Characterization of 28 unknown impurities in 16-membered macrolides by liquid chromatography coupled with ion trap/time-of-flight mass spectrometry.

In terms of risk assessment, the study of the impurity profile is important to ensure the safety and effectiveness of drugs in clinical application. Sixteen-membered macrolides are produced by microbial fermentation, and many closely related substances in the product make the components and impurities complicated. In this study, methods were developed to separate and identify the impurities in three representative 16-membered macrolides (josamycin, midecamycin and meleumycin) using a high-performance liquid chromatography coupled to high-resolution ion trap/time-of-flight mass spectrometry (IT-TOF MS). In total, 53 impurities were characterized in the positive mode of electrospray ionization, among which 28 novel impurities were found. The proposed structures of impurities were deduced based on MS/MS data, and the ultraviolet (UV) absorption behaviors of impurities were discussed. In addition to the impurities with maximum absorption wavelengths (λmax) of 231 nm and 280 nm, there was a new group of impurities with λmax of 205 nm in meleumycin, midecamycin and josamycin.