This study describes the development of a new stable isotope labeling method by carbon-13 in bacteria culture (SILCB) for the analysis of residual antibiotics via liquid chromatography with tandem mass spectrometry (LC-MS/MS). Stable isotope dilution (SID) LC-MS/MS with QuEChERS was employed to determine avermectin, particularly avermectin B1a (AV-a) and B1b (AV-b), based on completely 13C-labeled internal standards (13C-ISs) obtained from the SILCB. Our SILCB was developed from an optimal inorganic medium using 13C6-glucose for Streptomyces avermitilis (14893 strain). A rough extract containing 13C-ISs was purified via high-speed countercurrent chromatography with a volatile two-phase solvent system composed of n-hexane/ethyl acetate/methanol/0.5% formic acid in water (7/3/5/5/, V/V). The purified 13C-ISs were evaluated to confirm the presence of completely 13C-labeled ions with m/z 938 > 326 and m/z 923 > 309 for AV-a and AV-b, respectively. The QuEChERS approach with the 13C-ISs procedure achieved acceptable recovery rates in beef meat samples of 99.5 - 100.0% (RSD < 2.0%, n = 6). For the analysis of residual antibiotics in foodstuffs by SID-LC-MS/MS and QuEChERS, the SILCB represents a significant improvement over previous methods suffering from cumbersome sample preparation and matrix effects.
Keywords: LC-MS/MS; Stable isotope labeling-culture; avermectin; residue assay.