is more negative than the E1/2 of the cytochrome c, related to other similar systems [25-29] but is very similar to the value reported by Sagara et al., in the case of reduction of cytochrome c adsorbed on gold substrates [30]. A linear relation is observed between the reduction peak current and the scan rate, indicating a thin-layer like controlled process. This approach to incorporate biological molecules is interesting because in this way it is possible to increase the electrons exchange with the electrode surface otherwise impossible by using a solution casting approach [31]. These positive results can now open new possibilities and strategies in electroanalysis. A similar situation is observed in the case of Tosflex, an anionic perfluorinated ionomer, trademark of Tosoh Co. (Japan) (see Figure 6): As in the case of Nafion, the addition of strong electrolytes to the subphase changes drastically the shape of the isotherms.
Figure 6. Formula of Tosflex IE-SA 48
In general, the isotherms obtained for Tosflex seem to have a trend quite similar to those observed in the case of Nafion. This agrees with the similarity in the perfluorinated backbone skeleton present both in Tosflex and Nafion structures (see Figure 2 and Figure 6). On the other hand, in contrast, the fact that Tosflex is an anion exchanger while Nafion
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is a cation exchanger reflects in the evidence that, for Tosflex, the successful compression of the interfacial film depends on the nature of the anions added to the subphase. In the case of the isotherms reported in Figure 7, the condensation degree of the interfacial film changes following the sequence Fe(CN)63- > SO42- > COO- > NO3- > Cl- > CH3COO-. Interestingly, this sequence parallels the sequence of the ion-exchange selectivity of anion exchangers [25] so suggesting that the same factor (namely charge/radius ratio) rules both LB films condensation and ion exchange selectivity of Tosflex.
Figure 7. Pressurearea (A) isotherms of Tosflex in different electrolytes as shown in figure. Reprinted from Ref. [10], Copyright (2004) with permission from Elsevier.
Figure 8. CV of 10 Tosflex LB films deposited in subphase containing 10-4 M [Fe(CN)6]3- after transfer in 10-2 M NaCl; scan rates: 5, 10, 20, 50 mV s-1. Reprinted from Ref. [10], Copyright (2004) with permission from Elsevier.
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The preconcentration and ion-exchange characteristics of Tosflex LB films were investigated by using an anionic electroactive specie having electrochemical reversibility such as [Fe(CN)6]3-. The reduction peak at 250 mV is due to the one-electron reduction of [Fe(CN)6]3- (Figure not shown) incorporated in the film. In this case the peak current linearly scales with the square root of the scan rate, indicating a diffusion controlled process. Interestingly, the incorporation of the electroactive specie can be performed directly into the film by dissolution of the anion in the subphase. The related voltammogram recorded after transferring in a solution containing only the supporting electrolyte is reported in Figure 8. The shape of the voltammograms and the linear dependence of the peak current on v (for v= 100 mV s-1) indicates a thin-layer like behavior at relatively high scan. This fact can be ascribed to the higher condensation degree of the Tosflex LB films when Fe(CN)63- is dissolved in the subphase. To note, that the voltammetric signal of Tosflex LB films remains almost constant after several hours of dipping in supporting electrolyte. This fact is interesting for future application in electrocatalysis. An interesting application of the Langmuir-Blodgett technique has been recently proposed by Moretto et al. for another ionomeric polymer, namely Kodak AQ55 incorporating cytochrome c [4]. The pressure-area isotherms of AQ55 (see formula in Figure 9) in different electrolytes are shown in Figure 10.
Figure 9. formula of AQ 55
Even if the AQ55 isotherms are characterized by a different marked phase transitions than those of Nafion and Tosflex, their shapes shows some similarities with the trends observed previously. Curve a shows the isotherm of pure cytochrome c in subphase containing phosphate 0.1 M at pH 7. The isotherm is characterized by a broad trend with a collapse pressure of about 20 mN -1 m . This behavior is very similar to the trend of LB films of cytochrome p450 reported in literature [32]. Curve b shows the isotherm of AQ55: the behavior is quite different respect to those observed for Nafion and Tosflex isotherms. Even if the trend is steeper than the cytochrome c isotherm, the maximum surface pressure is 25 mN m-1, quite lower than the values recorded for Nafion and Tosflex isotherms. Also in this case, the addition of strong electrolytes is a necessary requirement to obtain stable monolayers at the air-water interface. The electrolytes added in the subphase may increase the solubility of the polymer and, at the same time, decrease the electrical repulsions due to the neutralization of the negative charge of the sulfonic groups. The isotherm of the mixed cytochrome c/A55 is reported in curve c; it evidences the typical behavior of mixture of two different compounds having limited miscibility [33]: two different collapse pressures are observed even if an interfacial film is formed between cytochrome c and AQ55. The lower trend observed for the AQ55 isotherms that those of Nafion and Tosflex is in agreement with the water solubility of AQ55 [3].

