OBJECTIVETo establish a high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) method for detection of nine penicillin residues in milk.

METHODSThe samples were precipitated with acetonitrile and degreased with n-hexane for pre-processing. The chromatography was performed by reversed-phase HPLC in a C₁₈ column and eluted gradiently on line for 11 min. The mobile phases were water with formic acid (pH 3.1) and acetonitrile-water with formic acid (pH 3.1). By optimizing the mass condition, the method was operated by multiple-reaction monitoring (MRM) and quantitated by external standard calibration.

RESULTSThe standard curves were linear in concentration range of 0.4 approximately 400 μg/L and all nine penicillins showed a good linear relationship in the milk matrix (r>0.990). The detection limit of the method was from 0.1 to 0.8 μg/L while the limit of quantification was from 0.3 to 2.6 μg/L. The average recoveries were higher than 80%. The within-day precisions were less than 8.5%.

CONCLUSIONThe established method is convenient, rapid and accurate and meets the requirement of monitoring penicillin residues in milk.