Synthesis, characterization, and antibacterial activity of N,O-quaternary ammonium chitosan

Article abstract of DOI:10.1016/j.carres.2011.08.002

N,N,N-Trimethyl O-(2-hydroxy-3-trimethylammonium propyl) chitosans (TMHTMAPC) with different degrees of O-substitution were synthesized by reacting O-methyl-free N,N,N-trimethyl chitosan (TMC) with 3-chloro-2-hydroxy-propyl trimethyl ammonium chloride (CHPTMAC). The products were characterized by ¹H NMR, FTIR and TGA, and investigated for antibacterial activity against Staphylococcus aureus and Escherichia coli under weakly acidic (pH 5.5) and weakly basic (pH 7.2) conditions. TMHTMAPC exhibited enhanced antibacterial activity compared with TMC, and the activity of TMHTMAPC increased with an increase in the degree of substitution. Divalent cations (Ba²⁺ and Ca²⁺) strongly reduced the antibacterial activity of chitosan, O-carboxymethyl chitosan and N,N,N-trimethyl-O-carboxymethyl chitosan, but the repression on the antibacterial activity of TMC and TMHTMAPC was weaker. This indicates that the free amino group on chitosan backbone is the main functional group interacting with divalent cations. The existence of 100 mM Na⁺ slightly reduced the antibacterial activity of both chitosan and its derivatives.

Full text of DOI:10.1016/j.carres.2011.08.002

Carbohydrate Research 346 (2011) 2445–2450
Contents lists available at SciVerse ScienceDirect
Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Synthesis, characterization, and antibacterial activity of N,O-quaternary
ammonium chitosan
a
a
a,
⇑
b
a
a
Tao Xu , Meihua Xin , Mingchun Li , Huili Huang , Shengquan Zhou , Juezhao Liu
a
College of Material Science and Engineering, Huaqiao University, Xiamen 361021, China
College of Chemical Engineering, Huaqiao University, Xiamen 361021, China
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 2 June 2010
Received in revised form 15 July 2011
Accepted 2 August 2011
Available online 11 August 2011
N,N,N-Trimethyl O-(2-hydroxy-3-trimethylammonium propyl) chitosans (TMHTMAPC) with different
degrees of O-substitution were synthesized by reacting O-methyl-free N,N,N-trimethyl chitosan (TMC)
with 3-chloro-2-hydroxy-propyl trimethyl ammonium chloride (CHPTMAC). The products were charac-
terized by H NMR, FTIR and TGA, and investigated for antibacterial activity against Staphylococcus aureus
and Escherichia coli under weakly acidic (pH 5.5) and weakly basic (pH 7.2) conditions. TMHTMAPC
1
exhibited enhanced antibacterial activity compared with TMC, and the activity of TMHTMAPC increased
Keywords:
Chitosan
N,N,N-Trimethyl chitosan
Quaternary ammonium chitosan
Antibacterial activity
2+
2+
with an increase in the degree of substitution. Divalent cations (Ba and Ca ) strongly reduced the anti-
bacterial activity of chitosan, O-carboxymethyl chitosan and N,N,N-trimethyl-O-carboxymethyl chitosan,
but the repression on the antibacterial activity of TMC and TMHTMAPC was weaker. This indicates that
the free amino group on chitosan backbone is the main functional group interacting with divalent
+
cations. The existence of 100 mM Na slightly reduced the antibacterial activity of both chitosan and
its derivatives.
Ó 2011 Elsevier Ltd. All rights reserved.
