INTRODUCTION:

Pharmaceutical carrier systems have been developed as the controlling delivery of drugs^1-3. Controlled drug delivery system improve efficiency, reduce toxicity followed by sustained release drug delivery⁴. The main purpose of preparation of aceclofenac microsphere⁵ is development of sustained release drug delivery^6-9. Microsphere is solid, nearly spherical, which has particle size ranging from 1-1000µm containing dispersed drug molecule in solution¹⁰. Microsphere enhance bioavaiability, reduce gastric irritation¹¹, enhance physical stability and gastric enzyme stability¹².

Microspheres have various applications in ophthalmic drug delivery, gene delivery, nasal drug delivery, buccal drug delivery, gastrointestinal drug delivery etc.^13-16. There are various types of microsphere such as bio-adhesive microsphere^17,18, magnetic microsphere^19-22, floating microspheres^23-26, radioactive microspheres^19,20, polymeric microspheres^21-24. There are various methods of preparation of microspheres such as solvent evaporation method²⁵, spray drying method^27-29, phase separation coacervation method, solvent extraction method³⁰ etc. Here this aceclofenac microspheres are prepared by Ionotropic gelation technique. Ionotropic gelation technique³¹ is based on the principle that polymer crosslinked³² with oppositely charged ion. This method is important because the degree of crosslinking and polymer concentration³² can be modified which makes it very useful in control drug delivery. Aceclofenac is an oxyacetic and non-steroidal anti-inflammatory drug which has a half life of 4 h³². Aceclofenac is used to reduce fever, inflammation of rheumatoid arthritis, traumatic pain etc. Sodium alginate is linear polysaccharide derivative of alginic acid³⁴. Sodium alginate is used as thickening agent, gelling agent texture improver in microsphere preparation³⁵. Guar gum is a galactomann polysaccharide which is extracted from guar bean that has thickening and stabilizing properties. Aceclofenac has higher anti-inflammatory action, highly protein binding and it acts by blocking the action of cyclo-oxygenase that is produced by prostaglandin.

MATERIALS AND METHODS:

Material:

Aceclofenac was purchased from Mankind Pharma, New Delhi. All formulation excipient like Sodium alginate (Loba Chemie Pvt. Ltd.), Guar gum (Loba Chemie Pvt. Ltd) and other excipient Calcium chloride, Sodium hydroxide, Potassium di- hydrogen Phosphate, Di- sodium hydrogen Phosphate) was obtained from Haldia Institute of Pharmacy, Haldia 721657, West Bengal.

Method:

The authentication and preformulation parameter of pure Aceclofenac drug maintaining their Quality, Purity and Standard.

Authentication parameter:

Melting Point Method:

Determination of melting point is one of the preformulation parameter where temperature is used for changes state from solid to liquid at atmospheric pressure. In this method solid and liquid can exist at equilibrium. The melting point of pure drug Aceclofenac was determined using two methods one is Conventional method and another is Digital method.

Partition Coefficient:

Partition Coefficient can be determined to check the solubility and permeability of drug. Then 1gm drug was added in separating funnel containing equal portion of 25ml benzene and 25ml water. The separating funnel was shaken for 25 – 30 min and stabilized the mixture for 5min. After stabilizing the mixture the aqueous phase was collected from separating funnel and filtrate. Then the filtrate was taken measure absorbance and calculate the log P.

Solubility Studies:

Solubility of a substance in a particular solvent is determined by concentration of a substance in a saturated solution in a particular temperature. At first different types of buffer solution PH 1.2 Acidic Buffer, PH 6.8 Phosphate Buffer, PH 7.4 Phosphate Buffer were prepared. Then dissolve drug material in above solution and water to form supersaturated solution .Next mixture can placed in Orbital Shaker for 24 hrs. After 24 hours the filtrate was taken and give absorbance. Then detect different concentration of drug soluble in different type of solution.

Calibration Curve of Aceclofenac

Weight 100mg of Aceclofenac (Mankind Pharma) was dissolve in phosphate buffer pH 6.8 to obtain 1000 ppm solution. From the above stock solution having concentration 100µg/ml various dilution were made to get concentration 2µg/ml, 4µg/ml, 6µg/ml, 8µg/ml, 10µg/ml by using phosphate buffer of pH-6.8. The absorbance of these solutions was measured using maximum wavelength (λmax), which was found to be 275nm by using UV-visible spectrophotometer (Analytical Technologies Limited). This absorbance was plotted on Y-axis against concentration on x-axis table 4.

