Journal of Environmental Management 91 (2010) 1915e1929

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Journal of Environmental Management

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Review

Decolorization of dye wastewaters by biosorbents: A review

Asha Srinivasan, Thiruvenkatachari Viraraghavan*
Faculty of Engineering and Applied Science, University of Regina, Regina, SK, Canada S4S0A2

a r t i c l e i n f o a b s t r a c t

Article history: Dye wastewater is one of the most difficult to treat. There has been exhaustive research on biosorption of
Received 26 October 2009 dye wastewater. It is evolving as an attractive option to supplement conventional treatment processes.
Received in revised form This paper examines various biosorbents such as fungi, bacteria, algae, chitosan and peat, which are
7 April 2010
capable of decolorizing dye wastewaters; discusses various mechanism involved, the effects of various
Accepted 2 May 2010
Available online 2 June 2010
factors influencing dye wastewater decolorization and reviews pretreatment methods for increasing the
biosorption capacity of the adsorbents. The paper examines the mismatch between strong scientific
progress in the field of biosorption and lack of commercialization of research.
Keywords:
Dye
! 2010 Elsevier Ltd. All rights reserved.
Fungi
Bacteria
Algae
Chitosan
Peat

1. Introduction
Dye wastewater from textile and dyestuff industries is one of the
most difficult industrial wastewaters to treat. The wastewater from
these industries is characterized by high alkalinity, biological
oxidation demand, chemical oxidation demand, total dissolved
solids with dye concentrations generally below 1 g/dm3 (Kaushik
and Malik, 2009). The synthetic origin and complex aromatic
structures of dyes make them stable and difficult to be biodegraded
(Fewson, 1998; Seshadri et al., 1994). Dyes are classified as anionic
(direct, acid and reactive dyes); cationic (basic dyes); and nonionic
(disperse dyes). The chromophores in anonic and nonionic dyes
mostly consist of azo groups or anthraquinone types. Anthraqui-
none based dyes are more resistant to degradation due to their
fused aromatic structures. The metal complex dyes are mostly
based on chromium. Azo dyes are the most widely used and
account for 60% of the total dye structures known to be manufac-
tured (Allen, 1971).
Dye wastewater is usually treated by physical or chemical
treatment processes. These include flocculation combined with
flotation, electroflocculation, membrane filtration, electrokinetic
coagulation, electrochemical destruction, ion-exchange, irradia-
tion, precipitation, ozonation, and katox treatment method
* Corresponding author. Tel.: þ1 306 585 4094; fax: þ1 306 585 4855.
E-mail address: t.viraraghavan@uregina.ca (T. Viraraghavan).
involving the use of activated carbon and air mixtures (Banat et al.,
1996). However, these technologies are generally ineffective in
color removal, expensive and less adaptable to a wide range of dye
wastewaters (Banat et al., 1996).
Adsorption has been observed to be an effective process for
color removal from dye wastewater. Use of activated carbon has
been found to be effective, but it is too expensive. Many studies
have been undertaken to investigate the use of low-cost adsorbents
such as peat, bentonite, steel-plant slag, fly ash, china clay, maize
cob, wood shavings, and silica for color removal (Ramakrishna and
Viraraghavan, 1997; Crini, 2006; Gupta and Suhas, 2009). However,
these low-cost adsorbents have generally low adsorption capacities
and require large amounts of adsorbents. Therefore, there is a need
to find new, economical, easily available and highly effective
adsorbents.
Recently, a number of studies have focused on biomaterials that
are capable of biodegrading and biosorbing dyes from wastewaters.
Biological materials such as peat, chitosan, yeast, fungi and bacte-
rial biomass, are used as biosorbents to concentrate and remove
dyes from solutions. The variations in dye structure and their
chemistries result in diverse interactions between dye and bio-
sorbent (Robinson et al., 2001; Crini, 2006). Adsorption of dyes is
dependent on dye properties such as molecular structure and type,
number and position of substituents in the dye molecule (Reife and
Freeman, 1996). There is limited information available on the
interactions between microbial biomass and dyes which can be
explained by the fact that decolorization by living and dead cells

0301-4797/$ e see front matter ! 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.jenvman.2010.05.003
1916 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929
involves several complex mechanisms such as surface adsorption,
ion-exchange, complexation (coordination), complexatione
chelation and micro-precipitation (Crini, 2006). Cell walls consist-
ing mainly of polysaccharides, proteins and lipids offer many
functional groups. The dyes can interact with these active groups
on the cell surface in a different manner. Adsorption is increased by
the presence of hydroxyl, nitro and azo groups in the dye molecule
but decreased by sulfonic acid groups (Reife and Freeman, 1996). It
is now recognized that the efficiency and the selectivity of
adsorption by microbial biomass are due to ion-exchange
mechanisms.
Peat has excellent ion-exchange properties, high cation
exchange capacity and because of its polar character, it can effec-
tively remove dyes from solution (Ramakrishna and Viraraghavan,
1997; Poots et al., 1976). For acid and basic dyes, the removal
performance was comparable with that of activated carbon, while
for disperse dyes, the performance was much better (Allen et al.,
1994; Ramakrishna and Viraraghavan, 1997). Chitosan has a low
affinity for cationic dyes while peat is shown to be a particularly
effective adsorbent for cationic dyes (Allen et al., 1994). The uptake
of dyes on chitosan may proceed through ion-exchange mecha-
nisms and intraparticle diffusion was found to play an important
role in sorption mechanism (Wu et al., 2000). The major adsorption
site of chitosan is a primary amine group which is protonated to
form eNHþ 3 in acidic solutions. Electrostatic interaction between
the eNHþ 3 groups and dye anions can be used to explain the
sorption mechanism (Chiou et al., 2004). The effectiveness of
treatment depends not only on the properties of the adsorbent and
adsorbate, but also on the following environmental conditions and
variables used for the adsorption process: pH, ionic strength,
temperature, existence of competing organic or inorganic ligands in
solution, contact time and adsorbent concentration (Crini, 2006).
This paper summarizes use of various biosorbents capable of
decolorizing dyes, reports on progress and discusses mechanisms
and the factors affecting the process.
2. Fungi
A wide variety of fungal organisms are capable of decolorizing
a wide range of dyes (Fu and Viraraghavan, 2001a, b). Many genera
of fungi have been employed either in living or inactivated form.
The use of white-rot fungi such as Phanerochaete chrysosporium in
decolorizing textile wastewater has been widely reported in liter-
ature (Bilgic et al., 1997; Cammarota and Sant Anna, 1992; Lankinen
et al., 1991; Tatarko and Bumpus, 1998; Young and Yu, 1997; Gomaa
et al., 2008; Sharma et al., 2009; Faraco et al., 2009). There are
various fungi other than white-rot fungi, such as Aspergillus niger
(Fu and Viraraghavan, 2000, 2001b, 2002a), Rhizopus arrhizus
(Zhou and Banks, 1991), Rhizopus oryzae (Gallagher et al., 1997)
which can also decolorize and/or biosorb diverse dyes. Information
on the use of living and dead fungi to decolorize dyes is presented
in Tables 1 and 2, respectively. For living cells, the major mechanism
is biodegradation because they can produce lignin modifying
enzymes, laccase, manganese peroxidase (MnP) and lignin perox-
idase (LiP) which due to their unspecific activity is the first step in
the process of mineralization of dyes (Raghukumar et al., 1996). The
relative contributions of LiP, MnP and laccase to the decolorization
of dyes may be different for each fungus. In addition to biodegra-
dation, a biosorption mechanism might also play an important role
in the decolorization of dyes by living fungi. For dead cells, the
mechanism is biosorption, which involves physico-chemical inter-
actions such as adsorption, deposition, and ion-exchange. Decol-
orization of dye wastewater by fungal biomass has been extensively
reviewed by Fu and Viraraghavan (2001a), Kaushik and Malik
(2009), and Singh (2006). Limited information is available on
interactions between dead fungal biomass and a variety of dyes
with complex molecular structures. Fu and Viraraghavan (2002b)
studied the roles played by functional groups such as carboxyl,
amino, phosphate and lipid fractions present in fungal biomass
from A. niger in biosorption of four different dyes. In biosorption of
Basic Blue 9 on A. niger, carboxyl and amino groups were found to
be the main binding sites while in biosorption of Acid Blue 29, only
amino group was a major site and electrostatic attraction was
believed to be the primary mechanism. In biosorption of Congo
Red, the amino, carboxylic acid, phosphate groups and lipid frac-
tions were all found to be important binding sites and in addition to
electrostatic attraction, other mechanisms were also believed to be
involved in biosorption. In biosorption of Disperse Red 1, physical
and chemical adsorption along with electrostatic attraction was
found to be the mechanism of biosorption while amino group and
lipid fractions were the major binding sites.
2.1. Yeast
Yeast decolorization and degradation of dyes has not been
extensively studied. Information on yeast decolorization of dyes
available in literature is given in Table 3. Biosorption of textile dyes
has been found to occur by biomass derived from yeast Kluyver-
omyces marxianus IMB3 (Bustard et al., 1998). K. marxianus IMB3
was also found to decolorize Remazol Black-B through physical
adsorption (Meehan et al., 2000). The oxidative yeasts Rhodotorula
sp. and Rhodotorula rubra were found to degrade crystal violet
completely in four days (Kwasniewska, 1985). An yeast strain
Candida zeylanoides was found to be able to degrade a number of
azo dyes, whose reduction product include metanilic acid for azo
dyes I and III, and sulfanilic acid for azo dyes II and IV. Saccharo-
myces cerevisiae has been found to be effective in removing dye in
molasses media (Aksu, 2003).
2.2. Fungal bioreactors for dye decolorization
Fungal bioreactors for dye decolorization and degradation have
been developed recently. Both color removal and degradation can
take place individually or simultaneously in the reactor. White-rot
fungi have been widely studied for its decolorization potential in
bioreactor systems. At present no information on commercial appli-
cations of fungal bioreactor systems are available. Rotating drum,
packed bed, fluidized bed, immobilized and membrane bioreactors
have been used as decolorizing bioreactors (Table 4). A membrane
bioreactor using Trametes versicolor combined with reverse osmosis
was effective for decolorization of dye wastewater (Kim et al., 2004). A
wood-rotting fungal strain F29 decolorized 95 to 99% Orange II in
a continuous packed bed and fluidized bed bioreactor systems (Zhang
et al.,1999). Immobilized bioreactors have been found to exhibit good
biological activities and abilities for long time operation (Singh, 2006).
Irpex lacteus immobilized in a pine wood (PW) reactor decolorized
Remazol Brilliant Blue R more rapidly than a polyurethane foam (PUF)
reactor (Kasinath et al., 2003).
2.3. Factors influencing fungal decolorization of dyes
2.3.1. Effect of nutrients on dye decolorization
Biodegradation of dyes can be enhanced by improving the initial
growth conditions. Glucose, starch, maltose, and cellobiose were
found to be good carbon sources for decolorization of cotton
bleaching effluent by white-rot fungus (Zhang et al., 1999). P.
chrysosporium showed superior performance at a glucose concen-
tration of 5 g/L and an ammonium chloride concentration of 0.05 g/
L in decolorizing Methyl Violet (Radha et al., 2005). A high dose of
nutrient nitrogen was found to inhibit decolorization of Congo Red
Table 1
Results of dye decolorization by live fungi.

