Development and Characterization of Mesalamine Microsphere for Colon Specific Drug Delivery

 

Komal S. Lavhate¹, Rahul S. Solunke¹, Kore K. J.², Rajkumar V. Shete³, Madhuri T. Deshmukh⁴

¹Department of Pharmaceutics, Rajgad Dnyanpeeth’s College of Pharmacy, Bhor, Pune 412206, MS India.

²Department of Microbiology, Rajgad Dnyanpeeth’s College of Pharmacy, Bhor, Pune 412206, MS India.  

³Department of Pharmacology, Rajgad Dnyanpeeth’s College of Pharmacy, Bhor, Pune 412206, MS India.

 

ABSTRACT:

Mesalamine Microsphere prepared by Ionic Gelation Technique by using sodium alginate and pectin as a release modifier and calcium chloride as a cross linking agent. In this formulation Eudragit-S-100 use a coating polymer that helps to releasing the drug at targeted site. The particle size of prepared Microsphere is less than 200µm that evaluated by the Scanning Electron Microscopy. The Mesalamine Microsphere has other characterization is in the term of Percentage Yield, Entrapment Efficiency, P- XRD, Differential Scanning Colorimetry, FT-IR and the In-vitro drug release. The In-vitro release pattern of Mesalamine studied in pH 7.4 phosphate buffers as 900ml media using USP dissolution apparatus II. The F3 batch of the formulation showed maximum release that is 99.75% at maximum time 9 Hrs. that is due to using pectin polymer that helps to retared the drug release at targeted site over the other formulation. F3 formulation showed better flow characterization by the term of Micromeritic Properties.

 

 

 

INTRODUCTION:

Microsphere are characteristically free flowing powder it is solid spherical smooth surface particle size is less than 200µm. Microsphere made up of protein and synthetic polymer which are biodegradable in nature other protective material that are biodegradable synthetic polymer and modified natural product that are (starch, gum, proteins, fats and waxes). Microsphere has been common technique in the production of controlled release dosage form [1].

 

Microsphere are multiparticulate drug delivery system which are prepared to obtain prolonged or controlled drug delivery to improve bioavailability, stability and to target the drug to specific site at a predetermined rate. To obtain maximum therapeutic efficacy, it’s most important to deliver the agent to the target tissue in the optimal amount in the right period of time there by causing less toxicity and minimal side effects. For the sustained release or controlled drug delivery system Microsphere using as a carrier for a drug its releases drug at a targeted site and shows action [2-4].

 

Microsphere is small in size so it occupies the larch surface area, it is expected to provide less inter and intra-individual variability more rapid and uniform gastric emptying. More uniform dispersion and reproducible transit through GIT tract. Site specific delivery of the drug to the target sites has the potential to reduce the side effects and improved pharmacological response and unwanted distribution of the drug to non-target organ can be avoided so the multiparticulate dosage form is more advanced than the single unit dosage form [5-7]. Mesalamine it is used in a treatment of ulcerative colitis. It contains 5-aminosalicylic acid. It is an inhibiter of cyclo-oxygenase pathway, it inhibiting the production of prostaglandin E2 in inflamed intestinal specimens [8-9].

 

MATERIALS AND METHODS:

Chemicals: Mesalamine, Eudragit-S-100 is gift sample from the Bills GVS Pharma. Sodium alginate, Pectin, calcium chloride from Chem Dyes Corporation Gujarat Rajkot.

 

Mesalamine Microsphere prepared by Ionic Gelation Technique:

All ingredients were weighed properly according to the formulas given in Table 1. For the preparation of the core microsphere weighed quantity of mesalamine dissolved in 0.1 N HCL [10]. At the same time, pectin dissolved in the water and kept aside for at least 1 hr. Then sodium alginate dissolved in 50ml of water to form a uniform paste solution. After 1 hr. when pectin solution was formed properly, pectin paste solution added to the container containing sodium alginate paste with continuous stirring to get uniform paste of required consistency. To this uniform paste, a clear solution of mesalamine in 0.1 N HCL was added. This mixture was ultrasonicated for 15 minutes for complete dissolution. Calcium chloride solution (5g in 100ml) was kept on a magnetic stirrer for continuous stirring. For the preparation of microspheres, polymer solution was added dropwise into calcium chloride solution through the syringe with a needle of 24 gauges. This complete solution allowed standing for 1 hr. for curing. Later separated, washed and dried in an oven at 500C for 24 hr. and store in plastic bags.

