苦苣苔科药用植物亲缘学研究
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摘要
苦苣苔科属于玄参目,分苦苣苔亚科和大岩桐亚科,有150属3700余种,我国有54属463种,都属于苦苣苔亚科。该科多种植物在我国广西、云南、贵州等南方省区民间广泛用于治疗跌打损伤、咳喘及各种疮疡肿毒等症,功效显著。为了准确掌握我国苦苣苔科药用植物资源、明确药用植物传统疗效、深入研究药用植物亲缘关系、筛选出可开发利用的优势种类,使我国苦苣苔科药用植物得到充分、合理利用,作者对我国苦苣苔科植物进行传统药物学调查的基础上综合运用植物分类学、化学分类学和分子生药学等研究方法对苦苣苔科药用植物显微特征、ITS序列、rbcL基因序列、trnH-psbA基因区间序列、苯乙醇苷类分布等方面进行了较全面研究,为可持续利用和发展提供理论依据。
主要研究结果如下：
1、通过查阅大量文献,对苦苣苔科植物的分类学研究、生药学研究、主要化学成分、药理活性、分子生药学研究五个方面的国内外研究进行了全面综述,对该科植物的全面、合理和可持续利用提供科学依据。
2、在广西大新、靖西、那坡及贵州威宁、兴义等10个地区进行了药用植物资源和传统疗效调查,结合标本馆原始标本记录和文献记载,系统整理、总结出苦苣苔科药用植物种类、分布及传统疗效,深入探讨了目前苦苣苔科药用植物存在的问题及产生的原因,并提出了建议。
3、利用石蜡制片法和水合氯醛透化法对苦苣苔科半蒴苣苔属(4种)、唇柱苣苔属(4种)、蛛毛苣苔属(3种)和吊石苣苔属(4种)4属15种药用植物进行了茎(或根状茎)和叶的横切面特征和粉末特征研究,确定了15种常用药用植物以石细胞的有无及形态、草酸钙结晶的有无及类型、维管束的连续与否为主要依据的显微鉴别特征,编写了属间和种间横切面检索表和粉末特征检索表。其中15种药用植物中除了吊石苣苔外其它14种药用植物的显微特征都是首次研究。
4、利用PCR技术等对苦苣苔科14属57种植物的ITS序列进行了比对和分析,通过DNAman、DNAstar和PAUP 4.0三种软件比较分析得苦苣苔科系统聚类树在属间聚类与传统分类学基本一致,说明利用ITS序列进行苦苣苔科属间亲缘关系研究是合适的。对药用植物比较多的半蒴苣苔属、唇柱苣苔属、蛛毛苣苔属和吊石苣苔属利用ITS序列进行了种间亲缘关系和分子鉴定研究,为合理用药提供指导。
利用PCR技术对苦苣苔科9属22种植物rbcL基因序列研究发现rbcL基因在苦苣苔科族间和属间序列相似度达到85%以上,同一属内种间序列相似度达到99%以上,说明苦苣苔科药用植物rbcL序列过于保守,不太适合进行属间或种间亲缘关系和分子鉴定研究。
利用PCR技术对苦苣苔科14属30种植物的trnH-psbA基因区间序列和对9属20种21个样品的rbcL--trnH-psbA-- ITS序列研究发现该科上述两个序列变异幅度比较大,是否可以进行属间或种间亲缘关系和分子鉴定研究还需深入研究。
苦苣苔亚科芒毛苣苔族的分类位置一直备受争议,有人认为长蒴苣苔族、芒毛苣苔族是两个独立的族,有人建议将芒毛苣苔族归入长蒴苣苔族中。作者通过对芒毛苣苔族和长蒴苣苔族的部分药用植物ITS序列、rbcL基因trnH-psbA序列及rbcL--trnH-psbA--ITS序列和系统发育树分析,支持将芒毛苣苔族归入长蒴苣苔族中。
5、用5个苯乙醇苷类化合物做对照品,采用HPLC-UV法对苦苣苔科10属24种药用植物进行了定性分析,结果表明阿克苷在在该科分布广泛,可作为研究属间或种间亲缘关系证据之一
本课题首次以药用植物亲缘学理论为指导,综合运用植物分类学、化学分类学和分子生药学等方法对苦苣苔科药用植物进行了较全面研究,创新点如下：
1、初步明确了半蒴苣苔属、唇柱苣苔属、蛛毛苣苔属和吊石苣苔属部分药用植物种类的亲缘关系,为扩大和可持续利用药用植物资源具有指导意义。
2、首次确定了半蒴苣苔属、唇柱苣苔属、蛛毛苣苔属和吊石苣苔属的15种药用植物的显微鉴别特征。
3、首次对苦苣苔科24种药用植物中苯乙醇苷类成分分布情况进行了分析。
Gesneriaceae, one group of Scrophulariales, can be divided into Cyrtandroideae and Gesnerioideae. There are 150 genera and 3700 species in all, and there are 54 genera and 463 species in China, which belong to Cyrtandroideae. Plants of this group are widely used for the treatment of traumatic injury, coughs and swelling sores in Southern China. In order to grasp accurately the source of Gesneriaceae in Chian, definite good functional results of medicinal plants of this group, thoroughly explore phylogenetic relationship of different plants, and select the priority species with fascinating prospects, on the basis of pharmacological investigation of plants of Gesneriaceae, the author has applied methods of plant taxonomy, chemotaxonomy, molecular pharmcognosy, and method so forth to make a microscopic representation, a thorough study on its ITS order, rbL gene order, trnH-psbA gene order, and distribution of phenylethanoid glycoside. This paper provides a theoretical basis to sustainable use and development of this kind of medicinal plants.
