Food Hydrocolloids 33 (2013) 186e191

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

Preparation and structural analysis of chitosan films with and

without sorbitol
Mei Liu, Yibin Zhou*, Yang Zhang, Chen Yu, Shengnan Cao
School of Tea and Food Technology, Anhui Agricultural University, 130 Chang Jiang West Road, Hefei 230036, China

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

Article history: The purpose of this study was to clarify the structural changes in chitosan films made with sorbitol. The
Received 24 January 2013 degree of deacetylation (DD) in the starting chitosan was 85% or 95%. The appearance and cross-sectional
Accepted 14 March 2013 characteristics of the films were analyzed by SEM. This showed sorbitol to be a good crosslinking agent,
generating the desired miscibility with both types of chitosan. XRD revealed an increase in crystallinity
Keywords: for films without sorbitol and a decrease for films spiked with sorbitol compared with the original
Chitosan film
chitosan. FT-IR showed that the addition of sorbitol did not change the chemical structure of chitosan and
Sorbitol
no new types of bonds were created during film formation. Changes in the chemical surrounding of
Structural analysis
carbon atoms analyzed by CP/MAS 13C NMR indicated that the acetic acid solution used partially
destroyed the chitosan powder and protonized some groups, enabling sorbitol to be inserted and to form
hydrogen bonds in the matrix.
Ó 2013 Elsevier Ltd. All rights reserved.

1. Introduction Up to now, most research has focused on the physicochemical

Environmentally friendly bio-based polymeric materials are
becoming increasingly popular due to their biocompatibility,
biodegradability, non-toxicity, antimicrobial activity and other
properties. These materials are usually made from biopolymers
such as polysaccharides, proteins, lipids and resins. Chitosan is
obtained by partial N-deacetylation of chitin (Ham, 1998) which is
the second most abundant natural polysaccharide after cellulose,
and is a high molecular weight polymer formed by b-(1-4)-2-
amino-2-deoxy-D-glucopyranose units. Chitosan has been widely
used in agriculture, biomedicine (Jayakumar, Menon, Manzoor,
Nair, & Tamura, 2010) and food (Agulló, Rodríguez, Ramos, &
Albertengo, 2003).
Studies have reported that films formed of pure chitosan can be
brittle and fragile, while films which incorporate plasticizers such
as glycerol, sorbitol (Thirathumthavorn & Charoenrein, 2007) or
PEG (Suyatma, Tighzert, & Copinet, 2005) have better physical
properties (Epure, Griffon, Pollet, & Avérous, 2011). Fakhoury et al.
(2012) reported that chitosan films formed with sorbitol possessed
better physicochemical properties than those containing glycerol.
Rotta et al. (2009) found chitosan films made with sorbitol to have
good physical properties such as transparency, water solubility,
wettability, and surface and thermal stability.
* Corresponding author. Tel./fax: þ86 551 5786342.
E-mail address: zhouyibin@ahau.edu.cn (Y. Zhou).
properties and applications of chitosan films, with few attempts to
investigate any potential structural differences between those
formed with and without sorbitol as the plasticizer. Examining the
structural characteristics of chitosan films can elucidate the cross-
linking relationship between chitosan and plasticizers such as
sorbitol. The purpose of this work was to analyze the structural
changes in different degrees of deacetylation chitosan transformed
from its powder form into films, with and without the addition of
sorbitol, and to provide basic data for the development of appli-
cations in the packaging, food processing and healthcare industries.
2. Materials and methods
2.1. Materials
Chitosan with two different degrees of deacetylation (DD), 85%
(MW ¼ 343.75 kDa) and 95% (MW ¼ 312.5 kDa), were purchased
from Zhejiang Golden-shell Biochemical Co. Ltd (China). Sorbitol
was purchased from Sigma (USA). All of the other chemicals used
were of analytical grade and were commercially available.
2.2. Preparation of chitosan films
Chitosan films were prepared using the film casting method with
some modifications (Epure et al., 2011). Samples of chitosan with
different degrees of deacetylation (2% wt/v), half of which were
spiked with sorbitol, were dissolved in 2% (v/v) acetic acid solution

