DESIGN, DEVELOPMENT AND CHARACTERIZATION OF pH SENSITIVE HYDROGEL FOR INTESTINAL DELIVERY OF PREDNISOLONE

ABOUT AUTHORS:
Patel Juhi D.*, Dr. A.K.Seth, Sachin P Chauhan, Nirmal Shah, Ankur Javia, Chintan Aundhia
Department of Pharmacy, Sumandeep Vidyapeeth University,
At & Po Pipariya, Ta- Waghodia, Dist. Vadodra-391760.(Gujarat) India
pateljuhi63@yahoo.com

ABSTRACT
In the present study pH sensitive hydrogel beads of prednisolone were prepared for intestine delivery system. The cross-linking reinforced chitosan- alginate complex beads were prepared by gelation of anionic sodium alginate, the primary polymer, with opposite charged counter ion to form beads which were further complexed with chitosan as a polyelectrolyte. The pH sensitive hydrogel beads were prepared by the unique ionotropic gelation method by applying 33 full factorial designs. In this present study, influence of various formulation parameters like drug: chitosan ration (1:0.5, 1:1, and 1:2), drug: sodium alginate ratio (1:1, 1:2, 1:3) and concentration of calcium chloride (3%, 4%, 5%) were studied. According to factorial design, total 27 batches were prepared and evaluated. The compatibility of drug with polymer was confirmed by Fourier transform infrared spectroscopy (FT-IR). All the formulation were subjected to percentage yield, drug entrapment efficiency, drug content, particle size, swelling study (In phosphate buffer pH 1.2 as well as in phosphate buffer pH 7.4), and in vitro drug release study. The data was found in the range of 72.00 ± 0.8 % to 92.00 ± 0.7 %,16.49 ± 0.2 % to 70.00 ± 0.2 %, 4.70 ± 0.4 to 13.30 ± 0.5,102.36 ± 2.27 to 138.48 ± 2.25 μm, , in phosphate buffer pH 7.4 between 109.00 ± 0.9 to 168.00 ± 0.4 and 106.60±0.6 to 136.00±0.3 in  phosphate buffer pH 1.2. From the results, B8 formulation was optimized. The prepared beads were enteric coated with 10% Eudragit S 100.  The surface morphology was studied by scanning electron microscopy (SEM) revealed spherical in shape with rough surface. It was reported that there was no swelling in acidic pH but increased in basic pH resulting in increased release of the drug in intestine. Hence it could be concluded that the prepared chitosan-sodium alginate hydrogel beads were satisfactory delivered to the stimulated intestinal fluid, which can be confirmed by further in vivo study.

REFERENCE ID: PHARMATUTOR-ART-2035

INTRODUCTION: Chitosan is a biopolymer, which could be used for the preparation of various polyelectrolyte complex products with polyanions such as carboxymethylcellulose, xanthan, alginate and gellan gum. Chitosan-polyanion complex have been widely investigated for application like drug and protein delivery, cell transplantation, enzyme immobilization. Among the chitosan –polyanion complexes, chitosan-alginate complex may be the important drug delivery system. The strong electrostatic interaction of alginate leads to the formation of chitosan-alginate complex.

Prednisolone is (11β)-11, 17, 21-trihydroxypregna-1, 4-diene-3, 20-Dione. It is well known corticosteroid that is used to treat inflammatory bowel diseases such as Crohn's disease and ulcerative colitis.  It is also used to treat arthritides, collagen disorder, and severe allergic reactions, autoimmune diseases, bronchial asthma, eye diseases and infective diseases.

This work focused on the preparation of novel chitosan-alginate beads with inner cross-linked core with calcium chloride and outer chitosan-alginate complex membrane. The one-stage procedure for the preparation of cross-linking reinforced chitosan-alginate beads was examined by dropping alginate solution into chitosan solution containing calcium chloride cross-linking agent. In order to increase the stability of chitosan-alginate complex, chitosan solution, consisting of calcium chloride, was used for the gelation of alginate. The presence of calcium salt leading to the competition of gelling reaction and polyelectrolyte complex results in the formation of  a more porous gel, allowing the diffusion of chitosan.

MATERIALS AND METHODOLOGY:
Prednisolone was gifted by Modi pharmaceutical (Ahmadabad), chitosan – Balaji drugs (Surat), Eudragit S 100 - Roehm Pharma polymer, (Germany), Sodium alginate - Sulabh laboratory, (Vadodara).

