SEPARATION AND IDENTIFICATION OF N-DEMETHYLVANCOMYCIN

More than ten years have passed, since Chinese vancomyein was produced by pharmaceutical factories. Comparing with typical vancomycin, however, Chinese vancomycin showes some differences in antibacterial activity and chromatographic behaviour. Reverse phase high-performance liquid chromatography was employed for the analysis of vancomycin. The two samples of vancomycin (commercial preparation of vancomycin Eli Lilly, USA and domestic preparation) were resolved by means of an ODS small particle (10 μg)column and a mobile phrase of methanol-ammonium phosphate dibasic. According to the chromatogramexamined, domestic vaneomycin, which has been identified as N-demethylvancomycin, was different from vancomycin(Eli Lilly, USA). By using vitamin B₁₂ as an internal standard, both vancomycin and the commercial preparation (Eli Lilly, USA) were determined. The CV of the quantitative determination of domestic vancomycin was found to be 1.01%. A gradient column Chromatography equipped with CM-sephadex C-50 for separation and purification of the components of domestic vancomycin was developed. The fractionation patterns of components were plotted. The main component was isolated and recrystallized. A single component was obtained and the purity was found to be 99.05%. The retention time of the main component of domestic vancomycin was found to be different from that of typical vancomycin. Based on the data of mielear magnetic resonance (¹HNMR and ¹³CNMR), fast atom bombardment mass spectrum and its hydrolized products, it was confirmed that there is a leucine rather than N-methyl-leucine in the molecule of Chinese vancomycin, but all the other residues are the same as that of vancomycin. Consequently, Chinese vancomycin is actually N-demetlaylvancomycin.