Development and Characterization of Repaglinide Loaded Floating Microparticles

Ajeet Singh^1,2, Ranjit Singh¹*

¹AVIPS, Shobhit University, Gangoh, Saharanpur, U.P., India – 247341.

²Department of Pharmaceutical Sciences, J.S. University, Shikohabad, Firozabad, U.P. India – 283135.

*Corresponding Author E-mail: ranjitsps@gmail.com

ABSTRACT:

Sustained release drug delivery has been successfully achieved using microparticles made from natural or synthetic polymers. The aim of this study is to develop and test floating microparticles of Repaglinide in order to improve drug bioavailability by extending gastric residence time. Repaglinide, an oral hypoglycemic, is a lipophilic drug that is rapidly absorbed from the stomach and eliminated with a half-life of just 1 h, so suitable to be formulated as floating drug delivery system for sustained release.Repaglinide floating microparticles were developed using an ionotropic gelation method that included calcium chloride as a cross-linking agent, sodium alginate, and different concentrations of hydroxypropyl methylcellulose (HPMC), ethyl cellulose (EC), and sodium bicarbonate (NaHCO₃). A three factor, three-level Box-Behnken design was used to study the effect of independent variables on dependent variables. In the formulation of microparticles, the amount of hydroxypropyl methylcellulose (X₁), ethyl cellulose (X₂), and sodium bicarbonate (X₃) were three independent variables, while percentage buoyancy (Y₁) and percentage drug release at 10 h (Y₂) were dependent variables. Micromeritic properties, percent yield, percent drug entrapment performance, surface morphology, percent buoyancy, in-vitro drug release, and drug excipient compatibility were all assessed in the formulations.FTIR studies revealed no interaction between the drug and the excipients.SEM for surface morphology studies revealed that their surface is spherical and smooth. The mean particle size of formulations was found to be between 415- 689 µm, the drug entrapment efficiency was found to be between 44.65% - 76.55% and percent buoyancy was noted to be between 63%- 78.33%. The results revealed that entrapment efficiency increased as polymer concentration was increased. The cumulative percent drug release after 10 h was noted to be between 76.87- 88.12%. Percent drug release decreased as polymers concentration was increased. The buoyancy was increased with increasing concentration of sodium bicarbonate. The developed microparticles could successfully retard the release of the drug.

KEYWORDS: Floating Microspheres, Repaglinide, Ionotropic Gelation Method, Drug delivery, Diabetes.

1. INTRODUCTION:

In recent years, much more attention has been drawn towards developing novel controlled release oral drug delivery systems to overcome the reported deficiencies of conventional dosage forms and to achieve the desired patient compliance and optimum clinical efficacy¹. Among the different routes of drug administration, the easiest, flexible and convenient is the oral route for the patient^2-5.

Drugs with narrow absorption window are desired to retain at the site of absorption for a longer period of time in order to obtain controlled release of drug^6,7. Several approaches have been devised to retain the drug delivery system at the gastrointestinal tract such as sedimentation, floatation, expansion, mucoadhesion and modified shape system^8-11. Drugs having short half-life are eliminated quickly from blood circulation and require frequent dosing; therefore, controlled release formulations of such drugs have been developed to increase the bioavailability¹². Floating drug delivery is useful for several categories of drugs, which act locally in the stomach, poorly soluble at an alkaline pH, having narrow window of absorption, unstable in the intestine or colonic environment and primarily absorbed in the stomach^13-17. Floating devices administered in a single unit form as hydro-dynamically balanced systems are unreliable in prolonging the gastric retention time owing to their all or none effect and thus may cause high variability in local irritation and bioavailability due to large amount of drug delivery at a particular site of GIT¹⁸. The aim of this study was to develop a multi-particulate floating Repaglinide drug delivery system using Ethylcellulose (EC) and Hydroxypropyl methylcellulose (HPMC) as release control polymers. Repaglinide is a non-sulfonylurea oral hypoglycemic agent of meglitinide class and has rapid onset, low bioavailability (50%) and short half-life (1h)¹⁹. Therefore, it was chosen as model drug for the development of floating microparticles to increase gastro-retention time.

2. MATERIALS AND METHODS:

2.1 Materials:

Repaglinide was obtained as gift sample from Sun Pharmaceutical Industries Ltd, S.A. S. Nagar, Punjab, India. Ethylcellulose (EC) and Hydroxypropyl methylcellulose (HPMC) (100 cps) were purchased from Himedia Chemicals, India. Sodium alginate was purchased from Loba Chemie Laboratory, Mumbai, Calcium chloride (CaCl₂) and Sodium bicarbonate (NaHCO₃) were obtained from Rankem Laboratories, New Delhi. The rest of the chemicals were of analytical grade.