 

Preparation of Eudragit S 100 coated Mesalamine Microsphere: [11]

Mesalamine Microsphere coated with Eudragit S-100 by solvent evaporation method. Microspheres were dispersed in 10ml of coating solution prepared by dissolution of Eudragit S-100 in ethanol: acetone (2:1). This organic phase was then poured in 70ml of light liquid paraffin containing 1% w/v Span 80. The system was maintained under agitation with speed of 1000rpm on magnetic stirrer at room temperature for 3 hours to allow for the evaporation of solvent. Finally, the coated Microspheres were filtered by whatman filter paper, washed with n-hexane and dried in desiccators.

 

Table No 1: Batch Specification of Formulation

Formulation code	Drug: polymer Ratio	Sodium Alginate (%W/V)	Pectin (%W/V)	Calcium Chloride (5% W/V)
F1	1:1	3	2	5
F2	1:2	3	2	5
F3	1:3	3	2	5
F4	2:1	3	1	5
F5	2:2	3	1	5
F6	2:3	3	1	5
F7	3:1	3	0	5
F8	3:2	3	0	5
F9	3:3	3	0	5

 

EVALUATION PARAMETER:

UV Visible Analysis:

0.01mg of Mesalamine was accurately weighed and transferred in to 10ml volumetric flask. It was dissolved in little amount of 0.1 N Hcl and in 10ml of 7.4 pH buffer solution and made up to the volume with 0.1 N Hcl for acidic medium and for alkaline medium volume made up with phosphate buffer 7.4 to give a stock solution of 1mg/ml (1000μg/ml con). This was served as blank. From the standard stock solution 1ml of solution was taken and diluted to 10ml to get a solution of 100μg/ml and this served as a standard solution .In to series of 1ml volumetric flask aliquots of standard solution i.e., 0.5ml, 1ml, 1.5ml, 2ml, 2.5ml were added and the volumes were made up to 10ml using 0.1 Hcl for acidic medium and for alkaline medium volume made up with phosphate buffer 7.4 respectively. The absorbance of these solutions was measured against the reagent blank at 303 nm using UV spectrophotometer. A standard curve was plotted with concentration on x-axis and absorbance on y- axis [12].

 

FTIR Study:

10mg of the sample and 400mg of KBr were taken in a mortar and triturated. A small amount of triturated sample was taken into a pellet maker and was compressed at 10kg/cm2 using a hydraulic press. The pellets was kept onto the sample holder and scanned from 4000 cm-1 to 400 cm-1 in SHIMADZU FT-IR Spectrophotometer.

 

Differential Scanning Colorimetry (DSC):

The DSC analysis of pure drug, polymer and the physical mixture of drug and polymer were carried out using differential scanning calorimeter (METTLER TOLEDO). Sample of about 5 mg was placed in a 50μl perforated aluminum pan and sealed. Heat runs for each sample were set from 5⁰C to 300⁰C using nitrogen as purging gas and samples were analyzed. [13].

 

Scanning Electron Microscopy (SEM):

The SEM analysis was carried out using a scanning electron microscope (SPPU-JEOL.). Prior to examination, samples were mounted on an aluminum stub using a double sided adhesive tape and making it electrically conductive by coating with a thin layer of gold (approximately 20nm) in vacuum. The scanning electron microscope was operated at an acceleration voltage of 5 kV and resolution of 4000.

 

P-XRD Analysis:

Powder XRD of Mesalazine, Eudragit-S-100, Formulation mixture of Mesalazine with Eudragit S-100, was recorded using Bruker AXS D8 Advance diffractometer with Si(Li) PSD detector. The operation data were: measuring circle diameter-435, 500 and 600 mm predetermined; angle range-360⁰; X-ray source-Cu, wavelength 1.5406A⁰.

