Chem. Pharm. Bull. 48(4) 579—581 (2000)
The Controlled Release of a Drug from Biodegradable Chitosan Gel Beads
Kyoko KOFUJI,* Tomohiro ITO, Yoshifumi MURATA, and Susumu KAWASHIMA
Faculty of Pharmaceutical Sciences, Hokuriku University, Ho-3, Kanagawa-machi, Kanazawa 920–1181, Japan.
Received October 25, 1999; accepted January 11, 2000
Chitosan (CS) forms a gel in solutions with a pH above 12, and the gelation occurs at pH of about 9 in 10%
amino acid solutions. In this paper, we investigated the enzymatic degradation and the drug release pro?le of this
novel CS gel beads. The degradability of the CS gel beads was affected by the CS properties, e.g. the degree of
deacetylation. The release of prednisolone (PS), as a model drug, from the CS gel beads was sustained signi?-
cantly compared with the gel prepared with NaOH only. However, the release was not able to be sustained by the
increment of NaOH concentration in the solution employed for the preparation of CS gel beads. We also investi-
gated the control of drug release from CS gel beads by application of a complex formed between chondroitin sul-
fate (Cho) and CS. The release of PS from the CS gel beads treated with Cho was prolonged, and the release pat-
tern was not affected by the treatment time. The time to 50% drug release was about 5 min with PS powder,
about 200 min in CS gel beads with 10% glycine (Gly) (pH 9.0), and about 330 min in the CS gel beads with 10%
Gly (pH 9.0) treated with Cho. Thus CS gel beads appear promising as a vehicle for sustained drug delivery, and
the degradation of CS gel beads may be controlled by the degree of deacetylation of CS.
chitosan; chondroitin sulfate; gel; biodegradation; amino acid
The polysaccharide chitosan (CS) is known to be an excel-
chased from Wako Pure Chemical Industries (Japan). The other reagents
lent material for drug preparation. CS is a plentiful natural
were obtained from Wako Pure Chemical Industries (Japan) and Nacalai
Tesque Inc. (Japan).
biopolymer and is also non-toxic, biocompatible and
Preparation of CS Gel Beads
CS gel beads were prepared by the fol-
biodegradable.1—5) These properties are important for materi-
lowing method. CS (1% w/w) was dissolved in 0.1 M acetate buffer (pH 4.5).
als that are implanted in the body, because such materials
Then 1% PS was suspended to the CS solution. One gram of the suspension,
must avoid the host’s defense system during their long-term
theoretically containing 10 mg of drug, was dropped slowly into 20 ml of
contact with living structures.6) CS has been studied as a
10% glycine (Gly) solution using a pipette and left at room temperature for
25 min. Hydrogel beads were formed spontaneously. The dried gel beads
unique vehicle for the sustained delivery of drugs; in addi-
were obtained by drying at 37 °C for 24 h on a dish, before being held under
tion, the preparation of CS microspheres has been ex-
vacuum in a desiccator in the presence of P O . The diameter of dried gel
amined.7—9) We have been investigating the preparation of a
bead with 10% Gly (pH 9.0) was about 3 mm. It consisted of Gly (50.7%),
suitable vehicle, for example micro-gel beads, for intra-artic-
PS (12.3%), CS (12.6%). Gly was determined according to the method of
ular injection in rheumatoid arthritis to allow sustained intra-
Watanabe and Imai.12)
Preparation of CS–Cho Gel Beads
CS hydrogel beads prepared by the
articular drug delivery. Lu et al.10) described that CS could
method described above were soaked in 20 ml of 1% Cho solution (1—48
act on the epiphyseal cartilage and wound healing of articular
h). The gel beads were removed from the Cho solution and dried at 37 °C for
cartilage. Therefore, we considered CS might be able to uti-
24 h on a dish, before being held under vacuum in a desiccator in the pres-
lize as a wound healing of articular cartilage after accom-
ence of P O .
plishment of a role as a vehicle for drug. In the previous
Enzymatic Degradation of CS Gel Beads
Lysozyme solution (20
mg/ml in 0.1 M phosphate buffer, pH 7.2, containing 0.2 M sodium chloride)
study, we found that CS could form sphere gel at a pH of
was incubated at 37 °C. The dried gel beads were added to 10 ml of the solu-
about 9 in amino acid solutions despite usually CS formed a
tion in a 100-ml sample bottle and incubated at 37 °C. The beads were re-
gel in solutions with a pH above 12.11) We consider that the
trieved periodically and dried at 37 °C for 24 h, before being held under vac-
phenomenon is coacervation. Preparations made at a lower
uum in a desiccator in the presence of P O . The weight of each dried gel
bead preparation was measured. The extent of degradation was expressed as
pH are preferable in terms of their effects on the solubility or
the percentage of residual weight of the gel beads after soaking in the
stability of the drug contained in the gel beads and on the tis-
lysozyme solution. In this experiment, CS gel beads was also soaked in 0.1 M
sue into which they are injected. Steroids have been applied
phosphate buffer (pH 7.2) without lysozyme for 30 min to remove Gly, and
for intra-articular injection therapy in rheumatoid arthritis.
their dry weight was adopted as the weight of CS gel beads that did not de-
Under such conditions, it is dif?cult to achieve a sustained
grade at all.
