Carbohydrate Polymers 134 (2015) 285–292

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Carbohydrate Polymers
journal homepage: www.elsevier.com/locate/carbpol

Effects of pH and temperature on colloidal properties and molecular

characteristics of Konjac glucomannan
Wenjie Jian a,b , Ka-Chai Siu a , Jian-Yong Wu a,∗
a
Department of Applied Biology & Chemical Technology, The Hong Kong Polytechnic University, Hung Hom, Kowloon, Hong Kong
b
Department of Pharmacy, Xiamen Medical College, Xiamen 361000, China

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

Article history: The hydrocolloidal characteristics of Konjac glucomannan (KGM) are important for its application as a
Received 18 May 2015 thickening and gelling agent for liquid foods. In this study, the rheological behavior and molecular proper-
Received in revised form 8 July 2015 ties such as molar mass, hydrodynamic radius and chain conformation of KGM in water were determined
Accepted 11 July 2015
at various pH levels (4.0–10.0) during heating from 20 to 80 ◦ C. Acidic and neutral conditions (pH 4.0–7.0)
Available online 31 July 2015
promoted the dispersion of KGM, and alkaline condition at pH 10 favored its aggregation in water, while
KGM maintained a random coil conformation in the whole pH range. Associated with the pH effects were
Keywords:
changes in the rheological behavior during heating from 20 to 80 ◦ C. The significant differences in the
Konjac glucomannan
Colloidal solution
colloidal and rheological characteristics were mainly attributed to alteration of intermolecular interac-
pH tion (attractive or repulsive) rather than deacetylation at various pH levels. Deacetylation occurred in
Temperature both acidic and alkaline condition. The second virial value was positive in acidic and negative in alka-
Light scattering analysis line condition. The results showed that hydrocolloidal characteristics of KGM in water were significantly
Molecular interaction affected by pH.
© 2015 Elsevier Ltd. All rights reserved.

1. Introduction & Nishinari, 2002; Nishinari & Takahashi, 2003; Solo-de-Zaldivar,

Konjac glucomannan (KGM), a plant polysaccharide derived
from the tuber of Amorphophallus konjac C. Koch, has been widely
applied in food industry as a gelling and thickening agent due
to its ability of thermal-irreversible gelling and effective viscos-
ity enhancement (Herranz et al., 2012; Tomczynska-Mleko et al.,
2014). Many recent studies have shown its various biological
activities including anti-tumor, immunomodulation, wound heal-
ing (Al-Ghazzewi, Elamir, Tester, & Elzagoze, 2015; Bo, Muschin,
Kanamoto, Nakashima, & Yoshida, 2013; Zhang, Chen, & Yang,
2014). Although the exact molecular composition may vary with
the source and production process, purified KGM has a polysaccha-
ride backbone composed of glucose and mannose at 1:1.5–1:1.6
molar ratio, with 5–10% acetyl substitution (Dave & McCarthy,
1997). It is generally believed that acetyl group on the KGM
polymer structure is the most important for its solubility and
gelling properties in water (Huang, Takahashi, Kobayashi, Kawase,
∗ Corresponding author. Fax: +852 2364 9932.
E-mail address: jian-yong.wu@polyu.edu.hk (J.-Y. Wu).
Tovar, Borderias, & Herranz, 2014). Through the formation of a
three-dimensional network of molecular chain by junction zones
after deacetylation, the aqueous solution of KGM is transformed
into a thermally stable gel (Williams et al., 2000). Exposure to alka-
line and heat treatment may cause the deacetylation of KGM and
significant reduction of solubility in an aqueous solution.
The afore-stated gelation mechanism suggests that thermal and
alkaline conditions are important factors affecting the gelling prop-
erties of KGM. Although many previous studies have evaluated the
rheological properties of KGM in aqueous solutions, most of these
have been carried out at neutral pH and room temperature (Jin
et al., 2014; Xu, Sun, Yang, Ding, & Pang, 2007). The characteristics
of KGM in acidic and alkaline pH conditions and at higher tem-
peratures are important for their applications in these conditions.
The evaluation and quantification of the related characteristics and
parameters is helpful to understanding the dynamic behavior and
interactions of KGM polymer chains, and their relationship to the
textural properties of liquid foods (Nishinari and Takahashi, 2003).
This study aims to examine the hydrocolloid characteristics of
KGM in water at various pH levels in response to heating, and
to elucidate the related gelling mechanisms. The parameters of