Preformulation studies

Drug- Excipient Compatibility Studies

Drug is an active part of dosage form and it is mainly responsible for therapeutic use and excipient which is included with drug for formulation is used for initiate, participate in physical and chemical interaction for drug stability. It is used for investigate new drug product and new drug delivery system also.

Procedure

The equal portion of Drug and Excipient (1:1) is taken in Ampules which is placed in Stability chamber for one week. After one week drug excipient study determined using TLC (Thin Layer Chromatography).

Method of Formulation

Aceclofenac (Mankind Pharma) microsphere is prepared by Ionotropic gelation technique. At first Aceclofenac (Mankind Pharma) (125mg fixed), a specific amount of Guar gum (Loba Chemie Pvt. Ltd.) and sodium alginate (Loba Chemie Pvt. Ltd.) (total polymer = 250mg ) was placed in a beaker. Then 10ml distilled water was added to the beaker with constant stirring. The mixture of drug and polymer was poured drop by drop in different concentration of CaCl2 as indicated in table1. After making all beads of formulation, particles were collected by filtration, washed, dried, and desiccated at room temperature.

Evaluation Parameter

a) Particle size analysis

Particle size was measured with a compound optical microscope (Olympus model HB, India), attached to a camera. The image obtained by the camera measured size with an in built scale in the software. Randomly 50 microspheres were taken from the each formulation and placed on glass slide and number of divisions of the calibrated eye piece was counted and corresponding particle diameter was recorded.

b) Micrometric properties:

1. Angle of repose:

The angle of repose of different formulation were measured according to: tan θ = h/r. Where θ = angle of repose, h= height of sample from the base, r = radius of the sample in the base.

2. Bulk density and tapped density:

Bulk and tapped density were measured by using graduated cylinder. At first known amount of formulation was poured into cylinder and measure the bulk volume. Then the cylinder tapped mechanically for 10 min and tapped volume was noted. Then bulk density and tapped density were calculated.

3. Carr’s index (%):

Car’s index of microspheres were calculated by following equation:

Tapped density – Bulk density

Car’s index = –––––––––––––––––––––––––– × 100

Tapped density

4. Hauser’s ratio:

Hausner’s ratio was determined by comparing tapped density to bulk density.

Tapped density

Hausner’s ratio = –––––––––––––

bulk density

c) Estimation of drug encapsulation efficiency (c37):

Known amount (10mg) of formulations (F_1, F₂, F₃, F₄, F_5, F₆) were taken individually and kept in a 100ml phosphate buffer of pH 6.8 for 24 hours. Then the solutions were sonicated for 10 min to extract the residual amount of drug from the microspheres and the solutions were filtered to remove the suspended polymeric debris and the filtrate was taken for the determination of Aceclofenac content spectrophotometrically by using UV spectrophotometer (Analytical Technologies Limited) at a wavelength of 275 nm against appropriate blank. In order to maintain accuracy, experiments were carried out in triplicate for all the formulations to check its reproducibility. DEE was calculated from the following equation.

Actual drug loading

% DEF = –––––––––––––––––––––– × 100

Theoritical drug loading

s) Swelling Study (c35):

pH dependent dynamic swelling studies of microspheres were performed using a light microscope Microspheres were allowed to swell individually in pH 6.8 and at different time intervals, microspheres were withdrawn. After removal of the adhered liquid with the help of a blotting paper, changes in diameter were monitored with the help of an ocular microscope. All the samples were analyzed in triplicate and results were shown in table 3 and fig;

Swelling Index = {(M_t-M_o) / M_o} × 100

Where

M_t= weight of bead at time ‘t’

M_o= weight of beads at time ‘t’ = 0

e) In vitro drug release study: (c34, c36)

In vitro drug release study of Aceclofenac loaded microspheres were carried out in simulated gastric and intestinal fluid to exhibit the phenomenal effect of matrix cross linking on release profile of drug from microspheres. In vitro drug release study was performed in pH 6.8 using a USP type II rotating paddle dissolution apparatus (Analytical Technologies Limited). Accurately weighed, 10mg of dried microspheres were placed in 900ml dissolution media and temperature was maintained at 37±0.5°C. at predetermined time interval, 10ml of aliquot sample was withdrawn from the dissolution media and same volume of fresh buffer was replenished immediately. The samples were analysed spectrophotometrically (Analytical Technologies Limited) at 275nm. Cumulative percent of drug release in pH 6.8 was plotted at a function of time. Each formulation was tested in triplicate.

