一株祼脚菇属菌株产抗菌活性物的基础液体培养碳、氮源等优化及其对柑橘青绿霉菌的作用

Optimization of Carbon and Nitrogen Source of Basic Liquid Culture of Gymnopus sp. Producing Anti-fungal Activity Substance and Its Effect on Green and Blue Mold of Citrus

江西是柑橘类水果的主要产区之一,青绿霉菌是导致柑橘采后腐烂的主要病害,化学杀菌剂可产生病原抗药性和残留,一株具有拮抗此病菌的祼脚菇属菌株0612-9被发现。为了探索其产活性物质的代谢培养和活性作用,采用菌丝生长速率法和显微观察法测定了其抗菌活性,单因素试验、Plackett-Burman、最陡爬坡试验和Box-Behnken试验响应面设计等优化出液体培养基组成和培养条件。结果显示:液体培养基组成和培养条件为:马铃薯279.94g/L,葡萄糖30g/L,麸皮8.87g/L,MgSO_4·7H_2O 0.47g/L,KH_2PO_42g/L,发酵周期10d,接种量5 ml/100 ml,转速180r/min,摇床温度26℃,装液量100ml/250ml,发酵液抗病菌活性从42.26%提高到55.77%。对意大利青霉和指状青霉的EC_(50)分别为69.25g/L和56.70g/L,最小杀菌浓度250μg/ml低于阳性对照物抑霉唑的500μg/ml,并且对病原菌菌丝体有一定的毒害作用。该祼脚属菌株有潜在开发成柑橘采后保鲜剂的价值。

Jiangxi is one of the main producing area of citrus fruits. Green and blue mold are the cause of major diseases of post-harvest decay of citrus fruits. Chemical fungicides have generated pathogen resistance and residual. A macrofungal strain Gymnopus sp. 0612-9 with activity of antagonistic the pathogens was found. In order to investigate the metabolic culture of producing active substance and its action against Green and blue mold, the anti-fungal The medium composi activity was determined by the method of mycelium growth rate and microscopic observation. tion and culture conditions were optimized by the single factor test, Plackett-Burman, steepest ascent test and the response surface Box-Behnken design. The results showed the liquid medium composition and culture conditions are potato 279.94g/L,glucose 30g/L, bran 8.87g/L, MgSO4 · 7H20 0.47g/L, KH2PO4 2g/L; 10 days of fermentation period, inoculation volume 5ml/100ml, shaking speed 180r/rain, culture temperature 26℃, liquid medium volume 100ml/250ml with the anti-pathogen activity of the fermentation broth increased from 42.26% to 55.77%. The EC50 value against Penicillium italicum and Penicillium digitatum are 69.25μg/ml and 56. 70μg/ml, respectively. The minimum fungicidal concentration （MFC） is 250 μg/ml,which was lower than the positive control of imazalil of 500μg/ml and it has some toxic effects on the mycelium of the pathogens. The strain of Gymnopus sp. has potential value to develop into antiseptic agent of citrus fruits.