1
. Introduction
that of chitosan. It increases with an increase in the chain length of
8
the alkyl substituent. A similar result against Escherichia coli
9
Quaternary ammonium chitosan, a typical kind of chitosan
derivatives, has attracted considerable attention recently as bacte-
(E. coli) has been reported by other researchers. N,N,N-Diethylm-
ethyl chitosan exhibits enhanced antibacterial activity against
E. coli in comparison with chitosan, and a decrease of pH value re-
1
2
3
riostatic agent, textile finishing agent, drug delivery carrier, and
so on. This is due to its enhanced water solubility and increased
number of positive charges compared to chitosan, which is a natu-
ral polysaccharide with many excellent properties, such as non-
toxicity, biodegradability, and biocompatibility.4
Generally, there are two common ways to carry out quaterniza-
tion of chitosan. One entails the introduction of a quaternary
ammonium moiety onto the chitosan backbone. N,N,N-Trimethyl
chitosan (TMC), the simplest form of quaternary ammonium chito-
san derivatives, is synthesized by reacting chitosan with methyl
1
0
sults in stronger activity. HTCC displays an increased antibacte-
rial efficiency against both E. coli and S. aureus relative to
1
1,12
13
chitosan.
On the contrary, Chi et al. found that HTCC had
14
no remarkable activity against E. coli. Qin et al. reported that
the antibacterial activity of HTCC was stronger under alkaline
conditions than under weak acidic conditions. These results point
to the possibility that quaternization does not always enhance
the antibacterial activity of chitosan and that the effect of pH on
the antibacterial activity of quaternary ammonium chitosan is
uncertain. When the antibacterial activities of methylated chitosan
derivatives containing aromatic moieties were evaluated, research-
ers found that N,N-dimethyl N-(4-N,N-dimethylaminobenzyl)
chitosan and N,N-dimethyl N-pyridylmethyl chitosan did not have
enhanced antibacterial activity against S. aureus compared with
TMC. However, N,N-dimethyl N-(4-N,N-dimethylaminobenzyl)
chitosan, with certain degree of quaternization, exhibited a slightly
5
iodide under alkaline conditions. The other evolves the introduc-
tion of a quaternary ammonium moiety on the outside of the chito-
san backbone. In this case, N-[(2-hydroxy-3-trimethyl ammonium)
propyl] chitosan (HTCC) is synthesized by the reaction of chitosan
with gycidyltrimethyl ammonium chloride (GTMAC) in water or
3
MAC) under alkaline conditions. Both methods could be used to
increase the antibacterial activity and water solubility of chitosan.
The antibacterial activities of TMC and N,N-dimethyl N-alkyl
chitosan against Staphylococcus aureus (S. aureus) are stronger than
6
-chloro-2-hydroxy-propyl trimethyl ammonium chloride (CHPT-
7
1
5
increased antibacterial activity against E. coli. After investigating
1
6
the activity of N-betainoyl chitosan,
O-methyl free N,N,N-
1
trimethyl chitosan, N-[1-carboxymethyl-2-(4-methylpiperazinyl)]
chitosan, N-[1-carboxymethyl-2-(4,4-dimethylpiperazinium)]
chitosan and N-[1-carboxymethyl-2-(1,4-dimethylpiperazini-
⇑
Corresponding author. Tel.: +86 595 2269 0819; fax: +86 595 22690917.
E-mail addresses: mcli@hqu.edu.cn, xutao@hqu.edu.cn (M. Li).
1
7
um)] chitosan, other researchers concluded that these groups
0
008-6215/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2011.08.002

2
446
T. Xu et al. / Carbohydrate Research 346 (2011) 2445–2450
O
(
CH ) N
3
3
Cl (CH ) N
Cl
3
3
Cl
HCl
OH
CHPTMAC
OH
O
OH
OCH2CH(OH)CH2N(CH3)3 Cl
O
O
HCHO
HCOOH
CH3I
NMP
NaCl
O
O
HO
O
HO
HO
N
NH2
NaOH
N
H3C
CH3
Cl
Cl
CH3
H3C
CH3
CH3
chitosan
TMC
Figure 1. Synthesis of chitosan quaternary ammonium derivatives.