RESULTS AND DISCUSSION:

Authentication parameter:

Melting Point Method:

The Melting Point of Aceclofenac is determined by Conventional and Digital Method and Melting Point of Aceclofenac is reported in table 2.

Partition Coefficient:

Log P value is determined by Partition Coefficient phenomenon and reported in table 2.

Solubility Studies:

The Solubility of Aceclofenac in Given Solution. (Water, PH 1.2 Acidic Buffer, PH 6.8 Phosphate Buffer, PH 7.4 Phosphate Buffer) is Reported in Table and Solubility of Pure drug Aceclofenac in Different solvent is shown in table 3.

Particle size:

Particle size of the prepared microspheres was measured by optical microscope and results were represented in table 5.Spherical nature of the particles were observed with size ranging from 330±10 to 210±11µm. particle size of F₁ was higher than that of F₂ and particle size of F₂ was greater than that of F₃. Particle size of microspheres increase as the amount of polymer increases.

Micromeritic properties of Aceclofenac microspheres:

All the formulation (F1-F6) shows angle of repose 20.32±0.120 to 23.00±0.31 which shows good flow property. The bulk density and tapped density of all formulations ranged from 0.483 to 0.563 and 0.592 to 0.723 shows good packing property. Carr’s index ranged from 10.56 to 22.13%. F6 formulation shows lowest carr’s index indicate good compressibility. Hausner’s ratio ranged from 1.1181 to 1.2841 show good flow property.

Swelling studies:

With the help of optical microscope, dynamic swelling studies were performed and change in microsphere diameter with respect to time was measured and represented in table 7. The equilibrium swelling was dependent on the content of carboxylic acid groups and on the content of hydrophobic monomer. The incorporation of hydrophilic units increased the swelling ratio in SIF (pH 6.8), but has had an inverse effect in SGF (pH 1.2) (Davara et al. 2001).

In vitro drug release study:

A very small amount of drug (minimum 5.25%, maximum 11.14%) was released from the prepared microspheres at pH 1.2 during the 2h study. After that there was a slow but steady release reaching 75-95% after 9 h (fig 2) (Chaurasia et al. 2006) observed 95% drug release from guar gum microspheres in cecal content media.

In vitro drug release study from the aceclofenac loaded microspheres were carried out in simulated gastric and intestinal fluid to exhibit the phenomenal effect of matrix crosslinking on release profile of drug from microspheres. In vitro drug release study was performed in sink condition by bathing the dissolving solid in solvent from time to time. Results of present cumulative release vs time graph for formulation in pH 6.8 F₁, F₂, F₃, F₄, F₅, F₆ were plotted to see the effect of different ratio of guar gum and sodium alginate on release profile.

Table 1: Ingredient used in different type of formulation of Microsphere:

Formulation code	Drug (mg)	Sodium alginate (mg)	Guar Gum (mg)	CaCl₂ solution (%w/v)
F1	125	200	50	10 %
F2	125	200	50	6.5%
F3	125	175	75	6.5%
F4	125	150	100	6.5%
F5	125	125	125	6.5%
F6	125	100	150	6.5%

Table 2: Melting Point of Aceclofenac

Sr. No.	Parameters	Experimental Result	Std.
1	Melting Point (⁰C)	155⁰C	152 -153⁰C
2	Log p Value	2.15	2.17

Table 3: Solubility of Aceclofenac in different solvents

Sr. No.	Medium	Concentration of drug Soluble (mg /ml)
1	Water	2.06 (mg /ml)
2	PH 1.2 Acidic Buffer	2.90(mg /ml)
3	PH 6.8 Phosphate Buffer	3.9 (mg /ml)
4	PH 7.4 Phosphate Buffer	3.78 (mg /ml)
Result	Class of drug	BCS Class II

