Studies on the Degradation Behavior of Chitosan-g-Poly (acrylic acid) Copolymers

Tamkang Journal of Science and Engineering, Vol. 5, No. 4, pp. 235-240 (2002) 235

Studies on the Degradation Behavior of Chitosan-g-Poly
(acrylic acid) Copolymers

Trong-Ming Don1, Chung-Yang Chuang2 and Wen-Yen Chiu2

1Department of Chemical Engineering
Tamkang University,
Tamsui, Taiwan 251, R.O.C.
E-mail: tmdon@mail.tku.edu.tw
2Department of Chemical Engineering
National Taiwan University,
Taipei, Taiwan 106, R.O.C.



Abstract

Graft copolymers of Chitosan and poly(acrylic acid) were
synthesized with a redox initiator, cerium ammonium nitrate. After 2 h
of reaction at 70 oC, a gel product was obtained. The swelling ratio of
the copolymer in water depends on the pH value and has a maximum
in the buffer solution at pH 7. This is because the morphological
structure of the copolymers in the water changes with the pH values.
In addition, the swelling ratio of the copolymer increases with the
feeding amount of acrylic acid monomer. The degradation behavior of
the copolymers was observed both in a lysozyme solution and an
active slurry solution. The degradation rate was higher in the
lysozyme solution and depended on the composition of the
copolymers.

Key Words: Chitosan, Poly(acrylic acid), Graft Copolymer,
Degradation, Lysozyme, Slurry

1. Introduction
solvent. Yet, chitosan can be dissolved in an acid
solution and becomes a cationic polymer because
Hybridization of natural polymers with
of the protonation of amino groups on the C-2
synthetic polymers is of great interest because of
position of pyranose ring.
their application to biomedical and biodegradable
In this study, cerium ammonium nitrate
materials [1]. One of the natural polymers that
(CAN) was used to initiate the graft copoly-
have attracted great attention recently is chitosan.
merization of acrylic acid (AA) onto chitosan.
Chitosan is a high-molecular-weight poly-
Hopefully, new material with desired properties
saccharide composed mainly of ?-(1,4) linked
can be achieved by the chemical combination of
D-glucosamine and partially of ?-(1,4) linked
natural and synthetic polymers. PAA has been used
N-acetyl-D-glucosamine. It is generally prepared
in the adhesives and super-absorbent polymers,
by the partial deacetylation of chitin in a hot alkali
because of the pendant carboxyl groups. Chitosan
solution. Chitin is the most abundant natural
is a material with several important advantages
polymer next to the cellulose and can be found in
including biocompatibility, biodegradability and
the skeletal materials of crustaceans and insects,
anti-bacterial. Both of them can absorb a great
and cell walls of bacteria and fungi. Chitin and
amount of water. Therefore, chemical combination
chitosan can be used in the fields of wastewater
of chitosan and PAA might have potential in the
treatment, food processing, cosmetics, pharmace-
use of the super-absorbent material with
uticals, biomaterials and agriculture [2,3]. With its
anti-bacterial and biodegradable properties. The
fibrous structure, chitin is hardly soluble in any
water absorption and degradability behavior, were

