川西獐牙菜的化学成分、药理作用和临床应用研究进展

川西獐牙菜是我国传统藏药"藏茵陈"原植物之一,含有呫吨酮、环烯醚萜苷、三萜等化学成分,具有保肝、抑菌等药理作用。其临床应用主要为各型肝炎的治疗,已被开发为多种剂型的成药,该植物其他方面的应用还有待于进一步研究。对该植物的化学成分、药理作用和临床应用研究进展做一综述,以期为开发和利用川西獐牙菜提供参考。     

藏药蒂达原植物在四川的地理分布与植物特征

目的 研究藏药蒂达原植物藏菌陈在四川地区的资源分布.方法 在野外考察的基础上,结合文献资料,总结了四川地区用作藏菌陈的植物种类,并研究其地理分布.结果 实地考察分析发现,川西獐牙菜和椭圆叶花锚作为蒂达原植物在四川地区分布广泛.结论 川西獐芽菜和椭圆叶花锚作为藏药蒂达原植物,应加快引种栽培.     

不同种獐牙菜中獐牙菜苦苷含量比较

目的比较不同品种獐牙菜植物中獐牙菜苦苷含量,为合理选择利用资源提供依据。方法甲醇超声提取獐牙菜植物中的活性成分,通过HPLC测定其中的獐牙菜苦苷含量。色谱条件：ZORBAX SB-C18（4.6mm×150mm,5μm）色谱柱,流动相为水-乙腈（92∶8）,流速1mL·min^-1,柱温25℃,检测波长为238nm。结果獐牙菜苦苷在检测范围内线性良好（r=0.9994）,加样回收试验的平均回收率为99.2%,重复性试验RSD为1.73%（n≥5）。几种獐牙菜中獐牙菜苦苷含量在0.58%～12.86%。结论獐牙菜苦苷含量在獐牙菜属植物中种间差别比较大。     

HPLC法测定川西獐牙菜中獐牙菜苷的含量

目的建立川西獐牙菜所含獐牙菜苷的HPLC定量测定方法。方法采用HPLC法。色谱柱:C18柱(200mm×4.6mm,5μm)柱;流动相:乙腈-0.1%磷酸(10∶90);流速为1.0mL·min-1;检测波长:237nm;柱温:30℃。结果獐牙菜苷在0.24～1.2μg范围内线性关系良好,平均回收率为102.6%,RSD为2.2%。结论所建立的检测方法可快速有效的测定獐牙菜苷。     

不同种獐牙菜中獐牙菜苦苷含量比较

目的 比较不同品种獐牙菜植物中獐牙菜苦苷含量,为合理选择利用资源提供依据。方法 甲醇超声提取獐牙菜植物中的活性成分,通过HPLC测定其中的獐牙菜苦苷含量。色谱条件： ZORBAX SB-C₁₈(4.6 mm×150 mm,5 μm)色谱柱,流动相为水-乙腈(92∶8),流速1 mL·min^-1,柱温25 ℃,检测波长为238 nm。结果 獐牙菜苦苷在检测范围内线性良好(r=0.999 4),加样回收试验的平均回收率为99.2%,重复性试验RSD为1.73%(n≥5)。几种獐牙菜中獐牙菜苦苷含量在0.58%~12.86%。结论 獐牙菜苦苷含量在獐牙菜属植物中种间差别比较大。     

11种獐牙菜及近缘植物中有效成分的高效液相色谱测定

建立了獐牙菜中3种苦味甙,即獐牙菜苦甙(swertiamarin)、獐牙菜甙(sweroside)、龙胆苦甙(gentiopicroside)及一种　酮甙──獐牙菜　酮甙(swertianolin)的高效液相色谱定量分析方法(μ-BondpakC₁₈柱,水—甲醇—异丙醇—四氢呋喃=65:30:5:1作流动相);对11种獐牙菜及近缘植物椭圆叶花锚、坚龙胆进行了含量测定,并对他们之间的异同进行了比较。     

高纯度獐牙菜苦苷的制备方法

目的从川西獐牙菜药材中制备高纯度獐牙菜苦苷。方法川西獐牙菜经醇提水沉后,上清液经HPD-300大孔吸附树脂吸附洗脱后得獐牙菜苦苷粗提物,再用闪式柱色谱对其进行分离。结果制备得到的獐牙菜苦苷,纯度在90%以上,工艺总收率为79.94%。结论本实验建立的制备高纯度的獐牙菜苦苷的方法,可操作性强,重现性较好,工艺收率和产品纯度均较为理想。     

