International Journal of Pharmaceutics 232 (2002) 225–234
Alginate/chitosan particulate systems for sodium diclofenac
release
M.L. Gonza´lez-Rodrı´guez a,*, M.A. Holgado a, C. Sa´nchez-Lafuente a,
A.M. Rabasco a, A. Fini b
a Departamento de Farmacia y Tecnologı´a Farmacace´utica, Facultad de Farmacia, C/Profesor Garcia Gonzalez s/n,
Uni�ersidad de Se�illa, 41012 Se�illa, Spain
b Istituto di Scienze Chimiche, Uni�ersita` di Bologna, 40127 Bologna, Italy
Received 25 June 2001; received in revised form 8 October 2001; accepted 11 October 2001
Abstract
Alginate/chitosan particles were prepared by ionic gelation (Ca2+ and Al3+) for the sodium diclofenac release. The
systems were characterized by electron microscopy and differential scanning calorimetry. The ability to release the
active substance was examined as a function of some technological parameters and pH of dissolution medium. The
release of sodium diclofenac is prevented at acidic pH, while is complete in a few minutes when pH is raised up to
6.4 and 7.2. The alginate/chitosan ratio and the nature of the gelifying cation allow a control of the release rate of
the drug. The release mechanism was briefly discussed. © 2002 Elsevier Science B.V. All rights reserved.
Keywords: Alginate/chitosan beads; Ca2+ and Al3+ ions; Sodium diclofenac; Alginate/chitosan ratio; Dissolution profiles
www.elsevier.com/locate/ijpharm
1. Introduction
The use of biodegradable polymeric carriers for
the drug delivery systems has gained a wide interest,
mainly for their biocompatibility and, among the
microparticulate systems, microspheres show a spe-
cial importance for providing local (Lorenzo-
Lamosa et al., 1998; Hari et al., 1996) as well as
temporal controlled release of the drug (Donbrow,
1992). Different types of polymers are encountered
in the literature used to this purpose. In the case
of the release of diclofenac are available: poly-3-
caprolactone (Alonso et al., 2000), poly-/vinylalco-
hol (Chawla et al., 2000), poly-lactide-co-glycolide
(Tuncay et al., 2000a,b). Also natural polymers
found their application in this field: albumin (Tun-
cay et al., 2000a,b), alginate (Gursoy and Cevik,
2000; Acikgoz et al., 1995), carboxymethyl-cellu-
lose (Arica et al., 1996), chitosan (Lim et al., 1997;
McLaughlin et al., 1998). The preparation and
characterization of the samples are quite similar in
most cases.
The anti-inflammatory drug (as sodium salt) is
dissolved in an aqueous solution containing the
soluble polymer and, in the case of carboxylate-
containing polymers (alginate, carboxymethyl
cellulose), the formation of microspheres was
* Corresponding author.
E-mail address: rabasco@fafar.us.es (A.M. Rabasco).
0378-5173/02/$ - see front matter © 2002 Elsevier Science B.V. All rights reserved.
PII: S0378 -5173 (01 )00915 -2
M.L. Gonza´lez-Rodrı´guez et al. / International Journal of Pharmaceutics 232 (2002) 225–234226
obtained by the addition of Ca2+ or Al3+: the
hydrophilic colloids interact with metal ions to
form crosslinked insoluble complexes, that precip-
itate incorporating the drug.
In this paper, we prepared microspheres of
alginate containing sodium diclofenac and exam-
ined the different influence of Ca2+ or Al3+ ions
on the microsphere morphology and the influence
of different amounts of chitosan on the release of
diclofenac. Among polyanionic polymers alginate
has been widely studied and applied for its possi-
bility to modulate the release, according to the
properties of its carboxyl groups as well as its
biodegradability and absence of toxicity. Also chi-
tosan finds wide applications in pharmaceutical
technology as tablet disintegrant, for the produc-
tion of controlled release solid dosage forms
(Miyazaky et al., 1994; Kas, 1997; Illum, 1998;
Sezer and Akbuga, 1999) or for improvement of
drug dissolution (Felt et al., 1998; Gupta and
Ravi Kumar, 2000).
