Vancomycin is an effective antibiotic used for the treatment of Gram-positive bacterial infections. During the analysis of vancomycin, an unknown impurity at the level of 0.5% was detected by high-performance liquid chromatography (HPLC). To characterize the structure of the impurity, a new two-dimensional preparative liquid chromatography (2D-Prep-LC) method was developed to separate the impurity from the vancomycin sample. After further analysis including liquid chromatography-mass spectrometry (LC-MS) and nuclear magnetic resonance (NMR) spectroscopy, the structure of the unknown impurity was identified as a vancomycin analog in which the N-Methyl-leucine residue on the side chain is replaced by an N-methylmethionine residue. In this study, we established a reliable and efficient method for separating and identifying vancomycin impurities, which will provide a valuable contribution to the field of pharmaceutical analysis and quality control.

万古霉素是一种有效的抗生素，用于治疗革兰氏阳性细菌感染。在万古霉素分析过程中，高效液相色谱法 (HPLC) 检测到 0.5% 的未知杂质。为了表征杂质的结构，一种新的二维开发了制备型液相色谱 (2D-Prep-LC) 方法来分离万古霉素样品中的杂质。经过包括液相色谱-质谱 (LC-MS) 和核磁共振 (NMR) 光谱在内的进一步分析，未知杂质的结构被鉴定为万古霉素类似物，其中侧链上的 N-甲基-亮氨酸残基被取代由 N-甲基甲硫氨酸残基。在本研究中，我们建立了一种可靠、高效的分离和鉴定万古霉素杂质的方法，将为药物分析和质量控制领域提供有价值的贡献。