Analytical methodology for enantiomers of salbutamol in human urine for application in doping control

Liquid chromatographic procedure with fluorimetric detection for chiral sep aration and quantification of salbutamol enantiomers in urine samples has b een developed. The extraction of free salbutamol from urine has been consid ered using liquid-liquid and solid-phase procedures. The effect of pH, salt ing-out effect and organic solvent has been studied in liquid-liquid extrac tion from aqueous and urine samples. For solid-phase extraction, different mechanisms (polar, non-polar, cation-exchange and interactions with a polym eric phase) have been tested and the effect of the urine matrix on the extr action recoveries has been considered. Bond-Elut Certify(TM) extraction car tridges provided the best specificity and good recoveries for salbutamol in urine. The sample is acidified, applied to the preconditioned cartridges a nd, after a washing step, salbutamol enantiomers are eluted with a mixture of chloroform and 2-propanol (80:20, v/v) containing 2% ammonia. Atenolol i s used as external standard. Enantioselective separation is accomplished wi th a Chirex(TM) 3022 stationary phase (urea type silica-bonded chiral phase ) using a mobile phase containing hexane-dichloromethane-methanol-trifluoro acetic acid (250:218:31:1, v/v) and fluorimetric detection with excitation and emission wavelengths set at 230 and 309 nm, respectively. The method pr oposed is rapid, selective and sensitive, and seems to be useful to differe ntiate between an authorized and a prohibited use of the drug in doping con trol. (C) 1999 Elsevier Science B.V. All rights reserved.