The main results as following:
1. This paper has made a review of research progress of Gesneriaceae, by means of consulting amount of documents and thoroughly research on plant taxonomy, pharmacognosy, molecular pharmcognosy, chemical constituents and pharmacology activity, this paper offers a theoretical basis to sustainable use and development of the kinds of medicinal plants.
2. An investigation of medicinal plants of Gesneriaceae was carried out in Da Xin (in Guang Xi Province), Jingxi County, Napo, Weining and Xingyi (in Gui Zhou Province) and other regions, together with consulting abundance of records of original samples and documents, we have summarized the categories, distribution and therapy effectiveness of the medicinal plants of Gesneriaceae and offered some suggestion to solving corresponding problems.
3. By means of paraffin sections and hydration chlorine and chloral hydrate, we have made a research on transverse and powder characteristics of roots, rootstocks and leaves of 4 genera and 15 species medicinal plants which separately belong to Hemiboea, Chirita, Paraboea and Lysionotus, at the same time we have defined microscopic features of these 15 species and compiled a plant key to intergeneric transverse section and interspecies powder characteristics. Except Lysionotus pauciflorus, other 14 medicinal gesneriaceae are first studied on their microscopic features.
4. We have studied the ITS order of 14 genera and 57 species of plants of gesneriaceae. Having applied soft wares of the DNAman、DNAstar and PAUP 4.0, we got the results that phylogenetic trees of plants of Gesneriaceae are almost the same according to intergeneric clustering and conventional taxonomy. That can prove that ITS order is appropriate for study on affinity of categories of Gesneriaceae. However, on account of distinction between intergeneric taxonomy and conventional taxonomy, the method of using ITS order to study affinity of intergeneric gesneriaceae still needs further study. Having used ITS order, we have studied affinity among the plants of Hemiboea, Chirita, Paraboea and Lysionotus. This research provides evidence for the guidance on proper using these medicinal plants.
Studying on the rbcL gene order of 9 genera and 22 species of plants of Gesneriaceae by means of PCR, we have found that the similarity of intergeneric gesneriaceae and intergeneric order comes up to over 85% and the similarity intrageneric order reaches up to 99%. It indicates that the order of medicinal plants of gesneriaceae is too compact to make a further research on intergeneric and intrageneric phylogeny.
Studying on trnH-psbA gene order of 14 genera and 30 species of plants and rbcL--trnH-psbA--ITS order of 21 samples of 9 genera and 20 species of gesneriaceae by means of PCR, we have found that the range of genetic variance of the two gene order are all wide. It indicates that we shoule do further research to identify them.
There is a debate about the way to classify---a subfamily of Gesneriaceae. Some scholars believe that Didymocarpeae and Trichosporeae are two independent families. Some scholars suggest that Trichosporeae should be included into the category of Didymocarpeae. Through analyzing of ITS order, rbcL gene order, trnH-psbA order, rbcL--trnH-psbA--ITS order and phylogenetic tree of some medicinal plants of Trichosporeae and Didymocarpeae, the author support the idea of classifying Trichosporeae into the category of Didymocarpeae.
5. By means of using 5 samples of phenylethanoid glycosides as reference compounds and qualitative research on 10 genera and 24 species of medicinal plants of Gesneriaceae by using the method of HPLC-UV, we have found that acteoside is widely distributing in this family. This result can be taken as one of the evidences to study intergeneric and intrageneric affinity.
At the first time we applied the methods of microscopic identification, molecule genetics identification and plant chemotaxonomy to have a research on their characteristics and affinity of medicinal plants of Gesneriaceae in our country. Significant innovations of this paper is following:
1. Initially elucidating the affinity of some plants of Hemiboea, Chirita, Paraboea, Lysionotus provides the guidance of sustainable use and development of this kind of medicinal plants.
