Food Hydrocolloids 24 (2010) 659e666

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Food Hydrocolloids
journal homepage: www.elsevier.com/locate/foodhyd

Flow behaviour, linear viscoelasticity and surface properties of chitosan

aqueous solutions
Nuria Calero, Jos Muoz*, Pablo Ramrez, Antonio Guerrero
Departamento de Ingeniera Qumica, Facultad de Qumica, Universidad de Sevilla c/P, Garca Gonzlez, 1, E41012 Sevilla, Spain

a r t i c l e i n f o

a b s t r a c t

Article history:
Received 11 December 2009
Accepted 18 March 2010

A study of rheological properties of chitosan solutions was carried out in terms of polymer concentration
(0.05e2.5 wt%) and temperature (5e25
C).
The chitosan solutions studied exhibited Newtonian behaviour at concentrations lower than 0.25 wt%.
Above this value, the rest of the systems showed shear-thinning behaviour.
The viscoelastic properties of chitosan solutions above 0.25 wt% were characterized by oscillatory
shear measurements under small-deformation conditions and the results showed uid-like viscoelastic
behaviour.
Time-concentration and time-temperature superposition methods were carried out to obtain an
overall master curve; where the rheological properties of chitosan solution at a specic temperature or
concentration can be predicted over a broad time scale.
The fact that the activation energy for steady shear ow was higher than for oscillatory shear
promoted deviations from the CoxeMerz rule, when temperature was decreased at the highest chitosan
concentration studied
In addition, their dynamic surface tension behaviour and surface viscoelastic properties were explored.
The time-dependent decrease in surface tension along with the limiting elasticity of the interface seems
to indicate that commercial chitosan may play the role of an emulsier as well as a stabiliser of O/W
emulsions.
2010 Elsevier Ltd. All rights reserved.

Keywords:
Chitosan solution
Viscoelastic properties
Flow properties
Surface viscoelasticity
Superposition methods

1. Introduction
Chitin is one of the most widespread of the naturally occurring
polysaccharides, specically the second most widespread, after
cellulose (Rebecca, Queen, Coughlin, & Krause, 2008). Chitin is
industrially produced through the use of chemical or enzymatic
treatments of crustacean shell waste, but is also found in mollusks,
insects, fungi and related organisms (Hudson & Jenkins, 2001).
However, its utility is limited in industrial applications due to its
poor solubility, which is provoked by the rigidity of its linear chains.
Nevertheless, chitosan, which is produced from chitin by Ndeacetylation and depolymerization reactions, is soluble in water
under acidic conditions because of its polycationic nature when its
degree of deacetylation is higher than 65e70%.
The main reason for the growing interest in chitosan is its
promising intrinsic properties (Berger et al., 2004). Indeed, chitosan
is nontoxic, biocompatible, biodegradable and bioactive (Akbuga,

* Corresponding author. Tel.: 34 954557179; fax: 34 954554567.

E-mail address: jmunoz@us.es (J. Muoz).
0268-005X/$ e see front matter 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.foodhyd.2010.03.009

1995). These properties merit further study for medical applications (Felt et al., 1999; Patashnik, Rabinovich, & Golomb, 1997) and
for the production of controlled drug-delivery systems (Diez-Sales,
Dolz, Hernndez, Casanovas, & Herraez, 2007; Li & Xu, 2002; Senel
et al., 2000; Thacharondi & Panduranga-Rao, 1995). Chitosan has
been reported to promote wound healing (Ueno, Mori, & Fujinaga,
2001) and to have bacteriostatic effects (Liu, Guan, Yang, Li, & Yao,
2001). Further applications have been reported as occurring in
agriculture, water & waste treatment and in cosmetics and toiletries (Rinaudo, 2006).
Chitosan is also known to possess numerous physiological
properties, such as prebiotic and fat binding capacity. It is also able
to reduce cholesterol and to favour loss of body weight by the
reduction of lipid absorption (Borderas, Snchez-Alonso, & PrezMateos, 2005; Muzzarelli, 2009)
Many food products contain both polysaccharides and proteins.
Proteins have an essential role as emulsifying and stabilizing agents,
whereas polysaccharides are mainly used as thickening and waterholding agents. The overall stability and texture of food colloids
depend on the functional properties of their ingredients and the
nature and strength of the protein-polysaccharide interactions

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N. Calero et al. / Food Hydrocolloids 24 (2010) 659e666

solution was pipetted into the capillary viscometer, which

was equilibrated at 25
C in a water bath for 30 min prior to
measurements.
Specic viscosity was determined using:

