INTRODUCTION:

Bioadhesion is the phenomenon between two materials, which are held together for extended periods of time by interfacial forces. It is referred as bioadhesion when interaction occurs between polymer and epithelial surface; mucoadhesion when occurs with the mucus layer covering a tissue. Generally bioadhesion is deeper than the mucoadhesion [10,22].

Flurbiprofen, a phenylalkanoic acid derivative, is one of the common drugs that are frequently used in the treatment of arthritic disorders. It is a non-steroidal anti-inflammatory, anti-pyretic, and analgesic agent that inhibits the enzymatic activity of cyclooxygenase, leading to the suppression of prostaglandin synthesis [1,8].

It is given orally in multiple doses (50–75 mg single dose for 3–4 times daily), because of its short elimination half-life (4 h), in order to achieve and maintain therapeutic concentration [21]. In addition, the oral administration of this drug causes toxicity at gastrointestinal level [22]. Because, the orally administered drugs are not able to achieve therapeutic concentration in joint cavity, therefore, a sustained release formulation of this drug for topical delivery is highly desired. Topical application of the drug prevents these side effects and offers potential advantage of delivering the drug at the site of action [6].

The U.S.P. defines gels as semisolids, either suspension of small inorganic particles or large organic molecules interpenetrated with liquid [8]. Gels are transparent or translucent semisolid formulations containing a high ratio of solvent/gelling agent. When dispersed in an appropriate solvent, gelling agents merge or entangle to form a three-dimensional colloidal network structure, which limits fluid flow by entrapment and immobilization of the solvent molecules. The network structure is also responsible for gel resistance to deformation and hence, its viscoelastic properties [15].

In this study, Flurbiprofen topical gels were formulated using natural bioadhesive polymer and were evaluated with different studies.

MATERIALS AND METHODS:

MATERIALS

Natural polymer was extracted from the ripe fruit of tamarind (Tamarindus indica) and Flurbiprofen was obtained from Sun Pharmaceutical Industries Ltd., India. Dimethyl sulfoxide and acetone was purchased from the SD Fine-chemical Ltd., Mumbai. Hydroxy ethylcellulose and Triethanolamine was purchased from High Media Pvt. Ltd, Mumbai.

METHODS

Extraction of Natural Bioadhesive Polymer

The mucilage from the natural source ripe fruit of tamarind was extracted following the method of Rao et al. The seeds of Tamarindus indica were washed thoroughly with water to remove the adhering materials. Then, the reddish testa of the seeds was removed by heating seeds in sand in the ratio of 1:4 (Seed: Sand). The testa was removed. The seeds were crushed lightly. The crushed seeds of Tamarindus indica were soaked in water separately for 24 h and then boiled for 1 h and kept aside for 2 h for the release of mucilage into water. The soaked seeds were taken and squeezed in a muslin bag to remove marc from the filtrate. Then, equal quantity of acetone was added to precipitate the mucilage. The mucilage was separated. The separated mucilage was dried at temperature 50°C, powdered and passed through sieve number 80. The dried mucilage was powdered and stored in airtight container at room temperature [4,6]..

Preparation of Topical Gel

Gels were prepared by cold mechanical method described by Schmolka et al. (1972) [14,15]. Required quantity of polymer (Natural polymer and Hydroxyethyl cellulose) was weighed and it was sprinkled slowly on surface of purified water for 2 hrs. After which it was continuously stirred by mechanical stirrer, till the polymer soaked in the water. With continuous stirring, triethanolamine was added to neutralize the gel and it maintains the pH of the gel. Now the appropriate quantity of DMSO (Dimethyl sulfoxide) was added to the gel, which behaves as the penetration enhancer, followed by the required quantity of methyl paraben as a preservative. Finally the drug Nimesulide was added to the gel with continuous stirring till drug get dispersed in gel completely. Six formulations of microparticulated intra-vaginal gel were prepared by using Natural polymer and Hydroxyethyl cellulose in different ratio. The prepared gel were packed in wide mouth glass jar covered with screw capped plastic lid after covering the mouth with an aluminum foil and were kept in dark and cool place [16, 17].

