Preparation and characterization of heterogeneous deacetylated

konjac glucomannan
Jing Li
a, b
, Ting Ye
a, b
, Xiaofang Wu
a, b
, Jian Chen
a, b
, Shishuai Wang
a, b
, Liufeng Lin
a, b
,
Bin Li
a, b,
*
a
College of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
b
Key Laboratory of Environment Correlative Dietology, Huazhong Agricultural University, Ministry of Education, China
a r t i c l e i n f o
Article history:
Received 10 October 2013
Accepted 1 February 2014
Keywords:
Konjac glucomannan
Heterogeneous deacetylation
Characterization
Solubility uevaluation
a b s t r a c t
Deacetylated konjac glucomannan (Da-KGM) powder was prepared in heterogeneous system. Kinetics of
heterogeneous deacetylation was investigated to predict the optimum reaction conditions. The results
indicated that: heterogeneous deacetylation was inuenced by the kind and amount of alkali, ethanol
concentration, temperature and reaction time. Meanwhile, it followed rst-order kinetics and the
apparent activation energy was 15.59 kJ/mol. The properties of Da-KGM were studied by Fourier-
transform infrared spectroscopy (FT-IR), differential scanning calorimetry (DSC) and X-ray diffraction
(XRD), which proved that there was no signicant difference of primary structure, thermal properties
and crystal properties among Da-KGMs. The solubility was also analyzed and evaluated. As deacetylation
degree (DD) increased, the solubility of KGM showed exponential decrease. In addition, lower temper-
ature was more effective to promote the dissolution of Da-KGM.
2014 Elsevier Ltd. All rights reserved.
1. Introduction
KGM is extracted from the tubers of Amorphophallus konjac C.
Koch (Nishinari, Williams, & Phillips, 1992) and regarded as a kind
of dietary ber, which has signicant health functions. It has been
widely used in food industry because of its good swelling, gelling
and other features. This natural material can be used for the
preparation of composite materials, edible lm, coating/packaging
lm, biodegradable lm, and controlled release matrix. KGM is a
water-soluble polysaccharide, composed of b-(1 / 4) linked D-
mannose and D-glucose in a molar ratio of 1.6:1 (Kato, 1969) or 1.4:1
(Dey & Dixon, 1985), with about 1 in 19 units being acetylated
(Corp., 1993; Katsuraya et al., 2003). It is widely accepted that the
presence of acetyl-substituted residues confers water-solubility to
the KGM in aqueous solution (Koroskenyi & McCarthy, 2001). In
addition, Da-KGM may be used as adsorbent in many elds. Car-
boxylic acid functionalized Da-KGM was synthesized with copoly-
merization of methyl acrylate and methyl methacrylate by free
radical graft, which showed a fast adsorption rate and high removal
efciency of Pb
2
and Cu
2
from aqueous solution (Liu, Luo, Lin,
Liang, & Chen, 2009). Furthermore, it was observed that Da-KGM
can be utilized as a low-cost and readily available biosorbent for
removal of tannin from aqueous solutions (Liu, Luo, & Lin, 2010).
Plenty of studies have been carried out to treat KGM with alkali
for deacetylation (Bin & Bi-jun, 2003; Cheng, Abd Karim, Norziah, &
Seow, 2002). The deacetylated KGM can form a heat-stable gel
(Perols, Piffaut, Scher, Ramet, & Poncelet, 1997). The gelation of
KGM was studied by rheological measurements with low-
amplitude oscillation experiments in the presence of different
salts (e.g. NaSCN, NaCl, Na
2
SO
4
), and compared with those of
methylcellulose, gelatin, acrylamide and pectin (Case, Kropp,
Hamann, & Schwartz, 1992). It was suggested that the gelation ki-
netics of KGM samples was governed by deacetylation rate (Gao &
Nishinari, 2004a). After that, a new study showed that a faster
gelation rate and a more elastic modulus were facilitated by
increasing the deacetylation degree (DD). The hydrophobic inter-
action and hydrogen bonding were both presented in KGM gela-
