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.     

青蒿内生真菌的抗肿瘤抗氧化活性

采用MTT法和DPPH法,分别测定已分离得到的68株贵州青蒿内生真菌乙酸乙酯粗提物的肿瘤细胞生长抑制率和DPPH自由基清除率。试验共筛选获得12株活性内生真菌,根据其形态特征进行鉴定,分别隶属于子囊菌亚门(Ascomycotina)的链格孢属(Alternaria)、刺盘孢属(Colletotrichum)、拟盘多毛孢属(Pestalotiopsis)和拟茎点霉属(Phomopsis);其中,有8株内生真菌至少对1种指示瘤株具有细胞毒活性,占总菌株数的11.8%;5株内生真菌具有不同程度的清除DPPH自由基活性,占总菌株数的7.4%;1株内生真菌同时具有细胞毒活性和抗氧化活性。     

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.     

一个新高产青蒿倍半萜合酶基因的克隆、表达和分析

用RACE方法从青蒿（Artemisia annua L.)高产株系001中克隆了一个新的1886bp的全长倍半萜合酶cDNA。克隆的倍半萜合酶氨基酸序列与烟草马兜铃烯合酶,莨菪岩兰螺旋二烯合酶,棉花杜松烯合酶的一致性分别为39％,38％和41％;与青蒿柏木脑合酶,紫穗槐二烯合酶和一个推测的倍半萜合酶克隆cASC125的一致性为50％,48％和59％。cDNA编码区序列被克隆进原核表达载体pET-30a,并在大肠杆菌（Escherichia coli)BL21(DE3)中诱导表达,但过量表达的蛋白主要是以不溶性蛋白形式存在。Northern blotting分析表明此基因在茎,叶,花中表达,在根中没有表达。     

一个新高产青蒿倍半萜合酶基因的克隆、表达和分析

用RACE方法从青蒿(Artemisia annua L.)高产株系001中克隆了一个新的1 886 bp的全长倍半萜合酶cDNA.克隆的倍半萜合酶氨基酸序列与烟草马兜铃烯合酶、莨菪岩兰螺旋二烯合酶、棉花杜松烯合酶的一致性分别为39%、38%和41%;与青蒿柏木脑合酶、紫穗槐二烯合酶和一个推测的倍半萜合酶克隆cASC125的一致性为50%、48%和59%.cDNA编码区序列被克隆进原核表达载体pET-30a,并在大肠杆菌(Escherichia coli)BL21(DE3)中诱导表达,但过量表达的蛋白主要是以不溶性蛋白形式存在.Northern blotting分析表明此基因在茎、叶和花中表达,在根中没有表达.     

Chemical Composition and Antipathogenic Activity of Artemisia annua Essential Oil from Romania

The essential oil extracted by hydrodistillation from Romanian Artemisia annua aerial parts was characterized by GC/MS analysis, which allowed the identification of 94.64% of the total oil composition. The main components were camphor (17.74%), α‐pinene (9.66%), germacrene D (7.55%), 1,8‐cineole (7.24%), trans‐β‐caryophyllene (7.02%), and artemisia ketone (6.26%). The antimicrobial activity of this essential oil was evaluated by determining the following parameters: minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC), minimal fungicidal concentration (MFC), and minimal biofilm eradication concentration (MBEC). Moreover, the soluble virulence factors were quantified with different biochemical substrates incorporated in the culture media. The reference and resistant, clinical strains proved to be susceptible to the A. annua oil, with MICs ranging from 0.51 to 16.33 mg/ml. The tested essential oil also showed good antibiofilm activity, inhibiting both the initial stage of the microbial cell adhesion to the inert substratum and the preformed mature biofilm. When used at subinhibitory concentrations, the essential oil proved to inhibit the phenotypic expression of five soluble virulence factors (hemolysins, gelatinase, DNase, lipases, and lecithinases). Briefly, the present results showed that the A. annua essential oil contained antimicrobial compounds with selective activity on Gram‐positive and Gram‐negative bacterial strains as well as on yeast strains and which also interfere with the expression of cell‐associated and soluble virulence factors.     