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236 Trong-Ming Don et al.
thus examined in this article.
Table 1. Reaction conditions and the sample codes of
various membranes
2. Experimental
System CAC510
CAC520
CAC540
2.1 Materials
CS (g)
5
5
5
Chitosan was obtained from Tokyo
AA (g)
10
20
40
Chemicals Inc., Tokyo. The degree of
CAN (mole)
0.01
0.01
0.01
deacetylation was found to be 86% by a colloid
titration method [4], and the viscosity average
H2O (g)
500
500
500
molecular weight was found to be 616,000 by a
Temperature (oC)
70 70 70
viscometric method, where the Mark-Houwink
constants k and ? were 1.38 x 10-2 and 0.85,
2.3 Swelling Studies
respectively [5]. AA monomer from Tedia
Chemical Company was distilled under reduced
Samples were immersed into several buffer
pressure. Only the distillate obtained at the
solutions with different pH values at 30 oC to observe
middle stage of distillation was used for
the swelling behavior. The swollen samples were
polymerization. CAN, (Ce(NH
removed at various time intervals, and the excess
4)2(NO3)6), was a
reagent-grade from Showa Chemical Inc. All the
water was removed from the sample surface with filter
other chemicals were analytical-grade or above
paper. The weight of swollen samples was measured,
and used as received without further purification.
and the samples re-immersed into the buffer solution.
Lysozyme was obtained from Merck (Germany)
The procedure was repeated until there was no further
and has an activity of 100,000 U/mg. The active
weight increase. The swelling ratio was calculated by
slurry was obtained from a local bakery
the weight of swollen sample at a specific time divided
company and its bacterial density was 2.4 x 107
by the initial weight in the dry state.
CFU/mL.
2.4 Degradation Behaviors
2.2 Synthesis of Graft Copolymer
Thermal degradation behavior was investigated
A specific amount of chitosan was first
with a method of thermal gravimetric analysis (TGA-7
dissolved in the aqueous solution of acrylic acid.
from Perkin-Elmer). The weight loss of sample was
The solution was purged with nitrogen and
measured in a temperature range of 30 to 550 oC with
heated to 70 oC in an isothermal water bath.
a heating rate of 10 oC/min. To investigate the
The cerium ammonium nitrate was dissolved in
enzymatic degradation of the copolymer sample, a
20 mL of water and pre-heated to 70 oC, before
buffer solution containing 0.025 M Na2HPO4/KH2PO4
it was poured into the solution. After 2 hours of
was prepared by adjusting the pH value to 7 with a 0.1
reaction, chitosan-g-PAA copolymer was
M HCl solution at 30 oC. Approximately 0.1-g sample
formed and the solution became a gel. The gel
membrane was put into a 30 mL of the above buffer
solution was taken out for dialysis to remove
solution. After the addition of 0.01 g of lysozyme, the
the residual ceric ion and all other small
sample membrane was taken out at several interval
molecules. To prepare the sample membranes
times, rinsed with the deionized water repeatedly and
for the test of swelling and degradation
then dried. The weight loss was measured to evaluate
properties, the gel was put into a stainless mold
the degradation. In another experiment to investigate
and then dried for 48 h in a circulation oven
the degradation behavior in active slurry, 0.1-g sample
followed by another 48 h in a vacuum oven at
membrane was put into a beaker with a 30-mL of TSB
60oC. The reaction conditions and the sample
solution, followed by the addition of 5 mL of the
codes of membranes are listed in Table 1. In
active slurry solution. The beaker purged with air was
addition, the conversion was calculated with the
set in an isothermal shaker at 100 rpm and 25 oC. After
following equation.
a certain period of time, the sample was taken out,
rinsed, dried and weighed.
W - W
X (%) ?
1
CS ×

100 (1)

W
3. Results and Discussion
AA
where WCS and WAA are the initial weight of
3.1 Synthesis of the Copolymer
chitosan and AA monomer, respectively, and W1 is
the final weight of drying product.
Ceric ion (Ce4+) is a strong redox initiator, which

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Studies on the Degradation Behavior of Chitosan-g-Poly(acrylic acid) Copolymers 237

can oxidize the pyranose ring of polysaccharide
resulting in the formation of a free radical on the ring
+ + +
-
-
[6-10]. Chitosan-g-PAA thus can be obtained by the
-
-
-
-
-
-
grafting of acrylic acid monomer onto the chitosan
-
-
-
-
-
-
following the traditional free radical polymerization
-
+ + +
procedure. In addition, poly(acrylic acid) homo-
polymer is also formed during the polymerization,
- -
through the chain transfer to monomer. The
- -
- -
conversion of acrylic acid monomer after 2 h of
- -
reaction increased from 80 % to 91 %, as the monomer
feed increased from 10 g to 40 g based on 5 g of
chitosan (Figure 1). The increase in conversion was
CS
PAA

Figure 2. Proposed structure for the copolymer gel
due to the decrease in solution viscosity when more
acrylic acid was added. In all the cases, a gel was
3.2 Swelling Behavior
formed after the reaction. A structure similar to the
cross-linked network is proposed for the reaction
Figure 3 shows the swelling behavior of the
product, Figure 2. When chitosan was dissolved in the
sample membranes (2-mm thickness) in several
acrylic acid solution, some of the amino groups
buffer solutions with different pH values. The
became positive-charged due to the reaction with the
swelling ratio depends on pH values and the
proton dissociated from the acrylic acid, NH
amount of monomer feed to the reaction system.
2 + H+ ?
NH +
For the pure chitosan, the swelling ratio increased
3 . At the same time, the grafted PAA became
negative-charged from the carboxylate groups, COOH
as pH value decreased. This is because in a more
? COO- + H+. As a result, both chitosan and grafted
acidic solution, more amino groups become
PAA chains were extended and more rigid because of
protonated. At pH 4, 95% of amino groups exist as
+
the repulsive force from the same charges along the
positive-charged NH3 [11]. These positive-
chains. However, when the extended PAA chains
charged groups along the chitosan chains exert
intersect with the chitosan chains, ionic force between
repulsive force, which consequently extend the
the negative-charged carboxylate group and the
chitosan chains. Therefore, it can absorb more
positive-charged amino group could hold them
water; since the structure loosen up. In addition,
together and served as a cross-linking point. Since both
the charges in the chitosan chains result in an
of them are hydrophilic polymers, they can absorb a
increase in the ion concentration and thus the
lot of water and lock these water molecules in this
osmotic pressure inside the structure, which makes
pseudo cross-linked structure.
the water easily to diffuse into the structure due to
the thermodynamic driving force. Because of these

reasons, the swelling ratio is higher in the acidic
100
solution.
For the copolymer samples, the swelling
behavior is also dependent on the structure of the
80
sample, which changes with pH value. It is
mentioned previously that the reaction product