tion, and hydrophobic interactions strengthen while hydrogen
bonding weaken with increasing DD (Du, Li, Chen, & Li, 2012).
So far, the deacetylation of KGM has been performed in homo-
geneous systems (mostly in water) by alkali treatment. However,
KGMsolution of large concentration cannot be carried out with this
method due to its high viscosity as well as the complicated pro-
cedures. To sum up, these disadvantages have severely limited the
* Corresponding author. College of Food Science and Technology, Huazhong
Agricultural University, Wuhan 430070, China. Tel.: 86 27 6373 0040; fax: 86 27
8728 8636.
E-mail address: libinfood@mail.hzau.edu.cn (B. Li).
Contents lists available at ScienceDirect
Food Hydrocolloids
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http://dx.doi.org/10.1016/j.foodhyd.2014.02.001
0268-005X/ 2014 Elsevier Ltd. All rights reserved.
Food Hydrocolloids 40 (2014) 9e15
applications of homogeneous deacetylation. Recent attention to
solid-phase reaction by the mechanochemical (MC) treatment has
been increased due to the growing economical and eco-logical re-
quirements (Pan, He, & Wang, 2008). However, considering the rate
of solid-phase reaction was very slow theoretically, the increasing
DD might be caused by ethanol washing during the later step
instead of the removal of acetyl group. As a result, the common
disadvantages of these two methods made it hard to obtain Da-
KGM with accurate deacetylation degree (DD). There were many
reports on heterogeneous deacetylation of chitin since the early
1930s (Bin & Bi-jun, 2003; Cheng et al., 2002; Nishinari et al., 1992).
The studies of Aiba et al. suggested that chitosan obtained by het-
erogeneous deacetylation had a block-type distribution of acetyl
group along the polymer chain (Aiba, 1991; Kurita, Sannan, &
Iwakura, 1977). Some other researchers comparatively studied the
heterogeneous deacetylation of a- and b-chitins fromshrimp, squid
pens and shells in a multistep process, which allowed them to
obtain chitosan with high molecular weight and high DD (Jiang &
Xu, 2006; Kurita, Ishii, Tomita, Nishimura, & Shimoda, 1994;
Lamarque, Viton & Domard, 2004). However, the heterogeneous
deacetylation of KGM has not been reported as yet. In this paper,
Da-KGM was obtained in a heterogeneous system on the basis of
heterogeneous deacetylation of chitin. This method was proposed
to obtain Da-KGM with precise and controllable DD, which would
provide the ideal material for the research of the gelation mecha-
nism and determining the role of acetyl group in gelation. More-
over, heterogeneous deacetylation was excellent for mass
production. In addition, kinetic prole was investigated to predict
the optimum reaction conditions. The physical properties of Da-
KGM were studied by FT-IR, DSC and XRD. The solubility was also
analyzed and evaluated.
2. Material and methods
2.1. Material
The raw KGM was purchased from Hubei Jianshi, Nongtai In-
dustrial Co., Ltd. The absolute value of degree of acetyl-substituted
residues in neat-KGM was 0.053. All chemicals were purchased
from Sinopharm Chemical Reagent Company. They were all of A.R.
grade and used without further purication.
2.2. Heterogeneous deacetylation
2.2.1. Effect of inuence factors on deacetylation reaction
Thirty grams (30.00 g) of puried KGM powder and 200 ml of a
certain concentration of ethanol solution was mixed in a 250 ml
conical ask. The mixed suspension was swelled in a constant
thermostat oscillator at a certain temperature for 30 min (150 rpm).
Subsequently, the suspension with alkali solution was reacted for a
certain period. After deacetylation, each sample was washed three
times with aqueous ethanol (50%, 75%, and 95%) to remove excess
alkali and nally washed with absolute ethanol. The excess of
ethanol was evaporated in a fume cupboard followed by vacuum
drying for 6 h at 40