Culture Dye Percent Experimental Mechanism Time Reference

removal conditions of contact
Rhizopus oryzae Rhodamine B 90 Initial dye concentration 100 mg/L; Chemical interaction, ionic 5h Das et al., 2006
(xanthine dye) pH 7.0; biomass dose 0.25 g/25 mL; interaction, physical forces
temperature 40 " C; point
of zero charge pH 3.5
Schizophyllum commune Acid Orange 7 44. 23a Initial dye concentration 100 mg/L; Bioaccumulation 72 h Renganathan et al., 2006
Acid Red 18 127.53a pH 2.0; temperature 30 " C
Reactive Black 5 180.17a
Pencillium oxalicum Reactive Blue 19 91 Initial concentration 100 mg/L; pH 2.0; Biosorption 80 min Zhang et al., 2003
biosorbent dose 0.25 g/100 mL
Isolate Aspergillus 1 Direct Yellow 72 Initial dye concentration range 1e100 mg/L Biosorption 24 h Abd El-Rahim et al., 2003
Direct Brown 85

A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929

Maxilon Red 49
Erio Red 21
Aspergillus ochrocous 5 Polar Red 96 Initial dye concentration 0.3 g/L Biosorption 8d Abd El-Rahim
and Moawad, 2003
Aspergillus niger 31 Polar Red 94 Initial dye concentration 0.3 g/L Biosorption 8d Abd El-Rahim
and Moawad, 2003
Umbelopsis isabellina Reactive Black 5 >99 Initial dye concentration 100 mg/L MnP 48 h Yang et al., 2003
Penicillium geastrivorus Reactive Black 5 >99 Initial dye concentration 100 mg/L Biosorption 48 h Yang et al., 2003
Funalia trogii Astrazon Red 92e98 Initial dye concentration range 0e1500 mg/L; Adsorption and 24 h Yesilada et al., 2002
initial pH 6e11; temperature 30 " C microbial metabolism
Funalia trogii Reactive Blue 19 >99 Inoculum contain 80 mL of Kirk’s Laccase, MnP 10 d Park et al., 2007
Reactive Blue 49 basal salts and 100 mg/L of dye; pH 4.5
Acid Violet 43
Reactive Black 5
Reactive Orange 16
Acid Black 52
Aspergillus sp. Reactive Blue 99 Initial dye concentration Biodegradation 24 h Mohandass et al., 2007
Reactive Black 75 100 mg/L; pH 3.0
Trichoderma harzianum Erioglaucine 76e88 Initial dye concentration 105 min Sadhasivam et al., 2007
range 10e50 mg/L;
biosorbent dose 1.5 g/50 mL; pH 4.0
Pleurotus florida Blue CA 93.54 Dye concentration 200 mg/L; Laccase 10 d Sathiya Moorthi et al., 2007
Trametes hirusta 92.17 nutrient salt media
Myrothecium sp. Remazol Brilliant Blue R 90 Initial dye concentration 80 mg/L; Biosorption and biodegradation 7d Zhang et al., 2007
pH 7.0; potato dextrose broth
Aspergillus niger Basic Blue 9 10 Intial dye concentration 50 mg/L; initial Biosorption 2d Fu and Viraraghavan, 2000
pH 5.1; biosorbent dose 0.2 g/75 mL
Aspergillus niger Acid Blue 29 80 Intial dye concentration 50 mg/L; initial Biosorption 30 h Fu and Viraraghavan, 2001b
pH 7.6; biosorbent dose 0.2 g/75 mL
P. chrysosporium Reactofix Gold Yellow 73 Cell mass concentration 1.4 mg/mL; pH of Lignin-degrading 3d Capalash and Sharma, 1992
growth medium 4.5; incubation at 39 " C system and adsorption
P. chrysosporium Reactive Red 22 92e100 Initial dye concentration 120e140 mg/L Lignin degrading system 30 h Wu et al., 1996
Pleurotus pulmonaris Congo Red 93 Initial dye concentration 200 ppm; glucose Biodegradation, 6d Tychanowicz et al., 2004
ammonium tartrate-corncob adsorption and laccase
solid state cultures
Irpex lacteus Methyl Red 56 Initial dye concentration 150 mg/g Lignin degrading system 14 d Novotny et al., 2001
C. versicolor Indigo Carmine 98 Initial concentration 23 mg/L Biodegradation and laccase 1h Levin et al., 2004
Trametes trogii Anthraquinone Blue 88 Glucose-asparagine and malt Laccase 4h Levin et al., 2001
extract/glucose medium
Geotrichum sp. Reactive Black 5 >99 Initial dye concentration 100 mg/L Biodegradation, MnP and laccase 10 d Maximo et al., 2003
Reactive Red 158 20 d
Reactive Yellow 27 20 d
Dichomitus squalens Orange G 95 Initial concentration 50 mM Laccase and MnP 14 d Eichlevora et al., 2005
(continued on next page)

