International Journal of Biological Macromolecules 72 (2015) 47–53

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International Journal of Biological Macromolecules

journal homepage: www.elsevier.com/locate/ijbiomac

Metal ion-induced alginate–locust bean gum IPN microspheres for

sustained oral delivery of aceclofenac
Sougata Jana ∗ , Arijit Gandhi, Subrata Sheet, Kalyan Kumar Sen
Department of Pharmaceutics, Gupta College of Technological Sciences, Asansol 713301, W.B., India

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

Article history: The alginate microspheres represent a useful tool for sustained oral delivery of drugs but exhibit sev-
Received 5 June 2014 eral problems associated with the stability and rapid release of drugs at higher pH values. To overcome
Received in revised form 25 July 2014 these drawbacks, alginate–locust bean gum (LBG) interpenetrating microspheres were prepared by cal-
Accepted 31 July 2014
cium ion (Ca+2 ) induced ionotropic gelation technique for prolonged release of aceclofenac. The drug
Available online 9 August 2014
entrapment efficiency of these microspheres was found to be 59–93%. The microspheres lied in the size
range of 406–684 ␮m. Scanning electron microscopy revealed spherical shape of the microspheres. No
Keywords:
drug–polymer interaction was evident after infrared spectroscopy analysis. The microspheres provided
Alginate
Locust bean gum
sustained release of aceclofenac in phosphate buffer solution (pH 6.8) over a period of 8 h. The drug release
Microspheres data were fitted into the Korsmeyer–Peppas model and the drug release was found to follow anomalous
Sustained drug delivery (non-Fickian) diffusion mechanism. Pharmacodynamic study of the microspheres showed a prolonged
Anti-inflammatory activity anti-inflammatory activity in carrageenan-induced rat paw model following oral administration.
© 2014 Published by Elsevier B.V.

1. Introduction their extensive application as food additives and their recognized

Microspheres refer to the micro-particulate polymer-based
carrier systems for drug delivery applications. They offer advan-
tages such as limited fluctuation of drug–plasma profile within a
therapeutic range, reduction in side effects, decreased dosing fre-
quency and improved patient compliance [1,2]. Over the past few
decades, several research papers have been published on micro-
spheres, composed of naturally occurring biodegradable polymers
[3]. Among naturally occurring biodegradable polymers, sodium
alginate and locust bean gum (LBG) are widely used polymer can-
didates for the designing and development of various drug delivery
systems. Sodium alginate, a linear anionic polysaccharide obtained
from brown algae is known to biodegradable and non-toxic [4].
Sodium alginate, the sodium salt of alginic acid, belongs to a
family of linear copolymer composed of two monomeric units, ␤-
d-mannuronic acid residue, ␣-l-guluronic acid residue and regions
of interspersed both the residues [5,6]. Alginates have been used
as matrix forming material in the design of various drug deliv-
ery systems to achieve sustained drug release over a prolonged
period due to its hydro-gel forming properties [7]. On the other
hand, locust bean gum is also derived from the endosperm of the
seeds of Ceretonia siliqua Linn belonging to the family Fabaceae [8].
Locust bean gum has a wide potential in drug formulation due to
∗ Corresponding author. Tel.: +91 9434896683.
E-mail address: janapharmacy@rediffmail.com (S. Jana).
lack of toxicity. It can be tailored made to suit the demands of appli-
cants in both the pharmaceutical and biomedical areas. This group
of polymers possesses a number of characteristics that makes it
useful as a formulation aid, both as a conventional excipient and
more specifically as a tool in polymeric controlled drug delivery. It
consists mainly of a neutral galactomanan polymer made up of 1,
4-linked d-mannopyronosyl units and every fourth or fifth chain
unit is substituted on C6 with a d-galactopyranosyl unit [9].
Sodium alginate have ability to undergo ionotropic gelation
in aqueous solution in presence of multivalent cations like Ca2+ ,
Zn2+ , Pb2+ , Cd2+ , Al3+ , etc. [10]. On the other hand, LBG also able
to undergo ionotropic gelation in aqueous solution in presence of
cations like Al3+ [11,12]. The mechanical stability of the ionotrop-
ically cross-linked gel systems is provided by multivalent cations.
In literatures, the combination of alginate and LBG as drug delivery
carrier was not found. Though drug loaded alginate microspheres
represent a useful tool for sustained oral drug delivery but show
several problems, mainly related to the stability, and rapid drug
release at higher pH values. To overcome these drawbacks, the
present investigation was undertaken to design IPN microspheres
using a dual combination of alginate and LBG for sustained drug
delivery.
Aceclofenac is chemically 2-[(2 ,6 -dichlorophenyl) amino]
phenylacetoxyacetic acid, as a non-steroidal anti-inflammatory
drug (NSAID) with short half-life (4 h) indicated for the symp-
tomatic treatment of pain and inflammation [13]. It is also used in