2. At the first time we have made a study on microscopic characteristics of 15 medicinal plants of Hemiboea, Chirita, Paraboea, Lysionotus.
3. At the first time we have made a study on distribution of constituents of phenylethanoid glycoside of 24 species of medicinal plants of Gesneriaceae.
主要研究结果如下：
1、通过查阅大量文献,对苦苣苔科植物的分类学研究、生药学研究、主要化学成分、药理活性、分子生药学研究五个方面的国内外研究进行了全面综述,对该科植物的全面、合理和可持续利用提供科学依据。
2、在广西大新、靖西、那坡及贵州威宁、兴义等10个地区进行了药用植物资源和传统疗效调查,结合标本馆原始标本记录和文献记载,系统整理、总结出苦苣苔科药用植物种类、分布及传统疗效,深入探讨了目前苦苣苔科药用植物存在的问题及产生的原因,并提出了建议。
3、利用石蜡制片法和水合氯醛透化法对苦苣苔科半蒴苣苔属(4种)、唇柱苣苔属(4种)、蛛毛苣苔属(3种)和吊石苣苔属(4种)4属15种药用植物进行了茎(或根状茎)和叶的横切面特征和粉末特征研究,确定了15种常用药用植物以石细胞的有无及形态、草酸钙结晶的有无及类型、维管束的连续与否为主要依据的显微鉴别特征,编写了属间和种间横切面检索表和粉末特征检索表。其中15种药用植物中除了吊石苣苔外其它14种药用植物的显微特征都是首次研究。
4、利用PCR技术等对苦苣苔科14属57种植物的ITS序列进行了比对和分析,通过DNAman、DNAstar和PAUP 4.0三种软件比较分析得苦苣苔科系统聚类树在属间聚类与传统分类学基本一致,说明利用ITS序列进行苦苣苔科属间亲缘关系研究是合适的。对药用植物比较多的半蒴苣苔属、唇柱苣苔属、蛛毛苣苔属和吊石苣苔属利用ITS序列进行了种间亲缘关系和分子鉴定研究,为合理用药提供指导。
利用PCR技术对苦苣苔科9属22种植物rbcL基因序列研究发现rbcL基因在苦苣苔科族间和属间序列相似度达到85%以上,同一属内种间序列相似度达到99%以上,说明苦苣苔科药用植物rbcL序列过于保守,不太适合进行属间或种间亲缘关系和分子鉴定研究。
利用PCR技术对苦苣苔科14属30种植物的trnH-psbA基因区间序列和对9属20种21个样品的rbcL--trnH-psbA-- ITS序列研究发现该科上述两个序列变异幅度比较大,是否可以进行属间或种间亲缘关系和分子鉴定研究还需深入研究。
苦苣苔亚科芒毛苣苔族的分类位置一直备受争议,有人认为长蒴苣苔族、芒毛苣苔族是两个独立的族,有人建议将芒毛苣苔族归入长蒴苣苔族中。作者通过对芒毛苣苔族和长蒴苣苔族的部分药用植物ITS序列、rbcL基因trnH-psbA序列及rbcL--trnH-psbA--ITS序列和系统发育树分析,支持将芒毛苣苔族归入长蒴苣苔族中。
5、用5个苯乙醇苷类化合物做对照品,采用HPLC-UV法对苦苣苔科10属24种药用植物进行了定性分析,结果表明阿克苷在在该科分布广泛,可作为研究属间或种间亲缘关系证据之一
本课题首次以药用植物亲缘学理论为指导,综合运用植物分类学、化学分类学和分子生药学等方法对苦苣苔科药用植物进行了较全面研究,创新点如下：
1、初步明确了半蒴苣苔属、唇柱苣苔属、蛛毛苣苔属和吊石苣苔属部分药用植物种类的亲缘关系,为扩大和可持续利用药用植物资源具有指导意义。
2、首次确定了半蒴苣苔属、唇柱苣苔属、蛛毛苣苔属和吊石苣苔属的15种药用植物的显微鉴别特征。
3、首次对苦苣苔科24种药用植物中苯乙醇苷类成分分布情况进行了分析。
Gesneriaceae, one group of Scrophulariales, can be divided into Cyrtandroideae and Gesnerioideae. There are 150 genera and 3700 species in all, and there are 54 genera and 463 species in China, which belong to Cyrtandroideae. Plants of this group are widely used for the treatment of traumatic injury, coughs and swelling sores in Southern China. In order to grasp accurately the source of Gesneriaceae in Chian, definite good functional results of medicinal plants of this group, thoroughly explore phylogenetic relationship of different plants, and select the priority species with fascinating prospects, on the basis of pharmacological investigation of plants of Gesneriaceae, the author has applied methods of plant taxonomy, chemotaxonomy, molecular pharmcognosy, and method so forth to make a microscopic representation, a thorough study on its ITS order, rbL gene order, trnH-psbA gene order, and distribution of phenylethanoid glycoside. This paper provides a theoretical basis to sustainable use and development of this kind of medicinal plants.