2.2 Compatibility study:

The pure drug and the mixture of drug with polymers were analyzed by FTIR spectroscopic method to study their compatibility. IR spectra were taken by IR spectrophotometer (Shimadzu 8400, Japan).

2.3Preparation of Repaglinide microparticles:

The drug's alginate microparticles were developed using the ionotropic gelation technique²⁰ with various polymers quantities, as shown in Table 3. Sodium alginate solution (2% w/v) was mixed to weighed amount of ethyl cellulose (1%- 2% w/v) dissolved in desired amount of ethanol. Weighed quantity of drug, Repaglinide (20mg) and Hydroxypropyl methylcellulose (1%- 2% w/v) were triturated to form fine powder followed by addition to the above solution. Sodium bicarbonate (50- 150mg), was added to this mixture and the resulting solution was mixed uniformly. The above solution was dropped into 100 ml of gently agitated calcium chloride (4% w/v) solution using a 26 G syringe needle to obtain microparticles. This solution having microparticles was stirred slowly for 10 minutes using magnetic bead. The prepared microparticles were further permitted to remain in the same solution for 20 minutes to enhance the mechanical strength. The devised microparticles were filtered, washed with distilled water and then air-dried at room temperature and finally stored in desiccators.

2.4 Experimental design for optimization:

To reduce the number of trials necessary to obtain maximum information on product properties, the screening was performed applying a three –factor, three –level Box- Behnken design. This design helps in optimizing a process using a small number of experimental runs. Hydroxypropyl methylcellulose concentration (X₁), Ethylcellulose concentration (X₂) and amount of Sodium bicarbonate (X₃) were three independent variables considered in the preparation of microparticles, while percent buoyancy (Y₁) and the percent drug release at 10 h (Y₂) were dependent variables. The statistical experimental design was produced and assessed using Design- Expert version 12 software (Stat-Ease Inc., USA). Table 3 shows the design matrix, which includes the examined responses, such as percent buoyancy and percent drug release at 10 h.

2.5 Characterization of the microparticles:

2.5.1 Micromeritic properties:

Microparticles were characterized for physical properties viz. particle size, loose bulk density, tapped bulk density, angle of repose, Carr’s index and Hausner’s ratio. Particle size was determined by sieving method and the mean particle size of microparticles was calculated. The flow characteristics were measured by angle of repose by fixed funnel method. Tapped density was determined by tapping method using bulk density apparatus and Carr’s index and Hausner’s ratio were estimated subsequently by using the following equations²¹:

Carr’s index (%) = {(TBD – LBD) X 100}/TBD

Hausner’s ratio = Tapped Bulk Density/Loose Bulk Density

2.5.2 Scanning Electron Microscopy (SEM):

A scanning electron microscope was used to investigate the shape and surface morphology of drug-loaded microparticles.

2.5.5 Floating behavior:

The microparticles (50mg) were agitated at 100rpm in simulated gastric fluid (pH 1.2, 100ml) containing Tween 20 (0.02% w/v). After 12 hours, the buoyant microparticles were separated from the rest by filtration. Both types of formulated microparticles were dried, and weighed. The following equation was used to calculate the buoyancy:

Wf/(Wf+Ws)x100 = buoyancy in percent

The weights of the floating and settled microparticles, respectively, are Wf and Ws²².

2.5.6 Entrapment efficiency and yield:

The microparticles were weighed (50mg), crushed and suspended in 10ml of ethanol and were kept for 12 h to dissolve the polymeric shell for extraction of drug.The drug content in the filtrate was spectrophotometrically scanned at 243nm after filtration and suitable dilution. The following formulas were used to calculate the percentage of drug entrapment and yield ²².

(Calculated drug content/Theoretical drug content) x100= Percent Drug entrapment

(Total weight of floating Microparticle/Total weight of drug and polymer) x100= Percent Yield

2.5.7 In-vitro release studies:

A paddle type six-station dissolution test apparatus was used to conduct in-vitro drug release tests (Veego, USP Standard). A weighed quantity of the microparticles, corresponding to 16 mg of the drug, was placed in 0.1N HCl (1.2 pH) containing Tween 20 (0.02% w/v) and kept at 37± 0.50°C and 100 rpm. The sink condition was maintained perfectly. At each 1h interval, a 5 ml sample was taken, filtered, and analysed at 243 nm. All of the experiments were carried out in triplicate.

