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

The multistep analytical procedure routinely applied in our laboratory for the detection of the aryl amine β-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 β-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 β-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.