Drug Content Determination

Drug content of gel was determined by dissolving accurately weighed 1gm of gels in 0.1N NaoH. After suitable dilution absorbance was recorded by using UV- visible spectrophotometer (UV - 1700, Shimadzu, Japan) at 247 nm. Drug content was determined using slope of standard curve. ^2,11The drug content was determined by using following equation:

Drug Content = (Concentration x Dilution Factor x Volume taken) x Conversion Factor

Spreadability study of Topical gel

Spreadability was determined by apparatus suggested by Mutimer et al (1956) [18] which was suitably modified in the laboratory and used for the study. It consists of a wooden block, which was provided by a pulley at one end. By this method, spreadability was measured on the basis of ‘Slip’ and ‘Drag’ characteristics of gels [12]. A ground glass slide was fixed on this block. An excess of gel (about 2 gm) under study was placed on this ground slide. The gel was then sandwiched between this slide and another glass slide having the dimension of fixed ground slide and provided with the hook. A 1 Kg weight was placed on the top of the two slides for 5 minutes to expel air and to provide a uniform film of the gel between the slides. Excess of the gel was scrapped off from the edges. The top plate was then subjected to pull of 80 gms. With the help of string attached to the hook and the time (in seconds) required by the top slide to cover a distance of 7.5 cm be noted. A shorter interval indicates better Spreadability [16-19].

Spreadability was then calculated using the following formula:

S = M x L/ T

Where, S = is the spreadability, M = is the weight in the pan (tied to the upper slide), L = is the length moved by the glass slide and T = represents the time taken to separate the slide completely from each other.

Extrudability Study of Topical Gel

It is a usual empirical test to measure the force required to extrude the material from tube. The method applied for determination of applied shear in the region of the rheogram corresponding to a shear rate exceeding the yield value and exhibiting consequent plug flow one such apparatus is described by wood et al [19].

In the present study, the method adopted for evaluating gel formulation for extrudability was based upon the quantity in percentage of gel and gel extruded from lacquered aluminum collapsible tube on application of weight in grams required to extrude at least 0.5 cm ribbon of gel in 10 seconds. More quantity extruded better was Extrudability. The measurement of extrudability of each formulation was in triplicate and the average values are presented [19]. The extrudability was than calculated by using the following formula [19]:

Extrudability = Applied weight to extrude gel from tube (in gm) / Area (in cm²)

Swelling Index Study of Topical Gel

Swelling of the polymer depends on the concentration of the polymer, ionic strength and the presence of water. To determine the swelling index of prepared topical gel, 1 gm of gel was taken on porous aluminum foil and then placed separately in a 50 ml beaker containing 10 ml 0.1 N NaoH. Then samples were removed from beakers at different time intervals and put it on dry place for some time after it reweighed. Swelling index was calculated as follows [16, 17]:

Swelling Index (S_W) % = [(W_t - W_o) / W_o] x 100.

Where, (S_W) % = Equilibrium percent swelling, W_t = Weight of swollen gel after time t, Wo = Original weight of gel at zero time.

In-vitro Drug Diffusion Study

Cellophane membrane obtained from sigma chemicals was used for this study. In Kiescary Chien (KC) diffusion cell, 1.0 gm of gel was kept in donor compartment. The entire surface of membrane was in contact with the receptor compartment containing 85 ml of 0.1 N NaoH. The receptor compartment was continuously stirred (100 rpm) using a magnetic stirrer. The temperature maintained was 37 ± 1°C. The study was carried out for 24 hrs with the interval of 0.5, 1, 2, 4, 6, 8, 10, 12 and 24 hrs. The sample was withdrawn at predetermined period of time and same volume was replaced with fresh 0.1 N NaoH. The absorbance of withdrawn sample was measured at 247 nm to estimate Flurbiprofen [11].

Ex–vivo Bioadhesive Strength Measurement of Topical Gel

A modified balance method was used for determining the ex-vivo bioadhesive strength [19]. Fresh goat hairless skin was obtained from a local slaughter house and used within 2 hours of slaughter. The skin was separated by removing the underlying fat and loose tissues. The membrane was washed with distilled water and then with 0.1 N NaoH [20,21].The modified Patel et al (2007) [20] method was used for the measurement of bioadhesive strength. The fresh skin was cut into pieces and washed with 0.1 N NaoH. Two pieces of skin were tied to the two glass slide separately from that one glass slide was fixed on the wooden piece and other piece was tied with the balance on right hand side. The right and left pans were balanced by adding extra weight on the left hand pan. 1 gm of topical gel was placed between these two slides containing hairless skin pieces, and extra weight from the left pan was removed to sandwich the two pieces of skin and some pressure was applied to remove the presence of air. The balance was kept in this position for 5 minutes. Weight was added slowly at 200 mg/ min to the left – hand pan until the patch detached from the skin surface. The weight (gram force) required to detach the gel from the skin surface gave the measure of bioadhesive strength [19-23]. The bioadhesive strength was calculated by using following:

Bioadhesive Strength = Weight required (in gms) / Area (cm²)

RESULTS AND DISCUSSION:

Formulation Design of Topical Gel

Topical gels were prepared by using cold mechanical method using Natural Polymer and Hydroxyethyl cellulose in different ratio with other ingredients and solvents as given in Table 1 [10, 11]. All the prepared topical gel formulations contain different drug: polymer ratio and coded as FBG1, FBG2, FBG3, FBG4, FBG5 and FBG6.