)
60
became a gel with a lot of water inside the
(%)
structure and a pseudo cross-linked structure was

r
s
i
on (%
proposed. However, in the buffer solution at pH 4,
40
ersion
onve
the dissociation of carboxylic acid in PAA is
C
nv
CAC510
suppressed, where only 20% of the carboxylic acid
CAC520
Co
CAC540
dissociate [12,13]. As a result, most of the PAA
20
chains are not as rigid as that in pH 7 solution and
also fewer cross-linking points is expected.
0
Therefore, the swelling ratio was lower than that in
0
30
60
90
120
150
the neutral solution. At high pH value as in the
Time (min)
Time (min)
basic solution, chitosan chains no longer bear
charges, though most carboxyl groups in PAA
Figure 1. The effect of AA monomer amount on the
dissociate. The pseudo cross-linked structure again
conversion as function of reaction time
collapses. Therefore, the copolymer sample at pH

---

238 Trong-Ming Don et al.
7 had the higher swelling ratio than those at pH 4
350 oC. The third-stage degradation behavior from
or pH 9. In addition, the swelling ratio increased
350 to 525 oC was mainly due to the chain scission
with the AA monomer feed, which reached 32
both in PAA and chitosan. It was found that the
when the monomer feed is 40 g based on 5 g of
higher the monomer feed, the lower the char yield
chitosan.
at 550 oC. This is because the char yield of pure
PAA is much lower than that of pure chitosan.

35

CAN system
100
30
pH 4
pH 7
pH 9
25
80
Chitosan flake
PAA
20
a
t
i
o
)
60
g ratio ng r


(
%
elli 15
ss (%) ss
w
S

t
lo
ht lo
e
ig
40
10
Swellin
W
Weigh
5
20
0
0
10
20
30
40
AA feed (g)
0
150
250
350
450
550
AA feed (g)

o
Temperature ( C)
Figure 3. The effect of pH value on the swelling ratio of
Temperature (?)
sample membrane as function of AA feed
Figure 4. The weight loss of pure chitosan and PAA as
function of temperature
3.3 Thermal Degradation Behavior
The thermal degradation behavior of pure

100
chitosan and PAA are shown in Figure 4.
CAC510d
Two-stage degradation behavior was observed for
CAC520d
PAA. The first stage, in the range of 250 to 350 oC,
80
CAC540d
was probably caused by the dehydration of
carboxylic acid and decarboxylation. The second
60
stage mainly after 350 oC was due to the chain
(%)
ss (%)
scission in the main chain. Chitosan started to

ht loss
t
lo
degrade at 250 oC and had a broad degradation
40
Weig
temperature range with a high char yield at 550 oC.
Weigh
The degradation mechanism is very complex
20
including the dehydration, deacetylation and chain
scission. For the copolymer samples, the ceric ion
0
initiator had to be removed by a dialysis method
150
200
250
300
350
400
450
500
550
before running any thermal degradation studies.
o
Temperature ( C)
Temperature (?)
Otherwise, it would oxidize chitosan and PAA at
high temperatures. The thermal degradation curves
Figure 5. The weight loss of several sample membranes
of various copolymer samples are shown in Figure
as function of temperature
5, where three stages were observed. The
first-stage degradation from 200 to 250 oC was
3.4 Enzymatic Degradation
attributed to the loss of water due to the amidation
Lysozyme can interact with chitosan and
from the carboxylate group in PAA with the
facilitate the hydrolysis of chitosan, especially in
associated positive-charged amino group in
an acid solution at pH 5-6. Therefore, lysozyme
chitosan. This amidation was evidenced by some
was used to evaluate the enzymatic degradation
other techniques such as Infrared spectro-
of the copolymer samples and the results are
photometer, which are not shown here [14]. Both
shown in Figure 6. The weight loss increased
the deacetylation of chitosan and the dehydration
with the degradation time for a CAC510 sample
as well as the decarboxylation of PAA chains
membrane until it reached a plateau value, which
caused the second-stage degradation from 250 to

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Studies on the Degradation Behavior of Chitosan-g-Poly(acrylic acid) Copolymers 239

was about one third of the initial weight after 4
sample, and the latter was the secreted enzymes
days. Yet, the rate was slower at the first two
by the microorganisms.
days, and then increased in the following two
days. After that, no more degradation occurred,
indicating that the remaining was not degradable

