Novel RP-UPLC Method Development and Validation for Simultaneous Quantification of Emtricitabine, Tenofovir and Efavirenz in Bulk and Tablet Dosage Forms

Abstract

Objective: The objective of this work is to develop a precise, accurate and validated reverse phase ultra-performance liquid chromatographic technique for effective simultaneous determination of Emtricitabine, Tenofovir and Efavirenz in bulk and tablet formulation. Method: Separation of the selected drugs was optimized after several trials including changing mobile phase and its composition, stationary phase, flow rate, column temperature, etc. Finally the separation of drugs was achieved on BEH C18 column using a mixture of methanol and phosphate buffer having pH 3.5 in the ratio of 65:35 v/v as mobile phase with flow rate of 0.3 ml/min and the analytes were detected at a wavelength of 260 nm. Results: The developed method was validated by determining the parameters like linearity, system suitability, recovery, precision, specificity, robustness, ruggedness, LOD, and LOQ as per the International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use (ICH) guidelines. The system suitability parameters were within the limits, retention times (Rt) for Emtricitabine, Tenofovir and Efavirenz were found to be 0.432, 0.671, and 2.772 min respectively. The method showed linearity between the concentration range of 10-50 μg/ml for Emtricitabine (r2= 0.9987), 15-75 μg/ml for Tenofovir (r2= 0.9983) and 30-150 μg/ml for Efavirenz (r2= 0.9982). The percentage recovery results at 50%, 100% and 150% of Emtricitabine, Tenofovir and Efavirenz were found to be in the range of 99.45 % - 100.15 %. Since there was no interference due to excipients and mobile phase, the method was found to be specific. The assay results of the combined tablet dosage form by the developed method were identified as in good agreement with the acceptance limit. Conclusions: The current method was proved to be effective for routine simultaneous determination of Emtricitabine, Tenofovir and Efavirenz in bulk and tablet formulation.