Formulation and Evaluation of Imatinib Mesylate Microspheres by Chemical Crosslinking Method

 

S.P. Senthil¹*, K.L. Senthilkumar², Sadasiva Reddy Chandi¹, R.P. Ezhilmuthu²,

M.M. Saravanan³. Nagesh R. Sandu²,

¹Department of Pharmaceutics, The Erode College of Pharmacy and Research Institute, Veppampalayam, Erode, Tamil Nadu, India-638112.

²Padmavathi College of Pharmacy and Research Institute, Dharmapuri, Tamil Nadu, India.

³Vinayaka Mission’s College of Pharmacy, Salem, Tamil Nadu, India-636008.

*Corresponding Author E-mail: senthilumasenthil@yahoo.co.in

 

The present research deals with formulation of microspheres containing an Anti-cancer drug Imatinib mesylate reduce the frequency of dosing. It is a protein-tyrosine kinase inhibitor, especially useful in the treatment of various types of cancer and can also be used for the treatment of atherosclerosis, thrombosis, restenosis, or fibrosis. Imatinib mesylate loaded microspheres were formulated by using both hydrophilic and hydrophobic polymers by Chemical Cross Linking method with Chitosan, and Ethyl Cellulose to develop a sustained release dosage form. The effect of concentration of cross-linking agent (Glutaraldehyde) on the microspheres properties like percentage of drug loading, biodegradability, drug release kinetics, particle size, encapsulation efficiency, angle of repose, bulk density, SEM, DSC and XRD were investigated in our study. Moreover, the kinetics of Imatnib mesylate released from different formulations of microspheres were analyzed using four different theoretical models, that is, Zero order, First order, Peppa’s, and Higuchi models. Microspheres prepared with Glutaraldehyde showed different release kinetics. Increasing the polymer concentration decreased the release rate of Imatinib Mesylate from microspheres because of formation of greater structural strength and more tightly texture with the drug. Besides, microspheres gave an adequate fit to either zero order or first order kinetic models, depending on the extent of cross linking reaction between drug and the cross linking agent.

 

 

INTRODUCTION:

The microparticles delivery system includes microcapsules, pellet, emulsions, microspheres, lipospheres, etc., generally given in the form of oral or topical solutions. Different types of coated particles may be obtained depending on the coating process used. The particles can be surrounded within a polymeric or proteinic matrix complex in either as solid aggregated state or a molecular dispersion, resulting in the formulation of microspheres. Alternatively, the particles can be coated by a solidified polymeric or proteinic envelope, leading to the formation of microcapsules. Imatinib Mesylate is currently registered in adults for two indications :( a) monotherapy in Chronic Myeloid Leukemia (CML) and (b) Metastatic Gastrointestinal Stromal Tumors (GISTs).

 

Microspheres are defined as homogenous, monolithic particles in the size range of about 1.0– 1000 μm and are widely used as drug carriers for Sustained release^{1, 4}. Administration of the drug in the form of microspheres usually improves the action by providing the localization of the active substance at the site of action and by sustaining the release of drugs. Glutaraldehyde was used as cross-linking agent to extent the release of the drug from the formulation. It may therefore be more advantageous to deliver this drug in a sustained release dosage form. The present study was focused on development of sustained release Imatinib mesylate using Chemical Cross linking method.

 

MATERIALS AND METHODS:

Materials:

Imatinib Mesylate was procured as a gift sample from Natco pharma limited, Mumbai (India).Chitosan² and Ethyl cellulose was obtained from LOBA chemicals, Kolkata. All chemicals were of analytical grade and were used without further purification.

 

Figure No.1 Cumulative Drug Release Graph

 

Method of preparation^{2, 3}:

A 4.0%(w/v) Polymer solution in aqueous acetic acid(5.0%) was prepared. This dispersed phase was added to continuous phase (100 mL) consisting of light liquid paraffin and 25ml of Petroleum ether containing 8.0mlof Span 80 in a beaker at room temperature. Stirring was continued at 2000rpm using a 3- blade half moon paddle for 5 minutes (Remi Equipments, Mumbai, India). A drop-by-drop solution of a measured quantity (2.5 mL each) of aqueous glutaraldehyde (25% v/v) saturated with toluene was added at 15, 30, 45, and 60 minutes. Stirring was continued for 2.0 hours and separated by centrifugation and washed, first with petroleum ether (60°C-80°C) four times, once with acetone and then thrice with distilled water to remove the adhered liquid paraffin and glutaraldehyde, respectively. The microspheres were then finally dried at room temperature and stored in vacuum desiccators.

 

Particle size analysis:

The microsphere size distribution was determined by the optical microscopy method using a calibrated stage micrometer (μm).