TMHTMAPC
did not contribute to activity at low pH. However, the N,N,N-tri-
methyl and N-[1-carboxymethyl-2-(1,4,4-trimethylpiperazinium)]
groups contributed to activity at pH 7.2. In our previous report,
we also found that modification of the free amino group of chitosan
2.2.1. Synthesis of TMC, CMC, and TMCMC
TMC72H (degree of N-trimethylation = 68.9%, degree of N-
dimethylation = 26.7%, DD = 95.6%), O-carboxymehtyl chitosan,
(CMC20H, degree of O-carboxymethylation = 58.8%), and N,N,N-tri-
methyl O-carboxymethyl chitosan (TMCMC20H, synthesized from
TMC72H, degree of O-carboxymethylation = 61.5%) were synthe-
18
backbone decreased its antibacterial activity.
+
At a concentration of 100 mM, Na ions reduce the antibacterial
activity of chitosan and divalent cations exhibited strong repres-
1
8
sized and characterized as our previous report.
2
+
2+
2+ 19
sion in the order of: Ba > Ca > Mg
.
At lower concentrations
+
20
(
10 or 25 mM), Na ions do not show detectable repression. How-
ever, Chung et al. reported that a high ionic strength (Na ) in-
creased the antibacterial activity of chitosan because of its
2.2.2. Synthesis of CHPTMAC
2
1
+
To synthesize CHPTMAC, 0.25 mol trimethylammonium aque-
ous solution was added to a three-neck flask and the solution
was chilled to 4 °C in an ice/water bath. 22 mL concentrated hydro-
chloric acid (37.5 wt %) and 8 mL anhydrous ethanol were added to
the cold solution. The pH of the solution was adjusted to 8.0 by
trimethylammonium, and then the temperature was raised to
10 °C, followed by adding 20 mL epichlorohydrin (0.24 mol) drop-
wise within half an hour. The two-phase solution was stirred for
1 h at 10 °C and another 1 h at 35 °C. Finally, the colorless transpar-
ent solution was vacuum distilled at 50 °C to dewater and remove
the unreacted epichlorohydrin, followed by washing with acetone
several times to obtain the white acicular crystal. The melting
point of CHPTMAC was 190–191 °C. The content of CHPTMAC
was 91%, determined by a potential titration method using silver
13
2+
enhanced solubility. Chi et al. found that divalent cations (Ca
2
+
and Mg ) had no remarkable impact on the antibacterial activity
of HTCC. Thus, the effect of the cations on the antibacterial activity
requires further investigation.
The commonly used method for the synthesis of TMC is always
accompanied with O-methylation, as Sieval et al. has previously
5
shown. This reduces the solubility of TMC and limited further O-
modification. Recently, a promising method for TMC synthesis that
2
2
avoids O-methylation and chain scission has been reported. In
this light, a N,O-di-quaternary ammonium chitosan, N,N,N-tri-
methyl O-(2-hydroxy-3-trimethylammonium propyl) chitosan
(
TMHTMAPC), was synthesized by reacting O-methyl-free TMC
2
3
with CHPTMAC (Fig. 1). The products were characterized by proton
nitrate.
1
nuclear magnetic resonance ( H NMR) spectroscopy, Fourier trans-
form infrared (FTIR) spectroscopy and thermogravimetric analysis
2.2.3. Synthesis of TMHTMAPC
(
TGA). This study aimed to investigate the antibacterial activity of
The synthesis of TMHTMAPC was similar to the synthesis of cat-
2
4
the synthesized compounds against a gram-positive bacterium, S.
aureus, and a gram-negative bacterium, E. coli, at pH 5.5 and pH
7
ionic amylopectin starch. Briefly, 1 g TMC72H was dissolved in
20 mL deionized water, followed by adjusting the pH value to 11
with 1 mM NaOH. The mixture was stirred for 1 h under nitrogen
protection. CHPTMAC (4 meq/GlcN) was added dropwise within
.2. The effect of the cations on the antibacterial activity of quater-
nary ammonium chitosan derivatives was also evaluated.
