Development and Validation of a Liquid Chromatography-Tandem Mass Spectrometry Analytical Method for the Concurrent Quantification of Cabotegravir and Rilpivirine in Rat Plasma

Objective: To establish and authenticate a precise and dependable liquid chromatography-tandem mass spectrometry (LC-MS/MS) technique for the concurrent quantification of Cabotegravir (CAB) and Rilpivirine (RIL) in rat plasma, specifically for pharmacokinetic studies. Methods: A protein precipitation extraction procedure was employed, followed by chromatographic separation on a Kromasil C18 column using ethanol and 0.1% triethylamine buffer (30:70, v/v, pH 2.5) as the mobile phase. Quantification was performed using an electrospray ionization interface coupled with a SCIEX QTRAP 5500 mass spectrometer in multiple reaction monitoring mode. The mass transitions for CAB (m/z 406.1240 → 218.3257), RIL (m/z 367.2103 → 174.1127), and the internal standard Raltegravir (m/z 445.1147 → 389.5261) were optimized for maximum sensitivity. Results: Witnessed excellent linearity (r2 > 0.999), precision (≤15% CV), and accuracy (85–115%) for both intra- and inter-day assays. Recovery rates for CAB and RIL ranged from 94% to 98%. The method exhibited negligible matrix effects, with lower limit of quantitation of 15 ng/mL for CAB and 10 ng/mL for RIL. Conclusion: This approach provides a robust and sensitive approach for the concurrent quantification of CAB and RIL in rat plasma, suitable for pharmacokinetic studies, offering a reliable tool for future clinical and pre-clinical research on these drugs.