Simultaneous determination and identification of penicillin residues by high-performance liquid chromatographic analysis

A new method was developed for simultaneous determination and identification of seven penicillins (amoxicillin, ampicillin, methicillin, penicillin G, oxacillin, cloxacillin, and dicloxacillin) in bovine serum. Each sample was simply extracted with acetonitrile, and identification of the reaction products of penicillins after treatment with 1,2,4-triazole-HgCl2 solution was then carried out by high-performance Liquid chromatography (HPLC) with on ultraviolet spectrophotometric detector at 325 nm. Separation of the penicillin reaction products by HPLC was carried out by using a mobile phase of 0.1 M phosphate buffer and acetonitrile in ratios of 85:15 and 60:40 (vol/vol) in a gradient flow. The retention times of the seven penicillins ranged from 4.3 to 24.6 min, and the detection limits of penicillin concentration ranged from 0.04 to 0.2 mu g/ml. The calibration curves for individual penicillins extracted from bovine serum were linear, and the correlation coefficients for each of the drugs were over 0.99. The determination of penicillins extracted from bovine serum spiked with each drug gave a recovery rate from 82.0 +/- 5.6% to 105.6 +/- 4.6%. This detection method may be useful for routine laboratory testing of residual penicillins.