中国乌头属植物药用亲缘学研究

毛莨科Ranunculaceae乌头属Aconitum L．植物作为有毒植物及药用植物一直受到广泛的关注。本属全世界约有300余种,其中超过半数分布在中国。本文在总结乌头属二萜生物碱的化学分类及其分布、特征性二萜生物碱及其分类价值以及二萜生物碱的生源关系及其分类学意义的基础上,结合其毒性和疗效讨论了国产乌头属植物亲缘关系、化学成分和疗效及毒性之间的相关性,亦即药用亲缘学的初步研究。发现牛扁亚属subgen．Lycoctonum是以牛扁碱和C18-二萜生物碱为主的类群,由于其毒性中等,因而是寻找镇痛、抗炎等新药的一个对象。从二萜生物碱化学成分来看,露蕊乌头亚属subgen．Gymnaconitum并不显得最为进化,对其分类位置尚难作出最后的结论。乌头亚属subgen．Aconitum亚属下系之间的化学分类表现出如下特征：（1）唐古特乌头系ser．Tangutica和圆叶乌头系set．Rotundifolia是以内酯型二萜生物碱为主的类群,毒性较小,是新药寻找的重点研究类群。保山乌头系ser．Bullatifolia以C20-二萜生物碱如光翠雀碱和宋果灵以及C19-二萜生物碱如乌头碱、滇乌碱和尼奥灵等为主。短柄乌头系see Brachypoda显示以3-乙酰乌头碱、乌头碱、伏乌碱等高度进化的乌头碱型二萜生物碱为主,胺醇类如尼奥灵次之,有时共存其他纳哌啉型C20-二萜生物碱的特征。准噶尔乌头系ser．Grandituberosa的化学特征是以高度进化的乌头碱型如乌头碱等和比较原始的胺醇如塔拉萨敏、尼奥灵等以及C20-二萜生物碱为主,均有较大毒性。褐紫乌头系ser．Brunnea则以C20-二萜生物碱如光翠雀碱和宋果灵为主,杂有高度进化的乌头碱型二萜生物碱如乌头碱等成分。化学分类上不支持其独立成为一个分支。以上各系组成乌头亚属的原始和中间过渡类群。其中唐古特乌头系和圆叶乌头系最为相近,褐紫乌头系可能是连接保山乌头系和短柄乌头系的“桥梁”,而准噶尔乌头系更靠近保山乌头系。（2）显柱乌头系set．Stylosa是以含大茴香酸酯基的乌头碱型二萜生物碱以及塔拉萨敏和查斯曼宁胺醇类为主的类群,是块根较大的“大乌头”的主要来源,具有很大的毒性。兴安乌头系ser．Ambigua以含大茴香酸酯基的乌头碱型和原始胺醇类如塔拉萨敏、尼奥灵等二萜生物碱为主。蔓乌头系ser．Volubilia是以含大茴香酸酯基／苯甲酸酯基的乌头碱型和塔拉萨敏胺醇类以及高度进化的乌头碱型二萜生物碱为主的中间过渡类群。乌头系ser．Inflata以含15-羟基的单酯、双酯或多酯以及胺醇类鸟头碱型二萜生物碱为主,且酯基中无大茴香酸酯基,此系是草乌的主要植物来源,具有很大的毒性,应十分慎用。这些系可能代表乌头亚属进化的类群。其中显柱乌头系与兴安乌头系可能较近缘。蔓乌头系可能是连接显柱乌头系／兴安乌头系与乌头系的中间类群。另外,保山乌头系、短柄乌头系和准噶尔乌头系可能是直接向显柱乌头系、蔓乌头系和乌头系过渡的较为原始的类群。（3）岩乌头系set．Racemulosa从化学分类角度来看是一个特殊的类群,支持其独立成一个分支。其化学特征以牛扁碱型和乌头碱型的胺醇二萜生物碱如牛扁碱、异塔拉萨亭定和C20-二萜生物碱为主。如果单纯从化学成分来看,它与牛扁亚属植物似乎有一定关系。     