獐牙菜属及近缘植物中齐墩果酸的高效液相色谱定量分析

目的:比较獐牙菜属及近缘植物中齐墩果酸的含量,为资源利用提供依据。方法:高效液相色谱法测定獐牙菜属及其近缘植物不同部位中的齐墩果酸含量,Shim-Pack VP-ODS(5 μm,4.6 mm×150 mm)柱,流动相:乙腈、水(78:22),检测波长215 nm,0.05 AUFS,流速1.0 mL·min~(-1),柱温40℃。结果:测定贵州獐牙菜中齐墩果酸的平均回收率为102.1％,重现性实验的RSD为1.15％,符合定量分析要求。分别测定了14种獐牙菜属及近缘植物不同部位齐墩果酸的含量。14种植物花、叶、茎中均含齐墩果酸,其中花中的含量最高,从0.937％到1.386％;叶与茎中的含量其次,且含量差异很大,从0.236％到1.041％;根中的含量为零。结论:本次实验说明这些在民间作为獐牙菜使用的植物中都含有保肝成分齐墩果酸,且在花中的含量明显高于其他部位。     

狭叶獐牙菜的两个新环烯醚单萜甙

本文报道狭叶獐牙菜(Swertia angustifolia)中两个新的环烯醚单萜甙的结构。分别命名为狭叶獐牙菜苦甙(angustiamarin,Ⅰ)和狭叶獐牙菜甙(angustioside,Ⅱ),此外还鉴定了獐牙菜甙(sweroside,Ⅲ),獐牙菜苦甙(swertiamarin,Ⅳ)和表优士特莫甙(epi-eustomoside,Ⅴ)。     

藏药二十五味珊瑚丸中獐牙菜藏药材名称的探讨

目的:探讨藏药二十五味珊瑚丸中獐牙菜药材名称的准确表述.方法:依据藏医药古籍及现代文献记载、现行药品标准和藏药习用品使用情况进行分析.结果:獐牙菜之名系多种藏药材的总药材名称,不能满足一药一物的要求,因此不宜在国家药品标准中使用.结论:藏药成方制剂中獐牙菜名称,应修订为桑蒂(川西獐牙菜),或甲蒂(印度獐牙菜),或帕蒂(...     

Authentication of stems of Dendrobium officinale by rDNA ITS region sequences

The rDNA ITS regions of five Dendrobium species were sequenced. Each Dendrobium species was found to have a unique sequence in the ITS region, so that they could be easily distinguished at the DNA level. The aligned 644 bp of the ITS region includes 235 bp ITS1, 163 bp 5.8S, and 246 bp ITS2. One hundred and eighty-nine sites are variable. The sequences of D. officinale could be easily distinguished from the other four adulterant species according to the sequence variation at 11 sites, 7 in ITS1, 1 in 5.8S, and 3 in ITS2. These could be used as molecular characters to distinguish the stems of D. officinale from the adulterants.     

Differentiation of Dendrobium species used as "Huangcao Shihu" by rDNA ITS sequence analysis

The genus Dendrobium Sw. is composed of 74 species and two varieties in China, and 32 species carry the name "Huangcao Shihu" on the herbal medicine market, making the identification of the origin of "Huangcao Shihu" difficult for consumers. Here, the ITS regions were sequenced and evaluated to differentiate the 18 Dendrobium species used as "Huangcao Shihu". Diversity in DNA sequences among various species was found with the inter-specific sequence divergence ranging from 3.2% to 37.9% in ITS1 and 5.0% to 26.6% in ITS2. Moreover, the variations within species were very low, ranging in sequence divergence from 0 to 3.0% in ITS1 and 0 to 4.0% in ITS2. Therefore, these species could be easily distinguished at the DNA level. Furthermore, based on the divergent ITS regions, five pairs of species-specific primers were designed and used for the rapid PCR identification of five Dendrobium species listed in the Chinese Pharmacopoeia.     