25
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b
20
Surface pressure / mN m-1
c
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a
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0 200
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600
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Trough area / cm
Figure 10. Pressurearea (A) isotherm of cytochrome c (a), AQ55 (b) and cytochrome c/AQ55 (c). Reprinted from Ref. [11], Copyright (2004) with permission from Elsevier.
Figure 11. Pressurearea (A) and surface potential isotherms of different PS-PMMA ionomers. Reprinted from Ref. [34], Copyright (2004) with permission from American Chemical Society.
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Recently, copolymer LB films of ionomers were utilized for the detection of dopamine [34]. In Particular, they use poly(styrene-co-methyl mathacrylate) (PS-PMMMA) ionomers with different grades of sulfonation. The results evidence the formation of stable Langmuir monolayer in the range between 6-8% of sulfonation. In Figure 11, the pressure-area isotherms and the surface potential curves are reported.
Figure 12. CVs of dopamine in 0.1 M HCl: bare ITO (a), 31 PS-PMMA 3% LB films and ascorbic acid (AA), (b), 31 PS-PMMA 3% LB films I the presence of AA. Reprinted from Ref. [34], Copyright (2004) with permission from American Chemical Society.
The calculated area per molecule for the copolymers in the condensed phase was found to increase when the degree of sulfonation decrease. The values calculated were found to be 18, 21 and 29 2/r.u. for PS-PMMA at 8, 6 and 3%, respectively. Higher percentage of sulfonation evidences material loss to the increase of the water solubility. This fact is
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reflected also in the surface potential behavior curve with the shift of the curve related to PSPMMA 8%. If there is solubility of material, the presence of the ions should decrease the solubility of the ionomer and then to increase the area per molecule accordingly to Osawa [35]. In reality, the presence of the salt in the subphase screens the electrostatic repulsion between the negative charges of the sulfonated groups, leading to a more coiled structure. These results are in agreement with previous report of Bertoncello et al. regarding Nafion LS films [8]. LB films of this copolymer were prepared and tested for the voltammetric determination of dopamine, an important molecule associated with Parkinsons disease [36]. These films prevent the interference of the anions similarly to the Nafion LS films [8]. For this reason the PS-PMMA ITO electrode was tested also in the presence of ascorbic acid, the main interference in this kind of application. In the bare ITO, it observes an oxidation and reduction peak at 0.68 V and -0.33 V, respectively. The peaks are related to the oxidation and then reduction of dopamine to dopaminequinone and viceversa and the participation of two electrons [37]. The addition of dopamine in the PS-PMMA electrode causes the appearance of the peaks at 0.31 V and -0.38 V. The oxidation peak is shifted to lower potential (about 0.3 V) evidencing that PS-PPMA acts as electrocatalytic agent for dopamine oxidation. No peaks are observed when 31 PSPMMA LB films were immersed in AA solution. When dopamine is added, it observes a shift of the peak potential due to the interaction of dopamine with ascorbic acid but maintaining the permselectivity respect ot the ascorbic acid. Recently, Ferreira et al. [38] fabricated LB films of polyaniline/ruthenium complexes as modified electrodes for dopamine detection. The way proposed allows avoiding the interference of the ascorbic acid. Dopamine is an important molecule of the mammalian nervous central system. Loss of dopamine inside neurons may result also in Parkinsons desease. They use a mixture of PANI and a ruthenium complex, namely mer-[RuCl3(dppb)(py)], were dppb= PPh2 (CH2) 4PPh2 and py= pyridine. In this case, the ruthenium complex acts as modifier electrodes and is used to detect dopamine.
Figure 13. CVs of 21 LB of PANI and PANI/Ruby 10%, scan rate: 0.05 V s -1, supporting electrolyte: 1 M. Reprinted from Ref. [38], Copyright (2004) with permission from Elsevier.
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The redox peak of PANI corresponds to the interconversion between the oxidation states leucoemeraldine to emeraldine. LB films of PANI/Ruby 10% evidences a drop of the peak current due to the decrease of the electroactivity operated by Rubpy. These peaks are attributed to the PANI because, in this range of potential (-0.6 V-0.4 V), rubpy is not electroactive [39]. The peak at about 0.1 V is attributed to quinone-like species [40]. The redox peaks at 0.23 and 0.00 V are associated to the oxidation/reduction of dopamine to dopaminequinone with the gain/loss of two electrons. The fact that the peak appears at lower potential than the bare ITO suggest that PANI and PANI/Rupy 10% are electrocatalysis agents for the oxidation of dopamine. Figure 14 (b) evidences that there is no variation of the peak position with the change of the scan rate. A plot of the peak current with the square root of the scan rate indicates that the electrochemical process is diffusion controlled. Instead, the peak current related to the PANI system at -0.22 V and -0.38 V changes linearly with the scan rate, indicating a thin layer or surface process, characteristic of adsorbed species at the electrode surface [41].
Figure 14. CVs of 21 LB PANI/Ruby 10% electrode; scan rate 0.04 V s-1, supporting electrolyte 1 M HCl, without dopamine (dotted line) and in presence of 4.8 10-4 M dopamine (dashed line) (a); CV as (a) at different scan rates (b);relation between anodic peak current and square root of the scan rate (c). Reprinted from Ref. [38], Copyright (2004) with permission from Elsevier.
The PANI/Rupy system was then employed to detect dopamine. The calibration curve (figure not shown) evidences a linearity in the range between 10-5 M to 10-3 M and a detection
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limit equal to 4 10-5 M. Further investigations demonstrated that the electrocatalytic effect is observed also in the PANI system: even if the addition of Rupy does not improve the electrocatalytic effect of PANI, the addition of Rupy give some advantages such as transferability and stability of PANI films [38, 39]. The system was tested also to understand the effect of the interferents such as inorganic anions (Cl-, Br-,), and many others [27]. The data evidence the possibility to detect dopamine with a concentration of ascorbic acid equal to three times the concentration of dopamine. This system is comparable to other different system reported in literature [36, 42-43] Another interesting way to immobilize cytochrome c by using LB technique has been recently proposed by Oh et al. [44]. Photosensitive polyimide films on gold substrated were fabricated by using Langmuir-Blodgett technique and then, micro-array pattern were obtained by lithographic technique. Cytochrome c was then immobilized by using decanethiol and mercaptoundecanoic acid. The procedure utilized is summarized in Figure 15.
Figure 15. Immobilization procedure of cytochrome c on polyimide patterned gold substrates. Reprinted from ref. [25], Copyright (2003), with permission from Elsevier
