Simultaneous determination of β-lactam antibiotics in milk by ion-pair liquid chromatography

Abstract A simple and rapid ion-pairing liquid chromatographic method was developed for the simultaneous determination of five penicillins (PCs), ampicillin (AB-PC), benzylpenicillin (PC-G), cloxacillin (MCI-PC), dicloxacillin (MDI-PC) and nafcillin (NF-PC) in milk. These PCs are most frequently used for the treatment of mastitis of cows. These antibiotics were extracted with acetonitrile from milk and cleaned up by solid-phase extraction with a C 18 cartridge. PCs were separated on a Kaseisorb LC ODS-300-5 column with a mobile phase (1 ml/min) of acetonitrile–methanol–0.05 M potassium dihydrogenphosphate (20:10:80, v/v/v) mixture containing 5 m M of sodium 1-decanesulfonate adjusted to pH 3.5 and UV detection at 210 nm. The average recoveries of five PCs from milk fortified at 0.5 and 1.0 μg/ml ( n =5) were 79.8–89.4% with relative standard deviations ranging from 2.7 to 7.2%. The detection limit of PCs in milk were 0.03–0.05 μg/ml.