A reverse phase liquid chromatography (LC) method was developed and validated for simultaneous estimation of pilocarpine nitrate and chlorbutanol in their gel dosage form. The isocratic LC analysis was performed on phenomenex gemini ODS C18 column (200 mm x 4.6 mm, 5 μ) using mobile phase composed of water (pH 4): methanol (50:50, v/v) at a flow rate of 1.0 ml/min. Quantitation was performed using UV detector at 260 nm. The retention times were found to be 3.273 min for pilocarpine nitrate and 4.807 min for chlorbutanol. The analytical method was validated according to ICH guidelines. The linearity was observed in the range of 20-60 μg/ml and 2.5-7.5 μg/ml with correlation coefficient, r=0.9998 and 0.998 for pilocarpine nitrate and chlorbutanol respectively. The accuracy (%recovery) was found to be 99.25 – 101.41% for pilocarpine nitrate and 98.92 – 101.505 % for chlorbutanol. The relative standard deviation values for repeatability and intermediate precision studies were less than 2%. The method was successfully applied for market sample analysis and mean percentage assay values were 99.34±0.796 and 99.527±1.083 for pilocarpine nitrate and chlorbutanol respectively.
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