 

Drug Entrapment Efficiency:

Microspheres were crushed using a glass mortar by pestle and equivalent to 5mg of Mesalamine weighed. These Microspheres were suspended in 25ml of phosphate buffer pH 7.4. After 24 h, the solution was filtered; 1ml of the filtrate was pipette out and diluted to 10ml and analyzed for the drug content using UV visible spectrophotometer at 303nm. The Drug Entrapment Efficiency was calculated using the following formula: [14]

 

Micromeritic Properties:

Accurately weighed Microspheres were poured gently through a glass funnel into a graduated cylinder exactly to 10mL mark. Initial volume was noted. Bulk density and tapped density were noted using tapping method using 10mL measuring cylinder. Angle of repose (ɵ), Hausner’s ratio (H) and Carr’s index (%C) were calculated to study the flow properties of Microspheres.

 

In-Vitro Drug Release:

Drug Release tests were performed by using LABINDIA dissolution tester (DS 8000) USP I apparatus (basket method) for each size fraction separately. Accurately weighed amounts (500mg) of Microspheres were introduced into 900 mL of PBS (phosphate buffer saline, pH 7.4) and stirred with 100 rpm at (37.0±0.5)⁰C. Five milliliters samples were withdrawn and filtered at selected time intervals. The concentration of Mesalamine was determined spectrophotometrically at 303 nm [16].

 

RESULT AND DISSCUTION:

UV Visible Analysis:

The absorbance of the sample solution of Mesalamine showed maximum absorption at wavelength 303nm in acidic medium (0.1N Hcl) regression coefficient is 0.9976 and in phosphate buffer pH 7.4 regression coefficient is 0.9949

 

FT-IR Analysis:

In this present investigation the FT-IR peak of Mesalamine evaluated by showing carboxylic acid stretch R-C=O-OH peak at 3425.69 cm-1 and 1357.93 cm-1. C=O stretch at 1728.48 cm-1. Aromatic ring stretch at 1620 cm-1. Drug and polymer mixture studies indicated that there were no kind of significant interactions between the polymer used with the drug in the formulation, as the principle peaks of the drug in the spectra obtained for the formulation was not altered. The peaks corresponding to the N—H stretching in drug showed a peak at 3394.83 cm-1. Aryl C-H shows the peak at 2908.75 cm-Benzene ring C6H5,–C-H shows 1728.28 and 1496.81 cm-1.

 

Differential Scanning Colorimetry (DSC):

The DSC studies was carried out to observe the thermal behavior of drug-loaded Microspheres whether the drug was encapsulated in them or not. The characteristic endothermic peak of Mesalamine pure drug is appeared at 288.38°C which was within a range of melting point. Similar as for Eudragit-S-100 peak appeared at 213°C. For sodium alginate the peak was at 252°C. Thermo-gram of the drug and polymer that also showed the thermal stability of the drug. [17]

 

Scanning Electron Microscopy (SEM):

The F3 formulation of the Mesalamine Microsphere revealed that the surface morphology was found to be spherical and rough due to higher concentration of drug and polymer. Significant particle size that is 100µm which is less the 200µm. hence the particle size of the F3 formulation within the range. [18]

 

XRD Analysis:

XRD patterns of coated mesalamine Microsphere with Eudragit S-100 showed a sharp intense peak at 15.167⁰ represented in the Fig no. 4 and indicating that the drug and the polymer existed in the crystalline state. And after the preparation of coated Microsphere the Mesalamine intense peak remains as it is but due to the sodium alginate, pectin and Eudragit-S-100 the additional peak was forms.

 

Drug Entrapment Efficiency:

The encapsulation efficiency is significantly increased with increase in core: coat ratio. When polymer concentration is increased increases in the drug entrapment of F3 batch 69.25±0.6% was observed in table no 2.

 

Angle of repose showed the flow property of the Microsphere the F3 formulation showed the excellent flow of the Microsphere also the Hausner’s ratio. Carr’s index, within the excellent range so that the prepared formulation is in a range.

