Preparation and In Vitro Evaluation of Aceclofenac Loaded Eudragit Microspheres

Pankaj Pimpalshende^1*, SS Burle¹ and C Vijay²

¹Dept of Pharmaceutics, Hi-Tech College of B Pharmacy, Chandrapur, (Maharashtra)

² Ultra college of pharmacy Madhurai, Tamilnadu

ABSTRACT

KEYWORDS:

INTRODUCTION:

Microspheres are homogenous, monolithic, spherical polymeric particle with size ranging from 0.1 to 1000 µm and are widely used as drug carriers for controlled release. Administration of drug in the form of microspheres usually improves the treatment by providing the localization of active substance at the site of action and by prolonging release of drug.¹Microcapsules are vesicular systems in which the drug molecules are surrounded by a membrane. Microspheres are matrix system in which the drug molecules are dispersed throughout the particle.² Microspheres are characteristically free flowing powder consisting of protein and synthetic polymer. The drug is released slowly at desired rate with reduced fluctuation in plasma drug concentration.³ The solvent evaporation is carried out by maintaining emulsion at reduced pressure or by stirring the emulsion so that the organic phase evaporates out the solid microspheres are obtained by filtration and washing. Solvent evaporation techniques offer a versatile, easy, practical method for manufacturing of microspheres.

Aceclofenac is a non-steroidal anti-inflammatory drug. NSAIDs are chemically heterogeneous group of compounds including the sub-class of Cox-2 inhibitor. The most available traditional NSAIDs act by inhibiting the prostaglandin synthases enzymes known as the cyclooxygenases. NSAIDs are considered to be the first line drugs in the symptomatic treatment of rheumatoid arthritis, ostoarthritis and ankylosing spondilitis. Aceclofenac is one of the emerging NSAIDs molecules for arthritis treatment. It is a newer derivative of diclofenac and has less gastrointestinal complications. The successful treatment of arthritis depends on the maintenance of effective drug concentration level in the body for which a constant and uniform supply of drug is desired. Sustained release dosage forms deliver the drug at a slow release rate over an extended period of time and achieve this objective. The short biological half life (about 4 hour) of Aceclofenac makes it an ideal candidate for sustained release.⁷ Aceclofenac is well tolerated, with adverse events being minor. The most common events include abdominal pain, gastritis, ulcerative stomatitis when given orally. Sustained and targeted delivery of Aceclofenac will reduce these toxicities considerably by maintaining a low and constant level of drug in blood. Hence it was planned to prepare microspheres using the pH – independent carrier Eudragit and evaluate them for sustained release.

MATERIALS AND MATHODS:

Aceclofenac as a gift sample from FDC Limited, Mumbai, Eudragit RL 100 (Rhome Pharma Industry), and all other reagents and chemicals used were of analytical grade, Electronic balance (Shimadzu), Magnetic stirrer (2MLH Remi Industries limited), Compound microscope (Binocular micron optic), Micrometer, Sieves (Jayant synthetic ltd., Mumbai), Vaccum oven (Scientific lab), Scanning electrone microscope (HITACHI) M 3000, UV–Visible spectrophotometer (Systronic 2201), Dissolution test apparatus (paddle type), (DISSO 2000) (LAB INDIA)

Preparation of Microsphares:¹¹^-13

Aceclofenac microspheres were prepared by solvent evaporation method. The drugs to carrier ratio (1:1, 1:2, 1:3) of carriers such as Eudragit RL 100 were used for preparation of three formulations. (Table No.1) Eudragit polymer were dissolved in 25 ml of acetone.1g of drug was added to this solution. The solution of organic phase was slowly poured in to 500 ml beaker containing 200 ml liquid paraffin and 1 % v/v span 80 as emulsifying agent with the help of 5 ml syringe and stirred in a magnetic stirrer for 3 hour at a controlled stirring speed of 300 rpm, at 40^oC temperature maintained. After the complete evaporation of solvent microparticles were collected by filtration, washed 3 times with petroleum ether and dried for 24 hour at room temperature and kept in desiccators.

Percent Yield:

Microspheres dried at room temperature were weighed and yield of microspheres preparation was calculated by using formula.

Size distribution of Microspheres:¹⁴^-16

Size distribution of the microspheres was determined by using standard test sieves. Microspheres retained on the sieves were collected and weighed and the distribution was analyzed based on the weight fraction on each sieve.

