紫外辐射增强对灯盏花内生细菌的影响初探

灯盏花根茎叶表面消毒法后,采用稀释平板法分离灯盏花根茎叶中的内生细菌,对分离菌株进行革兰染色和芽胞染色,结果表明:灯盏花根内生细菌的数量显著或极显著多于叶和茎,紫外辐射极显著减少灯盏花叶和茎中内生细菌的数量.76 %和54 %的灯盏花内生细菌为G~+细菌和含芽胞的细菌,紫外辐射增加灯盏花叶和茎中内生G~+细菌和含芽胞细菌的比例.     

药用植物灯盏花的研究进展

对药用植物灯盏花的特征特性、化学成分、药理作用及栽培技术,以及灯盏花的研究进展进行了综述,为进一步研究该植物提供参考．     

灯盏花素治疗脑梗死50例临床观察

目的：观察灯盏花素治疗脑梗死的临床疗效。方法交１００例急性脑死患者随机分为两组,治疗组５０例,应用灯盏花素治疗;对照组５０例,应用脉络宁治疗。结果治疗组总有效率为９２．００％,对照组照组总有效率为７２．００％,两组比较有显著性差异（Ｐ〈０．０５）。结论灯盏花素是治疗急性脑梗死较为理想的药物,值得临床推广。     

注射用灯盏花素滴注过程中产生混浊的原因分析

目的探讨注射用灯盏花索在滴注过程中产生混浊沉淀现象的原因。为临床提供合理配伍使用依据。方法取不同批次与厂家的注射用灯盏花素,在不同输液品种、不同pH值输液及不同注射用灯盏花素浓度等条件下,观察产生的混浊现象。结果注射用灯盏花素在溶液pH值〈3．6,发生混浊的现象加快;pH值在3．6以上．发生混浊现象延迟;pH值在4．0以上,观察360min未发生混浊现象。结论引起混浊现象的主要原因是溶液的pH值．葡萄糖注射液的pH值规定范围为3．2～5．5,一般pH值在4．0以下,可在滴注过程中产生混浊现象,因此,葡萄糖注射液不宜用作灯盏花素溶解稀释溶煤,而氯化钠注射液适用于灯盏花素溶解稀释溶媒。     

灯盏花雄性不育种质鉴定与花药发育的细胞学研究

对云南泸西栽培灯盏花群体进行调查,发现了灯盏花雄性不育种质个体,其出现频率约为1.06×10-4.对所发现的灯盏花不育株形态特征及其花药发育过程进行了观察,并对花粉活力进行鉴定.结果显示:(1)灯盏花不育株根、茎、叶形态与正常可育植株基本相似,管状花小,花丝短,花药瘦小,无花粉粒散出或花粉无活力.(2)灯盏花在其花药发育的小孢子母细胞时期、四分体时期、小孢子时期和单核早期,由于绒毡层细胞液泡化、提前解体,不能为小孢子或花粉发育提供所需物质,导致小孢子母细胞和四分体解体,产生无花粉的花药;或小孢子和单核花粉胞内降解,形成不同形状和外壁纹饰的败育花粉.研究认为,灯盏花花药绒毡层异常是其花粉败育的主要原因.     

灯盏花大小孢子形成与胚胎发育

用石蜡切片法对灯盏花从花原基分化到胚胎形成的全过程进行系统观察,结果表明:灯盏花从初孕花序至头状花序的直径(d)在2.8～3.1 mm时为花器官分化期,包括花原基分化期、花萼与花冠分化期、雌雄蕊分化期以及分化完成期4个阶段;灯盏花花药4室,药壁发育属双子叶型,绒毡层细胞属于变形绒毡层;小孢子母细胞减数分裂为同时型,成熟花粉为3-细胞型;子房下位,1室,单珠被,薄珠心,胚珠倒生,胚囊发育为蓼型;珠孔受精,属于有丝分裂前类型,胚乳发育为细胞型,胚胎发育应属紫菀型.     