FORMULATION OF HYDROGEL BEADS:
The hydrogel beads were prepared by the unique ionotropic gelation method by using33 factorial designs. Sodium alginate solution was prepared by dissolving in deionized water (50 ml) and heated at 60°C. The drug was dispersed uniformly in alginate solution below 40°C under continuous stirring. The stirring was continued until a uniform dispersion was obtained. The obtained dispersion was sonicated for 10-15 min. The resultant homogeneous bubble free slurry dispersion was dropped trough a 21G syringe needle into100 ml of calcium chloride solution containing chitosan as per the ratio. The solution kept under the stirring (1000-1200 rpm) to improve the mechanical strength of the beads. Immediate formation of small alginate beads took place after 5 min of curing time. The formed beads were collected by filtration and deride at 40°C.

CHARACTERIZATION OF HYDROGEL

Production yield:
The yield was calculated by dividing the weight of collected hydrogel beads by the weight of non-volatile compound used for the preparation of hydrogel beads and expressed in term in percentage.

Particle size measurement:
The prepared beads were subjected for particle size analysis using a microscope having accuracy of 0.001 mm. the average diameter of the 25 particles per batch was calculated.

Drug content:
The prepared hydrogel were powdered and finely hydrogel powder was subjected to sieving. The hydrogel particles were taken for drug content studies. A 20 mg of beads were taken in volumetric flask. To this 20 ml of pH 7.4 phosphate buffer was added and kept overnight. The final solution was filtered by using whatmann filter paper. From this 10 ml was pipette out in to 100 ml volumetric flask, made up the volume with 7.4 pH phosphate buffer and estimate for drug content.

Dynamic swelling study:
The dynamic swelling behavior of the beads was studied by mass measurement. The 50 mg beads were incubated with 25 ml phosphate buffer solution pH 1.2 and pH 7.4 separately at 37°c. The beads were taken out at different time intervals up to 12 hrs and blotted carefully without pressing hard to remove the excess surface liquid. The swollen beads were weighed using the electronic microbalance. The percent water uptake (Q) at different time intervals was calculated using the following equation:

Where, W1- Mass of the dry beads
W2-Mass o the swollen beads

Drug entrapment efficiency:
The entrap amount of drug prednisolone in hydrogel beads was estimated by using UV- visible spectrophotometer at 247nm.Known amount of beads (20 mg) were added to 20 ml USP phosphate buffer of pH 7.4 solution for complete swelling at 37°C. The beads were crushed in a glass mortar with pestle the solution was than kept for 2 hrs to extract the drug completely and centrifuge to remove polymeric debris. The clear supernatant solution was analyzed for drug content using UV- visible spectrophotometer at 247 nm.

In – vitro drug release study:
In vitro drug release study was carried out using a USP-XXII dissolution apparatus. The dissolution was measured at 37.0 ± 0.5 °C and 50 rpm paddle speed. Drug release from the beads was studied in 500 ml acidic medium (pH 1.2) for 2 hours and alkaline medium (pH 7.4 phosphate buffer) till end of the study. 0.1% SLS was added as a surfactant. At predetermined time intervals, 5 ml aliquots were withdrawn and replaced with the same volume of fresh solution. The amount of drug release was analyzed using UV- visible spectrophotometer at 247 nm.

Release Kinetics:
In order to investigate the mechanism of drug release from hydrogel beads of different ratios, the release data obtained from In vitro release studies were fitted to various kinetic equations.

Enteric coating of optimized formulation:
The beads prepared were transferred into acetone solutions of Eudragit S 100 at concentration of 10 %w/v and coated for 15 min under stirring. The resulting coated beads were filtered and air dried.

EVALUATION OF OPTIMIZED BATCH:

Swelling study of optimized batch:
The dynamic swelling behavior of the beads was studied by mass measurement. The 50 mg beads were incubated with 25 ml phosphate buffer solution pH 1.2 and pH 7.4 separately at 37°C. The beads were taken out at different time intervals up to 12 hrs and blotted carefully without pressing hard to remove the excess surface liquid. The swollen beads were weighed using the electronic balance. The percent water uptake (Q) at different time intervals was calculated using the following equation:

Where, W1- Mass of the dry beads
W2-Mass o the swollen beads

Determination of the Gastro-Resistance of enteric coated beads:
The prepared beads (B8) were placed in dissolution bath containing 0.1N HCl at 37 ± 0.2° C for 2 hour. During the acid step, sample was collected for quantification. Then, samples were collected at predetermined time intervals and analyzed spectrophotometrically at 247 nm.

Release study of optimized batch:
In vitro drug release study was carried out using a USP-XXII dissolution apparatus. The dissolution was measured at 37.0 ± 0.5 °C and 50 rpm paddle speed. Drug release from the beads was studied in 500 ml acidic medium (pH 1.2) for 2 hours and alkaline medium (pH 7.4 phosphate buffer) till end of the study. 0.1% Sodium lauryl sulphate (SLS) was added as surfactant. At predetermined time intervals, 5 ml aliquots were withdrawn and replaced with the same volume of fresh solution. The amount of drug release was analyzed using UV- visible spectrophotometer at 247 nm.

