Application of a sequential analytical procedure for the detection of the beta-agonist brombuterol in bovine urine samples

The multistep analytical procedure routinely applied in our laboratory for the detection of the aryl amine beta-agonists clenbuterol, mabuterol and mapenterol in bovine matrices has been extended to the analysis in urine samples of brombuterol, a new clenbuterol-like compound. In the screening steps, the urine samples were first enzyme-linked immunosorbent assay (ELISA)-tested, then the positive samples were analyzed by thin-layer chromatography (TLC) and the beta-agonists detected with the Bratton-Marshall color reaction. The TLC spots corresponding to the suspected compounds were scraped off the plates, collected and extracted separately with methanol. The beta-agonists in the extracts were detected by HPLC-Vis (limits of detection: 0.5 ng/g). In the confirmatory step the presence and the concentration of the compounds of interest in the samples were established by GC-MS with two different ionization techniques, EI and CI (limits of detection: 1.0 ng/g). The use of this procedure has made possible the detection of brombuterol in officially sampled bovine urine.