Figure 16 shows the voltammogram of cytochrome on 5 and 10 polyimide LB films patterned on gold substrate. The typical redox peaks of cytochrome c are clearly visible. Interestingly, a higher current is observed with 5 layers polyimide LB films than 10 layers. An interesting application involving the use of the LB technology to fabricate nanoorganized systems constituted by phospholipids incorporating redox molecules has been proposed by Mecheri et al. [45]. They fabricate LB films of a mixed system tetramethylbenzidine (TMB)/dipalmitoylphosphatidic (DPPA) acid. Langmuir monolayers were prepared by spreading TMB e DPPA (dipalmitoylphosphatidic acid): TMB in fact does not form stable LB films, consequently it is necessary to use a system such as DPPA, able to allow stable monolayers at the air-water interface. The isotherm of pure DPPA (figure not shown) evidences a value of the area per molecule of 0.45 nm2. This value is lower than the value obtained in the mixed DPPA-TMB
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system (about 1.10 nm2). This fact is due to the instability of the TMB LB film with consequent destabilization of the interfacial film at the air/water interface. Then, two possible approaches can be used to fabricate LB films: the first one by spreading TMB-DPPA mixture and transfer of the LB films onto gold electrodes, the second one by transfer the DPPA LB films on the gold electrodes followed by immersion of the DPPA gold modified electrodes in TMB solution. The electrochemical data demonstrated a strong dependence from the immobilization procedure. If the first approach allows a stable immobilization of the mediator inside the film but hindering the mobility, the second approach allowed to a higher mobility with consequent better electrochemical performances regarding the electrocatalytic oxidation of NADH. The voltammogram of TMB in solution (figure not shown) evidence two oxidation and reduction peaks, demonstrating that the oxidation of TMB involves two electrons: the first oxidation process generates a semiquinone-imine cation free radical that rapidly convert to a cationic systems [46]. In contrast, the TMB LB films (Figure 12b), shows only one oxidation and reduction peak. Taking into account that in the first step of the oxidation process there is a formation of a charge transfer complex between the reduced form and the oxidized form of TMB, this means that the first step of oxidation does not occur in the LB films. The oxidation process takes place directly at the electrode by formation of the double oxidized species: in fact, the immobilization of TMB prevents the dimer formation.
Figure 16. CVs of cytochrome immolized on patterned gold substrate. Reprinted from ref. [44], Copyright (2003), with permission from Elsevier
This system was then tested for the electrocatalytic study of TMB for NADH. The results evidence that the oxidation peak current of TMB increase in the presence of NADH, meanwhile the reduction peak current decrease, indicating a fast and efficient electrocatalytic effect. In 1999, Ram et al. demonstrated the possibility to fabricated conducting polymers Langmuir-Schaefer films [47]. They fabricate thin films of polyaniline derivatives chemically synthesized such as polyaniline (PANI), poly(o-toluidine) (POT), poly(o-anisidine) (POAS) and poly(o-ethoxyaniline) (PEOA). The doping of the LB monolayer by addition of acid in the subphase is a necessary step for the obtaining thin films highly ordered. Figure 18 reports the pressure-area isotherms in aqueous subphase at pH 1 for PANI, POT, POAS and PEOA.
191
The different shapes of the curves can be ascribed to the substituent in PANI backbones. The molecular area was estimated by using the repeat unit of each PANI derivatives. The values found were 26, 45, 55, 62 2/r.u for PANI, POT, POAS and PEOA, respectively and the cross-area was estimated 20 2 [48, 49]. The shapes of the isotherms change considerably, in particular when in subphase HCl at pH 1 is added. The doping with acids contributes to the stability of the system at the air-water interface by increase of the structure order. The area per molecule estimation is 20, 25, 21, 45 2/r.u for PANI, POT, POAS and PEOA. These variations can be ascribed to the change from emeraldine base to emeraldine salt at pH 1 [48, 49]. The differences observed for PEOA system are due to the ethoxy group pf PEOA. This substituent can promote different rearrangement at the air-water interface. The electrochemistry of such thin films was then investigated by using cyclic voltammetry. The related voltammograms are reported in Figure 19.
Figure 17. CVs of 3 TMB/DPPA LB films (dotted and solid line indicate the two different ways of TMB immobilization). Reprinted from ref. [45], Copyright (2004), with permission from Elsevier
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Figure 18. Pressure-area isotherms of PANI, POT, POAS and PEOA in deionized water (a) and in deionized water at pH 1 (b). Reprinted from ref. [47], Copyright (2004), with permission from Elsevier
The CVs show the redox characteristics of the individual polyaniline. The decrease of the redox potential respect to PANI can be explained in term of different substituent in the PANI derivatives. The shift to more positive potentials of the peak at about 200 mV is associated to the non polar conformations induced by the different substituents of the PANI derivatives. This fact causes the decrease of the conjugation along the polymer backbone and, at the same time, a decrease of the number of polarons/bipolarons states. The changes in the oxidation potentials can be ascribed to the higher electronic density states, which facilitate the protonation and oxidation of the amine groups. Different oxidation states can be detected: from leucoemeraldine to emeraldine, from emeraldine to pernigraniline and protonation from undoped base to doped salt form. LB films of PEOA were also used for the detection of heavy metals by using differential pulse voltammetry [50]. Figure 20 shows the voltammogram evidencing the presence of the peaks due to the different metals. A calibration plot obtained for Pb2+ (not shown, [31]) evidence a linear relation between current and lead concentration, confirming the possibility to use conducting polymers LB films as electrochemical sensors.
193
Figure 19. CVs of 30 PANI derivatives LB films deposited on glass/ITO electrodesin 1 M HCl, scan rate 50 mV s-1: PANI (a), POT (b), POAS (c) and PEOA (d). Reprinted from ref. [47], Copyright (2004), with permission from Elsevier
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Figure 20. Differential pulse voltammetry (DPV) of 50 PEOA LS films containing different metallic cations. Reprinted from ref. [50], Copyright (2001), with permission from Wiley-VCH
Ultrathin films of copolymers LB films based on PANI have been studied by Ram et al. polyaniline and poly(o-anisidine) copolymer was synthesized thin films were fabricated by using LS technique [51]. Figure 21 shows the pressure-area isotherms of PANI/POAS copolymer at different molar compositions in subphase at pH 1. As above mentioned, the doping is an important factor to improve the order of the film at the air-water interface and then the stability of the monolayer [52]. PANI evidences an higher collapse pressure due to the solvent effect (in this case NMP). The effect of the copolymerization is evidenced in curves 3-5: the calculated area per molecule was found 48, 32, 30 2/r.u. Based on these results it appears evident that the area per molecule values decrease with the increase of the ratio of poly(o-anisidine) in the copolymer. This fact can be explained in terms of better orientation of the fraction containing higher percentage of POAS respect to the fraction containing PANI.
Figure 21. Pressure-area isotherms of PANI, POAS and copolymers (PANI/POAS) at different molar composition as indicated in figure. Reprinted from ref. [51], Copyright (2001), with permission from Elsevier
195
