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【目的】获得炭团菌液体培养菌液中高效抑制樟子松枯梢病菌的活性物质。【方法】炭团菌液体培养菌液中活性物质的提取采用水提和酯提2种方式。水提方式采用直接提取法和超声波提取法2种方法,酯提方式选择正丁醇、乙酸乙酯、乙醇3种萃取剂。各提取物对病原菌的抑菌活性研究选用菌丝生长抑制率和孢子萌发抑制率2个指标,对病原菌菌丝生长的抑制测定采用生长速率法,对病原菌孢子萌发的抑制试验采用悬滴法。炭团菌液体培养菌液乙酸乙酯提取物稳定性的研究选用紫外线、温度、pH值、氧化剂与还原剂以及贮存时间5个因子。乙酸乙酯提取物中抑菌活性成分的分离采用柱层析、纯化采用薄层层析、结构鉴定采用紫外光谱(UV)、红外光谱(IR)、气质联用(GC-MS)和核磁共振(NMR)等分析测试技术。【结果】炭团菌液体培养菌液5种提取物对樟子松枯梢病菌均有一定的抑菌活性,其中乙酸乙酯提取物对病原菌菌丝生长的抑制率最高,达到73.20%;乙酸乙酯提取物、超声波提取物、正丁醇提取物对病原菌孢子萌发抑制率均超过90%。乙酸乙酯提取物对自然环境下的紫外线、温度具有很高的稳定性,并且具有一定的抗氧化还原能力和良好的耐热性,长期贮存不影响其抑菌活性。乙酸乙酯提取物中有3个组分,组分Ⅰ的抑菌率最高(72.94%)。组分Ⅰ含3种化合物,其中得率最高(57.952%)的为对甲氧基肉桂酸甲酯,化学式为C11H12O3。其他两种化合物为邻苯二甲酸二丁酯和邻苯二甲酸二异丁酯。本研究首次发现上述3种化合物的天然存在。【结论】炭团菌液体培养菌液乙酸乙酯提取物对樟子松枯梢病菌具有较高的抑菌活性,在自然环境下具有很高的稳定性,具有很高的开发价值和应用前景,其主要抑菌活性成分为对甲氧基肉桂酸甲酯。对甲氧基肉桂酸甲酯作为医药中间体主要用于化妆品中,作紫外防护吸收剂;邻苯二甲酸二丁酯和邻苯二甲酸二异丁酯均是聚氯乙烯最常用的增塑剂。上述3种化合物天然存在的发现,对于医药工业和化学工业具有极为重要的意义。
【Objective】 The objective of this study is to obtain the active substance that inhibits Pinus sylvestris var. 【Method】 The two kinds of methods of water extraction and ester extraction were used to extract the active substances in the liquid culture broth of the bacteria. Water extraction method using direct extraction and ultrasonic extraction of two methods, ester extraction method to choose n-butanol, ethyl acetate, ethanol extractants. Antibacterial activity of each extract on the pathogenic bacteria selection mycelial growth inhibition rate and spore germination inhibition rate of two indicators of the inhibition of pathogenic mycelial growth inhibition rate by growth rate method, the spore germination inhibition test using hanging drop method. Study on the Stability of Ethyl Acetate Extracts from Liquid Culture Broth of Elymophila. Five Factors: UV, Temperature, pH, Oxidant and Reducing Agent and Storage Time. The constituents of the antibacterial activity in the ethyl acetate extracts were separated by column chromatography and purified by thin-layer chromatography. The structures were identified by UV, IR, GC-MS and nuclear magnetic resonance (NMR) analysis and testing techniques. 【Result】 The results showed that all of the five extracts from liquid culture of mycobacterium tuberculosis had some antibacterial activity against Pinus sylvestris var. Mongolica and the inhibitory rate of ethyl acetate extract to mycelial growth of pathogen was the highest, reaching 73.20% Ethyl ester extract, ultrasonic extraction, n-butanol extract on spore germination of pathogenic bacteria were more than 90% inhibition rate. Ethyl acetate extract on the natural environment of UV, temperature has a high stability, and has some antioxidant capacity and good heat resistance, long-term storage does not affect the antibacterial activity. There are three components in the ethyl acetate extract, the highest inhibition rate of component I (72.94%). Component I contains three compounds, the highest yield (57.952%) of methyl p-methoxycinnamate, chemical formula C11H12O3. The other two compounds are dibutyl phthalate and diisobutyl phthalate. This study, for the first time, found the natural presence of these three compounds. 【Conclusion】 Ethyl acetate extract from liquid culture broth of Mycobacterium tuberculosis has high antibacterial activity against Shoot Pinus sylvestris var. Mongolica and high stability in natural environment, which has high development value and application prospect , The main antibacterial active ingredient is methyl p-methoxycinnamate. Methoxy cinnamate as a pharmaceutical intermediate is mainly used in cosmetics for UV protection absorbent; dibutyl phthalate and diisobutyl phthalate are the most commonly used plasticized PVC Agent. The natural discovery of these three compounds is of great importance to the pharmaceutical and chemical industries.