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1.
Identification of intermediates and enzymes involved in the early steps of artemisinin biosynthesis in Artemisia annua 总被引:4,自引:0,他引:4
Bertea CM Freije JR van der Woude H Verstappen FW Perk L Marquez V De Kraker JW Posthumus MA Jansen BJ de Groot A Franssen MC Bouwmeester HJ 《Planta medica》2005,71(1):40-47
An important group of antimalarial drugs consists of the endoperoxide sesquiterpene lactone artemisinin and its derivatives. Only little is known about the biosynthesis of artemisinin in Artemisia annua L., particularly about the early enzymatic steps between amorpha-4,11-diene and dihydroartemisinic acid. Analyses of the terpenoids from A. annua leaves and gland secretory cells revealed the presence of the oxygenated amorpha-4,11-diene derivatives artemisinic alcohol, dihydroartemisinic alcohol, artemisinic aldehyde, dihydroartemisinic aldehyde and dihydroartemisinic acid. We also demonstrated the presence of a number of biosynthetic enzymes such as the amorpha-4,11-diene synthase and the--so far unknown--amorpha-4,11-diene hydroxylase as well as artemisinic alcohol and dihydroartemisinic aldehyde dehydrogenase activities in both leaves and glandular trichomes. From these results, we hypothesise that the early steps in artemisinin biosynthesis involve amorpha-4,11-diene hydroxylation to artemisinic alcohol, followed by oxidation to artemisinic aldehyde, reduction of the C11-C13 double bond to dihydroartemisinic aldehyde and oxidation to dihydroartemisinic acid. 相似文献
2.
The time course of the levels of artemisinin, its biosynthetic precursors and the biosynthetically related sesquiterpenes was monitored during a vegetation period of Artemisia annua plants of different geographical origin. Considerable differences in contents of artemisinin and its direct precursors artemisinic acid and dihydroartemisinic acid were found between these A. annua's. For the first time the A. annua plants of different geographical origin were found to belong to different chemotypes. A chemotype with a high artemisinin level was found to have also a high dihydroartemisinic acid level but a relatively low artemisinic acid level. Reversibly, a chemotype with low levels of artemisinin and dihydroartemisinic acid contained a high artemisinic acid level. Artemisinic acid is considered to be the direct precursor of dihydroartemisinic acid in the biosynthetic pathway of artemisinin. The observed accumulation of artemisinic acid in one of the A. annua chemotypes may indicate the presence of a rate-limiting step in the biosynthetic pathway of artemisinin. The enzymatic reduction of artemisinic acid into dihydroartemisinic acid is probably a "bottle neck" in the biosynthetic pathway of artemisinin in varieties with high artemisinic acid and consequentially low artemisinin levels. After a night-frost period, the level of artemisinin was increased, in the Vietnamese A. annua plants, while the dihydroartemisinic acid level was decreased. This phenomenon is in accordance with our hypothesis that stress triggers the conversion of dihydroartemisinic acid to artemisinin. It is suggested that the presence of high levels of dihydroartemisinic acid may be an adaptation to stress conditions (e.g., night-frost), during which relatively high levels of 1O2 are formed. Dihydroartemisinic acid gives the plant protection by reacting with these reactive oxygen species yielding artemisinin as stable end-product. 相似文献
3.
Artemisinin is a sesquiterpene lactone endoperoxide and an important antimalarial drug produced in Artemisia annua. To unravel the diverse processes determining artemisinin yield in A. annua crops, artemisinin accumulation during the development of individual leaves was studied in two field experiments. During the life cycle of a leaf, artemisinin was always present. Quantities were low at leaf appearance and increased steadily. In leaves studied until after senescence, maximum quantities and concentrations were achieved after the leaf had turned brown. The total quantity of possible artemisinin precursors per leaf (dihydroartemisinic acid and other upstream precursors) was highest early in the leaf cycle when the leaf was still expanding. Dihydroartemisinic acid was more abundant than the other compounds and its quantity declined during leaf development whereas that of artemisinin increased. Dihydroartemisinic acid was not converted directly into artemisinin, because on a per leaf basis the decline in molar quantity of precursors in the earliest formed leaves was not compensated for by a simultaneous increase in artemisinin. Our results suggest that a (putative) intermediate such as dihydroartemisinic acid hydroperoxide temporarily may have accumulated in considerable quantities. The number of mature, capitate trichomes on the adaxial leaf side increased after leaf appearance until the end of leaf expansion, and then decreased, probably due to collapse of trichomes. Artemisinin production thus (also) occurred when trichomes were collapsing. Later formed leaves achieved higher concentrations of artemisinin than earlier formed leaves, because of a higher trichome density and a higher capacity per trichome. 相似文献
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Seeds of ARTEMISIA ANNUA L. originating from a wild population in Japan were cultivated in the garden planters of our Institute in 1995, and the seasonal and positional variations in content of artemisinin, artemisinic acid, arteannuin B and artemisitene, were measured. The content of artemisinic acid in leaves was high [max. 1.0% (dry wt basis)], as compared with other cultivars previously reported. This is the first report on a Japanese strain of ARTEMISIA ANNUA. 相似文献
6.
