INTRODUCTION:
Delivering medicine to the systemic circulation via skin is seen to be a desirable alternative for administrating it by mouth. The penetration across skin layer is a slow process due to the effect of the barrier properties. The skin, in particular the stratum corneum, possesses a barrier to drug penetration due to its high density (1.4 g/cm2 in dry state) and its low hydration of 15 to 20%. The barrier function is further facilitated by the continuous replacement of the stratum corneum.
It offers many important advantages over oral drug delivery, e.g., avoids gastrointestinal tract and hepatic first-pass metabolism, reduces variations in delivery rates, avoids interference due to the presence of food, controls absorption rate, increases patient compliance, suitable for unconscious patients, and enables fast termination of drug delivery, if needed.[1] The technique is generally non-invasive so it is well accepted by patients and can be used to provide local as well as systemic delivery over several days.[2] Limitations include slow penetration rates, drug formulation may cause skin irritation, patient may develop contact dermatitis, and be restricted to relatively low dosage drugs.
There are a number of routes by which a molecule can cross the stratum corneum, these are intercellular, transcellular, and transappendageal. Both polar and non-polar substances diffuse via transcellular and intercellular routes by different mechanisms. [3] The transappendageal route transports substances via sweat glands and the hair follicles with their associated sebaceous glands. This route is considered to be of minor importance because of its relatively small area. There has been much debate over the past decades on the route of penetration but experimental evidence suggests that, under normal circumstances, the predominant route is through the intercellular spaces. [4-6] Transdermal systems are ideally suited for diseases that demand chronic treatment. Hence, anti-diabetic agents of both therapeutic and prophylactic usage have been subjected to transdermal investigation.
Diabetes mellitus is a chronic metabolic disorder characterized by high blood glucose concentration hyperglycemia caused by insulin deficiency, often combined with insulin resistance. [4-6] Glibenclamide is an oral sulfonylurea hypoglycemic agent. It is currently available for treating hyperglycemia in Non-insulin dependent diabetes mellitus. This drug inhibits ATP-sensitive K+ channel in pancreatic beta cells. This inhibition causes cell membrane depolarisation, opening of voltage dependant calcium channels. Plasma half of Glibenclamide is 4-6 h which makes frequent dosing necessary to maintain the therapeutic blood level of the drug for long term treatment. Hence, control release transdermal preparation of Glibenclamide was prepared to provide sustained effect as compared to conventional multiple oral dosing [7-9]. HPMC 5cps and chitosan are used for preparing the matrix type patch because of their nature; HPMC is hydrophilic, natural polymer chitosan on the release pattern is studied.
Fig 1: A cross-section of human skin, showing various skin tissue layers and appendages.
MATERIALS AND METHODS:
Materials:
Glibenclamide was purchased from Alapati pharma. (ongole, India). Chitosan was the gifted sample from central Institute of Fischeries Technology, Cochin, India. HPMC 5cps, Acetic acid, Glycerine, Ethanol were purchased from Finar chemicals, (Ahemadabad, India). All other chemicals and reagents used were of laboratory or analytical grade.
Methods:
Drug Excipient interaction:
The infrared (IR) spectra were recorded using an FTIR spectrophotometer by the KBr pellet method in the wavelength region between 4000 and 400 cm-1. The base line correction was done using dried KBr. Infrared spectra of the mixture were taken over a wave number range. The spectra obtained for Glibenclamide, and physical mixtures of Glibenclamide with other excipients were compared to check the compatibility of the drug with other excipients.
Physico-chemical Characterization Study:
Physicochemical studies are usually associated with great precision accuracy and in the case of a new drug substance would include solubility and melting point.
A. Drug Characterization:
Solubility Analysis [10]
Slightly soluble in ethanol (95%) and methanol. Sparingly soluble in di-chloromethane. Insoluble in water and ether. Dissolves in diluted solutions of alkali hydroxides.
Procedure:
The solubility studies were performed in phosphate buffer solution, as the procedure described in USP. Excess amount of drug was added to the phosphate buffer pH 7.4 in each case and keeping it on a water bath shaker for 24hrs at 320C. After 24hrs, solutions were analyzed spectrophotometrically at 228 nm, which was the absorption maxima determined earlier.
B. Development of Calibration data curve of Glibenclamide:
Stock Solution:
10mg of Glibenclamide was accurately weighed and dissolved in a required quantity of methanol and make up 100 ml with phosphate buffer (100μg/ml).
Procedure:
Aliquots equivalence 0.2, 0.4, 0.6, 0.8, 1.0, 1.2, 1.4, 1.6, 1.8, 2μg/ml respectively were drawn from stock solution and made up to 10 ml by using phosphate buffer pH 7.4 These solutions were scanned and the absorbance was measured at 228nm against blank. The absorbance values thus obtained were plotted in graph of concentration versus absorbance.
FORMULATION OF DRUG LOADED FILMS[11-15]
Solvent Casting technique was employed in the present work for the preparation of drug films. Solution of plain Chitosan and Chitosan/ HPMC blend were prepared by dissolving the polymer in 1.0% w/v acetic acid solution, and the solution of HPMC was prepared by dissolving in a mixture of water and ethanol (8:2) respectively. To the above polymeric solution 20%, 30%, w/w (with respect to dry weight of polymer) of glycerol was added. Glycerol was used as a plasticizer in the preparation of films.40mg of Glibenclamide was added in small portions of chloroform and stirred for 20mins.Drug containing polymeric solution (50ml) were poured into a petridish (30.8cm2), and kept in an oven at 40◦c for complete drying. The dried films were removed from the Petri dish and stored in desiccators until use.