http://dx.doi.org/10.1016/j.ijbiomac.2014.07.054
0141-8130/© 2014 Published by Elsevier B.V.
48 S. Jana et al. / International Journal of Biological Macromolecules 72 (2015) 47–53

Table 1 weighing balance (Dhona 160 D, Dhona Instrument Pvt. Ltd.,

The percentage of polymers, drug (aceclofenac) and cross-linker for the preparation
India.) and the theoretical weight was calculated by taking into
of different aceclofenac-loaded microspheres.
consideration the weight of the drug and polymer employed
Formulation Sodium alginate LBG (% Aceclofenac CaCl2 (% during the preparation of microspheres. The percentage yields
code (% w/v) w/v) (% w/v) w/v)
of these microspheres were calculated using this following
F1 1 1 1 2 formula:
F2 1 1 1 4
F3 1 2 1 2 Percentage yield = (Weight of microspheres recovered/
F4 1 2 1 4
F5 1 3 1 2 Total weight of drug and polymers) × 100
F6 1 3 1 4
F7 2 1 1 2
F8 2 1 1 4
F9 3 1 1 2
F10 3 1 1 4 2.4. Estimation of drug entrapment efficiency
F11 2 2 1 2
F12 2 2 1 4
Accurately weighed 100 mg of prepared aceclofenac-loaded
microspheres from each batch were taken separately and were
the treatment of arthritis, osteoarthritis, rheumatoid arthritis and placed in 500 ml of phosphate buffer, pH 6.8, and kept it
ankylosing spondylitis [14]. Aceclofenac is reported to produce side overnight followed by sonication for 15 min in a sonicator
effects like gastric irritation, ulcer, particularly diarrhoea, nausea, (Frontline sonicator, FS-600, Frontline electronics and machinery
abdominal pain and flatulence, etc. as result of prolong treatment Pvt. Ltd., India). The polymer debris formed after disintegra-
[15,16]. Due to its short half-life, its recommended dose is consid- tion of beads was removed filtering through Whatman® filter
ered as 200 mg daily in divided doses. To reduce dosing frequency paper (No.40). The drug content in the filtrate was determined
and adverse effects during prolong treatment, sustained release using a UV–vis spectrophotometer (Shimadzu, Japan) by mea-
dosage of aceclofenac to deliver aceclofenac at a slow release rate suring absorbance at max of 274 nm. The drug entrapment
over an extended period of time is essential. Therefore, aceclofenac efficiency of microspheres was calculated using this following
was used as model drug in this investigation. formula:

2. Materials and methods Drug entrapment efficiency(%)

2.1. Materials = (Actual drug content in microspheres/

Aceclofenac was received as a gift sample from Cipla Pharma-
ceuticals, India; Sodium alginate was commercially purchased from
Merck Specialities Pvt. Ltd., India; LBG was procured from Hi-Media,
India; Calcium chloride was purchased from S.D. Fine chemicals Ltd,
India; all other reagents and chemicals used in this study were of
analytical grade.
2.2. Preparation of microspheres
The aceclofenac-loaded microspheres made of alginate–LBG
were prepared through interfacial ionotropic gelation technique.
Briefly, required amounts of sodium alginate and LBG were dis-
solved in deionized water (100 ml) using magnetic stirring of
300 rpm for 30 min separately. Both the polymer solution was
mixed with continuous magnetic stirring of 300 rpm for 30 min
again. Afterwards, aceclofenac was added to the mixture gels
of sodium alginate and LBG. Aceclofenac containing polymeric
gels were stirred using magnetic stirring of 300 rpm until they
became bubble free. The prepared homogeneous bubble-free drug-
polymeric solutions were extruded drop wise into counter-ion
solutions using a 25 ml hypodermic syringe (1 mm diameter) with
constant stirring. The counter-ion solutions contain different con-
centrations of calcium chloride. Added droplets were retained in
the counter-ion solutions for 5 min to complete the curing reaction
and to produce rigid microspheres. The wet microspheres were col-
lected by decantation, and washed two times with distilled water
and dried in room temperature for overnight. The dried micro-
spheres containing aceclofenac were stored in a desiccator until
used. Different microsphere formulations along with percentage
of polymers, drug (aceclofenac) and cross-linker are enlisted in
Table 1.
2.3. Determination of percentage yield
Theoretical drug content in microspheres) × 100
2.5. Particle size determination
The size of the prepared microspheres was measured by using
digital slide callipers (CD-6 CS, Mitutoyo Corporation, Japan). Hun-
dred particles were taken and inserted in between the space of two
metallic plates. Diameters of resultant particles were displayed in
the digital screen of the previously calibrated equipment.
2.6. Surface morphology analysis
The surface morphology of this formulated microsphere was
analyzed by scanning electron microscope (SEM) (JEOL, JSM-6360,
Japan). Microsphere were gold coated by mounted on a brass stub
using double-sided adhesive tape and under vacuum in an ion sput-
ter with a thin layer of gold (3–5 nm) for 75 s and at 15 kV to make
them electrically conductive and their morphology was examined.
2.7. Swelling behaviour measurement
Swelling behaviour measurements of prepared microspheres
were carried out in 0.1 N HCL (pH 1.2) and phosphate buffer of
pH 6.8. 100 mg microspheres were placed in vessels of dissolution
apparatus (Campbell Electronics, India) containing 500 ml respec-
tive media. The experiment was carried out at 37 ± 1 ◦ C under
50 rpm paddle speed. The swelled microspheres were removed at
predetermined time interval and weighed after drying the surface
by using tissue paper. Swelling index was determined using the
following formula:

Swelling index = [(Weight of microspheres after swelling

The total amount of microspheres for each formulation batch
was obtained by weighing these prepared microspheres in a − Dry weight of microspheres)/Dry weight of beads] × 100
 S. Jana et al. / International Journal of Biological Macromolecules 72 (2015) 47–53
2.8. Fourier transform-infrared (FTIR) spectroscopy
Samples were reduced to powder and analyzed as KBr pellets
by using a Fourier transform-infrared (FTIR) spectroscope (Perkin
Elmer Spectrum RX I, USA). The pellet was placed in the sam-
ple holder. Spectral scanning was taken in the wavelength region
between 4000 and 400 cm−1 at a resolution of 4 cm−1 with scan
speed of 2 mm/s.
2.9. In vitro release study
In vitro drug release from these prepared microspheres was
evaluated using dialysis bag diffusion technique. Accurately
weighed quantities of microsphere containing aceclofenac equiva-
lent to 100 mg aceclofenac were placed in one end of dialysis bag
(Cellophane membrane, molecular cut off 10,000–12,000 Da, Hi-
Media, India) containing 5 ml of phosphate buffer, pH 6.8. After that,
order side of the dialysis bag was tied and immersed in phosphate
buffer (pH 6.8) contained in the USP type II dissolution appara-
tus (Veego VDA-6D, Veego Instruments Co-operation, India). The
system was maintained at 37 ± 1 ◦ C under 50 rpm speed. The dialy-
sis bag acted as a donor compartment. The vessel of dissolution
apparatus acted as the receptor compartment. 5 ml of aliquots
was collected at regular time intervals, and the same amount of
fresh dissolution medium was replaced into dissolution vessel to
maintain the sink condition throughout the experiment. The col-
lected aliquots were filtered, and suitably diluted to determine the
absorbance using a UV–vis spectrophotometer (Thermo Spectronic
UV-1, USA) by measuring absorbance at max of 274 nm.
2.10. Analysis of in vitro drug release kinetics and mechanism
In order to predict and correlate the in vitro drug release
behaviour from these microspheres containing aceclofenac, it is
necessary to fit into a suitable mathematical model. The in vitro
drug release data were evaluated kinetically using various impor-
tant mathematical models like zero order, first order, Higuchi and
Korsmeyer–Peppas models [17,18].
Zero-order model: Q = kt + Q0 ; where Q represents the drug
released amount in time t, and Q0 is the start value of Q; k is the
rate constant.
First-order model: Q = Q0 ekt ; where Q represents the drug
released amount in time t, and Q0 is the start value of Q; k is the
rate constant.
Higuchi model: Q = kt0.5 ; where Q represents the drug released
amount in time t, and k is the rate constant.
Hixson–Crowell model: Q1/3 = kt + Q0 1/3 ; where Q represents the
drug released amount in time t, and Q0 is the start value of Q; k is
the rate constant.
Korsmeyer–Peppas model: Q = ktn ; where Q represents the
drug released amount in time t, k is the rate constant and n
is the diffusional exponent, indicative of drug release mech-
anism. Again, The Korsmeyer–Peppas model was employed in
the in vitro drug release behaviour analysis of these formu-
lations to distinguish between competing release mechanisms:
Fickian release (diffusion-controlled release), non-Fickian release
(anomalous transport) and case-II transport (relaxation-controlled
release).
When n is ≤0.43, it is Fickian release. The n value between 0.43
and 0.85 is defined as non-Fickian release. When n ≥ 0.85, it is case-
II transport [19].
2.11. Anti-inflammatory activity evaluation
The carrageenan-induced rat-paw oedema model was per-
formed to assess anti-inflammatory activity evaluation of the
49
aceclofenac microspheres [20]. Male Sprague Dawley rats (weights,
130–210 g) were used in the study in accordance with a protocol
approved by Institutional Animal house Ethics Committee (IAEC)
(Registration number 955/A/06/CPCSEA).
The acclimatized rats were kept fasting for 24 h with water ad
libitum. The fasted animals were divided into 3 groups (n = 6) and
treated as follows:
Group 1 (standard group): pure aceclofenac drug (10 mg/kg
body weight) with 0.5% sodium carboxymethyl cellulose (Na CMC)
aqueous solution;
Group 2 (test group): aceclofenac-loaded microspheres (equiv-
alent to aceclofenac 10 mg/kg body weight) with 0.5% Na CMC
aqueous solution; and
Group 3 (control group): 2.5 ml of 0.5% Na CMC aqueous solu-
tion.
After oral administration of samples using feeding needles, rats
of all groups were challenged by subcutaneous injection of 0.05 ml
of 1% (w/v) solution of carrageenean in saline into plantar site of the
right hind paw. Oedema paw volumes were measured before and
after carrageenan administration at different time intervals with a
plethysmometer (INCO, India). Mean increase in paw volume and
% inhibition was calculated for all time intervals by using following
formula:
%Inhibition = (1 − Dt /Dc ) × 100
where Dt is the difference in paw volume in drug treated group;
Dc is the difference in paw volume in control animals.
2.12. Statistical analysis
All measured data are expressed as mean ± standard deviation
(S.D.). The simple statistical analyses were conducted using Med-
Calc software version 11.6.1.0.
3. Results and discussion
The aceclofenac-loaded microspheres made of alginate–LBG
were prepared through ionotropic gelation technique. Due to short
half-life of aceclofenac, sustained release dosage of aceclofenac
over an extended period is essential for reduction of dosing fre-
quency and minimization of various adverse effects during prolong
treatment. In this context, aceclofenac-loaded microspheres made
of alginate–LBG for prolonged aceclofenac release were aimed.
3.1. Yield and drug entrapment efficiency
The percent yield and drug entrapment efficiency of vari-
ous aceclofenac microspheres were measured and presented in
Table 2. The yield was found to be in the range of 64.44 ± 2.18 to
91.64 ± 2.49% of total solid content employed during the formula-
tion of beads. The maximum amount of yield could be due to the
insolubility of alginate and locust bean gum in calcium chloride
solution and might result in minimal loss of the dispersion. It was
possible that the polymeric system became cross-linked with diva-
lent calcium ions, present in the gelation medium and resulted in
higher yield of the microspheres. Increasing the concentration of
LBG actually led to higher drug content in the microspheres due
to formation of thick surface and decreases loss of drug in the cur-
ing medium. The drug entrapment efficiency was found to in the
range 59.64 ± 2.60 to 93.25 ± 1.09%. The higher entrapment effi-
ciency was based on both the concentration of sodium alginate and
the ability of the calcium ions to cross link with sodium alginate.
The degree of cross linking is depending upon both the concen-
tration of the calcium chloride solution and the time of contact of
beads with this solution. The microspheres prepared using sodium
50 S. Jana et al. / International Journal of Biological Macromolecules 72 (2015) 47–53