(Dickinson, 1995). Chitosan is one of the polysaccharides used as

a stabiliser of food emulsions (Laplante, Turgeon, & Paquin, 2006;
Ogawa, Decker, & McClements, 2003), and due to their aforementioned physiological properties, its use contributes a value-added
benet to the nal emulsion. Furthermore, chitosan solutions have
been reported to show surface activity properties (Babak &
Desbrieres, 2004; Gargallo et al., 2004), which may enhance both
the formation and stability of emulsions. In addition, according to
the general proteinepolysaccharide interaction scheme described
by McClements (2005), mixtures of chitosan and protein above its
isoelectric point may form a complex that would result in enhanced
physical stability.
In general, commercially available chitosan samples exhibit
different functional properties due to variations in their degree of
deacetylation (range from 60 to 90%) and molecular weight (range
from 50 to 2000 kDa) (Cho, No, & Meyers, 1998; Del blanco,
Rodriguez, Schulz, & Agullo, 1999). The wide variety in the degree
of deacetylation and molecular weight found in commercial chitosan can be attributed to the fact that the N-deacetylation and
depolymerization reactions are rather sensitive to industrial processing variables.
For this reason, understanding the solution behaviour of
commercial chitosan is essential to predict, design, and characterize most of its applications. In particular, the linear viscoelastic
properties of aqueous solutions of chitosan are closely related to
their colloidal structure while the shear rate dependence of
viscosity to the structural modications under shear.
The main objective of this work was the study of bulk rheological properties of chitosan solutions in order to assess its
potential applications as a thickener and an emulsion stabiliser. A
further goal was to explore their possible surface activity; namely,
their dynamic surface tension behaviour and surface viscoelastic
properties, which are quite important in regards to their contribution to the emulsifying capacity when formulated with proteins.
To our knowledge this is the rst paper where results on the surface
viscoelastic properties of chitosan are reported.

The Haake Rheostress RS100 and MARS rheometers (ThermoScienticr, Germany), equipped respectively with K10 and C5P
Phoenix circulators (Thermo-Scientic, Germany) were used for
sample temperature control.
The oscillatory tests for chitosan solutions with different
concentrations (0.25e2.5) wt% at 25
C were carried out using
a double-cone (60 mm diameter, 1
angle) or a cone & plate (60 mm
diameter, 4
angle) geometry, depending on the consistency of the
chitosan solution. In addition, the inuence of temperature (5
C,
10
C, 15
C, 20
C and 25
C) was studied at a chitosan concentration of 2.5 wt%, using the aforementioned cone & plate sensor.
All measurements were repeated three times with each solution. Stress sweeps at a frequency of 1 Hz were performed for all
systems studied to estimate the linear viscoelastic range.
Steady state shear ow tests for chitosan solutions with
different concentrations were carried out at 25
C using a geometry
of concentric cylinders (Z40 DIN sensor system, Rj 20 mm,
Ra 21.7 mm) or the cone & plate sensor, depending on the solution viscosity. Meanwhile, chitosan solutions at different temperatures (ranging from 5
C to 25
C) were measured with the cone &
plate geometry.

2. Materials and methods

2.5. Surface tension and surface viscoelasticity measurements

2.1. Materials

Dynamic surface tension measurements for a 0.5 wt% CH solution were carried out with a pendant drop tensiometer (CAM 200
model, from KSV, Finland) at 25
C. In addition, its surface viscoelastic properties were studied by means of step perturbation tests.
First, the drop volume was increased by approximately 4 mL in ca.
0.5 s and the drop prole was recorded at high speed (0.1 s per
frame) for 20e25 s. Then, a reverse step of equal volume was made
and the surface tension was monitored for a given time.

Chitosan (CH) medium molecular weight (MW 190,000e

310,000 Da, 75e85% deacetylated). Anhydrous sodium acetate and
glacial acetic acid were used as supplied by SigmaeAldrich (St. Louis,
USA).
2.2. Preparation of chitosan solutions
Chitosan solutions with different concentrations (0.05%, 0.1%,
0.15%, 0.2%, 0.25%, 0.5%, 1%, 1.5%, and 2.5%, w/w) were prepared by
dissolving the proper amount of chitosan in a buffer solution (0.2 M
acetic acid and 0.05 M sodium acetate, pH 3.90). This pH value is
appropriate to enhance the microbiological stability of food stuffs.
The solution was stirred using an Ika-visc MR-D1 homogenizer, for
3 h at 700 rpm.
The system was left to stand for 24 h at 4
C for complete
hydration of the polymer and removal of bubbles.
0.1 wt% sodium azide was added to the solutions to prevent the
growth of microorganisms.
2.3. Intrinsic viscosity
The viscosity of diluted chitosan solutions (from 0.05 wt%
to 0.2 wt%) were measured with an Ubbelohde glass capillary
viscometer (Proton; size Inc, Barcelona, Spain) at 25
C. A volume of

hsp hsolution  hsolvent =hsolvent :