0268-005X/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.foodhyd.2013.03.003
 M. Liu et al. / Food Hydrocolloids 33 (2013) 186e191
(100 ml). For the chitosan samples with sorbitol, sorbitol was first
incorporated into the chitosan powder by manual mixing. The acetic
acid solution (2% v/v) was then added to the mixture. For the
unplasticized formulation, acetic acid solution (2% v/v) was added
directly to the chitosan powder. The mixtures were then stirred with
a magnetic stirrer at 60  C for 2 h, degassed under vacuum and then
poured into a glass pane (30  20  0.5 cm). The solution in the glass
pane was dried in an oven set to 60  C for 6 h and then further dried
at ambient temperature for 2 days, at about 57% relative humidity
(RH) and 25  C. The dried films were immersed in 2% (wt/v) sodium
hydroxide for half an hour to neutralize any residual acetic acid in
the films. These films were then washed thoroughly with distilled
water until the water ran neutral, and the prepared films were
stored at room temperature prior to testing.
2.3. Measurement of chitosan powder and films
2.3.1. Film microstructure examination by SEM
Micrographs of the samples were examined using a scanning
electron microscope (SEM, Sirion 200, Netherlands FEI) with an
accelerating voltage of 5 kV. Samples were coated with a gold layer
prior to observation and then observed at different magnifications.
Surface images were taken at a magnification of 1000 and 10,000,
and the cross-section topography at magnifications of 10,000 and
20,000.
2.3.2. Determination of the degree of crystallinity
X-ray diffraction (XRD) patterns were collected using a Philips
X’Pert PRO X-ray Diffractometer equipped with a Cu K-a radiation
source, operating at 40 kV and 200 mA. The relative intensity was
recorded in the scattering range of 3e50 at a scanning rate of 8 /
min in steps of 0.02 per second in continuous scanning mode. The
degree of crystallinity in the samples was calculated using the
follow equation.
Xc ¼ ½Fc=ðFc þ FbÞ  100%: (1)
where Fc and Fb represent the areas of crystalline and
non-crystalline regions, respectively (Tian, Liu, Hu, & Zhao, 2004).
2.3.3. Fourier transformed infrared (FT-IR) spectroscopy
Fourier transformed infrared spectroscopic examination of the
powder and films with and without sorbitol were recorded with a
Bruker infrared spectrometer (Vertex 80 and Hyperion 2000).
Characteristic absorption bands were used to analyze both the
powder and the films. Powder samples (around 1 mg) were
blended with 100 mg KBr, then ground thoroughly under a pres-
sure of 1.01  107 Pa. Films were placed on the steel plate and
measured directly in the wave number range 4000e400 cm1,
using 16 scans at a resolution of 0.02 cm1.
13
2.4. 4 CP/MAS C NMR
Solid-state 13C NMR spectra of the samples were obtained on a
Bruker Avance AV 400 MHz spectrometer. The spinning rate of the
rotor was 15 kHz. Spectra were measured with cross polarization
and magic angle spinning (CP/MAS) using a 4-mm probe at a
temperature of 20  C.
3. Results and discussion
3.1. Determination of film microstructure by SEM
SEM micrographs of the surface and cross-section of 85%
and 95% DD chitosan films containing sorbitol are presented in
187
Fig. 1AeH. The micrographs showed that the surface of both types
of sorbitol-spiked chitosan films was homogenous, continuous and
smooth. Thus, sorbitol offers good miscibility and compatibility
when incorporated into chitosan. A previous study has shown the
surface of pure chitosan films to be smooth, continuous and
compact (Pinotti, Garcia, Martino, & Zaritzky, 2007). Here, the
addition of sorbitol did not introduce discontinuities or porous
structures into the film. The cross-sectional micrographs of both
85% and 95% DD chitosan films made with sorbitol revealed them to
be continuous and compact, similar to those containing glycerol
examined in a previous study (Cervera et al., 2004).
3.2. Crystal transformation by XRD
XRD patterns and degree of crystallinity of the 85% and 95% DD
chitosan powder and films, with and without sorbitol, are shown in
Fig. 2 and Table 1. Both the 85% and 95% DD powders showed a
strong diffraction at 2q ¼ 20.5 and a weak reflection at 2q ¼ 10.5 .
This is in good agreement with the diffraction pattern of chitosan
reported by Salmon and Hudson (1997). Both unspiked films also
displayed two peaks at 20.5 and 10.5 , respectively. The intensity
of diffraction of the 85% and 95% DD chitosan unspiked films was
largely inferior to that of their corresponding chitosan powder,
while the degree of crystallinity of the films was respectively higher
than those of their powder (Table 1). During the process of
unplasticized film formation, the dispersed chitosan macromole-
cule in the solvent may be prone to forming more irregular crystals
(Kittur, Kumar, & Tharanathan, 2003).
The two spiked chitosan films also generated two peaks at the
same diffractive angles as those of the unspiked powders and their
pure films, but while the intensity of reflection increased remark-
ably at 10.5 and reduced markedly at 20.5 compared with original
powder. According to Fan, Hu, and Shen (2009), this result suggests
that the amorphous region of the film increased and a crystal
structure was partially formed with more extended chains during
the process of film formation. As shown in Table 1, the degree of
crystallinity of sorbitol-spiked films was lower than that in the
unspiked powders and films. This decrease may result from the
structural changes caused by partial chitosan destruction and the
variation in inter- and intra-molecular hydrogen bonds.
Additionally, the degree of crystallinity increased with
increasing deacetylation for both the powders and the films
(Table 1), in line with the results obtained by Chen, Lin, and Yang
(1994). This result may be explained by the fact that the chitosan
with the higher DD was more compact, resulting in a matrix more
favorable to hydrogen bonding formation which in turn would in-
crease the crystallinity (Ziani, Oses, Coma, & Maté, 2008).
3.3. Fourier transform infrared (FT-IR) spectroscopy
Fourier transform infrared (FT-IR) spectroscopy reveals infor-
mation about the molecular interactions of chemical components
and is useful to supplement microstructural characterization of
composite films. In order to investigate the structural difference
between 85% and 95% DD chitosan powders and films with and
without sorbitol, IR spectra were recorded.
Fig. 3A and B shows that the chitosan powder (CH), unspiked
chitosan film (CHF) and spiked film (CHFP) displayed similar ab-
sorption bands for both types of chitosan powder (85% and 95% DD)
but with variations in peak intensity, which was probably related to
the content of the chitosan. Increasing the degree of deacetylation
resulted in a decrease in the intensity of absorption at 1653 cm1
and 1322 cm1 and an increase at 1589 cm1 and 1594 cm1 for
powders and films, which can be attributed to the acetyl groups
(Ziani et al., 2008).
188 M. Liu et al. / Food Hydrocolloids 33 (2013) 186e191

