Accepted Manuscript

Title: Rheological study of chitosan acetate solutions

containing chitin nanofibrils

Author: Jana Mikešová Jindřich Hašek Galina Tishchenko

Pierfrancesco Morganti

PII: S0144-8617(14)00619-5
DOI: http://dx.doi.org/doi:10.1016/j.carbpol.2014.06.043
Reference: CARP 9008

To appear in:

Received date: 14-4-2014

Revised date: 6-6-2014
Accepted date: 11-6-2014

Please cite this article as: Mikešová, J., Hašek, J., Tishchenko, G., and Morganti,
P.,Rheological study of chitosan acetate solutions containing chitin nanofibrils,
Carbohydrate Polymers (2014), http://dx.doi.org/10.1016/j.carbpol.2014.06.043

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 1 Rheological study of chitosan acetate solutions containing chitin nanofibrils
2
3
4 Jana Mikešová1, Jindřich Hašek1,2, Galina Tishchenko1, Pierfrancesco Morganti3
5
1
6 Institute of Macromolecular Chemistry AS CR, v. v. i., Heyrovského nám. 2, 162 06
7 Praha 6, Czech Republic,
2
8 Institute of Biotechnology AS CR, v.v.i., Vídeňská 1083, 142 20 Prague 4,

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3
9 Mavi Sud Srl, Viale Industria 1, 04011 Aprilia, Italy

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10
11
12

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13 Abstract
14
15 Rheological properties of chitosan acetate solutions containing chitin nanofibrils

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16 (n-chitin) and biocompatible plasticizers intended for preparation of biodegradable
17 films are reported in the steady, oscillatory and transient shear flow. The experiments
18 were performed on slurries with an optimum proportion 65/35 wt. % between

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19 chitosan and n-chitin in the films which was determined from our results of
20 mechanical properties and absorption of water vapor.
21 The time-dependent dynamic experiments revealed the chitin nanofibrils as an
22 effective “gelling agent” of chitosan phase. The phenomenon is explained by a
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23 chitosan-like surface of n-chitin and by the interactions inducing orientational
24 cooperativity of chitosan molecules dissolved in close neighborhood of the
25 anisotropic chitin nanofibrils.
26 Additions of glycerol or poly(ethylene glycol), improving mechanical properties of
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27 the films, delay significantly the onset of gelation of chitosan/n-chitin slurries. The
28 effect is induced by an increase in viscosity of the slurries and by their enhanced
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29 chaotropic character.
30
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31 1. Introduction
32
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33 Nanocomposite biomaterials based on chitosan and n-chitin are widely

34 investigated for their antimicrobial activity, biocompatibility and biodegradability. They
35 are already successfully used in medicine, e.g. in wound healing (Muzzarelli et al.,
36 2007). The ability of these natural materials to suppress microbial growth is a
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37 promising also for their application as biodegradable food packaging (Morganti et al.,
38 2013).
39 Nanoparticles with a high aspect ratio and a highly developed surface area, such
40 as clays or carbon nanotubes, serve as reinforcing agents of various polymers
41 including chitosan (Sriupayo et al., 2005, Casariego et al., 2009). In recent studies, a
42 growing rigidity of chitosan nanocomposites with increasing content of incorporated
43 n-chitin was demonstrated (Morganti et al. 2013 and Yudin et al., 2014). The
44 reinforcing effect of n-chitin on chitosan phase is excellent due to good reciprocal
45 compatibility of n-chitin and chitosan, both being linear cationic (1-4)-2-amino-2-
46 deoxy-β-D-glucans, differing only in terms of degree of acetylation (DA) and physical
47 form (Muzzarelli, 2012). Since the enhanced rigidity of chitosan/n-chitin films leads as
48 a rule to a decrease of their elasticity, the proper component proportion in the slurry,
49 including a plasticizer, must be previously defined to ensure the balance between the
50 strength and elasticity of the films.
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51 In preparation of chitosan/n-chitin films by the casting technique from solution, the
52 study of flow properties gets a special importance. Rheological measurements give
53 necessary recommendations for forming a uniform and stable layer of the slurry on a
54 support. It has been reported that rheological properties of moderately concentrated
55 chitosan solutions (Hwang & Shin, 2000, Desbrieres, 2002, Cho et al., 2006) reflect
56 interactions between chains of chitosan dissolved in acetic acid solutions even at
57 acidic pH, at which the most amino groups were protonated. A tendency of chitosan
58 macromolecules to interact with each other was also detected in acidic aqueous

