High-Performance Liquid Chromatographic Determination of Penicillins following Derivatization to Mercury-Stabilized Penicillenic Acids

Abstract

Abstract A high-performance liquid chromatographic, pre-column derivatization procedure is described, allowing the determination of natural and semi-synthetic penicillins by absorption at 325nm. Derivatization with an imidazole/mercuric chloride reagent is followed by reverse-phase chromatography of the resulting mercury-stabilized penicillenic acids on an octade-cylsilane, chemically-bonded stationary phase. Gradient elution is employed with a mobile phase containing organic modifier, buffer and ethylenediaminetetraacetic acid. Precolumn reaction conditions including time and temperature are examined. Accuracy, reproducibility and detection limits of the method are discussed. The technique is applied to the analysis of fermentation media following sample preparation by centrifugation and deproteinization.