Diclofenac is a suitable candidate for incorpo-
ration into microspheres (Hosny et al., 1998; Go-
hel and Amin, 1999) to minimize its adverse effect
after oral administration; in fact, alginate micro-
spheres containing diclofenac start to release the
drug after the pH of the environment increases
above 7 (Sabnis et al., 1997), by-passing the gas-
tric environment and avoid direct contact between
the drug and the gastric mucosa.
2. Materials and methods
2.1. Materials
The following materials were obtained from the
indicated suppliers and used as received: sodium
alginate (low viscosity; viscosity of 2% solution
25 °C, �250 cps), chitosan (Practical grade from
crab shell; Sigma, Barcelona, Spain). Calcium
chloride hexahydrate, aluminum chloride hexahy-
drate (Sigma, Barcelona, Spain), di-sodium hy-
drogen phosphate anhydrous, potassium
di-hydrogen phosphate, hydrochloride acid 35%
and acetic acid glacial 100% (Merck, Barcelona,
Spain), were of the highest purity level available.
Sodium diclofenac was of pharmaceutical degree
(Farchemia, Italy).
2.2. Preparation of microspheres
Sodium diclofenac (0.25% w/v) was added to
aqueous solutions of sodium alginate (1.2% w/v)
and stirred up to complete dissolution. This solu-
tion was dropped using an hypodermic syringe
into a second solution, containing Ca2+ (or Al3+)
ions (1.3% w/v) and chitosan, previously dissolved
in acetic solution (0.5% v/v).
Microspheres formed immediately and were left
into the original solution for 24 h to ensure
internal gelification also. Then they were filtered,
washed and dried at room temperature.
The whole preparation was carried out at room
temperature. Table 1 lists the formulations
prepared.
2.3. Loading and yield of the process
To evaluate the amount of the drug inside the
microspheres, an indirect method was used (Fer-
na´ndez-Herva´s et al., 1998). Aliquots from the
filtered solutions remaining after removal of the
beads were assayed spectrophotometrically at 276
nm. The amount of sodium diclofenac entrapped
was calculated from the difference between the
total amount of drug added and the sodium di-
clofenac found in the filtered solution.
2.4. Thermal analysis
Thermograms were obtained using differential
scanning calorimeter (Mettler FP89HT), to evalu-
ate the state of the active agent inside the micro-
sphere of possible degradation.
All the samples were run at a scanning rate of
10 °C/min from 30 to 300 °C.
2.5. Scanning electron microscopy
Micrographs of the external surface were ob-
tained by SEM (Philips XL 30), depositing on the
sample a thin film of carbon. The shape and size
of the beads were determined using an image
analysis system (Scion Image) which was con-
nected to the microscope.
The following parameters were selected to char-
acterize the sodium diclofenac microspheres:
M.L. Gonza´lez-Rodrı´guez et al. / International Journal of Pharmaceutics 232 (2002) 225–234 227
� area (A), it is the selected surface;
� perimeter/length (l), it is the length around the
outside of the selection, or line length for line
selections;
� diameter of the equivalent area circle (ECD),
this is an equivalent diameter that is calculated
as follows:
Dcirl=2
�A
�
where A is the area of the closed boundary of
the particle.
� shape factor (s), this parameter is used to
measure object complexity, namely contour
complexity. The more the variation of the con-
tour, the more the shape factor parameter is
elevated. The value of s is determined as
follows:
s=
L2
4�A
where L and A are the perimeter and the area
of the closed boundary of the particle,
respectively.
2.6. Release tests
Dissolution assays were carried out in triplicate
for 6 h at 37�0.5 °C. For the first 2 h, pH of the
dissolution medium was buffered at pH 1.2 (HCl/
NaCl, to mimic the gastric district). pH was then
raised up to 6.6 (phosphate buffer) and main-
tained at this value for further 2 h. Finally, a little
higher pH (7.4) was used up to the end of the
experiment. The tests were performed into an
apparatus as described in USP 23 (Turu Grau,
mod. D-6).