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59 solutions with very low chitosan concentrations by means of static and dynamic light

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60 scattering (Anthonsen et al., 1994, Korchagina & Philipova, 2010). The conditions
61 necessary for formation of intermolecular reversible bonds, the nature of forming
62 aggregates and their stabilization are still under discussion. Montembault et al., 2005

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63 and Philipova et al., 2012, have concluded that the balance between the
64 intermolecular interactions through both hydrogen bonds and hydrophobic bindings
65 depends on the content of acetylated glucosamine groups in chitosan chains. In the

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66 case of low degree of acetylation (DA), the content of sites in chitosan chains
67 interacting through hydrophobic bindings is low and, therefore, hydrogen bonds
68 prevail in the reversible chains aggregation. Shamov et al., 2002, has concluded that

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69 acetate ions screen the protonated amino groups in chitosan chains in a great extent
70 and decrease the cationic property of chitosan having low DA.
71 In this work, new information about rheological properties of chitosan/n-chitin
72 slurries with and without bioplasticizers such as glycerol and poly(ethylene glycol)
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73 was obtained for optimizing the preparation of chitosan/n-chitin films. The effects of
74 chitin nanofibrils and plasticizers on gelation of the chitosan phase due to its self-
75 assembly are discussed in details.
76
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77 2. Experimental
78
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79 2.1. Materials
80 The commercial chitosan (HMC+ GmbH, Germany), molecular weight Mw 374 kDa,
81 degree of acetylation (DA) 11%, aqueous dispersion of chitin nanofibrils (Mavi Sud
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82 S.r.l., Italy) with DA 95% were used. The n-chitin dispersion was previously dialyzed
83 against distilled water through a 3 kDa-SpectraPor dialysis membrane to adjust pH to
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84 neutral values. After dialysis the n-chitin dispersion was concentrated by vacuum
85 evaporation to n-chitin concentration 0.042 g/g, pH 5.12. Biocompatible plasticizers
86 such as poly(ethylene glycol)(PEG-600) and glycerol (Aldrich, Germany) were used
as purchased.
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87
88
89 2.2. Preparation of solutions
90 The chitosan flakes (0.8 g) were dispersed in distilled water (40 ml), acidified with
91 99% acetic acid (0.8 ml) and mixed during 8 hours by mechanic stirring at room
92 temperature to prepare chitosan solutions. The homogeneous filled chitosan
93 solutions with chitosan/n-chitin proportion 65/35 wt. % were prepared by mixing the
94 homogeneous chitosan solution with the aqueous n-chitin dispersion during 2 hours.
95 The amount of plasticizers was equal to 30 wt. % of total content of chitosan and n-
96 chitin in the slurry. The component proportions (wt. %) in the measured solutions are
97 summarized in Table 1.
98
99
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100 Table 1
101 Composition [wt. %] of chitosan solutions under study
102
CS/n-chitin CS/n-chitin
SOLUTION chitosan CS/n-chitin
+ glycerol + PEG
chitosan (CS) 1.9 1.5 1.5 1.5
Q water 96.1 96.0 95.0 95.0
acetic acid 2.0 1.6 1.6 1.6

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n-chitin - 0.8 0.8 0.8

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glycerol - - 1.0 -
PEG-600 - - - 1.0

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103
104
2.3. Rheological characterization

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105
106 Rheological properties of the pure and filled chitosan solutions were measured at
107 room temperature using a rheometer Physica MCR 501 (Anton Paar GmbH, Austria)
108 equipped with anti-slipping parallel-plate geometry. The free surface of samples

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109 between the plates was covered by a micro-layer of silicone oil to avoid evaporation.
110 In the steady shear flow, the strain- and stress-controlled experiments were carried
111 out. In the oscillatory shear flow, the viscoelastic properties were measured in the
112 linear viscoelasticity region confirmed with strain tests at the frequency 1 Hz. The
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113 frequency- and time-dependent oscillatory tests were performed at the small
114 amplitude (2% strain) to minimize disturbances of the internal structure in the studied
115 solutions. The measurements with pre-shearing have been started after 3 s of the
116 steady shear at the rate 0.03 s-1. Three independent measurements were carried out
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117 for each studied solution.