1917
Table 1 (continued )

1918
Culture Dye Percent Experimental Mechanism Time Reference
removal conditions of contact
P. ostreatus Remazol Brilliant Blue R 100 Initial concentration 50 mM Laccase 9d Palmieri et al., 2005
P. chrysosporium Direct dyes 100 Initial concentration 120 mg/L Biodegradtion, 15 d Pazarlioglu et al., 2005a, b
adsorption and MnP
Trametes versicolor Direct Blue 1 63.2 Initial concentration 800 mg/L; Biosorption 6h Bayramoglu and Arica, 2007
pH 6.0; biosorbent dose 250 mg/50 mL
Daedalea flavida Coracryl Black 13.9 Initial concentration 30 mg/L; Cell free enzyme 3h Chander and Arora, 2007
Coracryl Pink 52.9 fungal culture grown for 6 days based decolorization 5h
Coracryl violet 26 3h
Coracryl Red 35.2 3h
Reactive Yellow 11.7 3h
Reactive Orange 4.8 3h
Reactive Red 39.8 3h

A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929

Rathidol Scarlet 39 3h
Dichomitus squalens Coracryl Black 69 Initial concentration 30 mg/L; Cell free enzyme 1h Chander and Arora, 2007
Coracryl Pink 100 fungal culture grown for 6 days based decolorization 2h
Coracryl violet 76 1h
Coracryl Red 45 1h
Reactive Yellow 79.9 2h
Reactive Red 76 1h
Rathidol Scarlet 79 3h
Irpex flavus Coracryl Black 67.3 Initial concentration 30 mg/L; Cell free enzyme 2h Chander and Arora, 2007
Coracryl Pink 100 fungal culture grown for 6 days based decolorization 3h
Coracryl violet 58 5h
Coracryl Red 50.2 2h
Reactive Yellow 79 1h
Reactive Orange 63.2 2h
Reactive Red 77 1h
Rathidol Scarlet 73.9 2h
Polyporus sanguineus Coracryl Black 67 Initial concentration 30 mg/L; Cell free enzyme 5h Chander and Arora, 2007
Coracryl Pink 64.9 fungal culture grown for 6 days based decolorization 3h
Coracryl Red 36.5 5h
Reactive Orange 19.3 2h
Reactive Red 59.2 5h
a
Milligram of dye adsorbed/g of biomass; percent removal data not available.
Table 2
Results of dye decolorization by dead fungi.

Culture Dye Percent Experimental Mechanism Time Reference

removal conditions
Aspergillus niger Acid Blue 29 99 Intial dye concentration 50 mg/L; initial pH 7.6; Biosorption 24 h Fu and
biosorbent dose 0.2 g/75 mL Viraraghavan, 2001b
Aspergillus niger (immobilized) Acid Blue 29 64.7a 4.5 g of beads; column dia 1.27 cm, 5.0 (min) Fu and
Basic Blue 9 8.3a height 40 cm; flow rate 6 mL/min 5.2 (min) Viraraghavan, 2003
Congo Red 1.1a 5.2 (min)
Disperse Red 1 0.1a 5.2 (min)
Aspergillus niger Congo Red 89.6 Intial dye concentration 50 mg/L; Biosorption 42 h Fu and
initial pH 6.5; biosorbent dose 0.2 g/75 mL Viraraghavan, 2002a
Aspergillus niger Synazol 88 Synazol Red 0.22%, Synazol Yellow 0.1% 18 h Khalaf, 2008
Aspergillus foetidus Reactive Black 5 >99 Initial dye concentration 100 mg/L; Biosorption 2h Patel and Suresh, 2008
pH 2e3; biosorbent dose 0.2 g/100 mL

A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929

P. chrysosporium Astrazone Blue FGRL 60 Initial dye concentration 50 mg/L; Adsorption 2h Asma et al., 2006
Cibracron Red 51 biosorbent 0.2 g/50 mL; without adjusting pH
Funalia trogii Astrazone Blue FGRL 48 Initial dye concentration 50 mg/L; Adsorption 2h Asma et al., 2006
Cibracron Red 38 biosorbent 0.2 g/50 mL; without adjusting pH
Cunninghamella elegans Direct Red 80 100 Initial dye concentration 1000 ppm, Biosorption 24 h Prigione et al., 2008
Reactive Blue 214 99 5000 ppm; biosorbent 3 g/30 mL
Reactive Blue 19 57e63
Rhizomucor pusillius Direct Red 80 100 Initial dye concentration Biosorption 24 h Prigione et al., 2008
Reactive Blue 214 >98 1000 ppm, 5000 ppm; biosorbent 3 g/30 mL
Reactive Blue 19 84e98
Rhizopus stolonifer Direct Red 80 100 Initial dye concentration 1000 ppm, Biosorption 24 h Prigione et al., 2008
Reactive Blue 214 98 5000 ppm; biosorbent 3 g/30 mL
Reactive Blue 19 99
Rhizopus arrhizus Reactive Orange 16 190a Initial concentration 0e500 mg/L; 20 h O’Mahony et al., 2002
Reactive Red 4 150a pH 2.0; biosorbent dose 1 g/L
Reactive Blue 19 90a
Rhizopus arrhizus Reactive Black 5 62.5 Initial dye concentration 800 mg/L; Adsorption 24 h Aksu and Tezer, 2000
biosorbent dose 1 g/L; pH 2.0
Rhizopus arrhizus Germazol 47.5 Initial initial dye concentration Physical adsorption 24 h Aksu and Cagatay, 2006
Torquoise Blue-G of 812.6 mg/L; pH 2.0;
biomass dosage of 0.5 g/L; temperature 45 " C
Rhizopus arrhizus Gryfalan Black RL 59 Initial concentration range 1000 mg/L; Adsorption and 400 min Aksu and
Trametes versicolor 37 biosorbent dose 1.0 g/L; pH 2.0; internal diffusion Karabayur, 2008
Aspergillus niger 37.5 Temperature 25 " C (pH 1.0 and
temperature 35 " C for A. niger)
Rhizopus nigricans Reactive Green 86 Initial dye concentration 50 mg/L; biomass loading: Adsorption 1h Kumari and
Reactive Blue 83 1 g% (w/v); pH 6; temperature: 29 # 1 " C Abraham, 2007
Penicillium chrysogenum Acid Orange 8 70.4 Polyethylenimine modified; Sorption 14 h Low et al., 2008
Acid Blue 45 39.2 Initial concentration 500 mg/L;
Reactive Orange 16 67.6 biosorbent dose 1 g/L; initial pH 6.8e7.2
Rhizopus stolonifer Bromophenol Blue 88 Initial concentration 800 mg/L; pH 2.0; Biosorption 20 h Zeroual et al., 2006
biosorbent dose 1 g/L
Neurospora crassa Acid Red 57 98.78 Initial concentration range 100e400 mg/dm3; Biosorption 40 min Akar et al., 2006
pH 1.0; temperature 20 " C; biosorbent dose 2 g/dm3
Trametes versicolor Direct Blue 1 95.2 Initial concentration 800 mg/L; Biosorption 6h Bayramoglu and
pH (Direct Blue 1) 6.0; Arica, 2007
biosorbent dose 250 mg/50 mL
Agaricus bisporus þ Thuja Reactive Blue 49 72.86a Initial concentration 150 mg/L; pH 1.0; point Biosorption 90 min Akar et al., 2009a
orientalis (mixed) of zero charge 1.5; biosorbent dosage 3.0 g/L
Thuja orientalis Acid Blue 40 48.5 Initial concentration 200 mg/dm3; pH 1.0; Biosorption 90 min Akar et al., 2008
temperature 20 " C; biosorbent dose 1 g/dm3
(continued on next page)

1919
 1920 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929

(Tatarko and Bumpus, 1998). Nitrogen was found to have no effect

on the decolorization of dyes by Cyathus bulleri (Vasdev et al., 1995).
As decolorization of dyes by P. chrysosporium occurs in secondary
Akar et al., 2009b

Wang et al., 2008

Xiong et al., 2010

Kiran et al., 2006

metabolic conditions, the enzyme LiP is released by fungal cells
under either carbon or nitrogen limitation (Zhen and Yu, 1998). pH
Reference

is an important factor for fungal growth. Fungi are generally found

to grow at low pH, normally ranging from 4 to 5. The optimum
growth temperature for most of the fungi is found to be at about
25e35 " C (Fu and Viraraghavan, 2001a).

