凤尾菇菌丝体及发酵液抗菌组分的分离及活性研究
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摘要
本研究以担子菌纲侧耳属的凤尾菇(Pleurotus sajor-caju)为实验材料,经液体深层发酵培养,得到凤尾菇菌丝体和发酵液。菌丝体经乙醇溶剂浸提,得菌丝体乙醇提取物,再经溶剂萃取。经活性检测,菌丝体粗提物及其萃取相抗细菌抗真菌活性较弱。发酵液经乙酸乙酯和正丁醇分级萃取,显示出较强的抗菌活性,并运用硅胶薄层层析、硅胶柱层析以及Sephadex-LH 20凝胶层析手段,分离得到了数个组分。通过生物活性跟踪实验,从中发现了一些具有较好抗细菌抗真菌活性组分,其中组分EPscl、Npscl对供试细菌的抑制效果较好。组分EPsc4-1对枇杷炭疽病菌菌丝生长和孢子萌发具有较强的抑制活性,半数抑制浓度IC50分别达到0.119mg/mL和0.323mg/mL。经GC-MS分析发现组分EPsc4-1主要活性成可能为萘并[1,2-d]咪唑。通过电镜,观察了组分EPsc4-1对枇杷炭疽病菌菌丝形态和结构的影响。扫描电镜观察,菌丝呈不规则的肿大和缢缩,明显畸形,菌丝生长明显受到抑制。透视电镜结果表明,菌丝细胞壁不规则增厚,细胞原生质不均匀,有较大的空腔形成,囊泡增多,部分菌丝组织坏死。
In this study, ethanol extracts of mycelia and broth of Pleurotus sajor-caju through submerged fermentation were prepared and extracted by solves. In the active detection, crude extracts and its different extraction of the mycelium showed weak antibacterial and antifungal activities, but the broth was extracted in turn with the ethyl acetate and n-butanol and was found to have strong activity. Several components were botained from the extractive fractions by the following ways:silica gel column chromatography, thin layer chromatography and Sephadex LH-20 gel chromatography. Antibacterial and antifungal activities were found in some components through biological activity, of which the component EPscl and Npscl showed strong antibacterial activity. The component EPsc4-1 can remarkably inhibit hyphal growth and spore germination of Colletotrichum gloeosporioides, with the IC50 of 0.119mg/mL to hyphal growth and the IC50 of 0.323mg/mL to spore germination. The GC-MS analysis showed that the main active ingredient of EPsc4-1 is naphtho[2,1-d]imidazole. The effect of EPsc4-1 on morphology and structure of hyphal cell of C. gloeosporioides was investigated under scanning electron microscope(SEM) and transmission electron microscope(TEM). The studies revealed that mycelial growth of C. gloeosporioides was inhibited obviously, irregular hyphal swelling and constriction was clearly deformed. The hyphal cell wall was irregularly thickened. Uneven cell protoplasm and the formation of a large cavity were observed in hyphal cell. In addition, vesicles increased in hyphal cell. There are some mycelium tissue putrescence.
In this study, ethanol extracts of mycelia and broth of Pleurotus sajor-caju through submerged fermentation were prepared and extracted by solves. In the active detection, crude extracts and its different extraction of the mycelium showed weak antibacterial and antifungal activities, but the broth was extracted in turn with the ethyl acetate and n-butanol and was found to have strong activity. Several components were botained from the extractive fractions by the following ways:silica gel column chromatography, thin layer chromatography and Sephadex LH-20 gel chromatography. Antibacterial and antifungal activities were found in some components through biological activity, of which the component EPscl and Npscl showed strong antibacterial activity. The component EPsc4-1 can remarkably inhibit hyphal growth and spore germination of Colletotrichum gloeosporioides, with the IC50 of 0.119mg/mL to hyphal growth and the IC50 of 0.323mg/mL to spore germination. The GC-MS analysis showed that the main active ingredient of EPsc4-1 is naphtho[2,1-d]imidazole. The effect of EPsc4-1 on morphology and structure of hyphal cell of C. gloeosporioides was investigated under scanning electron microscope(SEM) and transmission electron microscope(TEM). The studies revealed that mycelial growth of C. gloeosporioides was inhibited obviously, irregular hyphal swelling and constriction was clearly deformed. The hyphal cell wall was irregularly thickened. Uneven cell protoplasm and the formation of a large cavity were observed in hyphal cell. In addition, vesicles increased in hyphal cell. There are some mycelium tissue putrescence.
引文
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