Simultaneous determination of clenbuterol, chloramphenicol and diethylstilbestrol in bovine milk by isotope dilution ultraperformance liquid chromatography-tandem mass spectrometry.

A simple and sensitive analytical method was developed for the simultaneous determination of clenbuterol, chloramphenicol and diethylstilbestrol in bovine milk by isotope dilution ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Samples were directly purified through HLB cartridge. The organic phase was dried under nitrogen and residues were redissolved in mobile phase. Samples were analyzed by UPLC-MS/MS on an Acquity UPLC® BEH C(18) column with gradient elution. The samples were quantified using clenbuterol-D(9), chloramphenicol-D(5) and diethylstilbestrol-D(8) as internal standards. The proposed method was validated according to the European Union regulation 2002/657/EC determining specificity, decision limit (CCα), detection capability (CCβ), trueness, precision, linearity and stability. The method is demonstrated to be suitable for the determination of clenbuterol, chloramphenicol and diethylstilbestrol in bovine milk. The total time required for the analysis of one sample was about 50min.