Revista de Bioanálise e Biomedicina

High Performance Liquid Chromatographic Method for the Determination of Clobetasol in Rat Plasma and its Application to Skin Penetration

Abstract

Prashant B. Musmade, Praful B. Deshpande, Swapnil J. Dengle, Kranti B. Talole, Shriram M. Pathak, Adiga MNS, Krishnamurthy Bhat, Nayanabhirama Udupa and K. Laxminarayana Bairy

A simple a nd sensitive high per for ma nce liquid chromatography (HPLC) method was developed for qua ntification of clobeta sol (CLB) in r a t pla sma . Aripiprazole was used as an internal standard (IS). The present method uses protein precipitation with acetonitrile, followed by the liquid-liquid extraction with methyl tertiary butyl ether for extraction of the CLB from the matrix. Separation was carried out using Variance C 18 (250.0 × 4.6 mm, 5 μ m particle size) column and the effluent was monitored by an ultra violet (UV) detector at 240 nm. The mobile phase used was acetonitrile: phosphate buffer (pH 7.0; 25.0mM containing 0.2 %v/v triethylamine) (65: 35 % v/v) at a flow rate of 1.0 ml/min. The present method was va lida ted a s per the United States Food and Dr ug Administration (USFDA) guideline. This method was linear over the range of 25.0-1000.0 ng/ml with regression coefficient greater than 0.99. The mean recovery of CLB and IS were 70.8±1.9 and 83.32 ± 1.55 % respectively. The method was found to be precise, accurate and specific. The developed HPLC method was applied for the estimation of CLB in plasma after topical application of various zinc salts in rats to study the penetration of CLB through rat skin.