C and the powdered Da-KGM was obtained.
2.2.2. Preparation of KGM with different deacetylation degree
Thirty grams (30.00 g) of puried KGM powder and 200 ml of
50% (v/v) ethanol was mixed in a 250-ml conical ask. The mixed
suspension was swelled in a constant thermostat oscillator at 40

C
for 30 min (150 rpm). Subsequently, the suspension with Na
2
CO
3
solution was reacted at 40

C for 24 h. After deacetylation, each
sample was washed three times with aqueous ethanol (50%, 75%,
and 95%) to remove excess alkali and nally washed with absolute
ethanol. The excess of ethanol was evaporated in a fume cupboard
followed by vacuum drying for 6 h at 40

C and the powdered Da-
KGM was obtained. KGM with different DD was obtained by
changing the volume of Na
2
CO
3
, which were coded as Da0, Da1,
Da2, Da3, Da4, Da5, Da6 and the corresponding molar ratio of
Na
2
CO
3
to acetyl group was 0.5:8, 1:8, 2:8, 3:8, 4:8 and 8:8,
respectively.
2.2.3. Determination of deacetylation degree
Deacetylation degree (DD) is dened as the ratio of content of
acetyl groups been removed and total acetyl groups in KGM. This
method was based on the Eberstadt method including saponica-
tion and successive titration (Chen, Zong, & Li, 2006; Tanghe,
Genung, & Mench, 1963). Five grams (5 g) of the KGM powder
with 50 ml of 75% ethanol was mixed in a 250-ml conical ask. The
mixed suspension was then heated to 50

C in constant water bath
for 30 min before cooling down to room temperature. 5 ml of KOH
(0.5 mol/L) was added into the suspension for saponication in a
digital water bath oscillator for 48 h. The excess of alkali was back
titrated with 0.1 mol/L hydrochloric acid with phenolphthalein as
an indicator. The titration process for each sample was repeated
three times and the repeated results were calculated to obtain the
average value. DD was calculated by Eq. (1):
DD %
V
2
V
1
1 u
0

V
0
V
1
1 u
1

100% (1)
Where V
0
is the average value of the volume of hydrochloric acid
consumed for the blank in liters, V
1
is the average value of the
volume of hydrochloric acid consumed for the sample in liters, V
2
is
the average value of the volume of hydrochloric acid consumed for
the KGM powder in liters, u
0
is water content of KGM powder, u
1
is
water content of deacetylated KGM.
2.2.4. Kinetics of heterogeneous deacetylation of KGM
Deacetylation of KGM is actually a hydrolytic reaction of ester,
namely, a typical nucleophilic substitution reaction. Assuming that
the deacetylation of KGM is a pseudo rst order reaction of which
the reaction rate is only concerned with the content of acetyl group,
the dynamical equation could be expressed as Eq. (2) and Eq. (3):
ln1 DD kt (2)
ln k E
a
=RT B (3)
where k is reaction rate constant, t is reaction time, T is reaction
temperature (absolute temperature), E
a
is activation energy, R is gas
constant.
2.3. Characterization
2.3.1. FT-IR analysis
Fourier transform infrared (FT-IR) measurements of KGM sam-
ples were carried out at a FT-IR spectrometer (Nexus 470, Nicolet,
USA) at a resolution of 4 cm
1
in the range 400e4000 cm
1
. KGM
samples were prepared as KBr discs and were scanned against an
air background (Du et al., 2012).
2.3.2. DSC analysis
The thermal analyses of the KGM samples were carried out with
a model DSC 204 F1 (Netzsch, Germany). Temperature and
enthalpy of the instrument were calibrated by pure indium
(99.99%). 2.0 0.1 mg of the samples in an aluminumcrucible with
a reference sample of an empty crucible were analyzed in a dried N
2
gas atmosphere at a heating rate of 10

C min
1
in a temperature
J. Li et al. / Food Hydrocolloids 40 (2014) 9e15 10
range of 25e400

C. Sweep gas rate was 20 ml min

1
, protective gas
rate was 60 ml min
1
.
2.3.3. XRD analysis
X-ray diffraction patterns (XRD) were obtained by Rigaku D/
Max-IIIA X-Ray Diffractormeter. Samples were analyzed between
2q 4

and 2q 60

with a step size 2q 0.02

using a Cu Ka
1
radiation (l 1.54184), 20

C, 40 KV and 50 mA. The diffractometer

was equipped with 1

divergence slit and a 0.1 mm receiving slit.

