基于UPLC-Q-Exactive Orbitrap MS和质量亏损过滤技术的三叶青黄酮类化合物定性分析
Qualitative Analysis of Flavonoids in Tetrastigma Hemsleyanum Based on UPLC-Quadrupole/Exactive Orbitrap Mass Spectrometry and Mass Defect Filter Method
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摘要: 采用超高效液相色谱-四极杆/静电场轨道阱高分辨质谱（ultra-high performance liquid chromatography-quadrupole/exactive Orbitrap mass spectrometry，UPLC-Q-Exactive Orbitrap MS）和质量亏损过滤技术（mass defect filter，MDF）对三叶青黄酮类化合物进行定性分析。通过查阅文献，确定三叶青黄酮类物质母核作为MDF模板，根据取代基的数目和种类确定MDF的质量范围、质量亏损范围，采用UPLC-Q-Exactive Orbitrap MS采集—MDF-Orbitrap Traditional Chinese Medicine Library（OTCML）数据库检索—Mass Frontier验证的鉴定策略，对三叶青乙醇冷浸液所含的黄酮类物质进行定性分析，共鉴定出41种黄酮类化合物，其中19个为首次发现。该方法对于准确识别三叶青黄酮类化合物，进而科学评价三叶青质量、促进三叶青合理开发具有一定的指导意义。Abstract: A comprehensive qualitative analysis of flavonoids in the Tetrastigma hemsleyanum was developed by ultra-high performance liquid chromatography-quadrupole/exactive Orbitrap mass spectrometry (UPLC-Q-Exactive Orbitrap MS) and mass defect filter technique (MDF). UPLC-Q-Exactive Orbitrap MS is one of the powerful tools for analyzing complex systems of Traditional Chinese Medicine (TCM). The method has two main advantages, one is the higher resolving power relative to triplestage quadrupole mass spectrometers, the other is significantly reducing analysis time. MDF is a new data processing technique for high-resolution mass spectrometry data, which enables a majority of the interfering ions to be automatically filtered, allowing the original data to be analyzed more effectively, accurately, and rapidly. UPLC-Q-Exactive Orbitrap MS was employed to generate high-resolution mass spectrometry data combined with mass data processing strategy, which could be used for rapid discovery and structural identification of complex compounds of TCM. After the identification of the compounds, verification of the structure is also critical. Mass Frontier is software for managing, evaluating and interpreting mass spectrometry and chromatographic data. With its advanced algorithms and rich inventory data, it is very simple to analyze the mass spectrum and verify the structure of compound. According to the literature of the chemical constituents of Tetrastigma hemsleyanum, the nucleus of flavonoids in the Tetrastigma hemsleyanum was determined as a template for MDF, and the mass range and mass loss range of MDF were obtained on the basis of the number and type of substituents. Subsequently, the UPLC-Q-Exactive Orbitrap MS acquisition-MDF-Orbitrap Traditional Chinese Medicine Library (OTCML) database search-Mass Frontier verification identification process was employed to conduct a comprehensive qualitative analysis of the flavonoids contained in the ethanol extract of Tetrastigma hemsleyanum. 22 flavonoids were identified by applying the UPLC-Q-Exactive Orbitrap MS-OTCML-Mass Frontier validation strategy, while 41 flavonoids were identified by applying UPLC-Q-Exactive Orbitrap MSMDF-OTCML-Mass Frontier validation strategy. The results showed that this combination technology could effectively eliminate more interfering ions and excavate more potential flavonoids. This research enriches the basic research of chemical substances and provides significant guidance for evaluating the quality of Tetrastigma hemsleyanum, which will be beneficial to the rational development of Tetrastigma hemsleyanum. Above all, the MDF screening method combined with UPLC-Q-Exactive Orbitrap MS qualitative analysis demonstrates its feasibility and unique advantages on identification of the chemical constituent of TCM.
