Detection of beta-blockers in human urine by GC-MS-MS-EI: perspectives for the antidoping control

We have developed a general method for the detection of beta-blockers and/or of their metabolites in human urine. The method comprises a pretreatment procedure (enzymatic hydrolysis, liquid/liquid extraction and derivatization by pentafluoropropionic anhydride, PFPA), carried out on an initial aliquot of 2.5-5.0 ml of urine, and the instrumental analysis of the derivatives, performed by GC-MS-MS (ion trap) with electronic impact ionization (EI). The GC-MS-MS analysis allows to isolate and to characterize specific fragments of the original molecular structure, and particularly the fragments originating from parent ion clusters specific for all beta blocking drugs, giving rise to m/z = 366 and 202 ions respectively. MS-MS analysis of the parent ion allows checking for the presence of the above-mentioned peaks in the GC-MS chromatogram. The proposed method is capable of detecting a great variety of known (and possibly also of newly synthesized) beta-blockers, with an average sensitivity limit of 20 ng/ml of drug/metabolite in urine. The method is presently being evaluated as a general screening protocol to be followed by an antidoping laboratory to detect illicit beta-blockers administration to the athletes.