Authentication of medicinal Dendrobium species by the internal transcribed spacer of ribosomal DNA

Herba Dendrobii (Shihu) is a commonly used Chinese medicine derived from the stem of several orchid species belonging to the genus Dendrobium. It is rather expensive and adulteration is frequent. Proper authentication of the medicinal species is necessary to protect consumers and support conservation measures. DNA sequences of the internal transcribed spacer 2 (ITS 2) of 16 Dendrobium species were shown to be significantly different from one another by an average of 12.4% and from non-orchids and Pholidota (an adulterant of Shihu) by 29.8% and 18.8%, respectively. The intra-specific variation among the Dendrobium species studied was only about 1%. Therefore, ITS 2 regions could be adopted as a molecular marker for differentiating medicinal Dendrobium species from one another and also from non-orchids and adulterants.     

基于5S-rRNA 基因间区碱基序列鉴定繁缕

目的建立快速准确鉴定民间药繁缕(Stellariamedia)的方法。方法首次利用DNA分子鉴定技术对繁缕及混淆品牛繁缕(Myosotonaquaticum)进行了5SrRNA基因间区的PCR扩增和测序。结果DNA序列和限制性酶切谱可以用于鉴定繁缕及牛繁缕。同时对石生繁缕(S.vestita)、长叶繁缕(S.longifolia)及垂梗繁缕(S.radians)进行了5SrRNA基因间区的PCR扩增和测序,认为上述5种植物各自的DNA序列和限制性酶切谱可用于繁缕属分子系统学研究。结论该方法可用于药用植物繁缕的鉴定及繁缕属分子系统学研究。     

Characters of nrDNA ITS region sequences of fruits of Alpinia galanga and their adulterants.

The fruits of Alpinia galanga (L.) Sw. are used as a traditional Chinese medicine; but the dry fruits of A. conchigera, A. suishaensis, A. maclurei and A. polyantha are also used as the medicine in local areas. Because dry fruits of these related plants are similar to those of Alpinia galanga (L.) Sw. in odor, morphological characters and chemical components, and even anatomical characters, it is difficult to identify the medicine. Nuclear ribosomal DNA internal transcribed spacer (ITS) regions of the five taxa were directly sequenced using an automated sequencer. Sequence analysis showed that the ITS 1 ranges from 177 to 178 base pairs (bp), and the ITS 2 from 225 to 234 bp. The size of the 5.8S coding region is 164 bp for all species. Also, the pairwise sequence divergence is higher and some molecular markers were determined. According to these molecular markers, Alpinia galanga (L.) Sw. and the related species can easily be distinguished from each other. Therefore, evidence from nrDNA ITS sequence variation can identify the medicine at the DNA level.     

Molecular authentication of the traditional Tibetan medicinal plant Swertia mussotii

Swertia mussotii is an important species in Tibetan folk medicine. However, it is quite expensive and frequently adulterated, so reliable methods for authentication of putative specimens and preparations of the species are needed to protect consumers and to support conservation measures. We show here that the chloroplast (cp) DNA RPL16 intron has limited utility for differentiating S. mussotii from closely related species, since the cpDNA RPL16 sequences are identical in S. mussotii and two other species of Swertia. However, the rDNA internal transcribed spacer (ITS) sequences differ significantly between S. mussotii and all of 13 tested potential adulterants. Thus, the ITS region provides a robust molecular marker for differentiating the medicinal S. mussotii from related adulterants. Therefore, a pair of allele-specific diagnostic primers based on the divergent ITS region was designed to distinguish S. mussotii from the other species. Authentication by allele-specific diagnostic PCR using these primers is convenient, effective and both simpler and less time-consuming than sequencing the ITS region.     

Identification of Dryopteris crassirhizoma and the adulterant species based on cpDNA rbcL and translated amino acid sequences

Dry rhizome of Dryopteris crassirhizoma Nakai (Dryopteridaceae), also known as Guan Zhong, is a traditional Chinese herbal medicine used in the treatment of viral disease. But the dry rhizomes of Woodwardia JAPONICA (L. f.) Sm., OSMUNDA JAPONICA Thunb. and Cyrtomium fortunei J. Sm. are also used as Guan Zhong in local areas. The adulterants are similar to Dryopteris crassirhizoma. It is difficult to identify the botanical origin of these herbs. In our study, sequences of the cpDNA RBCL gene were determined and analyzed for Dryopteris crassirhizoma and adulterant species, where nineteen molecular markers had been determined. Also, amino acid sequences translated from the RBCL gene were analyzed and four important molecular markers were detected. Based on cpDNA RBCL and translated amino acid sequences, Dryopteris crassirhizoma can easily be distinguished from the other three fern species.     

