Abstract

A very simple and rapid bio-analytical method with high-throughput, appropriate sensitivity, selectivity, accuracy and precision has been developed and validated for the quantification of acetylsalicylic acid (ASA) and its main metabolite salicylic acid (SA) in human plasma using hydrochlorothiazide (HCTZ) as internal standard (IS) by high performance liquid chromatography tandem mass spectrometry (LC-MS/MS) with electronspray ionization (ESI). A simple technique of liquidliquid extraction was developed for plasma samples preparation of the analytes and the chromatographic separation was done under isocratic conditions by Phenomenex Kinetex C18 column (50 mm x 3 mm, 5 µm) with a run time of 1.8 min. The composition of mobile phase was acetonitrile and 10 mM ammonium formate in water (90:10, v/v). The precursor ion was deprotonated to product ion transitions for ASA, SA and HCTZ (IS) were measured on an operating in the multiple reaction monitoring (MRM) with negative ion mode by triple quadrupole mass spectrometer. The assay exhibited a calibration range of 10-15000 ng/mL for both ASA and SA. The bio-analytical validated method was successfully applied for bioavailability or bioequivalence studies to analyze human plasma samples.

Keywords: Acetylsalicylic acid, liquid chromatography, salicylic acid, tandem mass spectrometry.

Graphical Abstract