2.3.2. Characteristics of dye wastewater

120 min

120 min

The molecular structure of dyes is found to have an effect on the

120 h
Time

4h

extent of decolorization. Wong and Yu (1999) reported that dye

decolorization by T. versicolor was dependant on dye structures.
Anthroquinone dye was a laccase substrate while azo and indigo dyes
were not the substrates of laccase. Spadaro et al. (1992) observed that
Chemical ion-exchange

aromatic rings with substituents such as hydroxyl, amino, acetamido,

or nitro functions were mineralized to a greater extend than unsub-
Chemisorption

stituted rings in dye decolorization by P. chrysosporium. Dye

Mechanism

Biosorption

Biosorption

concentration also affects the efficiency of color removal. The decol-

orization efficiency by Coriolus versicolor decreased from 100% to 80%
when the dye concentration was increased from 100e500 mg/L to
700e1200 mg/L (Kapdan et al., 2000).
The ionic forms of the dye in solution and the surface electrical
charge of the biomass depend on solution pH. Therefore, solution
pH influences both the fungal biomass surface dye binding sites and
w2.0; biosorbent dose 3 g/L; temperature 40 " C

the dye chemistry in the medium. Fu and Viraraghavan (2000,

biosorbent dose 0.4 g/dm3; temperature 20 " C

2001b) reported that the effective initial pH of dye solution was 6

Initial concentration 150 mg/dm3; pH 1.0;
Initial concentration range 50e300 mg/L;

biosorbent dose 6 g/L, temperature 45 " C

and 4, respectively, for Basic Blue 9 and Acid Blue 29. At pH of 2.0,
Initial concentration 400 mg/L; pH 3.0;
pH 2.0; biosorbent dose 0.05 g/25 mL

no biosorption occurred for Basic Blue 9 due to the high concen-

tration of protons, while at pH of 12, no biosorption occurred for
Initial concentration 33.9 mg/L;

Acid Blue 29. Arica and Bayramoglu (2007) reported that as the pH
pH 2.0; point of zero charge

was decreased, the biosorption of Reactive Red 120 dye on the

fungal biomass Lentinus sajor-caju increased. Similarly, O’Mahony
et al. (2002) observed maximum removal of reactive dye Remazol
Black-B in the range of 1e2 with a sharp drop off at higher values.
Experimental

At lower pH values the fungal biomass will have a net positive

conditions

charge. These charged sites become available for binding anionic

groups such as reactive dyes.
Most textile and other dye effluents are produced at relatively
high temperatures and hence temperature will be an important
factor in real application of biosorption in future. Arica and
Milligram of dye adsorbed/g of biomass; percent removal data not available.

Bayramoglu (2007) found that biosorption of dye by Lentinus

removal
Percent

59.80a

sajor-caju increased with increasing temperature from 5 to 35 " C.

94.7

44.9

29.2

Aksu and Cagatay (2006) observed an increase in uptake of dye

with increasing temperature up to 45 " C for R. arrhizus showing
endothermic character of biosorption. Dyeing processes consume
large amounts of salt and hence the dye wastewater contains high
Reactive Brilliant

salt concentration. For this reason, ionic strength is an important

Direct Blue 199

factor. Zhou and Banks (1991, 1993) reported that high ionic
Acid Red 44

Acid Red 57
Red K-2BP

strength led to high biosorption of humic acid by R. arrhizus.

However, adsorption capacities of Lentinus sajor-caju biomass
Dye

showed no significant effect with increasing NaCl from 0 to 0.5 M

concentration (Arica and Bayramoglu, 2007).

2.3.3. Pretreatment
Research has shown that some pretreatment processes can
(carboxymethylcellulose

Cephalosporium aphidicola

increase the adsorption capacity of biomass. The pretreatment

immobilized beads)

methods include autoclaving, contacting with organic chemicals

Aspergillus fumigatus
Table 2 (continued )

such as formaldehyde or inorganic chemicals such as NaOH, H2SO4,

Agaricus bisporus

Aspergillus niger

NaHCO3 and CaCl2. Fu and Viraraghavan (2000, 2001b) used auto-

claving and contacted with chemicals such as 0.1 M NaOH, 0.1 M
HCl, 0.1 M H2SO4, etc. to pretreat the living A. niger biomass. It was
Culture

found that autoclaving increased biosorption capacity of Basic Blue

a

9 from 1.17 mg/g to 18.54 mg/g, while 0.1 M H2SO4 pretreatment

 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929
Table 3
Data on various yeast cells capable of dye decolorization.
Yeast Dye Percent
removal
Debaryomyces polymorphus C.I. Reactive black >90
dye (RB5), 200 mg/L
Kluyveromyces Remazol black-B, 200 mg/L 98
marxianus IBM3
Rhodotorula rubra Crystal violet, 10 ppm >99
Candida zeylanoides Azo dyes, 10e50 ppm 46e67
C. guilliermondii Remazol blue, 200 mg/L 41
Candida tropicalis Reactive Black 5, 100 mg/L >99
S. cerevisiae Reactive Green, 50 mg/L 83
Reactive Blue 38, 50 mg/L 83
Reactive Blue 3, 50 mg/L 91
enhanced biosorption capacity from 6.63 mg/g to 13.83 mg/g for
Acid Blue 29. They suggested that autoclaving could rupture the
fungal structure and expose the potential binding sites for the dye
while acid pretreatment could change the negatively charged
surface of fungal biomass to positively charged and thus increasing
the attraction between fungal biomass and Acid Blue 29, an anionic
dye. Arica and Bayramoglu (2007) observed heating the biomass
Lentinus sajor-caju at 100 " C for 10 min enhanced the biosorption
capacity while base-treatment with 0.1 M NaOH lowered the bio-
sorption capacity of the fungi to remove Reactive Red 120.
3. Bacteria
Most research has focused on studying the biodegradation/
decolorization potential of bacteria (Forgacs et al., 2004) and less
attention has been paid on employing dead bacterial biomass for
biosorption of dyes. With respect to bacterial dye biosorption, Won
et al. (2005) identified Corynebacterium glutamicum as a potential
biosorbent of Reactive Red 4, which can bind 104.6 mg/g at pH 1. Two
general types of bacteria exist, Gram-positive and Gram-negative.
Gram-positive bacteria are comprised of a thick peptidoglycan layer
(Beveridge, 1981; Dijkstra and Keck, 1996) connected by amino acid
bridges. Imbedded in the Gram-positive cell wall are polyalcohols,
some of which are lipid linked to form lipoteichoic acids. The cell
wall of Gram-negative bacteria is thinner, and composed of only
10e20% peptidoglycan (Beveridge, 1999). In addition, the cell wall
contains an additional outer membrane composed of phospholipids
and lipopolysaccharides (Sheu and Freese, 1973). The mode of solute
uptake by dead/inactive cells is extracellular; the chemical func-
tional groups of the cell wall play vital roles in biosorption.
Biodegradation processes may be anaerobic, aerobic or involve
a combination of the two. The ability of actinomycetes, particularly
Streptomyces species, to decolorize and degrade textile dyes has
been widely investigated.
3.1. Live bacteria
Initially, 14 streptomycetes were investigated for their ability to
decolorize two polymeric dyes, Poly B-411 and Poly R-478, as well
as the azo dye Remazol Brilliant Blue R (RBBR) (Pasti and Crawford,
1991). Further work on Streptomyces chromofuscus A11 confirmed
that decolorization was related to the ligninolytic capabilities of the
isolate. It was proposed that the peroxidases of the organism
converted the azo dye to a cation radical that was susceptible to the
nucleophilic attack of water or hydrogen peroxide. This resulted in
the simultaneous split of the azo linkage both symmetrically and
asymmetrically to produce intermediates which subsequently
undergo several redox reactions before producing more stable
1921
Mechanism Incubation Reference
Manganese dependant 24 h Yang et al., 2005
peroxidase
Physical adsorption 24 h Meehan et al., 2000
Enzymes 4d Kwasniewska, 1985
Adsorption and 22 h Martins et al., 1999
biodegradation
Adsorption 15 m Aksu and Donmez, 2003
MnP 48 h Yang et al., 2003
Adsorption 1h Kumari and Abraham, 2007
intermediates. Information on the use of various living bacteria to
decolorize dyes is presented in Table 5.
Nigam and Marchant (1995) and Nigam et al. (1996) demon-
strated that a mixture of dyes were decolorised by anaerobic
bacteria in 24e48 h, using free growing cells or in the form of
biofilms on various support materials. Ogawa and Yatome (1990)
also demonstrated the use of bacteria for azo dye biodegradation.
Kulla (1981) reported the ability of Pseudomonas strains to aero-
bically degrade certain azo dyes. An azoreductase enzyme was
responsible for the initiation of the degradation of the Orange II dye
by these strains and substituting any of the groups near the azo
group’s chemical structure hindered the degradation
(Zimmermann et al., 1982). Under anaerobic conditions, such as
anoxic sediments, many bacteria reduce azo dyes reportedly by the
activity of unspecific, soluble, cytoplasmic reductases, known as
azoreductases. Initially, the bacteria bring about the reductive
cleavage of the azo linkage, which results in dye decolorization and
the production of colorless aromatic amines. Concerns have long
been expressed over the threat that such compounds have on
human health, and their environmental impact is also troublesome
(McMullan et al., 2001).
Isolation of bacteria capable of the aerobic decolorization of
sulfonated azo dyes has proven difficult. Blhmel et al. (1998)
reported the isolation of an unidentified bacterial strain, S5
capable of utilizing the model sulphonated azo compound as a sole
carbon and energy source. Attempts to characterize the enzyme
responsible for the azo-bond cleavage in crude cell extracts have so
far proven unsuccessful.
Hu (1996) demonstrated the ability of bacterial cells isolated
from activated sludge process of a textile industry and soil to adsorb
11 reactive dyes including Reactive Blue, Reactive Red, Reactive
Violet, Reactive Yellow and Procion Red G. The cell wall portion of
Aeromonas sp. was found to have higher specific adsorption
capacity than the intact cells. Zhou and Zimmermann (1993) used
the actinomycete Streptomycetes BW130 as an adsorbent for the
decolorization of effluents containing anthroquinone, phtlocya-
nine, and azo dyes. It was found that decolorization was through
adsorption of these dyes to the cellular biomass without any
degradation. Other Cu-based azo dyes, such as formazan-copper
complex dyes, were completely decolorized through degradation
by the same actinomycete strains (Zhou and Zimmermann, 1993).
3.2. Dead bacteria
In a study by Hu (1996), three Gram-negative bacteria (Aero-
monas sp., Pseudomonas luteola and Escherichia coli), two Gram-
positive bacteria (Bacillus subtilis and Staphylococcus aureus) and
activated sludge (consisting of both Gram-negative and Gram-
positive bacteria) were used as biosorbents for the removal of
Table 4

