真菌提取物抑制烟草花叶病毒(TMV)研究进展

烟草花叶病毒(Tobacco mosaic virus,TMV)是一种具有寄主范围广、抗逆性强、在农业生产上造成危害大和难以防治等特点的植物病毒.从真菌中筛选并获得具有抑制TMV的活性物质是当前研究热点.本文综述了国内外关于真菌中活性物质抑制TMV的研究进展,内容包括3个方面:具有抑制TMV活性物质的真菌种类,主要为侧耳、白蘑、红菇等科中的真菌;具有抑制TMV作用的活性物质成分,主要为真菌多糖和蛋白质;抑制TMV的作用机理,主要包括通过钝化病毒和封闭侵染位点抑制病毒的侵染,干扰病毒蛋白质合成和病毒离子装配抑制病毒增殖或扩散,提高防御酶活性和激活抗性相关基因表达从而诱导植物产生抗性.此外对真菌提取物抗TMV田间应用缓慢的原因进行了讨论.随着对真菌中抗TMV的作用物质及其结构、理化性质、表达基因及抗病毒机制的研究深入,人们对环境与安全问题的日益重视以及化学农药的使用限制,真菌源抗TMV物质具有广阔的开发前景.     

金针菇中蛋白质含量的变化和其中一个蛋白质的生物活性

在对金针菇(Flammulinavelutipes)子实体活性蛋白质纯化过程中发现,金针菇在不同环境条件下不同蛋白质的表达量有差异.通过阴离子交换层析、凝胶层析和高压液相色谱,从金针菇中分离纯化获得了一种蛋白质Zb.经SDS-PAGE测定该蛋白分子量(Mr)为 30×103,等电聚焦电泳表明该蛋白质的等电点为 4. 7,且该蛋白质不含糖.N-端测序表明,该蛋白质N-端的 20个氨基酸为PQVKTSWEDLANLGWPIQQV.此外,在HepG2. 2. 2. 15细胞株上检测该蛋白质体外对肝炎病毒的抑制作用,发现对HBsAg抑制作用明显,抑制中浓度为 0. 117μg/mL,但对HBeAg几乎无作用.以胃癌细胞株MGC80-3为研究对象,检测表明,该蛋白质对MGC80-3抑制率在 50%时的蛋白质浓度约为 0. 75μg/mL.以起始浓度为 0. 296mg/mL的Zb检测它的血凝活性时发现,Zb对兔血红细胞和人A、B、O血红细胞的血凝滴度为 2-12.但同时观察到Zb处理后的兔血红细胞形态发生改变. 图 6表 1参 15     

裸蒴根的化学成分

首次对我国特有属药用植物裸蒴的化学成分进行研究.采用色谱方法分离化合物,以波谱技术鉴定其结构;并采用MTT法对其进行体外细胞毒活性测试.该植物根部的乙醇提取物中分离出4个化合物,分别鉴定为5-癸酰基-2-壬基吡啶(1)、豆甾烷-3,6二酮(2)、豆甾-4-烯-3,6二酮(3)和胡萝卜苷(4).其中化合物1~3为首次从裸蒴属植物中分离得到,这3个化合物对人肝癌细胞(SMMC-7721和Hep3B)的增殖均显示一定的抑制活性.     

凹叶厚朴中具有α-葡萄糖苷酶抑制活性的成分

凹叶厚朴是四川省人工栽种的一种用途广泛的常用中药.在发现凹叶厚朴乙醇提取物具有较强的α-葡萄糖苷酶抑制活性基础上,通过活性跟踪,对该提取物进行了分离,发现其中的生物碱显示了较强的抑制活性.通过溶剂萃取、树脂吸附和反复硅胶柱层析等分离方法从凹叶厚朴乙醇提取物中分离得到6个生物碱.用质谱和核磁共振等波谱方法分别鉴定为木兰箭毒碱,木兰花碱,鹅掌楸碱,蕃荔枝碱,罗默碱和Lysicamine,并应用小肠α-葡萄糖苷酶模型测定了它们对α-葡萄糖苷酶的抑制作用.其中,番荔枝碱和木兰箭毒碱的抑制活性较好,分别为60%和62%;其它4个生物碱成分活性几乎相当,鹅掌楸碱为46%,罗默碱为51%,Lysicamine为49%,木兰花碱为51%.图2参10     