(1)

Intrinsic viscosity, [h], is dened as the extrapolation of the reduced

viscosity, hred, to zero polymer concentration:

h

hsp =C


C/0

hred C/0 ;

(2)

2.4. Rheological characterization

3. Results and discussion

3.1. Flow behaviour
The intrinsic viscosity of the chitosan sample used in this
research turned out to be 12.15 dl/g at 0.2 M acetic acid and 0.05 M
sodium acetate and 25
C. This value is of the same order of
magnitude as those available in the literature (Hwang & Shin, 2000;
Kasaai, Arul, & Charlet, 2000; Lapasin & Pricl, 1995), which obviously depends on the degree of deacetylation, molecular weight,
temperature and solvent used. The intrinsic viscosity is dened as
the hydrodynamic volume occupied by a gram of an isolated
polymer in a given solvent. However the coil overlap parameter,
dened as the dimensionless product (C$h), is more frequently
used to indicate the volume occupied by the polymer molecule and
to estimate the so-called overlap concentration, which is the critical
polymer concentration (C*) locating the transition from the dilute

N. Calero et al. / Food Hydrocolloids 24 (2010) 659e666

661

100000

C[ ]
2
C[ ]+k'(C[ ])

10000

sp

1000

100

10

0,1
0,1

10

100

C[ ], C[ ]+k'(C[ ])

1000

Fig. 1. Dependency of the specic viscosity (hsp) on C[h] and C[h]k0 (C[h])2. T 25
C.
The continuous lines are guides for eyes.

to semi-dilute regions. C * $h is the space occupancy related to the

polymer concentration at the onset of coil overlapping and can be
estimated from the logelog plot of the specic viscosity vs. the coil
overlap parameter C$h (Fig. 1). The value of C * $h was estimated
to be around 3.1, which is similar to that found by Hwang and Shin
(2000). However, this value is lower than the average of 4.0
reported by Morris, Cutler, Ross-Murphy, Rees, and Price (1981) for
random-coil polysaccharides. The resulting value of C* was 0.26 g/
dl, which was supported by the value for which the dependence of
the specic viscosity on C$h kH C$h2 deviated from the slope
of 1 (Fig. 1) (Desbrieres, 2002).
The inuence of C$h on the specic zero shear viscosity followed a power-law equation with an exponent of 1.3 in the dilute
regime and 4.7 in the semi-dilute regime. In the latter regime the
exponents derived from theoretical treatments have been found to
vary between 3.75 and 4.25 (Drobynin, Colby, and Rubinstein,
1995). The exponent of the sample corresponding to this study in
the dilute regime is very similar to that reported by Hwang and
Shin (2000) for chitosan solutions. Congruently, the exponent of
the semi-dilute regime is in between the values reported by Cho,
Heuzey, Begin, and Carreau (2006) (4.1) and by Desbrieres (2002)
(5.2), who attributed the high value observed to the occurrence
of a large number of entanglements caused by hydrogen bonds and
hydrophobic interactions. However, it must be noted that Hwang
and Shin (2000) found a lower value (3.94). The value obtained in
our study coincides with that reported by Sittikijyothin, Torres, and
Gonalves (2005) for galactomannan gums, which form entanglement networks in aqueous solutions, much like chitosan. These
authors attributed this high value (4.7) to the high rigidity of the
backbones of the gums used. Morris, Cutler, Ross-Murphy, Rees, and
Price (1981) and Launay, Doublier, and Cuvelier (1986) found values
up to 5.1 for galactomannans.
Fig. 2 shows the ow properties for chitosan aqueous solutions as
a function of concentration (Fig. 2a) and temperature (Fig. 2b). The
system with the lowest chitosan content showed a Newtonian
behaviour within the experimental shear rate. The rest of the
systems studied exhibited a marked Newtonian region at low-shear
rate regime, followed by a decrease in viscosity (shear-thinning
behaviour) above a critical shear rate. Fig. 2 also illustrates the tting
quality of the results obtained to the Cross model (R2 > 0.996).

h0
1 g_ =g_ c p

(3)

Fig. 2. a. Steady shear ow curves as a function of chitosan concentration in the

(0.25e2.5) wt% range at 25
C. Continuous lines illustrate data tting to the Cross
model. CH stands for chitosan. b. Steady shear ow curves as a function of temperature
in the (5e25)
C for a solution with 2.5 wt% chitosan. Continuous lines illustrate data
tting to the Cross model.

