High diversity of Ligularia dictyoneura in chemical composition and DNA sequence

The chemical composition of root extracts of the title species collected at 20 different places in the Hengduan Mountains of China was examined. From these samples, a total of 17 eremophilane derivatives were isolated, three of which were new franoeremophilane derivatives: 3beta-acetoxy-6beta-(angeloyloxy)furanoeremophilan-10beta-ol, 1alpha-acetoxyfuranoeremophilan-15,6alpha-olide, and 6beta-[2-(hydroxymethyl)prop-2-enoyloxy]furanoeremophil-1(10)-ene. Based on the chemical composition, the samples could be classified into as many as seven types: one type containing non-furanoeremophilane derivatives and the other six containing furanoeremophilane derivatives with different oxidation levels. Results of DNA sequencing of the atpB-rbcL region and the internal transcribed spacers of the ribosomal RNA gene also indicated a high diversity in the species.     

Chemical and genetic differentiation of Ligularia tsangchanensis in Yunnan and Sichuan provinces of China

The intraspecific diversity in L. tsangchanensis collected in the Chinese Provinces Yunnan and southwestern Sichuan was studied by chemical and genetic approaches. The samples collected in Yunnan were found to contain cacalol (1) as the sole major component, while samples from Sichuan contained 7alpha- and 7beta-eremophila-9,11-dien-8-one (5 and 6) as well as the 3alpha-angeloyloxy derivative 7 as major components. In addition, the sequences of the internal transcribed spacers (ITSs) of the ribosomal RNA gene indicated that the Yunnan and the Sichuan samples constitute separate clades. These results demonstrate that L. tsangchanensis in Yunnan and Sichuan are distinct.     

Chemical and Genetic Differentiation of Ligularia hodgsonii in Japan and China

The chemical composition of the root extract and neutral base sequences of L. hodgsonii samples collected in Japan and China were examined. From the Japanese and the Chinese samples, 6β‐(angeloyloxy)furanoeremophilan‐15‐oic acid ( 2 ) and 3β‐acetoxy‐6β‐[(2‐methylbutanoyl)oxy]furanoeremophilan‐10β‐ol ( 3 ) were isolated as the major component, respectively. The sequence of the internal transcribed spacers (ITSs) of the ribosomal RNA gene was different in the Japanese and the Chinese samples. In contrast to the significant genetic difference, chemical diversity was limited to the positions of O‐functionalities on the furanoeremophilane skeleton, supporting our hypothesis that the production of furanoeremophilanes is ecologically advantageous.     

7种橐吾属植物的核型

研究了7种橐吾属（Ligularia）植物的染色体和核形态。干崖子橐吾（L．kanaitzensis）的核型为2n＝2x＝58＝26m＋28sm 4st;窄头橐吾（L.stenocephala）的核型为2n=2x=58=26m 32sm;细茎橐吾（L.hookeri）的核型为2n=2x=58=30m 26sm 2st;宽戟橐吾（Llatihastata）的核型为2n=2x=58=28m 26sm(2sat) 4st;网脉橐吾（L.dictyoneura）的核型为2n=2x=58=26m 28sm 2st 2t;蹄橐吾（L.hodgsonii）的核型为2n=2x=58=28m 28sm 2t;棉毛橐吾（L.vellerea）的核型为2n=2x=58=22cm 34sm 2t。虽然这7个种的染色体数目相同,2n=58,核型主要是由m和sm染色体构成,但各类的染色体数目在种间有差异。核的对称性高,着丝点端值（T．C）为61．45％－64．96％。除窄头橐吾和鹿蹄橐吾的染色体数与前人报道的相同外,其它5个种的染色体数目和核型为首次报道。     

