A simple high-performance liquid chromatography and ultraviolet spectrophotometric methods were developed and validated for the estimation of alprazolam and melatonion simultaneously in combined pharmaceutical solid dosage form. The wavelength fixed for Q-analysis measurement the two wavelengths fixed are 221.0 nm and 268.0 nm. The linearity obtained for alprazolam in the range of 2-10 μg/ml and 10-50 μg/ml for melatonin by spectrophotometric method with r square value greater than 0.999. The alprazolam and melatonin were analyzed by HPLC using reverse phase C18 column (150 x 4.6 mm i.d., 5 μm particle size), by isocratic mobile phase consisted of acetonitrile and water (80:20 v/v), at a flow rate of 0.8 ml/min in the detection wavelength of 223 nm. The linearity and range of alprazolam are 0.5-2.5 μg/ml and 3-15 μg/ml for melatonin by HPLC method with r square value greater than 0.998. Accuracy of all methods was determined by recovery studies and showed % recovery between 99 to 101%. Intraday and interday precision was checked for all methods and mean %RSD was found to be less than 2 for all the methods. The methods were successfully applied for estimation of alprazolam and malatonin in marketed formulation.
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