栽培灯盏花DNA指纹图谱研究及遗传多样性分析

目的:采用ISSR标记构建云南省栽培灯盏花DNA指纹图谱并进行遗传多样性分析.方法:从20对候选引物中筛选出7对引物对云南10个地方栽培灯盏花进行扩增,通过扩增带型的差异构建其DNA指纹图谱,利用Popgen32、NTSYS2软件进行遗传一致性系数和遗传多样分析.结果:7对引物在10份共扩增出48条谱带,其中33条具有多态性,占68.8％,表明各居群的遗传差异较大.10个地方种质资源被聚为2个大类,野生居群被聚为一类,栽培灯盏花聚为一类,其中栽培灯盏花有两大亚类.结论:通过构建DNA指纹图谱容易地把灯盏花种植资源相互区分鉴别出来,10个地方灯盏花种质资源遗传多样性丰富,栽培灯盏花与野生具有差异,栽培种质材料之间也有差异.     

灯盏花产黄酮内生放线菌筛选及产黄酮能力的初步研究

通过黄酮类物质特异颜色反应和可见分光光度法,筛选产黄酮的灯盏花内生放线菌,研究灯盏花产黄酮内生放线菌在PDA、高氏1号和淀粉3种培养基上的产黄酮能力。结果表明:59株灯盏花内生放线菌中,8株菌的镁粉+浓盐酸、氯化铝、浓氨水3种颜色反应均成阳性,能够产生黄酮。形态学观察初步鉴定这8株菌均为链霉菌属(Streptomyces)。3种培养基中,在PDA培养基上灯盏花产黄酮内生放线菌的菌丝体生物量、黄酮含量和产量较高。8株灯盏花产黄酮内生放线菌中,菌株RA′1-7生长较好,菌株ELA′3-2和RA2-1菌丝体黄酮含量较高,菌株ELA′3-2菌丝体黄酮产量较高。     

灯盏花内生真菌多样性、群落结构及生态功能预测

【背景】灯盏花(Erigeron breviscapus)是国内知名的传统中药材，但关于灯盏花内生真菌多样性、群落结构和生态功能研究报道比较缺乏。【目的】探究灯盏花不同药用部位内生真菌多样性、群落结构组成及生态功能。【方法】采用ITS序列的高通量测序技术对比研究云南道地药材灯盏花根、茎、叶和花的内生菌群落结构及生物多样性差异，并利用FUNGuild数据库预测真菌群落生态功能。【结果】12个样品共获得540个操作分类单元(operational taxonomic unit, OTU)，分属于5个门22个纲55个目114个科188个属。4个不同药用部位共有的OTU数目仅占14.45%，以根部独有OTU最多。各组织均以子囊菌门和担子菌门为优势菌门；其中，根部以子囊菌门为主，花部位以担子菌门为主。亚隔孢壳属(Didymella)为灯盏花植物的核心属，在各组织中均有分布；其余优势属尚有线黑粉菌属(Filobasidium)、Cystofilobasidium、织球壳属(Plectosphaerella)，灯盏花4个组织中优势属和特有属分布各不相同。α多样性分析表明，根部内生真菌丰度显著高于其他组织，但多样性方面组织差异不明显。PCoA结果表明，根部菌落结构相对独立，而叶与茎中菌落结构较为相似。利用FUNGuild数据库分析发现，腐生真菌在各组织中占比较高，并含有大量未知功能菌群。【结论】灯盏花不同药用部位内生真菌群落组成存在明显差异，具有组织偏好。以上研究完善了灯盏花内生真菌资源的生物信息，为灯盏花内生真菌资源的开发利用提供了理论依据。     

对九种天然产物清除自由基活性的理论评价

用量子化学方法计算了源自灯盏花、丹参、银杏和红花的九种天然产物--灯盏花乙素、焦袂康酸、3,5-二咖啡酰氧基奎宁酸、飞蓬酯B、丹酚酸B、丹参素、银杏双黄酮、银杏苦内酯B和羟基红花黄色素A的O-H键解离焓和电离势,并以此为理论指标评价了其清除自由基活性.结果发现,在非极性溶剂中3,5-二咖啡酰氧基奎宁酸、飞蓬酯B、丹酚酸B、丹参素等均具有较高的活性;而在极性溶剂中,相比其它八种化合物,灯盏花乙素显示出优良的清除自由基活性,提示这些化合物有望作为药物的有效成分治疗自由基引起的心、脑血管疾病.     

