青蒿倍半萜合酶(环化酶)研究进展

青蒿素是从中药青蒿中分离得到的抗疟有效单体,是含有过氧基团的新型倍半萜内酯化合物,是目前世界上最有效的疟疾治疗药物。青蒿素的生物合成途径属于类异戊二烯代谢途径中的倍半萜类分支途径,倍半萜合酶是该途径的关键酶之一,目前已从青蒿中克隆了多个倍半萜合酶基因。综述了青蒿中已克隆的几种倍半萜合酶基因的研究进展。     

Simple and rapid physico-chemical methods to examine action of antimalarial drugs with hemin: its application to Artemisia annua constituents

Malaria is a major health problem in many countries and according to an estimate of the WHO, more than 500 million infections occur per year. Artemisinin, a sesquiterpene from Artemisia annua L., has received considerable attention as a promising and potent antimalarial drug for its stage speciticity, its rather low toxicity, effectiveness against drug-resistant Plasmodium species and activity against cerebral malaria. From recent studies it seems that hemin is primarily involved in the antimalarial activity of the constituents of Artemisia annua L. Thus, the interaction of a compound with hemin may represent a crucial screening test to define its efficacy. In this study the interaction between artemisinin and hemin was investigated by UltraViolet/Visible (UV/Vis) spectrophotometry and High Performance Liquid Chromatography/Diode Array Detector/Mass Spectrometry (HPLC/DAD/MS). In addition, some flavonols isolated from Artemisia annua L. were also tested to investigate their possible role in the interaction between artemisinin and hemin. These two simple physico-chemical methods can be useful as rapid and widespread screening methods for the search of other alkylating antimalarial constituents from natural sources or for the evaluation of the activity of semisynthetic analogues of artemisinin.     

Microbial transformation of antimalarial terpenoids

The fungal and bacterial transformation of terpenoids derived from plant essential oils, especially the sesquiterpenoid artemisinin from Artemisia annua, has produced several new candidate drugs for the treatment of malaria. Obtaining new derivatives of terpenoids, including artemisinin derivatives with increased antimalarial activity, is an important goal of research in microbial biotechnology and medicinal chemistry.     

A cDNA clone for beta-caryophyllene synthase from Artemisia annua

An homology-based cloning strategy yielded a full-length cDNA from Artemisia annua that encoded a protein of 60.3 kDa which resembled a sesquiterpene synthase in sequence. Heterologous expression of the gene in Escherichia coli provided a soluble recombinant enzyme capable of catalyzing the divalent metal ion-dependent conversion of farnesyl diphosphate to beta-caryophyllene, a sesquiterpene olefin found in the essential oil of A. annua. In reaction parameters and kinetic properties, beta-caryophyllene synthase resembles other sesquiterpene synthases of angiosperms. The beta-caryophyllene synthase gene is expressed in most plant tissues during early development, and is induced in mature tissue in response to fungal elicitor thus suggesting a role for beta-caryophyllene in plant defense.     

Anti-Inflammatory Activities of Monoterpene and Sesquiterpene Glycosides from the Aqueous Extract of Artemisia annua L.

Artemisia annua L. is a Chinese medicinal herb, but the origin of its pharmacological properties, including its anti-inflammatory activity, remain unknown. In this study, five new monoterpene glycosides ( 1 – 5 ) and two new sesquiterpene glycosides ( 6 and 7 ) were isolated from the aqueous extract of the aerial parts of A. annua. The structures of these glycosides were determined using high-resolution electrospray ionization mass spectrometry, nuclear magnetic resonance spectroscopy, electronic circular dichroism calculations, and chemical hydrolysis methods. The anti-inflammatory activities of the isolated compounds were evaluated by down-regulating interleukin-6 (IL-6) in lipopolysaccharide-stimulated RAW 264.7 macrophages. Notably, all the new compounds significantly inhibited the expression of IL-6 in a dose-dependent manner.     