Surface morphology of Aceclofenac Microspheres:¹⁷^-19

Morphology of the microspheres was studied by scanning electron microscopy (SEM). The samples for SEM were prepared by lightly sprinkling the microparticles on a double adhesive tape stuck to a aluminum stub. The stubs were then coated with gold. The coated samples were then randomly scanned and photo-micrographs were taken with a scanning electron microscope (HITACHI MS 3000 H)

Drug Encapsulation:^20-21

Microspheres equivalent to Aceclofenac were weighed and crushed in mortar pastle and dissolved in 50 ml phosphate buffer solution pH (6.8) and stirred for 8 hour in magnetic stirrer. The solution was filtered and the absorbance of the filtered liquid was measured in a uv-spectrophotometer at 275 nm wavelength using pH 6.8 phosphate buffers as a blank solution and the amount of drug present in microspheres was determined. Caliberation curve was generated at same

In vitro drug release studies of the prepared aceclofenac microspheres:^{18, 19, 21}

The drug release studies of the microspheres were carried out upto 12 hours using dissolution test apparatus (The rotating paddle method). The drug release medium used for the studies was phosphate buffer of pH 6.8; 500 ml of phosphate buffer was taken as the dissolution medium. Microspheres containing equivalent of aceclofenac was accurately weighed and placed in the vessel. The paddle was rotated at 100 rpm. The dissolution medium was thermostatically controlled at 37^oC. Sample of 1ml was withdrawn from the dissolution medium at suitable time interval. The sample volume was replaced by equal volume of fresh medium. The withdrawn 1 ml samples were made upto 5 ml and analysed spectrophotometrically at a wave length of 275 nm. The release study of pure sample of aceclofenac was also carried out by the same manner.

RESULT AND DISCUSSION:

Preparation of Microspheres:

Solvent evaporation method was used to prepare aceclofenac micrsospheres. Three formulations of aceclofenac loaded microspheres were prepared using Eudragit RL 100 as carriers by solvent evaporation technique. The drug to carrier ratio 1:1, 1:2, 1:3 in that polymer.

Percent Yield:

The yields of preparation of aceclofenac microspheres are high for all microspheres obtained which are about 80%. It shows that the recovery of the material is good which is significant from the economical view. The yield of microspheres is also not dependent on polymer proportion selected or the polymer nature.

Particle size distribution:

Most of the microspheres were collected on sieve of 210 µm in all formulation. The particles weight size distribution and mean particle size of the aceclofenac microspheres (Lackman) are shown in Table 1 respectively.

Scanning Electron microscope analysis:

Shapes and surface characteristics of the microspheres were investigated and photographed using scanning electron microscope shown in Fig 1 morphological characteristic dependent on type of polymer used.Eudragit RL polymer used to prepared microsphere were spherical shape, its Micrographs show regular microparticles (x100). Microspheres were agglomerated tending to stick together owing to remaining residue of aceclofenac in their surface but redispersion characteristics were not affected the microparticles in wide size range could the observed in SEM Fig Almost spherical, porous, rough surface with the dispersed drug could be clearly noticed.

Fig 1 Scanning Electron microscope.

Table –2: In vitro drug release studies data for Batch 1 (1:1)

Sl. No.	Time	Amount released	Cumulative % released
1	0.5	3.4	17 ± 1.0
2	1	4.3	21.5 ± 1.5
3	1.5	4.9	24.5 ± 0.5
4	2	5.8	29 ± 1.0
5	3	6.9	34.5 ± 2.5
6	4	8.1	40.5 ± 2.5
7	5	8.3	41.5 ± 1.5
8	6	9.3	46.5 ± 4.5
9	7	10.5	52.5 ± 1.5
10	8	11.9	59.5 ± 1.5
11	9	13.7	68.5 ± 1.5
12	10	15.6	78 ± 1.0
13	11	17.1	85.5 ± 0.00
14	12	18.6	93 ± 1.0

In Vitro drug release study:

The dissolution study was carried out for 12 hours for all formulations. The drug released study was carried out by dissolution apparatus paddle method. Microspheres were studied at pH (6.8) phosphate buffer. The aceclofenac microspheres were analysed for in vitro drug release estimation the result are given in Table No 2, 3, 4. The drug release from microspheres was depending on polymer ratio used in preparation of microspheres, effect of drug: polymer ratio on the release. Aceclofenac release rate from microspheres was dependent on the ratio used a shown in Figure No 2. Aceclofenac release rates from Eudragit RL microspheres were near slow and the amount released in 12 hour reach 93% for Ratio 1:1 (Eudragit RL100), 89% for Ratio 1:2 (Eudragit RL100), 80% for Ratio 1:3 (Eudragit RL100). This is due to the higher water permeability of Eudragit RL100 having 10% of functional quaternary ammonium group.⁸ Since Eudragit RL is highly permeable drug release it’s also higher shows in fig 2.

