Submitted to: Journal of Chromatography A
Publication Type: Peer reviewed journal
Publication Acceptance Date: 8/31/2013
Publication Date: 9/8/2013
Publication URL: http://handle.nal.usda.gov/10113/57987
Citation: Lehotay, S.J., Mastovska, K., Lightfield, A.R., Nunez, A., Dutko, T., Ng, C., Bluhm, L. 2013. Rapid analysis of aminoglycoside antibiotics in bovine tissues using disposable pipette extraction and ultrahigh performance liquid chromatography - tandem mass spectrometry. Journal of Chromatography A. 1313:103-112. Interpretive Summary: Aminoglycoside antibiotics are commonly used as veterinary drugs in livestock production worldwide, and analytical methods are utilized in regulatory monitoring programs to ensure that excessive residue levels do not occur in the food animal tissues. The methods must achieve high quality results with high practical efficiency in the laboratory. A high-throughput qualitative screening and identification method for 9 aminoglycosides of regulatory interest has been developed, validated, and implemented for kidney, liver, and muscle tissues collected from slaughterhouses. The method was transferred to the USDA Food Safety and Inspection Service to replace their previous approach, and it has directly led to improved monitoring of veterinary drug residues in meat products in faster and less costly operations.
Technical Abstract: A high-throughput qualitative screening and identification method for 9 aminoglycosides of regulatory interest has been developed, validated, and implemented for bovine kidney, liver, and muscle tissues. The method involves extraction at previously validated conditions, cleanup using disposable pipette extraction, and analysis by a 3 min ultrahigh-performance liquid chromatography – tandem mass spectrometry (UHPLC-MS/MS) method. The drug analytes include neomycin, streptomycin, dihydrosptreptomycin, and spectinomycin, which have residue tolerances in bovine in the US, and kanamicin, gentamicin, apramycin, amikacin, and hygromycin, which do not have US tolerances established in bovine tissues. Tobramycin was used as an internal standard. An additional drug, paromomycin was also validated in the method, but it was dropped during implementation due to conversion of neomycin into paromomycin. Recoveries from spiking experiments at regulatory levels of concern showed that all analytes averaged 70-120% recoveries in all tissues, except hygromycin averaged 61% recovery. Lowest calibrated levels were as low as 0.005 ug/g in matrix extracts, which approximately corresponded to the limit of detection for screening purposes. Drug identifications at levels <0.05 ug/g were made in spiked and/or real samples for all analytes and tissues tested. Analyses of 60 samples from 20 slaughtered cattle previously screened positive for aminoglycosides showed that this method worked well in practice. The UHPLC-MS/MS method has several advantages compared to the previous microbial inhibition screening assay, especially for distinguishing individual drugs from a mixture and improving identification of gentamicin in tissue samples.