Validation of a microbiological method: the STAR protocol, a five-plate test, for the screening of antibiotic residues in milk

Abstract

The results of an in-house laboratory validation of a microbiological method for the screening of antibiotic residues in milk are presented. The sensitivity of this five-plate test, called Screening Test for Antibiotic Residues (STAR), was established by the analysis of milk samples spiked with 66 antibiotics at eight different concentrations. Ten different groups of antibiotics were studied: macrolides, aminoglycosides, cephalosporins, penicillins, quinolones, tetracyclines, sulphonamides, lincosamides, phenicolated and miscellaneous drugs. It was shown that 21 antibiotics were detected by the STAR protocol at or below the maximum residue limit (MRL), and that a further 27 drugs could be detected at levels from the MRL up to four times the MRL. The sensitivity of the STAR protocol was at or below the MRL for three macrolides, one tetracycline, two aminoglycosides, some sulphonamides, half of the beta-lactams, quinolones, lincosamides, trimethoprim and baquiloprim. Moreover, the STAR protocol was at least twice as sensitive as conventional methods for macrolides, quinolones and tetracyclines. The other antibiotics had limits of detection between four and 150 times the MRL. Each plate was preferentially sensitive for one or two families of antibacterials: the plate Bacillus cereus for tetracyclines, the plate Escherichia coli for quinolones, the plate Basillus subtilis for aminoglycosides, the plate Kocuria varians for macrolides, and the plate Bacillus stearothermophilus for sulphonamides and beta-lactams. This method has been used routinely on a day-to-day basis to direct the physicochemical confirmation towards one or two families of antibiotics. Considering the high cost of liquid chromatography coupled with tandem mass spectrometry detection analyses, the reduction of the range of antibiotics to test for confirmation is a significant gain in time and money.