# Chitosan Derivatives/Calcium Carbonate Composite Capsules Prepared by the Layer-by-Layer Deposition Method

Authors: Takashi Sasaki; Misato Shimizu; Yusong Wu; Kensuke Sakurai
Journal: Journal of Nanomaterials
 (2008-02-13)

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## Abstract

Core/shell capsules composed of calcium carbonate whisker core (rod-like shape) and chitosan/chitosansulfate shell were prepared by the layer-by-layer deposition technique. Two chitosan samples of different molecular weights (Mw=9.7×104 and 1.09×106g·mol-1) were used as original materials. Hollow capsules were also obtained by dissolution of the core in hydrochloric acid. Electron microscopy revealed that the surface of the shell is rather ragged associated with some agglomerates. The shell thickness l obeys a linear relation with respect to the number of deposited layers m as l=md+a(a>0). The values of d (thickness per layer) were 4.0 and 1.0 nm for the higher and lower Mw chitosan materials, respectively, both of which are greater than the thickness of the monolayer. The results suggest that the feature of the deposition does not obey an ideal homogeneous monolayer-by-monolayer deposition mechanism.  Shell crosslinked capsules were also prepared via photodimerization reaction of cinnamoyl groups after a deposition of cinnamoyl chitosan to the calcium carbonate whisker core. The degree of crosslink was not enough to stabilize the shell structure, and hollow capsule was not obtained.


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## Body

## 1. Introduction

Development of encapsulation
technique to fabricate nanosized core/shell or hollow particles is one of the
most important and intriguing issues in the field of nanotechnology, especially
because of its potential application to drug delivery systems. As for the shell material, biocompatibility
is also important for such biomedical applications. Various approaches to prepare
nanocapsules have been developed, and these may be classified into two
categories, that is, capsules are formed either (1) by taking advantage of the self-assembly
of amphiphilic molecules including micelles and vesicles, or (2) by using a
core particle as a template. The former includes shell formation by
polymerization and/or crosslinking of monomers in self-assembled system [1–4]. In
general, the approaches of (1) are rather tricky, requiring sophisticated
techniques to control the size and shape of the resulting capsule. In the latter
approaches, on the other hand, the size and shape are determined by those
of the core particles. In addition, if the core material is soluble in a
suitable solvent, hollow capsules can easily be obtained. For example,
ultrathin polymer shells have been formed on the surface of calcium carbonate particles
via emulsion polymerization to prepare rod-like core/shell and hollow capsules
[5, 6].The layer-by-layer
deposition method, which belongs to the category (2), is one of the most powerful
and versatile techniques to fabricate thin complex multilayers with controllable
thicknesses and geometries [7, 8]. By this technique, multilayered films can be
prepared by alternative adsorption of oppositely charged polyelectrolytes on a
substrate or particle surface. Various polyelectrolytes have been demonstrated
to give multilayered films on the surface of various particles [8–15]. Hollow
capsules can also be prepared by dissolution of the core materials in appropriate
solvents [9, 10, 14]. Also, it is
possible in some cases to substitute the original core for another material. For example, Shchukin et al. prepared ferrite
and magnetite particles inside hollow capsules that had been prepared by the
layer-by-layer method [16].In this paper, we
report preparations of biocompatible core/shell capsules by the layer-by-layer
technique. The shells were formed with alternative deposition of chitosan
(cationic polyelectrolyte) and chitosansulfate (anionic polyelectrolyte) on the
surface of calcium carbonate whisker as a core. The core can be removed being dissolved
in hydrochloric acid to obtain hollow capsules. As the whisker particles have a
rod-like shape, the obtained capsules are rod-like. Such a rod-like shape is
unique in the sense that the particle can be oriented by elongation or under flow.
Chitosan is obtained by acetylation of an abundant natural polysaccharide,
chitin. There has been a growing interest to chitosan as a biomedical material
because of its biocompatibility and biodegradability with little toxicity. To
stabilize the core/shell structure, we also tried to prepare shell crosslinked
capsules: cinnamoyl chitosan was deposited onto the whisker core, and the
crosslinking via photodimerization of the cinnamoyl groups was performed by UV
irradiation.