Table 2
Results of the percentage yield, drug entrapment efficiency, mean particle size and swelling index of the prepared formulations.

Formulation code Percentage yield (%)a Drug entrapment efficiency (%)a Average particle size(␮m)b Swelling index (%)a

In pH 1.2 In pH 6.8

F1 64.44 ± 2.18 59.64 ± 2.60 426 ± 12.26 30 ± 2.5 190 ± 2.8

F2 67.12 ± 1.69 63.22 ± 2.05 406 ± 10.18 26 ± 1.2 168 ± 4.6
F3 71.16 ± 2.20 70.16 ± 1.69 511 ± 22.34 32 ± 2.6 210 ± 3.2
F4 73.24 ± 2.17 78.24 ± 2.16 448 ± 18.74 28 ± 3.6 192 ± 3.7
F5 74.56 ± 1.81 82.22 ± 2.39 544 ± 15.26 33 ± 4.0 228 ± 1.4
F6 76.21 ± 2.04 83.92 ± 2.51 508 ± 19.28 30 ± 3.5 206 ± 2.6
F7 75.33 ± 1.06 70.28 ± 1.18 572 ± 21.74 38 ± 1.2 254 ± 4.3
F8 78.44 ± 2.78 72.28 ± 2.26 536 ± 20.16 34 ± 3.4 212 ± 4.8
F9 83.51 ± 1.74 80.56 ± 1.02 684 ± 23.36 44 ± 3.2 288 ± 1.6
F10 84.66 ± 1.56 86.21 ± 2.14 610 ± 24.14 41 ± 2.8 262 ± 2.2
F11 88.33 ± 2.16 89.59 ± 1.81 625 ± 16.28 42 ± 5.4 272 ± 4.8
F12 91.64 ± 2.49 93.25 ± 1.09 588 ± 18.74 39 ± 2.6 250 ± 2.8
a
Mean ± S.D.; n = 3.
b
Mean ± S.D.; n = 100.