Table 1: Composition of drug loaded films
|
Formulations |
Drug (mg) |
Glycerol |
Polymers |
Polymer 100ml (%) |
|
D1 |
40 |
20 |
Plain Chitosan |
2 |
|
D2 |
40 |
30 |
Plain Chitosan |
2 |
|
D3 |
40 |
30 |
Plain Chitosan+HPMC |
1+1 |
|
D4 |
40 |
30 |
Plain Chitosan+HPMC |
1.5+0.5 |
|
D5 |
40 |
20 |
Plain HPMC |
2.5 |
|
D6 |
40 |
30 |
Plain HPMC |
2.5 |
EVALUATION TESTS:
A. Physical Characterization:
1. Physical appearance:
All the transdermal films were visually inspected for color, clarity, flexibility and smoothness.\
2. Tensile strength[16]:
The films were evaluated using a texture analyzer (Instron Universal Model) equipped with a 500gm load cell. Film strip in 10 mm x 10 mm of dimension and free from air bubbles or physical imperfections, was held between two clamps positioned at a distance of 1 cm. During measurement, the film was pulled by top clamp at a rate of 10 mm/minutes. The force and elongation was measured when the films broke. Measurements were run four times for each film. The tensile strength and elongation at break were calculated as below:
Tensile strength (kg/mm2)=Breaking force (kg)/ cross section area of sample (mm2)
Elongation at break (%)=Increase in length at breaking point (mm)/ Original length (mm) x 100%
3. Swelling Index:
Weighed pieces 1x1 cm2 of film were immersed in distilled water; at 0.5, 1, 2, 4, 8 and 24 hours. Soaked films were removed from the medium at predetermined time, blotted to remove excess liquid and weighed immediately. The swelling index was calculated from the weight increase, as follows9:
Swelling Index = (w2-w1)/ w1
Where, w1 and w2 are the weight of the film before and after immersion in the medium, respectively.
4. Weight uniformity[17]:
The films of different batches were dried at 60◦ c for 4 hours before testing. Five patches from each batch were accurately weighed in a digital balance. The average weight and the standard deviation values were calculated from the individual weights
5. Thickness of the films:
The thicknesses of the drug-loaded polymeric films were measured at five different points using a digital micrometer. The average and standard deviation of five reading were calculated for each film.
6. Folding endurance[18]:
The folding endurance was measured manually for the prepared films. A strip of film (2x2 cm) was cut evenly and repeatedly folded at the same place till it broke. The number of times the film could be folded at the same place without breaking gave the exact value of folding endurance13.
7. Water Vapor Transmission (WVT)[19]:
The film was fixed over the glass vial with an adhesive containing 1 g of fused calcium chloride as a desiccant. Then the vial was placed in a desiccators containing saturated solution of potassium chloride 200ml (RH 84%). The cells were taken out and weighed after 1, 2, 3, 4, 5, 6 and 7 days of storage. Water vapor transmissions were calculated by taking the differences in the weight of the film before and after the study at regular intervals of 24 h for a total period of seven days.
8. Water Vapor Absorption (WVA):
Water vapor absorption was calculated by taking the differences in the weight of the film before and after the study at regular intervals of 24 h for a total period of seven days. For the determination of water vapor absorption studies of polymer films,3.14 cm2 areas was taken and weighed accurately and then placed on a wire gauge, which was kept in a desiccators containing a saturated solution of potassium bromide (200 ml). The humidity was found to be 84% RH. The films were taken out and weighed after 1, 2, 3, 4, 5, 6 and 7 days of storage.
9. Percentage moisture content:
The prepared films were weighed individually and kept in a desiccators containing calcium chloride at room temperature for 24 hours. The films were weighed repeatedly until they showed a constant weight. Values for the percentage of moisture content were calculated using the formula:
Percentage of moisture content=[Initial weight–Final weight/Final weight] x100
10. Percentage moisture uptake:
The weighed films were kept in desiccators at room temperature for 24 hours and then exposed to 84% RH using a saturated solution of potassium chloride. The films were weighed repeatedly until they showed a constant weight.A value for the percentage of moisture uptake was calculated using the formula:
Percentage of moisture uptake = [Final weight – Initial weight/Initial weight] x100
11. Flatness[20, 21]:
Longitudinal strips were cut out from each film, one from the center and two from either side. The length of each strip was measured and the variation in the length because of non-uniformity in flatness was measured by determining percent constriction, considering 0% constriction is equivalent to 100% flatness.
Percentage of constriction=L1-L2/ L2×100
Where,
L1 =Initial length of each strip,
L2=Finallength of each strip.
12. Drug content[22]:
Transdermal films of specified area (3.066 cm2) was cut into small pieces and taken into a 50 ml volumetric flask and 25 ml of phosphate buffer pH7.4 was added, gently heated to 45◦C for 15 minutes, and kept for 24 hours with occasional shaking. Then, the volume was made up to 50 ml with phosphate buffer of pH 7.4. Similarly, a blank was carried out using a drug-free patch. The solutions were filtered and the absorbance was measured at 228 nm.
13. In vitro drug release studies[23]:
A paddle over disc assembly (USP 23, Apparatus 2) was used for the assessment of release of drug. The TDDS patch was mounted on the disc and placed at the bottom of the dissolution vessel. The dissolution medium was 900 ml phosphate buffer of pH 7.4. The apparatus was equilibrated to 37±0.5◦Cand operated at 50 rpm. The samples (5 ml aliquots) were withdrawn at appropriate time intervals up to 8 hours and analyzed on a UV spectrophotometer at 228nm.
RESULTS:
PREFORMULATION STUDIES:
Drug excipient interaction study:
There were no changes in the major peaks of Glibenclamide in the presence of chitosan and HPMC 5cps.So the drug and the excipients are compatible with each other.