1922
Fungal bioreactors for dye decolorization.

Fungus Bioreactor configuration Dye Percent Enzyme Duration Reference

removal activity
Phanerochaete chrysosporium Rotating drum Poly R-478 19 MnP, LiP 15 m Dominguez et al., 2001
Trametes versicolor Stirred-tank reactor Poly R-478, 200 mg/L 80 Biotransformation 41 d Leidig et al., 1999
and adsorption
Phanerochaete sordida Immobilized on RBC Basic Blue 22, 200 mg/L 98 Lignolytic enzyme 24 h Ge et al., 2004
Coriolus versicolor Immobilized in form of biofilm, Eerzol Turquoise 82 Lignolytic enzyme 20 d Kapdan and
activated sludge, and wood ash Blue-G, 200 mg/L Kargi, 2002
P. chrysosporium Immobilized on polyurethane Poly R-478, 0.1 g/L 80 MnP 24 h Mielgo et al., 2002
foam, pulsed packed bed
Irpex lacteus Immobilized on PUF or PW, Remazol Brilliant 100 MnP and laccase 6d Kasinath et al., 2003
packed bed reactor Blue R, 150 mg/L
Irpex lacteus Immobilized on PW cubes, Remazol Brilliant 100 MnP and laccase 6d Novotny et al., 2004

A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929

packed bed reactor Blue R, 150 mg/L
Trametes hirsute Immobilized on stainless steel sponge, Indigo Carmine 100 Laccase activity 3d Rodríguez
fixed bed bioreactor Couto et al., 2004
P. chrysosporium Immobilized on ZrOCl2-activated Direct Blue 15, 20 mg/L 95e100 MnP Pazarlioglu
pumice, packed bed reactor et al., 2005a, b
P. chrysosporium Immobilized on mineral kissiris Methylene Blue, 60 mg/L 100 MnP 8d Karimi et al., 2006
T. versicolor Immobilized on birch wood, continuous RBC Reactive Blue 4, 200 mg/L 70 3d Nilsson et al., 2006
T. hirsute Immobilized on ground orange Indigo Carmine 94 3d Rodríguez
peelings, fixed bed reactor Methyl Orange 81.4 Couto et al., 2006
Poly R-478 46.9

Table 5
Dye decolorization by bacterial biomass.