无载体固定化黄花蒿细胞生产青蒿素

疟疾是一种严重危害人类健康的流行病,主要由疟原虫经蚊虫叮咬引起。目前,在临床上疟原虫对治疗疟疾的药物(如氯奎等)有较强的耐药性,并表现出明显的交叉耐药性。来自黄花蒿的青蒿素具有极其明显的抗疟活性,成为临床首选的药物,因此青蒿素的获取成为关键。本研究采用无载体固定化法培养黄花蒿生产青蒿素,初步研究了无载体固定化细胞的生长特性。检测发现,利用该方法生产的青蒿素是常规细胞培养法的9倍,因此该方法有望成为青蒿素生产的首选方法。     

中药青蒿的生态生理及其综合利用

中药青蒿即黄花蒿(Artemisia annua L.)是抗疟药的原料,青蒿素是其有效抗疟成分。本文对青蒿的生物学特性、资源分布、生长栽培和生理生态进行了分析,指出了提高青蒿素含量的可能途径及其综合利用的前景。     

五种新疆植物的抗真菌活性

对采自新疆的黄花蒿(Artemisia annua)、北艾(Artemisia vulgaris)、梭梭(Halaxylon ammodendron)、盐爪爪(Kalidium foliatum)和多枝柽柳(Tamarix ramosissima)抗植物病原真菌的活性进行了研究,植物病原真菌包括番茄灰霉病菌(Botrytis cinerea)、棉花枯萎病菌(Fusarium axysporum f.sp.vasinfectum)、稻瘟病菌(Magnaporthe grisea)、烟草黑胫病菌(Phytophthora parasitica var.nicotianae)和瓜果腐霉(Pythium aphani-dermatum),其中黄花蒿对真菌菌丝生长、多枝柽柳对稻瘟病菌孢子萌发表现出强的抑制活性。本研究为植物病害防治和新疆植物资源的开发和利用提供了依据。     

栽培青蒿中总黄酮提取工艺

利用超声波辅助技术,获得最大限度提取青蒿中总黄酮的新工艺。用正交设计理论,结合分光光度法,优化超声波辅助醇提法提取青蒿总黄酮工艺中的关键技术参数。最佳提取．工艺为：超声波频率59kHz,乙醇体积分数60％,提取时间40min,料液比1：40。超声波辅助提取法能够实现青篙中总黄酮的高效提取,产率达1．497％。     

种植青蒿与野生青蒿精油成分对比分析

用水蒸气蒸馏法和GC-MS法对来源于重庆云阳的栽培青蒿和野生青蒿精油的化学成分进行分析.从冬季和春季栽培的青蒿精油中分别鉴定出29和28个化合物,野生青蒿精油中鉴定出21个化合物.3种精油的化学成分差别明显,两种栽培青蒿精油有16个化合物相同.而栽培青蒿与野生青蒿精油仅有6个化合物相同.这表明基因型对青蒿精油的组成起决定作用;而气候因素对各成分的相对含量有重要影响,尤其对蒿酮和樟脑的积累影响较大.本文对进一步研究青蒿精油中萜类化合物之间的生源关系具有一定的参考价值.     

黄花蒿化感作用机理的初步研究

运用室内培养皿生物测定方法,对黄花蒿克生作用进行的研究结果表明,黄花蒿水浸提液对小麦幼苗生长有明显的抑制作用,且抑制强度随浸提液浓度升高而加强。其对小麦根的抑制作用比对叶的抑制作用强。对受体小麦根尖压片的观察统计表明,黄花蒿的水浸提液影响了小麦根尖分生区细胞有丝分裂的正常进行,并随着供体黄花蒿水浸提液浓度的升高,小麦根尖分生区分裂期细胞数目下降;受体材料小麦的一些生理生化测定指标表明,黄花蒿水浸提液使供试小麦根系活力稍有减弱,可溶性蛋白含量明显升高,使根的核酸含量升高而叶的核酸含量降低;使叶绿素含量有较明显降低。     

Effects of Overexpression of the Endogenous Farnesyl Diphosphate Synthase on the Artemisinin Content in Artemisia annua L.

