Ultraperformance liquid chromatography tandem mass spectrometric method for direct quantification of salbutamol in urine samples in doping control

Abstract

A fast and reliable quantitative method for salbutamol using direct analysis of the urine sample by ultraperformance liquid chromatography tandem mass spectrometry (UPLC/MS/MS) has been developed. Urine samples were spiked with salbutamol-d 6 (internal standard), and, then, they were diluted with ultrapure water (1:1, v/v). Aliquots of 1 μl of the mixture were directly analyzed by UPLC/MS/MS. The chromatographic separation was performed in a UPLC BEH C 18 (100 mm × 2.1 mm, 1.7 μm) column with a mobile phase contained 0.01% formic acid in ultrapure water (v/v) and 0.01% formic acid in acetonitrile (v/v), using gradient elution at 0.6 ml/min. The temperature of the column was set to 45 °C. The total run time was 3.2 min. Electrospray ionization in positive ion mode was used under multiple reaction monitoring (MRM) at different collision energies. Nitrogen and argon were used as desolvation and collision gas, respectively. The method was shown to be linear from 200 to 5000 ng/ml ( r 2 > 0.99). The limit of quantitation was estimated in 200 ng/ml. Intra-assay precision and accuracies, evaluated by using quality control samples containing 550 and 1100 ng/ml salbutamol, were always better than 8.4%. The intermediate precision was estimated to be in the range of 5.6–8.9%. The method was shown to be reliable when applying to routine samples, and the short analysis time resulting from a simple sample preparation and a fast instrumental analysis makes it of great interest for antidoping control purposes.