New Validated Liquid Chromatography Coupled with Tandem Mass Spectrometry Method Development for Quantification of Tenofovir in Human Plasma Co-administered with Dolutegravir and Lamivudine

Abstract

Aim: A selective and reproducible liquid chromatography coupled with tandem mass spectrometry method was developed for the quantification of Tenofovir in human plasma, in the presence of Dolutegravir and Lamivudine. Methodology: The method employed solid-phase extraction using strong cation exchange (SCX) cartridges and chromatographic separation on a ZORBAX SB C8 column with mobile phase 0.1% formic acid in water: acetonitrile (90:10, v/v). Detection was achieved via electrospray ionization in positive mode, monitoring multiple reaction monitoring transitions of Tenofovir and its deuterated internal standard. Results and Discussion: The retention time for Tenofovir was approximately 1.70 min, with a total run time of 3.0 min/sample. The calibration curve was linear over the range of 3.0–1055.7ng/mL, with correlation coefficients consistently above 0.9988 and an lower limit of quantification (LLOQ) of 3.052 ng/mL, demonstrating excellent sensitivity. Mean recovery for Tenofovir ranged from 59.88% to 62.11%, with a percent coefficient of variation below 1%, confirming the reproducibility of the extraction process. Conclusion: The method was validated as per regulatory guidelines, demonstrating acceptable precision, accuracy, recovery, and stability across a wide concentration range. This assay is suitable for pharmacokinetic, bioequivalence, and therapeutic drug monitoring applications in HIV patients receiving combination therapy.