Column-switching high-pressure liquid chromatographic method for the determination of a new carbapenem antibiotic, L-739,428, in rat and monkey plasma

Abstract

A column-switching HPLC method for determination of a new carbapenem antibiotic, L-739,428 (I), was developed that allowed direct injection of rat and monkey plasma. Following dilution with a pH 6.5 buffer, samples were injected without further cleanup into an extraction column dry-packed with 10-μm particle size Maxsil C 18 reversed-phase adsorbent. Endogenous plasma components were washed to waste for 6 min with a weak mobile phase of 0.025 M sodium phosphate, 0.005 M hexanesulfonate (pH 6.5). Compound I, retained on the extraction column, was then backflushed with a mobile phase which consisted of a mixture of the preceding buffer-ion pair solution and acetonitrile (96:4, v/v) into a Partisphere C 18 analytical column and detected by ultraviolet absorption at 299 nm. Chromatographic retention time was 11.5 min. Stability of I in plasma was maximized by use of a refrigerated autosampler which maintained plasma at 5°C until analyzed. The limit of quantification was 0.1 μg/ml using the equivalent of 75 μl plasma.