1
h, and then reacted for 5, 10, and 20 h, respectively. At these
points, 1 mM HCl was added to adjust the final pH to 7. The result-
ing solution was dialyzed against deionized water for three days by
changing buffer twice daily, and then lyophilized to obtain TMHT-
MAPC. The degrees of substitution (DS) for O-quaternization were
calculated from H NMR spectra according to the following
equation:
2
2
. Materials and methods
.1. Materials
1
Chitosan (Mw = 100 k Da, DD = 95.6% according to the manufac-
tory) was purchased from Zhejiang Aoxing Biotechnology Co., Ltd
China) and refined by dissolving in dilute acetic acid aqueous
(
O-quaternization% ¼ IHg=IHb ꢀ DSN-trimethylation
ð1Þ
water and then precipitated by adding NaOH aqueous solution fol-
lowed by filtration. Other reagents were commercially available
and used without further purification unless otherwise indicated.
.
The DS for O-quaternization increased to 22.4%, 33.1%, and
2.7%, when the reaction prolonged to 5, 10, and 20 h, respectively.
4
S. aureus (ATCC 6538) and E. coli (ATCC DH5a) obtained from
American Type Culture Collection (ATCC) were used for antibacte-
rial activity test.
2
.3. Characterization
IR spectra were recorded with KBr pellets on a Nicolet NEXUS-
70 spectrophotometer. H NMR spectra were measured at 313 K
2
.2. Synthesis of chitosan derivatives
1
4
on a INOVA-600 spectrometer. For chitosan, D
2
O/5% DCl was used
All the following samples are identified by the reaction time of
the last step, in which ‘H’ indicates hours.
as solvent, while for other products, D O was used as solvent.
2

T. Xu et al. / Carbohydrate Research 346 (2011) 2445–2450
2447
Thermogravimetric analysis was implemented using Universal
V2.4F TA thermal analyzer. The analysis was performed under a
continuous flow of dry nitrogen gas at a heating rate of 20 °C/min.
IHc/IH1 for TMC and TMHTMAPC), suggesting that CHPTMAC does
not react with dimethylamino group.
The FTIR spectra (Fig. 3) provide additional evidence for the suc-
cessful synthesis of the products. The FTIR spectra of chitosan and
its quaternary ammonium derivatives show peaks attributed to the
2
2
.4. Antibacterial test
ꢁ1
saccharide structure at 898 and 1154 cm , and a strong amino
ꢁ1
.4.1. Determination of minimum inhibitory concentration
characteristic peak at around 1614 cm . The absorption bands at
ꢁ1
The minimum inhibitory concentration (MIC) of chitosan and
1650 and 1320 cm are attributed to the amide I and III bands,
respectively. Compared with chitosan, the new bands at
its quaternary ammonium derivatives was determined via an agar
2
5
ꢁ1
plate method, with some modifications. In this method, samples
were prepared at a concentration of 1% (w/v), and then autoclaved
at 121 °C for 20 min. Duplicate two-fold serial dilutions of each
sample were added to a nutrient broth (at pH 5.5 or pH 7.2) to ob-
tain sample concentrations of 0.1, 0.05, 0.025, 0.0125, 0.00625, and
1490 cm in TMC and TMHTMAPC correspond to the asymmetri-
cal stretching of C–H in the methyl groups, which are characteristic
of the highly methylated chitosan quaternary salt. The N–H bend-
ꢁ1
ing (1614 cm ) of the primary amine disappears due to the
1
,6
change of the primary amine to the quaternary amine.
For
ꢁ1
0
.00313, 0.00157 and 0.00079% (w/v). The pH was adjusted with
TMHTMAPC, the peaks at 1050 cm
corresponding to C–O–C
as internal reference) in
ꢁ1
either 1% acetic acid or 1% NaOH. The culture of each bacterium
stretching increased (using 1650 cm
5
6
was diluted with sterile distilled water to 10 –10 CFU/mL. A loop
of each suspension was inoculated on a nutrient medium with
either a sample or control added, and then incubated at 35 °C for
comparison with TMC, indicating the successful O-quaternization
via ether bond.