Figure 22 reports the voltammograms related to PANI and copolymer PANI/POAS at different molar ratio compositions. A decrease of the redox potentials is observed. This fact is due to the substituents effect of the PANI system. In addition, there is a shift to less positive potential values for the oxidation peaks: this fact can be described in term of decrease of the number of charges states (polarons/bipolarons) due to the increase of the percentage of POAS in the system. The diffusion coefficients for the different copolymer systems were calculated by using the Bufler-Volmer equation [41]: the calculated values decrease from about 10-6 cm2 s-1 for PANI to about 5 10-8 cm2 s-1 for PANI/POAS (25:75). The diffusion coefficient decreases with the increase of the percentage of POAS in the system.
Figure 22. CVs of different polyaniline based systems: PANI (curve 1), PANI/POAS (75:25) copolymer (curve 2) , PANI/POAS (50:50) (curve 3), PANI/POAS (25:75) (curve 4). Reprinted from ref. [51], Copyright (2001), with permission from Elsevier
Figure 23. Pressure-area isotherm of Ru complex in subphase containing 10-5 M Prussian blue. Reprinted with permission from Ref. [53], Copyright (2000) American Chemical Society
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Recently, hybrid materials LB films such as Prussian blue and a surfactant derivative of the ruthenium tris(bipyridine) complex have been fabricated [53]. Figure 23 shows the pressure-area isotherm of Ru(bpy)[bpy(C17)2]22+. The isotherm does not present any plateau, suggesting that no phase transition occurs during the compression of the molecules. It observes a collapse pressure of about 40 mN m-1: this result is in agreement with previous report previously published [54]. Interestingly, the Prussian blue dissolved in the subphase does not modify the compression isotherm: this usually does not happen when positively charged molecules are dissolved in the subphase. Infrared spectra evidence the incorporation of Prussian blue inside the LB film, instead, X-ray diffraction evidence that some parts of the hybrid composite have lamellar structure [53].
Figure 24. Cvs of 8 LB Ru complex incorporating Prussian blue LB films at different scan rates. Reprinted with permission from Ref. [53], Copyright (2000) American Chemical Society
The Cvs show the typical reversible peaks corresponding to oxidation of Ru2+ to Ru3+ at about 1.1 V, the reduction of Prussian blue to Everitts salt (0.2 V) and the oxidation of Everitts salt to Prussian yellow (0.9 V). All these peaks increase with the number of layers (figure not shown), evidencing a homogeneous transfer. As observed in similar lamellar systems, the peak separation and the half-peak width increase with the LB films thickness [55]. This result can be explained in terms of partial hindrance of the diffusion of the charged species by the increasing of the long chain of the surfactant. Both a linear dependence of the peak current with the scan rate and with the square root of the scan rate are observed. These apparent opposite results must be correlated with the large diffusion effect due to the long chain of the surfactant layer [56]. A novel combination Langmuir-Blodgett tecnique with scanning electrochemical microscopy (SECM) has been recently proposed by Unwin et al. [57]. In this case the lateral conductivity of a LB film of polyaniline (PANI) is measured by using SECM. This application is really new and very important due to the importance of the electrically conducting polymers such as PANI in micro and nanoelectronics for applications as light emitting diodes, photovoltaic cells [58, 59]. In this way it has been possible to measure the electrical conductivity in situ avoiding the segregation effects that may happen after the PANI monolayer is transferred to a solid support. It has been possible to detect the electrical
197
transition from insulator to conductor of a PANI monolayer. With this approach, a biased microelectrode generates a flux of electroactive species, which may undergo a redox reaction at the interface (PANI monolayer), the extent of which depends on the surface conductivity. The schematic of SECM configuration is reported in Figure 25
Figure 25. Scanning electrochemical microscopy (SECM) configuration for measurements of the conductivity of a LB PANI monolayer. Reprinted with permission from Ref. [57], Copyright (2003) American Chemical Society
The conductivity of the PANI monolayer is investigated by using SECM in feedback mode in aqueous subphase contained 0.1 mM ferrocene monocarboxylic acid and HCl, to ensure that PANI monolayer is in the protonated (doped) emeraldine state. At low surface pressure it observes a decrease of the steady-state current when the distance between the microelectrode and the monolayer decrease. Instead, at high surface pressure it observes an increase of the steady-state current.
Figure 26. diagram of the charge transfer process occurring in the SECM feedback experiment. E refers to the potential difference at the PANI monolayer induced by the variation of the redox specie concentration in soluiton. Reprinted with permission from Ref. [57], Copyright (2003) American Chemical Society
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The current at the microelectrode is affected by two processes: the hindrance of the ferrocene diffusion while approaching the interface and the regeneration of the electroactive species at the interface. The fact that the feedback current changes from negative to positive as a function of surface pressure is therefore due to an insulator to conductor transition that is caused by compressing the film. In the SECM measurements, the lateral charge propagation in the monolayer is driven by the potential difference, established by variations in the concentration of the redox couple (Nernst relation) in the gap between the tip and the monolayer. The diffusion problem is treated by numerical simulation [57]. In Figure 26 is reported the diagram evidencing the charge transport process occurring in the SECM feedback experiment. The tip current Itip is the sum of two contributions: the component (known) due to hindered diffusion of ferrocene monocarboxylic acid Ihind and the current through the PANI monolayer, Imonolayer, which is deduced. Linear relation is obtained (data not shown) between E and Imonolayer, for each of the approach curves. For each surface pressure is then calculated the conductivity of the film taking into account the cylindrical geometry of the electrode and the thickness of the PANI LB films, determined by AFM and XPS. The results evidence an increase of the conductivity with the increase of the surface pressure, consequently, to obtain efficient lateral propagation the PANI LB film has to be highly compact. A remarkable increase in the conductivity is evidenced, beyond a threshold pressure of about 20 mN m-1, demonstrating that ultrathin 2D conducting polymer films must be highly compact to promote efficient lateral charge propagation.