118
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119 3. Results and discussion

120
121 Rheological properties of chitosan solutions containing chitin nanofibrils with and
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122 without the biocompatible plasticizers were measured in the steady, oscillatory and
123 time-dependent shear flow and compared with rheology of the pure chitosan solution.
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124 Our previous results on absorption of water vapor in chitosan/n-chitin films (Morganti
125 et al., 2013) revealed very small change of sorption, 2 wt. % only if the concentration
126 of chitin nanofibrils in the films increased from 24 wt. % to 36 wt.%. Based on these
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127 results, we conclude that an optimal n-chitin content in chitosan films should be about
128 30-40 wt. %. The 35 wt. % concentration of n-chitin in chitosan films was chosen for
129 the study of rheological properties of slurries using for films solution casting. Effects
130 of 20 wt. % additions of glycerol and poly(ethylene glycol) on mechanical and barrier
131 properties of prepared films with the content 35 wt. % of n-chitin were measured and
132 the results will be presented in a separate paper.
133 The addition of n-chitin into chitosan solution accelerates the process of self-
134 assembly of chitosan macromolecules resulting in faster gelation of the slurry. The
135 solid-like behavior is typical for filled chitosan solutions. It was characterized by
136 values of the yield stress in the steady shear and by appearance of a low frequency
137 plateau of the dynamic moduli in oscillatory flow. The rate of the self-assembly
138 process until the formation of reversible gel was considered as an increase in
139 elasticity of the studied solutions with time.
140

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141 3.1. Steady shear flow properties of pure chitosan solution
142 The pre-sheared 2 wt. % chitosan solution exhibited a Newtonian behavior at low
143 shear rates. The shear thinning (non-Newtonian) region appeared at higher shear
144 rates where the polymer chains were oriented in the direction of flow. The flow curve
145 of the pre-sheared solution is plotted in Fig.1 as the dependence of steady shear
146 viscosity η on the shear rate. The tendency of the chitosan chains to association was
147 found when the solution was kept in the rheometer upon the cessation of flow and
then the measurements were carried out repeatedly after 15 and 30 minutes. After

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148
the rest, the growth of viscosity appeared in the flow curves (Fig. 1). The increase in

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149
150 viscosity at very low shear rates (yielding) correlates with the content of chitosan
151 aggregates in the solution. The marked shear-thinning character at this region

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152 indicates that the aggregates begin to deform and break down in the shear. At the
153 shear rates higher 10 s-1, the viscosity becomes independent on the rest time due to
154 the breakage of aggregates. The used chitosan solution behaves as an associating

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155 polyelectrolyte. The obtained flow curve did not fit the frequency dependence of the
156 complex viscosity obtained from oscillatory tests at low shear rates, i.e. the frequently
157 used Cox-Merz rule (Cox & Merz, 1958) is not valid for the chitosan solution. The
158 observed rheological behavior corresponds to the results published by Cho et al.,

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159 2006.
160
161
2
10
M
Pa·s
d
1
10
te


0
p

10
ce

-1
10
-5 -4 -3 -2 -1 0 1 2 3
Ac

10 10 10 10 10 10 10 10 1/s 10
.
162 Shear Rate 
163
164 Fig. 1. Steady shear viscosity as a function of the shear rate of chitosan 2 wt. %
165 aqueous acetic acid solution measured at the various rest times after pre-shearing.
166 The pre-sheared solution (), after the rest time 15 min () and 30 min ().
167
168
169 3.2. Steady shear flow properties of chitosan solutions containing chitin nanofibrils
170 A small addition of chitin nanofibrils into chitosan solutions resulted in a reversible
171 gelation of the slurry at rest. The viscosity increased about 3-4 decades, a slip of the
172 filled solutions between the plates of rheometer at low shear rates as well as flow
173 instabilities at higher shear rates occurred. The slipping effects excluded standard

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174 measurements in the mode with increasing shear rate (step-up rate mode) until the
175 critical shear rate about 0.03 s-1. At the shear rates higher than this value, a physical
176 structure between chitosan chains and chitin nanofibrils was disturbed and at very
177 high shear rates, the non-Newtonian behavior became similar to the pure chitosan
178 solution. The experiments performed in the opposite mode with decreasing shear
179 rate (step-down rate mode) showed a self-recovery of the nanofibrils-chitosan
180 structure from the disturbed state into a gel structure. The similar behavior (slipping
181 and structure recovery) was observed also in the shear rate measurements of the

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182 filled solutions modified by glycerol or PEG plasticizers.