Antioxidant,Anti-Skin-Aging,Anti-Inflammatory,and Anti-Acetylcholinesterase Activities of Rourea oligophlebia Extracts

The objective of this study was to evaluate the antioxidant, anti-skin-aging, anti-inflammatory, and anti-acetylcholinesterase activities of the hexane (n-hex), AcOEt, BuOH, MeOH, and aqueous extracts from R. oligophlebia roots. The total phenolic and flavonoid contents (TPC and TFC) were determined using Folin-Ciocalteu and AlCl₃ colorimetric assays. The antioxidant capacity was examined by reducing power (RP), ferric reducing antioxidant power (FRAP), ABTS⋅⁺, and DPPH⋅⁺ radical cation assays. All extracts potentially exhibited antioxidant activity with IC₅₀ values ranging from 2.93 to 5.73 μg/mL for ABTS⋅⁺ and from 5.69 to 7.65 μg/mL for DPPH⋅⁺ except the n-hex extract. The BuOH, MeOH, and aqueous extract possess promising anti-skin-aging activities, as observed by an attenuation of UV-A toxicity on human keratinocytes. We proposed that these anti-skin-aging properties are possibly due to direct scavenging activity against reactive oxygen species and upregulate cellular antioxidant machinery. Moreover, we found that the antioxidant capacity was well correlated with anti-inflammatory capacity against nitric oxide (NO) production in terms of the n-hex, AcOEt, and BuOH extracts with IC₅₀ values from 23.21 to 47.1 μg/mL. In contrast, these activities were found to be poorly correlated with AchE activity. To the best of our knowledge, this is the first report of the antioxidant, anti-skin-aging, anti-inflammatory, and anti-acetylcholinesterase activities of the extracts of R. oligophlebia roots. These findings indicated that this species could be a potential source of natural antioxidant, anti-aging, and anti-inflammatory agents. Consequently, it may be suggested as a medicinal plant that prevents diseases related to oxidative stress and inflammatory responses.     

LC/MS/MS Phenolic Profile,Antioxidant Capacity,Angiotensin-Converting Enzyme (ACE) and Glutathione S Transferase (GST) Inhibitory Properties of Ultrasound-Assisted Propolis Extracts

Resinous beehive product propolis has many biological activities. It contains various aromatic substances that have great differences in their chemical composition depending on the natural flora. Thus, chemical characterization and biological properties of propolis samples is an important subject for the pharmaceutical industry. In this study, the propolis samples collected from three cities in Turkey were prepared as methanol (MEP), ethanol (EEP), chloroform (ChlEP), hexane (HxEP), and ethyl acetate (EAEP) extracts using an ultrasonic assisted technique. The antioxidant capacities of the samples were evaluated by free radical scavenging activity (DPPH), cation radical scavenging activity (ABTS), and reducing activity (CUPRAC) and (FRAP). The strongest biological activities were detected in ethanol and methanol extracts. Enzyme inhibition of the propolis samples were determined against the human glutathione S-transferase (GST) and angiotensin converting enzyme (ACE). IC₅₀ values of MEP1, MEP2, and MEP3 samples against the ACE were found as 13.9 μg/mL, 14.8 μg/mL, and 12.8 μg/mL, while against the GST IC₅₀ values of MEP1, MEP2, and MEP3 samples were as 5.92 μg/mL, 9.49 μg/mL, and 5.72 μg/mL. To know the possible causes of the biological test results advanced LC/MS/MS method was applied. trans-ferulic acid, kaempferol, and chrysin were found as the most abundant phenolic compounds in each sample. The propolis extracts obtained using the proper solvent have a good potential to be used in pharmaceuticals to treat the diseases related to oxidative damage, hypertension, and inflammation. Finally, the interactions between chrysin, trans-ferulic acid and kaempferol molecules with ACE and GST receptors were analyzed using molecular docking study. Selected molecules interact with active residues by binding to the active site of the receptors.     

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％。     

Isolation and Chemical Characterization of Antifungal,Antioxidant, and Anti-Inflammatory Compounds from Centrosema coriaceum using GC/MS,UFLC-QTOF-MS,and FACE

