The analysis of gentamicin by liquid chromatography using a column packed with poly(styrene-divinylbenzene) and pulsed electrochemical detection on a gold electrode is described. The mobile phase consists of an aqueous solution containing sodium sulfate, tetrahydrofuran, sodium 1-octanesulfonate and a phosphate buffer of pH 3.0. In contradistinction to methods previously published, this method not only allows a better separation of gentamicins C1, C1a, C2, C2a and C2b, but also the separation of several other, minor components, most of which were not identified. The effects of the different chromatographic parameters on the separation were also investigated. A number of commercial samples was analysed using this method, allowing sensitive detection of gentamicin without derivatization, and the results were compared with the results obtained with the European Pharmacopoeia method, prescribing pre-column derivatization.