ABSTRACT:
An accurate, sensitive, precise, rapid, reversed phase HPLC (RP-HPLC) method for the determination of related substances of Prulifloxacin in tablet dosage form has been developed and validated. The best separation was achieved using a Kromasil C18 250 x 4.6mm; 5µ HPLC column (Akzo Nobel) at 40°C employing a gradient elution. Mobile phase consisting of solvent A (solution containing 4 mL of Triethylamine in 1000 mL water, adjusted to pH 3.4 with ortho-phosphoric acid), solvent B (Acetonitrile) and solvent C (Methanol) was used at a flow rate of 1.0 ml min-1. UV detection was performed at 275 nm. This method can separate all known impurities and unknown impurities of Prulifloxacin in a run time of 60 minutes. The method was validated as per ICH guidelines, demonstrating to be accurate and precise (repeatability and intermediate precision level) within the corresponding linear range of known impurities The Specificity of the method was established from the peak purity indices obtained with the aid of PDA detection and satisfactory resolution between related impurities. The formulation was subjected to oxidation, hydrolysis, photolysis, and heat as stress conditions. Relevant degradation was found to take place under oxidative conditions. The method was linear over the concentrations ranging from LOQ to 200% of the limit concentration. Robustness against small modification in pH, column oven temperature, flow rate and percentage of the mobile phase composition was ascertained. All these results provided a validated method for determination of related substances of Prulifloxacin in tablet dosage form.