Direct and rapid quantitation of ephedrines in human urine by paper spray ionization/high resolution mass spectrometry

•A paper spray ionization/mass spectrometry method was developed for quantitative analysis of ephedrines in human urine.•The urine samples were directly analyzed without sample preparation and chromatographic separation.•Accuracies for WADA’s blind urine sample were ranged from 96.4 to 106.1%.•The present method could be used for the fast quantitation of ephedrines in doping control analysis. A rapid and direct paper spray ionization/mass spectrometry (PSI/MS) method was developed for quantitative analysis of ephedrine, pseudoephedrine, norpseudoephedrine, and methylephedrine in human urine. This method involves the use of a triangular filter paper and high-resolution mass spectrometry, where the molecular ions of ephedrines were generated by applying high voltage after loading the spray solvent to the paper which urine sample was pre-loaded. Small amounts (2μL) of urine spiked with an internal standard were directly analyzed for ephedrines. The PSI/MS method was validated for linearity, within- and between-run precision, accuracy, and limit of detection. The results showed good linearity (R2≥0.9928) and acceptable precision and accuracy. Furthermore, the accuracy of the method was assessed by analyzing a blind urine sample from World Anti-Doping Agency and comparing the measured concentrations with the nominal concentrations. This test resulted in accuracies ranging from 96.4 to 106.1%, indicating that the PSI/MS method has the potential to be an alternative technique for the fast quantitation of ephedrines in doping control analysis.