INTRODUCTION:

Oral route is the most convenient and extensively used route for drug administration. In recent years, controlled drug delivery formulations and the polymers used in these systems have become much more sophisticated, with the ability to do more than simply extend the release period of the drug. These are formulated to release the drug’s active ingredient gradually and predictably over a 12-hour to 24-hour period. They provide greater effectiveness in the treatment of chronic disease conditions through more consistent delivery of medication, reduced side effects, greater convenience and higher levels of patient compliance^1,2.

Controlled drug delivery occurs when a polymer, whether natural or synthetic is properly combined with a drug or other active agent in such a way that the active medicament is released in a predesigned manner. Though these are useful, there are some disadvantages with these products like less absorption, possible toxicity and production of undesirable by-products. To overcome such problems, the delivery systems with prolonged gastric residence time, such as floating drug delivery system was introduced³. This system belongs to the class of gastro-retentive dosage forms, which get retained in the stomach for longer periods of time, thus helping in absorption of drug for the intended duration of time. The widely employed method in this is the floating systems⁴. Mostly the floating systems were prepared by using the non-effervescent technique. Freeze-dried calcium alginate beads were employed which works on the basis of formation of a colloidal gel barrier. When microspheres come in contact with the gastric fluid, the gel formers like polysaccharides and polymers hydrate to form a colloidal gel barrier that controls the rate of fluid penetration into the device and consequent drug release. As the exterior surface of the dosage form dissolves, the gel layer is maintained by the hydration of the adjacent hydrocolloid layer. The air trapped by the swollen polymer lowers the density and confers buoyancy to the microspheres. However a minimal gastric content is needed to allow proper achievement of buoyancy⁵. Spherical beads of approximately 2.5 mm diameter can be prepared by dropping sodium alginate solution into aqueous solution of calcium chloride, causing precipitation of calcium alginate leading to formation of porous system, which can maintain a floating force for over 12 hours. These floating beads will give a prolonged residence time of more than 5.5 hours⁶. The floating drug delivery systems were used now-a-days in treating local inflammation, peptic ulcer and Helicobacter pylori associated ulcers. In chronic diseases associated with frequent and prolonged medication, this system can ensure promising drug delivery. Hollow microspheres of acrylic resins, eudragit, polyethylene oxide with cellulose acetate, polycarbonate and gelucire floating granules are the recent developments. Although different techniques like single emulsion, double emulsion, polymerisation, spray drying, solvent extraction were available, the phase separation co-acervation technique is used mostly. It is specially designed for preparing the reservoir type of system. The main principle involved in this process is formation of coacervates. Decreasing the solubility of polymer in the organic phase affects the formation of a polymer rich phase called coacervates. Different coacervation techniques include, solvent addition, non-solvent addition, addition of an incompatible polymer and change in pH.

In recent years, micro spheres have been proposed for treating many diseases needing a constant drug concentration in blood or drug targeting to specific cells or tissues. Microspheres are solid, spherical particles containing dispersed drug molecules either in solution or microcrystalline form with their size ranging from 50nm to 2mm. These are characteristically free flowing powders consisting of proteins or synthetic polymers, which are biodegradable in nature. Micro spheres can be targeted to a particular organ, a specific part of the organ or to a selective intracellular site⁷. Pantoprazole which is en effective agent in reducing the gastric acid secretion by inhibiting proton pump in the gastro-intestinal tract is used here⁸. The current work focused on the formulation of Pantoprazole sodium floating microspheres and evaluation of various characters of microspheres like invitro drug release, incorporation efficiency and buoyancy percentage.

MATERIALS AND METHODS:

Pantoprazole sodium was a gift sample from S.S. Medical agencies, Hyderabad, poly vinyl pyrrolidine and hydroxyl propyl methyl cellulose was gift samples obtained from Pellets Pharma Ltd., Hyderabad. Tween 80, sodium bicarbonate, sodium alginate, calcium chloride and glacial acetic acid were procured from S.D. Fine Chemicals, Mumbai. All other materials used were of analytical grade and procured commercially.

Analytical method

Estimation of Pantoprazole:

In the recent investigation, a simple, sensitive and more accurate spectrophotometric method was used for the estimation of Pantoprazole ⁹. The absorbance values of Pantoprazole were measured at a λ_max of 260 nm.

Preparation of Pantoprazole Standard Dilutions:

Aliquots of Pantoprazole stock solution (1 mg/ml) was transferred into 5 volumetric flasks and was further diluted with 1.2 pH and 6.8 pH phosphate buffers so as to get the standard dilutions of 2, 4, 6, 8 and 10 µg/ml. The absorbance values of the above dilutions were measured in ELICO double beam UV spectrophotometer at 260 nm using 1.2 and 6.8 pH phosphate buffers as blank separately. The absorbance values were plotted against concentrations of Pantoprazole and from the corresponding calibration curves, the R²values were obtained.

