International Journal of Pharmaceutical Research

2010, Volume 2, Issue 3, 33-38

ISSN 0975-2366

Research Article
Design and Development of Sustained Release Chitosan-Bora Rice Microspheres for
Targeted Drug Delivery to the Colon
Ramteke KH*. Bhattacharya A, Nath LK
Dept. of Pharmaceutical Sciences, DIbrugarh University, Assam.
*Corresponding Author: E-mail: kuldeep_mph@rediffmail.com
Received: 31/12/2010, Revised: 31/04/2010, Accepted: 30/07/2010
ABSTRACT
The purpose of this research was to formulate and systematically evaluate in-vitro performances of glipizide micro-
spheres for targeted delivery to the colon. Glipizide microspheres containing Chitosan-bora rice were prepared by simple
emulsification phase separation technique using glutaraldehyde as a crosslinking agent.
Results of preliminary trials indicate that polymer to drug ratio and speed of rotation affected characteristics of micro-
spheres. The size of microspheres was found to range from 420-723 μm. The microspheres exhibited high % drug entrap-
ment efficiency, best batch exhibited a high drug entrapment efficiency of 57.4% and swelling index ranges from 0.73 ± 0.03
to 1.35 ± 0.15. The amount of drug release after 28 hr was found to be 64.31% ± 1.26 in PBS pH 7.4 dissolution medium and
medium with 2% cecal material 85.53% ± 1.16 drugs released within 24 hrs.
The drug release of Glipizide significantly decreased with increasing bora rice concentration this may be due to the
higher % of amylopectin present. FTIR spectra confirmed the stable character of glipizide in drug loaded microspheres. DSC
study confirmed that drug is present in the molecular dispersion. SEM indicated the surface topology of microspheres. In
vitro data obtained for colon targeting microspheres of Glipizide showed sustained drug release. Hence Bora rice may be
used as sustained release polysaccharides for colon targeting drug delivery system.

KEY WORDS: Chitosan, Colon targeting microspheres, Glipizide, In vitro study

INTRODUCTION
Until recently, colon was considered as a site for wa-
ter reabsorption and residual carbohydrate fermentation.
However, it is currently being viewed as a site for drug
delivery. Moreover, colon transit time may last for upto 78
h, which is likely to increase the time available for drug
absorption. Glipizide shows pH dependent solubility so
there is need to formulate colon targeting drug delivery
system. Bora rice is a rich source of amylopectin which
shows sustained action of drug release. Several polysac-
charides are being investigated as carriers for colon-
specific drug delivery.
Objective
An objective of the present investigation was to pre-
pare and characterize chitosan – bora (CS-BR) rice micro-
sphere for colon targeting of Glipizide.
EXPERIMENTAL
Materials
Chitosan (CS) was obtained as a gift sample from
sigma fisheries, cochin. Bora rice (BR) purchased from
upper Assam region. Light liquid paraffin obtained from
Rankem. Gluteraaldehyde purchased from Qualigens. All
other chemicals/reagents used were of analytical grade.
Preparation of microspheres
Chitosan 3%w/v was dissolved in 20ml of 2% aq. v/v
acetic acid solution and stirred to get uniform solution.
This chitosan solution was centrifused at 2000 rpm for 2
min to form bubble free solution, dispersed required quan-
tity of glipizide to form uniform dispersion of drug. Add
required amount of bora rice powder (Table - 1) previously
passed through sieve no. 100. This dispersion was emulsi-
fied into liquid paraffin in presence of 0.5% span 80 using
Remi lab stirrer at various speed for 20 min. The mixture
of glutaraldehyde (5 ml) was added slowly and stirring
were continue for 3 hrs. The hardened microspheres were
separated by filtration and washed with n-hexane. The
microspheres were dried at 500C for 24 hr and stored in
desicator.
Assay of Glipizide
Glipizide was estimated by UV/Vis spectrophotome-
tric (Shimadzu UV-1601 UV/Vis double beam spectropho-
tometer) method (Figure 1). Aqueous solutions of glipizide
were prepared in phosphate buffer (pH 7.4) and absorbance
was measured on UV/Vis spectrophotometer at 276 nm.
The method was validated for linearity, accuracy and pre-
cision. The method obeys Beer’s Law in the concentration
range of 5-35 µg/ml.
Particle size analysis
The particle size of the microspheres was determined
by sieving method (Table 2). Microspheres were separated
into different size fractions by sieving for 10 min using
mechanical sieve shaker (Cuprit Electronic co. India) con-
taining std sieves having aperture of 1000, 710, 500, 355,
250 & 180 μm.
Micromeritic properties of drug loaded microspheres
The flow properties of microspheres were investigated
by determining the angle of repose, bulk density and
tapped density (Table 3). The angle of repose was deter-
mine by the fixed base cone method. Bulk and tapped den-
sities were measured in 10 ml graduated cylinder. The
sample contained in the cylinder was tapped mechanically,
tapped volume was noted down when it showed no change
in its volume and bulk density and tapped density was
calculated.

