SIMULTANEOUS IDENTIFICATION AND DETERMINATION OF RESIDUAL PENICILLINSBY USE OF HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH SPECTROPHOTOMETRIC OR FLUOROMETRIC DETECTORS

Using 7 penicillins (amoxicillin, ampicillin, methicillin, penicillin G, oxacillin, cloxacillin, and dicloxacillin), simultaneous and direct determination of residual penicillins in biological samples was carri ed out by use of bioassay and high-performance liquid chromatography w ith spectrophotometric or fluorometric detectors. By use of assay medi um seeded with penicillin-sensitive Micrococcus luteus (ATCC No. 9341) as a test organism, we were able to detect penicillins even at low co ncentrations. All penicillins treated with 10 U of penicillinase/ml di d not produce inhibition zones by disk testing, even at a concentratio n of 100 mu g of penicillin/ml/assay plate. Using a mobile phase of ac etonitrile:methanol:0.01M KH2PO4 (19:11:70, v/v/v; pH, 7.1), standard solutions of the penicillins were separated from each other by use of high-performance liquid chromatography analysis, producing symmetric p eaks without tailing, each of which had a characteristic retention tim e. Simultaneous detection of residual penicillins in bovine serum kidn eys, and liver, for the 5 penicillins for which analysis was possible by use of the UV method, yielded recovery rates from 71.4 to 102.3%; f or the 2 amino-penicillins, amoxicillin and ampicillin, which could on ly be detected by use of the fluorometric method, recovery rate ranged from 72.9 to 103%.