Non-alkaloidal composition of Ephedra Herb is influenced by differences in habitats

Journal of Natural Medicines - Ephedra Herb is a crude drug defined as the terrestrial stem of Ephedra sinica, E. intermedia, or E. equisetina. It is often used to treat headaches, bronchial...     

Genetic diversity of Ephedra plants in mongolia inferred from internal transcribed spacer sequence of nuclear ribosomal DNA

Ephedrae herba has been used for treating colds, relieving coughs and asthma from ancient times. We previously reported the distribution of Ephedra sinica, E. equisetina, E. przewalskii, E. regeliana, E. monosperma and Ephedra sp. in Mongolia, and among them E. sinica and E. equisetina were potential new resources of Ephedrae herba of Japanese pharmacopoeia grade, based on our field survey and subsequent molecular and chemical assessments. However, the Ephedra population in southwestern areas showed a high possibility of having hybrid origins. Further field surveys in southwestern areas, and sequence analysis of the partial nuclear internal transcribed spacer 1 (ITS1) region, besides trnK and 18S ribosomal RNA (rRNA) gene regions, were conducted in order to obtain detailed evidence of hybridization status. As a result, the distribution of E. glauca in western area and E. lomatolepis in western-most area was confirmed. The ITS sequences from all 8 Ephedra species collected in Mongolia were roughly divided into 5 types (types I-V). Type II sequence, having several additive nucleotides, was found in Ephedra sp., E. glauca, E. regeliana and E. sinica, which provided useful information for tracing hybrid origins. Morphological, genetic and distribution evidence suggested that the hybridization of Ephedra species occurred widely in southwestern Mongolia, and several Ephedra species including E. przewalkskii and E. intermedia were involved in these events. Integrated with our previous report, trnK-, 18S- and ITS-types from pure lines of each species are proposed. In addition, we propose a practicable method for detecting additive peaks on a direct sequencing electropherogram.     

Phylogenetic analysis of the DNA sequence of the non-coding region of nuclear ribosomal DNA and chloroplast of Ephedra plants in China

Twenty-four Ephedra plants belonging to 8 species grown in the northern and western parts of China were phylogenetically analyzed for their non-coding DNA sequences, internal transcribed spacers (ITSs) of nuclear ribosomal DNA as well as trnL intron and intergenic spacers between trnL and trnF (trnL/ trnF) of the chloroplast. Based on the ITS sequences, the 8 species could be divided into 3 groups: Group 1 (Ephedra intermedia, E. sinica, E. przewalskii), Group 2 (E. equisetina, E. monosperma, E. gerardiana), and Group 3 (E. likiangensis, E. minuta). The species classified into Group 1 grow mainly in the north, Group 3 in the south and Group 2 in the center, suggesting their genetic and geographic relationships. A specific primer set was designed to classify the 3 groups by routine PCR. Combined analysis of ITS and trnL/ trnF differentiated the 8 Ephedra species.     

国产麻黄的形态组织学研究——Ⅰ.北方主产的七种麻黄

据文献记载和作者的调查,我国产麻黄属(Ephedra)植物共有13种3变种1变型。本文报道我国北方生产的7种麻黄生药形态组织学比较研究结果。7种麻黄是:草麻黄Ephedra sinica Stapf、木贼麻黄E.equisetina Bunge、中麻黄E.intermedia Schrenk ex Mey.、膜果麻黄E.przewalskii Stapf,单子麻黄E.monosperma Gmel.ex Mey.、雌雄麻黄E.fedtschenkoae Pauls和细子麻黄E.reaeliana Florin。文中附有生药性状与组织构造特征比较表及生药组织图。     

十二种国产麻黄的品质评价

本文应用高效液相色谱法对我国24个产地所产的12种麻黄生药进行了六种生物碱的定量分析,这六种生物碱是:麻黄碱(ephedrine),伪麻黄碱(pseudoephedrine),去甲基麻黄碱(norephedrine),去甲基伪麻黄碱(norpseudoephedrine),甲基麻黄碱(methylephedrine)和甲基伪麻黄碱(methylpseudoephedrine)。根据分析结果,对这些麻黄生药的品质作出了评价。     

Herbological studies on Chinese crude drug Ma-huang. Part 1-On the botanical origin of Ma-huang in ancient China and the origin of Japanese Ma-huang

