High-performance liquid chromatographic analysis of aminoglycoside antibiotics

A simple precolumn derivatisation method for the determination of aminoglycoside antibiotics (AGs) is described. The stability of the o-phthalaldehyde (OPA) derivatives of the AGs obtain using beta-mercaptopropionic acid (beta-MP) was investigated by reversed-phase HPLC. One of the fluorescent derivatives of sisomicin was stable at least for 6 h in 50% methanol under the optimal conditions used (OPA concentration, pH and temperature). When plasma samples spiked with sisomicin were analysed the response was linear in the calibration range of 136-900 micrograms of sisomicin per injected volume (40 microliters). As little as 0.06 micrograms of sisomicin per 1 ml of plasma could be detected with a signal-to-noise ratio > or = 2. The method was also applied to whole blood samples from rabbit after a subcutaneous injection of 1 mg/kg of the AGs using dried blood spots (DBS) on the filter-paper punched discs. The detection of sisomicin and netilmicin in the DBSs on punched discs (10.1 micrograms of whole blood) were 0.053 and 0.50 micrograms per ml of whole blood, respectively (signal-to-noise ratio > or = 2). The method permits a simple collection of blood at the microlitre level and should prove particularly useful for monitoring the AGs in blood at therapeutic levels in geriatric and paediatric patients and could be also used for the preclinical study of the AGs blood levels of a number of mice or rats without killing. An RP-HPLC method using an on-line clean-up procedure for large sample-volume analysis of serum AGs is also described.