中国锦鸡儿属的分子系统发育

测定了中国锦鸡儿属Caragana各属下分类群20个代表种的ITS、trnL-F和trnS-G序列.基于3种DNA片段的单独分析所获得的系统发育树具有相似的拓扑结构;3种片段的合并分析提高了各分支的支持度,并获得了相似的系统发育树.落轴亚属subgen.Caragana的种类构成了一个在系统树上首先分化出来的单系分支,与形态特征和地理分布的研究一致.短齿系ser. Occidentales和长齿系ser. Bracteolatae的代表种构成了1个单独的分支,因此短齿系应被放入长齿系所属刺叶组sect.Longspina 而不是针刺组sect.Spinosae或sect.Pruinosa.分子系统学证据支持依据叶片宽窄在掌叶组sect.Frutescentes中再划分2个系的形态学研究结论;但Ser.Dasyphyllae和针刺系ser.Spinosae的亲缘关系较近,系统发育分析的结果似乎不支持在针刺组中单独划分2个系.宿轴类的物种聚成一个单系的分支,因此应被处理为一个组--鬼箭组sect.Jubatae;荚果里面被毛和无毛的种类各自构成2个小支,支持依据该特征在组下分系.系统树显示Sanczir定义的sect.Tragacanthoides然为多系类群,应将该组中所包含的刺叶组、针刺组、和鬼箭组的种类划分出来.基于ITS的遗传距离表明卷叶锦鸡儿C. ordosica与藏锦鸡儿C, tibetica应该是2个不同的种.     

论中国-喜马拉雅植物亚区乌头属植物地理分布特点

本文对乌头属Aconitum L．植物分布区内各地区的分布作了分析,统计了各地区不 同等级分类群的频度,认为中国-喜马拉雅植物亚区是乌头属植物地理分布最大的频度中心、 多样性中心和特有种的分布中心。 文中还讨论了乌头属内的演化关系,以及本属与邻近属的 系统发育关系,发现在中国-喜马拉雅植物亚区既有许多原始类群,又有大量的进化类群,提出 了本亚区不但是本属植物原始类群的保存中心,而且是活跃的分化中心。产生上述结果的原因可能与喜马拉雅山脉的抬升以及本亚区复杂的自然条件有着密切的关系。     

国产12种乌头属和18种翠雀属植物的细胞学研究

研究了12种乌头属Aconitum L．和18种翠雀属Delphinium L．植物的染色体。在12种乌头属植物中,除粗花乌头A．crassiflorum为四倍体(2n=4x=32)外,其他种类都为二倍体(2n=2x＝16),中甸乌头 A.piepunense中有B染色体存在,牛扁亚属Aconitum subgen．Lycoctonum的二倍体植物与乌头亚属Aconitum subgen．Aconitum 植物的染色体在大小和形态上有明显区别;所有18种翠雀属植物都为二倍体(2n＝2x=16),其染色体在大小和形态上极为相似,但与乌头亚属的染色体易于区别。翠雀属植物的核型不对称性程度明显高于乌头属植物,因此从染色体证据来看,翠雀属要比乌头属进化。     

基于不同DNA序列对四数獐牙菜系统位置分析

以扁蕾属植物为外类群,基于trnL-trnF序列、rpl16序列以及 ITS序列,采用最大简约法对四数獐牙菜的系统位置进行了分析.结果显示:尽管在ITS系统发育树、trnL-trnF系统发育树和rpl16系统发育树中所用的种和数量有所不同,所得到的3种系统发育树的拓扑结构也不一致,但四数獐牙菜的系统位置在3种系统发育树中的结果基本一致.在trnL-trnF系统发育树中,四数獐牙菜与花锚属的H.elliptica、H.brevicornis和H.weddelliana形成了一个单系群;而在ITS和rpl16系统发育树中,四数獐牙菜与歧散獐牙菜先聚在一起,再与花锚属的植物聚为一支支持率较高的单系支.研究表明,四数獐牙菜与Anagalidium属的歧散獐牙菜的亲缘关系最近,其次是与花锚属的植物,而与獐牙菜属植物的亲缘关系较远.本实验系统发育的结果支持将四数獐牙菜从獐牙菜属中分出而归入Anagalidium属的观点.     