## 2. Experimental Methods

### 2.1. Materials

Chitosan(Mw=9.7×104and1.09×106gmol−1) was
purified by dissolving it in 1.0% acetic acid aqueous solution, followed by
precipitation in 10% aqueous solution of sodium hydroxide. The precipitate was
washed with distilled water, methanol, acetone, and with N,N-dimethyl formamide
(DMF). Calcium carbonate whisker (BS-P, Maruo Calcium Co., Japan)
was supplied from the manufacturer, of which the diameter ranged from 300 to
600 nm and the length ranged from 1.0 to 3.0 μm. The surface of BS-P
whisker was covered with a thin layer of calcium phosphate via phosphoric acid
treatment.


### 2.2. Preparation of Chitosansulfate

Sulfonic
groups were introduced at either the hydroxyl groups or amino groups in
2-amino-2-deoxy-D-glucopyranose unit of chitosan in the following method. Into
a flask containing 25 ml of DMF was added dropwise 16.5 ml of chlorosulfonic
acid in an ice-water bath, and immediately added 2.5 g of the purified
chitosan. The mixture was stirred at room temperature for 24 hours. After the
reaction was completed, the mixture was poured into 400 mL of methanol with
stirring. The precipitate was separated and washed with methanol. The obtained
product was dissolved in distilled water, and the solution was neutralized with
20% aqueous solution of sodium hydroxide. Then, the solution was subject to
dialysis with distilled water for 36 hours. Final product chitosansulfate was
obtained by freeze-drying the dialyzed solution. IR (KBr) 810 (C–O–S), 1122 (SO2) cm−1.
Elementary analysis revealed the number of sulfonic groups per repeating unit of
chitosan to be 0.86.


### 2.3. Layer-by-Layer Deposition

Layer-by-layer
deposition of chitosan/chitosansulfate was executed according to the membrane
filtration method by Voigt et al. [17], that is, the suspended state of particles
were maintained throughout the deposition-filtration-lavage-filtration cycles
to avoid any damages of capsules due to particle aggregation. Into 500 mL of an aqueous solution consisted of sodium chloride (0.50 mol L−1) and
acetic acid (1.0% v/v) was dissolved 2.5 g of chitosan (referred to as A). An
aqueous solution of chitosansulfate (0.53 g L−1) that
contained 0.50 mol L−1 of sodium
chloride was prepared (referred to as B). One gram of calcium carbonate whisker
(BS-P) was suspended in 20 mL of distilled water with gentle sonication, and 5 mL of the chitosan solution A was added. The suspension was stirred for 5 minutes
for deposition of chitosan salt layer on the whisker surface. The suspended
particles were washed by adding distilled water continuously over a filter with
constant stirring so as to keep a well-suspended state even during the
filtration. Subsequently, 5 mL of the solution B was added to the suspension,
which was stirred for 5 minutes for deposition of chitosansulfate layer. The
suspension was then washed with distilled water in the same manner as above.
The cycle of deposition with solutions A and B was repeated 2–7 times to prepare capsules with different total
numbers of deposited layers m(m=4−14).


### 2.4. Preparation of Hollow Particles

The
core/shell capsules were converted to hollow particles by adding hydrochloric
acid to their aqueous suspensions (10 wt%) to dissolve the core part (calcium
carbonate). The acid suspensions were
stirred for 2 hours, of which the pH value was kept below 1.0. The particles
were separated by centrifugation and washed with water several times. The final products were dried under vacuum
for more than 48 hours at room temperature.