alginate:LBG of 1:1 showed low entrapment of aceclofenac. This
was due to the insufficient cross linking and large pore size per-
mitting the drug to diffuse out during and after gelation. From
the results, it was also observed that increasing calcium chlo-
ride concentration produced beads with higher levels of Ca2+ ions.
Consequently, the cross-linking of the polymer and compactness
of the formed insoluble dense matrices also increased, resulting
in more drug entrapment in the microspheres. Increasing in the
concentrations of LBG in formulations, the drug entrapment effi-
ciency progressively increased. It may be due to the formation of
dense matrix and uniform encapsulation of insoluble aceclofenac
within alginate-LBG interpenetrating network. The highest yield
(91.64 ± 2.49%) and drug entrapment efficiency (93.25 ± 1.09%)
were observed for the microspheres of formulation F12, prepared
using 2% (w/v) of sodium alginate, 2% (w/v) of LBG, 1% (w/v) ace-
clofenac and 4% (w/v) of CaCl2 .
3.2. Particle size analysis
The particle size of aceclofenac-loaded alginate–LBG micro-
spheres were measured and found within the range of 406 ± 10.18
to 684 ± 23.36 ␮m (Table 1). It was found that the particle size dis-
tribution was within a narrow size but the mean particle size was
different among the formulations. The results indicated that the
mean particle size of microsphere increased proportionally with
the increase in the amount of sodium alginate in the formula-
tions. This could be attributed to an increase in relative viscosity
at higher concentration of alginate and formation of large droplets
during addition of polymer solution to the gelling agent. Fur-
ther, an increase in concentration of calcium chloride significantly
decreased the mean particle size of microspheres. It has been stated
that when a drop of alginate solution comes in contact with calcium
ions, gelation occurs instantaneously. As Ca+2 ions, penetrates into
interior of droplets, water is squeezed out of the interior of droplets
resulting in contraction of beads.
followed by complete entrapment of drug into interior polymer
network.
3.4. Swelling behaviour
The swelling behaviour of microspheress of alginate–LBG con-
taining aceclofenac was evaluated in acidic buffer of pH 1.2 and
phosphate buffer of pH 6.8 up to 4 h, and presented in Fig. 2.
The Swelling index values of beads in pH 1.2 were within the
range between 26 ± 1.2 to 44 ± 3.2% and in pH 7.4 was 168 ± 4.6
to 288 ± 1.6%. The Swelling index of microspheres containing ace-
clofenac was lower in 0.1N HCl (pH 1.2) in comparison with that
of in phosphate buffer (pH 6.8). Under acidic conditions swelling of
calcium alginate beads occurs scarcely. The low swelling in acidic
media pH 1.2 was probably due to proton-calcium ion exchange
forming insoluble alginic acid regions and followed by solvent
penetration into the gel network. Being a polyelectrolyte, alginate
can exhibit swelling properties that are sensitive to the pH, ionic
strength and ionic composition of the medium. The equilibrium
swelling studies showed, with increase in the alginate concentra-
tion, swelling of beads was significantly increased in pH 6.8 at the
end of 4 h. It has been reported that the swelling can be enhanced
by the presence of phosphate ions in higher pH which displaces the
Ca2+ ions within the beads. The swelling behaviour of the formu-
lations also indicated that if concentration of calcium chloride was
increased, the swelling of the beads became reduced. The overall