灯盏花 chi 的克隆及其生物信息学分析

查尔酮异构酶（CHI）是调控黄酮生物合成的关键酶,分离和克隆这一酶的功能基因,对利用转基因技术进行灯盏花黄酮生物合成的调控具有重要意义。本研究采用RT-PCR和RACE技术,获得了chi cDNA全序列,GenBank登录号为GU208823.1,序列全长996 bp,开放阅读框为594 bp,编码197个氨基酸,3-Race有一个多聚腺苷酸加尾信号。应用软件预测该基因编码蛋白分子量约为21.6 kD,理论等电点为4.78。该基因编码的蛋白无跨膜结构域,其二级结构的主要构件为α-螺旋和随机卷曲。对其三级结构进行了建模,表明其结构与苜蓿chi的三级结构相似。同时根据灯盏花chi N端序列变化的特征,提出了灯盏乙素的合成可能与chi在细胞亚结构的定位及其与合成代谢相关酶形成复合酶的特异性有关。研究为利用基因工程定向改变灯盏花黄酮代谢产物奠定了基础。     

灯盏花chi的克隆及其生物信息学分析

查尔酮异构酶(CHI)是调控黄酮生物合成的关键酶,分离和克隆这一酶的功能基因,对利用转基因技术进行灯盏花黄酮生物合成的调控具有重要意义。本研究采用RT-PCR和RACE技术,获得了chi cDNA全序列,GenBank登录号为GU208823.1,序列全长996 bp,开放阅读框为594 bp,编码197个氨基酸,3-Race有一个多聚腺苷酸加尾信号。应用软件预测该基因编码蛋白分子量约为21.6 kD,理论等电点为4.78。该基因编码的蛋白无跨膜结构域,其二级结构的主要构件为α-螺旋和随机卷曲。对其三级结构进行了建模,表明其结构与苜蓿chi的三级结构相似。同时根据灯盏花chi N端序列变化的特征,提出了灯盏乙素的合成可能与chi在细胞亚结构的定位及其与合成代谢相关酶形成复合酶的特异性有关。研究为利用基因工程定向改变灯盏花黄酮代谢产物奠定了基础。     

Binding of Breviscapine Toward Serum Albumin Studied by Spectroscopic and Electrochemical Techniques

Breviscapine, a cerebrovascular drugs extracted from the Chinese herb Erigeron breviscapinus, has been frequently used to clinically treat cerebrovascular diseases such as cerebral thrombosis, cerebral infarction, and cerebral circulation insufficiency. In order to understand its pharmacology or toxicity, the binding mechanism of breviscapine to a model protein, human serum albumin (HSA), was probed by fluorescence, circular dichroism, Fourier transform infrared spectroscopy (FTIR), and electrochemical impedance spectroscopy approaches. The binding affinities and number of the drug with HSA were about 1.73 × 10⁴ M^?1 and 0.99 at 293 K, respectively. The conformation of the protein was slightly altered after interacting with breviscapine. The drug–protein complex was mainly stabilized by electrostatic forces.     

Two-step purification of scutellarin from Erigeron breviscapus (vant.) Hand. Mazz. by high-speed counter-current chromatography

Scutellarin, a flavone glycoside, popularly applied for the treatment of cardiopathy, has been purified in two-step purification by high-speed counter-current chromatography (HSCCC) from Erigeron breviscapus (vant.) Hand. Mazz. (Deng-zhan-hua in Chinese), a well-known traditional Chinese medicinal plant for heart disease. Two solvent systems, n-hexane-ethyl acetate-methanol-acetic acid-water (1:6:1.5:1:4, v/v/v/v/v) and ethyl acetate-n-butanol-acetonitrile-0.1% HCl (5:2:5:10, v/v/v/v) were used for the two-step purification. The purity of the collected fraction of scutellarin was 95.6%. This study supplies a new alternative method for purification of scutellarin.     

Separation of scutellarin from crude extracts of Erigeron breviscapus (vant.) Hand. Mazz. by macroporous resins

Scutellarin, a flavone glycoside, popularly used in the treatment of heart disease, has been efficiently separated using macroporous resins from crude extracts of Chinese medicinal plant Erigeron breviscapus (vant.) Hand. Mazz. HPD-800 resin offered the best adsorption and desorption capacity for scutellarin among the eight macroporous resins tested, and its adsorption data at 25 degrees C fit best to the Langmuir isotherm. The dynamic adsorption and desorption experiments have been carried out on a HPD-800 resin packed column to optimize the separation process of scutellarin from the crude extracts of E. breviscapus. After one run treatment with HPD-800 resin, the scutellarin content in the product was increased 15.69-fold from 2.61% to 40.96% with a recovery yield of 95.01%. The preparative separation process via adsorption-desorption method developed in this study provides a new approach for scale-up separation and purification of scutellarin for its wide pharmaceutical use.     