http://dx.doi.org/10.1016/j.carbpol.2015.07.050
0144-8617/© 2015 Elsevier Ltd. All rights reserved.
286 W. Jian et al. / Carbohydrate Polymers 134 (2015) 285–292
molecular properties, and chain conformation, and aggregation will
be derived through light scattering analysis as useful references for
the application of KGM in food processing.
2. Materials and methods
2.1. Purification and composition analysis of KGM
The initial Konjac glucomanan (KGM) material used for this
study was purchased from Shaotong Shanai Kojac Development
Co. (Yunnan, China). Before use, the KGM powder was purified by
alcohol precipitation followed as described in our previous study
(Jian, Yao, Wang, Guan, & Pang, 2010). Its weight-average molecu-
lar weight was about 1.14×103 kDa. This purified KGM was used
in this study and its composition was analyzed as follows. The
moisture content was determined after drying in an oven (105 ◦ C)
to constant weight, and the ash content determined in a muffle
furnace (550 ◦ C) according to methods from International Organi-
zation for Standardization (ISO 2291:1980, ISO 936:1998). The total
carbohydrate content was determined by phenol sulphuric acid
method using glucose as a standard (Chaplin & Kennedy, 1994) and
the total protein content by the Kjeldah method (Crisan & Sands,
1978). Monosaccharide constituents were analyzed by HPLC after
acid hydrolysis and 1-phenyl-3-methyl-5-pyrazolone (PMP) reac-
tion as reported previously (Chen, Siu, Cheung, & Wu, 2014). Acetyl
content was determined by titration using hydrochloride solution
according to the method of Laignel, Bliard, Massiot, & Nuzillard,
(1997).
2.2. Preparation of KGM solution
All experiments in this part were carried out at room temper-
ature as well as the storage of prepared solution. KGM powder
(0.1 g) was dissolved in super pure water (200 ml) by constantly
stirring for 24 h at room temperature. The KGM solution pH was
adjusted to 4.0, 7.0, 9.0, and 10.0 with 0.1 M and 1 M NaOH or HCl.
The volume of NaOH or HCl added for to the KGM solution for the
pH adjustment was very small, having a negligible effect on the
solution concentration. The solution was kept at room tempera-
ture for 12 h to allow the KGM to swell, followed by centrifuging
at 18,000 rpm (∼1814 × g) for 15 min to remove insolubles (aggre-
gates and gels formed at high concentrations, if any). The solution
(0.5 mg/ml) was directly used for measurement of dynamic light
scattering. For IR analysis, the solution was dialyzed with ultrafil-
tration membrane made of regenerated cellulose (1.0 kDa MWCO,
Millipore, USA), and then freeze-dried. As the KGM sample was
isolated by ethanol precipitation of the commercial KGM, dialy-
sis is a common step for the removal of low molecular weight
impurities such as small salts which may be attached to the KGM
polymer chains. Because of the high viscosity of KGM solution,
a low concentration solution (0.2 mg/ml) was prepared for the
GPC-MALLS measurement to assure complete passage of the dis-
solved KGM through the GPC-MALLS system. On the other hand,
a much higher concentration of KGM solution (5 mg/ml) was pre-
pared in the same way but without centrifuging for the rheological
measurement.
2.3. Measurement of dynamic rheological behavior
The rheological behavior of KGM was measured in various con-
ditions according to a reference (Herranz, Borderias, Solas & Tovar,
2012) with minor modifications. Dynamic rheology measurement
was performed of the KGM solution on an Anton Paar MCR302
Rheometer (Bohlin Instruments, Inc. Cranbury, NJ) with parallel-
plate pp50 (20 mm diameter and 1 mm gap). Preliminary tests