3.3. Morphological analysis

The morphological analysis of aceclofenac-loaded microspheres

made of alginate–LBG was visualized by SEM and is presented in
Fig. 1. They were found almost spherical in shape and dense with
thick polymer coat. Concentration of alginate influences the surface
morphology of particles. At higher concentration alginate formed
discrete and spherical shape with a rough outer surface and visible
large wrinkles has a sandy appearance because of the surface-
associated crystals of drug. Addition of LBG alters morphological
regularity due to formation of smooth surface; uniform sphericity
and formation of thick coat on outer surface of the alginate beads Fig. 1. SEM photograph of aceclofenac loaded alginate–LBG microsphere (F12).
 S. Jana et al. / International Journal of Biological Macromolecules 72 (2015) 47–53
Fig. 2. Comparison of the swelling behaviour of microspheres of alginate–LBG con-
taining aceclofenac in pH 1.2 and 6.8.
results suggested that the dried beads swelled slightly in the acidic
medium. When they were subsequently transferred to alkaline
medium, the particles began to swell to an appreciable level and
formed a thick diffusion layer surrounding the particles, thereby
sustaining the release of incorporated drug.
3.5. FTIR analysis
FTIR spectra analysis of pure aceclofenac, sodium alginate,
LBG and aceclofenac-loaded microspheres made of alginate-LBG,
is shown in Fig. 3. The FTIR spectra of sodium alginate showed
the characteristic peak at 3606 cm−1 due to stretching of OH,
1614 cm−1 due to the stretching of COO (asymmetric), 1417 cm−1
due to the stretching of COO (symmetric) and 1032 cm−1 due to
the stretching of C O C, respectively. In the FTIR spectrum of LBG
showed characteristic band around 3186 cm−1 due to stretching of
OH, band at 2915 cm−1 due to C H stretching, 1705 cm−1 indi-
cating C O stretching. The FTIR spectra of aceclofenac showed
that principal peaks at 3028 cm−1 due to aromatic C H stretching
vibrations and 2937 cm−1 due to aliphatic C H stretching vibra-
tions, a band at 1717 cm−1 due to C O stretching, a sharp band
at 1772 cm−1 due to C O stretching of carboxylic acid, band at
3320 cm−1 due to secondary N H rocking vibrations, and two sharp
peaks at 716 cm−1 due to 1,2 di-substituted C Cl stretching. These
Fig. 3. FTIR spectra analysis of pure aceclofenac, sodium alginate, LBG and aceclofenac loaded alginate–LBG microspheres.
51
peaks were also appeared clearly in the FTIR spectra of aceclofenac-
loaded microspheres of alginate–LBG (F12) without any significant
shifting of peaks. This observation confirmed that there was no drug
excipient interaction.
3.6. In vitro drug release
The in vitro drug (here, aceclofenac) release studies were car-
ried out for various microspheres of alginate–LBG containing
aceclofenac in phosphate buffer, pH 6.8. The cumulative per-
centage drug release from aceclofenac-loaded microspheres was
found sustained over a period of 8 h (Fig. 4). The percentage drug
released from aceclofenac-loaded alginate-LBG microsphere in 8 h
was within the range of 71.78 ± 2.14% to 89.12 ± 2.24%. As the poly-
mer concentration was increased, the release rate of aceclofenac
sodium from the microspheres decreased. The slower in the release
rate can be explained by the increase in the extent for swelling and
the gel layer thickness that acted as a barrier for the penetration
medium thereby retarding the diffusion of drug from the swollen
alginate beads. The results also explained that the initial burst
release of alginate was reduced by increasing the concentration of
LBG as a result of formation of diffusional bridges due to swelling
of hydrophilic linkage in pH 6.8 buffer. It was found that rate and
extent of drug release decreased significantly with increase of con-
centration of calcium chloride, because sodium alginate as a linear
copolymer consisting of ␤ (1→4) mannuronic acid and ␣ (1→4)
l-guluronic acid residues; a tight junction is formed between the
residues of alginate with calcium ions. However, in case of higher
calcium chloride concentration due to increased surface roughness
and porosity and also poor entry of dissolution medium into the
polymer matrix may be delayed drug release.
In vitro drug release data from various aceclofenac-loaded
alginate–LBG microsphere were evaluated kinetically using vari-
ous important mathematical models like zero order, first order,
Higuchi, Hixson-Crowell and Korsmeyer–Peppas models. The R2
values of these models were determined for evaluation of accuracy.
The result of the curve fitting into various mathematical models
is given in Table 3. When the respective R2 of these aceclofenac-
loaded alginate–LBG microsphere were compared, it was found to
follow the Korsmeyer–Peppas model (R2 = 0.9808 − 0.9876) over
a period of 8 h. The value of release exponent (n) determined
from in vitro aceclofenac release data of various aceclofenac-loaded
52 S. Jana et al. / International Journal of Biological Macromolecules 72 (2015) 47–53

Fig. 4. The in vitro drug release from aceclofenac-loaded alginate–LBG microspheres in phosphate buffer of pH 6.8 (mean ± S.D.; n = 3) of all formulations (a) for F1 to F6 and
(b) for F7 to F12.