4
8 h. Finally, the colonies were counted to obtain MIC value. The
3.2. Thermal stability of quaternized ammonium chitosan
derivatives
experiments were repeated trice. The MIC is defined as the lowest
concentration of chitosan or its derivates required to inhibit the
growth of bacteria. That is, the concentration at which no
microorganism colony or less than five colonies are visible within
The TGA thermograms of chitosan and its derivatives are pre-
sented in Figure 4. Generally, there are two distinct stages in the
degradation of chitosan and its derivatives. The first one starts at
about 80 °C with a weight loss of 5–12% due to the loss of adsorbed
1
9–38 h.
7
2
.4.2. Evaluation of the effect of cations on the antibacterial
and bound water. In contrast, the quaternary ammonium deriva-
activity
Chitosan or its derivative was added to an E. coli or S. aureus sus-
pension (10 mL, 10 CFU/mL), containing 25 mM BaCl , CaCl , NaCl
2 2
or 100 mM NaCl. After adjusting the pH to 5.5 by 0.1 mM HCl, the
mixture was incubated in ambient air at 35 °C. Control experi-
ments (without the addition of chitosan or its derivative) were also
tives exhibit more weight loss at this stage because quaternization
enhances their hydrophilicity. The second stage is registered at
nearly 260 °C for chitosan, 220 °C for TMC72H, and 200 °C for
TMHTMAPC20H. This is due to the decomposition of the chitosan
backbone chain and cleavage of substituent groups. The thermal
stability is poorer after further quaternization, and the decrease
follows this order: chitosan > TMC72H > TMHTMAPC20H. The de-
crease of the crystallinity due to random N-substitution and O-
substitution is believed to be the cause of this thermal instability.
7
2
6
conducted. After incubation for 1 h, a 50 lL aliquot or the dilutions
were spread on nutrient agar plates, which were incubated at 35 °C
for 24 h. Then the numbers of colonies were counted and the inhi-
bition rate was calculated which is defined as:
3
.3. Antibacterial activity of quaternized ammonium chitosan
Inhibition rateð%Þ
derivatives
¼
ðinitial cell number ꢁ cell number after treatmentÞ
According to the literatures,27,28 trimethylation of chitosan and
=
initial cell number ꢀ 100%
ð2Þ
quaternization of chitosan by GTMAC can enhance its antibacterial
activity against a number of gram-negative and gram-positive bac-
teria. In the present study, the antibacterial activities of the quater-
nary ammonium chitosan derivatives were compared with that of
chitosan. Table 1 shows MIC of chitosan and its derivatives with
various DS at pH 5.5 and 7.2 against S. aureus and E. coli. The qua-
ternary ammonium derivatives exhibit antibacterial behavior, and
all the derivatives of chitosan have stronger antibacterial activity
than the original chitosan at pH 5.5. After further O-quaternization,
TMHTMAPC exhibit stronger antibacterial activity than TMC, and
the activity of TMHTMAPC increases with an increase of DS at both
All the experiments were carried out in triplicate and average
values were reported.
3
3
. Result and discussion
.1. Synthesis and characterization of quaternized ammonium
chitosan derivatives
1
The H NMR spectra (Fig. 2) shows the successful synthesis of
CHPTMAC, TMC and TMHTMAPC. Compared with chitosan
þ
(
Fig. 2b), the new peak at 3.25 ppm is the signal for ꢁNðCH Þ,
pH 5.5 and 7.2. However, some researchers found that quaterniza-
3
tion of chitosan decreased its antibacterial activity.1
,16,17
This
the characteristic peak of TMC (Fig. 2c). The peak at 2.4 ppm is as-
signed to the hydrogen protons of –N(CH
dimethylation. In comparison with TMC, the new peaks at
.15 ppm and 3.4 ppm for TMHTMAPC (Fig. 2d) are attributed to
3
)
2
, indicating partial N-
maybe depends on the kinds of substituents. In the current study,
further O-modification of TMC by CHPTMAPC enhances its antibac-
terial activity, indicating 2-hydroxy-3-trialkylammonium propyl
groups are efficient to enhance its activity. Maybe modifications
with flexible side chains out of chitosan backbone to carry out
quaternization are more suitable to enhance the interaction be-
tween positive-charged chitosan derivatives and negative-charged
cell envelope.