Woerdenbag HJ Pras N Chan NG Bang BT Bos R van Uden W Van Y P Van Boi N Batterman S Lugt CB 《Planta medica》1994,60(3):272-275
The active principle of ARTEMISIA ANNUA L., artemisinin, is currently being developed to a registered antimalarial drug. For production purposes, plants with a high artemisinin content are required. We followed the development of the artemisinin content and of the biosynthetically related sesquiterpenes artemisinic acid, arteannuin B, and artemisitene in A. ANNUA plants, during a vegetation period in Vietnam, where this species is indigenous. In addition, the essential oil content and composition were studied. Samples of leaves, buds, flowers, or post-bloom flowers and fruits were taken at different stages: vegetative (5, 6, and 8 months old), at mass formation of buds (9 months), at full bloom (10 months), and post-bloom (10S months). The highest artemisinin content (0.86% dry wt) was present in the leaves of 5 months-old plants. At this stage also the highest leaf yield was found. Subsequently, the artemisinin content gradually dropped. At the age of 5 months the highest artemisinic acid and arteannuin B contents, 0.16 and 0.08% dry wt, respectively, were found as well. Artemisitene was present at all stages of development, ranging from 0.002 to 0.09% dry wt. With 1.9% v/w, the essential oil content was maximal just before flowering and was composed of 55% monoterpenes and 45% sesquiterpenes. At all other stages (0.4 - 1.0% v/w oil) this ratio was ca. 30%/70%. The main components of the oil were camphor and germacrene-D. 相似文献
7.
HPLC-UV-ELSD法同时测定青蒿中青蒿素、青蒿乙素和青蒿酸的含量 总被引:4,自引:0,他引:4
用HPLC-UV-ELSD法同时测定青蒿药材中青蒿素、青蒿乙素和青蒿酸的含量。采用Nucleodur C18色谱柱(250 mm×4.6 mm,5 μm ID);以乙腈-0.1%乙酸水(50∶50)为流动相;紫外检测波长209 nm,蒸发光散射检测器漂移管温度50 ℃。结果显示,青蒿素、青蒿乙素和青蒿酸能够达到很好分离。它们的线性范围分别为0.52~2.6 μg, r=0.999 4(n=5); 0.022~4.4 μg, r=0.999 9(n=5); 0.203~8.12 μg,r=0.999 8(n=5)。平均回收率分别为99.45%(RSD=2.3%, n=6);102.37%(RSD=1.7%, n=6);101.10%(RSD=0.79%, n=6)。本法简单、准确、快速,可同时测定青蒿药材中青蒿素、青蒿乙素和青蒿酸的含量。 相似文献
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A plant of Artemisia annua containing high amounts of artemisinic acid (0.8%) was isolated. the combined concentration of the sesquiterpene artemisinin, and its intermediates artemisinic acid and artemisinin-b in this plant was 1.35%. the plant has been cloned by micropropagation in tissue culture. 相似文献
9.
