Abstract

A new simple, precise, accurate and stability indicating isocratic reverse phase high performance liquid chromatographic (RP-HPLC) method has been developed on Kromasil RP-C18 column (250mm X 4.6mm; 5μm) for the simultaneous estimation of Clonidine and Chlorthalidone in combined pharmaceutical formulation. At ambient temperature, the separation was achieved by using the mobile phase methanol: acetonitrile: phosphate buffer in the ratio of 50:30:20 (v/v), at pH 5.8, with a flow rate of 1.0ml/min. By using UV detector quantization was achieved at 248nm over the concentration range of 0.3-1.8μg/ml for clonidine and 15-90μg/ml for chlorthalidone. The retention time of clionidine and chlorthalidone were 3.142 min and 4.890 min respectively. Method of validation was done with respect to linearity, accuracy, precision robustness and ruggedness. Linearity of the method was determined from the equation y=65653.x+67960(R²=0.999) for clonidine and y= 2911.x + 11360(R²=0.999) for chlorthalidone. Clonidine and chlorthalione exhibited good resolution of the order 4.95. The percentage recoveries of Clonidine and Chlorthalidone were within the range of 98.197-100.206 and 98.105-101.913 respectively. Relative standard deviation of precision and ruggedness were less than 2%. The stability of the molecule was measured under different stress conditions like acidic, basic, aqueous, peroxide, thermal, light and UV light. Percentage Assay of clonidine and chlorthalidone were found to be 98.01 and 99.45 respectively. Thus, due high resolution, linearity, precision, accuracy and stability, this method can successfully be applied for the simultaneous analysis of clonidine and chlorthalidone in combined pharmaceutical dosage form.