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[19] Huang, Q.; Lu, Z.; Rusling, J., Langmuir 1996, 12, 5472 [20] Galeska, I.; Chattopadhyay, D.; Moussy, F.; Papadimitrakopoulos, F., Biomacromolecules 2000, 1, 202 [21] Fresnadillo, D.G.; Marazuela, M.D.; Moreno-Bondi, M.C.; Orellana, G., Langmuir 1999, 15, 6451 [22] Yeo R.S.; Yeager H.L., 1985, in Modern Aspects of Electrochemistry, eds. Conway, B.E.; White R.E.; Bockris, J. OM., Plenum Press, New York, 1985, vol. 16, ch. 6 [23] Steck A.; Yeager, H.L., Anal.Chem. 1980, 52, 273 [24] Bertoncello, P.; Notargiacomo, A.; Nicolini, C., Langmuir 2005, 21, 172 [25] Majda, M., in Molecular design of electrode surface, Ed. Murray, R.W., Wiley, New York, 1992, ch.4 [26] Ugo, P.; Zangrando, V.; Moretto, L.M., Brunetti, B., Biosens.&Bioelectron. 2002, 17, 479 [27] Lojou, E.; Luciano, P.; Nitsche, S.; Bianco, P., Electrochim. Acta 1999, 44, 3341 [28] Eddowes, M.J.; Hill, H.A.O., J. Am. Chem. Soc. 1979, 101, 4461 [29] Ugo, P.; Pepe, N.; Moretto, L.M.; Battagliarin, M., J. Electroanal. Chem. 2003, 560, 51 [30] Sagara, T.; Murakami, H.; Igarasho, S.; Sato, H.; Niki, K., Langmuir 1991, 7, 3190 [31] Hahn C.E.W.; Hill, H.A.O.; Ritchie, M.D.; Sear, J.W., J. Chem. Soc. Chem. Comm. 1990, 125 [32] Nicolini, C.; Erokhin, V.; Ghisellini, P.; Paternolli, C.; Ram, M.K.; Sivozhelezov, V., Langmuir 2001, 17, 3719 [33] M.Petty, in Langmuir-Blodgett films an introduction, Cambridge University Press, 1996, Cambridge [34] Carvalho, A.J.F.; Ferreira, M.; Balogh, D.T.; Oliveira O.N.Jr.; Faria, R.m., J. Phys. Chem. B 2004, 108, 7033 [35] Osawa, F., in Polyelectrolytes, M. Dekker, New York, 1971, Ch. 7 [36] Lin, X.-Q.; Zhang, L.; Anal. Lett. 2001, 34, 1685 [37] Zhao, H.; Zhang, Y.; Yuan, Z.; Anal. Chim. Acta 2001, 441, 117 [38] Ferreira, M.; Dinelli, L.R.; Wohnrath, K.; Batista, A.A.; Oliveira O.N.Jr., Thin Solid Films 2004, 446, 301 [39] Ferreira, M.; Wohnrath, K.; Torresi, R.M.; Costantino, C.J.L.; Aroca, R.F.; Oliveira Jr, O.N.; Giacometti, R.A., Langmuir 2002, 18, 540 [40] Mello, S.V.; Mattoso, L.H.C.; Faria, R.M.; Oliveira Jr., O.N., Synth. Met., 1995 71, 2039 [41] Bard, A.J.; Faulkner, L., in Electrochemical Methods: Fundamental and Applications, 2nd edition, J. Wiley, 2001 [42] Ju, H.X.; Ni, J.G.; Gong, Y.; Chen, H.Y.; Leech, D.; Anal. Lett. 1999, 32, 2951 [43] Ciszewski, A.; Milczarek, G.; Anal. Chem. 1999, 71, 1055 [44] Oh, S.Y.; Park, Y.-K.; Ko, C.-B.; Choi, J.-W., Biosens.&Bioelectron. 2003, 19, 103 [45] Mecheri, B.; Piras, L.; Caminati, G., Bioelectrochemistry 2004, in press [46] Josephy, P.D.; Eling, T.; Mason, R.P., J. Biol. Chem. 1982, 257, 3669 [47] Ram, M.K.; Adami, M.; Sartore, M.; Salerno, M.; Paddeu, S.; Nicolini, C., Synth. Met.1999, 100, 249 [48] Ram, M.K.; Sundaresan, N.S.; Malhotra, B.D.; J. Phys. Chem. 1993, 97, 11580 [49] Ram, M.K.; Gowri, R.; Malhotra, B.D.; J. Appl. Polym. Sci. 1997, 13, 141 [50] Ram, M.K.; Bertoncello, P., Nicolini, C., Electroanalysis 2001, 13, 574
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[51] Ram, M.K; Sarkar, N.; Ding, H.; Nicolini, C., Synth. Met. 2001, 123, 197 [52] Ram, M.K.; Carrara, S.; Paddeu, S.; Maccioni, E.; Nicolini, C.; Langmuir 1997, 13, 2760 [53] Torres, G.R.; Dupart, E.; Mingotaud, C.; Ravaine, S., J. Phys. Chem. B 2000, 104, 9487 [54] Valenty, S.J.; Behnken, D.E.; Gaines, G.L., Inorg. Chem. 1979, 18, 2160 [55] Saliba, R.; Agricole, B.; Mingotaud, C.; Ravaine, S., J. Phys. Chem. B 1999, 103, 9712 [56] Aoki, K.; Tokuda, K.; Matsuda, H.; J. Electroanal. Chem. 1984, 160, 33 [57] Zhang, J.; Barker, A.L.; Mandler, D.; Unwin, P.R., J. Am. Chem. Soc. 2003, 125, 9312
INDEX
A
absorption, viii, 51, 56, 57, 69, 70, 71, 85, 86, 87, 88, 89, 92, 93, 94, 95, 96, 97, 98, 99, 114, 137, 140, 158 absorption spectra, viii, 51, 57, 69, 70, 71, 85, 86, 88, 89, 93, 94, 95, 96, 97, 98, 99, 137 absorption spectroscopy, 114 accessibility, 37 accuracy, 17, 54 acetonitrile, 143 acetylcholinesterase, 114, 133 acid, ix, 8, 14, 15, 16, 17, 18, 20, 23, 24, 26, 29, 30, 32, 37, 38, 39, 41, 42, 43, 44, 46, 48, 102, 103, 108, 111, 122, 125, 126, 127, 137, 143, 144, 145, 146, 151, 153, 155, 156, 159, 188, 190, 191, 198 activated carbon, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 28, 35, 39, 41, 42, 43, 44, 45, 46, 47, 48, 49 activation energy, 166 active site, 166, 168 actuality, 2 adhesion, 113, 114, 115, 121 adhesion force, 114, 115 adjustment, 33 adsorption, vii, viii, 1, 2, 3, 4, 5, 6, 7, 8, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 103, 112, 117, 133, 134, 140, 151, 152, 154, 156, 157, 159,160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 172, 173, 174 adsorption isotherms, 13, 16, 34, 166 advantages, 119, 189 AFM, viii, 101, 102, 103, 108, 109, 114, 115, 116, 117, 118, 119, 120, 122, 123, 128, 129, 140, 141, 142, 143, 144, 145, 199 aggregates, 179 aggregation, 52, 140, 179 agriculture, 42 AIBN, 153 aluminium, 2, 23, 37 amines, 133 amino acids, 135, 152 ammonium, 16 amorphous silicon, 130 anodization, 103 antibiotic, 117, 118 antibody, 113, 120, 127 antigen, 120 aqueous solutions, vii, ix, 1, 2, 13, 14, 15, 16, 17, 18, 19, 20, 21, 24, 25, 26, 27, 28, 29, 30, 31, 33, 34, 35, 36, 37, 39, 41, 42, 43, 44, 45, 46, 47, 48, 49, 152, 153, 161, 171, 172, 173 architecture, 52, 102 ascorbic acid, 187, 188, 190 atmospheric pressure, 112 atomic force, viii, 101, 102, 114, 121, 122, 123, 124, 128, 129, 130, 137, 141 atomic force microscope, 122, 123, 129, 130, 141 atoms, 142 attachment, 131, 152, 161
B
bandwidth, 69, 70, 85, 86, 87 barriers, 177 bending, 15, 158 benzene, 57 binding energies, 5 binding energy, 5, 167, 168, 169 biological activity, 113, 145 biomass, 15, 43, 45 biomaterials, vii, 1 biomedical applications, viii, 151 biosensors, vii, viii, ix, 101, 102, 125, 126, 145, 147, 177
202
biotechnology, viii, 101 bleaching, 37, 48 bonds, 2, 113, 142 boundary conditions, 9, 10
Index
composition, viii, 35, 72, 75, 76, 99, 101, 112, 131, 135, 140, 154, 196 compost, 14, 49 compounds, vii, viii, 29, 37, 52, 53, 54, 56, 57, 59, 66, 97, 112, 125, 131, 152, 185 compressibility, 105, 112 compression, 56, 57, 58, 59, 61, 63, 66, 67, 72, 74, 79, 81, 82, 83, 84, 85, 98, 99, 103, 104, 105, 106, 135, 136, 137, 138, 177, 178, 183, 197 condensation, 12, 179, 183, 184 conditioning, 27 conductivity, 198, 199 conductor, 198, 199 configuration, 198 conjugation, 193, 196 contact time, ix, 15, 16, 19, 20, 21, 23, 24, 25, 27, 28, 29, 31, 33, 34, 151, 156, 162 cooling, 54, 155 copolymerization, 195 copolymers, 186, 195, 196 copper, 14, 17, 20, 28, 31, 33, 34, 36, 37, 39, 42, 43, 44, 47, 48, 49 correlation, 8, 14, 24, 35, 39, 69, 78, 162, 164, 166, 169 correlation coefficient, 8, 35, 162, 164, 166 cost, vii, 1, 2, 12, 13, 14, 17, 19, 20, 21, 23, 25, 26, 28, 29, 32, 39, 44, 46, 49, 152 cost effectiveness, 39 covering, 108, 118 crystalline, 160 crystallinity, 160 crystallites, 179 crystals, vii, 51, 52, 53, 54, 57, 59, 60, 61, 66, 69, 71, 78, 95, 98, 143, 145 cycles, 107, 172, 173 cytochrome, 135, 139, 181, 182, 184, 185, 190, 191 cytochrome p450, 185
C
cadmium, 20, 24, 30, 31, 33, 36, 39, 41, 42, 43, 44, 45, 46, 47, 48, 49 calcination temperature, 26, 39 calcium, 2, 24 calibration, 189, 193, 196 capillary, 115 carbohydrate, 122 carbon, vii, 1, 2, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 28, 43, 44, 45, 46, 49 carbonization, 18, 21, 41 carcinogen, 45 casting, 183 catalysis, 142 catalyst, 48 catalytic activity, 142 cation, 2, 22, 30, 32, 34, 35, 37, 39, 157, 183, 191 cattle, 14, 49 CEC, 32, 34, 39 cell membranes, 120 cell surface, 143, 149 cellulose, vii, viii, 151, 152, 153, 156, 160, 173 CH3COOH, ix, 152, 172, 173 chemical bonds, 2 chemical structures, 133 chemisorption, 10, 12, 25, 46 China, 130 chloroform, 54, 69, 70, 72, 84, 87, 88, 103 cholesterol, 108, 110, 124, 143, 145 chromium, 13, 15, 16, 18, 19, 20, 23, 28, 31, 33, 37, 42, 43, 44, 45, 46, 47, 48 chymotrypsin, 127 class, ix, 177 clay minerals, 2, 21, 24, 26, 32 clean air, 138 cleaning, 54 cleavage, 33, 144 clusters, 139 coal, 18 coatings, 29 cobalt, 30, 45, 46, 49 color, iv compensation, 78 competition, 8, 29, 33, 34, 35 competitors, 142 complexity, 29, 152 complications, 113 composite, 197
D
damages, iv decomposition, 37, 94 defects, 108 deformation, 73, 115, 142 dehydration, 15 denaturation, 120, 133 density, 193, 196 deposition, 54, 56, 71, 87, 106, 107, 108, 111, 113, 114, 123, 131, 133, 136, 138, 139, 140, 145, 146, 178 depression, 118 derivatives, 32, 39, 110, 112, 130, 191, 193, 194, 195 desorption, ix, 10, 20, 35, 39, 140, 152, 156, 172
Index