核桃粕中抗氧化和糖苷酶抑制活性成分

为了发掘核桃粕潜在的利用价值,提取核桃粕的化学成分,测定其抗氧化活性和糖苷酶抑制活性并进行活性跟踪.结果显示,95%乙醇提取物具有一定的生物活性,抗氧化能力用DPPH法测定自由基清除率来表示,测得IC_(50)为1.482 8 mg/m L,抑制糖苷酶的能力用α-淀粉酶和α-葡萄糖苷酶的抑制活性表示,测得IC_(50)分别为6.679 8 mg/m L和0.3572 mg/m L.为了得到活性物质的最大提取效率,分别选用95%乙醇、纯水、70%乙醇、60%丙酮4种溶剂提取样品,并从提取得率、抗氧化活性、糖苷酶抑制活性等方面进行评估,最后选用70%乙醇为最佳提取溶剂.对70%乙醇粗提物进一步分相萃取,结合柱色谱、薄层色谱结晶等分离方式,从乙酸乙酯相中得到化合物Y-10-1,其清除自由基IC_(50)为0.0372 mg/m L,抑制α-淀粉酶和α-葡萄糖苷酶IC_(50)分别为0.017 9 mg/m L和0.617 3 mg/m L.抗氧化活性和糖苷酶抑制活性分别以维生素C和阿卡波糖作为阳性对照,测得阳性对照的抗氧化IC_(50)为0.100 2 mg/m L,α-淀粉酶抑制IC_(50)为1.462 mg/m L,α-葡萄糖苷酶抑制IC_(50)为0.165 1 mg/m L.经比较,Y-10-1的抗氧化活性和α-淀粉酶抑制活性均优于阳性对照.经液相和质谱鉴定,基本确定Y-10-1为没食子酸.本研究说明Y-10-1具有潜在的使用和开发价值.     

果胶酶激发子对黄瓜叶片病程相关蛋白及细胞壁物质的影响

以草酸青霉菌(Penicillium oxalicum)的固态发酵产物———果胶酶粗提液喷雾诱导黄瓜,研究了该激发子对黄瓜细胞内几丁质酶和-β1,3-葡聚糖酶2种病程相关蛋白(PR s蛋白)以及细胞壁物质的影响.结果表明,经激发子处理后,黄瓜叶片中几丁质酶和-β1,3-葡聚糖酶活性诱导升高,木质素及富含羟脯氨酸糖蛋白(HRGP)含量也诱导增强;同时,在激发子处理后接种黑星病菌的黄瓜中,上述2种PR s蛋白及细胞壁物质被大量诱导合成.其中,两种PR s蛋白均在接种后d 4达到最高值,分别为22.72 U/mg(pro)和268.16 U/mg(pro);木质素和HRGP含量在接种后d 3后达最高,分别比对照高196.12%和272.98%.表明果胶酶激发子在接种前期对黄瓜几丁质酶、-β1,3-葡聚糖酶、木质素及HRGP的诱导产生是导致病情降低的内在机制.图4参16     

TMV或宁南霉素处理对烟草叶片及类囊体膜蛋白组成的影响

对烟草花叶病毒(TMV)侵染、宁南霉素钝化处理的烟草叶片进行了叶片全蛋白多肽组分、类囊体膜蛋白组成的分析.结果表明,TMV侵染或钝化实验的烟草叶片中均出现TMV衣壳蛋白;但是与对照相比,TMV侵染叶片中Rubisco大亚基减少、小亚基增加,类囊体膜蛋白中PsaA、PsaB和Mr=33×103蛋白含量减少.表明TMV侵染要同时影响叶片细胞核基因组和叶绿体基因组的基因表达,从而影响光合作用碳代谢和类囊体膜光系统Ⅰ和Ⅱ.宁南霉素对TMV病毒短时间内具有钝化作用,而较长时间的作用效果不同,10 min钝化样品的蛋白组分保持较为完整,而30 min钝化样品的多肽缺失较为严重.图5表1参11     

芦竹抑藻物质的初步分离及抑制铜绿微囊藻的效果

采用乙酸乙酯提取芦竹(Arundo donax Linn.)得到高效抑藻粗提物,其对产毒铜绿微囊藻的半效应抑制浓度(EC50.6d)为50mg·1-1.粗提物经溶剂萃取分离得到中/碱性物质及酸性物质.活性分析表明,酸性物质有显著抑制铜绿微囊藻的活性.通过酸性氧化铝柱层析进一步分离酸性物质,得到三组弱极性抑藻物质及一组强极性抑藻物质.培养3d后,四组抑藻物质在10mg·1-1时的抑藻率分别为82%,83%,100%以及70%.     

