【作者】 李丽萍；

【导师】 谢响明；

【作者基本信息】 北京林业大学 ， 微生物， 2010， 硕士

【摘要】 本研究用70%的乙醇提取紫茎泽兰,将所得到的提取液旋转蒸发成浸膏,再依次用石油醚、乙醚、乙酸乙酯、氯仿、正丁醇、丙酮按照极性由小到大萃取,得到不同极性段的活性物质。然后用这些有机溶剂萃取液对5种供试菌进行抑菌试验,筛选出抑菌效果最好的萃取液,以及对这种萃取液最敏感的菌株。经试验得出紫茎泽兰氯仿萃取液对番茄青枯菌有较好的抑制作用,并从抑制剂在生理、生化、细胞形态等方面对番茄青枯菌的抑菌机理进行了深入研究,结果如下：1.紫茎泽兰70%乙醇提取液对金黄色葡萄球菌(Staphylococcus aureus)、枯草芽孢杆菌(Bacillus subtilis)、苏云金芽孢杆菌(Bacillus thuringiensis)、大肠杆菌(Escherichia coli)的最低抑菌浓度均为0.25g／ml,对番茄抑制作用最为显著,其抑菌浓度低达0.0625g/ml,以青枯菌为指示菌。研究了紫茎泽兰提取物的稳定性。结果表明,提取物的抑菌活性具有较好的热稳定性,在pH为9时抑菌效果较佳；2.石油醚和丙酮萃取液对所有供试菌均没有抑制作用,氯仿萃取液对番茄青枯菌(Rastonia solanacearum)的抑制作用较明显；3.氯仿萃取液中添加Zn2+和Mn2+,乙醚萃取液中添加Zn2+抑菌作用有所增加,其他处理抑菌效果没有明显改变；随着随着乙醚、乙酸乙酯、氯仿、正丁醇四种有抑菌活性的萃取液中加入的蔗糖和NaCl溶液浓度的增大。抑菌效果也逐渐增强；4.抑菌机理研究：通过SDS-PAGE电泳的方法检测发现,氯仿萃取液完全抑制或破坏了菌体小分子蛋白质的合成,对大分子蛋白质也有一定的抑制作用;透射电镜观察可知氯仿萃取液破坏了菌体形态；通过酶活测定可知氯仿萃取液抑制了番茄青枯菌的POD、CAT、SDH酶活,且随着加入萃取液浓度的增大,抑制作用也逐渐增强。总之,紫茎泽兰氯仿萃取液扰乱了菌体细胞内正常的代谢活动,导致细胞不再生长、繁殖,达到抑菌的目的。以上研究成果对于番茄青枯病防治的应用有一定的指导意义。更多还原

【Abstract】 In this study,70% ethanol extract of Eupatorium adenophorum Spreng was evaporated into paste. The paste was extracted with petroleum ether, ethyl ether, ethyl acetate, chloroform, butanol and acetone in accordance with the different polarity, meanwhile active substances of different polarity paste were obtained. Then, the antibacterial tests of organic solvent extracts were conducted against 5 experimented strains. Finally, the most sensitive strain and the best antibacterial extracts were selected. The experiment showed that chloroform extracts exhibited the best inhibitory effect against Rastonia solanacearum, and then the antimicrobial tests were carried out. The mechanisms of inhibition on Rastonia solanacearum by use of chloroform extracts was also explored in some aspects such as physiology, biochemistry and cell morphology.1.70% ethanol extracts of Eupatorium adenophorum Spreng had obviously antimicrobial activities, especially to Rastonia solanacearum with a MIC of 0.0625g/ml, and they also exerted the same function on the other 4 bacteria(Staphylococcus aureus, Bacillus subtilis, Bacillus thuringiensis, Escherichia coli)with 0.25g/ml. Moreover, the extracts of Eupatorium adenophorum Spreng were hot stability, antimicrobial activities were better at pH9;2. The petroleum extracts and acetone extracts had no inhibitory effect on all the experimented strains. The chloroform extracts had more distinctly antibacterial activities against Rastonia solanacearum.3. Zn2+,Mn2+ were added into chloroform extract and Zn2+ were added into ethyl ether, which can increase inhibitory effect remarkly. Other treatments didn’t show the similar trend. With the increase of the concentration of NaCl and sucrose, the inhibitory effect gradually improved.4.Inhibitory mechanisms:The result of SDS-PAGE indicated that chloroform extracts completely destroyed or inhibited small moleculars of Rastonia solanacearum while large moleculars also affected to some extent. Observation under transmission electron microscope revealed that the structure of experimented microbes was disrupted by chloroform extracts, experimented microbes’ morphology changed as well; chloroform extracts exhibited inhibitory effect against POD、CAT、SDH of Rastonia solanacearum. With the concentration of extract increasing, inhibition effect rose simultaneously. In conclusion, extracts from Eupatorium adenophorum Spreng disordered normal celluar metabolic process, leading to development and reproducibility ceased, ultimately the antimicrobial purpose was achieved. These researches will provide some guidence for the application of controlling Rastonia solanacearum.更多还原