 

Compatibility studies^{14, 15}:

One of the requirements for the selection of suitable carrier for pharmaceutical formulation was compatibility. Therefore in the present work a study was carried out by using FTIR spectrophotometer to find out if there are any possible chemical interactions between Imatinib mesylate, Ethyl Cellulose, Chitosan.

 

Angle of Repose¹¹:

The fractional force in the loose powder can be measured by the angle of repose. Angle of repose was calculated by static method using fixed funnel method. This was the maximum angle possible between the surfaces of the site of the powder to the horizontal plane.

 

Determination of Bulk Density¹¹:

The bulk density was determined by 3-tap method. Bulk density is defined as, “the mass of powder divided by the bulk volume”. The packing characteristics of the powder play an important role in determining physical properties of product.

 

Determination of Drug Content⁴:

Accurately weighed microspheres equivalent to 25mg of Imatinib mesylate, crushed in glass mortar and pestle and the powdered microspheres were suspended in 100 ml of 0.1N HCl. After 24 hours, the solution was filtered and the filtrate was analyzed for the drug content.

 

Encapsulation Efficiency¹²:

Encapsulation efficiency was calculated using the following formula;

 

In- Vitro Dissolution Studies^9,10:

The drug release study was performed using USP XXIII dissolution test apparatus, the capsules filled with equivalent amount of Imatinib mesylate 100 Mg was placed in a basket. The instrument was set at 100-rpm rotation and at 32⁰C, 900 ml of 0.1N Hcl was filled as dissolution medium for first 2 hours and phosphate buffer pH 7.4 from third hour onwards. Samples were withdrawn at predetermined intervals, from 0.5 – 24 hours filtered and analyzed spectrophotometrically at 230nm using corresponding medium as blank. After each withdrawal, the same quantity of fresh medium was replaced immediately.

Stability Studies¹⁶:

The Optimised preparation was divided into 3 sets and was stored at 4°C (refrigerator), room temperature and 40°C (thermostatic oven). After 15, 30, 60, 90 and 180 days drug content and SEM of the formulation was determined.

 

Kinetic Characteristics of the Drug Release^5-8:

To know the mechanism of the drug release from the microspheres, the results obtained from the In-vitro dissolution process were fitted into different kinetic equations as follows:

1.     Zero - order drug release: Cumulative % drug release Vs Time.

2.     First Order drug release: Log cumulative % drug retained Vs Time.

3.     Higuchi’s classical diffusion equation: Cumulative % drug release Vs Square root of time.

4.     Peppa’s Korsemeyer Exponential equation: Cumulative % drug release Vs Log time.

5.     ‘n’ values can be used to characterize diffusion release mechanism as :

n > 0.5	Fickian diffusion
0.5 < n < 1	Non-Fickian diffusion
n > 1	Class – II transport

 

Figure No.2 .SEM Photograph

 

 

Table No.1 Design of Formulation CFS1-CFS10

Formulation	Drug(mg)	Ethyl Cellulose	Chitosan	Glutaraldehyde(5ml each)
CFS1	400	200	--	2.5%
CFS2	400	400	--	2.5%
CFS3	400	600	--	2.5%
CFS4	400	800	--	2.5%
CFS5	400	1000	--	2.5%
CFS6	400	--	200	2.5%
CFS7	400	--	400	2.5%
CFS8	400	--	600	2.5%
CFS9	400	--	800	2.5%
CFS10	400	--	1000	2.5%

 

Table No.2 Evaluation of Formulation CFS1-CFS10

Formu lation code	Particle Size (µm)	Angle of Repose(0)	Bulk Density (gm/cm)	Tapped Density (gm/cm)	%  Yield (%)	Drug Content (in25mg)	Encapsulation Efficiency (%w/w)	Cum.% Release
CFS1	43.21	21.97	0.862	0.968	84.70	21.93	86.94	97.18
CFS2	41.36	23.04	0.821	1.006	85.89	21.56	85.73	97.76
CFS3	38.72	22.06	0.823	1.03	85.36	22.49	81.52	98.18
CFS4	38.93	22.36	0.786	0.964	86.10	21.34	82.05	99.12
CFS5	37.77	22.11	0.792	0.976	88.36	22.65	89.88	98.80
CFS6	37.27	22.66	0.805	1.015	85.80	22.72	84.95	98.38
CFS7	36.14	22.67	0.846	0.988	85.67	21.96	84.88	98.64
CFS8	39.04	23.64	0.852	1.019	86.16	21.64	91.81	99.44
CFS9	38.43	23.67	0.807	0.973	85.50	21.47	84.10	98.28
CFS10	39.51	23.81	0.820	0.970	86.41	21.44	84.07	98.02

n=3±S.D

 

 

Figure No.3 Optical Microscopic Image

 

Figure no.4. Semphoto Graph (After 40⁰)

 

RESULTS AND DISCUSSION:

The Sustained release microspheres of all batches were found to be spherical and moderate free flowing. FTIR studies said that there was no incompatibility between Drug and polymers. The Drug encapsulation efficiency of microspheres was found to be 81.52-91.81%w/w. It has been stated that, the formulation shows the bulk density values less than 1.2 gm/cm² indicating moderate flow characteristics of microspheres. The Angle of repose for the formulated microspheres found to be within the range and showed as moderate flow characters. Stability Studies stated that there was not much different in the drug content and moderate difference in the SEM analysis. The in-vitro drug release for all formulations was found to follow Non -Fickian Diffusion release kinetics.