Culture Dye Percent removal Mechanism Time Reference

Pseudomonas cepacia 13NA (immobilized) p-Aminoazobenzene, 10 mg/L 60e80 Azoreductase 10 h Ogawa and Yatome, 1990
Pseudomonas luteola Red G, 100 mg/L 37.4 Azoreductase 2d Hu, 1994
Pseudomonas luteola RBB, 100 mg/L 93.2 Azoreductase 2d Hu, 1994
Streptomyces BW130 Azo-reactive Red 147, 150 mg/L 29 Adsorption 14 d Zhou and Zimmermann, 1993
Streptomyces BW130 Azo-copper Red 171, 180 mg/L 73 Adsorption 14 d Zhou and Zimmermann, 1993
Bacillus subtilis IFO 3002 p-aminoazobenzene, 30 mg/L 80e90 Azoreductase 30 h Horitsu et al., 1977
Mixed anaerobic culture Diazo-linked chromophores 85 Azoreductase 2d Knapp and Newby, 1999
Pseudomonas putida (NCIMB 9776) Tectilon Blue (TB4R) 1000 mg/L 50 18% biosorption 24 h Walker and Weatherley, 2000
Bacterial consortium (mixture of Ochrobactrium sp., Reactive Black-B 92.3 mg/L þ Cr(VI) Dye 77 Cr (VI) 99 Bioaccumulation 2e4 d Kilic et al., 2007
Salmonella enterica and Pseudomonas aeruginosa) 78.6 mg/L
Corynebacterium glutamicum Reactive Black 5, 500 mg/L 94 Biosorption 12 h Vijayaraghavan and Yun, 2007a
Corynebacterium glutamicum (raw) Reactive Black 5, 500 mg/L 56 Biosorption 12 h ` Vijayaraghavan and Yun, 2007b
Corynebacterium glutamicum (immobilized) Reactive Black 5, 100 mg/L 78.6a Intraparticle diffusion 60 h Vijayaraghavan and Yun, 2007b
Streptomyces rimosus Methylene Blue, 50 mg/L 86 Biosorption 5 min Nacera and Aicha, 2006
a
Milligram of dye adsorbed/g of biomass; percent removal data not available.
 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929
reactive dyes such as Reactive Blue, Reactive Red, Reactive Violet
and Reactive Yellow. Dead cells of test genera showed a higher
uptake than living cells due to an increased surface area and Gram-
negative bacteria had a higher adsorption capacity than Gram-
positive bacteria due to higher lipid contents in the cell wall
portion.
The mode of solute uptake by dead/inactive cells is extracellular,
and the chemical functional groups of the cell wall play vital roles in
biosorption. Functional groups present on the bacterial cell wall
include carboxyl, phosphonate, amine and hydroxyl groups (van
der Wal et al., 1997). Several dye molecules, which exist as dye
cations in solutions, are also attracted towards carboxyl and other
negatively charged groups. Also, amine groups adsorb anionic dyes
via electrostatic interaction or hydrogen bonding. Vijayaraghavan
and Yun (2007a, b) observed that the amine groups of C. gluta-
micum were responsible for the binding of reactive dye anions via
electrostatic attraction. Carboxyl, amine, phosphonate, sulfonate
and hydroxyl groups have become well established as being
responsible for dye binding.
3.3. Factors affecting bacterial color removal
The most important factor to consider is the effect of oxygen on
cell growth and dye reduction. It has generally been observed that
efficiency of color removal by live bacterial cells decreases with
increase in concentration of oxygen. Increase in concentration of
oxygen may result in direct inhibition of the azoreductase enzyme
or preferential reduction of oxygen rather than the azo derivatives
(Semde et al., 1998). However, oxygen would be required to degrade
aromatic intermediates completely. Hence, a balance between the
anaerobic and aerobic stages in this treatment system must be
carefully controlled.
Hu (1996) investigated the effect of pH on the removal of six
reactive dyes by three gram-negative bacteria, P. luteola, E. coli, and
Aeromonas sp. and observed that biosorption capacity increased
significantly with decreasing pH for the all the cases. They
explained that at low pH, the dye anions are electrostatically
bonded to positively charged bacaterial cell surfaces. Biosorption of
Reactive Black 5 by C. glutamicum was also found to be found to be
favoured at pH value of 1 (Vijayaraghavan and Yun, 2007a, b). The
extent to which metal ions in solution undergo hydrolysis varies
with each metal and solution pH. In the case of live bacterial cells,
the optimum pH for color removal is often at a neutral pH value or
a slightly alkaline pH value (Pearce et al., 2003).
The temperature required by live bacterial cells to produce
maximum rate of color removal tends to correspond with the
optimum cell culture growth temperature of 35e45 " C (Pearce
et al., 2003). In the case of dead bacterial biomass, the highest
dye uptake for Methylene Blue by Streptomyces rimosus was
exhibited at 20 " C (Nacera and Aicha, 2006). Biosorption capacity
was found to decrease with an increase in temperature from 20 to
50 " C. The results showed that dye adsorption process by the
biomass was exothermic in nature. Vijayaraghavan and Yun (2007a,
b) however observed that the biosorption performance increased
with an increase in temperature up to 35 " C for uptake of Reactive
Black 5 by C. glutamicum.
Chemical pretreatments for bacterial biosorbents include acid,
alkaline, ethanol and acetone treatments of the biomass
(Vijayaraghavan and Yun, 2007a, b). In general, acidic pretreatment
has proven successful. Among the immobilization techniques
studied for bacterial biosorbents, polysulfone was found to be
a favorable immobilizing agent for C. glutamicum biomass
(Vijayaraghavan and Yun, 2007a, b). C. glutamicum in polysulfone
matrix showed the ability for the biosorption of reactive dyes over
20 successive sorptionedesorption cycles.
1923
4. Chitosan
There is abundant literature regarding the evaluation of
adsorption performances of raw chitosan, especially in terms of
adsorption capacity or uptake (Table 6). Numerous dyes, mainly
anionic dyes, have been so far been studied. In particular, chitosan
has been found to possess high removal capacities for anionic dyes
such as acid, reactive and direct dyes. This is due to the unique
polycationic structure of chitosan. Chitosan, produced from partial
deacetylation of chitin, is a polysaccharide composed of polymers
of glucosamine and N-acetyl glucosamine. Chitosan is of great
commercial interest due to its high percentage of nitrogen content
compared to synthetically substituted cellulose. Commercially
available chitosans are polymers with less than 25% acetyl content
while fully deacetylated product is rarely obtained. Chitosan is
soluble in acids and chemically more versatile than chitin and
cellulose (Crini and Badot, 2008).
4.1. Factors influencing decolorization by chitosan
4.1.1. Chitosan characteristics
The adsorption capacity of chitosan depends on its physical
structural parameters such as crystallinity, surface area, porosity,
particle type, particle size and water content. Crystallinity is high
for both chitin and fully deacetylated chitosan. Generally,
commercial chitosans are semi-crystalline polymers and the degree
of crystallinity is a function of the degree of deacetylation (DD).
Decrystallized chitosan is found to be much more effective in the
adsorption of anionic dyes (Trung et al., 2003). The case of dye
adsorption with crosslinked chitosan is a typical example of the
influence of particle size. Among the other parameters that have
a great impact on dye adsorption is particle type. Chitosan can be
presented as gels, flakes, powders and particles. Chitosan beads are
preferred since flake and powder forms of polymer are not suitable
for use as adsorbents due to their low surface area and lack of
porosity. Crini et al. (2008b) observed that compared to chitosan
flakes, chitosan beads exhibited a twofold or more increase in the
adsorption capacity for Basic Blue 9. One of chitosan’s most
promising features is its excellent ability to be processed into
nanostructures. Annadurai (2002) observed that the adsorption
capacity increased with a decrease in the particle size and the dye
molecules were preferably adsorbed on the outer chitosan surface.
The author suggested that this observation can be attributed to the
larger total surface associated with smaller particles. In contrast
Guibal et al. (2003) observed that the adsorption occurred not only
at the surface of the material due to rapid surface adsorption but
also in the intraparticle network of the polymer. The greater the
particle size, the greater the contribution of intraparticle diffusion
resistance to the control of the adsorption kinetics for materials of
low porosity. Intraparticle diffusion is found to greatly influence the
accessibility of dye molecules to internal sites. Adsorption perfor-
mance is also controlled by polymer porosity. Chitosan is known as
a non-porous polymer. It is characterized by a low surface area and
a low porosity that control the diffusion to the center of the
particles, especially with large molecules. These features generally
limit access to interior adsorption sites. So, polymer porosity may
affect the dye adsorption capacity of chitosan. The molecular
weight (MW) of chitosan is a key variable in adsorption properties
because it influences the polymer’s solubility and viscosity in
solution. Another important characteristic of chitosan is the degree
of N-acetylation (DA) or DD. The higher DD chitosan provided
a better adsorption. With an increase in DD, the number of amino
groups in the polymer increases, and with an increase of MW, the
polymer configuration in solution becomes a chain or a ball.
1924 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929

Table 6
Dye removal by chitosan.

Dye Chitosan Pretreatment Adsorption Mechanism Time of Reference

capacity (mg/g) contact
Reactive Blue 19 Beads Crosslinking 400 Physisorption 48 h Hasan et al., 2008
Remazol Black 13 Powder 91.5e130.0 Intraparticle diffusion 20 h Annadurai et al., 2008
Basic Blue 3 CHITOD 166.5 Adsorption, diffusion, 40 min Crini et al., 2008a
electrostatic interaction
Methyl Orange Powder Crosslinking 130 Electrostatic interaction, 10 h Morias et al., 2007
intraparticle diffusion
Remacryl Red TGL Powder Grafting 1.068 mmol/g Chemisorption 24 h Lazaridis et al., 2007
Acid Green 25 Crab shell 645.1 Adsorption 24 h Wong et al., 2004a,b
Acid Green 27 Nanoparticle 2103.6 Electrostatic interaction 24 h Hu et al., 2006
Acid Orange 7 Bead Crosslinking 1940 Chemisorption 6h Chiou et al., 2004
Reactive Red 141 Shrimp shell 156 Chemical and 24 h Sakkayawong et al., 2005
physical adsorption
Reactive Blue 222 Flake 199 Chemisorption 480 min Wu et al., 2001
Reactive Red 222 Bead (crab) 1106 Intraparticle diffusion 3d Wu et al., 2000
Reactive Red 222 Bead (shrimp) 1026 Intraparticle diffusion 3d Wu et al., 2000
Reactive Red 222 Bead (lobster) 1037 Intraparticle diffusion 3d Wu et al., 2000
Reactive Red 222 Flake (shrimp) 494 Intraparticle diffusion 3d Wu et al., 2000
Reactive Red 222 Flake (lobster) 398 Intraparticle diffusion 3d Wu et al., 2000