At prefixed time (15 min), 3 ml of solution were
withdrawn and spectrophotometrically assayed
for the diclofenac content (�=276 nm) (Hitachi,
mod. U-2000).
Dissolution tests were carried out taking into
account the following parameters: type of gelify-
ing ion and alginate/chitosan ratio.
3. Results and discussion
Side effects, mainly at the gastric level, are well
known, following oral administration of an
NSAID. Therefore the efforts of many researchers
have been concerned to solve these problems,
through a variety of techniques of protection of
the gastric mucosa or alternatively to prevent the
NSAID release in this district. In this paper we
evaluate the potential utility of natural materials,
such as alginate and chitosan in inhibiting sodium
diclofenac release in the gastric environment. And
since among the microparticulate systems, micro-
spheres have a special interest as carriers for
NSAID, mainly to extend the duration period of
the dosage form, we aimed to investigate the
Table 1
Composition of the different formulations used for the preparation of sodium diclofenac microspheres
Alginate/chitosanSodium alginate (%)Chitosan (%)CationBatch
L1 0.1Ca2+ 1.2 12
0.1Ca2+L2 151.5
Ca2+ 0.2L3 1.2 6
L4 Ca2+ 7.51.50.2
1.2 1.2Ca2+ 1L5
Al3+ 0.1 1.2 12L6
L7 1.5Al3+ 150.1
L8 1.2Al3+ 60.2
7.51.50.2L9 Al3+
1.2 1L10 Al3+ 1.2
Al3+ 0.6L11 1.2 2
31.20.4L12 Al3+
Al3+ 0 1.2 0L13
M.L. Gonza´lez-Rodrı´guez et al. / International Journal of Pharmaceutics 232 (2002) 225–234228
Table 2
Encapsulation efficiency (EE) and shape parameters of the different formulations; ECD, equivalent circular diameter
Area (mm2) Perimeter (mm)Batch Shape factorEE (%) ECD (mm)
99.65�5.28L1 11.35�2.10 10.82�3.03 0.825�0.03 3.80�0.52
11.98�3.58 10.94�2.8599.63�7.99 0.798�0.05L2 3.91�0.99
99.66�7.21L3 9.30�5.81 10.21�4.11 0.897�0.01 3.44�1.01
99.63�10.02L4 10.51�2.78 10.54�4.08 0.847�0.05 3.66�0.21
8.70�3.13 10.10�3.6698.82�5.21 0.938�0.03L5 3.33�0.09
99.87�4.11L6 14.37�9.54 11.62�3.85 0.752�0.01 4.28�0.24
14.41�4.51 11.62�2.10L7 0.749�0.0299.85�10.32 4.28�0.54
12.99�5.41 11.40�5.9299.68�7.41 0.801�0.05L8 4.07�0.65
99.72�3.33L9 13.14�5.99 11.56�5.21 0.813�0.02 4.09�0.54
10.01�2.10 9.10�2.45 0.662�0.01L10 3.57�0.8199.07�6.52
10.22�5.47 9.09�2.6699.3�11.45 0.648�0.03L11 3.61�0.77
10.30�2.33 9.32�1.48 0.675�0.05L12 3.62�0.6399.49�7.41
10.04�4.10 9.20�3.21 0.675�0.0499.76�5.23 3.58�0.41L13
possible applicability of chitosan treated alginate
beads as a controlled release system for soluble
salts. We prepared microspheres containing
sodium diclofenac starting from natural polysac-
charides by ionotropic gelation method and exam-
ined the effects of various factors
(alginate/chitosan ratio, electrolyte concentration
and nature of bead).
3.1. Microsphere characterization
3.1.1. Morphology
Alginate/chitosan microspheres containing
sodium diclofenac were evaluated for particle size,
yield and encapsulation efficacy and surface mor-
phology. The ionic gelation method gave beads
with a high diameter ranging from 2 to 4 mm
(Table 2). The drug encapsulation yield was more
than 98% in all the cases, and the efficacy was
neither affected by the alginate amount nor the
crosslinking ion used. So, this method is useful to
encapsulate ionic drugs with a high water
solubility.