In recent years, natural products with biological activities have been increasingly researched. The elucidation of phytoconstituents is necessary for the development of drugs as a natural alternative for the treatment of various diseases. The work aimed to evaluate in vitro and in silico bioactivities of hexane (CCHE) and methanol (CCME) fractions of ethanolic extract from Centrosema coriaceum Benth (Fabaceae) leaves and elucidate their phytoconstituents. CCHE and CCME showed antifungal activity for Candida glabrata (MIC of 1000 μg/mL) with fungistatic effect and action in cell envelope by sorbitol and ergosterol assays. CCHE and CCME presented promising antioxidant activity against the DPPH radical with IC₅₀ of 13.61±0.50 and 6.31±0.40 μg/mL, respectively, and relative antioxidant activity (RAA%) of 45.77±3.61/ 28.53±2.25 % for CCHE and 82.18±2.25/51.99±3.23 % for CCME when compared to rutin and quercetin, respectively. Moreover, these fractions demonstrated promising results for the inhibition of lipid peroxidation by β-carotene/linoleic acid assay. For anti-inflammatory and cytotoxicity activities, CCHE and CCME significantly inhibited the production of nitric oxide and TNF-α, without toxicity on murine intraperitoneal macrophages, respectively. Esters, alkanes, steroids, tocopherols, and terpenes were identified in CCHE by GC/MS. Flavonoids, phenolic acids, and disaccharides were detected in CCME by UFLC-QTOF-MS and FACE. Furthermore, rutin was purified from CCME. In silico predictions evidenced that compounds present in both fractions have high affinity to the fungal membrane besides antioxidant and anti-inflammatory activities. Based on these observations, CCHE and CCME have a noteworthy potential for the design of novel antifungal and anti-inflammatory agents that should be explored in future studies.     

微生物发酵青蒿叶和叶渣的研究

为扩大青蒿原料的应用途径,延伸青蒿产业链,对青蒿叶和叶渣进行发酵研究.拟开发可用于动物保健的青蒿来源的产品.采用微生物发酵青蒿及青蒿叶渣,检测枯草芽孢杆菌、酿酒酵母菌、植物乳杆菌等菌株发酵青蒿叶和叶渣后其粗蛋白、粗脂肪、粗纤维素以及青蒿素、青蒿乙素、双氢青蒿酸、青蒿酸含量变化.青蒿叶发酵产物及功效成分含量与对照组比较,...     

青蒿鲨烯合酶基因的克隆、结构分析与大肠杆菌表达

用RT-PCR方法从青蒿(Artemisia annua L.)中克隆了一个1539bp全长鲨烯合酶cDNA。青蒿鲨烯合酶氨基酸序列与拟南芥、烟草、人类、酵母鲨烯合酶的一致性分别为70％、77％、44％和39％。青蒿鲨烯合酶基因组DNA结构很复杂,包括14个外显子和13个内含子。全长的或C末端截短的鲨烯合酶cDNA被克隆进原核表达载体pET30a并在大肠杆菌(Escherichia coli)BL21(DE3)中诱导表达。但在含有全长的鲨烯合酶cDNA的大肠杆菌中并没有观察到预期大小的鲨烯合酶表达,而C末端截短疏水区30个氨基酸的鲨烯合酶可在大肠杆菌中过量表达。     

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.     

不同土壤环境对黄花蒿生长和青蒿素含量的影响研究

通过田间小区试验,比较研究了施肥与不施肥条件下,4种土壤环境（沙土、旱地土、水稻土和棕色石灰土）对黄花蒿的生长、生物量分配和青蒿素含量的影响。结果表明：黄花蒿对土壤养分的适应性较强,在沙土、旱地土、水稻土和石灰土上均能生长发育,养分水平低时,分配更多的生物量到根,根生物量分数和根/冠比增大;养分水平高时,分配更多的生物量到叶,叶生物量分数增加。黄花蒿的生长和青蒿素含量显著受土壤养分的影响,不施肥时,石灰土和水稻土栽培黄花蒿的株高、地径、总生物量、叶生物量和青蒿素含量显著大于旱地土,而旱地土又显著大于沙土。但在施肥条件下,以上参数不同土壤间无显著差异,且显著高于不施肥。因此,只要根据土壤养分状况合理施肥,黄花蒿在不同养分土壤栽培均能获得较高的青蒿素产量。     

青蒿鲨烯合酶基因的克隆、结构分析与大肠杆菌表达

用RT-PCR方法从青蒿(Artemisia annua L.)中克隆了一个1 539 bp全长鲨烯合酶cDNA.青蒿鲨烯合酶氨基酸序列与拟南芥、烟草、人类、酵母鲨烯合酶的一致性分别为70%、77%、44%和39%.青蒿鲨烯合酶基因组DNA结构很复杂,包括14个外显子和13个内含子.全长的或C末端截短的鲨烯合酶cDNA被克隆进原核表达载体pET30a并在大肠杆菌(Escherichia coli) BL21(DE3)中诱导表达.但在含有全长的鲨烯合酶cDNA的大肠杆菌中并没有观察到预期大小的鲨烯合酶表达,而C末端截短疏水区30个氨基酸的鲨烯合酶可在大肠杆菌中过量表达.     