detection, 102, 112, 120, 125, 186, 188, 189, 193, 196 deviation, 75, 167 dielectric constant, 55, 62 diffraction, 141, 160, 197 diffusion, ix, 9, 11, 13, 14, 16, 18, 19, 20, 21, 24, 25, 27, 28, 29, 31, 32, 35, 40, 44, 46, 128, 152, 162, 163, 164, 165, 166, 173, 181, 184, 189, 196, 198, 199 diffusion process, 46 dimethylformamide, 142 dipole moments, 78, 79, 80, 91, 92, 93 direct observation, 65 discs, 159 disorder, 170 displacement, 173 distilled water, 153, 154 distortion, 115 DMF, 142, 143 DNA, 45, 120, 129, 142 DNA damage, 45 domain structure, 102, 138 dopamine, 186, 187, 188, 189 doping, 191, 192, 195 dosage, 16, 17, 25, 28, 161 double bonds, 111 drug delivery, viii, 101, 102, 117, 129 drug interaction, 117, 118 drugs, 117, 135, 147, 152 duality, 139 dyes, vii, 42, 51, 52, 53, 54, 55, 72, 75, 76, 77, 78, 79, 84, 85, 86, 87, 88, 93, 94, 95, 96, 98, 99
203
England, 54 entropy, ix, 11, 12, 26, 30, 32, 41, 152, 169 environmental conditions, 112 enzymatic activity, 125 enzyme immobilization, 127 enzymes, 101, 102, 112, 113, 125, 127, 133, 134, 142, 143, 146 epoxy groups, 153, 154, 155, 156, 158 equilibrium, ix, 2, 3, 5, 7, 8, 9, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 23, 24, 26, 28, 30, 31, 34, 35, 36, 39, 40, 42, 43, 44, 45, 47, 49, 57, 65, 137, 151, 162, 165, 166, 167, 169, 171, 173 erosion, 117 ester, 158 ethanol, 86, 87, 88, 125, 133, 143, 153 evaporation, 135 excitation, 90, 91 exciton, 89, 90, 91, 92 exertion, 104 experimental condition, vii, 1, 15, 35, 133, 137 extinction, 85 extraction, 36
F
fabrication, viii, ix, 54, 101, 132, 133, 135, 177 fatty acids, 52, 103, 110, 133, 135, 137, 152, 177, 178 feedback, 133, 138, 198, 199 fiber, 56 fiber bundles, 56 fibers, 19, 22 film formation, 58, 102, 139 film thickness, 140, 181 films, vii, viii, ix, 51, 52, 53, 54, 56, 57, 58, 59, 60, 61, 62, 65, 69, 70, 71, 72, 73, 74, 75, 77, 78, 79, 86, 87, 88, 89, 90, 93, 94, 95, 96, 97, 98, 99, 101, 102, 103, 106, 108, 110, 112, 113, 114, 116, 117, 119, 120, 122, 123, 124, 125, 126, 127, 129, 130, 131, 132, 134, 135, 136, 137, 138, 139, 143, 145, 146, 147, 148, 177, 178, 179, 180, 181, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200 Finland, 54 fixed rate, 106 fluid, 2, 112, 115, 128 fluorescence, 66, 102, 108, 114, 128 folding intermediates, 142 formamide, 142 formula, 92, 179, 184 fragility, 145 fragments, 113, 127 free energy, 4, 11, 12, 26, 28, 30, 32, 40, 104, 126, 169, 170
E
effluent, 3, 39, 171 effluents, 2, 19 electric charge, 161 electrical conductivity, 198 electrical properties, viii, 124, 131 electrocatalysis, 184, 189 electrochemistry, 192 electrode, 182, 187, 188, 189, 191, 199, 200 electrodes, 125, 132, 179, 188, 190 electrolyte, 25, 30, 105, 180, 184, 188, 189 electrolytes, 179, 183, 184, 185 electron, 124, 131, 137, 142, 143, 144, 156, 160, 181, 184 electron microscopy, 142, 144 electrons, 182, 188, 189, 191 electroplating, 20 endothermic, ix, 14, 18, 20, 23, 24, 25, 27, 30, 31, 33, 35, 36, 37, 152, 166, 168, 170, 173 engineering, 11, 39, 48, 125, 133
204
friction, 114, 116 FTIR, ix, 15, 17, 114, 140, 145, 151, 156, 157, 158 FTIR spectroscopy, 140, 156 fuel, 179 functionalization, 112 fusion, 103, 117, 128 future, 184
Index
immobilization, 41, 125, 126, 135, 144, 191, 192 immobilized enzymes, 132 immunodeficiency, 117 immunoglobulin, 102, 121, 125, 126 immunoglobulins, 113 impregnation, 13, 16 impurities, 143 incidence, 56, 57, 70, 71, 95, 96, 102, 128 incubation time, 117, 118 India, 1, 24, 31, 151, 153, 173 infrared spectroscopy, 37 insulin, 135 integration, 9, 12 intercepts, 164 interface, viii, 51, 52, 56, 57, 59, 62, 63, 69, 72, 78, 79, 81, 82, 83, 84, 85, 97, 98, 101, 103, 104, 105, 106, 108, 110, 112, 113, 114, 121, 123, 128, 132, 133, 134, 135, 137, 138, 144, 145, 177, 179, 185, 190, 192, 195, 198, 199 interference, 8, 66, 68, 80, 187, 188 intermolecular interactions, 52, 76, 135 inversion, 33 ion adsorption, 18, 43, 45, 49 ion-exchange, 2, 21, 27, 28, 33, 34, 183, 184 ionic strength, 15, 23, 24, 26, 30, 31, 32, 33 ions, vii, ix, 1, 2, 8, 11, 12, 13, 14, 17, 18, 19, 20, 21, 23, 24, 25, 26, 27, 29, 30, 31, 32, 33, 35, 36, 37, 38, 39, 41, 42, 43, 44, 45, 47, 48, 49, 152, 170, 173, 186 iron, 2, 34, 37 isotherms, viii, ix, 2, 3, 16, 20, 24, 25, 27, 28, 29, 30, 31, 33, 34, 36, 48, 51, 58, 59, 60, 61, 62, 67, 72, 73, 74, 75, 77, 79, 81, 82, 83, 84, 85, 98, 99, 105, 112, 139, 152, 166, 179, 183, 184, 185, 186, 191, 192, 193, 195
G
gel, 136 Gibbs energy, 27, 41 glow discharge, 125 glucose, 125, 126, 127, 143, 145 glucose oxidase, 125, 127, 143, 145 glutathione, 45 glycerol, 108, 116 gold, 182, 190, 191 GPC, 47 grades, 16, 186 grazing, 102, 128
H
Hamiltonian, 89, 90 heavy metals, vii, 1, 2, 12, 14, 21, 22, 23, 28, 33, 34, 35, 36, 38, 39, 42, 46, 48, 179, 193, 196 height, 115, 116, 118 heme, 181 hemicellulose, 158 heterogeneity, 117, 167 hexane, viii, 51, 53, 58 histidine, 113 HIV, 122 homogeneity, 71, 95, 139 host, 74, 143 hybrid, 6, 125, 197 hydrogen, 2, 35, 142, 152, 158 hydrogen bonds, 142 hydrophilicity, 110 hydrophobic properties, 110 hydrophobicity, 110 hydroxide, 23, 35 hydroxyl, 158 hysteresis, 57, 59, 135
J
Japan, 156, 183
K
KBr, 159 kinetic constants, 162 kinetic equations, 8 kinetic model, vii, 1, 8, 10, 13, 14, 18, 23, 24, 25, 29, 30, 36, 40, 41, 44, 163, 164 kinetic parameters, 164, 166 kinetic studies, 8, 18, 30, 41 kinetics, ix, 2, 8, 10, 11, 13, 14, 15, 18, 19, 20, 22, 25, 27, 28, 29, 30, 32, 34, 35, 39, 42, 43, 44, 46, 47, 49, 102, 121, 152, 164, 173 kinks, 105 Korea, 35
I
ideal, 6, 75, 103, 115, 135, 170 image, 56, 108, 115, 116, 120, 143, 144, 160 images, viii, 16, 51, 56, 65, 66, 67, 68, 79, 81, 82, 83, 84, 85, 98, 102, 109, 114, 115, 116, 117, 118, 119, 122, 124, 126, 129, 138, 144, 145, 160 imitation, 102 immersion, 106, 107, 108, 113, 190
Index
205
L
labeling, 102 leaching, 36 lecithin, 124 lens, 66, 69, 98 life sciences, 117 lifetime, 101, 125 light, 198 light beam, 56, 57 light emitting diode, 198 light emitting diodes, 198 linear dependence, 181, 184, 198 linearity, 189 lipids, viii, 101, 110, 113, 117, 133, 139, 144, 145, 146, 147 liposomes, 114, 128 liquid crystals, vii, 51, 52, 53, 54, 58, 59, 61, 62, 66, 69, 70, 71, 72, 74, 75, 77, 78, 84, 88, 95, 96, 98, 99 liquid interfaces, 123 liquid phase, 57, 65, 109 liquid-air interface, 134 liquids, 3, 45, 130 lithography, 103 luciferase, 127 Luo, 122
M
macromolecules, 113, 130, 133, 147 magnesium, 2, 37 majority, 131 manganese, 26, 31, 33, 43, 49 manipulation, 102 manufacture, 14 manure, 14, 49 Marx, 148 materials science, ix, 177 matrix, 72, 99, 113, 135, 145, 169 maximum sorption, 33, 35, 36 media, 30, 62 membranes, viii, 101, 103, 110, 112, 114, 119, 121, 124, 125, 126, 127, 128, 129, 135 mercury, 28, 42, 46, 47, 49 metals, vii, 1, 2, 20, 25, 32, 33, 34, 35, 36, 37, 39, 152, 179, 193, 196 methanol, 133, 142, 143 micelles, 179 microporous materials, 43 microscope, 56, 66, 98, 130, 156
microscopy, viii, 51, 65, 101, 102, 108, 114, 117, 118, 121, 122, 124, 126, 128, 129, 130, 137, 138, 140, 141, 143, 198 miniaturization, 52 mixing, 23, 75 mobility, 191 model system, 102, 114, 117, 119 modeling, 16 modelling, 45 modification, 14, 24, 26, 37, 39, 47, 118, 125, 157, 159 modulus, 114, 136, 140 moisture, 44 molecular orientation, 97, 119, 144 molecular oxygen, 45 molecular sensors, 123 molecular structure, vii, 51, 53, 54, 59, 69, 72, 75, 93, 94, 98, 99, 102, 126 molecules, viii, 2, 5, 7, 11, 51, 52, 53, 54, 57, 59, 61, 63, 68, 69, 70, 71, 72, 74, 75, 78, 79, 80, 84, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 101, 103, 104, 105, 106, 110, 112, 113, 114, 115, 117, 131, 132, 133, 134, 135, 138, 139, 143, 145, 160, 161, 166, 168, 177, 179, 181, 182, 190, 197 monitoring, 139, 147 monoclonal antibody, 102, 113 monolayer, viii, ix, 4, 6, 7, 14, 15, 17, 19, 24, 25, 26, 27, 28, 30, 32, 51, 54, 55, 56, 57, 58, 61, 62, 63, 65, 66, 69, 72, 74, 75, 78, 79, 81, 82, 83, 84, 85, 87, 95, 97, 98, 99, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 112, 113, 121, 123, 127, 132, 133, 134, 135, 136, 137, 138, 139, 143, 144, 145, 146, 147, 152, 166, 173, 177, 178, 179, 186, 191, 195, 198, 199 monolayers, 178, 185, 190 monomers, 69, 90, 93, 94, 95, 97, 99, 114 Morocco, 37 morphology, 56, 65, 102, 118, 120, 138, 139, 143, 160 multi-component systems, 21 multilayer films, viii, 123, 131, 135 multilayered structure, 143 myoglobin, 135