2.4. Solubility experiments
2.4.1. Effect of deacetylation degree on solubility of KGM
A certain amount of sample powder (0.10 g) with 24.90 g ice was
dispersed in a beaker. The mixture in an ice bath (0 0.5

C) was
stirred at a lowspeed until all the ice melted. After the mixture was
centrifuged for 20 min at 4500 rpm, 10.00 g of supernatant in a
weighed ask was dried to constant weight at 105

C. The process
for each sample was repeated for 3e5 times and the repeated re-
sults were calculated to obtain the average value. Solubility was
calculated by Eq. (4) and water content was calibrated (Xie, Liu, &
Chen, 2007).
Solubility%
m2:5
W
100% (4)
Where m is the dry matter content in the ask, W is the total mass
of the sample.
2.4.2. Effect of temperature on solubility of Da-KGM
A certain amount (0.1 g) of Da6, Da5 or Da3 sample powder with
24.90 g deionized water was dispersed in a beaker. The mixture was
put in 85

C, 55

C, 25

C, 10

C, 4

C water baths and 0

C, 4

C ice
salt baths, respectively. After stirring for 1 h, the mixture was
centrifuged for 20 min at 4500 rpm. Subsequently, 10.00 g of su-
pernatant in a weighed ask was dried to constant weight at 105

C.
The process for each sample was repeated for 3e5 times and the
repeated results were calculated to obtain the average value. Sol-
ubility was also calculated by Eq. (4).
3. Results and discussions
3.1. Heterogeneous deacetylation of KGM
3.1.1. Effect of the ratio of alkali to acetyl groups on deacetylation
degree of KGM
It wasprovedthat theremoval of acetyl groupin1gKGMneededto
consume 3.45 10
4
mol of OH

, which was identical to Maekajis

report (Maekaji, 1978b). The molar ratios necessary to remove all the
acetyl groups were calculated as mKGM:mNaOH 77:1,
mKGM:mKOH 56:1, mKGM:mCa(OH)
2
82:1, mKGM:mNa
2
CO
3
57:1. The experiment was set up such that molar ratio of OH

and acetyl group was 1:8, 2:8, 4:8, 6:8, 8:8 and 16:8, respectively.
It can be seen from Fig. 1a that higher alkali amount gave
benet to the deacetylation, but it was still very hard to remove
all acetyls unless excessive alkali was added (nNaOH/
KOH:nacetyl 16:8). The effect of the alkalis on deacetylation
ranked as follows: NaOH > Ca(OH)
2
> Na
2
CO
3
. KOH was gener-
ally second only to NaOH with a couple of exceptions. This result
could be attributed to the fact that deacetylation was a hydrolytic
reaction and the basic strength determines hydrolytic ability.
Here, hydrolytic ability of KOH was weaker than NaOH, which
might be explained by the degree of ionization of KOH compared
with NaOH in alcohol medium. However, the reason was still
unclear.
3.1.2. Effect of ethanol concentration on deacetylation degree of
KGM
KGM can be soluble in water while insoluble in ethanol. There
were two advantages of deacetylation in ethanol medium. One was
that ethanol could slowdown the penetration of alkali by hindering
the swelling of KGM. The other was to promote deacetylation by
improving alkalinity of the system. The latter was commonly
mentioned in studies of deacetylation of chitin which can be
explained by generation of sodium ethoxide facilitated deacetyla-
tion (Cho, Jang, Park, & Ko, 2000). As can be seen in Fig. 1b, DD
increased moderately and then declined slightly as the concentra-
tion increased, certainly because before the concentration of
ethanol reached 80%, deacetylation was dominated by the increase
of alkali, yet when the concentration of ethanol was higher (>80%),
deacetylation was mainly subjected to the effect that ethanol could
slow down the penetration of alkali by hindering the swelling
of KGM.
3.1.3. Effect of reaction temperature on deacetylation degree of KGM
Fig. 1c indicated that deacetylation was facilitated by the
increasing temperature. This could be related to the fact that higher
temperature caused increasing thermal motion of OH

and facili-
tated penetration to internal macromolecules, which accelerated
attacking frequency towards the ester group.
3.1.4. Effect of reaction time on deacetylation degree of KGM
Fig. 1d showed that DD increased as time extended. This was
dependent upon the basic principles of deacetylation of KGM: with
the prolongation of time, the reaction time of OH

