Analysis of neomycin sulfate and framycetin sulfate by high-performance liquid chromatography using evaporative light scattering detection

A rapid and simple method for the determination of main components and related substances of both neomycin sulfate and framycetin sulfate by HPLC and evaporative light scattering detection (ELSD) is described. The method was also used to determine the neomycin B and the sample sulfate content. Detection and quantitation of aminoglycoside antibiotics are problematic because of the lack of UV absorbing chromophore. The use of a universal detector avoids the need for sample derivatization or use of specific detector based on pulsed amperometry described to be difficult in routine assays. Separation was performed with a Polaris C18 150 mm × 4.6 mm i.d., 3 μm reversed-phase column with a solution of 170 mM trifluoroacetic acid (TFA) mobile phase at a flow rate of 0.2 mL/min. The chromatographic parameters were optimized with the help of experimental design software. Mass spectrometry (MS) was employed to confirm the ELSD profile. The final method was validated using methodology described by the International Conference of Harmonization in the field of Active Pharmaceutical Ingredients. Commercial samples of different sources were analyzed and results were in good agreement with specifications of the European Pharmacopoeia.