国产7种乌头属植物的核型研究

本文报道了国产7种乌头属植物（两色乌头（Aconitum alboviolaceum Kom.,牛扁 A.barbartum var.puberulum Ledeb.,蔓乌头A.volubile Pall.,展毛蔓乌头 A.ciliare DC.,北乌头 A.kusnezoffii Reichb.,蒿叶乌头 A.artemisiaefolium Bar.et Skv.,细叶乌头 A.macrorhgnchum Turcz.）的核型。其中展毛蔓乌头、蒿叶乌头和细叶乌头的核型为首次报道。本文还首次指出乌头属中具多年生根状茎的牛扁亚属植物的核型没有明显的二型性,而具二年生块根的乌头正属植物的核型的二型性十分明显。此外还讨论了细叶乌头及展毛蔓乌头的系统位置,认为它们与蔓乌头有较近的亲缘关系。     

国产乌头属的化学分类

根据二萜生物碱的类型、生物合成、在乌头属植物中的分布,并参照国产乌头属的系统分类、形态演化和地理分布,本文讨论国产乌头属的化学分类。 1．以牛扁碱型成分为主的牛扁亚属和以乌头碱型成分为主的乌头亚属可能在乌头属进化的初期阶段就已分化,各自沿着独立的道路发展。2．乌头亚属包括以下类群：(1)以阿替生型、维特钦型、乌头碱型胺酵和 酯碱为主的保山乌头系,主要分布于国产乌头属近代发展分化中心之一的横断山脉和金沙江流域,可能国产乌头组的近代分化就是由该系发展而来; (2)以乌头碱、中乌头碱、下乌头碱为主的乌头系,为进化程度较高的群,所含乌头碱及尼奥灵显示了该系与保山乌头系的亲缘关系; (3)以乌头碱和松果灵为主的准噶尔乌头系,所含乌头碱和松果灵显示了该系与保山乌头系的亲缘关系; (4)以滇乌碱类酯碱为主的显柱乌头系和蔓乌头系,为进化程度较高的群,其酯碱有别于乌头碱类。3．以阿替生及C₁₉内酯型为主的甘青乌头系及圆叶乌头系,可能为进化早期形成的高山特化类群。4. 以阿替生和C₁₉乌头碱型胺醇为主的露蕊乌头亚属,可能为特化类群。     

西藏乌头属二新种

描述了自西藏自治区发现的毛茛科乌头属二新种,新腋花乌头（Aconitum novoaxillare）和截基乌头（A．basitruncatum）,并给出此二新种与近缘种的区别特征。     

基于ITS序列对中国簇毛黄耆亚属（黄耆属）系统学问题的初步研究

测定了簇毛黄耆亚属（Pogonophace)4组8种和外类群Caragana roborovskyi的ITS序列,从GenBank中调出相关12属47种的ITS序列,组成数据距阵,应用PAUP程序中的最大简约法构建了系统发育树状图。扁荚组（Sect.Sesbanella)与亚属其余类群在系统树上处于不同的分支,亲缘关系较远这个亚属不是一个单系类群;膨果组（Sect.Bibracteola)。背扁组（Sect.Phyllolobium)和袋果组（Sect.Trichostylus)作为一个单系类群能得到ITS序列的支持,但与分组以及膨果组下系的划分都得不到ITS序列分析的支持。     

一株淡水多甲藻的形态观察和多甲藻属系统发育分析

通过形态学方法将中国淡水藻种库一株编号为FACHB-329的甲藻鉴定为楯形多甲藻不等变种Peridinium umbonatum var.inaequale Lemmermann,并且描述了此甲藻所产生的孢囊形态。还对该藻的SSU和LSU rDNA序列进行测序,系统发育分析的结果也支持形态鉴定。系统发育研究结果表明多甲藻属是多系起源的,本研究所用的多甲藻属类群可以形成两个大的分枝——Peridinium umbonatum类群形成的分枝和狭义多甲藻属类群（Peridinium sensu stricto）形成的分枝。并且形态学和分子数据都显示了P.umbonatum类群与狭义多甲藻属类群之间的差异。P.umbonatum类群形成的淡水单系类群虽然没有高度的自展支持但获得了不同的系统发育分析的支持,而狭义多甲藻属类群形成的单系类群获得了高度的支持。淡水多甲藻P.aciculiferum和P.wierzejskii则与海洋斯氏藻属关系密切。     