FIG 2: IN VITRO DRUG RELEASED GRAPH OF ACECLOFENAC WITH HIGUCHI PLOT

Table –3: In vitro drug release studies data for batch 2 (1:2)

Sl. No.	Time	Amount released	Cumulative % released
1	0.5	3.9	19.5 ± 1.5
2	1	5	25 ± 4.0
3	1.5	5.6	28 ± 3.0
4	2	6.7	33.5 ± 4.5
5	3	7.6	38 ± 5.0
6	4	8.1	40.5 ± 5.5
7	5	9.7	44 ± 4.0
8	6	10.4	53 ± 7.0
9	7	11	59.5 ± 6.0
10	8	11.9	68 ± 4.5
11	9	13.6	69 ± 6.0
12	10	15.2	76 ± 4.0
13	11	16.5	82.5 ± 4.5
14	12	17.9	89.5 ± 5.5

Table –4: In vitro drug release studies data for batch 3 (1:3)

Sl. No.	Time	Amount released	Cumulative % released
1	0.5	2.2	11 ± 1.0
2	1	3.3	16.5 ± 2.5
3	1.5	4.2	21 ± 4.0
4	2	5.1	25.5 ± 5.5
5	3	5.9	29.5 ± 6.5
6	4	6.8	34 ± 6.0
7	5	7.5	37.5 ± 5.5
8	6	8.9	44.5 ± 7.5
9	7	10.2	48.5 ± 7.5
10	8	11.2	56.5 ± 6.0
11	9	12.3	61.5 ± 4.5
12	10	13.7	68.5 ± 1.5
13	11	14.8	75 ± 4.0
14	12	16.1	80.5 ± 1.5

Mechanism of drug release:

Constant drug release over time could not be obtained as the drug release is through the matrix microcapsules. This is proved by the straight line Higuchi plots. Obtained with Higuchi upto 12. After this time an increased drug release was observed which may be due to the erosion of microspheres. Hence it can be concluded that the release is diffusion with erosion after shows. This will be helpful in compensating the reduced drug release rates at the end.

CONCLUSIONS:

Aceclofenac microspheres were prepared successfully using solvent evaporation technique. Different drug : polymer ratio and stirring speed of the system were important to obtain almost spherical particles, also depend on solubility of polymer in organic solvent also the production yield of preparation of microspheres and encapsulation efficiencies were very high, but polymer ratio increased the encapsulation efficiencies were increased. Shape and size of microspheres was determined using scanning electron microscopy with photographs.

The drug : polymer ratio all Batch’s selected as a best ratio because its percent yield of that ratio nearly achieve 83% and drug encapsulation efficiency was very high and drug release should be up to 93%, to achieved microspheres for controlled released of aceclofenac microspheres formulation reduced dosing frequency, decrease side effect and improved patient compliance.

ACKNOWLEDGEMENT:

Authors thank to principal, ultra college of pharmacy for providing facilities to carry out present research work

REFERENCE:

1. Donald L. Wise, Hand book of pharmaceutical controlled release technology, 1^st edition, Marcel Dekker 2005, p. 329.

2. Remington, the science and practice of pharmacy, 20^th edition, Philadelphia, 2000, p. 924.

3. S.D. Vyas and Roop Khar: Targeted and controlled drug delivery system, Satish Kumar Jain, 2002, p. (418)³ (419-427)³⁸.

4. Linazhang, Release behavior of microspheres from cross-linked. N-methylated chitosan encapsulated ofloxacin, Carbonhydcate polymers 65(2006) 288-295.

5. Tanwar Y.S., Floating Microspheres : Development, characterization and application, www.pharminfo.net 2006

6. Roopkhar, S.P. Vyas, controlled drug delivery, concepts and advances, 1^st edition, MK Jain, 2002, p. 98.

7. Mutali. S, preparation in vitro, preclinical and clinical evaluation of once daily sustained release tablets of Aceclofenac, Arch pharma res vol. 30, No.2, 2007 pp. 222-234.

8. Indian pharmacopocia, controller of publication, Delhi, 1996, Vol. (2) p. A144.

9. Prof. Saraf. S, The pain management is always a problem for a physician and the search for a safe and effective option in still on, 2006, www.pharminfo.net.