4.1.2. Process variables and solution chemistry
Increasing chitosan dose had a dramatic positive impact on color
removal and there was an approximately linear relationship
between chitosan dose and color removal of the dye (Crini et al.,
2008a). The amount of the dye adsorbed onto chitosan was found
to increase with an increase in the initial concentration of dye
solution if the amount of adsorbent was kept unchanged (Park
et al., 1995; Knorr, 1983). At low initial concentration there is
possibility of the formation of monolayer coverage of the molecules
at the outer interface of the chitosan. At higher concentrations,
however, the number of available adsorption sites becomes lower
and subsequently the removal of dyes depends on the initial
concentration. The diffusion of exchanging molecules within chi-
tosan particles may govern the adsorption rate at higher initial
concentrations. Contact time is another important factor and most
authors seem to agree on a figure in the range 3e5 days for most
dye molecules (Crini and Badot, 2008). The contact time and
adsorption rate are dependent on the initial dye concentration.
Gibbs et al. (2003) observed that increasing the initial dye
concentration increased the time required to achieve complete
recovery of the dye. It has been observed that the equilibrium time
increases with the crosslinking ratio (Cestari et al., 2004; Chiou and
Li, 2003; Kim and Cho, 2005). It has been observed that the
molecular size of the dye was a major factor in adsorption char-
acteristics. Smith et al. (1993) noted that small, low molecular
weight dyes adsorbed best on chitosan.
The ability of the anionic dyes to adsorb onto chitosan beads is
often attributed to the surface charge which depends on the pH of
the operating batch system. Chitosan is polycationic in acidic
medium; the free amino groups are protonated and the polymer
becomes fully soluble and this facilitates electrostatic interaction
between chitosan and the negatively charged anionic dyes.
Decreasing the pH makes more protons available to protonate the
amine group of chitosan with the formation of a large number of
cationic amines. This results in increasing dye adsorption by chi-
tosan due to increased electrostatic interactions. Gibbs et al. (2004)
noted that, at low pH, chitosan’s free amino groups are protonated,
causing them to attract anionic dyes, demonstrating that pH is one
of the most important parameters controlling the adsorption
process. The free amine groups in chitosan are much more reactive
and effective for chelating pollutants than the acetyl groups in
chitin, and there is no doubt that amine sites are the main reactive
groups for (anionic) dye adsorption, though hydroxyl groups may
contribute to adsorption. The optimum pH is frequently reported in
the literature to be around pH 3.0e6.0.
4.1.3. Pretreatment
Treatment of chitosan with acid produces protonated amine
groups along the chain and this facilitates electrostatic interaction
between polymer chains and the negatively charged anionic dyes,
as previously observed by Maghami and Roberts (1988). Chemical
grafting of chitosan with specific ligands has been advantageous in
removing dyes (Jayakumar et al., 2002). It is known that the only
class for which chitosan (Chao et al., 2004) and crosslinked chitosan
(Hebeish et al., 2004) have low affinity are basic (cationic) dyes.
Crini and Badot (2008) and Crini et al. (2008b) suggested the use of
N-benzyl mono- and disulfonate derivatives of chitosan in order to
enhance its cationic dye hydrophobic adsorbent properties and to

Table 7
Dye decolorization by algal biomass.

Culture Dye Percent removal Mechanism Time Reference

Spirogyra sp.
Chlorella vulgaris
Enteromorpha prolifera
Azolla filiculoides
Caulerpa scalpelliformis
Cosmarium sp.
Azolla rongpong
Azolla filiculoides
Synazol (red 0.22%, yellow 0.1%) 85 18 h Khalaf, 2008
Remazol Black-B, 800 mg/L 52.4 Physical adsorption 24 h Aksu and Tezer, 2005
Acid Red 274, 250 mg/L 96.4 Surface adsorption and 120 min Ozer et al., 2005
intraparticle diffusion
Acid Red 88, 1000 mg/L 43.6 5e6 h Padmesh et al., 2005
Acid Green 3, 1000 mg/L 53.4
Acid Orange 7, 1000 mg/L 43.8
Basic Yellow, 150 mg/L 90 Physical adsorption 4h Aravindhan et al., 2007
Malachite Green, 10 ppm 74 Biodegradation 210 min Dhaneshwar et al., 2007
Acid Green 3, 1000 mg/L 30.7 Biosorption 12 h Padmesh et al., 2006a
Acid Blue 15, 1000 mg/L 43.8 12 h Padmesh et al., 2006b
 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929 1925

Table 8 such as hydroxyl, carboxylate, amino and phosphate found on the

Dye biosorption by peat. algal cell surface are considered to be responsible for sequestration
Dye Biosorption Reference of contaminants from wastewater. The dye removal, especially by
capacity (mg/g) using algae, may be attributed to the accumulation of dye ions on
Basic Blue 9 9.9 Ramakrishna and the surface of algal biopolymers and further to the diffusion of the
Viraraghavan, 1997 dye molecules from aqueous phase onto the solid phase of the
Acid Blue 29 8.6 Ramakrishna and
biopolymer (Ozer et al., 2006). Extracellular polymers consist of
Viraraghavan, 1997
Acid Red 91 0.6 Ramakrishna and surface functional groups, which enhance sorption of the dye
Viraraghavan, 1997 molecules onto the surface of the polymer (floc) during dye
Disperse Red 1 9.0 Ramakrishna and removal process (Mohan et al., 2002; Shukla et al., 2002). The
Viraraghavan, 1997 released metabolic intermediates (long chain biopolymers) which
Basic Blue 3 375 Allen et al., 1994
Basic Red 22 314 Allen et al., 1994
have excellent coagulation capacity along with the dye remaining
Basic Blue 3 390 Allen et al., 1988 in the aqueous phase tend to adsorb onto the surface of the poly-
Basic Yellow 21 300 Allen et al., 1988 mers and settle (biocoagulation) (Mohan et al., 2002). The infor-
Basic Red 22 240 Allen et al., 1988 mation on use of algae to decolorize dye wastewater is given in
Acid Blue 25 8.9e16.3 Poots et al., 1976
Table 7. Mohan et al. (2002) studied the removal of Reactive
Yellow 22 dye by active Spirogyra sp. and reported that the dye
improve its selectivity. Researchers have suggested use of chitosan removal mechanism can be explained by biosorption, bioconver-
chemically modified with succinic anhydride (Lima et al., 2006), sion and biocoagulation. While, removal of Acid Red 274 dye using
enzymatic grafting of carboxyl groups onto chitosan (Chao et al., inactivated Spirogyra rhizopus system was attributed to biosorption
2004), novel chitosan-based materials by reacting chitosan with and biocoagulation (Ozer et al., 2006).
a higher fatty acid functionalized with a glycidyl moiety in order to A study by Ozer et al. (2006) revealed the potential ability of the
introduce long aliphatic chains (Shimizu et al., 2005) to adsorb algae S. rhizopus to decolorize synthetic wastewaters containing initial
dyes. Carboxymethylated chitosan is a rather better adsorbent than concentration of dye Acid Red 274 from 25 to 1000 mg/L. Almost
raw chitosan for acidic dyestuffs (Uzun and Guzel, 2005). Raw complete removal of Acid Red 274 dye from synthetic wastewater
chitosan powders have disadvantages such as unsatisfactory with final concentration lower than 25 mg/L was achieved by using S.
mechanical properties, poor heat resistance and are soluble in rhizopus resulting from biocoagulation and biosorption.
acidic media, which can be overcome by developing crosslinked
chitosan beads. After crosslinking, these materials maintain their 5.1. Factors affecting algal decolorization
properties and original characteristics (Cestari et al., 2005),
particularly their high adsorption capacity, although this chemical Dhaneshwar et al. (2007) observed that for microalga Cosma-
modification results in a decrease in the density of free amine rium sp. an increase in pH from 4.0 to 6.0 led to a threefold increase
groups at the surface of the adsorbent in turn lowering polymer in decolorization rate of Malachite Green, which reached
reactivity towards metal ions (Gibbs et al., 2004). In general, the a maximum value of 92.4% at the pH of 9.0. Similarly, Aravindhan
adsorption capacity depends on the extent of crosslinking and et al. (2007) observed that uptake of Basic Yellow dye by Caulerpa
decreases with an increase in crosslinking density. scalpelliformis increased from 17 to 27 mg/g for an increase in pH
from 3.0 to 8.0. At lower pH, the Hþ ions compete effectively with
dye cations, causing a decrease in color removal efficiency. At
5. Algae higher pH, the surface of biomass gets negatively charged, which
enhances the positively charged dye cations through electrostatic
Algae have been found to be potential biosorbents because of force of attraction. However, Aksu and Tezer (2005) observed
their availability in both fresh and saltwater. The biosorption maximum uptake of different reactive dyes at pH 2.0 and then
capacity of algae is attributed to their relatively high surface area declined sharply with further increase in pH. It is expected that
and high binding affinity (Donmez and Aksu, 2002; Tien, 2002). nitrogen containing functional groups such as amines or imada-
Cell wall properties of algae play a major role in biosorption; zoles in the biomass will also be protonated at acidic pH values.
electrostatic attraction and complexation are known to take place Higher uptakes obtained at lower pH values may be due to the
during algal biosorption (Satiroglu et al., 2002). Functional groups electrostatic attractions between these negatively charged dye

Table 9
Adsorption capacities of biosorbents and activated carbon.