Phylogeny of the Aconitum delavayi complex (Ranunculaceae ) based on evidence from nuclear ribosomal ITS sequences

Internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA (nr DNA) we re used to investigate the phylogeny of the Aconitum delavayi complex that was found mainly in Hengduan Mountains, SW China. Based on maximum parsimony and neighbor -joining     

Phylogenetic inference of the genus Bupleurum (Apiaceae) in Hengduan Mountains based on chromosome counts and nuclear ribosomal DNA ITS sequences

pleurum of Apiaceae in China. This paper reports chromosome numbers of six species and two varieties of Bupleurum, and for four species and two varieties their chromosome numbers are reported for the first time. The phylogeny of Bupleurum was investigated based on the ITS region of the nuclear ribosomal DNA (nrDNA) of 14 taxa from the Hengduan Mountains, 3 taxa from the North China (Hebei and Heilongjiang), and 16 taxa from Africa and the Mediterranean region. Varia-tions in chromosome numbers and the ITS sequences were used to infer phylogenetic relationships between Bupleurum species in Hengduan Mountains. The results showed that the Hengduan Mountains might represent one of the frequency and diversity centers for Bupleurum. The ancestors of Bupleurum species in the Hengduan Mountains may be related to the woody B. fruticosum in North Africa, or the species in the western Mediterranean region. It is postulated that the ancestral population migrated into Hengduan Mountains through the Middle East and the Caucasus. Furthermore, the neo-endemic B. mundtii in South Africa appeared to be a close relative of the species in the Hengduan Mountains. In the trend of basic chromosome number evolution, x = 8 should be regarded as the ancestral basic number, while x = 6, 7 as the derived ones. The Bupleurum species in the Hengduan Moun-tains have been undergoing changes in the basic chromosome numbers or the ploidy level. The ITS phylogenetic tree showed that the Chinese species were divided into two clades: one with the basic chromosome number x = 8, and the other with x = 6, 7. The results rejected the previous infrageneric classification of Bupleurum in China. We further suggested to raise B. marginatum var. stenophyllum to species rank based on the combined evidence from morphology, karyology, pollen morphology, and the ITS phylogenetic tree.     

基于染色体计数和ITS序列初步探讨横断山区柴胡属植物(伞形科)的系统发育

横断山区是中国柴胡属Bupleurum植物的分布中心。本文对横断山区6个种2变种进行了染色体记数报道,其中4个种2变种是首次报道。对横断山区的10个种4个变种、中国北方（河北和黑龙江）的3个种的nrDNA ITS进行测序,同时从GenBank里面下载同属的来自非洲和地中海西部的16个nrDNA ITS序列数据,结合染色体数目变化结果,初步探讨了横断山区柴胡属植物的系统发育。结果表明横断山区可能是现代柴胡属植物的频度中心和多样分布中心之一。它们的祖先种可能是非洲北部的木本柴胡属植物B.fruticosum,或者是地中海西部的柴胡属植物,推测是通过中东和高加索扩散而形成的,其中与非洲南部特有种B.mundtii的亲缘关系也较近;染色体基数演化趋势是：8是较原始基数,6和7是次生基数,其染色体异基数变异和多倍化可能是物种形成、进化以及向外扩散的主要方式;在ITS系统发育树中,中国柴胡属植物染色体基数为8的种类聚为一支,染色体基数为6和7的种类聚为了一支,不支持舒璞等（1998）关于中国柴胡属的属下分类系统。结合已有的形态学、细胞学、孢粉学证据和ITS系统发育树,建议窄竹叶柴胡B.marginatum var.stenophyllum独立成种。     