聚花野丁香的化学成分及其抗氧化活性

为研究聚花野丁香(Leptodermis glomerata)的生物活性,综合利用多种色谱方法,对该植物95%乙醇提取物的化学成分进行了分离、纯化,从中共得到15个化合物.通过波谱数据分析及比对,将其分别鉴定为杨梅素(1)、(2R,3R)-二氢杨梅素(2)、槲皮素(3)、pinostrobin(4)、1,3-二甲氧基-2-羟基蒽醌(5)、2-甲氧基-1,3-二羟基蒽醌(6)、1,3,4-三甲氧基-2-羟基蒽醌(7)、1,2,3,4-四甲氧基蒽醌(8)、1,2,3-三甲氧基蒽醌(9)、5,7-二羟基色原酮(10)、3S-faramol(11)、(2R,4S)-catalponol(12)、(2S,4S)-catalponol(13)、3β-乙酰基齐墩果酸(14)、齐墩果酸(15).采用DPPH、ABTS、NO三种自由基清除活性测试模型,对上述化合物的抗氧化活性进行测试.结果表明,(2R,3R)-二氢杨梅素(2)、pinostrobin(4)和5,7-二羟基色原酮(10)表现出显著的自由基清除活性:当浓度为1 mg/mL时,化合物2对DPPH、NO自由基的清除率为91.2%、67.8%;当浓度为2 mg/mL时,化合物4对DPPH自由基的清除率为102.8%;当浓度为1mg/mL时,化合物10对ABTS、NO自由基的清除率分别为104.7%、95.7%.本研究表明聚花野丁香富含多羟基酚类化合物,其中部分化合物显示较强的抗氧化活性,具有开发利用价值.(图1表1参23)     

QuEChERS-超高效液相色谱-串联质谱法测定鱼血清中的精神活性物质

本文建立了一种同时测定鱼血清中19种精神活性物质的分析方法。该方法基于QuEChERS方法对样品进行提取和净化,待测物采用超高效液相色谱-串联质谱法(UPLC-MS/MS)进行分析,检测的19种精神活性物质均采用同位素内标法定量。该方法的方法检测限(MDL)范围为0.02—0.06 ng·mL^-1,基质效应处于76.7%—127.7%之间,相对回收率处于81.1%—123.9%之间,方法相对标准偏差均≥18.0%。该方法成功应用于对鲫鱼血清中19种精神活性物质的残留检测。     

Antibacterial,antiviral and antioxidant activities of aerial part extracts of Peganum harmala L. grown in Tunisia

Ethyl acetate, chloroform, butanol, and methanol extracts from the aerial part of Peganum harmala were tested for antibacterial, antioxidant, and antiviral activities. The antibacterial activity was evaluated by the determination of minimum inhibitory concentration (MIC) using the solid medium technique. Oxacillin, amoxicillin, ticarcillin, cefotaxim, and amphotericin were used as control agents. The antiviral activity was evaluated against human cytomegalovirus (HCMV) strain AD-169 (ATCC Ref. VR 538) and Coxsackie B virus type 3 (CoxB-3) using a cytopathic effect (CPE) reduction assay. The antioxidant activity was evaluated using two tests: 1,1-diphenyl-2-picrylhydrazyl (DPPH) free-radical scavenging and ammonium thiocyanate methods. Among tested extracts; the chloroform extract displayed a higher antibacterial activity against Gram-positive than Gram-negative bacteria. The butanol extract demonstrated the highest antioxidant activity. The methanol extract showed significant antiviral activity against CoxB-3 virus. The chloroform extract may be an important source of bactericidal compounds against Gram-positive bacteria.     

