METHOD DEVELOPMENT AND METHOD VALIDATION BY RP-HPLC OF LOPINAVIR AND RITONAVIR

Kalal Sri Ganga*, Rizwana Begum, Allakonda Rajamani and Dr. Arumugam Yasodha

ABSTRACT

A reverse phase liquid chromatographic method for estimation of Lopinavir and Ritonavir in bulk drugs and marketed pharmaceutical dosage form was developed and validated. The chromatographic conditions to achieve the highest performance parameters using Sunfire C 18 (4.6×250mm) 5μm Column with guard filter were optimized. The separation was carried out using a mobile phase containing Acetonitrile: Water (40:60% v/v) in the ratio of 40:60% v/v pumped at a flow rate of 0.9 mL/min with detection at 220 nm. The method was shown to be linear in 5–25 μg/mL and 75–375 μg/mL concentration range (regression coefficients of 0.9995 and 0.9982) for Lopinavir and Ritonavir respectively. The limit of detection (LOD) and limit of quantification (LOQ) was found to be 0.7μg/ml and 2.1μg/ml & 13.8μg/ml and 41.8μg/ml for Lopinavir and Ritonavir respectively. The accuracy of the method was assessed by adding fixed amount of pre- analyzed sample to different standard solutions (50%, 100%, and 150% of the tested concentration) in triplicate. The percentage mean recoveries were found to 98%-102%. The method was found to be precise with %RSD value was found to be within the limits for intraday and interday precision study, respectively. The method specificity and robustness were also established. New and sensitive RP-HPLC method for estimation of Lopinavir and Ritonavir has been developed, in respect to the reviewed analytical methods.

Keywords: Lopinavir and Ritonavir, RP-HPLC, Accuracy, Precision, Robustness.