and ester
extended, giving rise to increasing reaction probability. The DD
increased sharply within the rst hour (from 0 to 64.06%), while in
the following time (1e72 h), DD increased gently with time pro-
longing. This was because deacetylation is a typical nucleophilic
substitution reaction which was related to not only the concen-
tration of substrate, but also concentration of the product. At the
beginning, DD of the product was low, leading to a quick reaction.
As the DD increased with time, there was a drop of the number of
acetyl group in the product, which caused the difculty for the
nucleophilic substitution reaction to carry on.
3.1.5. Kinetics of heterogeneous deacetylation
Fig. 2a indicated that reaction rate increased with reaction
temperature from 0 to 10 min with the apparent reaction rate
constant being 4.88 10
2
at 30

C, 8.54 10
2
at 50

C and
9.97 10
2
min
1
at 70

C, respectively. Assuming that the reaction

was rst order reaction, according to Empirical Arrhenius: ln
k E
a
/RT B, Fig. 2b showed that the apparent reaction rate
constant was 15.59 kJ/mol, lower than 49.32 kJ/mol in homoge-
neous system (Maekaji, 1978a), suggesting that ethanol promoted
the deacetylation. From10 to 60 min, the reaction still followed rst
order reaction, but the apparent reaction rate constant decreased
signicantly being 6.3 10
3
at 30

C, 5.7 10
3
at 50

C and
6.1 10
3
min
1
at 70

C, respectively. There was not marked
difference as temperature changed.
3.1.6. FT-IR spectrum analyses
The deacetylation degree of sample Da0, Da1, Da2, Da3, Da4, Da5
and Da6 was 0.00%, 16.87%, 32.58%, 51.90%, 67.32%, 80.51% and
98.28%, respectively. As can be seen from Fig. 1 of our other article
(Du et al., 2012), the KGM backbone was not changed by deacety-
lation and the remarkable difference among the FT-IR spectra of
KGM with different DD was due to the decrease or elimination of
the characteristic absorption bands of KGM at approximately
1730 cm
1
. The absorption at 1730 cm
1
has been assigned to C]O
(carbonyl group) by many authors (Jacon, Rao, Cooley, & Walter,
J. Li et al. / Food Hydrocolloids 40 (2014) 9e15 11
1993; Maekaji, 1974) and its removal from the KGM backbone ini-
tiates the polymer chain interactions to form a gel (Zhang et al.,
2001). Moreover, the peak of CeC(]O)eO at 1252.1 cm
1
showed the same change trend, yet with smaller change scale.
However, the peak of eOH at 3434.5 cm
1
showed a somewhat
wider tendency with DD increasing, which is distinct from the
earlier studies (Koroskenyi & McCarthy, 2001; Pan et al., 2008). It is
probably due to the fact that content of acetyl group in raw KGM
was so low that the content of generated eOH through deacetyla-
tion cannot be high. In spite of all this, other absorption peaks did
not change which indicated that the molecular backbone of Da-
KGM was basically consistent with the raw material.
3.2. DSC analyses
The result of DSC analysis was listed in Table 1. The strong hy-
drophilicity of KGM is attributed to the large number of hydroxyl
groups and it was hard to remove all moisture by drying. The
endothermic peak was attributed to evaporation of water mole-
cules residing in the KGM, which roughly reected water holding
capability. It can be seen clearly that Da-KGM showed a lower
temperature of endothermic peak as DD increased, suggesting the
decrease of water holding capacity. But there were exceptions,
which were explained by the water-holding capability of the
sample being also closely related to its states, especially the way
and degree of dehydration in sample preparation.
There are numerous structural hydrogen bonds in poly-
saccharides. As a result, the reaction of decomposition, chain
breaking, oxidation or dehydration cross-linking took place
frequently which caused exothermic effect when increasing
temperature to break hydrogen bonds (Yang, Xiong, & Zhang,
2002). In DSC thermograms of KGM with different DD, two un-
equal exothermic peaks appeared between 200