烟草花叶病毒siRNA设计及其植物表达载体构建

RNA干扰技术已广泛应用于沉默基因表达的研究.本文分析烟草花叶病毒(TMV)基因序列,选择设计编码小分子发卡结构RNA(hpRNA)的cDNA;并根据农杆菌双元载体质粒p2355多克隆位点区限制性酶切位点,两端分别加入XbaI和BamHI酶切位点;分别合成单链DNA复性后插入到p2355上,经PCR、测序验证,表明已成功构建了具有潜在表达烟草花叶病毒siRNA的植物表达载体.图3表1参26     

Induction of host specificity in larvae of Manduca sexta: chemical dependence controlling host recognition and developmental rate

Summary. The tobacco hornworm, Manduca sexta L. (Lepidoptera: Sphingidae) is a specialist on Solanaceae. This host specificity is induced as the larva feeds on solanaceous foliage, so that solanaceous-reared larvae will refuse to feed on other plants. Experiments were designed to determine the role of dietary constituents on the induction of host specificity and the effects of these on development of M. sexta. Choice assays using leaf discs of cowpea, Vigna sinensis (Fabaceae), were used to monitor the isolation of relevant chemical cues from foliage of potato. An aqueous extract of potato foliage strongly stimulated feeding. This extract was partitioned with n-butanol under alkaline conditions to obtain a highly active butanol extract. Reversed phase flash chromatography with a water-methanol gradient gave an active fraction that was used as a supplement for wheat germ-based artificial diet. Larvae reared on this s-diet became dependent on potato allelochemicals for initiation and continuation of feeding activity. These larvae also developed faster than larvae reared on the control p-diet, but no effect on adult mass was detected. Further flash chromatography of the active fraction under alkaline conditions provided a highly active sub-fraction, and semi-preparative HPLC using gradients of water and acetonitrile resulted in the isolation of a single compound. Bioassays indicated that this compound alone can account for host recognition by solanaceous-reared larvae. The results suggest that the mechanism of induced host specificity in M. sexta involves development of dependence on this compound. Received 21 December 1999; accepted 14 March 2000     

In vitro antibacterial and anti-inflammatory properties of seaweed extracts against acne inducing bacteria, Propionibacterium acnes

This study was conducted to evaluate the antimicrobial activities of common seaweeds from the coast of South Korea against the etiologic agents of acne vulgaris. Fifty-seven species of seaweed were screened for potential antimicrobial activity. Methanol extracts of 13 species (22.8%) showed inhibitory effects against Propionibacterium acnes. The aqueous extracts of only two species (3.5%) showed antimicrobial activity. When tested with the agar disk diffusion method, Ecklonia cava, E. kurome, Ishige sinicola, and Symphyocladia latiuscula had the strongest inhibitory effects. However, these four seaweed extracts showed no antibacterial activity against Staphylococcus epidermidis at 5 mg disk-1. The minimum inhibitory concentration (MIC) values of E. cava and E. kurome were both 0.31 mg ml-1 and the MIC values of l. sinicola and S. latiuscula were 0.26 and 0.21 mg ml-1, respectively. Among whole plants of E. cava and E. kurome, extracts of the pinnate blade had the highest inhibitory activity on bacterial growth. In cytotoxicity assays, methanol extracts of E. cava, E. kurome, and I. sinicola showed no effect on cell viability at concentrations of 200 microg ml-1. However, the methanol extracts of S. latiuscula reduced cell viability rates to 50% at the same concentration. Additionally, methanol extracts of E. cava, E. kurome, and I. sinicola potently inhibited the in vitro production of nitric oxide. These results suggest that the methanol extracts from these three species may be useful in the development of therapeutic agents for acne vulgaris. Further investigations to determine the bioactive compound are in progress.     