### 2.5. Preparation of Cinnamoyl Chitosan

Acylation
of chitosan with cinnamoyl chloride was performed according to the method
described in [18]. The cinnamoyl group was introduced at either of the two
hydroxyl groups in the repeating unit of chitosan. To 30 mL of methanesulfonic
acid was added 2 g of the purified chitosan. The mixture was cooled in an
ice-water bath, and was added 0.4 g of cinnamoyl chloride. Subsequently, the
solution was stirred at room temperature for 2 hours, and then, it was poured
into 500 mL of methanol. The precipitate was separated by filtration, washed
with methanol and with acetone. The product was then reprecipitated in ammonium
hydroxide aqueous solution (pH = 8.0). Finally, the separated product was
washed with water, methanol, and acetone, followed by filtration and drying
under vacuum at room temperature for 24 hours. IR (KBr) 1705 (ester carbonyl), 1636
(disubstituted alkene) cm−1.
Quantitative analysis of UV absorption at 280 nm in DMF/water (1/1) containing
acetic acid (1.0%v/v) solution revealed that the number of cinnamoyl groups was
0.15 per repeating unit of chitosan.


### 2.6. Deposition of Cinnamoyl Chitosan on the Whisker Surface

Into
500 mL of an aqueous solution consisted of sodium chloride (0.50 mol L−1) and
acetic acid (1.0%v/v), was dissolved 2.5 g of cinnamoyl chitosan. The solution
was added to an aqueous suspension of BS-P (50 g L−1), and it
was stirred for 10 minutes for deposition of cinnamoyl chitosan single layer on
the whisker surface. Subsequently, UV irradiation (λ=254nm) to the
suspension was executed for 48 hours at room temperature with constant
stirring.


### 2.7. Microscopy

To
investigate the morphology and sizes of the capsules, transmission electron
microscopy (TEM) was done by using a Jeol JEM-2000FXII operated with an
acceleration voltage of200kV. No shadowing was applied. Scanning electron
microscopy (SEM) was also done by using a Hitachi S-2300.


### 2.8. Evaluation of the Core/Shell Ratio

Compositions
(core/shell ratio) of the capsules were evaluated by volumetric analysis. A
finite mass of the capsule was immersed in hydrochloric acid with stirring
until the core material was completely dissolved. Hydrochloric acid and water
in the obtained solution were evaporated and residual precipitate was redissolved
in deionized water. Calcium ion content in this solution was determined by
chelatometric titration, and the ratio of the mass of the shell to that of the whole
capsulems/mt was evaluated.


## 2.1. Materials

Chitosan(Mw=9.7×104and1.09×106gmol−1) was
purified by dissolving it in 1.0% acetic acid aqueous solution, followed by
precipitation in 10% aqueous solution of sodium hydroxide. The precipitate was
washed with distilled water, methanol, acetone, and with N,N-dimethyl formamide
(DMF). Calcium carbonate whisker (BS-P, Maruo Calcium Co., Japan)
was supplied from the manufacturer, of which the diameter ranged from 300 to
600 nm and the length ranged from 1.0 to 3.0 μm. The surface of BS-P
whisker was covered with a thin layer of calcium phosphate via phosphoric acid
treatment.


## 2.2. Preparation of Chitosansulfate

Sulfonic
groups were introduced at either the hydroxyl groups or amino groups in
2-amino-2-deoxy-D-glucopyranose unit of chitosan in the following method. Into
a flask containing 25 ml of DMF was added dropwise 16.5 ml of chlorosulfonic
acid in an ice-water bath, and immediately added 2.5 g of the purified
chitosan. The mixture was stirred at room temperature for 24 hours. After the
reaction was completed, the mixture was poured into 400 mL of methanol with
stirring. The precipitate was separated and washed with methanol. The obtained
product was dissolved in distilled water, and the solution was neutralized with
20% aqueous solution of sodium hydroxide. Then, the solution was subject to
dialysis with distilled water for 36 hours. Final product chitosansulfate was
obtained by freeze-drying the dialyzed solution. IR (KBr) 810 (C–O–S), 1122 (SO2) cm−1.
Elementary analysis revealed the number of sulfonic groups per repeating unit of
chitosan to be 0.86.