 

IN- VITRO DRUG RELEASE STUDIES:

A coating of Eudragit-S-100 the drug showed slow release or no release in acidic media. Also the pectin used as release modifier that helps to retard the release of the drug so the 1:3 formulation is shown the best release upto 9 hrs. In other formulation the premature drug release is happed so the pectin helps to protect the premature drug release in the buffer.

 

Micromeritic Properties:

Table No 2:  Flow Characteristics of Prepared Microspheres:

Formulation code	Angle of Repose	Hausner’s Ratio	Carr’s Index (%)	% Yield	% Entrapment Efficiency
F1	29.52±0.12	1.10±0.36	7.2±0.43	77.46±0.96	57.24±0.12
F2	33.45±0.65	1.11±0.52	10.2±0.46	80.46±0.65	65.25±0.23
F3	27.52±0.63	1.9±0.28	9.4±0.39	84.23±0.84	69.25±0.6
F4	34.52±0.59	1.16±0.31	11.2±0.41	69.42±0.91	52.42±0.52
F5	38.42±0.68	1.25±0.27	14.5±0.55	75.82±0.37	58.43±0.13
F6	32.51±0.91	1.12±0.26	24.52±0.41	79.25±0.37	64.57±0.51
F7	40.52±0.56	1.15±0.29	16.45±0.42	64.76±0.65	46.35±0.36
F8	42.15±0.52	1.28±0.28	18.75±0.56	68.47±0.91	48.25±0.23
F9	41.25±0.65	1.14±0.22	21.89±0.39	75.74±0.63	52.56±0.14

 

Table No 3: % Drug Release:

 

The regression coefficients of determination (R²) were listed in from Table no 4. The regression coefficient of determination indicated that the release data was best fitted with zero order kinetics. The entire kinetics model proved that the release of the Mesalamine microsphere is followed the zero order kinetic models so the prepared formulation showed the sustained release flow of the drug it is considered by the analysis of the model.

 

Table No 4: Regression Coefficient (R²) value of Different Kinetic Model

Formulation Code	Zero Order	First Order	Higuchi Profile	Korsmeyer-peppas
	R2	R2	R2	R2
F1	0.9981	0.8852	0.9753	0.9704
F2	0.9987	0.8402	0.9725	0.9705
F3	0.9975	0.7203	0.9694	0.9759
F4	0.9949	0.8104	0.9668	0.9612
F5	0.997	0.8486	0.9923	0.9933
F6	0.9892	0.9071	0.9968	0.9975
F7	0.9993	0.8412	0.985	0.9819
F8	0.9942	0.9051	0.9929	0.9873
F9	0.9951	0.8891	0.9962	0.9917

 

CONCLUSION:

Mesalamine microsphere prepared by Ionic Gelation method and coating with eudragit-s-100 by solvent evaporation method. The pectin is played an important role it is used as a release modifier that helps retard the Drug Release rate it has been observed that the drug release rate decreases with time due to increasing concentration of polymer. The reason attributed to this fact is a slow erosion of the coating layer and pectin. Pectin microspheres are effective for controlling the release of a drug within the GIT that increases the therapeutic efficiency. In this formulation observed that the formulation F3 containing polymers mixture sodium alginate and pectin in ratio was able to protect the formulation from premature Drug Release up to 9 hr. when compared to those formulations containing concentration of pectin. The finalize batch characterize by the Drug Release test was found to be 99.75%. All the formulations fitted in the kinetics model according to this the resulted formulation follow zero-order kinetic model. The F3 batch has a smaller particle size characterized by SEM that occupies the large surface area resulting in increased absorption. The F3 batch shows excellent flow property so the reticuloendothelial barrier decreases and site-specificity increased. Therefore the developed of mesalamine formulation proves that used in the treatment of colon-specific drug delivery and facilitating in the management of UC.

 

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Received on 16.07.2019         Modified on 16.11.2019

Accepted on 10.10.2019         © RJPT All right reserved