Impurity profile and spectrum characteristics of the isomers in cefamandole nafate using high‐ performance liquid chromatography/high‐performance size exclusion chromatography tandem ion trap/time‐of‐flight mass spectrometry.

Rationale: Reversed‐phase, high‐performance liquid chromatography (RP‐HPLC) and high‐performance size exclusion chromatography (HPSEC) methods were developed to effectively separate unknown impurities and polymerized impurities in cefamandole nafate. The liquid chromatography‐tandem ion trap/time‐of‐flight mass spectrometry (LC‐IT‐TOF‐MS) was applied to characterize the structures of the impurities. Ultraviolet (UV) spectrum characteristics and mass spectrum characteristics of △3‐isomer and 7‐epimer in cefamandole nafate were studied to distinguish the isomers. Methods: RPLC‐IT‐TOF‐MS was used to characterize the structures of unknown impurities and polymerized impurities eluted from the C18 column. On this basis, the two‐dimensional (2D) HPSEC‐IT‐TOF‐MS was used to confirm the structures of polymerized impurities eluted from the TSK‐gel G2000SWxl column. Complete fragmentation patterns of impurities were studied and used to obtain information about the structures of the impurities. Results: The structures of 19 unknown impurities in cefamandole nafate were elucidated based on the high‐resolution MSn data with both positive and negative modes, assisted by the UV spectra and stress testing, of which 2 impurities were polymerized impurities. Cefamandole nafate produced a series of degradation impurities, and another principal component cefamandole acid also produced a series of similar degradation impurities. The disciplines between mass fragmentation pattern/UV spectrum and structure for △3‐isomer and 7‐epimer were presented to distinguish their structures. Conclusions: The results of this study provided a scientific basis for the improvement of official monographs in pharmacopoeias to effectively control the impurities and ensure drug safety for the public. This study also revealed the formation mechanisms of degradation impurities in cefamandole nafate, which may guide industry to improve the manufacturing process and storage conditions to reduce the content of impurities in products. [ABSTRACT FROM AUTHOR]