3
the methyl hydrogen protons and methylene hydrogen protons
þ
of ꢁCH
2
NðCH Þ , respectively, indicating successful O-quaterniza-
3
3
tion. Other signals for O-quaternization overlap with those for
chitosan, giving difficulties for identification. Moreover, partial 3-
O-quaternization and glycedyl-O-quaternization may occur, but
with lower DS than 6-O-quaternization due to steric hindrance,
These results also reveal that the antibacterial activity of both
chitosan and its quaternary ammonium derivatives are stronger
under weakly acidic conditions than under weakly basic condi-
1
since no obvious characteristic peaks are observed in H NMR spec-
tra. Additionally, the DS of dimethylation remains the same during
the reaction of O-quaternization (indicated by the same value of
1
,28
tions, which coincide well with published reports.
Furthermore,

2
448
T. Xu et al. / Carbohydrate Research 346 (2011) 2445–2450
Figure 2. 1H NMR spectra of (a) CHPTMAC, (b) chitosan, (c) TMC72H, and (d) TMHTMAPC20H. Chitosan was dissolved 2% DCl/D
O, while others were dissolved in D O.
2 2
Figure 4. TGA thermograms of (a) chitosan, (b) TMC72H, and (c) TMHTMAPC20H.
Figure 3. IR spectra of (a) chitosan, (b) TMC72H, and (c) TMHTMAPC20H.
both chitosan and its quaternary ammonium derivatives exhibit
stronger antibacterial activity against S. aureus than against
E. coli. This can be attributed to the different cell envelopes of S.
aureus and E. coli. The cell envelope of a gram-positive bacterium
thin layer of peptide polyglycogen and an outer lipopolysaccharide
(LPS) layer. The outer LPS layer is a potential barrier against foreign
molecules with high molecular weights. Hence, chitosan and its
quaternary ammonium derivatives have more difficulties penetrat-
ing the outer membrane of E. coli. However, although S. aureus
and E. coli are used as representative bacteria to evaluate the
1
3
(S. aureus) is fully composed of peptide polyglycogen. In contrast,
a gram-negative bacterium (E. coli) cell envelope is made up of a

T. Xu et al. / Carbohydrate Research 346 (2011) 2445–2450
2449
lent cations (Ba2+ and Ca2+) significantly decreases the antibacte-
Table 1
MIC of quaternary ammonium chitosan derivatives against E. coli and S. aureus at pH
rial activity of chitosan, which is in good agreement with
5
.5 and pH 7.2
19,21
previous reports.
The decrease is caused by the chelating of
Material
E. coli
S. aureus
pH 7.2
metal ions with chitosan, where the amino group acts as the
2
9
pH 5.5
pH 7.2
pH 5.5
main chelating site. Since the free amino group is also the anti-
3
0
bacterial functional group, the chelation results in a decrease of
the antibacterial activity of chitosan. However, the antibacterial
activity of the quaternary ammonium chitosan derivatives shows
less decrease with the addition of divalent cations relative to that
of chitosan, indicating that the quaternary ammonium groups of
TMC and TMHTMAPC have weaker interactions with metal ions
than the free amino group. Compared with chitosan, the antibac-
terial activity of CMC20H and TMCMC20H shows more decrease
with the addition of divalent actions, indicating carboxymethyla-
tion contributes to its adsorption ability of metal ions. This is in
agreement with published reports on the metal ions adsorption
TMC72H
0.025
0.1
0.05
0.025
0.00625
0.0125
0.05
0.025
0.00625
0.00313
TMHTMAPC5H
TMHTMAPC10H
TMHTMAPC20H
0.0125
0.00625
0.00313
0.00625
0.00313
0.00157
antibacterial activity of chitosan and its derivatives, these results
with only one type and one strain of each type of bacteria are
not sufficient to conclude that chitosan and its derivatives have a
stronger activity against G bacteria than G bacteria. Further
investigation against more strains of bacteria of both G and G
+
ꢁ
+
ꢁ
31
2+
capacity of carboxymethylated chitosan. Moreover, Ba has
stronger repressive effects on the antibacterial activity of chito-
is required.
