RESUMO
A rapid, sensitive and selective HPLC method with post-column derivatization was proposed for the determination of salinomycin and related products in fermentation broths and premixes. The solvent extracts of samples were analysed on a reversed-phase monolithic type column. The mobile phase consisted of methanol/zinc acetate (0.05 M) adjusted to pH 4.0 with acetic acid (85/15, v/v). Post-column derivatization with vanillin at 85 degrees C was used for simultaneous, selective detection of salinomycin at 520 nm and related products at 460 nm. Optimal ratio of mobile phase/reagent flow rate was 2:1. Alternatively, pre-column derivatization of salinomycin and related products with three different reagents (2,4-dinitrophenylhydrazine, p-bromophenacyl bromide and p-nitrobenzoyl chloride) was examined. Suitable derivates for HPLC separation and UV detection were prepared using p-nitrobenzoyl chloride. Extraction ability of various solvents for extracting of salinomycin and co-products from premix samples was also tested. Acetone, ethanol and pyridine were found to be the best extraction solvents for these compounds.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Piranos/análise , Streptomyces/metabolismo , Ácido Acético/química , Acetona/análise , Benzaldeídos/química , Química Farmacêutica/métodos , Cromatografia , Indústria Farmacêutica/métodos , Etanol/análise , Fermentação , Concentração de Íons de Hidrogênio , Metanol/química , Modelos Químicos , Modelos Moleculares , Piranos/química , Piridinas/análise , Reprodutibilidade dos Testes , Espectrofotometria Ultravioleta/métodos , Temperatura , Raios Ultravioleta , Acetato de Zinco/químicaRESUMO
A simple and rapid high performance liquid chromatography method for the determination of salinomycin in fermentation media of Streptomyces albus strains and in premixes has been developed. This method involves reverse-phase separation of the component analysed with UV detection at 210 nm using methanol and 0.2 M acetate buffer pH 5.8 (100:10, v/v) as the mobile phase. The reliability of the method was confirmed by validation. A linear relationship was obtained within range 0.2-2.0 mg ml(-1) (r=0.9999). The relative standard deviation of methods within-laboratory reproducibility was 1.6%. The estimated quantitation limit of assay was about 32.5 microg ml(-1). The method has been successfully used in the determination of salinomycin content in testing production processes and premixes of different commercial brands.
