基于LC-MS分析4,5-二咖啡酰奎宁酸在大鼠血浆和尿液中的代谢产物Analysis of metabolites of 4,5-dicaffeoylquinic acid in rat plasma and urine based on LC-MS
詹美榕;贾益群;陈龙;王夏雷;王雨;
ZHAN Mei-rong;JIA Yi-qun;CHEN Long;WANG Xia-lei;WANG Yu;Experiment Center of Science and Technology, Shanghai University of Traditional Chinese Medicine;
摘要(Abstract):
应用超高效液相色谱串联高场轨道阱质谱(UPLC-Orbitrap Elite-MS/MS)分析大鼠灌胃给予4,5-二咖啡酰奎宁酸后,血浆和尿液中的代谢物质组。采用Thermo C_(18)色谱柱(2.1 mm×100 mm, 1.9μm),以0.1%甲酸水-乙腈为流动相进行梯度洗脱。在负离子模式下采集生物样品的质谱数据,通过比对空白与给药样品,利用Compound Discovery 2.1软件对数据进行提取处理,比对化合物的精确相对分子质量以及质谱碎裂信息,计算二级碎片拟合比,进而归属代谢产物。结果在大鼠血浆中检测到15种代谢产物,在尿液中检测到16种代谢产物。涉及到的代谢反应有甲基化反应、水合反应、去水合反应、还原反应、葡萄糖醛酸化反应、硫酸酯化反应。血和尿中的代谢产物互为补充,初步揭示了此化合物在体内的移行形式和排泄形式。该研究建立起生物样本前处理、高分辨液质联用仪器采集数据、定性软件鉴定代谢产物结构的方法。为之后中药有效成分、体内药效形式的研究打下了基础。
Ultra high performance liquid chromatography tandem high field orbital trap mass spectrometry(UPLC-Orbitrap Elite-MS/MS) method was applied in this paper to analyze the metabolites of 4,5-dicaffeoylquinic acid in rat plasma and urine after oral administration. A gradient elution was performed by using Thermo C_(18) column(2.1 mm×100 mm, 1.9 μm), with 0.1% formic acid solution-acetonitrile as the mobile phase. Mass spectral data of biological samples were collected in negative ion mode. The data were extracted by Compound Discovery 2.1 software. Then the blank group samples and the drug samples were compared for exact molecular weight and the mass fragmentation information, and the secondary fragment fitting ratio was calculated to finally attribute the metabolites. As a result, 15 metabolites were detected in rat plasma, and 16 metabolites were detected in urine. The involving metabolic reactions included methylation, hydration, dehydration, reduction, glucuronide conjugation, and sulfation reaction. The metabolites in plasma and urine complemented each other and initially revealed the migration and excretion patterns of this compound in the body. A method for pre-processing biological samples, high-resolution LC-MS instrumentation data, and qualitative software was established in this study to identify metabolite structures, laying the foundation for the study of the active ingredients and in vivo pharmacodynamics forms of Chinese medicines.
关键词(KeyWords):
4,5-二咖啡酰奎宁酸;UPLC-Orbitrap Elite-MS/MS;代谢产物
4,5-dicaffeoylquinic acid;UPLC-Orbitrap Elite-MS/MS;metabolites
基金项目(Foundation): 上海中医药大学青年教师培养计划项目(A1-U1820501040282)
作者(Authors):
詹美榕;贾益群;陈龙;王夏雷;王雨;
ZHAN Mei-rong;JIA Yi-qun;CHEN Long;WANG Xia-lei;WANG Yu;Experiment Center of Science and Technology, Shanghai University of Traditional Chinese Medicine;
DOI: 10.19540/j.cnki.cjcmm.20190516.501
参考文献(References):
- [1] 朱乃亮,彭平,赵丽敏,等.植物中常见咖啡酰奎宁酸类化合物研究进展[C].北京:中华中医药学会中药化学分会学术年会,2013.