Table 3
Results of curve fitting of the in vitro drug release profile of various alginate-LBG microparticles containing aceclofenac.

Formulation code Zero order model (R2 ) 1st order model (R2 ) Higuchi model (R2 ) Hixson–Crowell model (R2 ) Korsmeyer-Peppas model

R2 n

F1 0.8422 0.9226 0.9276 0.8468 0.9864 0.68

F2 0.8756 0.8744 0.9268 0.9694 0.9808 0.53
F3 0.9264 0.9346 0.8774 0.9674 0.9832 0.63
F4 0.8634 0.9428 0.9254 0.9588 0.9828 0.75
F5 0.8766 0.9567 0.9456 0.8732 0.9823 0.66
F6 0.9145 0.9345 0.9742 0.9248 0.9826 0.74
F7 0.8746 0.8784 0.9124 0.9658 0.9816 0.58
F8 0.8560 0.8956 0.9411 0.9564 0.9876 0.64
F9 0.8720 0.9122 0.9724 0.9684 0.9866 0.56
F10 0.8314 0.8568 0.9132 0.8846 0.9854 0.62
F11 0.9194 0.9342 0.9466 0.8962 0.9838 0.69
F12 0.9126 0.9386 0.9762 0.9172 0.9872 0.72

alginate–LBG microsphere ranged from 0.53 to 0.75, indicating

anomalous (non-Fickian) diffusion mechanism for drug release. The
anomalous diffusion mechanism of drug release demonstrates both
diffusion controlled, and swelling controlled drug release.

3.7. Anti-inflammatory activity

Anti-inflammatory effect of the F12 microspheres composed

of alginate–LBG containing aceclofenac was evaluated using
carrageenan-induced rat-paw oedema model. After oral adminis-
tration of pure aceclofenac at a dose of 10 mg/kg body weight and
F12 microspheres containing aceclofenac at a dose of equivalent
to aceclofenac 10 mg/kg body weight, rats of all groups (control,
standard and test) were challenged by a subcutaneous injection of
0.05 ml of 1% (w/v) solution of carrageenan in saline into the plan- Fig. 5. The percentages inhibition of paw oedema swelling for the standard (pure
aceclofenac) and the test (F12 microspheres containing aceclofenac) at various time
tar site of left hind paw. The percentages inhibition of paw oedema
intervals.
swelling for the standard (pure aceclofenac) and the test (F12
microspheres containing aceclofenac) were calculated from mean
paw oedema volumes (ml) at various time intervals were calculated 4. Conclusion
and presented (Fig. 5). The pure aceclofenac (standard) exhibited
comparatively rapid inhibition of paw oedema swelling than that Aceclofenac-loaded alginate–LBG microspheres for prolonged
of the F12 microspheres tested indicating rapid anti-inflammatory aceclofenac release were prepared through ionotropic gelation
activity up to 3rd hour and after that, this was decreased. The F12 technique and evaluated. The drug entrapment efficiency of
microspheres containing aceclofenac (test) showed slower inhibi- these microspheres was found 59.64 ± 2.60% to 93.25 ± 1.09% and
tion of paw oedema swelling and maintained increasing inhibition their average particle sizes were 406 ± 10.18 to 684 ± 23.36 ␮m.
of paw oedema over 5 h after oral administration indicating slower FTIR analysis confirmed the compatibility of the aceclofenac
and sustained action of aceclofenac. with the polymer blend used to prepare microspheres of
 S. Jana et al. / International Journal of Biological Macromolecules 72 (2015) 47–53 53

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