Seasonal variations of Artemisinin and its biosynthetic precursors in tetraploid Artemisia annua plants compared with the diploid wild-type 总被引:2,自引:0,他引:2
Using colchicine we induced tetraploidy in Artemisia annua L. plants. During a vegetation period we monitored the time course of the levels of artemisinin, its direct precursors, the biosynthetically related sesquiterpenes and the essential oil content in the diploid (wild-type) and tetraploid A. annua plants. The averaged artemisinin level in tetraploids was 38% higher than that of the wild-type as measured over the whole vegetation period. In contrast, the averaged essential oil content of the tetraploids over this period was 32% lower. This might suggest a reciprocal correlation between artemisinin (sesquiterpenes) and the essential oil content (monoterpenes). The averaged biomass of the leaves of the tetraploid plants was lower compared to the wild-type plants. Therefore, the artemisinin yield per m2 tetraploids was decreased by 25%. Although the tetraploid plants were smaller than the wild-type plants, certain individual organs like the leaves were considerably larger, and seeds obtained by cross pollination between tetraploid A. annua plants had a spectacular size. In principle, tetraploid A. annua can be a useful starting material for a breeding program in order to obtain larger and faster growing plants, which produce higher levels of artemisinin. 相似文献
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(+)-Deoxoartelinic acid (13), a new hydrolytically stable, water-soluble, and potent non-acetal-type antimalarial drug candidate, was successfully prepared from artemisinic acid by using sulfur ylide and photooxygenative cyclization in seven steps. This compound showed superior in vitro antimalarial activity against the chloroquine-resistant K1 strain of Plasmodium falciparum and higher suppression (98.7%) than arteether in vivo against Plasmodium chabaudi infected mice. (+)-Deoxoartelinic acid also showed remarkable stability with a half-life of 258.66 h, 23 times more stable than clinically useful arteether in simulated stomach acid, and improved solubility, 4 times more soluble than artemisinin in water. 相似文献
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M Jung X Li D A Bustos H N elSohly J D McChesney W K Milhous 《Journal of medicinal chemistry》1990,33(5):1516-1518
(+)-Deoxoartemisinin (2), a new and more active antimalarial agent, was successfully prepared from artemisinin in one step using NaBH4 and BF3.Et2O in THF. (-)-Deoxodeoxyartemisinin (5), a potential metabolite of deoxoartemisinin, was also prepared either from 2 or from artemisinic acid. 2 shows 8-fold increased antimalarial activity in vitro against chloroquine-resistant malaria as compared to artemisinin (1). Compound 2 possesses superior in vivo antimalarial activity to 1. 相似文献
13.
Xiaoxin X. Zhu Lan Yang Yujie J. Li Dong Zhang Ying Chen Petra Kostecká Eva Kmoníèková Zdenìk Zídek 《Pharmacological reports : PR》2013,65(2):410-420
BackgroundIn addition to recognized antimalarial effects, Artemisia annua L. (Qinghao) possesses anticancer properties. The underlying mechanisms of this activity are unknown. The aim of our experiments was to investigate the effects of distinct types of compounds isolated from A. annua on the immune-activated production of major mediators of angiogenesis playing a crucial role in growth of tumors and formation of metastasis.MethodsIncluded in the study were the sesquiterpene lactones artemisinin and its biogenetic precursors arteannuin B and artemisinic acid. The semi-synthetic analogue dihydroartemisinin was used for comparative purposes. The flavonoids were represented by casticin and chrysosplenol D, the coumarin type of compounds by 4-methylesculetin. Their effects on the lipopolysaccharide (LPS)-induced in vitro production of nitric oxide (NO) were analyzed in rat peritoneal cells using Griess reagent. The LPS-activated production of prostaglandin E2 (PGE2) and cytokines (VEGF, IL-1β, IL-6 and TNF-α) was determined in both rat peritoneal cells and human peripheral blood mononuclear cells using ELISA.ResultsAll sesquiterpenes (artemisinin, dihydroartemisinin, artemisinic acid, arteannuin B) significantly reduced production of PGE2. Arteannuin B also inhibited production of NO and secretion of cytokines. All NO, PGE2 and cytokines were suppressed by flavonoids casticin and chrysosplenol D. The coumarin derivative, 4-methylesculetin, was ineffective to change the production of any of these factors.ConclusionsThe inhibition of immune mediators of angiogenesis by sesquiterpene lactones and flavonoids may be one of the mechanisms of anticancer activity of Artemisia annua L. 相似文献