- [2] 柯细彤.灯盏细辛中咖啡酰奎宁酸类成分代谢产物及其抗缺血性卒中活性研究[D].昆明:昆明理工大学,2017.
- [3] 倪付勇,刘露,宋亚玲,等.金银花中抗补体活性酚酸类成分的研究[J].中国中药杂志,2015,40(2):269.
- [4] Nguyen M T,Awale S,Tezuka Y,et al.Xanthine oxidase inhibitory activity of vietnamese medicinal plants[J].Biol Pharm Bull,2004,27(9):1414.
- [5] dos Santos M D,Chen G,Almeida M C,et al.Effects of caffeoylquinic acid derivatives and C-flavonoid from Lychnophora ericoides on in vitro inflammatory mediator production[J].Nat Prod Commun,2010,5(5):733.
- [6] Nam S Y,Han N R,Rah S Y,et al.Anti-inflammatory effects of Artemisia scoparia and its active constituent,3,5-dicaffeoyl-epi-quinic acid against activated mast cells[J].Immunopharm Immunot,2017,40(1):1.
- [7] Fiamegos Y C,Kastritis P L,Exarchou V,et al.Antimicrobial and efflux pump inhibitory activity of caffeoylquinic acids from artemisia absinthiumagainst gram-positive pathogenic bacteria[J].PLoS ONE,2011,6(4):e18127.
- [8] Soh Y,Kim J A,Sohn N W,et al.Protective effects of quinic acid derivatives on tetrahydropapaveroline-induced cell death in C6 glioma cells[J].Biol Pharm Bull,2003,26(6):803.
- [9] Lee H J,Lee J Y,Kim S M,et al.Inhibitory effects of dicaffeoylquinic acids from Artemisia dubia on aldo-keto reductase family 1b10[J].J Korean Soc Appl Bi,2010,53(6):826.
- [10] 李亮,陈笑艳,钟大放,等.液相色谱-质谱联用技术在药物代谢产物鉴定中的应用[J].质谱学报,2017,38(4):375.
- [11] 王慧英,王锐,杨华,等.基于质谱技术的药物代谢产物鉴定策略进展[J].药学进展,2018,42(3):193.
- [12] 温菁,方平飞.液-质联用技术分析体内药物代谢物的研究进展[J].中国药学杂志,2017,52(21):1879.
- [13] 张晶,温泉,徐旭,等.基于UHPLC-QTOF-MS/MS技术的铁筷子中3个香豆素在大鼠体内代谢研究[J].中国中药杂志,2018,43(21):4330.
- [14] 王喆.复杂体系中多目标化合物定性和定量分析技术和策略研究[D].北京:北京协和医学院,2016.
- [15] 冯健男,杜守颖,白洁,等.生物样品前处理的研究进展[J].中国中药杂志,2014,39(21):4143.
- [16] Weber G F.Drug metabolism[M].Berlin:Springer,2015.
- [17] Venkatakrishnan K L,Von M,Greenblatt D.Human drug metabolism and the cytochromes P450:application and relevance of in vitro models[J].J Clin Pharmacol,2013,41(11):1149.
- 4,5-二咖啡酰奎宁酸
- UPLC-Orbitrap Elite-MS/MS
- 代谢产物
4,5-dicaffeoylquinic acid - UPLC-Orbitrap Elite-MS/MS
- metabolites
- 詹美榕
- 贾益群
- 陈龙
- 王夏雷
- 王雨
ZHAN Mei-rong- JIA Yi-qun
- CHEN Long
- WANG Xia-lei
- WANG Yu
- Experiment Center of Science and Technology
- Shanghai University of Traditional Chinese Medicine
- 詹美榕
- 贾益群
- 陈龙
- 王夏雷
- 王雨
ZHAN Mei-rong- JIA Yi-qun
- CHEN Long
- WANG Xia-lei
- WANG Yu
- Experiment Center of Science and Technology
- Shanghai University of Traditional Chinese Medicine