The main results as following:
1. This paper has made a review of research progress of Gesneriaceae, by means of consulting amount of documents and thoroughly research on plant taxonomy, pharmacognosy, molecular pharmcognosy, chemical constituents and pharmacology activity, this paper offers a theoretical basis to sustainable use and development of the kinds of medicinal plants.
2. An investigation of medicinal plants of Gesneriaceae was carried out in Da Xin (in Guang Xi Province), Jingxi County, Napo, Weining and Xingyi (in Gui Zhou Province) and other regions, together with consulting abundance of records of original samples and documents, we have summarized the categories, distribution and therapy effectiveness of the medicinal plants of Gesneriaceae and offered some suggestion to solving corresponding problems.
3. By means of paraffin sections and hydration chlorine and chloral hydrate, we have made a research on transverse and powder characteristics of roots, rootstocks and leaves of 4 genera and 15 species medicinal plants which separately belong to Hemiboea, Chirita, Paraboea and Lysionotus, at the same time we have defined microscopic features of these 15 species and compiled a plant key to intergeneric transverse section and interspecies powder characteristics. Except Lysionotus pauciflorus, other 14 medicinal gesneriaceae are first studied on their microscopic features.
4. We have studied the ITS order of 14 genera and 57 species of plants of gesneriaceae. Having applied soft wares of the DNAman、DNAstar and PAUP 4.0, we got the results that phylogenetic trees of plants of Gesneriaceae are almost the same according to intergeneric clustering and conventional taxonomy. That can prove that ITS order is appropriate for study on affinity of categories of Gesneriaceae. However, on account of distinction between intergeneric taxonomy and conventional taxonomy, the method of using ITS order to study affinity of intergeneric gesneriaceae still needs further study. Having used ITS order, we have studied affinity among the plants of Hemiboea, Chirita, Paraboea and Lysionotus. This research provides evidence for the guidance on proper using these medicinal plants.
Studying on the rbcL gene order of 9 genera and 22 species of plants of Gesneriaceae by means of PCR, we have found that the similarity of intergeneric gesneriaceae and intergeneric order comes up to over 85% and the similarity intrageneric order reaches up to 99%. It indicates that the order of medicinal plants of gesneriaceae is too compact to make a further research on intergeneric and intrageneric phylogeny.
Studying on trnH-psbA gene order of 14 genera and 30 species of plants and rbcL--trnH-psbA--ITS order of 21 samples of 9 genera and 20 species of gesneriaceae by means of PCR, we have found that the range of genetic variance of the two gene order are all wide. It indicates that we shoule do further research to identify them.
There is a debate about the way to classify---a subfamily of Gesneriaceae. Some scholars believe that Didymocarpeae and Trichosporeae are two independent families. Some scholars suggest that Trichosporeae should be included into the category of Didymocarpeae. Through analyzing of ITS order, rbcL gene order, trnH-psbA order, rbcL--trnH-psbA--ITS order and phylogenetic tree of some medicinal plants of Trichosporeae and Didymocarpeae, the author support the idea of classifying Trichosporeae into the category of Didymocarpeae.
5. By means of using 5 samples of phenylethanoid glycosides as reference compounds and qualitative research on 10 genera and 24 species of medicinal plants of Gesneriaceae by using the method of HPLC-UV, we have found that acteoside is widely distributing in this family. This result can be taken as one of the evidences to study intergeneric and intrageneric affinity.
At the first time we applied the methods of microscopic identification, molecule genetics identification and plant chemotaxonomy to have a research on their characteristics and affinity of medicinal plants of Gesneriaceae in our country. Significant innovations of this paper is following:
1. Initially elucidating the affinity of some plants of Hemiboea, Chirita, Paraboea, Lysionotus provides the guidance of sustainable use and development of this kind of medicinal plants.
2. At the first time we have made a study on microscopic characteristics of 15 medicinal plants of Hemiboea, Chirita, Paraboea, Lysionotus.
3. At the first time we have made a study on distribution of constituents of phenylethanoid glycoside of 24 species of medicinal plants of Gesneriaceae.
引文
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