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183
184 3.3. Determination of the yield stress in solutions containing n-chitin

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185 The experiments in the steady shear flow revealed a solid-like behavior of filled
186 chitosan solutions at rest. The solutions started to flow at a critical shear rate. This
187 typical phenomenon of medium and concentrated suspensions is characterized with

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188 the yield stress – the minimum stress needed to initiate flow. Stress controlled
189 experiments were carried out to determine these flow characteristics. Fig. 2 shows
190 the results as a dependence of the shear viscosity on shear stress. The filled solution
191 behaves as an elastic solid below the yield stress and a sharp drop in viscosity after

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192 reaching the yield stress reflects the destruction of a physical network. The yield
193 stress was evaluated as the stress corresponding to the maximum value of shear
194 viscosity. For the filled solution, the value of the yield stress equal to 6.8 Pa was
195 determined.
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196
4
10
Pa·s
d

3
10
te

2
10
p

 1
10
ce

0
10
Ac

-1
10
0 1 2 3
10 10 10 Pa 10
197 Shear Stress 
198
199 Fig. 2. Effects of the plasticizers on the yield stress value of the chitosan/n-chitin
200 slurries. Steady shear viscosity vs. shear stress of the pure chitosan solution () and
201 the slurries: chitosan/n-chitin (), chitosan/n-chitin/glycerol (), chitosan/n-
202 chitin/PEG (□).
203
204 The reversible physical network formed in the n-chitin filled chitosan acetate
205 solutions is induced by strong intermolecular interactions between chitosan chains
206 and the interactions among chitosan molecules and chitin nanofibrils. The nature of
207 interactions lies in mechanical entanglements of chitosan chains, and formation of
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208 temporary hydrogen bonds between the hydrogen bond donors and remaining
209 acceptors. Excellent solubility of chitosan in an acidic solution is well known.
210 However, the properties of n-chitin fibrils in aqueous solutions are more complex.
211 The interior of chitin nanofibrils is formed by a rigid α-chitin crystalline phase formed
212 by antiparallel chitin molecules well stabilized by intermolecular H-bonds between
213 acetyl groups of neighboring chains. The degree of deacetylation of the used n-chitin,
214 determined by NMR, is very low (DA~95%). Supposing that the deacetylation agents
215 cannot easily penetrate into the depth of the α-chitin crystalline phase, we can

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216 conclude that several layers of chitin nanofibrils near the surface are partly

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217 deacetylated. The strong hydrogen bridges between acetyl groups of neighbouring
218 antiparallel chitin molecules are responsible for insolubility of the chitin crystalline
219 phase. The acetyl groups of chitin chains exposed to solvent which were converted

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220 to amino groups (chitosan-like surface) determine the excellent stability of aqueous
221 dispersion of huge elongated rigid chitin nanofibrils for many months. Each nanofibril
222 is composed of thousands of macromolecules safely anchored by their chitin

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223 segments in the crystalline core of nanofibrils. The deacetylated parts of all these
224 molecules near the fibril surface form numerous hydrogen bonds with water in acidic
225 solutions via their charged amine groups.

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226 Addition of the chitin nanofibrils into the chitosan solution results in a gel formation.
227 In the absence of an external force, the system is in a deep potential minimum,
228 because the intermolecular interactions between chitosan chains and chitosan-
229 nanofibrils are strong. The strong interactions of each highly anisotropic n-chitin
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230 nanoparticle with many thousands of chitosan molecules uniformly distributed in a
231 large volume of solution ensure its stability and solid-like behavior under static
232 conditions. Thus, the following model can be used to explain the observed
233 rheological behavior of the system. The sparse polycation matrix of chitosan
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234 molecules homogeneously occupies the whole volume of the solution. It is connected
235 by strong attractive and repulsive forces with a network of equally uniformly
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236 distributed chitin nanoparticles with high capacity for accumulation of kinetic energy.
237 The plasticizers PEG-600 and glycerol added to the filled chitosan solutions
238 decreased the yield stress value by about one half. The modification of solutions with
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239 glycerol and PEG reduced their yield stresses similarly to the values 3.9 and 3.1 Pa,
240 respectively. Effect of the plasticizers, at their concentration comparable with of n-
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241 chitin, on the filled solution is explained by shielding the electrostatic interactions
242 between positively charged segments, shifting a character of solvent to the
243 chaotropic side and by lowering a number of free water molecules in solution.
244
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245 3.4. Linear viscoelastic behavior of chitosan solutions containing n-chitin (small-
246 amplitude oscillatory shear tests)
247 The rheological investigation of the filled solutions in a small-amplitude oscillatory
248 shear has a great advantage against the steady shear measurements. These
249 experiments are performed at very low deformations and therefore no slip of the filled
250 chitosan slurries is observed. The deformation amplitude during the oscillatory
251 experiments is located inside the linear viscoelastic region. This region defines
252 maximum deformation that can be applied to the filled solution without destructing the
253 internal structure. The highly sensitive characteristic of the network structure in
254 solution is the storage modulus G', which is the measure of elasticity. Rheological
255 properties of the filled chitosan solutions in oscillatory flow are shown in Fig. 3 where
256 the linear viscoelastic characteristics, the storage modulus G', and loss modulus G"
257 are plotted against angular shear frequency ω. Similar to the steady shear flow, the

