基于液质联用技术的中国苦苣苔科药用植物特征化合物类群的表征

【作者】 胡峻；

【导师】 刘勇；

【作者基本信息】 北京中医药大学 ， 中药化学， 2016， 硕士

【摘要】 研究目的苦苣苔科(Gesneriaceae)为双子叶植物纲菊亚纲的玄参目的一科,常分为苦苣苔亚科和大岩桐亚科,目前在全世界共有150属3700余种。我国共有58属约470余种,全部属于苦苣苔亚科,其中28属中国为中国所特有。该科许多种植物在贵州、广西、云南等省区少数民族地区广泛用于治疗痢疾、跌打损伤、毒蛇咬伤、关节炎等,疗效十分显著。然而该科植物多岩生或地生,植物的生存受环境影响较大,部分植物濒临灭绝;绝大部分我国特有属、种植物的化学成分研究尚处于空白,已经揭示本科所含有的化学成分包括黄酮类、苯乙醇苷类、萜类、醌类等,并且都具有很强的药理活性。为了对我国苦苣苔科药用植物的资源、药用植物的化学成分有一个明确的认识,充分及合理利用我国苦苣苔科药用植物资源,本论文进行了以下的研究:(1)对常见的苯乙醇苷类成分对照品进行液质联用分析,归纳总结各类苯乙醇苷类的裂解特征;(2)对采集的不同种属苦苣苔科样品进行化学成分提取、液质测定,建立苦苣苔科植物化学成分的测定方法,结合苯乙醇苷类成分质谱裂解规律,对苦苣苔科样品化学成分进行整体表征;(3)对吊石苣苔属植物和半蒴苣苔属植物整体化学成分进行主成分分析(PCA),OPLS-DA分析两者差异成分,来探讨两个属之间的化学亲缘关系。研究方法1.系统性地查阅相关文献,对苦苣苔科植物的分类学、化学成分、药理活性的国内外研究进行了全面的综述。2.运用超高效液相串联四级杆飞行时间质谱联用仪器(UPLC-Q-TOF-MS)建立了对8种苯乙醇苷类对照品建立合适的液质分析方法,使得所有成分具有良好的分离度,优化质谱条件使得各成分裂解碎片稳定,裂解信息全面。3.根据化合物液相色谱信息,一级、二级质谱信息,结合化合物数据库、DAD特征光谱,化合物ClogP值等信息,对24个种的苦苣苔科样品进行了整体成分表征及鉴定。4.运用主成分分析(PCA),OPLS-DA分析吊石苣苔属植物和半蒴苣苔属植物差异成分。结果与结论1.对8种苯乙醇苷类对照品的一二级质谱信息进行分析,归纳了不同类型苯乙醇苷类成分质谱裂解规律;对咖啡酰基在C-4’位和C-6’位取代的苯乙醇苷类进行了有效的区分;对含葡萄糖或鼠李糖的苯乙醇单糖苷类成分进行了区分,对含鼠李糖-葡萄糖、鼠李糖-芹糖的苯乙醇二糖苷进行了区分,对糖取代基在C-3’和C-6’位置的苯乙醇苷类进行了区分。2.对样品进行UPLC-Q-TOF-MS分析。根据对照品裂解规律、裂解碎片及相关资料推测可能的裂解途径,通过自带的数据库以及Chemspider、Scifinder得到可能的分子式及候选化合物。根据二级质谱碎片裂解途径,结合DAD特征光谱化合物ClogP值等信息,以及苦苣苔科已报道的相关研究进行成分鉴定。结果显示苦苣苔科植物中含有苯乙醇苷类、黄酮等成分,不同种属之间苯乙醇苷类成分类型及含量存在一定的差异。3.以吊石苣苔属植物与半蒴苣苔属主要生理活性组分为信息基础,利用主成分分析法(PCA)进行聚类分析,OPLS-DA模型进一步分析确定吊石苣苔属植物与半蒴苣苔属所含化合物指纹图谱的差异,同时对数据进行Pareto标度化处理,得到两个属潜在化学标记物(Marker)13个,主要为苯乙醇苷类成分。结果表明,吊石苣苔属植物化学成分与半蒴苣苔属植物化学成分存在一定的类似性,但区分两个属之间的次生代谢物为苯乙醇苷类成分,其含量差别也比较明显。更多还原