Release of PS from the various types of CS gel beads
intra-articular drug level only on the basis of drug solubility.
into 0.1 M phosphate buffer (pH 7.2) was determined. The dried gel beads
During in vivo degradation, the drug release will be governed
corresponding to 1 g of hydrogel were added to 500 ml of the dissolution
by both diffusion and biodegradation of the matrix. There-
medium in a JP XIII dissolution test apparatus (paddle method, 100 rpm,
fore, in this study, we investigated the biodegradability and in
37 °C). A 4-ml aliquot of the solution was removed periodically for analysis
vitro drug release pro?les from this novel CS gel beads. Fur-
and replaced with 4 ml of the dissolution medium (pre-warmed to 37 °C) to
maintain a constant volume. The absorbance of each sample was determined
thermore, we attempted to control the drug release from the
on a Hitachi model 200-20 spectrophotometer at 246 nm. All the dissolution
CS beads by modifying the structure of gel matrix.
tests were performed in triplicate.
Results and Discussion
CS with various degrees of deacetylation [70% (7B), 80%
Enzymatic Degradation of CS Gel Beads
(8B)] was purchased from Katokichi Co. Ltd. (Japan). The molecular
mucopolysaccharides are degraded by enzymatic hydrolysis;
weights of the CS were 2210000 (7B) and 2140000 (8B).2) Prednisolone
(PS) was purchased from Nacalai Tesque Inc. (Japan). Egg-white lysozyme
the degradation of CS has been reported.1—6) In a previous
(activity 0.8 mg/mg) and chondroitin sulfate C sodium salt (Cho) were pur-
study, the viscosity of CS solutions became gradually lower
? To whom correspondence should be addressed.
© 2000 Pharmaceutical Society of Japan
Vol. 48, No. 4
Effect of Deacetylation of CS on the Enzymatic Degradation of CS
Effect of NaOH Addition to 10% Gly Solution on the Release of
PS from CS (7B) Gel Beads
Effect of Amino Acid Addition in the Preparing Solution on the
Release of PS from CS (7B) Gel Beads
Effect of Cho Treatment of CS Gel Beads on the Release of PS
in the presence of lysozyme. The degradation was affected by
within the beads was released gradually. However, in the case
the CS properties, e.g. the degree of deacetylation, and 100%
of the gel beads prepared using 1 N NaOH, the PS release
deacetylated CS was not degraded.11) The degradation of CS
was faster because the beads could not maintain a spherical
gel beads in 0.1 M phosphate buffer containing lysozyme was
shape. Also, as shown in Fig. 3, PS was released faster from
investigated. The weight of dried gel beads on CS (7B) began
CS gel beads prepared using the above 2 N NaOH solution,
to decrease after 3 d. However, on CS (8B), the loss of weight
despite the presence of 10% amino acid, because the gel
of the dried gel beads was slight (Fig. 1). Tomihata and
beads could not maintain a spherical shape in the process of
Ikada6) described that the in vitro and in vivo degradations of
removal from the solutions or drying. The PS release pattern
the CS ?lms, prepared by the solution casting method, oc-
was virtually unaffected by increasing the preparation time or
curred less rapidly as their degree of deacetylation became
changing the species of CS (7B, 8B). Thus, the sustained re-
higher, and the ?lms which were more than 73% deacety-
lease was not attributable to the insolubility of the drug, but
lated CS showed slower biodegradation. The degradability of
to its retention in the CS gel matrix.
the gel may differ with the gel preparation method. However,
Pro?le of PS Release from CS–Cho Gel Beads
the degradability of the CS gel beads prepared with 10%
ing the gel matrix by utilizing complex formation between
amino acid solutions (pH 9.0) was similar to the gel prepared
CS and other mucopolysaccharides, such as Cho or
with NaOH only or by the solution casting method. Thus it
hyaluronic acid (HA), may be more effective for controlling
may be possible to control the degradability of CS gel beads
the release of a drug because the amino group of CS may
by taking advantage of the differences in degradability of the
form an electrostatic complex with the carboxyl group of
various species of CS with different degrees of deacetylation.
Cho or HA.13,14) As shown Fig. 4, the release of PS from the
Retention of PS in CS Gel Beads
CS solutions sus-
CS–Cho gel beads, which had been soaked for 6 h in 1% Cho
pending less than 5% PS were able to maintain the spherical
solution, was inhibited. We concluded that the complex be-
shape of the gel beads. The amount of PS retained in the
tween CS and Cho was formed the surface of CS gel beads,
beads decreased according to the increase in the soaking time
because the pattern of drug release was not changed by
or the amount of NaOH in 10% Gly solution. PS was re-
changing the soaking time. The retention of PS in the gel
tained in the gel beads, which maintained a spherical shape,
beads decreased as the soaking time increased. However,
to about 85—100% of the theoretical total amount.
about 70% of the theoretical total amount of PS was retained
Pro?le of PS Release from CS Gel Beads
in the gel beads, even after the beads had been soaked for 6 h
lease of PS from CS gel beads prepared with 10% Gly (pH
in the Cho solution.
9.0) was observed. As shown in Fig. 2, all the PS retained
Thus CS gel beads appear promising as a vehicle for sus-
tained drug delivery, and the degradation of CS gel beads
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