Scanning Electron Microscopy (SEM):
The surface morphology of beads was investigated by using scanning electron microscopy (SEM) (JEOL JSM-5610LV). The beads were mounted onto stub using double sided adhesive tape and sputter coated with platinum using a sputter coater. The coated beads were observed under SEM instrument at the required magnification at room temperature. The acceleration voltage used was 15 KV with the secondary electron image detector.

STABILITY STUDY OF OPTIMIZED BATCH:
Various ICH storage conditions are available which are as 25°C ± 2°C (60% ± 5%RH), 30°C ± 2°C (65% ± 5%RH) and 40°C ± 2°C (75% ± 5%RH). The best formulation were placed in screw capped glass container and stored at various ICH storage condition for a period of 60 days. The samples were analyzed for physical appearance and for the drug content at regular interval of 15 days.

RESULT AND DISCUSSION:
The formulation compositions of various batches are shown in table no: 1. The Percentageyield (%),Drug Entrapment Efficiency (%),Drug content, Mean particle size, Dynamic swelling study are summarized in table no: 2. The drug entrapment efficiency was in the range of 16.49 ± 0.2 to 70.00 ± 0.2.  It is indicating that the beads prepared with lower concentration of polymer were lowest as compared to those prepared with higher concentration of polymer. The high concentration of polymers resulted in intense cross-linking. Hence, as the polymer concentration increases, it increases the drug entrapment. The test for uniformity of content was carried out to ascertain that the drug was uniformly distributed in the formulation and the result indicates that the prepare hydrogel was capable of producing hydrogel with uniform drug content. The mean particles size of hydrogel beads was found to be in a range of 102.36 ± 2.27 to 138.48 ± 2.25 µm. The surface morphology of drug loaded microspheres was studied by scanning electron microscopy and study revealed that hydrogel beads were spherical in shape and uniform in size.

The release of the entrapped drug from the hydrogel depends on the swelling behavior, because swelling is directly propositional to drug release in case of hydrogel. As the hydrogel swells, the pores of network open and release of the entrapped solute occurs. Therefore, the dynamic swelling study of the prepared beads was carried out in both phosphate buffer pH 1.2 and pH 7.4 and the results are shown in the range between 104.45 ± 0.3 to 145.92 ± 0.6 and 109.00 ± 0.9 to 168.00 ± 0.4 respectively.The swelling behavior of beads was expressed as the ratio of initial weight of beads to the final weight of swollen beads as a function of time. The swelling of the beads depends upon the concentration of polymer. The swelling of the beads increased with an increasing amount of polymer in beads.

Table No 1: Factorial Design

Factor

Level

   Low             Medium                       High         

X1

 3%                4%                          5%

X2

1:0.5                1:1                          1:2

X3

1:1                   1:2                            1:3

Transformed values

-1                      0                              1

Where,

X1: Concentration of CaCl2

X2: Drug: chitosan ration

X3: Drug: sodium alginate ratio

Amount of drug has been kept constant.

The in vitro drug release of the prepared hydrogel beads was found to be dependent on drug – polymer ratio, and concentration of cross linking agents. The chitosan was used as pH sensitive polymer, as a result chitosan was controlled the rate of drug release from hydrogel beads. In the drug release from these beads was characterized by an initial phase of high release (burst effect). The initial bust effect was considerably reduced with the increase the amount of polymer. The % cumulative drug release was in range between 62.10 ± 0.8to 90.58 ±0.2.

Based on the results of the percentage yield, entrapment efficiency, drug content, particle size, swelling study and in – vivo release profile study, best formulation (i.e. B -8) has been selected from the all prepared formulations for the further analysis. The optimized batch was subjected to enteric coating with Eudragit S 100.

Table No: 2 Characterization of hydrogel beads

Batch No

Percentage Yield

(%)

Drug Entrapment Efficiency (%)

Drug content

(%)

Mean particle size

(µm)