2.5.8 In vitro release kinetics:

The kinetics of drug release were investigated by applying different kinetic models to in vitro release results, including First order (log cumulative percent drug remaining vs. time), Zero order (cumulative percent drug released vs. time), Hixson Crowell model (cube root of percent drug remaining vs. time), Higuchi model (cumulative percent drug released vs. square root of time), and Korsmeyer Peppas model (log cumulative percent drug released vs. log time).For the linear curves obtained through regression analysis ,regression coefficient (R²) values were computed^{23, 24}.

3. RESULTS AND DISCUSSION:

3.1 Compatibility study:

The compatibility of drug and polymers was studied by FTIR (figures 1-2). The findings revealed that there was no interaction between the drug and the excipients, as the same peaks were seen in the spectra of the pure drug and the mixture (table 1, 2).

Figure 1: FT-IR spectra of Repaglinide

Figure 2: FTIR of Drug and excipients

Table 2: Interpretation of the IR spectraof Repaglinide

Functional group	Observed peaks at Wave Number (cm^-1)
NH stretching	3391
OH stretch	2917
CH stretching	2856
C=O stretch	1634

3.2 Preparation of Repaglinide Microparticles:

The alginate microparticles containing Repaglinide were prepared by ionotropic gelation technique using calcium chloride aqueous solution as cross linking agent. When dispersion mixture of sodium alginate, EC, HPMC and Repaglinide was dropped into the solutions containing the calcium ions, ionotropically gelled alginate beads containing Repaglinide were formed instantaneously. In presence of divalent calcium ions in the cross linking solution, the calcium was ionically substituted at the carboxylic site present the alginate strands and attaches two alginate strands together. This led to the formation of solid gel²⁵.

3.3 Optimization of formulated Repaglinide microparticles:

The main challenge in pharmaceutical product development is to find the appropriate combination of variables that will yield the product with optimal quality. The conventional method of pharmaceutical optimization based on changing one variable at a time while keeping others fixed is laborious and time consuming²⁶. This method also requires a complete series of experiments for every variables of interest. Moreover such a method does not provide means of observing possible interactions between variables. Compared with the contemporary optimization method utilized, Box- Behnken Design bears the main advantages of figuring out the potential interactions between parameters and also time saving by reducing the number of experiments²⁷. The response surface methodology usually indicates the mutual interaction behavior of factors through three- dimensional response surface graphs and two- dimensional contour graphs relating measured responses.

Table 3: Observed values of responses for Box-Behnken design

Formulation Code	HPMC % (X₁)	EC % (X₂)	NaHCO₃mg (X₃)	Buoyancy % (Y₁)	Drug Releaseat 10h % (Y₂)
F₁	1.5	1.0	50	67.00	81.12
F₂	2.0	1.5	50	63.00	76.87
F₃	1.0	1.0	100	68.66	88.12
F₄	1.0	2.0	100	66.66	84.37
F₅	1.5	2.0	50	67.33	80.62
F₆	2.0	2.0	100	66.12	76.87
F₇	1.0	1.5	150	75.66	88.12
F₈	1.5	2.0	150	78.33	81.67
F₉	2.0	1.5	150	76.64	78.75
F₁₀	2.0	1.0	100	70.00	77.24
F₁₁	1.5	1.5	100	65.33	82.49
F₁₂	1.0	1.5	50	66.66	86.47
F₁₃	1.5	1.0	150	75.73	80.24

3.4 Evaluation of the microparticles

3.4.1Micromeritic properties

Micromeritic properties of the microparticles were assessed and the observations are depicted in table 4.

Table 4: Micromeritic properties of the microparticles

Formulation code	Particle size (µm)	Bulk density (g/cm³)	Tapped density (g/cm³)	Angle of repose (ø)	Carr’s Index (%)	Hausner’s Ratio
F₁	618±0.59	0.542±0.016	0.601±0.011	26.54±1.558	9.83±1.135	1.11±0.014
F₂	425±0.90	0.536±0.020	0.611±0.023	26.16±1.858	11.35±0.563	1.14±0.017
F₃	415±0.74	0.545±0.018	0.604±0.027	27.18±1.808	9.69±1.186	1.10±0.014
F₄	548±1.41	0.558±0.009	0.607±0.018	25.68±1.462	8.05±1.380	1.04±0.093
F₅	489±0.81	0.561±0.010	0.609±0.013	22.33±1.452	8.43±0.359	1.09±0.045
F₆	689±2.10	0.551±.018	0.623±0.010	28.30±2.133	11.02±1.847	1.12±0.019
F₇	453±2.31	0.559±0.022	0.607±0.015	27.60±1.593	7.94±1.503	1.09±0.014
F₈	645±1.30	0.531±0.016	0.585±0.027	27.59±2.364	9.20±1.419	1.10±0.017
F₉	646±1.55	0.534±0.010	0.609±0.020	24.56±1.678	12.25±1.175	1.14±0.014
F₁₀	564±0.37	0.556±0.011	0.634±0.013	21.53±2.824	12.27±0.747	1.14±.009
F₁₁	578±1.21	0.594±0.022	0.617±0.019	27.44±2.204	11.86±0.907	1.13±0.011
F₁₂	526±0.91	0.551±0.009	0.618±0.019	23.63±1.401	10.82±1.244	1.12±0.015
F₁₃	612±2.16	0.550±0.009	0.611±0.016	28.16±1.928	10.24±1.414	1.11±0.017

