The analysis of dihydrostreptomycin sulfate using a column packed with base deactivated reversed phase silica gel and ultraviolet detection at 205 nm is described. The mobile phase consists of an aqueous solution containing 4 g/l of sodium sulfate, 1.5 g/l of sodium octanesulfonate, 100 ml/l of acetonitrile and 50 ml/l of a 0.2-M phosphate buffer at pH 3.0. The method allows separation of streptidine, dihydrostreptomycin B, streptomycin, dihydrostreptomycin and deoxydihydrostreptomycin, as well as some other components which were not identified. The total time of analysis is 55 min. The effects of the different chromatographic parameters on the separation were also investigated. A number of commercial samples were analyzed using this method.