Determination of vancomycin related substances by gradient high-performance liquid chromatography.

A gradient high-performance liquid chromatographic method for the determination of vancomycin related substances is described. This method was developed to profile vancomycin, co-fermentation products, and degradation products. The resultant chromatograms confirm the multifactored nature of vancomycin, separating a number of peaks from the main component. The development of acceptable chromatographic performance is described, with the final method intended for use as a control assay. Quantitation of total related substances is made by comparing the relative area of the main peak to total peak area in a pair of chromatograms from a stock solution and a twenty-five fold dilution. A rapid-scan UV detector was used to demonstrate the similar spectral characteristics of the vancomycin related substances, confirming the validity of the relative area approach. Statistical method validation data are included, evaluating the use of this method for quantitative applications. Example applications demonstrate the effectiveness of this method.