N
NaCl, 27, 109, 172, 180, 184 NADH, 191 nanodevices, viii, 101 nanoelectronics, 198 nanogravimetry, 137 nanolithography, 122 nanometer, viii, 101, 102, 103, 114, 115, 117, 119, 141, 143
206
Index
phosphatidylcholine, 112, 130 phosphatidylethanolamine, 112, 115, 129 phospholipids, 102, 103, 110, 112, 114, 117, 124, 128, 133, 135, 136, 137, 138, 139, 143, 145, 146, 178, 190 photobleaching, 102, 114 photonics, 52 photovoltaic cells, 198 physical chemistry, 48 physicochemical properties, viii, 101, 103, 106 physics, 52 pith, 18, 45, 46 plasma membrane, 121 plasticity, 28 platinum, 45, 137, 145 PMMA, 186, 187, 188 Poland, 51, 54, 100, 131 polar groups, 78 polarization, 114 polyacrylamide, 145 polyimide, 190 polyimide film, 190 polymer, 125, 136, 152, 179, 184, 185, 193, 196, 199 polymer films, 199 polymerization, 152, 153 polymers, ix, 130, 131, 139, 141, 177, 179, 191, 193, 196, 198 polypeptide, 135 polyvinyl alcohol, 25, 48, 153 porosity, 16, 164 potassium, 2, 13, 16, 19 precipitation, 23, 25, 26, 27, 32, 35 probe, 114, 115, 120, 130, 141 promoter, 108 propagation, 199 proportionality, 181 protein engineering, 133 protein folding, 142 protein structure, 120, 135 protein-protein interactions, 102 proteins, viii, 101, 102, 103, 112, 113, 114, 117, 126, 132, 133, 135, 139, 142, 143, 147, 149, 152, 161, 168, 172, 179 proteolytic enzyme, 143 protons, 25 prototype, 125 pulp, 18, 22, 41, 47 pure water, 56, 105 purification, 30, 31, 54, 143, 153 PVA, 25, 153 pyrolysis, 18
nanometer scale, viii, 101, 102, 103, 114, 117 nanostructures, vii, viii, 52, 101, 102, 103, 113, 116, 122 nanosystems, viii, 101 nanotechnology, ix, 149, 177 NCS, viii, 51, 57, 98 neurons, 188 next generation, 120 nickel, 19, 22, 31, 33, 37, 42, 44, 45, 47, 48, 49 Nigeria, 25, 33 nitrate, 38 nitrogen, 17, 18 nitrogen gas, 18 NMR, 37 noise, 56 nonlinear optics, vii, viii, 53, 131 North Africa, 23, 38 nucleation, 137
O
oil, 27 olive oil, 14 optical microscopy, 102, 108, 116, 124, 130 optical properties, 52 organic compounds, 29, 174 organic solvents, 133, 142, 143 organism, 143 organizing, vii, viii, 131 orientation, 177, 195 oscillation, 140 osmotic pressure, 152 oxidation, 18, 19, 181, 188, 189, 191, 193, 196, 198 oxygen, 2, 179
P
parallel, 56, 70, 71, 92, 95, 96, 114 particle mass, 33 particle size distribution, 24 partition, 29, 102 penicillin, 145 pepsin, 113, 132 peptides, 101, 117, 129 percolation, 117 performance, 12, 14, 16, 19, 20, 26, 46, 112 permission, iv, 109, 115, 116, 118, 119, 178, 180, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199 permit, 2 permittivity, 55 pesticides, 145 phase behavior, 102, 105, 121 phase transitions, 105, 123, 127, 185
Index
207
Q
quartz, viii, 24, 32, 51, 53, 56, 60, 71, 72, 87, 95, 96, 98, 139, 140 quinone, 188
R
radiation, 56 radius, 183 raw materials, 19 reaction rate, 8 reactions, 44, 140 reagents, 153 real time, 102, 119, 130, 140 reality, 186 recognition, 102, 112, 113, 132, 135, 139 recommendations, iv recrystallization, 144 red shift, 92 reduction, 181, 182, 184, 188, 189, 191, 198 reflectivity, 65, 138 refractive index, 140 regeneration, 21, 156, 172, 199 regression, 162 regression analysis, 162 relative size, 116 relaxation, 137, 152 reliability, 147 remediation, 43 replacement, 18, 30 residues, 135 resistance, 57, 135, 164 resolution, 56, 102, 103, 108, 112, 114, 115, 116, 118, 119, 120, 122, 126, 129, 130, 141, 142, 143, 144 response time, 112, 145 reusability, 156 rheology, 146 rice husk, 15 rings, 66, 68, 80, 158 room temperature, 13, 16, 24, 28, 104 roughness, 143 Royal Society, 123 rubber, 20 ruthenium, 188, 197
S
saturation, 3, 168 Saudi Arabia, 23, 38, 41 sawdust, 13, 20, 21, 22, 45 scanning electron microscopy, 140 scarcity, 39
scattering, 114, 128, 160 Schrdinger equation, 90, 91 seed, vii, 1, 16, 22, 46 segregation, 114, 198 selectivity, 19, 24, 31, 179, 183 self-assembly, 132, 133, 147, 148 self-organization, 119 SEM micrographs, 159 sensing, 112, 113, 131, 140, 145, 147 sensitivity, 112, 140 sensitization, 127 sensors, 112, 125, 145, 146, 193, 196 Serbia, 25, 48 serine, 116 serum, vii, 121, 133, 152 serum albumin, vii, 121, 133, 152 shape, vii, 51, 66, 85, 93, 95, 108, 166, 183, 184 shear, 140 signals, 158 silica, 2 silicon, 2, 106, 123, 143, 144 simulation, 199 Singapore, 48 SiO2, 125, 126 skeleton, 183 sodium, ix, 2, 16, 24, 25, 30, 37, 48, 151, 153, 154, 172 sodium hydroxide, 30, 37 solid phase, 106 solid surfaces, 8 solid waste, 20, 46 solubility, 17, 26, 54, 110, 131, 133, 179, 185, 186 solubility in water, 179 solution, 177, 183, 184, 188, 191 solvent, 195 solvents, 131, 142, 143 sorption experiments, 24 sorption isotherms, 22 sorption kinetics, 156 sorption method, 37 sorption process, 36, 37, 44, 156 species, 2, 14, 22, 29, 37, 96, 105, 133, 135, 140, 141, 143, 145, 161, 188, 189, 191, 198, 199 specific surface, 17, 32, 164 spectrophotometer, 56, 57, 156 spectroscopy, 37, 54, 114, 137, 140 speed of response, 131 spontaneity, 26, 170 stabilization, 113 steroids, 133, 152 sterols, 103, 135 streams, 15 stretching, 15, 158
208
stroke, 56, 106 strong interaction, 89 styrene, 186 substitution, 123 sugar beet, 18, 41, 47 sugar industry, 19 sugarcane, 18 sulfuric acid, 13, 15, 16, 20, 24 Sun, 174, 175 surface area, 14, 15, 16, 17, 18, 19, 21, 26, 32, 34, 35, 37, 39, 48, 103, 110, 132, 138, 157, 160 surface energy, 166 surface layer, 134, 144 surface modification, 19, 21, 160 surface structure, 115, 142 surface tension, 54, 104, 105, 132, 135, 137, 146 surfactant, 16, 46, 197, 198 surfactants, 177, 179 survey, vii, 1, 39 suspensions, 37 sustainability, 39 symmetry, 26, 115, 144 synthesis, 18, 153 synthetic polymers, 135
Index
U
ultrastructure, 144 unconditioned, 27 uniform, 4, 5, 44, 57, 65 United Arab Emirates, 25, 38 United Kingdom, 177 urea, 125, 127
V
vacuum, 139, 153 valence, 2 vanadium, 33 vapor, 106 variations, 192, 199 vector, 71, 92 versatility, 119, 141 vesicle, 103, 114, 128 vibration, 15, 158 viruses, 139 viscoelastic properties, 114 viscosity, 178 visualization, 120, 129, 138
T
tactics, 113 talc, 38 temperature dependence, 33 tension, 54, 104, 105 testing, 2, 112 texture, 102 Thailand, 36 thermal analysis, 26 thermal stability, 143 thermodynamic calculations, 25 thermodynamic parameters, 12, 23, 24, 30, 36, 169 thermodynamics, vii, 1, 11, 20, 30, 42, 44, 46, 48 thin film, ix, 177, 181, 191, 192, 195 thin films, vii, viii, ix, 103, 119, 122, 131, 133, 137, 140, 146, 147, 148, 177, 181, 191, 192, 195 third dimension, 66, 104 topology, 130 total internal reflection, 114, 128 toxicity, 39 transducer, 112 transition temperature, 136 transport, 11, 47, 152, 199 tunneling, 137, 140 Turkey, 27, 43
W
waste, vii, 1, 13, 14, 15, 16, 18, 19, 21, 27, 31, 37, 42, 43, 44, 46, 48 wastewater, 2, 8, 12, 15, 17, 19, 20, 23, 27, 28, 29, 31, 35, 39, 41, 42, 43, 46, 47, 48 water, 177, 178, 179, 185, 186, 190, 192, 193, 195 water resources, 2 wavelengths, 69, 92, 93, 96 weight gain, 156 wood, 13, 14, 17, 20, 22, 41, 46, 48 workers, 145, 165, 169
X
XPS, 140, 199 X-ray, 26, 37, 102, 128, 137, 140, 143, 156, 160, 197 X-ray analysis, 160 X-ray diffraction, 26, 37, 102, 137, 140, 156, 197 X-ray photoelectron spectroscopy (XPS), 140 XRD, ix, 43, 151, 158, 160
Z
zeolites, 42 zinc, 14, 17, 28, 30, 31, 33, 36, 37, 39, 41, 42, 43, 46, 47, 48 zirconium, 32