Table 5: Percent yield, % buoyancy, drug content and drug entrapment efficiency of themicroparticles

Formulation Code	Theoretical drug content(mg)	Actual drug content(mg)	%Drug entrapment efficiency	% Buoyancy	% Yield
F₁	20	10.08±0.677	50.40±3.387	67.00±0.91	76.05
F₂	20	13.23±1.003	66.15±5.014	63.00±0.86	65.34
F₃	20	8.93±0.255	44.65±1.429	68.66±0.90	66.09
F₄	20	11.77±0.345	58.85±1.729	66.66±3.55	68.41
F₅	20	12.68±0.513	63.40±2.56	67.33±1.33	71.47
F₆	20	15.31±0.343	76.55±1.710	66.12±0.81	69.40
F₇	20	9.21±0.452	46.05±2.260	75.66±0.94	70.59
F₈	20	14.99±0.791	74.95±5.226	78.33±1.41	73.42
F₉	20	13.36±0.219	66.80±1.096	76.64±0.39	75.17
F₁₀	20	12.91±0.075	64.55±0.288	70.00±1.57	68.83
F₁₁	20	11.46±0.425	57.3±2.126	65.33±0.28	68.18
F₁₂	20	9.64±0.452	48.20±2.264	66.66±0.81	71.83
F₁₃	20	11.60±0.520	58.00±2.627	75.73±2.04	70.39

Table 6: Release kinetics of Repaglinide floating microparticles

FormulationsCode	Zero order(R²)	First order(R²)	Higuchi(R²)	Hixson – Crowell(R²)	KorsmeyerPeppas (R²)
F₁	0.974	0.987	0.981	0.929	0.977
F₂	0.980	0.979	0.982	0.972	0.990
F₃	0.989	0.901	0.959	0.950	0.978
F₄	0.979	0.970	0.978	0.954	0.985
F₅	0.983	0.986	0.996	0.931	0.996
F₆	0.980	0.991	0.992	0.955	0.995
F₇	0.983	0.942	0.992	0.874	0.993
F₈	0.993	0.961	0.965	0.992	0.952
F₉	0.969	0.976	0.972	0.970	0.978
F₁₀	0.956	0.961	0.973	0.971	0.981
F₁₁	0.993	0.928	0.966	0.975	0.984
F₁₂	0.992	0.903	0.951	0.995	0.954
F₁₃	0.998	0.957	0.970	0.992	0.975

4. CONCLUSION:

In the present study, the Repaglinide floating microparticles were successfully formulated by ionotropic gelation method. The microparticles were optimized using the Box-Behnken Design. The prepared microparticles were optimized on account of % buoyancy and % drug release. The optimized formulation (F₈) exhibited 81.67% drug release in 10 h. It was noticed that % drug release was decreased as polymer concentration increased and drug entrapment efficiency was increased with increasing polymer concentration. The buoyancy increased with increasing concentration of sodium bicarbonate. FTIR studies recommended that there was no interaction between drug and polymer. SEM confirmed spherical shape of formulated microparticles. The in-vitro drug release after 10 h ranged between 76.87- 88.12%. The in-vitro drug release indicated sustained release of the drug and the release pattern stick to the Zero order model. As a result, the developed novel Repaglinide dosage form is able to prolong the release and improve bioavailability, making it ideal for sustained release.

5. ACKNOWLEDGEMENT:

Authors are grateful to Sun Pharmaceutical Industries Ltd, S.A.S. Nagar, Punjab, India for providing Repaglinide as a gift sample. Authors are also thankful to Central drug research institute Lucknow, india for getting facilities to perform SEM studies.

6. CONFLICT OF INTEREST:

Authors have none to declare.

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Received on 12.05.2021 Modified on 27.05.2021

Research J. Pharm. and Tech 2021; 14(12):6573-6578.

DOI: 10.52711/0974-360X.2021.01137