were performed at 25 ◦ C with angular frequency of 6.28 rad/s and
strains in the range of 0.1–100% to find the linear viscoelastic region
for pH 4.0, 7.0, 9.0 and 10.0 corresponding to the strains of 30%,
35%, 20% and 1.5%, respectively. Accordingly, the shear storage
modulus (G ) and loss modulus (G ) were measured at angular fre-
quency of 6.28 rad/s and 1% strain as a function of temperature
varying from 20 ◦ C to 80 ◦ C at 2 ◦ C/min and data collection every
0.25 min. All tests were performed in triplicate, and the results were
averaged.
2.4. Infrared and Raman spectral measurements
Infrared (IR) measurement was performed on a Fourier trans-
forms IR spectrometer (Nicolet 6700, Thermo Fisher, USA) at a
resolution of 4 cm−1 in the range of 400–4000 cm−1 (Xu et al.,
2007). Freeze-dried KGM samples were prepared as KBr slices and
scanned against air background for 32 times with each sample to
attain an average. Raman spectra of KGM samples (freezed-dried
powder) were recorded from 100 cm−1 to 3000 cm−1 on a HR800
Raman microscope (Horiba Jobin Yvon), equipped with a 488 nm
laser operated at 180 mW and spot size of excitation laser about
1 ␮m (Synytsya, Copikova, Matejka, & Machovic, 2003).
2.5. Dynamics light scattering measurement
Dynamics light scattering (DLS) measurement was performed
to attain the hydrodynamic radius (Rh) and to estimate molar
mass (Mw-R) with a DynaPro NanoStar instrument (Wyatt, USA)
(Sagou, Rotureau, Thomas, & Duval, 2013), Before loading, the KGM
sample solution in water was filtered through a Millex-HN nylon
clarification kit with a pore size of 0.2 ␮m (Fisher Scientific). The
temperature was varied from 25 ◦ C to 80 ◦ C at 0.5 ◦ C/min ramp
rate. The signal acquisition period was set to 10 s and the aver-
aged result of 10 acquisitions was taken as a measurement. The
refractive index increment for KGM was set to 0.138. The molar
mass Mw-R was estimated from Rh by the model of linear polymer
and random coil conformation (Prawitwong, Takigami, & Phillips,
2007).
2.6. GPC-MALLS analysis
Gel permeation chromatography with on-line multi-angle laser
light scattering (GPC-MALLS) was applied to determine the weight-
average molecular weight (Mw ) and conformation characteristics
of KGM dissolved in water (Prawitwong et al., 2007). The GPC-
MALLS system consisted of a Waters e2695HPLC system and a
Waters 2414 refractive index detector, and a Wyatt DAWN HELEOS
II detector (18 angles). Sample solution (0.2 mg/mL) was injected
into the system at 100 ␮L. Two GPC columns were used in a series
for the separation, TSK-GEL G4000PWxl and G5000PWxl, (Tosoh
Bioscience, Tokyo, Japan). Sodium nitrate solution (50 mM with
0.02% w/v sodium azide) was used as the mobile phase at a flow
rate of 0.4 ml/min and 25 ◦ C. Normalization was performed with
dextran standards (Sigma Aldrich, Mw 30 kDa). Both GPC data and
MALLS data were collected and analyzed using Astra6.0 software
package.
2.7. Determination of second virial coefficient by MALLS analysis
The second virial coefficient was determined through MALLS
analysis with a Wyatt Dawn Heleos II detector linked with
a Wyatt Calypso automating delivery system on batch mode
(Ma et al., 2015). Before the measurement, the initial KGM
solution at 0.5 mg/ml was diluted into a series of gradient con-
centrations (0.1, 0.2, 0.3 and 0.4 mg/ml) with the respective pH
buffer solution. Meanwhile, 1 mg/ml solution of dextran standard
 W. Jian et al. / Carbohydrate Polymers 134 (2015) 285–292
Fig. 1. Change of storage modulus and loss modulus of KGM solution (5 mg/ml) with temperature at various pH levels.