黄帚橐吾挥发物的化感作用及其主要成分分析

研究了青藏高原东部高寒草场上广布的菊科毒杂草--黄帚橐吾(Ligularia virgaurea)挥发物的化感作用,并运用气相色谱-质谱方法对其挥发物主要化学成分进行了分析.结果表明,黄帚橐吾挥发物对所有5种受试牧草种子的萌发速度指数和最终萌发率均产生抑制作用,其中对垂穗披碱草和大雀麦的种子萌发抑制较为显著,而对早熟禾和羊茅影响较小.黄帚橐吾挥发物中主要含有2-甲基-庚烷(9.84%)、3-甲基-庚烷(8.25%)、庚烷(7.93%)、4-甲基-1-异丙基-双环[3,1,0]己-2-烯(7.79%)、3-甲基-己烷(6.38%)、2-甲基-己烷(5.54%)、D-苧烯(.70%)等18种化合物,占总挥发物的68.24%,其中单萜类化合物含量占总挥发物的16.58%,因此萜类化合物是其主要的化感活性物质,并通过挥发途径释放后影响到受体植物种子的萌发.黄帚橐吾挥发物的化感作用对增强其生存竞争力和种群增长起着重要作用.     

Chemical and genetic study of Ligularia duciformis and related species in Sichuan and Yunnan Provinces of China

The chemical constituents of the root extracts and the evolutionarily neutral DNA base sequences were studied for 28 samples of Ligularia duciformis, L. kongkalingensis, and L. nelumbifolia collected in Sichuan and Yunnan Provinces of China. The samples could be classified into four chemotypes (1-4). Sesquiterpenoids having eremophilane and oplopane skeletons were isolated from two (Chemotype?1) and three (Chemotype?2) samples, respectively. Two new oplopane derivatives were isolated and their structures were determined. In 18 samples, phenylpropenoids were the major components (Chemotype?3). In five samples, neither phenylpropenoids nor sesquiterpenoids were found (Chemotype?4). Despite this large chemical variety, no correlation was found between the chemotype and the morphological criteria of species identification. The analysis of the evolutionarily neutral DNA regions also indicated that the samples were not separated into distinct clades and that introgression was extensive.     

Chemical Constituents in Hybrids of Ligularia tongolensis and L. cymbulifera: Chemical Introgression in L. tongolensis

Two samples with morphologies intermediate between Ligularia tongolensis and L. cymbulifera were collected in Desha, Sichuan Province, and one, in Pachahai, Yunnan Province, P. R. China. The DNA sequencing confirmed that the samples were hybrids of the two species. Tetradymol ( 1 ), the major compound of L. cymbulifera not found in L. tongolensis, was isolated from the hybrid samples collected at both locations, while furanoeremophilan‐15‐oic acid derivative 4 , a compound characteristic to L. tongolensis, was found in the Pachahai hybrid but not in the Desha hybrids. Thus, the chemical consequence of hybridization can be variable. In addition, analysis of L. tongolensis samples at Pachahai indicated that introgression has been a mechanism of generating chemical diversity in the plant. Eleven compounds including three new ones were isolated.     

Direct MS measurement of the extract of Ligularia virgaurea collected in Yunnan and Sichuan provinces of China

Introduction – There are numerous Ligularia species in the eastern Qinghai–Tibet Plateau and adjacent areas. L. virgaurea has been used as a traditional folk medicine for the treatment of stomachache and nausea. Objective – To analyse chemical constituents of L. virgaurea, grown in Yunnan and Sichuan provinces of China. Methodology – The direct MS measurement of the crude extract of plant samples was used for grouping of this species. As the main compounds were available in pure form, the main peaks were analysed by LCMS. Results – An easy and speedy method of analysis using MS spectra was developed. On the basis of the findings, L. virgaurea could be divided into two groups. The genetic studies also supported this grouping. Type 1 mainly includes virgaurenones and virgaurenolides. The MS of type 2 is quite different because it includes mainly ligularol and its congeners. Both MS were easily distinguished. Conclusion – The crude extracts of 11 L. virgaurea samples already collected in recent years were analysed and it was possible to identify them as type 1 or 2. This method was applied to three samples collected in 2009 to successfully classify them as either type 1 or 2. Copyright © 2010 John Wiley & Sons, Ltd.     