The botanical origin of a Chinese crude drug Ma-huang in ancient China and the origin of Japanese Ma-huang were herbologically studied. The results showed that the plants of Ephedra sinica Stapf, E. intermedia Schrenk & C. A. Meyer and E. equisetina Bunge were used as Ma-huang in China, and the first species was considered to be of high quality. The characters of Mao-zhou Ma-huang and Tong-zhou Ma-huang printed in Tu-jing-ben-cao, published in the Song Dynasty in China, were identified as E. likiangensis Florin and E. intermedia, respectively, and both species were recognized as excellent Ma-huang in the Ming Dynasty. The word origin of Katsune-kusa, the Japanese name for Ma-huang in the Heian Era, was etymologically considered as meaning the plant having reddish brown roots. In Japan, the plant of Equisetum ramosissimum Desf. var.japonicum Milde, of the family Equisetaceae, was substituted for Ma-huang in the middle of Edo Era, and it was designated that this action was based on the confusion of Ephedra plants and Equisetum plants those days in China.     

Survey on resources of Ephedra plants in Xinjiang

The resources of wild Ephedra plants in the Xinjiang Uygur Autonomous Region were surveyed. Ephedra plants mainly grow on the fringes of the Taklimakan Desert and Gureban-tonggute Desert. We found six genotypes of Ephedra przewalskii growing widely in Xinjiang. Three genotypes of Ephedra intermedia were limited to the northern and eastern parts, and Ephedra regeliana scattered in the northern part of Xinjiang. These Ephedra specimens were analyzed for DNA sequences of nuclear ribosomal DNA, internal transcribed spacers 1 and 2, chloroplastic DNA, trnL intron and trnL-trnF intergenic spacer. Intraspecific variation of the nucleotide sequence in E. przewalskii was found in different habitats. Norephedrine, ephedrine, pseudoephedrine, and methylephedrine contents of the specimens were determined. Although Ephedra intermedia of all three genotypes contained ephedrine alkaloids, ephedrine alkaloids were not detected in E. regeliana and E. przewalskii.     

Studies on cultivated ephedra plants in inner mongolia autonomous region and ningxia hui autonomous region

Progression of the desertification in northern China has been causing damage to wild Ephedra plants on which we depend for most of supply of the traditional herbal medicine, "Ma huang." The Chinese government encourages the cultivation of Ephedra plants, and Ephedra fields have been reclaimed in the original Ephedra habitats in recent years. We surveyed 7 Ephedra fields that have been recently developed in the Inner Mongolia Autonomous Region and Ningxia Hui Autonomous Region to collect information on Ephedra plant cultivation, especially pertaining to crop species. Specimens taken from those Ephedra fields were genetically and morphologically analyzed, and their ephedrine alkaloid content was examined. DNA analyses of Ephedra specimens, including DNA sequencing of ITS (internal transcribing sequence of nuclear ribosomal DNA) and trn L/F (intron of trnL and intergenic spacer between the trnL and trnF of chloroplast DNA) region and species-specific amplification of trn L/F were conducted to identify Ephedra species. Based on the results of DNA sequencing and morphological determination, the crops grown in 6 fields ware identified as Ephedra sinica, while co-planting of E. sinica and E. intermedia was found in one field where a higher appearance rate of plants with varied morphology from wild Ephedra plants was observed. Furthermore, direct sequencing of the PCR product of the trn L/F region of some specimens from the field and their species-specific PCR showed ambivalent result. Cloning and sequencing of the PCR product of the trn L/F region of those specimens DNA suggested their heteroplasmy, containing both E. sinica- and E. intermedia-type chloroplasts. On the other hand, the profile of the ephedrine alkaloid content was clearly correlated with the result of direct sequencing of the trn L/F region; the specimens showing the E. sinica-type sequence contained more ephedrine than pseudoephedrine, and the specimens of the E. intermedia-type more pseudoephedrine.     

国产麻黄的形态组织学研究——Ⅰ.北方主产的七种麻黄

据文献记载和作者的调查,我国产麻黄属(Ephedra)植物共有13种3变种1变型。本文报道我国北方生产的7种麻黄生药形态组织学比较研究结果。7种麻黄是:草麻黄Ephedra sinica Stapf、木贼麻黄E.equisetina Bunge、中麻黄E.intermedia Schrenk ex Mey.、膜果麻黄E.przewalskii Stapf,单子麻黄E.monosperma Gmel.ex Mey.、雌雄麻黄E.fedtschenkoae Pauls和细子麻黄E.reaeliana Florin。文中附有生药性状与组织构造特征比较表及生药组织图     

Survey of <Emphasis Type="Italic">Ephedra </Emphasis>resources in the Northern Areas of Pakistan and their genetic diversity