(30 kDa) was prepared with each respective pH buffer solu-
tion, and used as the normalization standard for the following
measurement.
The instrument system was first flushed with a pH buffer solu-
tion to obtain the initial baseline light scattering intensity, followed
by injection of 1 mg/ml dextran solution to normalize the system.
KGM sample solution of gradient concentrations (0.1–0.5 mg) was
then injected, followed by a buffer flush to retain the baseline. The
light scattering intensity reading was taken 5 min after each injec-
tion. Zimm plot was generated with the light scattering data based
on the Rayleigh theory, from which the second virial coefficient
was determined by the slope of the concentration versus the excess
Rayleigh ratio plot.
2.8. Zeta-potential measurement
Zeta-potential of the KGM solutions (0.5 mg/ml) was measured
by dynamic laser light scattering (DLS) analysis on a Malvern
Zetasizer Nano-ZS90 instrument using a folded capillary cuvette
DTS1060 at room temperature, with five repeating measurements
taken for each sample (Hunter, 2013).
2.9. Statistical analysis
All experiments were carried out in triplicate and the results
were represented by means and standard deviations. Analysis
of variance (ANOVA) was computed using SPSS 16.0 (SPSS Inc.,
Chicago, IL).
3. Results and discussion
3.1. Chemical composition of KGM
According to analysis, KGM was composed of 93.4% total car-
bohydrate, 1.87% total protein, 1.62% acetyl, 8.49% moisture, and
287
1.53% ash (all by mass) and of mannose and glucose in 2.3:1 molar
ratio. As KGM is extracted from the tuber of Amorphophallus konjac
C. Koch, the purity and composition depend on the source plant
material as well as the extraction process (Nishinari, 2000), and its
monosaccharide composition varies with the plant species (Pang,
Lin, Zhang, Tian, & Sun, 2003). Most of the above composition values
of KGM were within the ranges commonly reported in the litera-
ture (Kato & Matsuda, 1969; Koroskenyi & McCarthy, 2001; Smith
& Srivastava, 1959), except for the relatively higher molar ratio of
mannose to glucose, which is commonly in the range of 1.5:1–1.6:1
(Koroskenyi & McCarthy, 2001; Maeda, Shimahara, & Sugiyama,
1980). Therefore, the KGM sample used in this study was repre-
sentative of those widely used in the food industry and its property
data can be useful references for general situation.
3.2. pH and temperature effects on rheological behavior of KGM
in water
Fig. 1 shows the changes viscoelastic modulus of KGM solu-
tion with temperature increase (20–80 ◦ C) at various pH levels.
The rheological measurements showed a high reproducibility, with
errors less than 3%. KGM presented significantly different rheolog-
ical behaviors with the pH changes as shown by Fig. 1. At pH 4
(Fig. 1a) and pH 7.0 (Fig. 1b), the storage modulus (G ) was close to
the loss modulus (G ) at 20 ◦ C, and as the temperature increased,
G became higher than G . This tend suggests that KGM at 20 ◦ C was
in the colloidal solution-to-gel transition state and remained in a
colloidal solution as the temperature exceeded 20 ◦ C in acidic and
neural conditions. Another trend was the linear decline of G and G
with temperature increase to a very low level at 80 ◦ C, implying the
weakening of inter- and intra-molecular attraction forces rapidly
by heating, which may involve the degradation of polymer chain
and dissociation of polymer aggregates. At pH 9.0 (Fig. 1c), the ini-
tial moduli at 20 ◦ C were significantly higher, and dropped sharply
288 W. Jian et al. / Carbohydrate Polymers 134 (2015) 285–292