## 2.3. Layer-by-Layer Deposition

Layer-by-layer
deposition of chitosan/chitosansulfate was executed according to the membrane
filtration method by Voigt et al. [17], that is, the suspended state of particles
were maintained throughout the deposition-filtration-lavage-filtration cycles
to avoid any damages of capsules due to particle aggregation. Into 500 mL of an aqueous solution consisted of sodium chloride (0.50 mol L−1) and
acetic acid (1.0% v/v) was dissolved 2.5 g of chitosan (referred to as A). An
aqueous solution of chitosansulfate (0.53 g L−1) that
contained 0.50 mol L−1 of sodium
chloride was prepared (referred to as B). One gram of calcium carbonate whisker
(BS-P) was suspended in 20 mL of distilled water with gentle sonication, and 5 mL of the chitosan solution A was added. The suspension was stirred for 5 minutes
for deposition of chitosan salt layer on the whisker surface. The suspended
particles were washed by adding distilled water continuously over a filter with
constant stirring so as to keep a well-suspended state even during the
filtration. Subsequently, 5 mL of the solution B was added to the suspension,
which was stirred for 5 minutes for deposition of chitosansulfate layer. The
suspension was then washed with distilled water in the same manner as above.
The cycle of deposition with solutions A and B was repeated 2–7 times to prepare capsules with different total
numbers of deposited layers m(m=4−14).


## 2.4. Preparation of Hollow Particles

The
core/shell capsules were converted to hollow particles by adding hydrochloric
acid to their aqueous suspensions (10 wt%) to dissolve the core part (calcium
carbonate). The acid suspensions were
stirred for 2 hours, of which the pH value was kept below 1.0. The particles
were separated by centrifugation and washed with water several times. The final products were dried under vacuum
for more than 48 hours at room temperature.


## 2.5. Preparation of Cinnamoyl Chitosan

Acylation
of chitosan with cinnamoyl chloride was performed according to the method
described in [18]. The cinnamoyl group was introduced at either of the two
hydroxyl groups in the repeating unit of chitosan. To 30 mL of methanesulfonic
acid was added 2 g of the purified chitosan. The mixture was cooled in an
ice-water bath, and was added 0.4 g of cinnamoyl chloride. Subsequently, the
solution was stirred at room temperature for 2 hours, and then, it was poured
into 500 mL of methanol. The precipitate was separated by filtration, washed
with methanol and with acetone. The product was then reprecipitated in ammonium
hydroxide aqueous solution (pH = 8.0). Finally, the separated product was
washed with water, methanol, and acetone, followed by filtration and drying
under vacuum at room temperature for 24 hours. IR (KBr) 1705 (ester carbonyl), 1636
(disubstituted alkene) cm−1.
Quantitative analysis of UV absorption at 280 nm in DMF/water (1/1) containing
acetic acid (1.0%v/v) solution revealed that the number of cinnamoyl groups was
0.15 per repeating unit of chitosan.


## 2.6. Deposition of Cinnamoyl Chitosan on the Whisker Surface

Into
500 mL of an aqueous solution consisted of sodium chloride (0.50 mol L−1) and
acetic acid (1.0%v/v), was dissolved 2.5 g of cinnamoyl chitosan. The solution
was added to an aqueous suspension of BS-P (50 g L−1), and it
was stirred for 10 minutes for deposition of cinnamoyl chitosan single layer on
the whisker surface. Subsequently, UV irradiation (λ=254nm) to the
suspension was executed for 48 hours at room temperature with constant
stirring.


## 2.7. Microscopy

To
investigate the morphology and sizes of the capsules, transmission electron
microscopy (TEM) was done by using a Jeol JEM-2000FXII operated with an
acceleration voltage of200kV. No shadowing was applied. Scanning electron
microscopy (SEM) was also done by using a Hitachi S-2300.