Although the encapsulation of the drug was
approximately 100% in all the formulations, the
use of different ions, such as calcium or alu-
minum, determines the mechanical properties of
alginate gels and the egg-box structure of the
beads, which gives it the spherical morphology.
Fig. 1 shows a microsphere prepared adding
Ca2+, exhibiting acceptable sphericity and a nota-
ble surface porosity, with a shape factor greater
than 0.80 in all the cases. This morphology was
found independent of the starting composition,
provided that Ca2+ ions were the gelifying agent.
Due to the adhesive properties of chitosan, micro-
spheres tend to agglomerate (Lim et al., 1997;
Ganza-Gonza´lez et al., 1999; Murata et al., 1999).
The aspect and morphology of the particulates
prepared with Al3+ ions is different: no formula-
tion enabled the formation of a spherical mor-
phology; on the contrary, the particles are
flattened, disk-shaped with a collapsed center. The
surface appears smooth and little porous (Fig. 2),
with a shape factor less than 0.80. The trivalent
ions cause more points of aggregation between
two contiguous alginate chains, binding them so
strictly and quickly that, as a consequence, there
is no time to get spherical forms, during their
formation.
Fig. 3 recalls the feature of a drop touching a
water surface, suggesting that instantaneous gelifi-
cation blocks the situation of the very first contact
between the two solutions, the one containing
alginate and the other one the inorganic ions.
3.1.2. Differential scanning calorimetry
Thermograms of both types of microspheres do
not offer clear peaks of identification (Figs. 4 and
5) (Ford and Timmins, 1989).
Calcium chloride shows two endotherm peaks
in the temperature range 180–220 °C; while
M.L. Gonza´lez-Rodrı´guez et al. / International Journal of Pharmaceutics 232 (2002) 225–234 229
Fig. 1. Scanning electron micrographs of sodium diclofenac microspheres formulated with calcium chloride.
sodium alginate decomposes at about 240 °C
with a broad exotherm. Peaks of the single com-
ponents are not visible when combined into a
microsphere, whose thermogram shows only a
broad and small endotherm, probably related to
dehydration, present at a temperature about
120 °C. The same applies to Al-microspheres: in
this case the endotherm peak present in the ther-
mogram of AlCl3·6H2O is also absent. This can be
interpreted as follows: that a chemical reaction
occurred in solution, modifying the starting
reagents and that a strong interaction is present
among all the compounds inside the formulation;
this fact additionally acts as a stabilizer, since
degradation endotherms both for alginate and
sodium diclofenac are absent at the highest tem-
peratures of the thermogram.
3.1.3. Effect of the ion on the release process
For the preparation of chitosan treated alginate
beads containing sodium diclofenac, we dissolved
sodium diclofenac in the aqueous solution of
sodium alginate. The addition of the divalent (or
trivalent) ions produced (Kondo, 1979) a partial
neutralization of carboxylate groups present on
the alginate chain, forming an insoluble (but per-
meable) transitory thin gelatinous film:
2nNaAlginate + nCa2+
� nCa(Alginate)2 + 2nNa+
Ionic gelation is the result of the formation of
an ‘egg box’ between facing units of two different
chains and depends on the inorganic ion/alginate
ratio. When the solution has a polycationic chain
(e.g. protonated chitosan) this acts as a crosslink-
ing thus improving the microsphere hardness.
Fig. 2. Scanning electron micrographs of sodium diclofenac
microspheres formulated with aluminum chloride.
M.L. Gonza´lez-Rodrı´guez et al. / International Journal of Pharmaceutics 232 (2002) 225–234230
Fig. 3. Scanning electron micrographs of the structure formed from a drop touching a water surface, during the microsphere
fabrication process. These sodium diclofenac microspheres were formulated with aluminum chloride.
