Development And Validation of Stability Indicating RP-HPLC Method For Simulataneous Determination of Darunavir Ethanolate and Ritonavir in Tablet Dosage Form and Forced Degradation Study by QBD Approach

Abstract

A simple, precise and rapid stability indicating RP-HPLC method has been developed for the simultaneous estimation of Darunavir ethanolate and Ritonavir in Pharmaceutical dosage form in presence of degradation products. It involved Zorbax bonus, 150 mm x 4.6 mm of 3.5 μm particles packing stationary phase. The separation was achieved using The mobile phase A consisted of mixture of ammonium acetate buffer and methanol -50:50(%v/v) Mobile phase B-ammonium acetate buffer and ACN-45:55(%v/v) at flow rate of 1.2 ml/min. The effluent is monitored at 240 nm. The retention time of Darunavir ethanolate and Ritonavir was found to be 7.512 and 16.654 respectively. The total run time was 35 minutes within which two active compounds and their degradation products were separated by applying QbD approach. The optimized method was applied for forced degradation study by Quality by Design. Various factors affecting the method were optimized using a d-optimal design. Point verification of actual versus optimized predicted trials was performed. A Design space in which the method was robust could be generated successfully. Acid, alkali, and Peroxide degradation was carried out and significant degradation product was achieved .The method was found to be specific enough to separate degradation products from main analytes. The method was linear in the range of 240-360 μg/ml (R²=0.999) and 30-70 μg/ml (R² =0.999) for Darunavir ethanolate and Ritonavir, respectively. The described method was validated with respect to system suitability, specificity, linearity, accuracy, precision and robustness. Result of each parameter was met with its acceptance criteria Key words: Darunavir Ethanolate, Ritonavir, RP-HPLC, validation, stability and Degradation products