Preparation of Pantoprazole sodium sesquihydrate microspheres

In the present study, Pantoprazole sodium sesquihydrate microspheres were prepared by co-acervation technique as employed by many of the researchers¹⁰. 50 mg of Pantoprazole was dissolved in 5 ml of distilled water. Then a polymer solution was made by mixing 810 mg of sodium alginate with 90 mg of required polymer (HPMC, PVP e.t.c) in 30 ml of distilled water. Then these two solutions were mixed. To this solution, sodium bicarbonate was added in required concentration and stirred well with the help of a magnetic stirrer for 15 minutes at medium speed. The solution was allowed to set for 10 minutes, so that the air bubbles were subsided. This solution was poured into a solution of 1% calcium chloride in 10% glacial acetic acid slowly with the help of a syringe and kept aside for some time. The microspheres were washed thoroughly to remove excess of acid and dried at room temperature or by using a hot air oven and stored in a well closed container. By using these microspheres, various formulations were made using different polymers in increasing concentrations which was indicated in table 1.

Table 1: Composition of Pantoprazole microspheres

S. No	Formulation code	Polymer ratio (PVP / HPMC)
1	P1	1:0.5
2	P2	1:1
3	P3	1:1.5
4	H1	1:0.5
5	H2	1:1
6	H3	1:1.5

Characterisation of Pantoprazole floating microspheres

Pantoprazole microspheres of various polymeric concentrations were prepared by coacervation technique. Various physico-chemical properties that are evaluated for the Pantoprazole microspheres containing various concentrations of the polymers (PVP and HPMC) include particle size, dissolution, incorporation efficiency and buoyancy percentage.

Particle size analysis

Particle size analysis was carried out by using the optical microscopy method with the help of a calibrated eye piece micrometer¹¹. The size of around 200 particles was measured and an average diameter was calculated.

Invitro dissolution studies¹²

Dissolution studies were carried out for the determination of rate of drug release and solubility. A USP dissolution test apparatus Type-I was used to determine the dissolution profiles of the prepared Pantoprazole microspheres. The dissolution medium used was 900 ml phosphate buffer of pH 1.2 for the first 2 hours. After 2 hours, the microspheres were filtered and the dissolution process was continued by placing these filtered microspheres in a freshly prepared 900 ml phosphate buffer of pH 6.8 up to 12 hours equilibrated to 37⁰C. The basket was rotated at a speed of 100 rpm. From the dissolution flask, 5 ml samples were withdrawn at various time intervals such as 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12 hours. Concentration of Pantoprazole in the samples was determined by UV-Visible spectrophotometer at 260 nm. The amount of Pantoprazole dissolved was calculated from the concentration.

Incorporation efficiency [IE]

To determine the incorporation efficiency, microspheres (100 mg) were, thoroughly crushed by triturating in mortar and pestle and suspended in a minimal amount of ethanol for dissolving coat shell of microspheres. The suspension was suitably diluted with water and filtered to separate the shell fragments. The drug content was analysed after suitable dilution spectrophotometrically at 260 nm. The amount of drug incorporated in microspheres was calculated by the following formula

Buoyancy percentage¹³

An in vitro buoyancy study was conducted by spreading 500 mg microspheres over the surface of a USP XXIV dissolution apparatus (Type II) filled with 900 ml of 0.1 M acidic solution containing 0.02% tween 80 as a dispersing medium. The medium was agitated with a paddle rotating at a speed of 100 rpm for 12 hours. After each time interval, the floated microspheres were collected, dried and weighed. Buoyancy percentage was calculated using the formula;

RESULTS AND DISCUSSION:

The calibration curve of Pantoprazole showed a regression value of 0. 999 in both media of phosphate buffer with pH 1.2 and 6.8, which indicate the regression, are within the range.

Characterisation of Pantoprazole sodium floating microspheres

Particle size analysis

The particle size of floating microspheres varied among the formulations due to variations in the composition of formulations. Formulations P3 and H3showed relatively large size, whereas the formulations P1 and H1showed relatively small size floating microspheres. The results were indicated in table 2.

Table 2: Particle size analysis of the Pantoprazole microspheres

S. No	Formulations	Particle size range (µm)
1	P1	20 – 30
2	P2	25 – 40
3	P3	40 – 60
4	H1	15 – 30
5	H2	20 – 40
6	H3	30 – 60

Invitro dissolution studies

Microspheres were subjected to in vitro release studies using USP dissolution apparatus type I in 900 ml phosphate buffer solution of pH 1.2 for about 2 hours and 900 ml of phosphate buffer solution of pH 6.8 up to 12 hours. The P2 formulation showed sustained release of the drug and the drug release was found to be approximately linear. Furthermore, the drug release from the floating microspheres matrix was controlled by the polymer. As the polymer content was increased and the drug loading was decreased, the release of the drug was decreased significantly. The results of dissolution profiles of Pantoprazole microspheres with PVP in different concentrations were shown in table 3 and indicated in figure 1.

