HPLC-DAD-ESI-MS/MS法用于枳实提取物的化学成分分析及多组分同时定量测定

摘要

目的:建立适用于枳实提取物的化学成分分析及多组分同时定量测定的准确、灵敏的分析方法,并用于该提取物的质量研究.方法:采用高效液相色谱-二极管阵列-电喷雾-串联质谱法(HPLC-DAD-ESI-MS/MS).色谱分离采用C18色谱柱,以乙腈-2％甲酸为流动相进行梯度洗脱.定性分析采用负离子全扫描和多级扫描模式,多组分同时定量测定采用DAD检测器于283 nm检测.结果:从枳实提取物中鉴定了18个化合物,包括12个黄酮,5个香豆素和1个柠檬苦素,并建立了同时定量测定其中12个主要化合物的定量分析方法.另外还研究了该12个化学成分在ESI-MS/MS中的裂解规律,为该提取物化学成分分析提供依据.该提取物的全面深入研究尚未见报道.结论:方法学验证表明所建立的方法快速、准确,可有效用于枳实提取物的化学成分与多组分同时定量测定.本研究工作将为枳实提取物及相关药物制剂的质量研究提供技术支持.%AIM:To develop a sensitive and accurate analytical method for the profiling,identification and simultaneous quantification of the chemical constituents in Aurantii Fructus Immaturus extracts.METHODS:A high performance liquid chromatography-diode array detection-electrospray ionization tandem mass spectrometry (HPLC-DAD-MS/MS) method was established for the characterization and quantification of multi-constituents in Aurantii Fructus Immaturus extracts.Chromatographic separation was achieved on a C18 column using a detailed gradient program with 2％ formic acid and acetonitrile as the mobile phase.Mass spectrometry was in the negative mode for identification,and a diode array detector was set at 283 nm for quantification.RESULTS:Eighteen compounds were identified,or tentatively identified,by comparing their retention times,UV and MS spectra with those of authentic compounds or literature data,including 12 flavonoids,5 coumarins and a limonoid in Aurantii Fructus Immaturus extracts.Twelve of the major compounds were determined simultaneously by HPLC-DAD.The fragmentation pathways of the authentic references were also investigated for the structural identification of the constituents in the extracts.There have been no reports on the simultaneous and comprehensive analysis of both the major and minor compounds in Aurantii Fructus Immaturus extracts.CONCLUSIONS:The method validation indicated that the developed analysis method could be employed as a rapid,effective technique for the structural characterization and simultaneous quantification of multi-constituents in this botanical extract.This work is expected to provide comprehensive information for the quality evaluation of Aurantii Fructus Immaturus extracts,which should be a valuable support for the further study and development of this raw material and its related medicinal products.