 

CONCLUSION:

From the study, it was concluded that the Imatinib Mesylate (α-form) loaded microspheres prepared with cross-linking agent Glutaraldehyde, with different polymers like Ethyl cellulose, Chitosan showed decreased in the release rate of drug from the formulation depending on the concentration of the polymers. When the concentration of the polymers influenced the drug release pattern. The formulation with drug and polymers in 1:3 ratio (CFS8) with cross linking agent maximum concentration was considered best because, it showed retard drug release in pH 7.4 buffer was found to be 99.44 % almost complete and uniform after the 24hour release study. Drug encapsulation efficiency for formulation (CFS8) was 91.81% w/w and all the remaining parameters were within the prescribed limit.

 

REFERENCE:

1.       Garg Tarun, Murthy RSR, Patented Microencapsulation Techniques And Its Application, Journal of Pharmacy Research, 2011,4(7),2097-2102

2.       Dhara B. Patel, Natural excipients in controlled drug delivery systems, Journal of Pharmacy Research 2009, 2(5),900-907.

3.       K.G.Parthiban, Novel drug delivery system: Formulation and characterization of exemestane microspheres by chemical cross linking method, Research Journal of Pharmaceutical, Biological and Chemical Sciences, October – December 2010 1(4) Page No.83- 90.

4.       S.S.Bansode,S.K.Banarjee,D.D.Gaikwad,S.L.Jadhav,R.M.Thorat,Microencapsulation : A Review, International Journal of Pharmaceutical Sciences Review and Research, Volume 1, Issue 2, March – April 2010; Article 008,page no 38-43.

5.       New Spectrophotometric Methods for the Determination of Imatinib in Bulk Drug and in Pharmaceutical Formulations,Research Journal Pharmacy and Technology-page no.578.

6.       Korsmeyer R. W., Gurny R. Peppas, “Mechanism of Solute Release From Porous Hydrophilic Polymers.”, Int. J. Pharm. 1983,Pg. 25-35.[15-17]

7.       Higuchi T., “Mechanism of Sustained Action Medication:Theoretical Analysis of Rate of Release of Solid Drug Dispersed in Solid Matrix.” J Pharm.Sci, 1963, Pg. 1145-1149. [15-17]

8.       Alfred Martin. Diffusion and Dissolution. In: “Physical Pharmaceutics”, 4th edition, Lippincott Williams and wilkins. Maryland. USA. 2001. Pg. 324 - 361. [15-17]

9.       Paulo Costa, Jose Manuel Sousa Lobo, “Modeling and comparison of dissolution profiles”,  Eur.J.Pharm., 2001; 13:123-133. [13-17]

10.     Yung-Kwan Chun, Hongkee Sah, Hoo-Kyun Choi, “Preparation of microspheres containing antimicrobial agents for eradication of H.Pylori.” Int.J.Pharm. 2005; 297:172-179. [13-14].

11.     Alferd Martin, Pillar Bustamante, A H E Chun, Micromeritics, in “Physical Pharmacy”, 4th Edition,2001; Lippincott Williams and Wilkins; pg. no. 427-429. [12].

12.     VS Subramanyam, Micromeritics, in “Physical Pharmaceutics”, 2nd Edition, 2000; Vallabh prakashan-Delhi; pg.no. 180-210.[11].

13.     Leon Lachman, Herbert A Lieberman, Joseph L Kanig,Micromeritics and coating of  capsule, in “Theory and

Practice of Industrial Pharmacy”, 2nd edition; Lea and Febiger, Philadelphia;pg.no:425-436.[10].

14.     Skoog, Holler,” Principles of Instrumental Analysis” fifth edition,pg. No: 380-426.[5]

15.     Applications of Absorption Spectroscopy of Organic compounds by John.R.Dyer, Indian edition pg no: 33-38.[6]

16.    Harshad Parmar, Sunil Bakliwal, Nayan Gujarathi, Different  Methods of Formulation and   Evaluation of  Mucoadhesive Microsphere, International Journal of Applied Biology and Pharmaceutical Technology. Page:1165.

 

 

Accepted on 19.06.2012      © RJPT All right reserved

Research J. Pharm. and Tech. 5(7): July 2012; Page 934-937