50
by the lysozyme. In the same Figure, it was
found that the weight loss of the CAC510 was
only slightly smaller than the pure chitosan. It is
40
assumed that the weight loss was mainly caused
or at least initiated from the degradation of
30
chitosan, since the experimental result shows
ss (%)
that pure PAA does not degrade by the lysozyme.

e
ight loss (%)
If the weight loss was re-calculated based only
t
lo W 20
on the chitosan component in the copolymer (the
Lysozyme
weight loss based on the whole sample divided
CS
Weigh 10
CAC510d
by the weight content of the chitosan component,
Table 2), a value of 85% was obtained for the
0
CAC510 sample. This means that 85% of the
0
1
2
3
4
5
6
7
chitosan component degraded away, which is
Time (day)
much higher than the pure chitosan. If this were
Time (day)

true, a conclusion would be reached that the
Figure 6. The weight loss of pure chitosan CS and a
degradation rate of chitosan was higher in the
sample CAC510 under enzymatic degrada-
copolymer than the pure form, which is not
tion of lysozyme as function of time
necessary true. Further investigation from the

FTIR and elemental analysis of the dissolved
Table 2. Weight loss of various samples at 48 and 96 h
fragments in the solution, the evidence of PAA
of lysozyme degradation
chain was actually found. Therefore, it is more
Sample CAC510
CAC520
CAC540
plausible to assume that some of the grafted
CS content in
PAA chains, especially the short ones, are easily
38.2 %
22.4 %
12.2 %
the copolymer
dissolved into the water accompanied with the
Weight loss
degraded chitosan fragments. This would
12.3 %
4.7 %
3.6 %
(%) at 48 h
increase the weight loss of the copolymer sample.
Weight loss
In addition, if the monomer feed was increased,
32.8 %
9.8 %
6.1 %
(%) at 96 h
chance of forming short grafted PAA chains was
less. Therefore, the weight loss for these systems
should be smaller as can be seen in Table 2, if

50
one compares CAC510, CAC520 and CAC540
samples.
Active slurry
CS
There are many microorganisms in the
40
CAC510d
active slurry and some of them can contact and
adhere to the surface of the sample membranes.
) 30
The enzymes secreted from these micro-
ss (%) s (%

organisms may degrade the copolymer samples
t
lo
20
e
ight los
and, the degradation fragments can be served as
W
the nutrition or carbon source for the micro-
Weigh
10
organisms. Figure 7 shows the weight loss of the
copolymer samples in an active slurry solution
increased with the degradation time. The weight
0
0
2
4
6
8
10
12
14
16
loss was higher for the copolymer samples than
Time (days)
the pure chitosan under the same conditions.
Time (day)

However, compared with the results in the
Figure 7. The weight loss of pure chitosan CS and a
directly enzymatic degradation, the degradation
sample CAC510 under degradation in active
rate was slower. This is because the former was
slurry solution as function of time
the direct use of the enzyme to degrade the

---

240 Trong-Ming Don et al.
4. Conclusions
[10] Don, T.-M., Hsu, S.-C. and Chiu, W.-Y., J.
Appl. Polym. Sci., Vol. 86, pp. 3057 (2002).
Chitosan-g-poly(acrylic acid) copolymers
[11] Kienzle-Sterzer, C. A. Ph. D. Thesis,
were successfully synthesized with the ceric ion.
Massachusetts Institute of Technology,
After 2 h of reaction at 70 oC, a gel product was
Cambridge, U.S.A. (1984).
obtained and a pseudo cross-linked structure was
[12] Nagasawa, M., Murase, T. and Kondo, K., J.
proposed. The swelling ratio of the copolymer has
Phys. Chem., Vol. 69, p. 4005 (1965).
a maximum value in the buffer solution at pH 7. In
[13] Ricka, J. and Tanaka, T. Macromolecules,
addition, the swelling ratio of the copolymer
Vol. 17, p. 2916 (1984).
increases with the feeding amount of acrylic acid
[14] Chuang, C.-Y., M.S. thesis, National Taiwan
monomer. For the sample with 40 g of AA
University, Taipei, Taiwan (2001).
monomer feed based on 5 g of chitosan, the

swelling ration reached 32 at pH 7. The thermal

degradation of the copolymer samples started at

200 oC and exhibited three stages of degradation.
Manuscript Received: Oct. 16, 2002
The degradation behavior of the copolymers was
Revision Received: Nov. 6, 2002
observed both in a lysozyme solution and an active
and Accepted: Nov. 11, 2002
slurry solution. Yet, the degradation rate was

higher in the lysozyme solution and depended on

the composition of the copolymers.


Acknowledgment

The authors wish to express their appre-

ciation for the financial support of the National

Science Council, Republic of China (Project No:

NSC 90-2313-B-032-001-).


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Studies on the Degradation Behavior of Chitosan-g-Poly(acrylic acid) Copolymers 241

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