On the addition of inorganic cations into the
solution containing anions (alginate and diclofe-
nac) two different reactions can start: the gelation
of alginate chain and the (possible) precipitation
of an insoluble diclofenac salt. However, the pre-
cipitation of diclofenac as an insoluble salt should
make its recovery difficult during the dissolution
test, even at varying pH. On the contrary, when
the experimental conditions are favorable, diclofe-
nac is released from the microspheres and appears
in solution. Therefore it can be assumed that
diclofenac is still present in a soluble form inside
the microsphere, as a sodium salt or as a complex
with the polycation chitosan readily available. In
fact, since inorganic ions were added dropwise,
they were always present in the solution in highly
deficit concentrations with respect to the alginate
anion units. At the composition used, alginate
carboxylate groups strongly exceed those present
in diclofenac; furthermore, the polyanion rapidly
reacts with each cation blocking it into a thermo-
dynamically very stable ‘egg box’. This capture of
inorganic ions prevents competition towards di-
clofenac anion, which could not react with Ca2+
or Al3+ and form insoluble products. The mutual
neutralization between oppositely charged algi-
nate and chitosan decreases the solubility of the
whole system; the same could occur in the forma-
tion of a complex between chitosan cations and
diclofenac anions. Therefore the driving force for
the microsphere formation is a decrease of solu-
bility. This mechanism explains the high efficiency
of the microsphere formation in depleting diclofe-
nac from the solution, irrespective of the different
experimental conditions examined. The precipitate
inside a microsphere has a complex composition
and structure.
The alginate/chitosan microsphere represent an
efficient system for controlling the release of di-
clofenac (Yotsuyanagi et al., 1991). At acidic pH,
the release is low for 2 h: the amount of the
released diclofenac did not exceed 6–7%, indepen-
dent of the microsphere size. At acidic pH, algi-
nate is protonated into the insoluble form of the
alginic acid: this displays properties of swelling
that explains the low aliquot of the release. In this
case the release is hindered by chitosan: its posi-
tively charged groups strongly interact with algi-
M.L. Gonza´lez-Rodrı´guez et al. / International Journal of Pharmaceutics 232 (2002) 225–234 231
nate and diclofenac ions, both reducing swelling
and release and, possibly, the protonation of their
carboxylated groups. At pH 6.4 a rapid increase
of the release rate was observed up to 100%. The
deprotonation of the alginic acid causes the disin-
tegration of the microsphere systems and the com-
plete release of diclofenac as soluble ions.
At increasing pH, the increasing deprotonation
of chitosan weakens the extent of the interactions
inside the microsphere; moreover, diclofenac is
soluble as anions and can be brought into the
solution. The release is complete in a few minutes.
The role of the phosphate anions present in the
buffer of the dissolution medium in sequestering
the Ca2+ ions and taking away a factor for the
insolubility of alginate chains cannot be excluded.
This aspect should, however, play differently in
the presence of Ca2+ or Al3+ ions, even though
the release profiles overlap perfectly in both cases.
All the formulations examined show a dissolution
profile as that reported in Fig. 6.
3.1.4. Effect of alginate/chitosan ratio on the
release process
The case of the alginate/chitosan ratio is differ-
ent. The presence of chitosan increases the control
of the release from the microsphere, since, at
increasing concentration, it can form a network of
bondings between the two polymer chains.
This was expected, since at increasing chitosan
amount into the formulations, interactions be-
tween the two polymers should have been in-
Fig. 4. Thermograms corresponding to the sodium diclofenac microspheres with calcium chloride.
M.L. Gonza´lez-Rodrı´guez et al. / International Journal of Pharmaceutics 232 (2002) 225–234232
Fig. 5. Thermograms corresponding to the sodium diclofenac microspheres with aluminum chloride.
creased, forming a closer network, which should
decrease the diffusion of the drug outwards of the
bead. Therefore, to deeper examine this parameter
the alginate/chitosan ratio was changed to value
1, 2 and 3 w/w. In these cases, differences were
more evident. At the ratio 1:1, while Ca2+ offered
a dissolution profile as the previous ones, in the
case of Al3+ microspheres, the release is lowered
below 50%. This result confirms what was re-
ported in the literature that increasing chitosan
concentrations decr