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从中国东北部产黄花蒿中分离得到主成分青蒿酸(1).通过分析青蒿酸对溴苯酰甲酯的X-衍射结晶学数据,确定了青蒿酸的绝对构型.另外,从同植物中还分离得到少量青蒿素(2),以及青蒿乙素(3),谷甾醇和豆甾醇. 相似文献
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HPLC-UV-ELSD联用测定黄花蒿叶片中青蒿素及相关倍半萜的含量 总被引:2,自引:0,他引:2
目的HPLC-UV-ELSD联用建立黄花蒿叶片中青蒿素及其生合成相关倍半萜的含量测定方法。方法采用Venusil XBP-C18色谱柱(200 mm×4.6 mm,5μm),流动相:乙腈-水梯度洗脱,检测波长:203 nm,流速:1.0 mL.min-1,ELSD载气压力:0.35 MPa,喷嘴温度:35℃,漂移管温度:60℃,检测器:UV-ELSD联用,测定黄花蒿叶片中青蒿素、青蒿素B、3α-羟基-1-去氧青蒿素含量。同一色谱柱条件下流动相改为乙腈-水-甲酸(体积比为60.00∶40.00∶0.08),检测波长:215 nm,流速:1.0 mL.min-1,检测器:UV检测器,测定黄花蒿叶片中青蒿酸含量。结果4种倍半萜在各自的线性范围内线性关系良好(r≥0.9990),RSD<3%,平均加样回收率分别为98.5%(RSD=2.3%)、96.5%(RSD=1.8%)、97.5%(RSD=3.6%)、95.4%(RSD=2.5%)。结论该方法可作为黄花蒿中青蒿素及其生合成相关倍半萜的含量测定方法。 相似文献
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Podophyllotoxin is a pharmaceutical compound found in leaves and rhizomes of American mayapple (P. peltatum L.), a species being investigated as an alternative to that of the Indian mayapple (P. emodi). Leaves alone can serve as a renewable source of podophyllotoxin (and other lignans) leaving rhizomes undisturbed to produce leaf biomass in subsequent years. It is not known, however, how frequently or severely plants can be defoliated without adversely affecting future plant growth, lignan content, or podophyllotoxin yield (g.m(-2)). This study compared harvest strategies that were mild to severe in frequency and timing of leaf removal. A wild population in full sun was subjected to leaf removal treatments of varying frequency (every year, every 2nd or 3rd year) and timing (early or late). Control plots not previously harvested were included every year. Plots were 1.0 m2 and established during spring of 2001. Duration of the study was four years. P. peltatum plants did not tolerate the most severe harvest treatment: annual harvest frequency in combination with early harvest time. Early annual harvests reduced total leaf dry mass and total leaf area in a consistent and linear manner. In contrast, plants tolerated annual harvests when conducted late in the growing season and tolerated early harvests when conducted every 2nd or 3rd year. The number of sexual shoots was reduced to zero by early annual harvests. Podophyllotoxin content was 2.7 to 6.5 times greater in leaves harvested early compared to those harvested late, though content was significantly greater in only two out of four years. In conclusion, we can recommend leaf removal every year from well-established P. peltatum populations grown in full sun if harvests are conducted late in the growing season. This harvest strategy ensures maximum podophyllotoxin yield without jeopardizing future leaf biomass yield. Leaves harvested early appear to have greater podophyllotoxin content, but we discourage early harvest every year. Instead, our results indicate that leaves can be harvested early every other year without reducing long-term performance of P. peltatum populations. 相似文献
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目的:研究低温处理对青蒿素及其他萜类合成通路的影响。方法:以4℃为胁迫条件,利用高效液相色谱法测定青蒿素含量;硫酸钛沉淀法和N,N-二甲基-对-亚硝基苯胺漂白测定法分别检测黄花蒿叶片过氧化氢(H2O2)和单线态氧(1O2)含量;紫外分光光度法检测过氧化氢酶(CAT)活性;实时荧光定量PCR技术定量分析青蒿素合成途径及竞争途径关键酶基因的表达。结果:4℃处理4h后黄花蒿叶片中1O2和H2O2含量升高,并伴随青蒿素含量和CAT活性逐步提高,4、24和48h后青蒿素含量分别提高20%、65%和80%;4℃处理24h后,青蒿素合成相关基因[3-羟基-3-甲基戊二酰辅酶A还原酶基因(HMGR)、法呢基焦磷酸合酶基因(FPS)、紫穗槐二烯合酶基因(ADS)、细胞色素P450单加氧酶基因(CYP71AV1)、细胞色素P450氧化还原酶基因(CPR)、青蒿醛Δ11(13)双键还原酶基因(DBR2)]的表达普遍上调,其中ADS的表达提高16倍;而青蒿素合成竞争途径的β-石竹烯合酶基因(CS)表达则下调近20倍。结论:低温刺激可能通过产生高浓度活性氧(ROS)促进青蒿素合成前体转化、上调青蒿素合成相关基因表达并抑制竞争途径基因表达等途径促进青蒿素合成。 相似文献
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Agrobacterium rhizogenes-mediated transformation of Artemisia annua: production of transgenic plants
Transgenic hairy roots were induced in the leaves of Artemisia annua by treatment with the LBA 9402 strain of Agrobacterium rhizogenes. The axenic hairy root cultures were found to produce the sesquiterpenes artemisinic acid and arteannuin B. The hairy root cultures were observed to spontaneously regenerate into plantlets on solid hormone-free MS medium. The regenerated plants had phenotypic characteristics typical to the transformed plants. Among the plants of the age of one month in culture, the transgenic plant was bigger (2.643 g/plant) than the normal (0.856 g/plant). Both these kinds of in vitro plants carried sesquiterpenes-artemisinic acid and arteannuin B. 相似文献