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CHEMICAL ENGINEERING METHODS AND TECHNOLOGY

LANGMUIR MONOLAYERS IN THIN FILM TECHNOLOGY

CHEMICAL ENGINEERING METHODS AND TECHNOLOGY

NANOTECHNOLOGY SCIENCE AND TECHNOLOGY

CHEMICAL ENGINEERING METHODS AND TECHNOLOGY

LANGMUIR MONOLAYERS IN THIN FILM TECHNOLOGY

Nova Science Publishers, Inc.

Copyright 2011 by Nova Science Publishers, Inc.

NOTICE TO THE READER

Published by Nova Science Publishers, Inc. New York

In: Langmuir Monolayers Editors: Jennifer A. Sherwin

ISBN: 978-1-61122-461-0 2011 Nova Science Publishers, Inc.

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

2. ADSORPTION ISOTHERMS, KINETICS AND THERMODYNAMICS

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

If R L >1, the isotherm is unfavorable

RL =1, the isotherm is linear

where P3, P4, P5 and P6 are isotherm constants.

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

q m kCe (C0 Ce )[1 (k 1)(Ce / C0 )]

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

2.2. Adsorption Kinetics

Carbons and Clays for Heavy Metals Removal

2.2.3. Pseudo-Second-Order Model Pseudo-second-order rate equation is expressed as (Ho, 2006a,b):

In the linear form Eq. (2.27) becomes,

k2qe2. Now, Eq. (2.28) can be rearranged to obtain:

qt ; then when t approaches 0, h will become t

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

3. ADSORBENTS FOR HEAVY METALS

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

Carbons and Clays for Heavy Metals Removal

CONCLUSIONS AND RECOMMENDATIONS

John U. Kennedy Oubagaranadin and Z.V.P. Murthy

Carbons and Clays for Heavy Metals Removal

John U. Kennedy Oubagaranadin and Z.V.P. Murthy