Chemical and genetic diversity of Ligularia vellerea in Yunnan, China

Intra-specific diversity in Liularia vellerea growing in the northwestern to central Yunnan province of China was studied by chemical and genetic approaches. Samples collected in the Jianchuan, Lijiang, and Zhongdian areas contained 6,15-dioxygenated furanoeremophilanes as their major components (type A); whereas samples from the Luguhu area accumulated 1,6-dioxygenated furanoeremophilanes (type B); a sample from near Kunming, however, contained 6,15-dioxygenated eremophilanolides (type C). 11 beta H- and 11 alpha H-6 beta-angeloyloxy-15-carboxyeremophil-7-en-12,8-olides (eremofarfugins D and E) were also isolated and their structures were determined. A correlation between the composition and the DNA sequence was observed in the ITSs.     

Chemical and Genetic Diversity of Ligularia kanaitzensis in the Hengduan Mountains Area. Chemical Relationship with L. subspicata

Root chemicals and the sequences of the internal transcribed spacers (ITSs) were analyzed for 9 Ligularia kanaitzensis and 3 L. subspicata samples collected in northwestern Yunnan and southwestern Sichuan, China. Subspicatins A and C were isolated from two L. kanaitzensis samples. Introgression of genes responsible for these compounds from L. subspicata was suggested by their strong connection with L. subspicata/L. lamarum and the geographical proximity of the samples to L. subspicata. DNA analysis of a set of 27 L. kanaitzensis samples including those analyzed previously showed that they belong to two clades, designated A and B. Together with the presence/absence of furanoeremophilane, the 27 samples were sorted into three groups: clade A/furan, clade B/furan, and clade B/non-furan. The ancestral plant presumably belonged to clade B/non-furan, because furanoeremophilanes are biosynthesized from eremophilan-8-ones. 1β-Angeloyloxyfukinone, a likely intermediate between fukinone and subspicatin C, was isolated for the first time. This finding allowed us to propose plausible biosynthetic pathways of subspicatins A and C.     

Chemical and Genetic Studies on Hybrid of Ligularia subspicata and Ligularia cyathiceps Collected in Yunnan Province of China

Two morphologically ambiguous Ligularia samples (samples A and B), and samples with morphology of Ligularia subspicata (sample C), Ligularia lamarum (sample D), or Ligularia cyathiceps (sample E), were collected at Tianchi Pond, Shangrila County, Yunnan Province, China. Analysis of the nucleotide sequence of the internal transcribed spacers (ITSs) in the nuclear ribosomal RNA gene cluster indicated that not only sample B but also sample D was a hybrid of L. cyathiceps and L. lamarum/L. subspicata. Although the morphology of sample A suggested that it was also a hybrid, the ITS sequence of sample A was that of L. cyathiceps. Twenty compounds were isolated from the five samples, and the structures of two new compounds 7 and 14 were determined. Furanoeremophilanes typical of L. lamarum/L. subspicata were detected not only in samples C and D, but also in samples A and B. These results indicate that the ability to produce root chemicals can spread through introgression.     

掌叶橐吾挥发油化学成分分析

掌叶橐吾[Ligularia przewalskii (Maxim.)Diels]为菊科(Compositae)橐吾属(Ligularia Cass.)植物,多年生草本,高30～130 cm,生于海拔1 100～3 700 m的河滩、山麓、林缘、林下及灌丛,产四川、青海、甘肃、宁夏、陕西、山西和内蒙古[1].橐吾属植物药用种类较多,有止咳化痰、活血化瘀、清热解毒、催吐、利尿、利胆退黄等功效[2],许多种类的根及根茎作为中药"紫菀"使用或作为其代用品.掌叶橐吾的根可用于化痰、止咳、平喘,在青海,甘肃天水和文县,陕西宝鸡、眉县和太白等地为当地所习用[3,4].     