Nuclear ribosomal ITS sequences and phylogeny in East AsianAconitum subgenusAconitum (Ranunculaceae), with special reference to extensive polymorphism in individual plants

Internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA (nrDNA) were used to examine the phylogeny of East Asian aconites. Individual aconites were discovered to contain as many as eight different ITS sequences after cloning and PCR-SSCP (single-stranded conformational polymorphisms) analysis. We identified eight putative ITS pseudogenes from four taxa with low predicted secondary structure stability and high substitution rates. Maximum likelihood (ML) and neighbor-joining (NJ) methods were used for phylogenetic reconstruction. The ITS trees agree with the previous chloroplast DNA (cpDNA) tree for the vast majority of the taxa. We found two East Asian clades in the ITS trees: 1) a clade with the Chinese diploid,Aconitum volubile and East Asian tetraploids, and 2) a clade of East Asian diploids and Siberian tetraploids. In the former clade, most tetraploid taxa appear to be polyphyletic; sequences from individual plants did not correspond to recognized taxonomic units. This indicates a recent divergence of the East Asian tetraploids.     

A phylogenetic analysis of the Schisandraceae based on morphology and nuclear ribosomal ITS sequences

Cladistic analyses were conducted using morphological and ITS sequence data to examine phylogenetic relationships within the Schisandraceae. The most parsimonious trees obtained from the morphological data consistently show a dichotomy, with clades corresponding with the two currently accepted genera, Kadsura and Schisandra . In the ITS trees S. propinqua consistently groups with the Kadsura clade, and the position of S. glabra is uncertain. The trees resulting from the analysis of combined data are polytomous, due to the variable positions of K. scandens, K. coccinea and S. propinqua . The monophyletic status of Kadsura subgen. Kadsura sect. Kadsura and Schisandra subgen. Pleiostema are strongly supported, although the monophyly of other supraspecific taxa is unclear. Further studies are required before an unambiguous phylogeny is achieved for the family.     

Molecular evaluation of nominal species in the Ceratitis fasciventris, C. anonae, C. rosa complex (Diptera: Tephritidae)

Ceratitis fasciventris, C. anonae and C. rosa form a complex of economically important fruit fly pests infesting a variety of crops in African countries. Hitherto only adult males of these species can be distinguished easily by morphological characters. Other stages cannot, and for some taxa the taxonomic interpretation and species boundaries remain unclear. In order to clarify phylogenetic relationships and taxonomic status of these species, sequences of mitochondrial (16S, COI, ND6) and nuclear markers (period, ITS1) were analysed in specimens of the three morphospecies throughout the distribution of the complex. Maximum likelihood trees did not recover monophyletic groups corresponding to the morphospecies. Conversely, ND6 and COI divided West African C. fasciventris specimens in two consistent and bootstrap supported clades, involving specimens from Benin and from Mali/Ivory Coast, while the nuclear gene fragments per and ITS1 recovered a well-supported clade corresponding to C. fasciventris from Kenya/Uganda. Hence, the phylogenetic relationships and taxonomic interpretation of the complex appear more intricate than previously hypothesised. The current molecular data do not allow to identify C. fasciventris, C. anonae and C. rosa as distinct phylogenetic species but rather suggest that the morphospecies C. fasciventris is itself a complex of cryptic taxa.     

Molecular phylogeny of selected Old World Astragalus (Fabaceae): incongruence among chloroplast trnL-F, ndhF and nuclear ribosomal DNA ITS sequences