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258 added nanofibrils into the chitosan solution moved the both dynamic moduli up 3-4
259 decades higher. However, more interesting is the principal change in the rheological
260 behavior of the slurries. The pure chitosan solution behaves as a viscoelastic fluid
261 because of G" > G'. On the other hand, in the case of filled solution, the storage
262 modulus overwhelms the loss modulus (G' > G"), the elasticity is dominated and the
263 solution shows the properties as a soft solid. This solid-like phenomenon is
264 manifested also with the frequency independent G' and G", the typical feature of
265 strong gels (rubber-like behavior). The both phenomena are attributed to a physical

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266 network, formed due to chitin-chitosan interactions in the filled solutions and, as a

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267 rule, indicate an improvement of mechanical and barrier properties of the chitosan/n-
268 chitin nanocomposites.
269

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4 4
10 10

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Pa Pa

2 2
10 10

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1 1
10 10
G' G''
0 0
10 10
M
-1 -1
10 10
-2 -2
10 10
d
-3 -3
10 10
-2 -1 0 1 2
10 10 10 10 rad/s 10
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270 Angular Frequency 

271
272 Fig. 3. Storage (G') and loss modulus (G") as a function of the angular frequency of
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273 the pure chitosan solution and the chitosan/n-chitin slurries. The full marks are used
274 for the storage modulus values. The pure chitosan solution (,), and the slurries:
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275 chitosan/n-chitin (,), chitosan/n-chitin /glycerol (▼,), chitosan/n-chitin/PEG

276 (■,□).
277
The effects of the used plasticizers on the physical gel formation in chitosan/n-
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278
279 chitin slurries are shown in Fig. 3. As accepted, the elasticity of the modified slurries
280 drops and the rubber-like region is observed at low frequencies only. The impact of
281 both PEG and glycerol on the viscoelastic characteristics was similar and the
282 obtained results are confirmed by the reduced yield stresses. Otherwise, the modified
283 filled chitosan solutions showed the convenient properties for films processing
284 because of the decreased values of slurries viscosity and elasticity.
285
286 3.5 Observation of the self-assembly gelling process in chitosan solutions containing
287 n-chitin (sol-gel transition)
288
289 A self-assembly gelling process in chitosan/n-chitin slurries was examined using
290 time-dependent experiments in the oscillatory shear flow. To avoid a destruction of
291 the internal structure of slurries, the measurements were carried out at very small

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292 deformation. The results of the tests are depicted in Fig. 4 where the time
293 dependences of the storage (G') and loss (G") moduli are plotted. The dynamic
294 moduli increase with time for both the pure and filled chitosan solutions. In the case
295 of chitosan/n-chitin solutions and those modified with plasticizers, both moduli
296 intersect in a crossover point (gel point) where the state of slurries was changed from
297 a viscoelastic fluid to a viscoelastic solid phase. The gel point indicated the formation
298 of a physical network between chitosan macromolecules and chitin nanofibrils due to
299 the self-assembly process described in 3.3. The added plasticizers retarded the

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300 formation of gel that was confirmed by the results of yield stress and oscillatory tests.