where ho is the low-shear Newtonian viscosity, g_ c is the critical

shear rate for the onset of shear-thinning response and p is
a parameter related to the slope of the power-law region.
The values of these parameters are shown in Tables 1 and 2 as
a function of concentration and temperature, respectively. The lowshear Newtonian viscosity, ho, showed an exponential growth with
chitosan concentration (equation (4)).



c  cref
0:34

h0 0:132$exp

(4)

Table 1
Flow curve tting parameters for the Cross model. T 25
C.

h0 (Pa s)
g_ c (1/s)
p

2.5 wt%

1.5 wt%

1.0 wt%

0.5 wt%

0.25 wt%

10.8
13.8
0.63

0.86
148
0.77

0.15
643
0.76

0.09
1130
0.76

0.007 (a)
e
e

Standard deviation of the mean (3 replicates) for h0 < 3%.

Standard deviation of the mean (3 replicates) for g_ c < 3:5%.
Standard deviation of the mean (3 replicates) for p < 3.5%.
a
Newtonian behaviour along the whole shear rate range covered.

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N. Calero et al. / Food Hydrocolloids 24 (2010) 659e666

Table 2
Flow curve tting parameters for the Cross model for a solution with 2.5 wt% of
chitosan.

h0 (mPa s)
g_ c (1/s)
p

25
C

20
C

15
C

10
C

5
C

10.8
13.8
0.63

14.4
8.4
0.61

19.6
5.1
0.55

25.2
3.5
0.60

31.8
2.6
0.57

Standard deviation of the mean (3 replicates) for h0 < 3%.

Standard deviation of the mean (3 replicates) for g_ c < 3:5%.
Standard deviation of the mean (3 replicates) for p < 3.5%.

where c stands for chitosan concentration (wt%); cref stands for

a chitosan concentration of reference (1 wt%).
As for the inuence of temperature, ho followed an Arrheniustype equation, leading to an activation energy for the viscous ow
of 34.9 kJ/mol.

h0 105 exp

4191
T


(5)

A typical behaviour was found for the critical shear rate since it
decreased with chitosan concentration and conversely increased
with temperature. The parameter p values were not markedly
inuenced by temperature; hence an average value of 0.59 could be
calculated. However, a decrease in chitosan concentration led to
a slightly higher p value that did not show any signicant variation
in the 0.5 wt% to 1.5 wt% range. It should be taken into account that
the experimental ow properties for these systems are still far from
the fully developed power-law decay ow region. Thus, the aforementioned average value may also be used to obtain reasonable
ttings at the lowest chitosan concentrations. This fact suggested
that an overall master ow curve, including the effects of concentration and temperature, could be obtained by normalizing the
apparent viscosity with ho and using a horizontal shift factor which
can be estimated from the values of the critical shear rate. As Fig. 3
shows, all the ow curves collapsed fairly well into the master
curve corresponding to 2.5 wt% chitosan at 25
C.
The values for the time-concentration and timeetemperature
shift factors are shown in Table 3.
The values of ac tted an exponential growth function as
follows:

(6)

2,0

lg /

1,0

aC
aT

2.5 wt%

1.5 wt%

1.0 wt%

0.5 wt%

0.25 wt%

1.00
25
C
1.00

0.10
20
C
1.77

0.02
15
C
2.86

0.01
10
C
4.37

0.007
5
C
6.68

where c stands for chitosan concentration (wt%); cref stands for

a chitosan concentration of reference (1 wt%).
As for the dependence of aT with temperature, an Arrheniustype equation was found:



7779
T

aT 4:97  1012 exp

(7)

The activation energy derived from this equation was 64.7 kJ/mol.
It is interesting to note, from a practical point of view, that the
master ow curve calculated by superposition methods provided
information below and above the experimental shear rate range
covered. Superposition of ow curves determined at different
chitosan concentrations enabled a more precise denition of the
low-shear Newtonian zone to be obtained. In addition, the superposition of ow curves at different temperatures provided the
information required to gain information at high shear rates.
Obtaining information at high shear rate is quite relevant for some
applications but cone and plate geometries cannot give reliable
experimental data at such high shear rates due to migration of
sample out of the sensor system.
The fact that all the ow curves obtained in the (0.25e2.5) wt%
and (5e25)
C ranges superimposed revealed that the mechanism
governing the shear-induced disentanglement of chitosan macromolecules in the solution conditions used was quite similar in all
cases. However, conclusions on a possible simple thermo-rheological behaviour, indicating the lack of conformational transitions
or formation of supramolecular structures must be drawn from
small amplitude oscillatory shear (SAOS) experiments better than
ow curve tests.
3.2. Small amplitude oscillatory shear (SAOS)



c  cref
ac 0:029 exp
0:43

1,5

Table 3
Values of the timeeconcentration and timeetemperature shift factors used to obtain
the overall master ow curve of a 2.5 wt% chitosan solution at 25
C.