IJPR | July-September | 33
 Ramteke & Bhattacharya / International Journal of Pharmaceutical Research 2010 2(3) 33-38

Table 1: Bach specification for prepared microsphere.

Polymer Ratio Drug:Polymer Volume of

Batch Temperature Time of Cross-
(CS:BR) ratio Cross-Linking
Code (0C) Linking (min)
wt/wt wt/wt agent (ml)
A1 a 1:2 1:2 30 - 40 10 5
A2 a 1:4 1:2 30 - 40 10 5
A3 a 1:6 1:2 30 - 40 10 5
A4a 1:8 1:2 30 - 40 10 5
B1 b 1:2 1:2 30 - 40 10 5
B2 b 1:4 1:2 30 - 40 10 5
B3 b 1:6 1:2 30 - 40 10 5
B4 b 1:8 1:2 30 - 40 10 5
Stirring Rate: a = 500 rpm; b = 800 rpm

Drug entrapment efficiency

Microspheres (50 mg) were crushed in a glass mortar-
pestle and the powdered microspheres were suspended in
50 ml phosphate buffer (pH 7.4). After 24 h the solution
was sonicate for 1 hr, filtered and the filtrate was analyzed
for the drug content. The drug entrapment efficiency was
calculated as per the following formula:
Practical drug content/Theoretical drug content x 100.
Swellability
A known weight (100 mg) of various glipizide loaded
chitosan/bora rice microspheres were placed in phosphate
buffer, pH 7.4 and allowed to swell for the required period
of time at 37ºC ± 0.5ºC in the dissolution. The micro-
spheres were periodically removed and blotted with filter
paper; then their change in weight (after correcting for drug
loss) was measured until attainment of equilibrium. The
swelling ratio (SR) was then calculated using the following
formula:
double adhesive tape, which struck to an aluminum stub.
The stubs were then coated with gold to a thickness of
~300 0A using a sputter coated and viewed under the scan-
ning electron microscopy (Figure 4).
In vitro dissolution study
The drug release study was carried out using USP
XXIV paddle type apparatus (Electrolab, TDT-06T, India)
at 37 ± 0.5° and at 75 rpm using 900 ml of phosphate buf-
fer (pH 7.4) as a dissolution medium (Figure 4). Micro-
spheres equivalent to 5 mg of glipizide were used for the
test. Five ml of sample solution was withdrawn at prede-
termined time intervals, filtered through a 0.45 µm mem-
brane filter, diluted suitably and analyzed spectrophotome-
trically. Equal amount of fresh dissolution medium was
replaced immediately after withdrawal of the test sample.
Percentage drug released at different time intervals was
calculated using the Lambert-Beer’s equation (y = 32.0624
- 0.0212, R2=0.9997) described above.

