炭团菌提取物对樟子松枯梢病菌的抑菌活性及稳定性

Antifungal sctivities and stability of extracts from culture liquid of Hypoxylon perforatum to Sphaeropsis sapinea

【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] Our research objective is to obtain the active substances from culture liquid of Hypoxylon perforatum with inhibitory effect on Sphaeropsis sapinea growth and germination. [ Methods] Water and ester were used for extracting active substances from culture liquid of Hypoxylon perforatum, either by extracting directly from cultural liquid, or extracting under ultrasonic. Growth-inhibiting rate and germination-inhibiting rate were used as the index of antifungal activity. [ Results] Five extracts from culture liquid of Hypoxylon perforatum showed antifungal activities to Sphaeropsis sapinea growth and germination. [Conclusion] Ethyl acetate-extract from culture liquid of Hypoxylon perforatum has a highly antifungal activity to Sphaeropsis sapinea, high stability in the natural environment. So it has a highly development of the value and application prospect. The mainly inhibiting active substance is methyl p-methoxy cinnamate, which as medicine intermediate was used in cosmetic to be UV- protection absorbefacient. Diisobutyl phthalate and Dibutyl phthalate are all the common plastic-enhancer for PolyvinylChloride （PVC）. It is very important in medicinal industry and chemistry industry to discovering the natural presence of the three compounds.