Forced Degradation Study for Tenofovir DF, Lamivudine and Efavirenz in Triple Combination Anti-Retroviral Tablets and Development of Validated Stability Indicating Assay Method by UPLC

Abstract

Introduction: Tenofovir Disoproxil Fumarate/Lamivudine/Efavirenz Tablets are a pharmaceutical dosage form indicated for the treatment of Human Immunodeficiency Virus (HIV) infection. Methods: A simple, time efficient and stability indicating Reverse Phase Ultra Performance Liquid Chromatography (RP – UPLC) method was developed for the simultaneous determination of the three drugs present in the tablets. Mobile phase-A is 0.1% Trifluoro Acetic acid in water and mobile phase-B is acetonitrile in gradient elution mode. Flow rate at 0.4 mL/min was fixed in the method. Acquity BEH Phenyl (100 mm × 2.1 mm), 1.7 µ column was used for the separation of analytes at 260 nm wavelength. The developed method has been validated as specified in International Conference on Harmonization (ICH) guideline. Forced degradation study was conducted for Tenofovir Disoproxil fumarate (TDF), Lamivudine (LAM) and Efavirenz (EFA) pharmaceutical tablets to identify the degradation behaviour of individual drugs under stress study. Results: The method was observed to be linear from 6-90 µg/mL concentration for TDF & LAM and 12-180 µg/mL for EFA with correlation coefficient values of more than 0.999. The method was found to be precise with RSD (Relative standard deviation) values of below 2% for replicate measurements. Recovery results were found to be between 99.0 and 100.8%. LOQ values for TDF, LAM & EFA were 0.041 µg/mL, 0.034 µg/mL & 0.053 µg/mL, respectively. Specificity of the analytical method was checked by injecting the stressed samples and evaluating the homogeneity of peaks using Photo Diode Array (PDA) detector. Peak purity results indicated that no interference was observed from degradation impurities for the quantification of TDF, LAM and EFA in the dosage form. The method was found to be robust under the deliberately modified chromatographic parameters. Conclusion: The developed UPLC method is stability indicating, successfully validated and suitable for quantification purpose.