Determination of atomoxetine and its metabolites in conventional and non-conventional biological matrices by liquid chromatography–tandem mass spectrometry

A procedure based on liquid chromatography–tandem mass spectrometry (LC–MS/MS) is described for determination of atomoxetine (ATX) and its metabolites 4-hydroxyatomoxetine (4-OH-ATX) and N-des-methylatomoxetine (N-des-ATX) in plasma, urine, oral fluid and sweat using duloxetine as internal standard. Analytes were extracted from 0.5 mL biological fluids and sweat patch with 2 mL aliquots of tert-butyl methyl ether. The organic layer was evaporated and redissolved in mobile phase. Chromatographic separation was carried out on reverse-phase column and an isocratic mobile phase formed by 40% water and 60% 5 mM ammonium acetate, 47.2 mM formic acid, 4 mM trifluoroacetic acid in acetonitrile–water (85:15, v/v) at a flow rate of 0.5 mL/min. Separated analytes were identified and quantified by positive electrospray ionization tandem mass spectrometry and in multiple reaction monitoring acquisition mode. Limits of quantifications for the three analytes were 0.5 ng/mL plasma and oral fluid, 10 ng/mL urine and 1 ng/patch using 0.5 mL biological fluids or one sweat-patch per assay. Calibration curves were linear over the calibration ranges with r2 > 0.99. At three concentrations spanning the linear dynamic range of the assay, mean recoveries in different biological matrices were always higher than 65%. This method was applied to therapeutic monitoring of ATX and its metabolites 4-OH-ATX and N-des-ATX in conventional and non-conventional biological matrices from individuals in drug treatment.

► LC–MS/MS procedure for determination of atomoxetine and its metabolites in plasma, urine, oral fluid and sweat.
► Rapid and simple extraction of analytes from different biological matrices.
► Minimum handling and time required and high assay sensitivity.
► Method feasible for routine therapeutic monitoring of ATX and its metabolites in pharmaco-toxicological and clinical laboratories.