## 2.8. Evaluation of the Core/Shell Ratio

Compositions
(core/shell ratio) of the capsules were evaluated by volumetric analysis. A
finite mass of the capsule was immersed in hydrochloric acid with stirring
until the core material was completely dissolved. Hydrochloric acid and water
in the obtained solution were evaporated and residual precipitate was redissolved
in deionized water. Calcium ion content in this solution was determined by
chelatometric titration, and the ratio of the mass of the shell to that of the whole
capsulems/mt was evaluated.


## 3. Results and Discussion

TEM observations confirmed
that thin shells of chitosan/chitosansulfate are formed by the layer-by-layer
technique. Figures1, 2, and 3 show TEM images for the core/shell and hollow
capsules prepared by the layer-by-layer deposition. For core/shell particles, thin
outer shells are discernible around the dark core part. Also, formation of
hollow capsules is clearly seen, confirming that the deposition of the outer
layer occurred. It is also noticed that the surface of the layer seems to be rather rough and ragged, and that
the layer contains some agglomerates. These features are more clearly seen for
the hollow samples. Occasionally for some of the core/shell
particles, the core part was lost resulting in partially or totally hollow
particles as shown in Figures 1(d), 2(c) and 2(d). This phenomenon was
observed more frequently for samples with higher m. This may be due to slight
solubility of the calcium carbonate in the acidic aqueous solvent (acetic acid aqueous
solution) that was used in the deposition process.Transmission electron micrographs for
capsules prepared from the higherMw  chitosan/chitosansulfate by the layer-by-layer deposition method. (a) Core/shell capsules with m=4, (b) hollow capsules with m=4, (c)-(d) core/shell capsules with m=14, and (e) hollow capsules with m=14.
(a)(b)(c)(d)(e)Transmission electron micrographs for capsules
prepared from the lowerMw chitosan/chitosansulfate by the layer-by-layer deposition method. (a)
Core/shell capsules with m=4, (b)  m=8, and (c)-(d) m=12.
(a)(b)(c)(d)Transmission electron micrographs for
core/shell capsules of (a)m=4 with
the higher Mw, (b)  m=14 with the higher Mw, (c)  m=4 with the lower Mw,
and (d) m=12  with the lower Mw.
(a)(b)(c)(d)Hollow particles were
successfully obtained by the treatment with hydrochloric acid only for the
higher molecular weight(Mw)
chitosan/chitosansulfate (1.09×106gmol−1). The shell prepared from the lower Mw chitosan/chitosansulfate (9.7×104gmol−1) was
probably lost by dissolution into the hydrochloric acid aqueous solution during
the core-removing process. However, the unintentional hollow capsules mentioned
above (yielded without the hydrochloric acid treatment) were partially obtained
even for the lower Mw material.Table1 shows the layer
thickness l and diameter D of the obtained core/shell and hollow
capsules. The values presented in this table were obtained by averaging over a
few hundreds of sampling points in TEM micrographs obtained with higher
magnifications as typically shown in Figure 3. The standard deviations for l are fairly large: this is due to the ragged feature of the shell. On the other hand, the large standard deviations
for D are due to the original size
distribution of BS-P as we reported previously [5].Table 1
Shell thicknessl, diameter D, and
core/shell composition of the capsules prepared by the layer-by-layer deposition
method.
Mw(1)ml (nm)σ1(2) (nm)D (nm)σD(3) (nm)ms/mt(4)Core/shellH43012397540.09HollowH4424409132—Core/shellH8489320550.11HollowH834832855—Core/shellH10631432357—HollowH10401334548—Core/shellH146912380390.16HollowH14511840864—Core/shellL4144——0.08Core/shellL8186——0.07Core/shellL12228——0.10(1)L:Mw=9.7×104 g mol−1, H:Mw=1.09×106 g mol−1.(2)Standard deviation for l.(3)Standard deviation for D.(4)Ratio of the mass of the shell to that of the whole capsule evaluated by
chelatometric titration.The layer thicknessl increases
with increasing the total number of depositions (deposited layers) m. However, l is not directly proportional to m, and an expression l=md+a(a>0) describes the data well.
This means that the deposited amount in the first deposition cycle is greater than