Neuroprotective effects of scutellarin against hypoxic-ischemic-induced cerebral injury via augmentation of antioxidant defense capacity

An increasing number of studies has indicated that hypoxic-ischemic-induced cerebral injury is partly mediated via oxidative stress. Recent researches have focused on searching for drug and herbal manipulations to protect against hypoxic-ischemic-induced oxidative cell damage. Scutellarin is a flavonoid derived from the Erigeron breviscapus (vant.) and has been reported to exhibit neuroprotective properties. However, its precise mechanism, particularly its antioxidation mechanism, remains elusive. In the present study, we investigated the neuroprotective effects of scutellarin on middle cerebral artery occlusion (MCAO)-induced brain damage in rats, and oxygen-glucose deprivation (OGD)-induced toxicity in primary culture of rat cortical neurons. In vivo, intraperitoneal injections of scutellarin (20 and 60 mg/kg) improved the neurological score and diminished the percentage of brain infarct volume. At the same time, scutellarin significantly increased superoxide dismutase (SOD), catalase (CAT) activities and glutathione (GSH) level in ischemic brain tissues, enhancing endogenous antioxidant activity. Moreover, pretreatment of scutellarin (25, 50 and 100 μM) protected neurons against lethal stimuli, decreased the percentage of apoptotic cells and inhibited reactive oxygen species (ROS) generation in OGD-induced primary cortical neurons in vitro. These results suggest that the preventive and therapeutic potential of scutellarin in cerebral injury patients is, at least in part, ascribed to augmentation of cellular antioxidant defense capacity.     

Plant regeneration of <Emphasis Type="Italic">Erigeron breviscapus</Emphasis> (vant.) Hand. Mazz. and its chromatographic fingerprint analysis for quality control

An efficient micropropagation system for Erigeron breviscapus (vant.) Hand. Mazz., an important medicinal plant for heart disease, has been developed. Shoot organogenesis occurred from E. breviscapus leaf explants inoculated on a medium supplemented with a combination of plant growth regulators. On average, 17 shoots per leaf explant were produced after 30 days when they were cultured on MS basal salts and vitamin medium containing 5 μM 6-benzylaminopurine (BAP) and 5 μM 1-naphthaleneacetic acid (NAA). All the regenerated shoots formed complete plantlets on a medium containing 2.5–10 μM indole-3-butyric acid (IBA) within 30 days, and 80.2% of the regenerated plantlets survived and grew vigorously in field conditions. Based on the variation in common peaks and the produced amount of the most important bioactive component, scutellarin, a high performance liquid chromatography (HPLC) fingerprinting system was developed for quality control of these micropropagated plants. Chemical constituents in E. breviscapus micropropagated plants varied during plant development from regeneration to maturation, the latter of which showed the most similar phytochemical profile in comparison with mother plants. The regeneration protocol and HPLC fingerprint analysis developed here provided a new approach to quality control of micropropagated plants producing secondary metabolites with significant implications for germplasm conservation.     

Scutellarin promotes in vitro angiogenesis in human umbilical vein endothelial cells

Angiogenesis is critical to a wide range of physiological and pathological processes. Scutellarin, a major flavonoid of a Chinese herbal medicine Erigeron breviscapus (Vant.) Hand. Mazz. has been shown to offer beneficial effects on cardiovascular and cerebrovascular functions. However, scutellarin’s effects on angiogenesis and underlying mechanisms are not fully elucidated. Here, we studied angiogenic effects of scutellarin on human umbilical vein endothelial cells (HUVECs) in vitro. Scutellarin was found by MTT assay to induce proliferation of HUVECs. In scutellarin-treated HUVECs, a dramatic increase in migration was measured by wound healing assay; Transwell chamber assay found significantly more invading cells in scutellarin-treated groups. Scutellarin also promoted capillary-like tube formation in HUVECs on Matrigel, and significantly upregulated platelet endothelial cell adhesion molecule-1 at both mRNA and protein levels. Scutellarin’s angiogenic mechanism was investigated in vitro by measuring expression of angiogenic factors associated with cell migration and invasion. Scutellarin strongly induced MMP-2 activation and mRNA expression in cultured HUVECs in a concentration-dependent manner. Taken together, these results suggest that scutellarin promotes angiogenesis and may form a basis for angiogenic therapy.