Intra-specific diversity of the chemical composition of Ligularia lamarum in the Hengduan Mountains,China: The structures of four new eremophilanes and a new seco-eremophilane

Chemical compositions and internal transcribed spacer (ITS) sequences of five samples of Ligularia lamarum collected in Sichuan Province, China, were analyzed. Fourteen compounds, including four new eremophilanes and one new seco-eremophilane, were isolated and their structures were elucidated by spectroscopic methods. Intra-specific diversity in the chemical composition was found to be higher than previously known. The result of DNA analysis suggested that one of the samples was introgressed, although its chemical composition was typical of L. lamarum.     

棉毛橐吾和穗序橐吾的自然杂交研究

在棉毛橐吾（Ligulariavellerea）和穗序橐吾（L．subspicata）同域分布的居群中存在形态特征介于两者之间的个体。经分析,这些个体有3种不同的形态类型,可能是这两个物种之间的杂交后代。为检验这一假设,对所采集的假设亲本和中间个体材料进行了核糖体内转录间隔区ITS4—5和叶绿体atpB．rbcL测序。假设杂交个体的ITS直接测序结果具明显的叠加性,之后的克隆测序也分离出两种与假设亲本种相同的序列类型．因此。这些假设杂交个体得到了证实。据母系遗传的叶绿体atpB—rbcL测序结果,穗序橐吾同是它们的质体贡献者,即虽然这些杂交个体具不同的形态特征,但都是以棉毛橐吾和穗序橐吾为亲本且以穗序橐吾为质体贡献者的杂交后代。     

五种橐吾属植物的核型研究

首次报道了5种国产橐吾属植物的核型,结果如下：东俄洛橐吾(Ligularia tongolensis)核型为2n=58=32m 14sm 12st;侧茎橐吾(L.pleurocaulis)四川稻城居群核型为2n=58=24m 32sm 2st 3～5B,云南中甸居群核型为2n=58=36m 22sm;云南橐吾(L．yunnanensis)核型为2n=58=28m 30sm;叶状鞘橐吾(L．phyllocolea)核型为2n=58=30m 24sm 4st 1B;浅苞橐吾(L．cynthceps)核型为2n=58=24m 34sm。5种橐吾染色体数目都为58。在叶状鞘橐吾和四川稻城产侧茎橐吾中发现有B染色体存在,这在以前对橐吾属及近缘属植物的核型研究中未见报道。     

河南蹄叶橐吾根的化学成分研究

从蹄叶橐吾根乙醇提取物中分离得到11种化合物,经理化和波谱分析分别鉴定为原儿茶醛(1)、7-羟基-色原酮(2)、咖啡酸(3)、阿魏酸(4)、1,5-二咖啡酰奎宁酸(5)、当归酸(6)、β-谷甾醇(7)、胡萝卜苷(8)、蜂斗菜素(9)、异蜂斗菜素(10)及正三十五烷(11)。其中酚类化合物1～5、倍半萜类化合物9～10及脂烃11为首次从该植物中分离得到。     

蹄叶橐吾根的化学成分研究

从蹄叶橐吾（Ligularia fischeri）的根中分离得到13个化合物,通过波谱学等方法鉴定为2-hydrox-platyphyllide（1）,liguhodgsonal（2）,nsujapone（3）,6’-亚油酰基田-胡萝卜苷（4）,Luped（5）,6,7-二羟基香豆素（6）,3-谷甾醇（7）,胡萝卜苷（8）,（25,3S,4R,12E）-N-[2’-（R）-羟基二十二碳酰基]-1,3,4-三羟基-2-氨基-二十碳-12-烯（9）,单硬脂酸甘油酯（10）,α-羟基二十四烷酸（11）,1,3-二棕榈酸甘油酯（12）,二十七烷醇（13）。其中化合物1—6为首次从该种植物中分离得到。     