More than 400 species of medicinal plants grow in the Northern Areas of Pakistan, including Ephedra plants. To investigate the wild Ephedra plant resources in the area, we surveyed the medicinal plants and collected 71 specimens from 18 collecting sites to analyze their genetic variation. The DNA sequences of the internal transcribed spacers 1 and 2 (ITS 1 and 2) of nuclear ribosomal DNA and a noncoding sequence of chloroplast DNA (trn L/F) were analyzed. This DNA data analysis and external morphological features were used to confirm the species of the specimens, and it was found that E. intermedia was the major species in the area and that E. gerardiana and E. przewalskii were present sporadically. Although it inhabits a relatively small area in comparison with the northwestern Chinese provinces, the DNA sequence of E. intermedia in the Northern Areas of Pakistan was significantly more heterogeneous than the same species grown in those neighboring regions. Most of the E. intermedia specimens contained more than 0.7% ephedrine alkaloids, fulfilling the requirement of the Japanese Pharmacopoeia; thus, the Ephedra plants in the area are a genetic and medicinal resource of great importance.     

Environmental and soil characteristics in Ephedra habitats of Uzbekistan

Journal of Natural Medicines - In Uzbekistan, Ephedra distachya L., E. equisetina Bunge, E. foliata Boiss. ex C. A. Mey., E. lomatolepis Schrenk, and E. strobilacea Bunge show species specificity...     

国产麻黄的形态组织学研究——Ⅱ.西南主产种及其它种

本文报道我国西南产麻黄——丽江麻黄Ephedra likiangensis Florin、匍枝丽江麻黄E.likiangensis f.mairei(Florin)C.Y.Cheng、藏麻黄E.saxatilis Royle ex Florin、山岭麻黄E.gerardiana Wall、垫状山岭麻黄E.gerardiana Var.congesta C.Y.Cheng、矮麻黄E.minuta Florin和异株矮麻黄E.minuta var.dioeca C.Y.Cheng,以及形态组织特征较特殊的宁夏产斑子麻黄E.lepidosperma C.Y.Cheng、新疆产窄膜麻黄E.lomatolepis Schrenk,西藏产西藏中麻黄E.intermedia var.tibetica Stapf的生药形态组织学研究结果。并根据对国产麻黄的生药形态组织学的系统研究结果,分别编写了各种国产麻黄(包括13种3变种1变型)的生药性状和生药显微特征检索表。     

人工种植麻黄中左旋麻黄碱、右旋麻黄碱含量及相关性研究

目的　对不同产地、不同生长期人工种植麻黄生物形态和土壤特征指标进行了同期跟踪测定 ,依次探讨麻黄人工规范化种植的技术条件 ,为麻黄现代化种植 ,提高技术水平打下一定的基础。方法　采用高效液相色谱法 ,测定麻黄中左旋麻黄碱、右旋麻黄碱含量。结果　基地生长期一年半的中麻黄的总碱含量均已超过药典规定标准 ,达到收购标准 ,草麻黄亦接近收购标准。结论　植株含量测定结果表明 ,秋季移栽麻黄的麻黄碱含量较春季移栽麻黄高 ,而传统的麻黄移栽期为春季 ,本研究对指导生产具有一定的现实意义。     

HPLC-ESI-TOF-MS法优选麻黄中麻黄碱的提取工艺

目的优选麻黄中麻黄碱的最佳提取工艺。方法水煎法获得麻黄药材总提取物,反相高效液相色谱-电喷雾-飞行时间串联质谱选择离子对法测定总提物中麻黄碱的含量,正交设计优选最佳提取工艺。结果方法检测限为0.05μg·mL-1,线性范围为0.005～0.5mg·mL-1;麻黄碱最佳提取工艺为:加30倍量水,75℃加热40min,提取4次。结论该检测方法灵敏、准确,可用于复杂体系中麻黄碱的测定;水煎法经济、绿色、无污染,可作为麻黄碱等生物碱类物质的提取方法。     

近红外漫反射光谱法结合CP-ANN和PLS高通量分析草麻黄药材

目的:建立草麻黄药材的近红外漫反射光谱高通量分析方法。方法:测量草麻黄样品的近红外漫反射光谱(near infra-red diffuse reflectance spectra,NIRDRS),应用化学计量学技术进行光谱处理和数据预处理,分别建立并验证草麻黄药材的产地和采摘时间判别对向传播人工神经网络(counter-propagation artificial neural network,CP-ANN)模型及麻黄碱和伪麻黄碱含量预测偏最小二乘(partial least square,PLS)模型。结果:草麻黄药材的产地和采摘时间判别CP-ANN模型的验证样品预测准确率分别为100.0%和80.0%;麻黄碱和伪麻黄碱含量预测PLS模型的验证样品预测均方根误差(root mean square errors ofprediction,RMSEPs)小,分别为1.12和0.236,预测值与参考值的相关系数(correlation coefficients)大,分别为0.9721和0.9309。结论:采用所建方法能同时对草麻黄药材的产地和采摘时间进行准确判别,对其麻黄碱和伪麻黄碱的含量进行准确预测。该方法准确、快速,无需特殊的样品处理,也不使用化学试剂。     