【Abstract】 Objective:Gesneriaceae,one of the important groups of Scrophulariales,can be divided into Cyrtandroideae and Gesnerioideae,with 150 genera and 3700 species altogether.There are 58 genera and 470 species,with 28 genera specialty in china,which belong to Cyrtandroideae.Plants of this family are widely used to treat dysentery,bruises,snake bites,arthritis in minority areas of Guizhou,Guangxi,Yunnan provinces.However,most of the plants grow on rocks or ground,and their life is affected by the environment,which caused parts of plants approaching extinction.The chemical constituents of the majority plants are still in the blank.The revealed chemical constituents included flavonoids,benzene glycosides,terpenoids,quinones,etc.and they possess strong pharmacological activity.In order to grasp our resources of Gesneriaceae medicinal plants,define its chemical constituents,screene exploitable species and make good use of the resource.The studies are conducted as follows(1)Analysing phenylethanoid glycosides with liquid chromatography-mass spectrometry analysis to summarized the characteristics of various types of cleavage of phenylethanoid glycosides.(2)Making extraction and LC-MS determination of the collected sanmple,then establish the chemical constituents analytical method,combing the cleavage of phenylethanoid glycosides to characterize chemical constituents of Gesneriaceae,phenylethanoid glycosides(3)Using PCA and OPLS-DA to analyze of the difference component of Lysionotus D.Don and Hemiboea Clarke to explore the chemical relationship between them.Method1.By consulting large number of documents,this paper make a comprehensive review of the chemical composition and pharmacological activity of Gesneriaceae plants.2.An appropriate method is established for the analysis of 8 kinds of phenylethanoid glycosides using ultra performance liquid quadrupole time of flight mass spectrometry instruments(UPLC-Q-TOF-MS),so that all the ingredients have a good separation,optimizing MS conditions so that information of cleavage fragments of each component is stable.3.According to primary,secondary mass information,combined with the compound database,DAD characteristic spectrum,compound ClogP value and other information,we make an overall characterization and identification of 24 Gesneriaceae samples.4.PCA and OPLS-DA were used to analyze of the difference component of Lysionotus D.Don and Hemiboea Clarke.Results and conclusion1.Analysing the primary,secondary mass information of the 8 phenylethanoid glycosides standard and summarizing fragmentation way different types of phenylethanoid glycosides;making a valid distinction of C-4’ position and C-6’ position Caffeoyl in phenylethanoid glycosides,phenylethanoid single glycoside containing glucose or rhamnose;phenylethanoid glycoside containing rhamnose-glucose or rhamnose-apiose,C-3’position and C-6’ position glycosylation in phenylethanoid glycosides.2.Gesneriaceae plant samples were analysed through UPLC-Q-TOF-MS.According to the reference fragmentation way,database,and Chemspider,Scifinder to get possible formulas candidate compound.Then through secondary mass information,DAD characteristic spectrum and compound ClogP value to make identification of the compounds.Results show that Gesneriaceae plants contain phenylethanoid glycosides,flavonoids and other types of compound,differences of typesand contents of phenylethanoid glycosides exists between different species of gesneriaceae.3.Using PC A and OPLS-DA to analyze the difference component of Lysionotus D.Don and Hemiboea Clarke.and 13 markers were selected between them,most of which are phenylethanoid glycoside.The results indicate that chemical components of Lysionotus D.Don and Hemiboea Clarke have similarities,but the distinctions between them are obvious as well.更多还原