SR= (wg – w0)/w0 Preparation of 2% rat cecal material

SR= Swelling ratio
wg = Final weight of microspheres
w0 = Initial weight of microsphere
FTIR Study
FTIR spectra of Glipizide, blank microspheres and
drug loaded microspheres were obtained in KBR pellets
using a JASCO model 5300 spectroscope in the ranges
4000 to 400 cm-1 (Figure 2).
Male albino rats weighing 200 – 250 g and maintained
at normal diet were used for the study. Six rats were as-
phyxiated using carbon dioxide. The abdomens were
opened, the ceci were traced, legated at both ends, dis-
sected, and immediately transferred into pH 7.4 phosphate
buffered saline (PBS), previously bubbled with CO2. The
cecal bags were opened and the contents were individually
weighed, pooled and the suspended in PBS to provide 2%
w/v dilution. Because the cecum is naturally anaerobic, all
of these operations were carried out under CO2.

Thermal Studies In vitro drug release study in the presence of colonic

Thermogram of samples was obtained by a Prkin-
Elmer Differential ScanningColorimeter (Figure 3). Sam-
ples of 10mg were accurately weighed into aluminum pans
and then hermetically sealed with aluminum lids. The
thermograms of samples were obtained at a scanning rate
of 100C/ min over a temperature range of 50 to 2500C.
Shape and Surface morphology
Surface and shape morphology of CS-BR micro-
spheres were evaluated by means of scanning electron
microscopy (HITACHI S-3600N). The samples of SEM
were prepared by lightly sprinkling the microspheres on a
fluid containing 2% rat cecal material
The drug release study was carried out using USP
XXIV paddle type apparatus (Electrolab, TDT-06T, India)
at 37 ± 0.5° and at 75 rpm using 900 ml of phosphate buf-
fer (pH 7.4) containing 2% rat cecal material as a dissolu-
tion medium. Microspheres equivalent to 5 mg of glipizide
were used for the test (Figure 5). 5 ml of sample solution
was withdrawn at predetermined time intervals. Equal
amount of fresh dissolution medium was replaced imme-
diately after withdrawal of the test sample. The withdrawn
samples were pipetted into a series of 10 ml volumetric
flask, diluted suitably and centrifuged. The supernatant was

34 | IJPR | July-September
 Ramteke & Bhattacharya / International Journal of Pharmaceutical Research 2010 2(3) 33-38

filter through 0.45 µm membrane filter and filtrate was

subject to UV analysis as described previously. Bulk Tapped Angle
Batch Sphericity of
Density Density of Re-
Code Microspheres
RESULT AND DISCUSSION (g/cc) (g/cc) pose
Microspheres were prepared by the emulsification
0.842 ± 1.45 ± 38.54 ±
method using chitosan/bora rice (Table 1). The fabricated A1 Very irregular
0.065 0.12 2.47
microspheres were spherical in shape and exhibited a range
of sizes within each batch. 0.835 ± 1.36 ± 36.55 ±
Microspheres were prepared using gradually increas- A2 Very irregular
0.09 0.086 2.13
ing BR concentration in combination with a fixed concen-
tration of CS to assess the effect of polymer concentration 0.865 ± 1.49 ± 38.74 ±
on the size of microspheres. The mean particle size of the A3 Very irregular
0.057 0.083 1.95
microspheres significantly increased with increasing BR
concentration and was in the range of 420 to 723 µm (Ta- 0.81 ± 1.47 ± 39.62 ±
A4 Very irregular
ble -2). It may be due to high molecular weight. The par- 0.078 0.08 2.12
ticle size was reduced as agitation speed increases. Formu-
lation A1 to A4 forms lumps and they are irregular in 0.551 ± 0.610 ± 28.34 ± Spherical free
B1
shape due to low agitation speed i.e 500 so we selected the 0.034 0.076 1.95 flowing
batches B1 to B4 for the further study.
0.551 ± 0.614 ± 27.56 ± Spherical free
B2
Table 2: Various formulation parameters for micro- 0.019 0.026 1.87 flowing
spheres
0.559 ± 0.620 ± 28.42 ± Spherical free
B3
0.045 0.033 2.06 flowing
Mean
Entrap- 0.556 ± 0.612 ± 29.12 ± Spherical free
Batch Yield Particle Degree of B4
ment effi- 0.025 0.023 1.76 flowing
Code (%) size swelling
ciency
(μm)
Tapped density of the formulation batches A1 to A4
A1 90.12 57.2 ± 2.5 570 1.35± 0.15 was high as compare to B1 to B4 it may be due to the irre-
gular in size and hence it shows very poor flow property.
A2 91.25 55.6 ± 4.8 592 1.31±0.15 Batches B1 to B4 showed good flow property due to its
spherical nature.
A3 90.68 57.7 ± 3.6 661 1.27±0.16