2
+
19
san than Ca , which is in accordance with a previous report.
3
.4. Effect of cations on the antibacterial activity of chitosan
The same trend was observed for the quaternary ammonium
chitosan derivatives.
and its quaternary ammonium derivatives
Chung et al. reported that the antibacterial activity of chitosan
According to previous reports,19,21 cations in the dispersion
can affect the antibacterial activity of chitosan. In the present
study, Ba (25 mM), Ca (25 mM) and Na (25 and 100 mM)
were used to investigate the effect of cations on the antibacterial
activity against E. coli and S. aureus (Fig. 5). The addition of diva-
+
increased with an increase in the ionic strength (Na ) of the solu-
2
1
tion. In the present study, however, we found that the existence
2
+
2+
+
+
of 100 mM Na caused a slightly repressive effect on the antibacte-
+
rial activity of chitosan, although the addition of 25 mM Na exhib-
ited no remarkable impact. This coincides with a previous report.19
Figure 5. The inhibition rate with 25 mM Ca2+ and 25 mM Ba2+ ions added against (a) E. coli and (b) S. aureus, with 25 mM and 100 mM Na+ added against (c) E. coli and (d) S.
aureus, at pH 5.5 after incubation for 1 h. The values represent means ± SD (n = 3).

2
450
T. Xu et al. / Carbohydrate Research 346 (2011) 2445–2450
The existence of 100 mM Na+ also shows a slight repression of
the antibacterial activity of quaternary ammonium chitosan
derivatives. As proposed by Tsai and Su,19 Na and chitosan, at high
concentration, form a complex which reduces the binding force be-
Acknowledgments
+
This research is supported by the National Natural Science
Foundation of China (No. 20574024), the Provincial Natural Science
Foundation of Fujian (E0810019, 2009J01029, and 2011J01312),
and the Key Projection of Science and Technology of Fujian Prov-
ince (2009H0030).
+
tween chitosan and the cell surface. In the same way, Na
(
100 mM) and quaternary ammonium chitosan derivatives in the
current study may have formed a complex. Moreover, the addition
of salt decreases the viscosity of the polymer solution due to the
reduction of the electrostatic repulsion by the positively charged
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2,33
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2
0
respectively. Moreover, the quaternization of chitosan increases
the hydrophilicity of the chitosan derivatives, which leads to the
difficulty of obtaining anhydrous materials. This can be proven by
the first stage of thermal degradation (Fig. 4). Therefore, it is dif-
ficult to compare the current results with other quaternary
ammonium chitosan derivatives precisely. Only the relative trend
of the antibacterial activity of chitosan and its derivatives can be
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4
. Conclusion
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ity was also investigated. All the quaternary ammonium chitosan
derivatives showed stronger antibacterial activity than chitosan.
TMHTMAPC exhibited enhanced antibacterial activity compared
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the repression was weaker in TMC, and TMHTMAPC. This indi-
cated that the quaternary ammonium groups of the quaternary
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+
2+
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metal ions than with the free amino group. The existence of
+
1
00 mM Na caused a slight repression of the antibacterial activ-
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+
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because Na and the chitosan derivatives formed a complex that
3
reduced the binding force between the materials and the cell
surface.
2