高效液相色谱-离子肼质谱串联技术在线鉴定氨氯地平原料药中未知杂质的结构

目的: 基于高效液相色谱-离子阱质谱串联技术(HPLC-MS/MS)对原料药氨氯地平中未知杂质进行在线结构鉴定。方法: 以甲酸铵水溶液-甲醇为流动相,经Waters X-Bridge C₁₈柱(150 mm ×4.6 mm,5 μm)分离,通过ESI离子阱质谱获得相关色谱与一级与二级质谱谱图。结果: 首先通过对氨氯地平主化合物及已知杂质的质谱裂解行为进行研究,提出了以二氢吡啶环为中心的retro-Diels-Alder(RDA)裂解机理。在此基础上,将未知杂质的质谱裂解谱图与已知化合物的相应谱图比较,基于质谱裂解的异同判断结构的异同,从而鉴定各未知杂质的结构。3个未知杂质分别为二甲酯式氨氯地平、甲酯异丙酯式氨氯地平、异丙酯乙酯式氨氯地平。结论: 本研究提出了一条基于HPLC-MS/MS在线鉴定药物中未知杂质的研究思路与策略,对其他药物中杂质的研究同样具有借鉴作用。

Objective: To perform online identification of unknown impurities in the drug substance of amlodipine based on high performance liquid chromatography-ion trap mass spectrometry(HPLC-MS/MS). Methods: For HPLC separation, the ammonium formate buffer-methanol was used as the mobile phase, and a Waters X-Bridge C₁₈ column(150 mm×4.6 mm, 5 μm) was adopted.The scan MS and the fragmentation MS experiments were performed based on trap ion mass spectrometer in the negative ESI mode. Results: The MS fragmentation behavior of the main compound and one known impurity was studied in details and an informative retro-Diels-Alder(RDA) fragmentation mechanism of the dihydropyridine ring was observed.Based on the former process, the mass spectrogram of fragmentation was compared with the corresponding spectrogram of the known compound to determine the difference in the structure based on the difference in spectrometric fragmentation, thus the structure of each unknown impurity was identified.The three unknown impurities were successfully identified as dimethylamlodipine, methylisopropylamlodipine, and ethylisopropylamlodipine, respectively. Conclusion: The strategy proposed based on the online identification of unknown impurities by HPLC-MS/MS in this study is valuable, and can be adopted in studies of other drug impurities.

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