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[1] 侯艳婷，王晓明，张家伟，潘桂湘. 高分辨质谱数据MDF技术的进展[J]. 天津中医药，2016，33(12)：765-768.HOU Yanting, WANG Xiaoming, ZHANG Jiawei, PAN Guixiang. Progress in MDF technology of high resolution mass spectrometry data[J]. Tianjin Journal of Traditional Chinese Medicine, 2016, 33(12)：765-768(in Chinese). [2] 张丹，王彩虹，金滢，王喆，张金兰. 高效液相色谱-高分辨质谱法研究药物代谢产物的数据处理策略[J]. 质谱学报，2017，38(4)：410-416.ZHANG Dan, WANG Caihong, JIN Ying, WANG Zhe, ZHANG Jinlan. Study on data processing strategy of drug metabolites by high performance liquid chromatography-high resolution mass spectrometry[J]. Journal of Chinese Mass Spectrometry Society, 2017, 38(4): 410-416(in Chinese). [3] YANG L, SUN H, SUN W J, ZHAO L, MENG X C, WANG X J. Rapid and global detection and characterization of aconitum alkaloids in Yin Chen Si Ni Tang,a traditional Chinese medical formula,by ultra performance liquid chromatography-high resolution mass spectrometry and automated data analysis[J]. Journal of Pharmaceutical and Biomedical Analysis, 2010, 53(3): 421-431. [4] XIE T, LIANG Y, HAO H P, JIYE A, XIE L, GONG P, DAI C, LIU L S, KANG A, ZHENG X, WANG G J. Rapid identification of ophiopogonins and ophiopogonones in Ophiopogon japonicus extract with a practical technique of mass defect filtering based on high resolution mass spectrometry[J]. Journal of Chromatography A, 2012, 1227: 234-244. [5] FU L L, DING H, HAN L F, LI J, YANG W Z, ZHANG C X, HU Y, ZUO T T, GAO X M, GUO D A. Simultaneously targeted and untargeted multicomponent characterization of Erzhi Pill by offline two-dimensional liquid chromatography/quadrupole-Orbitrap mass spectrometry[J]. Chromatogr A, 2019, 1 584: 87-96. [6] ZHANG J Y, WANG F, ZHANG H, LU J Q, QIAO Y J. Rapid identification of polymethoxylated flavonoids in traditional chinese medicines with a practical strategy of stepwise mass defect filtering coupled to diagnostic product ions analysis based on a hybrid LTQ-Orbitrap mass spectrometer[J]. Phytochemical Analysis, 2014, 25 (5): 405-414. [7] 潘惠勤. 基于液相-高分辨质谱的多基原钩藤生物碱系统表征研究[D]. 上海：中国科学院大学(中国科学院上海药物研究所),2018. [8] 王放. 基于LC-HR-MSn的苦碟子及血栓通注剂化学成分研究[D]. 北京：北京中医药大学,2014. [9] LI H, LI Y, ZHANG Y, SHI H, HU W, ZHANG Z. Comparison of refluxing, ultrasonic-and microwave-assisted extraction of of dihydromyricetin from Ampelopsis grossedentata[J]. J AOAC Int, 2008, 91(6): 1278-1283. [10] ZHANG Y S, ZHANG Q Y, LI L Y, WANG B, ZHAO Y Y, GUO D A. Simultaneous determination and pharmacokinetic studies of dihydromyricetin and myricetin in rat plasma by HPLC-DAD after oral administration of Ampelopsis grossedentata decoction[J]. J Chromatogr B Analyt Technol Biomed Life Sci, 2007, 860 (1): 4-9. [11] 崔传文，孙翠玲，陈全成，邹秀红，黄学敏，陈海峰. 乌蔹莓化学成分的初步探究[J]. 中国中药杂志，2012，37(19)：2906-2909.CUI Chuanwen, SUN Cuiling, CHEN Quancheng, ZOU Xiuhong, HUANG Xuemin, CHEN Haifeng. A preliminary study on the chemical constituents of black berry[J]. China Journal of Chinese Materia Medica, 2012, 37(19): 2906-2909(in Chinese). [12] 史国茹. 岩五加生物活性化学成分研究[D]. 天津医科大学，2012. [13] 曾婷. 石猴子化学成分的研究[D]. 