Determination of gentamicin in urine samples after inhalation by reversed-phase high-performance liquid chromatography using pre-column derivatisation with o-phthalaldehyde

Gentamicin and netilmicin (internal standard) were extracted from urine using C, solid-phase extraction cartridges (94.3% recovery) and then derivatised with o-phthalaldehyde and 3-mercaptopropionic acid. The derivative was stable for >6 h. The mobile phase, methanol-glacial acetic acid-water (800:20:180, v/v), contained 0.02 M sodium heptanesulfonic acid, pH 3.4, and was passed at 1.0 ml min(-1) through a C, column with fluorescence detection (excitation 340 nm, emission 418 nm). The four main components of gentamicin (C-1, C-1a, C-2, C-2a) and netilmicin, the internal standard, were separated. Using the C, gentamicin peak, linearity was demonstrated from 0.5 to 10 mug ml(-1) and the limit of detection was 75 mug l(-1). Following 80-mg oral, 40-mg intravenous and 80-mg nebulised administration, the mean (SD) gentamicin urinary excretion was zero, 38.27 (0.96) and 1.93 (0.28) mg, respectively. Despite the relatively low lung deposition following inhalation of gentamicin the assay developed can be used to quantify the low urinary concentrations. Using this assay it should be possible to carry out urinary pharmacokinetic studies to identify the relative lung deposition of gentamicin following different methods of inhalation. (C) 2002 Elsevier Science B.V. All rights reserved.