Dye Adsorbent Adsorption capacity Adsorption capacity of Reference

of biosorbents (mg/g) activated carbon (mg/g)
Reactive Blue 2 Chitosan 2498 217 Chiou et al., 2004
Reactive Red 2 Chitosan 2422 712 Chiou et al., 2004
Direct Red 81 Chitosan 2383 241 Chiou et al., 2004
Reactive Yellow 86 Chitosan 1911 127 Chiou et al., 2004
Direct Red 28 Dead fungi (A. niger) 14.7 16.8 Fu and Viraraghavan, 2002a. b
Reactive Red 5 Alga 556 278 Aksu and Tezer, 2005
Basic Blue 9 Peat 9.9 6.4 Ramakrishna and
Viraraghavan, 1997
Acid Blue 29 Peat 8.6 10 Ramakrishna and
Viraraghavan, 1997
Acid Red 91 Peat 0.6 9.1 Ramakrishna and
Viraraghavan, 1997
Disperse Red 1 Peat 9.0 4.9 Ramakrishna
and Viraraghavan, 1997
Astrazone Blue basic dye Yeast (S. cerevisiae) 70 18.5 Farah et al., 2007
1926 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929
anions and positively charged cell surface. Hydrogen ion also acts as
a bridging ligand between the alga cell wall and the dye molecule.
Aksu and Tezer (2005) observed maximum biosorption capacity at
35 " C and Ozer et al. (2005) obtained maximum biosorption
capacity at 30 " C, both indicating that the process is exothermic.
6. Peat
Peat can be described as partially fossilized plant material that
occurs in wet areas where there is a lack of oxygen (Viraraghavan and
Mihial, 1995). It is a complex material, with lignin and cellulose as
major constituents (Viraraghavan and Rana, 1991; Rana and
Viraraghvan, 1987). The polar functional groups of lignin, which
include alcohols, aldehydes, ketones, acids, phenolic hydroxides and
ethers are involved in the formation of chemical bonds. The lignin
and humic fractions contain predominantly p-hydroxyl groups
(Couillard, 1994). Because of their polar character, the specific
adsorption for dissolved solids such as transition metals and polar
organic molecules is reported to be high (Coupal and Lalancette,
1976). Partially decomposed peat has a porosity of approximately
95% and a specific area of 200 m2/g. Thus, peat is a highly polar and
porous material. The pH influences the structure and properties of
peat. Peat normally has a pH of around 4.0, due to the presence of
humic acids. The structure of peat degrades at pH > 9.0; below pH
3.0, its chelating capacity decreases. Natural peat may be used for the
removal of contaminants without further pretreatment, but has
certain disadvantages; low mechanical strength, a high affinity for
water, poor chemical stability and a tendency to shrink and/or swell.
To overcome these problems, thermal or chemical pretreatment has
been employed. Chemical pretreatment of peat using acids such as
phosphoric and sulfuric have been employed (Couillard, 1994). Sun
and Yang (2003) found use of the modified peat-resin particle by
mixing modified-peat with polyvinylalcohol (PVA) and formalde-
hyde to be effective in removing basic dyes.
Adsorption of basic and acidic dyes onto peat has been studied
(Allen et al., 1994; Viraraghavan and Mihial, 1995; Ramakrishna and
Viraraghavan, 1997) and the information is presented in Table 8.
The adsorption of basic dyes onto peat from single component and
multi-component solutions have been reported by Allen et al.
(1988, 2004). The adsorption was presented in the form of equi-
librium isotherms and the Freundlich, Langmuir and Red-
lichePeterson isotherm equations were fitted to the results and the
isotherm constants were obtained. The mechanism of the sorption
process may be explained as a chemical reaction in which the
functional groups of peat form a cation exchange reaction, due to
the high cation exchange capacity of the peat. The presence of polar
functional groups causes the peat surface to be negatively charged
and hence they have a high adsorption capacity for cationic (basic)
dyes. The maximum affinity for these dye cations can be expected
at higher pH values, because of fewer anionic adsorption sites, at
lower pH values. Ramakrishna and Viraraghavan (1997) observed
a maximum dye adsorption for Basic Blue 9 on peat at a pH of 6e7
with no significant increase in removals beyond pH 7. Similar
observations were made by Viraraghavan and Mihial (1995). Ho
and McKay (1998) observed that degree of agitation, initial dye
concentration and temperature influences the sorption rate of dyes
on the peat surface. The sorption of dyes, namely, Basic Blue 69 and
Acid Blue 25 onto peat was favoured at higher concentrations of dye
solution, and high temperatures for Basic Blue 69. However in the
case of Acid Blue 25, sorption was favoured at lower temperatures.
7. Summary of review
It is clear from the literature that biosorbents have the potential
to remove a wide variety of dyes. The biosorption process will
provide an attractive alternative if manufactured biosorbents are
available and economical. Pretreatment techniques such as auto-
claving, drying, chemical treatment and crosslinking are required to
improve the biosorption capacity of the adsorbent. There is a need
to develop biosorbents in a simple, inexpensive way and have
a high biosorption capacity. Decolorization by living cells involves
complex mechanisms and dependant on operational conditions,
nutrient requirements and toxicity. The use of non-viable fungi,
bacteria or algae is easier and effective. The effectiveness of bio-
sorption depends on the characteristics of the adsorbent, adsor-
bate, process variables and solution chemistry. Several pollutants
would co-exist in an industrial effluent and performance of bio-
sorbents to decolorize wastewater under actual industrial condi-
tions needs to be investigated (Crini, 2006). Also, complex structure
of dye molecules influences biosorption and further research is
needed to establish the relationships between dye molecule
structure and biosorption (Fu and Viraraghavan, 2001a). The
differences in the physical and chemical charecteristics of the
adsorbent and their dependence on process variables and solution
chemistry make it difficult to compare between one another. Table
9 compares the adsorption capacities for activated carbon and
other biosorbents. Adsorption capacities of chitosan and S. cer-
evisiae were higher than that of activated carbon while the
adsorption capacity of fungus (A. niger) was comparable to that of
activated carbon. Of all the biosorbents reviewed, chitosan, algal
and fungal biomass were found to have excellent biosorption
capacities.
8. Application to practice
Earlier reviews on biosorption (Fu and Viraraghavan, 2001a;
Gupta and Suhas, 2009; Kaushik and Malik, 2009; Gadd, 2009)
have shown that a lot of scientific information is already available
on the use of a number of biosorbents for dye removal; however no
detailed economic and market analyses are available. There is still
reluctance in opting for non-viable immobilized biomass-based
systems compared to the use of biological reactors with living cells,
although advantages of the former systems are well established.
There have been many attempts in the past at commercializing
immobilized biomass biosorbents such as alga_SORB, AMT-bio-
claim, B.V. Sorbex’s biosorbents and Bio-fix, but none have made
a successful commercial entry in the market (Tsezos, 2001; Wase
and Forster, 1997; Wang and Chen, 2009). Peat is considered as
the most successful biosorbent in use either in natural state or in
a modified form; however it is not regarded as the best biomaterial
for commercialization because it is a finite resource and it is also
not available everywhere in the world (Wase and Forster, 1997).
The challenge with biosorption as with many in the biotech-
nology industry is to move this process to an industrial scale. It is
relatively less difficult to demonstrate it in a laboratory; it is a little
more challenging to demonstrate it at a pilot scale, but to really
scale it up to a large scale would call for a significant financial and
technological effort. This mismatch between scientific progress in
biosorption research (biosciences) and stagnation in industrial
biotechnology innovation needs to be corrected through trans-
lational research and technology transfer with a push for
commercialization of research. Universities can play an active role
in this process through more formalized approach to technology
transfer and protection of intellectual property (West and
Nightingale, 2009).
9. Conclusions
Biosorbents capable of decolorizing dye wastewater have been
reviewed. The presence of variety of functional groups in the
 A. Srinivasan, T. Viraraghavan / Journal of Environmental Management 91 (2010) 1915e1929
biosorbents makes them selective and highly capable of bio-
degrading and biosorbing dyes from wastewater. The extensive
research conducted on various biosorbents show that they are
emerging as a promising alternative to conventional treatment
systems. Algal and fungal biomasses and chitosan have shown
excellent color removal capabilities. However, use of biosorbents to
remove color in a dye wastewater is still in the research stage. Efforts
are needed to commercialize this research through (a) selection of
suitable biosorbents based on economic and market analysis, (b)
pilot-scale studies with actual wastewaters and (c) full-scale
demonstration systems.
Acknowledgements
The authors would like to thank the Natural Sciences and
Engineering Research Council of Canada for partial funding through
a grant to the second author.
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