Artemisinin is a novel effective antimalarial drug extracted from the medicinal plant Artemisia annua L. Owing to the tight market and low yield of artemisinin, there is great interest in enhancing the production of artemisinin. In the present study, farnesyi diphosphate synthase （FPS） was overexpressed in high-yield A. annua to Increase the artemisinin content. The FPS activity in transgenic A. ennue was twoto threefold greater than that In non-transgenic A. annua. The highest artemisinin content in transgenic A. annua was approximately 0.9% （dry weight）, which was 34.4% higher than that in non-transgenic A. annua. The results demonstrate the regulatory role of FPS in artemisinin biosynthesis.     

青蒿过氧化物酶的克隆及其在体外对青蒿素生物合成的影响

用RACE方法从青蒿(Artemisia annua L.)高产株系001中克隆了一个过氧化物酶.将此基因在大肠杆菌BL21(DE3)pLysS细胞中进行原核表达得到重组蛋白(APOD1),表达的蛋白分别以抗坏血酸、愈创木酚为底物进行过氧化反应,结果显示,APOD1催化愈创木酚的活力是抗坏血酸的1.8倍左右,由此表明,克隆的APOD1类属于植物经典过氧化物酶(第三大类过氧化物酶).经与其他植物过氧化物酶同源性比较分析,推测APOD1的氨基酸序列与白羽扇豆(Lupinus albus)、辣根菜(Armoracia rusticana)、小麦(Triticum aestivum)、烟草(Nicotiana tabacum)和蕃茄(Lycopersicon esculentum)的一致性分别为42.0%、36.2%、38.9%、33.6%和32.8%.Northern杂交分析表明,此基因在青蒿的根、茎和叶中均有表达.加入APOD1至青蒿细胞提取液有利于青蒿酸向青蒿素的生物转化,但APOD1并不能直接以青蒿酸作为氧化底物.     

Deoxyarteannuin B, dihydro-deoxyarteannuin B and trans-5-hydroxy-2-isopropenyl-5-methylhex-3-en-1-ol from Artemisia anuua.

The amorphane sesquiterpenes, deoxyarteannuin B and dihydro-deoxyarteannuin B, were isolated from Artemisia annua and their structures mainly determined by two-dimensional NMR spectoscopic analyses. The irregular monoterpene, trans-5-hydroxy-2-isopropenyl-5-methylhex-3-en-1-ol, was also characterized in the same way, and its structure was confirmed by synthesis from lavandulol. All of these natural products are suggested to be formed by autoxidation reactions. Full assignments of the 1H and 13C resonances for the known natural products epi-deoxyarteannuin B and isoannulide, determined by the same methodology, are also reported.     

青蒿素对蔬菜种子发芽和幼苗生长的化感效应

试验以菜豆、豇豆、大白菜和小白菜为对象,用不同浓度的青蒿素浸种,研究了黄花蒿产生的化感物质??青蒿素对蔬菜种子发芽及幼苗生长的影响。结果表明,青蒿素对蔬菜种子发芽和幼苗生长的化感作用表现出浓度效应和品种差异,即浓度越高,抑制作用愈强,尤以豇豆种子发芽率和小白菜生长的表现最为明显,前者的发芽率可降低75.00%,后者的苗高降幅高达88.37%,且胚根停止生长。青蒿素抑制同季和后季作物的种子发芽和幼苗生长,有利于扩大黄花蒿的生存空间,增强生存竞争优势。在黄花蒿?蔬菜种植体系中,选择抗化感作用较强的大白菜和菜豆可提高土地利用率和整体生产水平。用青蒿素浸种后,蔬菜幼苗的根系活力降低,菜豆和豇豆叶绿素含量提高,而大、小白菜降低,均可视为妨碍生长的生理原因。此外,青蒿素浸种还提高蔬菜种子可溶性糖和游离氨基酸含量,推测青蒿素对种子水解酶活性的影响较小,但抑制合成酶催化的生化反应,导致代谢紊乱,抑制幼苗生长。     