青藏高原东缘同域分布的2种橐吾属杂草的合子前生殖隔离

生殖隔离是阻止种间杂交、保持物种完整性和独立性的关键,对生物多样性的产生和维持有重要意义.不同物种间的生殖隔离形成阶段、方式和强度不完全相同,因而对物种间隔离机制的研究有助于揭示物种形成和维持机制.对青藏高原东缘同域分布的2种橐吾属杂草黄帚橐吾和箭叶橐吾的花期物候观察、访花昆虫观察及花粉-柱头亲和性进行研究,分析了2种橐吾多种合子前生殖隔离机制的特点及其在生殖隔离中的作用.结果表明: 黄帚橐吾种群花期比箭叶橐吾早20 d左右,虽然盛花期存在差异,但是2种群花期存在10 d左右的重叠期,并且在重叠期内2种橐吾花粉和柱头均具有活性.2种橐吾共享传粉昆虫,为泛化传粉模式,并且传粉者对2种植物的访花行为相似,存在2种植物间交叉访问.尽管2种橐吾花期不存在完全的时间隔离,也不存在完全的传粉者隔离,但2种橐吾柱头对异源花粉的亲和性差,表现为柱头对异源花粉有明显的排斥反应,异源花粉管在柱头上的生长过程明显受阻.这种花粉-柱头排异形式很有可能是2种橐吾属植物重要的合子前隔离机制,从而避免了种间的自然杂交,更好地维持了自然种群的多样性和遗传结构的稳定性.     

不同植物对高寒草场主要杂草箭叶橐吾的化感作用

用13科31种植物地上部分的水浸液(0.05 g DW.mL-1)对高寒草场主要杂草箭叶橐吾(Ligularia sagi-tata)和同域牧草垂穗披碱草(Elymus nutans)进行种子萌发实验,旨在筛选能够抑制箭叶橐吾的植物种类。结果表明:有23种供体材料对箭叶橐吾种子萌发具较强抑制作用,并降低了萌发指数(M<-0.50);有9种供体植物显著抑制箭叶橐吾,同时对牧草的伤害较少甚至有促进作用。综合比较对2种植物的综合化感效应指数(M)差异,毛茛(Ranunculus japonicus)、高乌头(Aconitum sinomontanum)、五爪金龙(Ipomoea cairica)、露蕊乌头(Aconitumgymnandrum)和乳白香青(Anaphalis lactea)作用效果显著(其M差值分别是:-0.87、-0.82、-0.71、-0.68和-0.62),这5种植物有望开发新型植物源除草剂。     

Development of Wheat-alien Lines with Added Leymus racemosus Chromosomes and 6VS/6AL Translocation Chromosomes

By chromosome C-banding and bi-color fluorescence in situ hybridization (FISH) using digoxigenin-labelled total genomic DNA of Leymus racemosus (Lam.) Tzvel. and biotinylated total genomic DNA of Haynaldia villosa (L.) Schur as probes, three wheat-alien lines with L. racemosus Lr.7 addition and H.　villosa 6VS/6AL translocated chromosomes, and eight lines with L.　racemosus Lr.14 addition and H.villosa 6VS/6AL translocated chromosomes were respectively identified from DALr.7×T6VS/6AL (93G51-4×P64) and DALr.14×T6VS/6AL （94G15×P64）F2 or F3 hybrids. Fluorescein-isothiocyanate-conjugated avidin and rhodamine-conjugated sheep anti-digoxigenin Fab fragment were used in bi-color FISH detection. The chromosomes of L.racemosus and 6VS fragment of H.　villosa were simultaneously detected by their red and green fluorescence. Powdery mildew and scab resistance were also evaluated. The result showed that the obtained plants had high resistance to these two diseases. The potential usage of bi-color FISH in identifying chromatin of L.racemosus and H.villosa was discussed.