转基因烟草中赖氨酸含量的提高

采用叶盘法将高赖氨酸蛋白基因导入到烟草中.GUS组织化学染色、PCR扩增、Southernblot分析表明, 该基因已经整合到烟草基因组中.测定 10株转基因烟草叶片赖氨酸含量,大部分植株有着明显的提高,最高幅度达到了49. 89%. 图 6参 11     

长期继代培养马铃薯愈伤组织的植株再生

对影响马铃薯愈伤组织发生、生长和长期继代培养的愈伤组织再生植株的几种因素进行了研究。马铃薯茎段的愈伤组织发生频率高于叶片的愈伤组织发生频率。Ｄｉｃａｍｂａ诱导茎段和叶片的愈伤组织发生频率在９０％以上；而２，４－Ｄ的诱导效果不佳并诱发外植体生根。再生培养其中加入０．５ｇ／Ｌ的酶解酷蛋白是必需的；蔗糖的浓度以２％－４％为宜；附加ＺＴ２．０ｍｇ／Ｌ、ＧＡ０．１ｍｇ／Ｌ可使长期继代的愈伤组织的植株再生频率     

壳寡糖诱导转反义mapk基因烟草的TMV抗性和PAL活性研究

壳寡糖具有诱导植物对病原物防御反应的作用,而PAL活性的升高通常被认为是发生抗性反应的标志之一.本文以mapk基因受到反义抑制的转基因枯斑三生烟草为试材,比较了壳寡糖诱导后转基因型和野生型烟草的PAL活性变化及对TMV抗性的变化.由实验结果初步推测,在壳寡糖诱导的植物抗性信号通路中,mapk可能为PAL的一个重要的上游基因,同时mapk在烟草抗TMV信号转导中亦为一个重要组分.图4表2参13     

Synthesis and medicinal properties of plant-derived vilangin

This article reports for the first time the antibacterial, antifungal, antioxidant and anticancer activities of vilangin synthesised from embelin isolated from Embelia ribes berries. E. ribes is widely used as traditional herbal medicine in India. E. ribes is used as appetiser, carminative, anthelmintic, alexiteric, laxative and alterative. This plant is also used to treat tumour, ascites, bronchitis, mental diseases, dyspnoea, heart diseases, urinary discharges, jaundice, hemicrania, and worms in wounds. Here the major compound, embelin, was isolated from the fruits of E. ribes. Vilangin was synthesised by condensation of embelin with formaldehyde in acetic acid solution. The biological activity of vilangin is unknown so far. Hence, we studied antibacterial, antifungal, antioxidant and anticancer activities of vilangin. The minimum inhibitory concentration value of vilangin was 62.5 μg/mL against Staphylococcus epidermidis and Salmonella typhimurium. The radical scavenging activity was maximum at 1,000 μg/mL. Ferric reducing antioxidant power assay of vilangin showed a twofold higher value compared to the standard. Vilangin showed 61.95 % of cytotoxic activity in vitro against A549 lung adenocarcinoma cancer cell line at a dose of 500 μg/mL with half-maximal inhibitory concentration (IC₅₀) of 400 μg/mL. Vilangin can thus be further tested as a new drug.     

Ozon: Bioindikation von Photooxidantien mit Bel W3-Tabak

Results of an ozone-biomonitoring program performed from May to October 1994 at five different stations on exposed Bel W3 tobacco plants in the southwest of Germany (region of Trier) are presented. No linear dose-response relationship could be deduced between ambient ozone concentration and leaf necroses. Gas-exchange rates of tobacco plants during typical summer days would have to be determined to evaluate the stomatal uptake of photo-oxidant gases. Nevertheless, monitoring with the aid of Bel W3 tobacco plants permits an evaluation of qualitative differences in the ambient ozone concentrations. More severe leaf damage were clearly related to periods with higher radiation and was consequently found more frequently in sites located at higher altitudes. Microclimatic differences, however, may also be responsible in part for the ozone-related leaf damage seen at these sites. For this reason, only general conclusions concerning the relative levels of pollution can be deduced from biomonitoring studies using the reactivity of the indicator plant, which strongly restricts the reliability of such field studies. Pigment levels and leaf patterns in tobacco plants which still demonstrate no necrosis cannot be used as early ozone indicators.     

烟根腐解物对烟草生长的影响

烟草生产过程中会产生大量烟草残体,为了研究烟草残体对烟草生长的影响,将烟根、水稻土和无菌水按一定比例混合后,在适当条件下培养得到烟根腐解物.将烟根腐解物按不同的施用量设置6个处理组,并以未施用烟根腐解物组作为对照,分别在烟草生长的团棵期、旺长期和成熟期测定烟草的农艺性状、根系活力、硝酸还原酶(nitrate reduc...