14种菊科植物提取物对菜青虫的杀虫活性

用胜红蓟、黄花蒿、青蒿、艾蒿、鬼针草、三叶鬼针草、鱼眼草、小鱼眼草、紫茎泽兰、千里光、希签草、万寿菊、肿柄菊、苍耳等１４种菊科植物的石油醚、氯仿、丙酮、乙醇、甲醇和水提取物对菜青虫５龄幼虫的生测结果表明,黄花蒿、青蒿、艾蒿、鱼眼草、小鱼眼草、紫茎泽兰、万寿菊等７种菊科植物甲醇和乙醇提取物具有较高的杀虫活性。此结果为进一步从菊科植物中提取,分离和筛选高活性杀虫活性成分奠定了基础。     

采用薄层层析─紫外分光光度法测定广西不同产地黄花蒿中青蒿素含量

采用薄层层析─紫外分光光度法测定不同地区黄花蒿中的青蒿素含量,并对照测定野生与人工栽培样品,结果人工栽培的黄花蒿中青蒿素含量较野生高。     

Structural and functional characterization of HMG-COA reductase from Artemisia annua

Plants synthesize a great variety of isoprenoid products that are required not only for normal growth and development but also for their adaptive responses to environmental challenges. However, despite the remarkable diversity in the structure and function of plant isoprenoids, they all originate from a single metabolic precursor, mevalonic acid. The synthesis of mevalonic acid is catalysed by the enzyme, 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG- CoA reductase). The analysis of the amino acid sequence of HMG-CoA reductase from Artemisia annua L. plant showed that it belongs to class I HMG-CoA reductase family. The three dimensional structure of HMG-CoA reductase of Artemisia annua has been generated from amino acid sequence using homology modelling with backbone structure of human HMG-CoA reductase as template. The model was generated using the SWISS MODEL SERVER. The generated 3-D structure of HMG-CoA reductase was evaluated at various web interfaced servers to checks the stereo interfaced quality of the structure in terms of bonds, bond angles, dihedral angles and non-bonded atom-atom distances, structural as well as functional domains etc. The generated model was visualized using the RASMOL. Structural analysis of HMG-CoA reductase from Artemisia annua L. plant hypothesize that the N and C-terminals are positioned in cytosol by the two membrane spanning helices and the C-terminals domain shows similarity to the human HMG-CoA reductase enzyme indicating that they both had potential catalytic similarities.     

北京市延庆采砂迹地型湿地植被群落分类与演替分析

采用TWISPAN分析方法,对北京市延庆采矿迹地湿地40个样地51种植物进行了分类排序,将群落类型划分为7个类型,即茵陈蒿-狗尾草-黄花蒿群落、野艾蒿-茵陈蒿-尖叶胡枝子-狗尾草群落、野艾蒿-茵陈蒿-狗尾草-野大豆-苣荬菜群落、狗尾草-苍耳-黄香草木犀-虎尾草群落、狼拔草-野艾蒿-旋覆花-大籽蒿-芦苇群落、香蒲-芦苇-茭白-野大豆-鬼针草群落和竹节灯心草-褐穗莎草-稗群落;同时运用双变量主坐标分析(DPCoA)对51种植物进行了排序分析,研究结果显示研究区存在两种演替规律,即由喜湿旱生耐淹水植物演替为喜旱旱生植物和由湿地挺水植物向湿生植物演替两种类型.对于采砂基地型湿地植被恢复不仅恢复土壤营养条件,调控湿地水分条件也是十分关键的.研究结论将为采砂迹地型湿地恢复提供科学支持.