Submitted to: Journal of Agricultural and Food Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/22/2004
Publication Date: 12/7/2004
Citation: Schneider, M.J. 2004. A rapid flourescence screening assay for enrofloxacin and tetracyclines in chicken muscle. Journal of Agricultural and Food Chemistry. 52. pp. 7809-7813.
Interpretive Summary: The fluoroquinolones and tetracyclines are groups of antibiotics which are used in both medical and veterinary applications. Use of these antibiotics in animals produced for food has raised concerns as the presence of these residues in food may lead to increased microbial resistance in humans. Among fluoroquinolones, only enrofloxacin (ENRO) is currently approved for use in chickens in the U.S., with a tolerance level of 300 ng/g. Three tetracyclines, consisting of tetracycline (TC), oxytetracycline (OTC) and chlortetracycline (CTC), are approved as well, with a tolerance of 2000 ng/g. Given the large number of chickens produced in the U.S., it is important to have efficient methods for monitoring levels of these antibiotic residues in chicken. We have now developed a simple, rapid fluorescence screening assay for both ENRO and the tetracyclines in chicken muscle at their tolerance levels. This method requires only one extraction step, rather than one for each antibiotic, thus allowing both types of residues to be analyzed more efficiently. No overlap was observed between fluorescence of control samples and those fortified at the tolerance levels for ENRO and CTC. The utility of this method was illustrated by analysis of blind fortified samples. The efficiency of this analysis should facilitate its use by regulatory agencies in screening for ENRO and the tetracyclines in chicken muscle.
Technical Abstract: A simple, rapid fluorescence assay was developed for screening both enrofloxacin (ENRO) and tetracyclines in chicken muscle at the U.S. tolerance levels (300 ng/g and 2000 ng/g, respectively). Screening for both classes of antibiotics is accomplished using one extraction, thus simplifying and expediting the process. The method requires an initial extraction of chicken muscle with 1% acetic acid in acetonitrile, centrifugation, and analysis of the supernatant for ENRO fluorescence. After addition of ammonium hydroxide, magnesium chloride and methanol, followed by centrifugation and filtration, the supernatant can be measured for tetracycline fluorescence. Comparison of the fluorescence of control and tolerance level-fortified samples of both ENRO and chlortetracycline (CTC) shows no overlap. Setting a threshold as the average fortified fluorescence minus 3 sigma allows for successful screening, as illustrated with blind samples as controls or fortified with ENRO and/or CTC over a range of concentrations. This method can provide an alternative or supplemental approach to currently used microbial screening assays.