10. Hand book of pharmaceutical excipient, fourth edition 4^th p. (462-468)⁹ p. (234-241)^10b, p (591-595)^10c

11. Jamagondil L.N., Patil V; Pokharkar V: studies on microcapsulation of metformin HCl using ethyl cellulose, Indian Journal of pharmaceutical science, 2004, p. 494.

12. Jain G.K; Preparation and Evaluation of Ethambutol Microspheres, Indian journal of pharmaceutical science, 1990, p. (153-154).

13. Asha Patel, Subhabrata Ray, Ram Sharnagah Thakur, Invitro evaluation and optimization of controlled release floating drug delivery system of metformin hydrochloride, DARU vol.14, No.2, 2006, p.(57-64).

14. Jayakrishnan .A; A floating type oral dosage form for piroxicam based on hallow polycarbonate microspheres in vitro an in vivo evaluation in rabbits, Journal of controlled release 79(2002) 71-79.

15. Afred Martin, Jamer swarbrick, Arthur commarate; physical pharmacy, 3^rd edition, Varghese publication 1991, p.503.

16. Lachman .L, the theory and practice of Industrial pharmacy, 3^rd edition, Varghese publication, 1997, p. 27-29.

17. Negro .S; Vitamin A palmitate and aciclovic biodegradable microspheres for intraocular sustained releae; International journal of pharmaceutics 326 (2006), p. 100-106.

18. Kawashima Yashiaki; Microspheres design for the colonic delivery of 5-fluorouracil. Journal of controlled release 90(2003) p. (313-322)

19. Muthuswami K.; preparation and evaluation of lansoprazole floating micropheres, Indian journal of pharmaceutical science 67(1), 2005, p. 75-79.

20. Shrivastavo .A.K; Floating microspheres of limetidin; formulation characterization and in vitro evaluation, Acta pharma, vol. 55, 2005, p. (277-285)

21. Kas, H.S; Diclofenac sodium incorporated PLGA (50:50) microspheres: formulation consideration and in vitro / in vivo evaluation. International journal of pharmaceutical 195 (2000) p.179-188.

22. Agarwal G.P; Calcium cilicate based microspheres of repaglinide for gastroretenltive floating drug delivery system: preparation and in vitro characterization, Journal of controlled release 107 (2005) p. (300-309).

23. Fell .T; Amoxicillin releae froma floating dosage form based on alginates, International journal of pharmaceutics, 210(2005) p.45-49.

24. Chowdary K.P.R; Ethylcellulose Microspheres of glipizide : Characterization, in vitro and in vivo evaluation, Indian Journal of pharmaceutical sciences 2004, 66(4) : 412 – 415.

25. Sweta Shah, Roula Qurish, Vijay patel and Mansoor Amiji, Evaluation of the factors influencing stomach, specific delivery of Antibacterial Agents for Helicobacter pylori infaction, J. Pharm, Pharmacol. 1999 51: (667-672).

26. Marchetti, J.M, Nimusulide PLA microspheres as a potential sustained release system for the treatment of inflamatory disease, international journal of pharmaceutics 295(2005) 201-211.

27. Pothal R.K, preparation and Evaluation of mucoadhesine microcapsules of theophylline, The Indian pharmacist 2004 (74-78)

28. Nagoji, Comparative study of in vitro drug release from Ethyl cellulose coated spherical matrices and micro capsules o Nimesulide, Eastern pharmacist – 2000 (119 – 120).

29. Nokhodchi Ali, The microsponge delivery system of benzoyl peroxide: prepartion characterization and release studies, International journal of pharmaceutics 308(2006) 124-132.

30. Jamagondi .L Patil V, studies on micro encapsulation of Metformin HCL using ethyl cellulose, Indian Journal of pharmaceutical science 2004, (494).

Received on 24.02.2010

Accepted on 14.04.2010

Research Journal of Pharmaceutical Dosage Forms and Technology. 2(2): March –April. 2010, 169-172

Batch no	Drug:Polymer Ratio	%V/V SPAN80	% yield	Ave size in µm	Drug content	%cumulative Release	R2 value for 12 hour
1	1:1	1%	82%	231 µm	84%	93 ±1.0	0.9718
2	1:2	1%	83%	225 µm	80%	89.5±5.5	0.9635
3	1:3	1%	77%	227 µm	82%	80.5±1.5	0.9824