药用植物束花石斛、流苏石斛及其形态相似种的PCR-RFLP鉴别研究药用植物束花石斛、流苏石斛及其形态相似种的PCR-RFLP鉴别研究

目的建立简易的采用rDNA ITS区作为分子标记对中药石斛的基源植物进行分子鉴定的技术。方法 用聚合酶链反应—限制性片段长度多态性(PCR-RFLP)方法获得ITS片段的限制性图谱。束花石斛及其形态相似种的PCR扩增产物用Cla I和Apa LI酶切,流苏石斛及其形态相似种的PCR扩增产物用Sph I酶切。结果根据预测的PCR-RFLP图谱,可以鉴别药用植物束花石斛、流苏石斛及它们的形态相似种。用这种方法鉴定了市场上收集的25件商品束花石斛、流苏石斛新鲜药材的原植物。结论rDNA ITS的PCR-RFLP可用于鉴定石斛药材的原植物,具有简便、费用低的优点。     

ITS2+psbA-trnH复合序列鉴定徐长卿、白薇和白前

目的：建立以ITS2+psbA-trnH复合序列鉴定徐长卿、白薇和白前及其同属近缘混伪品的DNA条形码鉴定方法。方法：搜集徐长卿、白薇、白前及其近源混伪品,采用改良的CTAB法提取DNA,通过实验分别获得ITS2和psbA-trnH序列,将同一样本的ITS2序列与psbA-trnH序列整合得到43条ITS2+psbA-trnH复合序列,从GenBank数据库中下载来源于同一样本的ITS2序列与psbA-trnH序列整合得到14条ITS2+psbA-trnH 网上复合序列,用MEGA 6.05软件分析徐长卿、白薇、白前及其近源混伪品的复合序列变异位点,计算种内、种间的K2P距离,并构建NJ系统聚类树。结果：徐长卿、白薇和白前药材ITS2+psbA-trnH复合序列比对后产生17个变异位点;徐长卿、白薇和白前基源物种种内最大K2P距离均小于其与混伪品的种间最小K2P距离;系统NJ树能准确将徐长卿白薇和白前及其近缘混伪品。结论：该研究建立了应用ITS2+psbA-trnH复合序列鉴定徐长卿、白薇、白前及其近缘混伪品的DNA条形码鉴定方法。     

基于matK和ITS2及二级结构对药材香薷及其混伪品的鉴别研究

目的 快速、准确鉴别药材香薷及其混伪品，保障香薷的药材质量和用药安全。方法 收集石香薷、江香薷和香薷植物材料分别进行matK和ITS2序列的扩增与测序，测序结果经Codon Code Aligner软件校对，同时从GenBank下载石香薷、江香斋及其易混品种海州香薷、香薷、密花香薷、牛至等物种的matK和ITS序列。其中，ITS序列经隐马尔可夫模型去除两端的5.8S和28S序列，共得到16个物种的ITS2序列50条；经Clustal软件校对共获得9个物种的matK序列28条。通过Mega7.0软件分析matK和ITS2序列，计算所有物种种内和种间遗传距离，构建邻接法(neighbor joining，NJ)聚类树，通过ITS2 Database预测ITS2二级结构，采用4Sale软件比对二级结构，通过ProfDistS软件构建基于联合ITS2一级序列及其二级结构的剖面邻接(profile neighbor-joining，PNJ)系统发育树。结果 基于matK和ITS2序列的遗传距离均表明香薷正品与其各种混伪品之间存在明显barcoding gap。NJ和PNJ进化树的拓扑关系一致，可以区分药材香薷及其混伪品。香薷的ITS2二级结构与其各混伪品具有显著差异。结论 建议matK和ITS2序列均可以作为鉴别香薷与其混伪品的DNA条形码，ITS2二级结构信息的加入可丰富鉴定结果，为香薷药材的准确鉴别、香薷属与石荠苎属植物的科学分类提供参考。