DEVELOPMENT AND VALIDATION OF QUANTITATIVE DETERMINATION METHOD OF DIHYDROSTREPTOMYCIN SULPHATE AND PROCAINE BENZYLPENICILLIN IN INJECTION SUSPENSIONS

Abstract

The aim of the work was to develop and validate a method for the simultaneous identification and quantification of dihydrostreptomycin and benzylpenicillin in injectable suspensions. The method was validated by testing two preparations in the form of injectable suspensions containing benzylpenicillin 108–144 mg/ml and dihydrostreptomycin 180–220 mg/ml. Test samples were dissolved in purified water P, and standard samples: benzylpenicillin - in methanol (up to a concentration of 126 μg/ml), dihydrostreptomycin - in purified water P (up to a concentration of 200 μg/ml). The maximum allowable total uncertainty of the analysis was 1.64%, which is within the recommendations of SFU 2.0. The samples were separated on a Dionex Ultimate 3000 chromatograph equipped with a Luna C18 (2) 250 × 4.6 mm, 5 μm chromatographic column. The mobile phase was a mixture of acetonitrile and a solution of 0.01 M sodium heptanesulfonate with 0.05 M sodium phosphate trisubstituted, acidified with 0.1 M phosphoric acid to pH 6.0, in a volume ratio of 2: 8. Under mentioned conditions, dihydrostreptomycin and benzylpenicillin were completely separated. The established parameters of the chromatographic system did not exceed the limits specified in the FDA recommendations. The calibration curves were linear in the recommended SFU 2.0 range (80–120% of the nominal concentration of the corresponding active substance). The ratio of the amount of standard sample added to the test samples with its subsequent detection in the preparation was 99.35–100.79% for benzylpenicillin and 99.49–100.12% for dihydrostreptomycin, which does not exceed the limits recommended in SFU 2.0. The precision criterion was 0.07 for dihydrostreptomycin and -0.08 for benzylpenicillin, which is within the limits recommended in SFU 2.0. At the same time, the results of the study by different analysts at different times differed by 1.3% for dihydrostreptomycin and 0.98% for benzylpenicillin, which is well within the limits adopted in the recommendations of the FDA and SFU 2.0. Therefore, the method developed and validated by us for the simultaneous determination of dihydrostreptomycin sulfate and benzylpenicillin procaine in injectable suspensions can be considered suitable for routine analysis.