Simultaneous determination of aceclofenac and its three metabolites in plasma using liquid chromatography–tandem mass spectrometry

A new LC/MS/MS-based method allows simultaneous determination of aceclofenac and its three metabolites (4′-OH-aceclofenac, diclofenac, and 4′-OH-diclofenac) in plasma. After acetonitrile-induced precipitation of proteins from the plasma samples, aceclofenac, 4′-OH-aceclofenac, diclofenac, 4′-OH-diclofenac, and flufenamic acid (an internal standard) were chromatographed on a reverse-phase C18 analytical column. The isocratic mobile phase of acetonitrile/0.1% formic acid(aq) [80:20 (v/v)] was eluted at 0.2 mL/min. Quantification was performed on a triple–quadrupole mass spectrometer employing electrospray ionization, and the ion transitions were monitored in multiple reaction-monitoring mode. The monitored transitions for aceclofenac, diclofenac, 4′-OH-diclofenac, 4′-OH-aceclofenac and flufenamic acid were m/z 352.9 → 74.9, 296.1 → 251.7, 311.8 → 267.7, 368.9 → 74.9, and 279.9 → 235.9, respectively. The coefficient of variation of the assay precision was less than 6.5%, and the accuracy ranged from 93% to 103%. The limits of detection were 2 ng/mL for aceclofenac and 0.2 ng/mL for both diclofenac and 4′-OH-diclofenac. This method was used successfully to measure the concentrations of aceclofenac and its three metabolites in plasma from healthy subjects after administration of a single 100-mg oral dose of aceclofenac. This analytic method is a very simple, sensitive, and accurate way to determine the pharmacokinetics of aceclofenac and its metabolites.