Anti-Inflammatory,Antioxidant and Enzyme Inhibition Activities in Correlation with Mycochemical Profile of Selected Indigenous Ganoderma spp. from Balkan Region (Serbia)

The aim of this research work was to study the bioactivity potentials (antioxidant, anti-inflammatory, and enzyme inhibitory) of ethanol (EtOH), water (H₂O) and chloroform (CHCl₃) extracts of G. applanatum, G. lucidum, G. pfeifferi and G. resinaceum as well as their mycochemical profile: the total content of phenolics (TP) and sugars (TS) and LC/MS/MS detection of phenolics. LC/MS/MS profile showed that p-hydroxybenzoic and protocatechuic acids were mostly found. The highest ABTS and DPPH activities were detected in polar G. applanatum extracts (159.84±0.59 mg TE/g d.w., IC₅₀=0.85±0.30 μg/mL, respectively), while G. resinaceum CHCl₃ extract was the most potent in NO assay (IC₅₀=41.21±0.18−81.89±0.81 μg/mL). The highest TP and TS were generally determined in G. applanatum EtOH extracts. Enzyme inhibitory effects were determined in H₂O extracts. Generally, CHCl₃ extracts showed the most powerful anti-inflammatory potential. These results suggest that analyzed species are a promising source of bioactive compounds and may be considered as candidates for new food supplements or drug formulations.     

不同氮素水平下入侵种豚草与本地种黄花蒿、蒙古蒿的竞争关系

已有研究表明,土壤氮素增加可提高外来植物的入侵性,降低本地植物的竞争力.为揭示全球氮沉降对入侵种与本地种之间竞争关系的影响,我们于2010年5-8月在中国科学院北京森林生态系统定位研究站温室内,采用取代系列实验方法(standard replacement experiment),研究了3个氮素水平下入侵种豚草(Ambrosia artemisiifolia)与本地种黄花蒿(Artemisia annua)、蒙古蒿(Artemisia mongolica)的生长特征及种内、种间竞争关系的变化.实验采用双因素-随机区组设计,设置了低氮、中氮和高氮3个氮素水平,每一氮素水平分别设置豚草和黄花蒿、豚草和蒙古蒿组成的竞争实验,生长90 d后测量株高和生物量.结果表明:单栽情况下,随氮素水平的增加3个物种的株高均增加,而生物量均无显著变化;混栽情况下,3个物种株高和生物量随氮素水平的增加变化各异,豚草呈极显著增加趋势,而黄花蒿无明显变化,蒙古蒿则先增加后减少.豚草的快速生长使其在竞争中处于优势地位,对本地种黄花蒿和蒙古蒿产生明显的竞争效应.但不同氮素水平下,豚草对本地种的竞争力不同:低氮素水平下,豚草＜两个本地种;中氮素水平下,黄花蒿＜豚草＜蒙古蒿;高氮素水平下,豚草＞两个本地种.氮素添加显著提高了豚草的种间竞争力,改变了豚草与本地种之间的竞争关系,使竞争有利于入侵种.据此推测,在全球变化的背景下,氮沉降的增加将会促进外来种豚草的入侵,增加本地群落的可入侵性.     

青蒿法呢醇合酶原核表达、纯化与功能鉴定

将经RACE方法克隆到的青蒿倍半萜合酶cDNA(AF304444) 开放阅读框插入到原核表达载体pET30a(+)的NcoⅠ和BamHⅠ酶切位点之间,构建N端和C端均携带有HIS₆表达标签的重组表达载体pET30SESQ。将pET30SESQ转入大肠杆菌BL21(DE3), IPTG(Isopropyl-beta-D-thiogalactoside)诱导蛋白表达,表达产物经镍琼脂糖柱纯化。纯化蛋白加入酶促反应体系(FPP),GC-MS分析酶促反应体系的正己烷萃取物,结果显示此重组酶可以催化FPP向法呢醇的转化。     

适于青蒿芽生长和青蒿素积累的光、温和培养方式探讨

探讨了光照、温度和培养方式对青蒿芽生长和青蒿素合成的影响。适宜芽生长和青蒿素积累的光照强度约为３ ０００ ｌｘ,照光时间为２０ ｈ／ｄ ;芽生长和青蒿素积累的最适温度分别为２５ ℃和３０ ℃,通过先２５ ℃（２５ ｄ）后３０ ℃（５ ｄ） 的温度转变二步培养法可以提高青蒿素的产量;青蒿芽生长和青蒿素积累的最佳培养方式为非浸没低转速摇瓶培养。     

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.