Drug Content, Spreadability study, Extrudability study and Bioadhesive strength measurement of Topical Gel

From these data we have found that topical gel prepared from natural polymer having greater drug content, spreadability, extrudability and bioadhesive strength mostly NMG3 as compare to topical gel prepared from Hydroxyethyl cellulose. Table 2 shows the data for the drug content, spreadability, Extrudability, and bioadhesive strength measurement of topical gel.

Table 1. Formulation Design for the Preparation of Topical Gel

Ingredients	FBG1	FBG2	FBG3	FBG4	FBG5	FBG6
Flurbiprofen	100	100	100	100	100	100
T. Indica	100	200	300	____	____	_____
Hydroxyethyl cellulose(mg)	___	____	____	100	200	300
Triethanolamine	0.25	0.25	0.25	0.25	0.25	0.25
Dimethyl s.(gm)	1.5	1.5	1.5	1.5	1.5	1.5
Methylparaben(mg)	20	20	20	20	20	20
Distilled Water	Upto 100 gm	Upto 100 gm	Upto 100 gm	Upto 100 gm	Upto 100 gm	Upto 100 gm

Table 2. Drug Content, Spreadability study, Extrudability study and Bioadhesive strength measurement of Topical Gel

Formulation code	Drug content	Spreadability	Extrudability	Bioadhesive strength
NMG1	1.23	14.35	15.42	1.24
NMG2	1.26	15.42	16.78	1.56
NMG3	1.34	15.02	17.06	1.89
NMG4	1.16	14.67	14.43	1.07
NMG5	1.10	13.45	15.30	1.12
NMG6	1.02	13.08	16.20	1.18

Swelling Index Study of Topical Gel

From these data we found, topical gel prepared from natural polymer has greater percent swelling index mostly NMG3 as compare to topical gel prepared from Hydroxyethyl cellulose. Table 3 shows swelling index study data and figure1 shows graphical representation of swelling index study.

Table 3. Swelling Index Study of Topical Gel

Times (hrs)	% Swelling Index
Times (hrs)	FPG1	FPG2	FPG3	FPG4	FPG5	FPG6
1.0	9.56	11.24	13.48	8.65	9.24	11.20
2.0	13.24	18.67	22.05	20.40	22.30	23.06
4.0	24.45	38.90	40.65	32.89	35.02	38.56
6.0	38.90	45.23	56.87	40.60	44.86	47.07
8.0	57.34	58.34	69.52	56.06	59.78	63.67
10.0	70.52	77.39	90.42	68.80	76.45	80.20

Table 1. Swelling Index Study of Topical Gel

In—vitro Drug Diffusion Study

From these data we have found that the prepared topical gel FBG3 releases 82.46 % of drug over a period of 24 hrs. Table 4 shows the data for the in-vitro drug diffusion study of prepared topical gel. Figure 2 shows the graphical representation of in-vitro drug diffusion study of topical gel.

Table 4. In vitro Drug Release Study

Times (hrs)	% Swelling Index
Times (hrs)	FPG1	FPG2	FPG3	FPG4	FPG5	FPG6
0.0	0.0	0.0	0.0	0.0	0.0	0.0
1.0	8.56	13.24	15.48	9.65	10.24	14.20
2.0	15.24	19.67	21.05	22.40	25.30	27.06
4.0	28.45	38.90	30.65	35.89	37.02	39.56
6.0	40.90	45.23	46.87	42.60	48.86	47.07
8.0	57.34	58.34	59.52	57.06	59.78	63.67
10.0	70.52	77.39	68.42	68.80	76.45	80.20
12.0	82.05	86.90	70.78	84.85	87.56	86.90
24.0	95.60	98.35	82.46	92.80	96.89	99.82

Figure 2. In–vitro Drug Release Study of Topical Gel

CONCLUSION

Topical gels of Flurbiprofen were successfully prepared by using cold mechanical method using Natural Polymer. We have concluded that the topical gel prepared from the natural polymer having good spreadability, Extrudability and bioadhesive strength. So the topical gel prepared from natural polymer will be greatly for making an ideal topical preparation. FP3 has the greater swelling index properties in comparison of others it means topical gel prepared from natural polymer having the greater swelling tendency. From the In - vitro drug diffusion study we have concluded that the gel prepared from the natural polymer, controls the release of drug for longer period of time which will be helpful to avoid the more fluctuation and also reduces the cost of therapy.

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