Fig. 1. SEM micrographs of surface and cross section of films made with chitosan with different degrees of deacetylation (DD) spiked with sorbitol. A) Surface (1000) of 85% DD; B)
surface (10,000) of 85% DD; C) surface (1000) of 95% DD; D) surface (10,000) of 95% DD; E) cross section (10,000) of 85% DD; F) cross section (20,000) of 85% DD; G) cross
section (10,000) of 95% DD; H) cross section (20,000) of 85% DD.

Characteristic peaks of the CH, CHF and CHFP are summarized in
Table 2. A broad absorption band at 3440 cm1 resulted from the
overlapping of hydroxyl group and amino group stretching vibra-
tions. Bands at 2915 cm1 and 2875 cm1 represented CH2 sym-
metric and anti-symmetric stretching vibrations, respectively. The
band at 1653 cm1 could be attributed to the amide I (C]O)
stretching vibration, while the amide III band (CeN) was evident at
1322 cm1 for both powders and films. For amide II (NeH), the
absorption of the powder was evidenced by a broad peak at
1594 cm1 while CHF and CHFP generated a sharp peak at a lower
wave number (1589 cm1) (Pawlak & Mucha, 2003; Rivero, García,
& Pinotti, 2010; Xu, Kim, Hanna, & Nag, 2005). The weak peak at
1154 cm1 was assigned to the CeOeC symmetric stretching vi-
bration, the relatively broad peak at 1082 cm1 was assumed to be
 M. Liu et al. / Food Hydrocolloids 33 (2013) 186e191 189