C and 400

C
(Table 1), which resulted from the pyrolysis of KGM.
Generally speaking, smaller sample crystallinity led to better
water-solubility and lower exothermic peak temperature. Theo-
retically, decreased acetyl group content of KGM contributed to
decreased steric hindrance, strengthened intermolecular in-
teractions and risen difculty of thermal decomposition. It can be
seen clearly that the Da-KGM showed a higher temperature of
exothermic peak than the native KGM. Yet the temperature of
exothermic peak of KGM with different DD did not increase regu-
larly with the increase of DD. Moreover, as can be seen in Table 1,
the relationship between endothermic or exothermic peak and DD
was not linear, nor did the relationship between heat enthalpy
changes and DD, which was not consistent with our expectations. A
linear relationship was only found in a very limited range. Overall,
thermal characteristics of each sample were similar. Due to
deacetylation in ethanol medium, KGM samples only swelled
slightly so that there wasnt molecule elongation and rearrange-
ment, which led to the fact that DD has no marked effect on mo-
lecular structure aggregation as was observed in the homogeneous
system.
3.3. XRD an1alyses
Fig. 3 showed the X-ray diffraction patterns of Da-KGM. All
samples exhibited a broad diffuse scattering at about 20

C (2q)
which corresponds to d 4.57

A, and there is only very subtle
distinction in the position and strength of the diffuse scattering.
Fig. 1. (a) Effect of the ratio of alkali to acetyl groups on DD of KGM (C
Ethanol
50%; 40

C; 4 h); (b) Effect of ethanol concentration on DD of KGM (nOH:nacetyl groups 8:8; 0

C; 4 h);
(c) Effect of reaction temperature on DD of KGM (nOH:n acetyl groups 8:8; C
Ethanol
50%; 24 h); (d) Effect of reaction time on DD of KGM (nOH:n acetyl groups 8:8; C
Ethanol
50%;
40

C).
J. Li et al. / Food Hydrocolloids 40 (2014) 9e15 12
The result indicated that deacetylation inuenced crystal structure
of KGM just in a small range. Overall, both the Da-KGM and native
KGM presented amorphous state, suggesting that molecular inter-
action was very weak and molecular arrangement was looser. This
could be attributed to the fact that KGMwas composed of two kinds
of monosaccharide with acetyl group on the chain segments, which
made the molecule chain lack steric regularity (Dave, Sheth,
McCarthy, Ratto, & Kaplan, 1998). The crystalline properties could
be described as unordered in the long-range, ordered in the short-
range (Brindley, 1980).
3.4. Solubility experiments
3.4.1. Effect of deacetylation degree on solubility of KGM
Fig.4 showed a very interesting phenomenon: solubility of KGM
decreased with DD increasing at 0

C. However, was it the real
Table 1
Data from DSC of KGM with different deacetylation degree.
Sample Endothermic peak The rst exothermic
peak
The second exothermic
peak
Peak value
(