100 100
(a) (b)

Transmittance (%)
Transmittance (%)
80 80

60 60
Acetyl
2930
40 1650 40

3430
20 20
3600 2800 2000 1200 400 3600 2800 2000 1200 400
-1
Wavenumber (cm-1) Wavenumber (cm )
100 100
(c) (d)
Transmittance (%)

Transmittance (%)
80 80

60 60

40 40

20 20
3600 2800 2000 1200 400 3600 2800 2000 1200 400
-1 -1
Wavenumber (cm ) Wavenumber (cm )

Fig.2. FT-IR spectra: (a) Native KGM; freeze-dried KGM dissolved in water at (b) pH 4.0, (c) pH 9.0, and (d) pH 10.0.

as the temperature was increased to 21 ◦ C, and then followed the

similar trends to those at the lower pH.
Most interesting and dramatic changes were observed at pH 10.0
(Fig. 1d). The initial G and G at 20 ◦ C were much lower than those
at the lower pH, the difference between G and G was relatively
large. G and G followed very different trends with the tempera-
ture increase from 20 to 80 ◦ C. G declined gradually in the whole
temperature range except at 60 ◦ C, it jumped to a notably higher
level. In contrast, G showed a small decline with the temperature
increase from 20 to about 40 ◦ C and remained steady till 60 ◦ C; at
about 60 ◦ C, it jumped to a much higher level, and then increased
sharply with further increase of temperature to 80 ◦ C. These trends
were indicative of a colloidal solution undergoing gelation, i.e. at
pH 10.0, the KGM solution turned to a gel as the temperature was
increased to 60 ◦ C and the gel strength increased rapidly with fur-
ther temperature increase. The gelation process at pH 10.0 due to
heating was consistent with the alkaline-induced gelation of KGM
reported previously (Herranz, Borderias, Solas et al., 2012).
Therefore, the influence of temperature on the rheological
behavior of KGM solution was strongly dependent on the pH range.
In the acidic and weakly alkaline range (pH 4.0–9.0), lower tem-
peratures (below 21 ◦ C) favored the gelling of KGM, and higher
temperatures (above 21 ◦ C) promoted its dispersion in water.
In highly alkaline pH range, lower temperatures favored solu-
tion stability, and higher temperatures (above 60 ◦ C) promoted
gelling. Besides the rheological measurement, further analysis and
quantification of the structural, conformation and aggregation
properties of KGM in solution will be helpful for characterization
and elucidation of the changes in different pH ranges with temper-
ature.

3.3. Infrared spectral characteristics

Fig. 2 shows the FT-IR spectra of KGM samples from var-

ious pH solutions. On the spectrum at neutral pH 7.0 (Fig. 2a), Fig. 3. Raman spectra: (a) Native KGM; (b) freeze-dried KGM dissolved in water at
the strong vibration at 3430 cm−1 is ascribed to the hydroxyl ( OH) various pH levels.
 W. Jian et al. / Carbohydrate Polymers 134 (2015) 285–292 289

Fig. 4. Temperature sweep of hydrodynamics radius (a) and weight-average molecular mass (b) of KGM solution at various pH levels.

group of polysaccharide, peak at 2930 cm−1 to the vibration
of methyl ( CH3 ), and the small peak at 1730 cm−1 to the acetyl
group (CH3 CO ) as the characteristic substituted group in KGM
(Zhang et al., 2001). The KGM in acidic solution (pH 4, Fig. 2b) and
alkaline solutions (pH 9, Fig. 2c; pH 10 Fig. 2d) showed the similar
IR spectra except for the absence of acetyl (1730 cm−1 ) and methyl
(2930 cm−1 ) peaks. This implied that acidic and alkaline treatment
caused the deacetylation but no change in the backbone of KGM.
Fig. 3a shows the Raman spectrum of native KGM. As the acetyl
group (CH3 CO ) of KGM is attached with hydroxyl at C-6 posi-
290 W. Jian et al. / Carbohydrate Polymers 134 (2015) 285–292

tion to form acetic acid ester linage, the weak peak at 1740 cm−1
is attribute to the vibration of carbonyl(C O) and the peak at
1473 cm−1 to the vibration of methyl (CH3 ) in the acetyl group (De
Gelder, De Gussem, Vandenabeele, & Moens, 2007; Synytsya et al.,
2003). In comparison of the Raman spectra before (Fig. 3a) and after
treatment (Fig. 3b), there was a significant decrease in Raman inten-
sity after acidic and alkaline treatment and also the elimination of
the characteristic for acetyl peaks at 1740 cm−1 and 1473 cm−1 .
The findings from the Raman spectra confirmed the results of FT-IR
analysis.
Similar to the changes observed from the above IR and Raman
spectra of KGM, deacetylation has also occurred to KGM as a
result of combined alkaline and heat treatment in previous studies
(Herranz et al., 2012; Solo-de-Zaldivar et al., 2014; Williams et al.,
2000). Deacetylation in KGM could significantly affect its solubility
and gelling properties (Du, Li, Chen, & Li, 2012). However, previ-
ous studies have mostly focused on alkaline deacetylation of KGM
and ignored acidic conditions. Our present study has shown that
deacetylation also occurred in acidic conditions, similar to the acid
hydrolysis of xylan-associated acetyl groups in dilute sulfuric acid
solution (Maloney, Chapman, & Baker, 1985).
As the acetyl group of KGM can be hydrolyzed in both acidic
and alkaline conditions, deacetylation is not an essential condition
for gelation of KGM. Meanwhile, although the degree of acetyla-
tion has been shown to affect the solubility of KGM in neutral
water (pH 7.0) (Du et al., 2012; Huang et al., 2002), it may be much
less important than temperature and pH in affecting the colloidal
and rheological properties KGM. In other words, the changes of
rheological behaviors with pH and temperature can be associated
with alternation of other molecular properties than deacetylation.
The findings are in agreement with the view of Williams et al.
(2000) that gelation of native KGM solution may also occur without
deacetylation.