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301 The chitosan/n-chitin and chitosan/n-chitin /PEG slurries reached the gel point in 28
302 min and 80 min, respectively, after pre-shearing. In pure chitosan solution, the
303 increase in both moduli with time reflected networking between chitosan

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304 macromolecules. Nevertheless, in the absence of large rigid stabilizing particles such
305 as n-chitin, the long-distance structure was not achieved even after 3.5 h. The similar
306 slopes of the elasticity increase (see modulus G' in Fig. 4) show that kinetics of the

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307 self-assembly in the pure and filled chitosan solutions should be analogous at the
308 beginning of the process. This fact encourages us to suppose that the formation of
309 aggregates between chitosan macromolecules in solution is the primary process also

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310 in the presence of chitin nanofibrils. The real reason of gelation after n-chitin addition
311 is the ability of each single n-chitin particle to influence simultaneously thousands of
312 chitosan molecules penetrating homogeneously the whole volume of the solution.
313
M
2 2
10 10
d
Pa Pa
te

1 1
G' 10 10 G''
p
ce
Ac

0 0
10 10
0 2 000 4 000 6 000 8 000 10 000 s 12 000
314 Time t
315
316 Fig. 4. Physical gelation of the chitosan/n-chitin slurries: time dependence of the
317 dynamic moduli of the pure chitosan solution - (G' (), G"()) and the slurries:
318 chitosan/n-chitin – (G' (■), G"()), chitosan/n-chitin /PEG - (G' (□), G"()).
319
320 4. Conclusions
321
322 The incorporation of the highly anisotropic chitin nanofibrils into chitosan solutions
323 changed their rheological properties significantly. The chitosan/n-chitin slurries
324 revealed a solid-like behavior and their steady shear flow was limited due to the
325 slipping on the plates of rheometer and the flow instabilities. The independence of

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326 the dynamic moduli on frequency showed that the rubber-like behavior is typical for
327 the chitosan/n-chitin solutions under study. The self-assembly processes in both the
328 pure and filled chitosan solutions were elucidated by the time-dependent
329 measurements of both dynamic moduli up to reaching the gel-point.
330 The chitin nanofibrils were recognized as a strong “gelling agent”. It was found that
331 at the beginning of the self-assembly, the kinetics is similar in both the pure and the
332 filled solutions. The gelling time is prolonged and the yield stress values decrease
333 when plasticizers are added into chitosan/n-chitin slurries.

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334 The rheological behavior of the system is explained by a tough and elastic

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335 interconnection between two physical networks - the network of chitosan chains
336 homogeneously penetrating the whole volume of sample and the homogeneous
337 network of chitin nanofibrils with enormous capacity of kinetic energy.

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338 The plasticizer molecules dissolved in the aqueous phase of chitosan/n-chitin
339 solution increase the viscosity of slurry and protect water to form polarized structures
340 around the charged surfaces, i.e. they give the solution more chaotropic character.

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341 The observed decrease of the yield stress values after plasticizers addition into
342 chitosan/n-chitin slurries is thus explained both by an increase in viscosity and a
343 decrease in repulsive forces between polycation segments.

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344
345
346 5. Acknowledgements
347
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348 The authors gratefully acknowledge the financial support of the European Union
349 (Grant 315233).
350
351
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352 6. References
353
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354 Anthonsen, M. W., Varum, K. M., Hermansson, A. M., Smidsrod, O., & Brant, D. A.
355 (1994). Aggregates in acidic solutions of chitosans detected by static laser light
356 scattering. Carbohydrate Polymers, 25, 13-23.
p

357 Casariego, A., Souza, B. W. S., Cerqueira, M. A., Teixeira, J. A., Cruz, L., Diaz, R.,
358 & Vicente, A. A. (2009). Chitosan/clay films’ properties as affected by biopolymer and
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359 clay micro/nanoparticles’ concentrations. Food Hydrocolloids, 23, 1895-1902.

360 Cho, J., Heuzey, M-C., Bégin, A., & Carreau, P. J. (2006). Viscoelastic properties of
361 chitosan solutions: Effect of concentration and ionic strength. Journal of Food
362 Engineering, 74, 500-515.
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363 Cox, W. P. & Merz, E. H. (1958). Correlation of dynamic and steady flow viscosities.
364 Journal of Polymer Science, 28, 619-622.
365 Desbrieres, J. (2002). Viscosity of semiflexible chitosan solutions: Influence of
366 concentration, temperature, and role of intermolecular interactions.
367 Biomacromolecules, 3, 342-349.
368 Hamdine M., Heuzey M.-C., & Bégin A. (2005). Effect of organic and inorganic acids
369 on concentrated chitosan solutions and gels. International Journal of Biological
370 Macromolecules 37, 134-142.
371 Hwang, J. K., & Shin, H. H. (2000). Rheological properties of chitosan solutions.
372 Korea-Australia Rheology Journal 12, 175-179.
373 Korchagina, E. V., & Philippova, O. E. (2010). Multichain aggregates in dilute
374 solutions of associating polyelectrolyte keeping a constant size at the increase in the
375 chain length of individual macromolecules. Biomacromolecules, 11, 3457–3466.