A 100

80 CH
a

b
Intensity

60
CHFP
c

T/%
d 40
CHF
e
20

0
0 10 20 30 40 50 4000 3500 3000 2500 2000 1500 1000 500
2 theta (deg) -1
wavenumber/cm
Fig. 2. XRD of chitosan powder and films with different degrees of deacetylation (DD): B
(a) DD 85% CH; (b) DD 95% CH; (c) DD 85% CHF; (d) DD 95% CHF; (e) DD 85% CHFP; (f) 100
DD 95% CHFP. CH, CHF and CHFP represent chitosan powder, and chitosan films
without and with sorbitol, respectively. CH
80

the CeO stretching vibration in the C3eOH group, and the weak CHFP
peak at 1033 cm1 was assigned to the CeO stretching vibration of 60
C6eOH group (Feng, Liu, Zhao, & Hu, 2012; Rivero et al., 2010). The
assignment of these absorption bands was in agreement with those
T/%

CHF
reported previously for chitosan powder and films (Abugoch, Tapia,
40
Villamán, Yazdani-Pedram, & Díaz-Dosque, 2011; Rivero et al.,
2010).
The corresponding CH, CHF and CHFP showed the same char-
20
acteristic peak positions for both types of (DD) powder tested,
respectively (Table 2). This indicated the powders and the films had
the same group absorption frequency for different degrees of
0
deacetylation. Some groups of the spiked and unspiked films 4000 3500 3000 2500 2000 1500 1000 500
exhibited red shift and blue shift compared with the powder. The
peak at 2875 cm1 blue-shifted and those located at 1594 cm1,
-1
wavenumber/cm
1421 cm1, 1381 cm1, 1263 cm1 and 1154 cm1 red-shifted. These
Fig. 3. IR spectra of powder and films made with chitosan with different degrees of
shifts may have been caused by different interactions in the matrix, deacetylation (DD): (A) DD 85%, (B) DD 95%. CH, CHF and CHFP represent chitosan
such as inter-molecular rearrangement and variations in the powder, and chitosan films without and with sorbitol, respectively.
configuration of the main chain (Gu & Wu, 2007; Piermaria et al.,
2011). The infrared spectra curves exhibited by CHFP changed MAS 13C NMR spectra of the 85% and 95% DD powders and spiked/
only in intensity compared with those of CHF. Thus, the addition of unspiked films are shown in Fig. 4. The assignment of the chemical
sorbitol appears not to have changed the chemical structure of shift of carbon atoms on the main chain is summarized in Table 3,
chitosan, nor were new types of bonds created during film for- and is in agreement with previous reports (Fan et al., 2009; Feng
mation (Martínez-Camacho et al., 2010; Ziani et al., 2008). A pre-
vious study also attributed the differences in the spectra of chitosan Table 2
powder and films to film-formation processes and an increase in Assignments of FT-IR characteristic absorption peaks of chitosan with different de-
grees of deacetylation (DD) in powder form (CH), and in films with (CHF) and
the order of the network structure (Kurek et al., 2012).
without (CHFP) sorbitol.

13 Characteristic peak (cm1) Samples

3.4. CP/MAS C NMR study
DD85% DD95%
Solid-state NMR spectroscopy is an effective and powerful tool CH CHF CHFP CH CHF CHFP
for the characterization of mixed materials and has been widely eOH, eNH2 3440 3440 3440 3440 3440 3440
used to investigate compound structural conformation changes. CP/ CH2 symmetric stretching 2915 2915 2915 2915 2915 2915
CH2 anti-symmetric stretching 2875 2880 2880 2875 2880 2880
C]O (amide I band) 1652 1652 1652 1652 1652 1652
NeH (amide II band) 1594 1589 1589 1594 1589 1589
Table 1
CH2 bending 1421 1417 1417 1421 1417 1417
Degree of crystallinity (DC) in chitosan with different degrees of deacetylation (DD)
CH bending and CH3 deformation 1381 1377 1377 1381 1377 1377
in powder form (CH), and in films with (CHF) and without (CHFP) sorbitol.
CeN (amide III) 1322 1322 1322 1322 1322 1322
Samples DD85% DD95% OeH bending 1263 1258 1258 1263 1258 1258
CeOeC symmetric stretching 1154 1150 1150 1154 1150 1150
CH CHF CHFP CH CHF CHFP CeO of C3eOH 1082 1082 1082 1082 1082 1082
DC (%) 34.3 37.8 30.8 40.8 44.6 34.3 CeO of C6eOH 1033 1033 1033 1033 1033 1033
190 M. Liu et al. / Food Hydrocolloids 33 (2013) 186e191

