Objective: The present research work aims to develop and validate a selective and highly sensitive method for the determination of apixaban in human plasma using liquid chromatography-tandem mass spectrometry (LC-MS/MS).
Methods: 200 µl of sodium heparin plasma samples were acidified and clean-up was performed by using solid-phase extraction (SPE). Apixaban 13C D3 was used as an internal standard (deuterated) to lower the relative matrix effects and a single step SPE was employed for sample clean up. 10 µl of SPE eluent was loaded onto Hypersil Beta Basic C18, 100×4.6 mm, 5 µ column for highly selective chromatographic separation using an isocratic mobile phase. 2 mmol ammonium acetate in water and acetonitrile were delivered by using a quaternary low-pressure gradient pump without premixing at a minimum flow rate of 0.50 ml/min.
Results: LC-MS/MS method was successfully developed and validated to demonstrate the lowest detection limit of 0.05 ng/ml and a linear dynamic range from 1-250 ng/ml with r2>0.99. Method development and validation results proved that the method is selective and highly sensitive for the determination of apixaban in human plasma using LC-MS/MS.
Conclusion: Current method can be applied for both therapeutic drug monitoring (TDM) and pharmacokinetic (PK) study analysis.
METHOD DEVELOPMENT AND VALIDATION OF SELECTIVE AND HIGHLY SENSITIVE METHOD FOR DETERMINATION OF APIXABAN IN HUMAN PLASMA USING LIQUID CHROMATOGRAPHY-TANDEM MASS SPECTROMETRY