3.4. Changes of hydrodynamic radius and molar mass

As shown in Fig. 4a, hydrodynamic radius (Rh) presented differ-

ent trends with temperature increase at different pH levels. At pH
4.0 and 7.0, Rh decreased sharply from about 120 nm to 40 nm with
temperature increasing from 25 ◦ C to 50 ◦ C; at pH 10.0 Rh increased
sharply at 60 ◦ C; at pH 9.0, Rh was not affected by temperature
change. The molar mass (Mw-R) of KGM showed the same trends
with temperature as Rh at various pH ranges (Fig. 4b). Both Rh and
Mw-R were significantly affected by the pH change (P < 0.01). The
results suggested that the changes in Rh with temperature were
mainly attributed to the aggregation or disaggregation of the poly-
mer chains. These results have shown the significance of pH level
on the thermal stability and aggregation of KGM. At pH 4.0 and 7.0,
increasing temperature favored its dispersion and colloidal stability
in solution, while at pH 10, temperature had an opposite effect. Such
trends were consistent with the above rheological changes with pH
and temperature, indicating that pH and temperature affected KGM
rheological behavior through variation of intermolecular aggrega-
tion or disaggregation.
3.5. Molecular characteristics from GPC-MALLS analysis
Besides the Rh and Mw-R obtained from DLS, the weight-
average molar mass (Mw), root-mean-square radius (Rm) and chain
conformation are also important molecular characteristics which
are derived from GPC-MALLS analysis (Fig. 5). The profile at pH
10.0 (Fig. 5a) was significantly different from those at other pH lev-
els, indicating the Mw changes at pH 10. The Mw decreased from
1463 to 1318 kDa with the pH decrease from pH 7.0 to pH 4.0, and
to 1385 kDa with pH increase from pH 7.0 to 9.0, but increased to
Fig. 5. Chromatograms of KGM at various pH levels obtained from GPC-MALLS anal-
ysis: (a) GPC-MALLS spectra; (b) RMS radius; (c) molar mass (0.2 mg/ml KGM in
water).
2035 kDa with the pH increase to 10.0. The number-averaged molar
mass (Mn) as well as the dispersity index (Mw/Mn) followed the
similar trends of change with temperature (Table 1). The Mw and
Rm values at pH 7.0 obtained in this study were close to the those
(1170 kDa and 120.3 nm reported by Prawitwong et al. (2007)), but
significantly different from those (Mw 247.6 kDa and Rm 100 nm)
by Xu et al. (2013) due probably to the different raw material prop-
erties and preparation methods.
The changes in Mw and other related molecular properties
of KGM could be attributed to different mechanisms. At pH
4, the decrease in Mw was probably caused by acidic degra-
dation of glycoside linkage; at pH 9.0, the slight decrease in
Mw was probably a result of deacetylation. The significant Mw
increase at pH 10.0 was attributed to the aggregation of KGM
in alkaline conditions. The changes of the root mean square
radius (Rm) with pH also followed the similar trends with Mw
(Table 1).
In spite of the significant changes in Mw and particle (aggregate)
size with pH, the conformation of KGM remained in the random coil
as seen from the relatively small change in the slope of conforma-
tion plot (Rm vs Molar mass) from 0.41 to 0.49 (Table 1). Therefore,
it is concluded that pH range had a significant effect on the molec-
 W. Jian et al. / Carbohydrate Polymers 134 (2015) 285–292 291

Table 1
Molecular characteristics of KGM solution (0.2 mg/ml) at pH 4.0, pH 7.0, pH 9.0 and 10.0 obtained from GPC-MALLS.

pH Mn (kDa) Mw (kDa) Rm (nm) RMS plot slope

4.0 1024.9( ± 3.10%) 1318(±1.60%) 123.6(±0.70%) 0.49(±0.09%)