A4 92.51 56.4 ± 5.3 723 1.21±0.12 Calibration curve of Glipizide:

B1 93.22 56.1 ± 4.5 420 0.98±0.04

B2 91.35 57.4 ± 1.9 445 0.92±0.02

B3 90.45 56.8 ± 4.9 479 0.86±0.04

B4 91.68 57.3 ± 2.8 487 0.73±0.03

Calibration Curve
1

0.9

0.8

0.7
A b s o rb a n c e

0.6

0.5

0.4

0.3

0.2

0.1

0
0 5 10 15 20 25 30 35
Concentration (mgm/ml) Figure 2: FTIR spectra of pure Glipizide (GLP), Blank
Microspheres (BMS), Drug Loaded Microspheres (DLM),
Figure 1: Calibration curve for Glipizide in PBS 7.4 Bora Rice (BR), Chitosan (CS).
Table 3: Micromeritic properties of drug loaded micro- Glipizide showed prominent peaks at 1651, 2943, and
spheres. 1529 due to the presence of C=N aliphatic groups, C-H2

IJPR | July-September | 35
 Ramteke & Bhattacharya / International Journal of Pharmaceutical Research 2010 2(3) 33-38

aliphatic C-H aliphatic. The same peaks were also ob-

served in the formulation indicating the stable nature of the
drug during encapsulation.

Figure 3: Differential Scanning calorimetry thermogram of (GLY) Glipizide, (DLM) Drug loaded microsphere, (BRB)
Blank microsphere and (BR) Bora Rice powder.

(A) (B)

(C) (D)

Figure 4: A: Blank CS-BR microspheres, B: Surface of Blank microspheres, C: Surface of Drug loaded microspheres, D:
Surface of drug loaded microspheres after dissolution.

36 | IJPR | July-September
 Ramteke & Bhattacharya / International Journal of Pharmaceutical Research 2010 2(3) 33-38

DSC is very useful in the investigation of the thermal release in the simulated colonic fluid with cecal content
properties of microspheres, providing both qualitative and may be a result of the combined effect of diffusion and
quantitative information about the physicochemical state of erosion.
drug inside the microspheres. Prominent melting endo-
therm of pure glipizide was found at 218.50C. Drug loaded CONCLUSION
microspheres doesn’t show any endotherm may be due to The results of our study clearly indicate that there is a
the drug was present in the molecular dispersion or solid great potential in delivery of glipizide to the colonic re-
solution state in the polymeric microspheres loaded with gion. Study showed that the manipulation of polymer con-
drug. centration and stirring rate influence particle size of micro-
spheres, sphericity and flow property of microspheres.
Scanning Electron Microscopy From the above study it concluded that high concentration
Scanning electron microscopy revealed that CS-BR of Bora Rice will retard the drug release, may be due to
microspheres were discrete and spherical in shape with high content of amylopectin present in the bora rice. For-
rough outer surface because of the surface associated with mulation B4 is the best formulation for sustaining the drug
crystals of drug (Figure 4C). After dissolution study the release to the colon. Hence from the above study it con-
surface showed erosion (Figure 4D) cluded that high amylopectin containing bora rice, natural
polysaccharide may be used as sustained release polymer
for colon targeting drug release study.
Drug release profile
ACKNOWLEDGEMENT
The authors greatly acknowledge M/s Stadmed Pri-
100
vate Ltd, Kolkata, India, for the supply of glipizide free of
80 charge. The authors are also grateful to the Oil India, Dhu-
% drug release

B1
60
liajan, India for help in performing characterization studies.
B2
B3
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