A CHEMICiZI, METHOD FOR THE DETERMINATION OF STREPTOMYCIN IN BLOOD ,4ND SPINAL FLUID*

The clinical application of streptomycin, as for any other potent drug, requires reliable information on blood levels of the drug. Microbiological assays will give estimates subject to wide variations unless a sufficient number are performed to permit statistical evaluation. The need for a reasonably simple and reproducible chemical assay for streptomycin in blood is apparent. We have previously described chemical methods for the assay of streptomycin in clinical preparations, nutrient broth, and urine (l), which were based on the formation of malt01 from st,reptomycin by treatment with alkali. These methods have satisfactory specificity and reproducibility, but the lower limit of sensitivity was 30 y of streptomycin. Since the concentration of streptomycin in the blood which is of clinical interest is in the order of magnitude of 1 to 30 y per ml., a more sensitive method is necessary. The early work on the structure of streptomycin indicated the presence of a reactive carbonyl group (2) and of guanido groups (3). On acid hydrolysis (4) streptomycin is cleaved into streptidine, containing the strongly basic guanido groups, and streptobiosamine, which contains the carbonyl group. The determination of streptomycin described in this paper is based on the presence of the reactive carbonyl group together with the strongly basic groups on 1 molecule. Since the presence of these two functional groups in the same molecule is quite rare in biological materials, the method is highly specific. In general, a substituted hydrazine can be used as a carbonyl reagent for the assay of streptomycin if the following conditions are fulfilled: The streptomycin hydraeone must be measurable by direct or indirect color formation, fluorescence, or ultraviolet absorption. The excess reagent together with the hydrazones of acidic, neutral, and weakly basic compounds must be separable from the strongly basic streptomycin hydrazone. Of particular importance in blood assays is the separation from the hydrazones of reducing sugars, amino sugars, keto acids, and from the possible cleavage products of streptomycin such as N-methyl-n-glucosamine (5) and streptobiosamine. The reagent should also be reasonably stable.