C)
Enthalpy
(J/g)
Peak value
(

C)
Enthalpy
(J/g)
Peak value
(

C)
Enthalpy
(J/g)
Da0 102.7 228.9 243.2 61.33 320.5 14.49
Da1 101.8 206.2 288.0 34.41 328.5 18.02
Da2 98.32 196.1 286.3 33.34 330.5 9.648
Da3 101.6 193.4 292.3 18.28 328.2 13.84
Da4 97.42 178.0 294.1 31.95 329.9 4.178
Da5 82.22 254.3 286.7 62.54 329.1 6.867
Da6 76.98 125.7 280.6 32.18 330.5 29.84
Fig. 3. XRD proles of KGM with different deacetylation degree.
Fig. 2. Effect of temperature on deacetylation of KGM (a); and the curve of ln k to 1/RT (b).
J. Li et al. / Food Hydrocolloids 40 (2014) 9e15 13
relationship between DD and solubility? Was it possible that
deacetylation only occurred in part of KGM which led to the fact
that the completely deacetylated section was insoluble while the
natural fraction was soluble? Maybe the results conveyed only an
average picture. For this reason, plenty of soluble fraction and
insoluble fraction in mixture were collected followed by using acid-
base titration and FT-IR to determine acetyl group content. Fig.5
showed the FT-IR results of several typical samples. No signicant
difference was found in the acetyl group content between soluble
and insoluble fractions. Therefore, it can be maintained that the
relationship between DD and solubility was an intrinsic attribute
of KGM.
It is worth noting that most of non-substituted b-(1 /4) con-
nected glucomannan and dextran are insoluble because of the
intermolecular hydrogen bonding (Dea & Morrison, 1975; Ratcliffe,
Williams, Viebke, & Meadows, 2005). Yet KGM is soluble as a
member of the family. Dea and Morrison (1975) indicated that KGM
was soluble for that steric effect of acetyl group led to a high degree
of entropy between molecular chains. However, Brownsey et al.
didnt agree with this view. They simply considered that acetyl
group hindered the formation of intermolecular hydrogen bonding
(Ridout, Cairns, Brownsey, & Morris, 1998). Whatever the reason is,
according to previous studies and experimental results, we partly
assign solubility of KGM to the presence of acetyl group.
Now, a clear impression was that acetyl group endowed KGM
with solubility, yet there was a scarcity of reports on the quanti-
tative relationship between acetyl group content and solubility.
Koroskenyi and McCarthy (2001) indicated that solubility of KGM
exponentially declined as DS (degree of substitution) increased.
However, some other researchers clearly pointed out that the
increasing content of acetyl group promoted the dissolution of KGM
(Gao & Nishinari, 2004b; Huang, Takahashi, Kobayashi, Kawase, &
Nishinari, 2002).
It seemed that the two inferences failed to cohere. But when
searching to the root, we discovered that the DS of the former was
higher (0.5e3), while the latter was comparatively lower (0.05e
Fig. 4. Effect of deacetylation degree on solubility of KGM.
Fig. 5. Comparison of FT-IR spectra of acetyl group from soluble (upper curves) and
insoluble (lower curves) fractions (From left to right: Da1, Da3 and Da5, respectively). Fig. 6. Effect of temperature on solubility of Da-KGM: (a) Da3; (b) Da5; (c) Da6.
J. Li et al. / Food Hydrocolloids 40 (2014) 9e15 14
0.42). Further relating it with our study, we speculated that: KGM
showed good solubility only within a very narrow range of acetyl
group content which changed slightly compared with the acetyl
group content of natural KGM.
The speculation could be attributed to the fact that acetyl group
was a kind of hydrophobic group. High contents resulted in
reduction in solubility, which was consistent with Koroskenyis
study. But when the acetyl group content was moderate, strong
steric effect promoted the dissolution by hindering molecular chain
association. This was in conformity with the result of Huang et al.
(2002) and Gao and Nishinari (2004b). To conclude, our results
can be well explained that as acetyl group content changed from
moderate to low, steric effect decreased, so did the solubility.
3.4.2. Effect of temperature on solubility of Da-KGM
Effect of temperature on solubility of Da-KGM was shown in
Fig.6. It was indicated that lower temperature promoted dissolu-
tion, which further proved that solubility of KGM decreased with
DD. If ice instead of water was used to dissolve KGM, the promoting
effect would be even more obvious. Maekaji (1973) pointed out that
peptization of KGM gel was easier to be carried out at lower tem-
perature. Later, Perols et al. (1997) embedded enzyme into the gel
and released at 4

C. If KGM gel and Da-KGM powder were
considered as the same substance (the strength of intermolecular
interaction is different), the dissolution of Da-KGM was in accor-
dance with the peptization of KGM gel.
4. Conclusion
The optimal conditions have been determined to prepare het-
erogeneous deacetylated KGM. Heterogeneous deacetylation fol-
lowed rst-order kinetics and its apparent activation energy
(15.59 kJ/mol) was lower than that of the homogeneous system
(49.32 kJ/mol), suggesting ethanol promoted the deacetylation. FT-
IR analyses showed that the remarkable difference of KGM with
different DD was decrease or elimination of the peak at approxi-
mately 1730 cm
1
. The Da-KGM displayed a lower temperature of
endothermic peak and higher temperature of exothermic peak, yet
without showing a regular change with the increasing DD. It can be
deduced that the crystalline properties of Da-KGM had no
remarkable distinction from the native KGM. Solubility of KGM
exponentially declined as DD increased. In addition, lower tem-
perature was helpful for dissolution.
Acknowledgments
The authors gratefully acknowledge nancial support from
contract grant sponsors: The National Natural Science Foundation
of China (Grant No. 31371841).
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