25C
20C
15C
10C
5C

0.25 wt% CH
0.5 wt% CH
1 wt% CH
1.5 wt% CH
2.5 wt% CH

0,5
0,0
-0,5
-1,0
-1,5
-2,0
-3,5 -3,0 -2,5 -2,0 -1,5 -1,0 -0,5

0,0

0,5

1,0

1,5

2,0

2,5

3,0

-1

lg ( /s )+lg a
Fig. 3. Master ow curve at the reference concentration of 2.5 wt% chitosan and
reference temperature of 25
C. CH stands for chitosan.

Fig. 4 shows the mechanical spectra of chitosan solutions as

a function of their concentration (Fig. 4a) and temperature (Fig. 4b).
The storage modulus values are not shown at concentrations lower
than 1.5 wt% because they were below the accuracy limit of the
rheometer, which indicates the low viscoelastic response of these
solutions. These results reect a uid-like behaviour, typical of
random-coil entanglement networks, showing a tendency to reach
a crossover between G0 and G00 at high concentration and low
temperature. All the mechanical spectra shown in Fig. 4b tend to
reach similar characteristic low frequency slopes for G0 (and also for
G00 ) in the temperature range studied. The values of those slopes,
derived from the time-temperature master mechanical spectrum
(graph not shown) turned out to be 1.70 and 0.96, respectively,
which are close to those typical of a Maxwellian uid (Malkin and
Isaev, 2006)
Similar to the normalization analysis carried out for the ow
curves, a master curve for the frequency dependence of the
complex viscosity, including the effects of concentration and
temperature, is plotted in Fig. 5 which shows an excellent superposition of all the experimental values. It must be pointed out that
the values of ac used in this master curve are the same as those used
for the master ow curve (Table 3). The aT required to superimpose
the complex viscosity values are shown in Table 4.

N. Calero et al. / Food Hydrocolloids 24 (2010) 659e666

663

2,0

1000
G'

G''

10

25C
20C
15C
10C
5C

2.5 wt% CH
1.5 wt% CH
1 wt% CH
0.5 wt% CH
0.25 wt% CH

1,5

2.5 wt% CH
1.5 wt% CH
1 wt% CH
0.5 wt% CH
0.25 wt% CH

100

1,0
0,5

lg( / 0)

0,1

0,0

G', G''/Pa

-0,5

0,01
-1,0

1E-3
-1,5

1E-4
-2,0

1E-5
1E-3

-4,0 -3,5 -3,0 -2,5 -2,0 -1,5 -1,0 -0,5

0,01

0,1

10

100

/rad/s

100

GG/Pa

10

G', 25C
G', 20C
G', 15C
G', 10C
G', 5C

G'', 25C
G'', 20C
G'', 15C
G'', 10C
G'', 5C

0,01
0,01

0,1

0,5

1,0

1,5

2,0

2,5

Fig. 5. Master curve for the frequency dependence of the complex viscosity at the
reference concentration of 2.5 wt% chitosan and reference temperature of 25
C. CH
stands for chitosan.

1000

0,1

0,0

lg ( /(rad/s))+lg a

1000

10

100

1000

/rad/s
Fig. 4. a. Inuence of chitosan concentration on the mechanical spectra at 25
C. Shear
stress amplitude: 2 Pa (2.5 wt% chitosan), 1 Pa (1.5 wt% chitosan), 0.3 Pa (1 wt% chitosan), 0.15 Pa (0.5 wt% chitosan), 0.1 Pa (0.25 wt% chitosan). CH stands for chitosan. b.
Inuence of temperature on the mechanical spectra for a solution with 2.5 wt% chitosan. Shear stress amplitude: 2 Pa.

