Quantitative multi-residue determination of beta-agonists in bovine urine using on-line immunoaffinity extraction-coupled column packed capillary liquid chromatography-tandem mass spectrometry.

This report demonstrates the potential of on-line immunoaffinity extraction and coupled column packed capillary liquid chromatography-ion spray tandem mass spectrometry for multi-residue determination of five beta-agonists, clenbuterol, mabuterol, mapenterol, methylclenbuterol, and tolubuterol, in bovine urine using an automated column switching system. Trace enrichment and preliminary sample cleanup was performed on-line using bovine urine diluted with phosphate-buffered saline. The column switching process involves trapping the target analytes onto a mini-bore immunoaffinity column, whereupon the target analytes are released from the immunoaffinity column onto a trapping column and subsequently eluted onto a packed capillary analytical column. The latter packed capillary column was used to provide the optimum sensitivity for ion spray LC-MS-MS analyses. The three-column system consists of a 2.0 mm I.D. immunoaffinity column, a 1 mm I.D. reversed-phase trapping column and a 320 microm I.D. packed capillary analytical column. Both qualitative and quantitative results are presented for the multi-residue determination of the target beta-agonists from the complex urinary matrix. Using tolubuterol as an internal standard, the quantitative data showed good linear response within the concentration ranges studied. Lower levels of quantitation were 50 part per trillion (ppt) for clenbuterol and methylclenbuterol, 20 ppt for mabuterol and 10 ppt for mapenterol. The bovine renal elimination is described using the technique for one of the beta-agonists, clenbuterol. The concentration of clenbuterol was detectable 15 days after the cessation of oral administration.