A simple, accurate, precise sensitive, repeatable and stability indicating RP-UPLC method was developed for simultaneous determination of metronidazole (MET) and ofloxacin (OFL) in combine dosage form. The method was developed by using reverse phase C18 Purosher star (100 mm × 2.1 mm id, 2µm particle size) as stationary phase with phosphate buffer: acetonitrile (70:30 %v/v) as a mobile phase, pH was adjusted to 2.5 by ortho-phosphoric acid at a flow rate of 0.5 ml/min and and column temperature maintained at 45°C. Quantification of eluted compound was achieved with PDA detector at 317 nm. The combination drug product is exposed to thermal, acid/base hydrolytic and oxidative stress conditions, and the stressed samples were analyzed by proposed method. Metronidazole and ofloxacin followed linearity in the concentration range of 5-35 µg/ml and 2.5-17.5 µg/ml with r2=0.999 (n=6) respectively. Limit of detection (LOD) and limit of quantification (LOQ) values for Metronidazole was 0.162 and 0.492 µg/ml and for ofloxacin was 0.067 and 0.203 µg/ml respectively. Chromatographic peak purity data of MET and OFL indicated no co-eluting peaks with the main peaks of drugs which demonstrated the specificity of assay method for their estimation in presence of degradation products.The validation study is carried out as per International Conference on Harmonization (ICH) guidelines. This method was also successfully applied for estimation of metronidazole and ofloxacin in pharmaceutical formulation.
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