This study reports maximum parsimony and Bayesian phylogenetic analyses of selected Old World Astragalus using two chloroplast fragments including trnL-F and ndhF and the nuclear ribosomal internal transcribed spacer (nrDNA ITS). A total of 52 taxa including 34 euploid Old World and New World Astragalus , one aneuploid species from the Neo-Astragalus clade as a representative and 14 other Astragalean taxa, plus Cheseneya astragalina and two species of Caragana as outgroups were analyzed for both trnL-F and nrDNA ITS regions. ndhF was analyzed in 30 taxa and the same number for the combination of these three datasets were examined. In general, the trnL-F dataset and the ndhF and nrDNA ITS datasets generated more or less the same clades within Astragalus . However, in the trnL-F and ndhF phylogenies, Astragalus species are not gathered in a single clade, the so-called Astragalus s.s., as indicated by the nrDNA ITS tree. Visual inspection of these three phylogenies revealed that they were inconsistent regarding the position and relationships of Astragalus hemsleyi , A. ophiocarpus , A. annularis–A. epiglottis / Astragalus pelecinus, A. echinatus and A. arizonicus . Incongruence length difference test suggested that the trnL-F , ndhF and nrDNA ITS datasets were incongruent. In spite of this, phylogenetic analyses of the combined datasets as one unit or as three partitions generated trees that were topologically similar as a mix of the cpDNA and the nrDNA ITS trees. However, the combined dataset provided more resolved and statistically supported clades. The recently described A. memoriosus appeared closely related to A. stocksii (both from sect. Caraganella ) based on both trnL-F and nrDNA ITS sequences.     

Comparative analysis of phylogenetic relationships of grain amaranths and their wild relatives (Amaranthus; Amaranthaceae) using internal transcribed spacer, amplified fragment length polymorphism, and double-primer fluorescent intersimple sequence repeat markers.

The most economically important group of species in the genus Amaranthus is the A. hybridus species complex, including three cultivated grain amaranths, A. cruentus, A. caudatus, and A. hypochondriacus, and their putative wild progenitors, A. hybridus, A. quitensis, and A. powellii. Taxonomic confusion exists among these closely related taxa. Internal transcribed spacer (ITS) of nuclear ribosomal DNA, amplified fragment length polymorphism (AFLP), and double-primer fluorescent intersimple sequence repeat (ISSR) were employed to reexamine the taxonomic status and phylogenetic relationships of grain amaranths and their wild relatives. Low ITS divergence in these taxa resulted in poorly resolved phylogeny. However, extensive polymorphisms exist at AFLP and ISSR loci both within and among species. In phylogenetic trees based on either AFLP or ISSR or the combined data sets, nearly all intraspecific accessions can be placed in their corresponding species clades, indicating that these taxa are well-separated species. The AFLP trees share many features in common with the ISSR trees, both showing a close relationship between A. caudatus and A. quitensis, placing A. hybridus in the same clade as all grain amaranths, and indicating that A. powellii is the most divergent taxon in the A. hybridus species complex. This study has demonstrated that both AFLP and double-primer fluorescent ISSR have a great potential for generating a large number of informative characters for phylogenetic analysis of closely related species, especially when ITS diversity is insufficient.     

A systematic study of North American Angelica species (Apiaceae) based on nrDNA ITS and cpDNA sequences and fruit morphology

Angelica is a taxonomically complex genus widespread throughout the North Temperate Zone. Previous phylogenetic studies of the genus have focused primarily on its East Asian species. The relationships among its North American members, the monophyly of these species, and the value of fruit morphology in circumscribing its taxa have yet to be examined. This study represents the most comprehensive sampling of Angelica to date (100 species) and includes all 26 species in North America. Relationships are inferred using Bayesian inference, maximum likelihood, and maximum parsimony analyses of ITS sequences and, for multiple accessions of each North American species, cpDNA ndhF-rpl32, rpl32-trnL, and psbM-psbD sequences. The fruit morphological characters examined were those considered phylogenetically important in East Asian Angelica. The results revealed that the North American species fell into three major clades: North American Angelica clade, Archangelica clade, and the Eurasian Angelica clade. Angelica dawsonii has affinities with Lomatium brandegeei. Fourteen species within the North American Angelica clade were strongly supported as monophyletic. Two paraphyletic species resulted in new combinations in A. lineariloba and A. venenosa. Conflict between the ITS-derived and cpDNA-derived phylogenies and the lack of resolution in portions of the trees may be due to chloroplast capture and rapid species radiation. Fruit morphology supported some interspecific relationships based on molecular data, and relationships revealed by ITS and cpDNA data were roughly in accordance with fruit classification type and geographic distribution region, respectively. A diagnostic key based on fruit morphology is provided for the identification of the North American Angelica taxa.