The Arrhenius-type temperature dependence of these values of

aT gives an activation energy of 31.7 kJ/mol, which is close to the
value of the activation energy calculated from the temperature
dependence of the low-shear Newtonian viscosity. This is in
agreement with the denition of aT for uncross-linked polymers of
low molecular weight in the frame of the modied Rouse theory
(Ferry, 1980). The aT values used for SAOS results (Table 4) did not
coincide with the previous values given for the master ow curve
(Table 3). This departure was also observed for the temperature
dependence of the shift factors as demonstrated by the different
activation energy values obtained. Thus, the value obtained from
ow properties (64.7 kJ/mol) is about twice the value calculated
from SAOS (31.7 kJ/mol). This can be related to the fact that the
critical shear rate which locates the onset of the power-law region
is lower than the corresponding critical frequency. In fact, the
values of the shift factors (ac and aT) used for ow curves are rather
coincident with the values of g_ c;ref =g_ c , where g_ c;ref stands for the
critical shear rate of the system selected as the reference (2.5 wt% at
25
C). A similar analysis can be made for the SAOS results. The
different values of aT and activation energy derived from ow and
SAOS properties were a rst indication that the CoxeMerz rule was
not fully followed as temperature decreased.

The assessment of the CoxeMerz rule as a function of chitosan

concentration and temperature is shown in Fig. 6a and b, respectively. It is interesting to note that the CoxeMerz rule was clearly
followed at 25
C in the concentration range studied, while it failed
at the higher shear rates for the 2.5 wt% solution when temperature
was decreased below 20
C. Departures from the CoxeMerz rule
have been attributed to different factors, such as the occurrence of
a) specic physical polymer interactions, leading to the formation
of high density entanglements (galactomannan hyperentanglements) (Morris et al., 1981); b) structured inter and intramolecular associations (Yu & Gunasekaran, 2001) c) aggregates and
ordered association structures, like liquid crystals (Lapasin & Pricl,
1995); d) physical and/or chemical interactions resulting in gellike structures or dispersions (Metzger, 2006), e) materials with
some structural heterogeneity (Rao, 2007). The increasingly higher
strain values that a sample undergo when conducting an upward
ow curve experiment is markedly greater than the strain values
applied in SAOS. A similar situation would take place when conducting a downward ow curve test. High strain values provoke
a severe disruption of the initial structured sample, which hardly
can fully recover in a short time. Conversely, samples which exhibit
predominant uid-like viscoelastic properties are prone to undergo
a fast stress relaxation process. The fail of the CoxeMerz rule seems
to be enhanced by the development of elastic properties in viscoelastic materials. The results obtained in the present study conrm
that the higher the elasticity (the longer the characteristic relaxation time) the greater departure from the rule is likely to occur as
also suggested by the results reported for concentrated dispersions
of Acacia tortuosa gum exudates (Muoz et al., 2007) and by the
behaviour of the so-called Boguer uid, a paradigmatic example of
liquid with marked elasticity (Al-Hadithi, Barnes, & Walters, 1992).
It must be noted that the inuence of temperature was analysed in
this study at the highest concentration tested (2.5 wt%), where
strong overlapping and compression among chitosan macromolecules will occur. The fail of the CoxeMerz rule at the lower

Table 4
Values of the timeetemperature shift factors used to obtain the master curve for the
frequency dependence of the complex viscosity at the reference concentration of
2.5 wt% and at the reference temperature of 25
C.

aT

25
C

20
C

15
C

10
C

5
C

1.00

1.26

1.58

2.00

2.51

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N. Calero et al. / Food Hydrocolloids 24 (2010) 659e666

a 1000

2,0
I I
0.25 wt% CH
0.5 wt% CH
1 wt% CH
1.5 wt% CH
2.5 wt% CH

10

lg( / 0), lg( */ 0)

, */Pas

100

I I

0.25 wt% CH
0.5 wt% CH
1 wt% CH
1.5 wt% CH
2.5 wt% CH

1,5
1,0
0,5

2.5 wt% CH
1.5 wt% CH
1 wt% CH
0.5 wt% CH
0.25 wt% CH

2.5 wt% CH
1.5 wt% CH
1 wt% CH
0.5 wt% CH
0.25 wt% CH

0,0

-0,5
0,1

-1,0
0,01

-1,5
1E-3

25C
20C
15C
10C
5C

-2,0
0,01

0,1

10

-1

100

1000

-4,0 -3,5 -3,0 -2,5 -2,0 -1,5 -1,0 -0,5 0,0 0,5 1,0 1,5 2,0 2,5 3,0

/s , /rad/s

-1

lg ( /s )+lg a; lg ( /(rad/s))+lg a
Fig. 7. Overall master curve from ow behaviour and SAOS measurements at the
reference concentration of 2.5 wt% chitosan and at the reference temperature of 25
C.
CH stands for chitosan.

