Liquid chromatographic determination of gentamicin in serum with spectrophotometric detection

A simple, accurate, and specific liquid chromatographic procedure is described for the determination of gentamicin in 50 muL of serum. Gentamicin and sisomicin (IS) were converted into their trinitrophenyl derivatives by reaction with a water soluble derivatizing agent at 70 degrees C for 30 minutes. The derivatives were extracted from the crude mixture with chloroform, which was separated, evaporated, and the residue reconstituted in acetonitrile. Gentamicin was separated into three major isomers (C1, C2, and C1a) on a reversed-phase octyl column by eluting with a mobile phase. The eluted compounds were detected at 340 nm and quantified from their peak heights or peak areas. Chromatography was complete in less than 11.0 minutes. The lower limit of detection for gentamicin was less than 0.5 mg/L, analytical recoveries varied from 96.6% to 99.3%, linearity extended to 15.0 mg/L, and day-to-day precision was between 2.2% to 2.9%.