中麻黄ISSR—PCR反应体系的建立和优化

目的：建立适用于中麻黄的简单重复序列区间．聚合酶链反应（ISSR—PCR）最佳反应体系。方法：以凝胶成像系统下特异谱带特征为评价指标,以Mg2＋、dNTPs、TaqDNA聚合酶、引物和DNA模板为考察因素,采用L16（4^5）正交试验优选条件;通过单因素试验对反应体系中各种影响因素进行筛选。结果：中麻黄ISSR-PCR最佳反应体系（20p1）为30ngDNA模板、0．25mmol／LdNTPs、2．0mmol／LMg2＋（10xPCRBuffer）、0．4μmol／LISSR引物、1．5UTaqDNA聚合酶;并筛选出了12条ISSR-PCR引物的最佳退火温度。结论：所建中麻黄ISSR—PCR反应体系经过10份中麻黄材料检验证明该体系稳定可靠,可用于中麻黄遗传分析。     

Dimeric proanthocyanidins from the roots of Ephedra sinica

Two new dimeric proanthocyanidins, ephedrannin B ( 2) and mahuannin E ( 4), along with two known congeners, ephedrannin A ( 1) and mahuannin D ( 3), were isolated from the roots of Ephedra sinica Stapf. Based on NMR, MS, and CD analysis, the structures of the new compounds were deduced to be 5,7,4'-trihydroxyflavan-[4(alpha)-->8,2(alpha)-->O-->7]-kaempferol ( 2) and 5,7,4'-trihydroxyflavan-[4(beta)-->8,2(beta)-->O-->7]- EPI-afzelechin ( 4). The compounds 1 - 4 were evaluated for cytotoxicity against three tumor cell lines (SGC-7901, HepG2, and HeLa), and 2 was found to be significantly active.     

RP-HPLC法测定麻黄配方颗粒中麻黄碱和伪麻黄碱的含量

目的：建立用反相高效液相色谱法测定麻黄配方颗粒中麻黄碱和伪麻黄碱含量。方法：采用BonChrom—C18柱。流动相为乙腈-0．02mol/L磷酸二氢钾-磷酸（5：95：0．1）,柱温为35℃,检测波长为207nm,流速为1．0ml／min。结果：盐酸麻黄碱线性范围0．03305-0．3966μg,r=0．9999,平均回收率为100．42％,RSD为1．78％（n=6）;盐酸伪麻黄碱的线性范围为0．023～0．276μg,r=0．9997,平均回收率为99．27％,RSD为2．05％（n=6）。结论：本法重复性好,可作为麻黄配方颗粒中麻黄碱和伪麻黄碱的定量分析方法。     

The use of polymerase chain reaction (PCR) for the identification of ephedra DNA in dietary supplements

As part of a continuing research effort to develop chemical and genetic authentication profiles of botanicals, an investigation was performed with the goal to detect, identify and verify Ephedra sinica Stapf DNA in dietary supplements such as plant mixtures and tablets/capsules. We amplified and sequenced the chloroplast psbA-trnH spacer from 21 Ephedra spp. and from two of their closest relatives, Gnetum gnemon L. and Welwitschia mirabilis Hook. Based on sequence comparisons, we identified regions unique to all of the Ephedra spp. samples analyzed. We concluded that the psbA-trnH spacer sequence could be used as a molecular marker. Based on this spacer sequence, we designed Ephedra spp.-specific primers that can help to identify Ephedra spp. DNA in plant mixtures containing as little as 1/1,000 part of Ephedra spp. tissue. We used a DNA extraction method that allows for quick DNA isolation from plant mixtures for PCR analysis.     

Survey of crude drugs effective in eliminating superoxides in blood plasma of mice

The effect of crude drugs on superoxide dismutase activity in the blood plasma of mice was examined. The ethanol extract of each crude drug was intraperitoneally administered. Ephedrae herba (Ephedra sinica Stapf, Ephedraceae), Nelumbinis folium (Nelumbo nucifera Gaertner, Nymphaeaceae) and Scutellariae radix (Scutellaria baicalensis Georgi, Labiatae) were significantly and dose dependently effective in eliminating superoxides in the plasma. Most of the extracts (except 2000 mg/kg of Scutellariae radix) showed no activity after oral administration.