100
*

I I

, */Pas

25C
20C
15C
10C
5C

master curve (Fig. 7) showed rheological information over a larger

range of shear rates or frequencies than could be studied using
a single rheometer for a 2.5 wt% chitosan solution at 25
C.
3.3. Surface properties of chitosan

1
0,1

10

100

-1

/s , /rad/s

Fig. 6. a. CoxeMerz plot at different chitosan concentrations at 25
C. CH stands for

chitosan. b. CoxeMerz plot at different temperatures for a solution with 2.5 wt%
chitosan.

temperatures may be attributed to the formation of high density

entanglements with a reduced contribution of Brownian forces to
counterbalance the effect of shear and to assist in the recovery
process after cessation of shear. Previous reported results supported that chitosan solutions followed the CoxeMerz rule up to
2 wt% (Martnez-Ruvalcaba, Cornet, & Rodrigue, 2004) and that the
opposite was true at a substantially higher concentration (4.17% w/
v) (Cho et al, 2006). Our results report a critical concentration
(2.5 wt%) for the onset of departures from the CoxeMerz rule.
Despite that the CoxeMerz rule was not obeyed below 20
C, it
was possible to obtain a reasonable superposition of all the curves
obtained from the ow behaviour and SAOS measurements,
including the effects of both chitosan concentration and temperature as demonstrated in Fig. 7. The ac values used in this general
master curve were the same as those used to superimpose the ow
curves (Fig. 3) and the frequency dependence of the complex
viscosity (Fig. 5). This result seems to indicate that the behaviour of
these chitosan solutions was not far from that of a thermo-rheologically simple material. Superposition of steady shear and SAOS
results as a function of chitosan concentration and temperature
provided a calculated extended ow curve for the 2.5 wt% solution
at 25
C. The shear rate and frequency ranges widened up to 7
decades, improving the information derived from either individual
series of ow curve or SAOS tests. Last but not least, the general

Fig. 8 shows a decrease in the surface tension value as a consequence of the adsorption of the chitosan sample on the airewater
surface at the experimental conditions. The surface activity of
chitosan macromolecules in aqueous solutions was attributed by
Schulz, Rodrguez, Del Blanco, Pistonesi, and Agull (1998) to their
amphiphilic nature on account of the relative amount of high
deacetylated degree molecules to that of low deacetylated degree.
A similar interpretation was used later by Babak, Auzely, and
Rinaudo (2007) to explain the surface activity of a puried chitosan sample. However, the surface tension reduction observed in
this study could be also attributed to the cooperative effect of
protein contamination. In fact, the interpretation of the molecular
origin of chitosan surface activity is still a controversial matter as
pointed out recently by McClements (2009). The dynamic surface
tension results obtained are typical of the adsorption of

70

65

60
-1

25C
20C
15C
10C
5C

(mNm )

I I
25C
20C
15C
10C
5C

55

50

45

40
100

1000

10000

t/s
Fig. 8. Dynamic surface tension for a solution with 0.5 wt% chitosan at 25
C.

N. Calero et al. / Food Hydrocolloids 24 (2010) 659e666

0,006

50

0,004

30

(rads)

40

E0(mNm-1)

macromolecules, showing a lag period characterized by a very low

decrease of surface tension due to the low surface density of the
adsorbed species. The surface tension reduction taking the value
obtained for the buffer solution used (69 mN/m) as a reference,
suggested the formation of a chitosan lm at the interface, which
could result in the occurrence of viscoelastic properties. These were
explored by step perturbations of the pendant drop. Fig. 9 shows
the area perturbation as explained above in the experimental
section and the system response. A nearly instantaneous jump was
observed from the initial surface tension, corresponding to an
adsorption time of 3 h, to a peak value. The aforementioned jump
denes the so-called initial surface tension change Dg0 (Hansen,
2008). This is related to the limiting elasticity, E0, which would
correspond to the response of an insoluble monolayer with a purely
elastic behaviour, according to the following equation:

665

20
0,002
10

0
0

2000

4000

6000

0,000
8000 10000 12000 14000 16000 18000 20000

t(s)

A Dg
E0 0 0
DA

(8)

where E0 limiting elasticity; A0 initial drop area before perturbation; Dg0 initial surface tension change; DA surface area step
The fall of surface tension after the peak value is due to relaxation processes at the interface. Namely, the relaxation processes
can be attributed to diffusion of new surfactant molecules towards
the freshly formed interface and to the rearrangement of previously
adsorbed chitosan macromolecules.
The experimental data tted the following equation as derived
by Miller, Loglio, Tesei, and Schano (1991) for a diffusion-controlled
model:

Dg1 Dg0 exp2u0 terfc

p
2u0 t

(9)

where u0 is the reciprocal of the characteristic relaxation time. For

the reverse step perturbation the following equation was used
(Hansen, 2008; Miller, Loglio, Tesei, & Schano, 1991):