that in the later cycles. The deposition mechanism onto the surface of BS-P may
be different from that of the succeeding deposition, reflecting the different
chemistry of the surface from that for the deposited chitosan/chitosansulfate
surface. Linear regression analysis revealed average shell thicknesses per
layer to be d=4.0 and 1.0 nm for
the higher and lower molecular weights of the original chitosan, respectively. These values are much
higher than the thickness of a chitosan monolayer (a few angstroms). These
results suggest that the deposition does not occur by
a homogeneous monolayer-by-monolayer mechanism and that the opposite charges in the deposited
layer are not distributed uniformly, which may lead to some aggregation in the
succeeding deposition process. Also, the estimated values of d suggest that the aggregation during
the deposition occurs to less extent for the lower Mw material than for the higher one.The larged values seem to
be consistent with the TEM observation showing some agglomerates in the
deposited layer. Our previous circular dichroism study revealed that the
chitosan chains are rather rigid in aqueous acidic solution, taking a helical conformation
[18], which might prevent to form a homogeneous flat layer with uniformly
distributed charges upon depositing. The helical conformation may have tendency
to form an ordered structure, which also promotes the molecular aggregation
[19].The values of the core/shell compositionms/mt in the capsules are qualitatively consistent with the shell thickness (Table 1). However, thickness calculated from the obtained ms/mt values is much greater than the d values evaluated from the TEM observation, for example, for the higher Mw sample with m=14, the calculated thickness is 130 nm. This is probably due to residual chitosan and chitosansulfate that was not
deposited on the whisker particles. It seems to be rather difficult to remove
such excess polymer impurities by the present preparation method.From the above results, we conclude that the present deposition
mechanism is far from an ideal process of a monolayer-by-monolayer accumulation
caused exclusively by non-bonded interactions between the oppositely charged
sites. Nevertheless, the layer-by-layer method was revealed to be available to
fabricate capsules possessing chitosan/chitosansulfate shell.Slightly larger
apparent diametersD were observed for
the hollow samples than for the core/shell ones as shown in Table 1. This may
be due to deformation of the shell on removing the core. If the hollow shell
collapsed into a completely flat geometry, the observed width (apparent
diameter) would be greater than the diameter of the original core/shell
particle by a factor of π/2≅1.57. However, Table 1 shows much less increase
in D for the hollow particles,
suggesting that they are not completely flat. This can be confirmed by SEM
images presented in Figure 4. Such appearance of the hollow particles indicates
that the shell has some rigidity to retain the original shape of core/shell
capsule to some extent in contrast to the hollow shells composed of other
synthetic polyelectrolytes such as poly(sodium 4-styrenesulfonate) (PSS) and
poly(allylamine hydrochloride) (PAH) system, of which the appearance looks like
a deflated balloon with wrinkles [10, 11]. The rigid feature of this chitosan/chitosansulfate
shell may be originated from the inherent rigidity of the polymer chain, that
is, a polysaccharide chitosan has rather high chain rigidity. In addition, once the chitosan and
chitosansulfate chains are deposited on the surface, no swelling with the
medium may occur, thus, the vitrification of shell immediately occurs after the
deposition, resulting in rigidity of the shell.Scanning electron micrographs for hollow
capsules prepared from the lowerMw chitosan/chitosansulfate by the layer-by-layer deposition method (m=12). These hollow particles were unintentionally obtained without the
treatment with hydrochloric acid (probably the core was accidentally dissolved
away during the deposition processes).
(a)(b)Another cause for the less
increase inD on removing the core could
be shrinkage of the whole capsule. Significant shrinkage has been reported to
