中药复方制剂八宝丹中化学成分的UHPLC-Q-TOF/MS分析

UHPLC-Q-TOF-MS in analyzing chemical constituents of traditional Chinese herbal preparation Babao Dan

目的 采用超高效液相色谱-电喷雾串联四级杆飞行时间质谱（UHPLC-Q-TOF/MS）分析技术对中药复方制剂八宝丹中的化学成分进行快速分析鉴别。方法 采用安捷伦“Formula-Database Generator”软件自建包括547个化合物的八宝丹化学成分数据库,运用UHPLC-Q-TOF/MS技术采集八宝丹提取物的总离子流色谱图,根据各化学成分的精确质荷比,通过自动比对的方法进行分析鉴别;进一步调节四级杆飞行时间质谱（Q-TOF/MS）的二级碎片电压,根据二级裂解碎片离子对八宝丹中的化学成分进一步确证。色谱分离采用Waters XBridge BEH C18（2.1mm×100mm,2.5μm）液相色谱柱;流动相为乙腈（A）和0.5%甲酸水（B）,梯度洗脱：0-5min,5%A;5-25min,5%-95%A;25-30min,95%A。柱温25℃,流速0.8mL/min,柱后分流比为2∶1。质谱分析采用四级杆串联高分辨飞行时间质谱,电喷雾离子源正、负离子模式下扫描（范围均为m/z100-1 500）,其毛细管电压分别为3 500V和4 000V,二级质谱碰撞能量为10-40eV。结果 在正、负离子模式下,共鉴别出八宝丹中的化学成分78个,包括皂苷、胆汁酸、氨基酸等。结论 建立了基于UHPLC-Q-TOF/MS的分析方法,可对中药复方制剂八宝丹中的化学成分快速鉴别,阐明了八宝丹的化学物质基础,为八宝丹的质量控制及药理机制研究奠定了基础。

Objective To rapidly identify the chemical components in a traditional Chinese compound herbal preparation Babao Dan by ultra-high performance liquid chromatography-quadrupole-time of flight mass spectrometry（UHPLC-Q-TOF/MS）. Methods A formula database including 547 compounds was developed by Agilent software ＂Formula-Database Generator＂.The total ion chromatograms of the Babao Dan extracts were obtained by UHPLC-Q-TOF/MS,and the chemical components were identified by automatic matching method according to the exact mass-to-charge ratio of each chemical component.The chemical components of Babao Dan was further verified using the secondary fragment ion obtained by adjusting the secondary monitor voltage of Q-TOF/MS.The separation was performed on a Waters Xbridge BEH C18column（2.1mm ×100mm,2.5μm）.The mobile phase composing of acetonitrile（A）and 0.5% formic acid aqueous solution（B）was used for gradient eluting（0-5min,5% A;5-25 min,5%-95% A;25-30 min,95% A）.The temperature of column was 25℃;the flowing rate was 0.8mL/min;and the post-column split ratio was 2∶1.The Q-TOF/MS and electro spray ion source（ESI）were applied for qualitative analysis under positive and negative ion mode,with the mass scan range being m/z100-1 500.The capillary voltage was 3 500 Vunder positive ion mode and 4 000 Vunder negative ion mode,and the range of collision energy ofMS2 was 10-40 eV.Results UHPLC-Q-TOF-MS identified a total of 78 chemical components from Babao Dan,including saponins,bile acids,amino acids and so on.Conclusion A rapid and efficient method for identifying the chemical components of Babao Dan by UHPLC-Q-TOF-MS has been established,and the chemical components of Babao Dan has been clarified,which lays a foundation for the quality control and further pharmacological study of Babao Dan.