Dg2 Dg1 t  Dg1 t  t1

(10)

where t1 represents the time when the reverse step was made. This
procedure was applied at discrete times along the adsorption
process (Fig. 8) in order to obtain a sort of picture of the evolution of
the surface viscoelasticity. E0 and u0 were determined from equations 8 and 9, respectively. Their values are shown in Fig. 10 as
a function of the adsorption time. The lm became more and more

24

75

-1

65
22

60

A(mm )

(mNm )

70

55
50

20

45
40
18

35
30
0

10

20

30

40

50

t(s)
Fig. 9. Step relaxation response for a solution with 0.5 wt% chitosan after an adsorption time of 3 h (squares). Droplet area as function of the adsorption time (plus sign).
The theoretical curves (line) were calculated from equations (9) and (10) with
E0 40 mN m1 and u0 0.004 rad/s. T 25
C.

Fig. 10. Limiting elasticity, E0 (triangles) and reciprocal of the characteristic relaxation
time, u0 (squares), as function of the adsorption time. T 25
C.

elastic as chitosan progressively adsorbed on the surface; reaching,

after 3 h, the limiting values of 0.0045 (rad/s) and 41 mN/m for
parameters u0 and E0, respectively. These values are consistent with
those published by Babak, Desbrires, and Tikhonov (2005) for
a hydrophobically modied chitosan sample, although in order to
reach a surface elasticity of the same order of magnitude, the latter
required a lower concentration than the sample used in this study.
This can be clearly attributed to the enhanced emulsifying properties of the hydrophobically modied sample.
4. Concluding remarks
The rheological behaviour of chitosan solutions was studied as
a function of polymer concentration and as a function of temperature, the most concentrated solution studied being 2.5 wt% chitosan. In comparison with previous studies by other authors;
chitosan solutions exhibited a similar behaviour in the dilute
regime (C < C*). However, remarkable differences concerning the
exponent of the characteristic slope of the semi-dilute regime
(C > C*) were found, which may be attributed to the fact that the
different buffers, degree of deacetylation, and molecular weight of
chitosan samples result in different polymer-polymer interactions
in the entanglement network.
The chitosan solutions studied exhibited Newtonian behaviour
at a concentration of 0.25 wt% and below. Conversely, above this
value all the systems showed shear-thinning behaviour which
tted the Cross Model.
The timeeconcentration (TCS) and timeetemperature (TTS)
superposition methods could be applied to obtain an overall master
ow curve. The values for the timeeconcentration and timeetemperature shift factors followed an exponential growth function and an Arrhenius-type equation, respectively.
The linear viscoelastic properties obtained from SAOS measurements for the chitosan solutions at higher concentrations than
0.25 wt% demonstrated random-coil-type viscoelastic behaviour. In
the same way as the analysis carried out for the ow curves, a master
curve was obtained for the frequency dependence of the complex
viscosity, which included the effects of concentration and temperature. The values for the timeeconcentration shift factors, ac, were
coincident with those used in the master ow curve. However, the
values for the timeetemperature superposition shift factors, aT,
were different from the previous values given for the master ow
curve. Therefore, the aT shift factors used for SAOS obeyed the
Arrhenius-type equation with a different activation energy value.
This discrepancy showed that the CoxeMerz rule was not entirely

666

N. Calero et al. / Food Hydrocolloids 24 (2010) 659e666

valid when temperature decreased, indicating the occurrence of

a slight change in the polymerepolymer interactions. Interestingly,
despite the fact that the CoxeMerz rule was not fully obeyed below
20
C, it has been shown that TTS and TCS may be used to produce an
overall master curve from the ow behaviour and SAOS measurements. This master curve included the effects of both chitosan
concentration and temperature by assuming different activation
energies for ow and complex viscosity behaviour.
The studies of the surface properties of chitosan revealed that
this biopolymer was able to reduce the surface tension and to form
a viscoelastic lm at the airewater interface. This behaviour is of
great interest in order to prepare stable emulsions since chitosan
solutions may contribute to the emulsifying capacity when
formulated with proteins.
Finally, we may conclude that chitosan solution fulls important
requirements to be used in emulsion processing; not only as an
emulsion stabiliser agent, due to its role as a thickener, but also
contributing its own potential emulsifying activity. This potential of
chitosan solutions should be explored in the near future, whether
used by itself or in combination with other emulsifying agents such
as proteins.
Acknowledgments
The results presented are part of a research project (reference:
P08-AGR-03974) supported by Consejera de Innovacin, Ciencia y
Tecnologa (Junta de Andaluca, Spain). The authors kindly
acknowledge their nancial support.
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