occur for the polymer shells deposited to the BS-P whisker surface via emulsion
polymerization [5]. This is associated with a thickening of the shell. For the
present capsules with chitosan/chitosansulfate shells, however, shell
thickening is not observed clearly in Table 1. It is likely that no significant
shrinkage occurs for the present capsules. The absence of shrinkage may be
partially due to the rigidity of the shell as mentioned above. There may be an
essential difference in the deposition mechanism between the seed emulsion
polymerization [5] and the present layer-by-layer method: in the former, some
configurational instability is frozen in during polymerization-vitrification
process and relaxation (stress release) occurs when the core is removed, while
in the latter process, stable configuration could be achieved to some extent in
the solution state prior to deposition, and therefore, no structural relaxation
occurs on removing the core.Figure5 shows transmission
electron micrographs for the capsules with the UV irradiated cinnamoyl chitosan
shells. The deposition was successful, and the mean layer thickness of the
core/shell capsules was 33±18 nm. This is even higher than
the thickness 30 nm of the shell prepared by the layer-by-later method with m=4 (Table 1). It is not clear whether this is due to an
error (standard deviation is 18 nm) or to an intrinsic deposition feature
characteristic to cinnamoyl chitosan. Unfortunately, no hollow capsules were obtained
even for the higher Mw material.
This may be due to insufficient degree of crosslink of the shell, that is, the
crosslinking photoreaction did not occur enough to stabilize the shell structure,
and the cinnamoyl chitosan was probably dissolved away in the hydrochloric acid
in the core-removing process.Figure 5
Transmission electron micrograph for shell crosslinked
capsules prepared by UV irradiation after deposition of cinnamoyl chitosan. The
core was not removed.In an acetic acid
aqueous solution(1.0%v/v) of cinnamoyl chitosan, the photodimerization
reaction was confirmed by a significant decrease in UV absorption at 280 nm
(characteristic to cinnamoyl group) and by an IR absorption band at 1458 cm−1 (cyclobutane ring) after UV irradiation (λ=254nm) for 24 hours
at room temperature. However, for the
deposited shell, only a slight signal was observed at 1458 cm−1,
indicating that the reaction could occur to less extent than in the solution
state. Thus, it may be difficult to yield a high degree of crosslink by the
present photoreaction in the solid shell. Once the cinnamoyl chitosan is
deposited on the whisker, its conformational motion is frozen, and photodimerization
reaction occurs only for the pairs of cinnamoyl groups which accidentally take
conformations suitable for the reaction. In addition, the reaction contains
intrachain reactions, which results in no crosslink.


## 4. Conclusions

We
have successfully prepared core/shell nanocapsules possessing biocompatible
outer shells of chitosan/chitosansulfate by the layer-by-layer deposition method.
Hollow capsules were obtained only for the higherMw chitosan/chitosansulfate shells. Electron microscopy
revealed that the thickness of the shell l increases almost linearly with increasing m though it is not directly proportional to m,
and that the thickness per layer is greater than that of the chitosan
monolayer. These results as well as the ragged surface morphology of the
deposited layers with agglomerates
suggest that the deposition mechanism is not exactly an ideal
monolayer-by-monolayer process. The rigidity of chitosan chain
may be responsible for the above feature of deposition. Nevertheless, our
results have demonstrated a great possibility to fabricate nanocapsules
possessing outer shells of rigid polysaccharides such as chitosan/chitosan
derivatives by the layer-by-layer deposition method. The photo-crosslinking of
cinnamoyl chitosan shell by UV irradiation was not enough to stabilize the
shell structure. An alternative method using interfacial reactions by
crosslinking monomers such as diisocyanate compounds [4] may be preferable to
achieve higher degree of crosslink.



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*Source: 185632-2008-03-16.xml*