C3,5 configurations have different 13C NMR spectra. The b configuration

typically has a chemical shift at around 97 ppm, so the carbon atom
peaks measured here at 97.32 ppm and 97.33 ppm were assumed to
A C6 C 2 indicate that the chitosan in our study was in b configuration form.
C1 C4
The C1 and C4 chemical shifts of CHF changed more significantly
CH C=O C1 ' CH 3 than those of the other carbon atoms for both the 85% and 95% DD
chitosan tested compared with CH (powder form). This finding can be
attributed to the IR measurement being highly sensitive to the
CHF conformational change in b-1, 4-linked carbohydrates (Heux,
Brugnerotto, Desbrieres, Versali, & Rinaudo, 2000; Saito, Tabeta, &
Ogawa, 1987). The CH3 and C]O junctions of the 95% DD CHF shif-
CHFP ted higher field than those of the 85% DD CHF, which is probably
related to the higher degree of deacetylation (Table 3). C1, C3, C4, C5 and
C6 atoms showed similar chemical shifts for both forms of DD CHF.
C1, C4 and CH3 exhibited high-field shifts while C2, C3, C5 and C6
200 180 160 140 120 100 80 60 40 20 0 (ppm) showed low-field shifts for both forms of DD CHF tested, compared
with the original powder. These shifts have been described previ-
C3,5
B ously as the changes in electron intensities around C1 and C4 caused
C1 C6 C by the breaking of b-1, 4 glycosidic bonds (Gu & Wu, 2007). The
C4 2
C=O CH3 acetic acid solution used here is likely to have reacted with the
CH C1 '
amino group of the chitosan, causing CH3 to shift high-field.
Conversely, the intra- and inter-molecular hydrogen bond forma-
tions around C2, C3, C5 and C6 would lead them to shift low-field.
CHF In CHFP, the chemical shift of the carbon atoms changed to a
certain extent compared with the corresponding CHF. C2 and C]O
showed high-field shifts, likely due to the sorbitol hydroxyl and
CHFP chitosan amino groups forming hydrogen bonds. During the pro-
cess of film formation, the b-1, 4 glycosidic bond in the chitosan
would have been partly ruptured by the acetic acid solution,
resulting in the hydroxyl groups of the sorbitol being attracted by
200 180 160 140 120 100 80 60 40 20 0 (ppm) the exposed carbonium ion and leading to C1 and C4 migration to
Fig. 4. 13C NMR spectra of powder and films made with chitosan with different degrees
low-field (Epure et al., 2011). The low-field shifts of C3 and C5 were
of deacetylation (DD): (A) DD 85%, (B) DD 95%. CH, CHF and CHFP represent chitosan probably related to the hydrogen bonds between the hydroxyl
powder, and chitosan films without and with sorbitol, respectively. groups on C3 and C5, sorbitol and the amino group on chitosan.
Moreover, as shown in Table 3, when sorbitol was added the
chemical shifts of the carbon atoms altered more significantly than
et al., 2012; Rinaudo, Desbrieres, Le Dung, Thuy Binh, & Dong, for the unspiked film. For instance, C1, C3 and C5 showed more
2001). marked low-field shifts. This finding demonstrated that the sorbi-
The chitosan powders and films had similar 13C NMR spectra for tol, as a plasticizer, participated in the formation of the structure
the same DD, though with variations in signal intensity and slightly and changes in the surrounding environments of the carbon atoms.
different chemical shifts (Fig. 4 and Table 3). The signal intensity of The result of CP/MAS 13C NMR examinations suggested that the
85% DD CH was higher than that of 95% DD CH, especially in powder acetic acid solution mainly reacted with the amino group of the
form. A previous study has shown that the signal intensity of chi- chitosan and resulted in the b-1, 4 glycosidic bonds of chitosan
tosan spectra depends strongly on the molecular and segmental partly cracking, leading to exposed groups. Sorbitol principally
motions (Bajdik et al., 2009). Here, the molecular and segmental reacted with the exposed groups, such as the hydroxyl and amino
motions of 85% DD CH were more intense than those of 95% DD CH. groups on chitosan, after chitosan was partly dissolved by the acetic
This result was consistent with that found with X-ray diffraction acid. This observation agrees with previous reports that such
discussed previously. Furthermore, 85% and 95% DD CH appeared as exposed groups furnished the re-polymerization in systems and
weak peaks at 97.32 ppm and 97.33 ppm, respectively. According to formed dense network structures (Bajdik et al., 2009). After the
a previous study (Nishino, Matsuda, & Hirao, 2004), chitosan has chitosan chain was ruptured, sorbitol was easily inserted in the
three kinds of configuration, a, b and g, where different spaces of the molecular chain polymer and led to the film structure