Table 3: Dissolution data of Pantoprazole microspheres obtained from different concentrations of PVP

Time (hours)	Cumulative % drug release
Time (hours)	P1	P2	P3
0	0	0	0
1	1.6	2.8	0.96
2	6.7	8.2	3.4
3	12.8	15.4	8.9
4	19.4	20.1	14.4
5	21.6	25.6	20.4
6	29.8	33.8	26.8
7	37.2	46.2	39.9
8	48.3	58.3	48.2
9	59.2	64.4	54.9
10	67.9	76.2	66.2
11	76.6	83.4	74.3
12	84.6	92.8	80.2

Fig 1: Comparative drug release profiles of Pantoprazole microspheres with different concentrations of PVP

By performing the in vitro dissolution studies, it was found that among the prepared formulations, the H2 formulation showed sustained release of the drug and the drug release was found to be approximately linear. As the polymer content was increased and the drug loading was decreased, the release of the drug was found to be decreased significantly and the results were indicated in table 4 and figure 2.

Table 4: Dissolution data of Pantoprazole microspheres obtained from different concentrations of HPMC

Time (hours)	Cumulative % drug release
Time (hours)	H1	H2	H3
0	0	0	0
1	0.8	1.9	0.6
2	4.5	6.6	2.1
3	12.6	13.4	10.3
4	20.1	22.6	16.4
5	28.4	30.6	24.6
6	32.6	41.4	32.8
7	41.4	50.3	40.3
8	46.2	55.2	48.4
9	50.1	63.9	53.2
10	56.4	72.8	61.9
11	60.2	80.7	66.4
12	68.9	88.8	72.8

Fig 2: Comparative drug release profiles of Pantoprazole microspheres with different concentrations of HPMC

Incorporation efficiency [IE]

Incorporation efficiency tests were carried out for the prepared formulations. Among them, P2 and H2showed higher percentage of incorporation efficiency. As smaller the microspheres, more will be the buoyancy percentage and more sustained release of the drug was achieved. While larger the size, less will be the buoyancy percentage and there was no sustained release of the drug. The results were indicated in table 5.

Table 5: Percentage Incorporation Efficiency (IE) of Pantoprazole floating microspheres

S. No	Formulations	% Incorporation Efficiency (IE)
1	P1	81.4
2	P2	91.2
3	P3	70.4
4	H1	62.2
5	H2	82.6
6	H3	66.4

Buoyancy percentage

The floating test was performed to investigate the floatability of the prepared microspheres. Good in vitro percentage buoyancy was observed for formulations P2 and H2. This may be attributed to the low tapped density of the microspheres. Microspheres of formulations P2 and H2showed best floating ability (72% and 69%) when compared to other formulations. The buoyancy percentage of various Pantoprazole formulations was given in table 6.

Table 6: Buoyancy percentage of Pantoprazole floating microspheres

S. No	Formulations	Buoyancy percentage
1	P1	63.4
2	P2	72.3
3	P3	60.9
4	H1	58.1
5	H2	69.2
6	H3	52.3

CONCLUSION:

For better absorption and enhanced bioavailability of some drugs, prolonged retention time of the dosage form in the stomach is essential. This problem can be solved by the preparation of gastro – retentive drug delivery systems. An attempt was made to prepare floating microspheres of Pantoprazole using polymers of PVP and HPMC by coacervation technique. From the obtained results, it was concluded that the drug release from the floating microspheres matrix was controlled by the polymer. When the polymer proportion in the formulation was increased with decrease in drug loading, drug release was decreased significantly. The nature of polymers and their concentration influenced the physical and floating behaviour of prepared microspheres. In vitro release data obtained from buoyant microspheres showed good buoyancy and prolonged drug release for formulations P2 and H2. The prepared microspheres have different size and incorporation efficiency. Hence the formulations P2 and H2 showed appropriate balance between buoyancy and drug release rate. Diffusion was found to be the main release mechanism. Thus the prepared microspheres proved to be potential candidates for multiple–unit delivery devices adaptable to any intra gastric conditions.

ACKNOWLEDGEMENTS:

The authors are thankful to M/s. S. S. Medical Agencies, Hyderabad for their generous supply of Pantoprazole pure drug as a gift sample. The authors thank the management of Sri. P. Rami Reddy Memorial College of Pharmacy, Kadapa for their sheer support to conduct this work.

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Received on 10.07.2014 Modified on 05.08.2014

Res. J. Pharm. Dosage Form. and Tech. 6(3):July- Sept. 2014; Page 218-222