RP-HPLC method development and validation for the simultaneous estimation of dolutegravir, emtricitabine, and tenofovir alafenamide in tablet dosage form

Abstract

Antiretroviral combination therapy regimens comprising of a nucleoside reverse transcriptase inhibitor (Emtricitabine, EMC), integrase inhibitor (Dolutegravir, DTG), and nucleotide reverse transcriptase inhibitor (Tenofovir Alafenamide, TAF) are frequently prescribed for HIV patients. The objective of this research was to establish and validate a precise and reliable method for the simultaneous estimation of DTG, EMC, and TAF, respectively in a marketed tablet dosage form using RP-HPLC system. RP-HPLC (Jasco) method was developed using Qualisil-5 BDS C18 column (250 mm × 4 mm, 5 μm). The mobile phase consists of acetonitrile: orthophosphoric acid (0.1%) adjusted in water pH 4.7 with triethylamine in the composition of 43:57 v/v at a flow rate of 1.2 mL/min using UV detection at 271 nm at ambient column temperature, keeping the injection volume 20 μL. The retention times of DTG, EMC, and TAF were found to be 8.321 mins, 2.210 mins, and 4.089 mins, respectively. All the criteria for the validation (linearity, accuracy, precision, and robustness) were observed to be within the acceptation range. The described method was linear over a concentration range of 2-12 μg/mL, 8-48 μg/mL, and 1-6 μg/mL for the assay of DTG, EMC, and TAF, respectively with r2 values of 0.999 in each case. High recovery of ~99.5% was observed in all cases with a % RSD of <2.This method was successfully developed to estimate the concentration of DTG, EMC, and TAF in tablet dosage form simultaneously owing to high precision, reproducibility, and accuracy attributes.. Keywords: RP-HPLC, Dolutegravir, Emtricitabine, Tenofovir Alafenamide, Simultaneous estimation, Validation.