Table 3
13
Signal assignment of the C NMR spectra of chitosan with different degrees of deacetylation in powder form (CH), and in films with (CHF) and without (CHFP) sorbitol.

Carbon atoms DD85% DD95%

CH CHF CHFP CH CHF CHFP

eCH3 22.97 ppm 22.95 ppm 22.95 ppm 22.94 ppm 22.87 ppm 23.122 ppm
C-2 57.01 ppm 57.03 ppm 56.98 ppm 56.98 ppm 57.08 ppm 57.041 ppm
C-6 60.72 ppm 60.79 ppm 60.76 ppm 60.72 ppm 60.93 ppm 60.745 ppm
C-3/C-5 74.90 ppm 74.93 ppm 74.97 ppm 74.89 ppm 74.90 ppm 74.952 ppm
C-4 82.47 ppm 81.97 ppm 82.05 ppm 82.36 ppm 82.15 ppm 82.246 ppm
C0 -1 97.32 ppm 97.21 ppm 97.19 ppm 97.33 ppm 97.35 ppm 97.315 ppm
C-1 104.64 ppm 103.92 ppm 104.24 ppm 104.65 ppm 103.90 ppm 104.225 ppm
C]O 173.66 ppm 173.47 ppm 173.29 ppm 173.33 ppm 173.45 ppm 173.431 ppm
 M. Liu et al. / Food Hydrocolloids 33 (2013) 186e191
and variations in formation elucidated here. The different compo-
sitions of the spiked and unspiked films would result in differences
in various properties.
4. Conclusion
The films spiked with sorbitol presented a homogenous and
smooth appearance as observed by SEM. This demonstrated that
sorbitol had good compatibility with 85% and 95% DD chitosan.
Results from XRD revealed a decrease in chitosan crystallinity in
spiked films and an increase in crystallinity in unspiked films
compared with the original powder. The addition of sorbitol to the
chitosan matrix can thus lead to a more orderly structure,
compared with films devoid of sorbitol that had a more irregular
crystalline structure. FT-IR analysis showed the chemical structure
of chitosan not to be affected by sorbitol, since both spiked and
unspiked films exhibited peaks at roughly the same wave numbers.
During film formation, molecules were rearranged along with the
main chain to form a more stable structure. Analysis by CP/MAS 13C
NMR showed that the breaking of chitosan bonds by acetic acid
enabled sorbitol to be inserted into the polymer molecular chain,
forming many hydrogen bonds in the process.
Varying the formation and structure of chitosan films can alter
properties such as water vapor permeability, tensile strength and
elongation. More work needs to be carried out to develop a range of
applications for chitosan films.
Acknowledgments
We gratefully acknowledge the staff of the University of Science
and Technology of China for their support and comments during
the preparation of this manuscript. This work was supported by
a grant from the National Science Foundation of China (No.
31271960).
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