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Page 9 of 12
376 Montembault, A., Viton, Ch., & Domard, A. (2005). Rheometric study of the gelation
377 of chitosan in aqueous solution without cross-linking agent. Biomacromolecules, 6,
378 653-662.
379 Morganti, P., Tishchenko, G., Palombo, M., Kelnar, I., Brozova, L., Spirkova, M.,
380 Pavlova, E., Kobera, L., & Carezzi, F. (2013). Chitin nanofibrils for biomimetic
381 products: Nanoparticles and nanocomposite chitosan films in health care. In: Marine
382 Biomaterials: Characterization, Isolation and Applications. Chapter 35, 681-716. Ed.
383 Se-Kwon Kim, CRC Press, Taylor & Francis Group, ISBN: 9781466505643.

t
384 Muzzarelli, R. A. A., Morganti, P., Morganti, G., Palombo, P., Palombo, M., Biagini,

ip
385 G., Belmonte, M. M., Giantomassi, F., Orlandi, F., & Muzzarelli, C. (2007). Chitin
386 nanofibrils/chitosan glycolate composites as wound medicaments. Carbohydrate
387 Polymers, 70, 274-284.

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388 Muzzarelli, R. A. A. (2012). Nanochitins and nanochitosans, paving the way to eco-
389 friendly and energy-saving exploitation of marine resources. Polymer Science:
390 A Comprehensive Reference, 10, 153-164.

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391 Philippova, O. E., Korchagina, E. V., Volkov, E. V., Smirnov, V. A., Khokhlov, A. R.,
392 & Rinaudo, M. (2012). Aggregation of some water-soluble derivatives of chitin in
393 aqueous solutions: Role of the degree of acetylation and effect of hydrogen bond

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394 breaker. Carbohydrate Polymers, 87, 687-694.
395 Shamov, M. V., Bratskaya S. Yu., & Avramenko V. A. (2002). Interaction of
396 carboxylic acids with chitosan: Effect of pK and hydrocarbon chain length. Journal of
397 Colloid and Interface Science, 249, 316-321.
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398 Sriupayo, J., Supaphol, P., Blackwell, J., & Rujiravanit, R. (2005). Preparation and
399 characterization of α-chitin whisker-reinforced chitosan nanocomposite films with or
400 without heat treatment. Carbohydrate Polymers, 62, 130–136.
401 Yudin, V. E., Dobrovolskaya, I. P., Neelov, I. M., Dresvyanina, E. N., Popryadukhin,
d

402 P. V., Ivan’kova, E. M., Elokhovskii, V. Yu., Kasatkin, I. A., Okrugin, B. M., Morganti,
403 P. (2014). Wet spinning of fibers made of chitosan and chitin nanofibrils.
te

404 Carbohydrate Polymers, 108, 176-182.

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405
406 Table 1
407 Composition [wt. %] of chitosan solutions under study
408
CS/n-chitin CS/n-chitin
SOLUTION chitosan CS/n-chitin
+ glycerol + PEG
chitosan (CS) 1.9 1.5 1.5 1.5
Q water 96.1 96.0 95.0 95.0

t
acetic acid 2.0 1.6 1.6 1.6

ip
n-chitin - 0.8 0.8 0.8
glycerol - - 1.0 -

cr
PEG-600 - - - 1.0
409
410

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411

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411 Keywords

412 Rheology, chitosan solutions, chitin nanofibrils

413

414

t
ip
415 Highlights

cr
416  Chitin nanofibrils (n-chitin) change flow properties of chitosan solutions dramatically.

417  Chitosan/n-chitin slurries show solid- and rubber-like behavior.

us
418  Chitin nanofibrils are recognized as a strong “gelling agent”.

419  The beginning of gelation is delayed by bioplasticizers.

420
421
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