Swelling  study in pH 1.2  Phosphate buffer sol

Swelling study  in pH 7.4 Phosphate buffer sol

B1

92.28±0.2

25 ±0.3

10.44±0.6

120.40±0.5

136.00±0.3

138.15±0.3

B2

90.85±0.4

16.49±0.2

4.70±0.4

131.33±0.2

125.00±0.8

130.00±0.5

B3

88.88±0.7

42.27±0.7

9.30±0.7

120.40±0.7

124.15±0.5

136.20±0.2

B4

83.33±0.3

34.54±0.4

11.40±0.3

78.48±0.4

131.00±0.9

140.00±0.8

B5

63.40±0.6

43.20±0.3

10.8±0.5

135.10±0.2

109.60±0.2

122.20±0.6

B6

89.14±0.3

29.00±0.5

6.87±0.2

78.48±0.8

113.20±0.5

119.00±0.3

B7

62.90±0.5

40.80±0.7

10.20±0.8

131.33±0.6

122.90±0.7

135.00±0.5

B8

92.00±0.7

70.00±0.2

13.30±0.5

84.40±0.3

145.92±0.6

168.00±0.4

B9

73.06±0.2

53.30±0.5

8.84±0.2

120.40±0.5

110.95±0.3

121.00±0.7

B10

78.24±0.9

29.00±0.8

11.6±0.7

78.48±0.7

112.23±0.7

149.15±0.3

B11

88.57±0.4

36.10±0.4

10.30±0.4

135.10±0.3

124.55±0.4

109.00±0.9

B12

93.30±0.6

39.77±0.2

8.78±0.6

131.40±0.9

108.10±0.2

121.50±0.6

B13

79.86±0.3

28.18±0.6

9.30±0.3

84.40±0.3

121.30±0.5

129.00±0.2

B14

89.92±0.7

58.00±0.5

11.61±0.9

120.4±0.7

120.10±0.9

140.10±0.3

B15

87.71±0.2

37.02 ± 0.5

8.72 ±  0.2

78.48±0.2

126.35±0.3

139.00±0.5

B16

72.00±0.8

32.00 ± 0.3

8.20 ± 0.5

84.40±0.7

129.60±0.4

141.50±0.2

B17

81.60±0.5

47.50 ± 0.8

9.52 ± 0.3

78.48±0.3

125.60±0.5

141.10±0.7

B18

74.66±0.9

59.39 ± 0.4

9.82 ± 0.7

131.33±0.2

106.60±0.6

115.50±0.4

B19

82.88±0.3

16.62 ± 0.7

6.69 ± 0.4

120.40±0.6

112.70±0.7

120.00±0.6

B20

73.48±0.7

34.21 ± 0.3

9.76 ± 0.8

78.48±0.2

104.45±0.3

121.50±0.3

B21

89.30±0.4

56.50 ± 0.5

11.30 ±0.3

135.10±0.2

118.62±0.5

126.00±0.8

B22

85.20±0.2

31.21 ± 0.5

10.30 ±0.4

120.40±0.5

113.45±0.3

134.20±0.6

B23

89.00±0.5

32.40±0.4

8.16±0.7

78.48±0.1

127.25±0.7

140.90±0.4

B24

90.40±0.3

54.50±0.2

10.90±0.3

131.33±0.7

124.50±0.2

137.15±0.3

B25

84.70±0.6

38.00±0.3

9.52±0.6

84.40±0.4

119.65±0.5

131.80±0.9

B26

88.80±0.4

49.75±0.5

9.95±0.2

120.40±0.2

108.10±0.4

136.50±0.3

B27

68.00±0.7

39.19±0.3

9.21±0.5

84.40±0.4

113.00±0.7

129.00±0.5

The swelling study of optimized batch was shown that there was no swelling found after the enteric coating with Eudragit S 100. So the drug should not release in stomach and the degradation of drug in acidic pH was controlled. As expected, the beads remain intact during the acid step because the degree of ionization of carboxylic acid groups in the Eudragit S 100 increased with pH. Eudragit S 100 is fully dissolved and rapidly releases the drug from the core beads, whereas, at pH 1.2, the one is almost intact and retard the release of the drug. The drug release from the optimized batch was 92.61 ± 0.6 which shows that the drug releases from the enteric coated hydrogel beads provide a system of low permeability and a good barrier against drug diffusion under pH conditions, at which protection is required. The presence of enteric coating has a little effect on drug release but efficiently protect the acid labile drug from highly acidic environment of stomach. The drug release profile of optimized batch was shown in table No: 3.

Table No: 3 Release study of optimized batch

Time (hrs)

Batch 8

1

0

2

0

3

36.81 ± 0.4

4

54.43 ± 0.7

5

68.30 ± 0.5

6

78.58 ± 0.4

7

86.25 ± 0.3

8

92.61 ± 0.6

The surface morphology of beads was investigated by using scanning electron microscopy     (SEM) (JEOL JSM-5610LV). The surface morphology of hydrogel swollen beads was spherical in shape having dense surface. The minute pores were also found on the surface of the beads.

The release kinetics of drug release was studied by applying various pharmacokinetics models to in – vivo release profile. The data were plotted for zero order, first order, and higuchi’s model and for Korsemeyer’s – Peppas model. From the regression value for each model, it can be observed that release pattern follows the zero order kinetics rather than first order. From the results of Higuchi model and Korsemeyer’s – Peppas model, it can be observed that release mechanism of drug from hydrogel was followed anomalous diffusion/non fickinian refers to combination of both diffusion and erosion controlled release.

Release from the hydrogel kinetics of drug beads