7.0 1103.2( ± 2.80%) 1463.3(±2.60%) 127.2(±0.80%) 0.44(±0.12%)
9.0 1155.6( ± 1.60%) 1385.2(±1.50%) 123.3(±0.60%) 0.44(±0.11%)
10.0 1345.8( ± 3.90%) 2035.1(±4.90%) 129.9(±2.70%) 0.41(±0.27%)

tend to form aggregates or gels at pH 10.0. This pH-dependent

dispersion or aggragation of KGM in water is in close agreement
with that dervied from the results of rehological and molecular
weight measurements.
The colloidal and gelling properties of biopolymers in solution
are strongly dependent on short-range inter-molecular inter-
actions such as hydrgen bonding and hydrophobic interaction
(Nishinari & Zhang, 2004; Yin, Zhang, Huang, & Nishinari, 2008). The
second virial coefficient value (A2 ) is a useful index for the degree of
intermolecular interactions of biopolymers in dilute solutions due
to several molecular forces including ionic, hard-sphere, van der
Waals and other short-range interactions (e.g., hydrophobic inter-
actions) (Lehermayr, Mahler, Mader, & Fischer, 2011). Negative A2
values are indicative of attactive intermolecular interactions for
aggregate formation while positive A2 values represent repulsive
intermolecular interactions for colloidal dispersion and stability.
Zeta potential as a measure of electrostatic charges is another
useful parameter for molecular interactions through electrostatic
force (Saito et al., 2013). As shown in Fig. 6c, the KGM solu-
tion exhibited negative zeta potentials (negatively charged) at pH
3.0–10.0, and the negative zeta potential increased from −4.12
to −6.12 mv with the pH increase from pH 7.0 to 10.0. However,
according to the DLVO theory, the absolute magnitude of zeta
potential between 10 and 30 mv indicates an unstable dispersion,
and the value below 5 mv a strong aggregation (Bhattacharjee,
Elimelech, & Borkovec, 1998). In view of the low absolute zeta
potential of KGM, the electrostatic potential was not sufficient to
maintain its stable disepersion. It furter suggests that electrostatic
interactions in KGM molecules were in much less significant than
other molecuar intreactions in affecting its colloidal stability and
gelation behavior.

4. Conclusions

The results have shown that the colloidal and rheological

Fig. 6. Zimm plot of KGM at pH 7.0 and 25 ◦ C (a) and the corresponding second
virial coefficient (b) and zeta potential (c) of KGM solution (0.5 mg/ml) at various
pH levels.
ular weight and particle size but not on the chain conformation of
KGM in water, and acidic condition (pH 4.0) promoted the degra-
dation of KGM molecular chain, and alkaline condition favored its
aggregation.
3.6. Molecular interactions
behaviors of KGM in water were significantly affected by pH and
temperature. At room or relatively low temperatures, the pH level
strongly affected the average molecular weight and particle size
of KGM but not the chain conformation. Acidic conditions pro-
moted the degradation and disaggregation of molecular chain
whereas alkaline conditions favored its aggregation. KGM remained
in random coil conformation in all pH conditions. The pH range sig-
nificantly affected the rheological behaviors of KGM upon thermal
treatment. In relatively acidic conditions, thermal treatment pro-
moted the dispersion of KGM or the formation of a stable solution in
water. In alkaline conditions, KGM presented the typical gelation
process at temperature over 60 ◦ C. These effects of pH and tem-
perature on the colloidal and rheological behaviors were mostly
attributed to the changes in intermolecular interactions instead of
deacetylation.

Fig. 6a is the Zimm plot of KGM solution at pH 7.0 and 25 ◦ C

obtained by static laser scattering measurement (Zimm plots at
other pH levels not shown). The second virial coefficient values
(A2 ) derived from the Zimm plots are positive at pH 3.0–9.0 and
negative at pH 10.0 (Fig. 6b). The results indicate that KGM retains
a stable dispersion in acidic and slightly alkaline conditions and
Acknowledgements
This work was supported by Hong Kong Scholar Program
(XJ2013031) and The Hong Kong Polytechnic University, Hong
Kong Government GRF grant (PolyU 5033/11P), China Postdoctoral
292 W. Jian et al. / Carbohydrate Polymers 134 (2015) 285–292
Science Foundation (2012M510199), and the Natural Science Foun-
dation of Fujian Province (2012J05046).
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