基于UHPLC-LTQ/Orbitrap MS法的苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z的大鼠肝微粒体体外代谢产物鉴定

苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z是苦碟子(Ixeris sonchifolia(Bunge)Hance)中2种重要的倍半萜内酯类成分。为研究它们在大鼠肝微粒体中的代谢产物,采用Waters Acquity UPLC BEH C18色谱柱(2.1mm×50mm×1.7μm)分离,以0.1%甲酸水(A)-乙腈(B)为流动相进行梯度洗脱,采用高效液相色谱-线性离子阱静电轨道场质谱(UHPLC-LTQ/Orbitrap MS)技术,在ESI负离子模式下分析样品。筛选出18个化合物(包括苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z,及其16个代谢产物),并对其中15个代谢产物进行结构鉴定。结果表明,苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z经大鼠肝微粒体主要发生了羟基化、加氢和脱氢的Ⅰ相代谢反应。该结果初步阐明了苦荬菜内酯Z和11,13α-二氢苦荬菜内酯Z在大鼠肝微粒体中的代谢规律,可为新药、毒理学研究提供支持。

Profiling and Identification of the Metabolites of Ixerin Z and 11,13α-Dihydroixerin Z in Rat Liver Microsomes by UHPLC-LTQ/Orbitrap MS

Ixerin Z and 11,13α-dihydroixerin Z are the important ingredients of sesquiterpene lactones in Kudiezi. In this study, the metabolites of Ixerin Z and 11,13α-dihydroixerin Z in rat liver microsomes were screened and identified using high performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry (UHPLC-LTQ/Orbitrap MS) at negative ion mode. The separation was conducted on Waters Acquity UPLC BEH C18 column (2.1 mm×50 mm×1.7 μm) with the gradient elution of 0.1% formic acid aqueous solution (A) and acetonitrile (B). As a result, Ixerin Z, 11,13α-dihydroixerin Z and 16 metabolites were selected out, among which 15 metabolites were characterized based on the retention time, accurate mass measurements and fragmentation patterns. The results showed that there were two possible metabolic pathways of Ixerin Z in rat liver microsomes: 1) Metabolites 2, 5 and 12 were generated from Ixerin Z after hydroxylation; 11,13α-dihydroixerin Z was generated from Ixerin Z after hydrogenated on its double bond, then metabolites 8, 14 and 15 were generated from 11,13α-dihydroixerin Z after hydroxylation. 2) The metabolites were generated from the prototype via addition reaction, such as cysteine combination (1 and 4), glutathione combination (7 and 9). There were also two possible metabolic pathways of 11,13α-dihydroixerin Z in rat liver microsomes: 1) Ixerin Z was generated from 11,13α-dihydroixerin Z after hydrogenated on its double bond, then the metabolites 12 was generated from Ixerin Z after hydroxylation. Metabolites 3, 8, 10, 13, 14 and 15 were generated from 11,13α-dihydroixerin Z after hydroxylation, followed by hydrogenation to produce 6 and 11. 2) Metabolite 7 was generated in conjugation reaction. The results demonstrate that they mainly undergo oxidation, reduction, hydrolysis via phase I in rat liver microsomes, which will help to clarify the metabolic pattern of Ixerin Z and 11,13α-dihydroixerin Z, and can also provide support for further new drug and toxicology research.