赣南师范学院，2013. [14] 冯正权，倪克锋，何煜，丁志山，朱峰，吴良村，沈敏鹤. 三叶青黄酮诱导SGC-7901胃癌细胞凋亡的实验研究[J]. 中国临床药理学与治疗学，2006(06)：669-672.FENG Zhengquan, NI Kefeng, HE Yu, DING Zhishan, ZHU Feng, WU Liangcun, SHEN Minhe. Experimental study on apoptosis of SGC-7901 gastric cancer cells induced by Clover Flavonoids[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2006(06): 669-672(in Chinese). [15] 倪克锋，金波，蒋福升，黄挺，冯正权，林胜友，吴良村，黄伶. 三叶青黄酮对H_(22)荷瘤小鼠TIMP-2mRNA表达的影响[J]. 中国中医药科技，2009，16(03)：195-196.NI Kefeng, JIN Bo, JIANG Fusheng, HUANG Ting, FENG Zhengquan, LIN Shengyou, WU Liangcun, HUANG Ling. Effects of flavonoids on the expression of TIMP-2 mRNA in H_(22) tumor-bearing mice[J]. Chinese Traditional Chinese Medicine Science and Technology, 2009, 16(03): 195-196(in Chinese). [16] 余纳，吴晓尉，汪芳裕. 三叶青总黄酮抗肿瘤作用的研究进展[J]. 中国中西医结合消化杂志，2015，23(09)：670-672.YU Na, WU Xiaoyu, WANG Fangyu. Research progress in anti-tumor effect of total flavonoids from Trifolium repens L[J]. Chinese Journal of Integrated Traditional and Western Medicine on Digestion, 2015, 23(09): 670-672(in Chinese). [17] 何煜舟，丁美萍，汪云开，徐缨，黄小民，张卓一，陈珺，寿旗扬. 山萘酚对缺血再灌注脑损伤大鼠的保护作用[J]. 中华中医药学刊，2009，27(08)：1673-1675.HE Yuzhou, DING Meiping, WANG Yunkai, XU Wei, HUANG Xiaomin, ZHANG Zhuoyi, CHEN Wei, SHOU Qiyang. Protective effects of Kaempferol on rats with cerebral ischemia-reperfusion injury[J]. Chinese Journal of Traditional Chinese Medicine and Pharmacy, 2009, 27(08): 1673-1675(in Chinese). [18] 刘昕，林琳，倪雅娟，魏瑾，张超英. 木犀草素可减轻异丙肾上腺素诱导的心力衰竭大鼠心肌细胞线粒体的损伤[J]. 中华老年多器官疾病杂志，2019，18(09)：693-698.LIU Yu, LIN Lin, NI Yajuan, WEI Wei, ZHANG Chaoying. Luteolin can alleviate mitochondrial damage in myocardial cells of rats with heart failure induced by isoproterenol[J]. Chinese Journal of Multiple Organ Diseases in the Elderly, 2019, 18(09): 693-698(in Chinese). [19] CHUAN L I, XI Y, CHENG C, CAI S X, HU J B. Isorhamnetin suppresses colon cancer cell growth through the PI3K-Akt-mTORpathway[J]. Mol Med Rep, 2014, 9(3): 935-940. [20] 宋慧婷，李长印，万瑶瑶，丁选胜，谭喜莹，戴国梁，刘史佳，居文政. 应用基于LC-TOF-MS分析的质量亏损过滤方法筛选黄芪注射液中的皂苷类成分[J]. 中国中药杂志，2017，42(04)：686-695.
SONG Huiting, LI Changyin, WAN Yaoyao, DING Xuansheng, TAN Xiying, DAI Guoliang, LIU Shijia, JU Wenzheng. Application of mass loss filtering method based on LC-TOF-MS analysis to screen saponins in Astragalus injection[J]. Chinese Journal of traditional Chinese Medicine, 2017, 42(04): 686-695(in Chinese).[21] 白海玉，刘宝忠，霍金海. 基于液质联用技术的野菊花中黄酮成分分析[J]. 中国中医药科技，2018，25(06)：826-830.BAI Haiyu, LIU Baozhong, HUO Jinhai. Analysis of flavonoids in wild chrysanthemum based on liquid chromatography-mass spectrometry[J]. Chinese Journal of Traditional Medical Science and Technology, 2018, 25(06): 826-830(in Chinese). [22] 郑成凤，潘裕添，蔡文燕，苑小宁，高飞. HPLC-UV-MS/MS法对金线莲中黄酮类组分的鉴定和测定[J]. 天然产物研究与开发，2013，25(10)：1381-1386.ZHENG Chengfeng, PAN Yutian, CAI Wenyan, YUAN Xiaoning, GAO Fei. Identification and determination of flavonoids in anoectochilus chinensis by HPLC-UV-MS/MS[J]. Natural Product Research and Development, 2013, 25(10): 1381-1386(in Chinese). -
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