Table 4: Calibration of Aceclofenac in Water

Concentration (µg/ml)	Absorbance
5	0.029
10	0.048
15	0.076
20	0.105
25	0.131
30	0.155
35	0.18

Fig 1: Standard calibration curve for Aceclofenac pH 6.8

Table 5: Micromeretics properties of different microspheres

Sl No	Formulation	Angle of repose	Bulk density	Tapped density	Carr’s index (%)	Housners Ratio	Particle size (µm)
1	F1	21.57	0.500	0.601	16.80	1.202	332±10
2	F2	20.32	0.510	0.642	20.56	1.2588	291±8
3	F3	21.42	0.520	0.621	16.26	1.1942	267±5
4	F4	22.32	0.563	0.723	22.13	1.2841	251±7
5	F5	23.00	0.483	0.592	18.41	1.2256	223±11
6	F6	21.16±0.06	0.550±0.62	0.615±0.05	10.56	1.1181	332±10

Table 6: TLC of Drug and Drug Excipient

Sr. No	Samples (Pure form of Drug material) (Drug + Excipient mixture)	Retention factor of drug before the stability chamber	Retention factor of drug after the stability chamber
1	Pure drug of Aceclofenac	0.77	0.80
2	Aceclofenac + Sodium Alginate	0.79	0.81
3	Aceclofenac + Guar Gum	0.81	0.82

Table 7: Evaluation of Aceclofenac Microspheres

Formulation	Encapsulation efficacy (%)	% Swelling	In vitro drug release (pH 6.8) After 9 hour
F1	66.5%	160.5	70.56%
F2	74.2%	172	72.33%
F3	70.1%	182.5	75.23%
F4	79.2%	151.5	88.6%
F5	86.5 %	139	95.40%
F6	79.0%	304	86.10%

Table 8: Results of curve fitting of the in vitro release data from different formulations

Formulation	Correlation Coefficient (R²)				KorsmeyerPeppas	n
Formulation	Zero order	First order	Hixson Crowell	Higuchi	R²	n
F₁	0.9661	0.972	0.9732	0.9346	0.9373	0.856046797
F₂	0.9678	0.9851	0.9851	0.9477	0.9451	0.838391167
F₃	0.9391	0.9698	0.9631	0.9663	0.9388	0.669569257
F₄	0.9436	0.9819	0.9731	0.972	0.9533	0.7338518
F₅	0.9593	0.9892	0.9943	0.9519	0.9739	0.637860105
F6	0.982	0.9856	0.9959	0.9642	0.9501	0.743335716

Fig 2: In vitro release profile of different type of formulation in phosphate buffer solution of pH-6.8

CONCLUSION:

Aceclofenac loaded microsphere were prepared and evaluated to find out particle size, entrapment efficiency, swelling index and drug release profile. In wet condition the size of the beads were measured and found bigger than dry condition. The entrapment efficiency and particle size were found to be satisfacted. After evaluating the swelling index and dissolution profile both had shown similar result. It can be concluded that it is possible to modify the release rate of aceclofenac from microsphere by choosing appropriate drug polymer ratio and suitable electrolyte concentration. When compared the data of cumulative % drug release we found out that formulation F5 released 95.40% after 9hours and follows Hixson Crowell and First order release kinetics and also follows diffusion mechanism as it’s n value is 0.64 for KorsmeyerPeppas.

Aceclofenac was microencapsulated with guar gum, sodium alginate using ionotropic gelation technique. The drug loaded microsphere were lightly yellowish in colour, free flowing and almost spherical in shape. Aceclofenac microspheres were subjected to particle size, entrapped efficiency, swelling study as well as in vitro drug release, among others. The drug loaded microspheres showed drug entrapped efficiency 15.39%-38.07% and particle size 210±11 - 330±10. The swelling study showed swelling index of139-304. At pH 6.8 phosphate buffer. In vitro drug release studies were carried out up to 6 h in 0.02M phosphate buffer (pH 6.8).

CONFLICT OF INTEREST:

The authors have no conflicts of interest regarding this research.

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Received on 10.09.2022 Modified on 12.02.2023

Res. J. Pharma. Dosage Forms and Tech.2023; 15(3):163-168.

DOI: 10.52711/0975-4377.2023.00027