探討本土性黑殭菌MA-126所含二次代謝物及其抗登革熱病毒之研究

黑殭菌(Metarhizium anisopliae)為一種對昆蟲具有致病能力之昆蟲寄生性真菌，其分布於世界各地。主要用途是作為預防病蟲害保護農作物的生物性農藥。根據先前研究，黑殭菌能產生相當有潛力的二次代謝物，包含抗癌與抗病毒活性。本研究中將黑殭菌MA-126分別以固、液態發酵培養，進而用乙酸乙酯和水進行分配萃取後，取乙酸乙酯萃取物針對登革熱病毒之NS2B蛋白進行抗病毒測試，發現該萃取物於濃度為50μg/ml時對NS2B蛋白分別具有35%、10%的抑制率。兩者乙酸乙酯層使用LH-20管柱層析、高效能液相層析進行純化，從液態發酵培養萃取物中分離出4個天然物，(5S,8S,9R)-8-benzoyl-9-hydroxy-8-methoxy-3-methyl-2-(5-methylfuran-2-yl)-1-oxa-7-azaspiro[4.4]non-2-ene-4,6-dione(1),(3S,3aR,7aS)-3a,7a-dihydroxy-3-((S)-1'-hydroxy-4'-methylpent-3'-en-1'-yl)-4-methoxy-3-methylhexahydroisobenzofuran-5(3H)-(2),chlovalicin(3), (4S,4aR,5S,8aR)-4,4a,8a-trihydroxy-5-methoxy-4-methyl-3-(3'-methylbut-2'-en-1'-yl)octahydro-6H-isothiochromen-6-one(4)，從固態發酵培養萃取物中分離出4個天然物，Destruxin A(5), Destruxin B(6), Destruxin E chlorohydrin(7)和Destruxin E diol(8)。上述天然化合物的結構鑑定，均採用各種核磁共振圖譜技術，包括1H、13C、DEPT、COSY、HMQC、HMBC和NOESY配合物理數據 (質譜、旋光值、紅外線光譜)，並且對照文獻上相關化合物的數據來定立。其中化合物1為新化合物，化合物1-4是首次於黑殭菌中分離出的天然化合物。

關鍵字

黑殭菌；固態發酵培養；液態發酵培養； NS2B蛋白

並列摘要

Metarhizium anisopliae is a kind of entomopathogenic fungi on insects, it is distributed all over the world. The main application for this fungi is as a pest control for crops, acting as a biological pesticide. In previous studies, Metarhizium anisopliae was able to produce a large quantity of secondary metabolites containing anticancer and antiviral activities. In this study, Metarhizium anisopliae MA-126 was cultured into solid-state fermentation and liquid state fermentation. After partition extraction, the NS2B protein had 35% and 10% inhibition rate when both of them have 50μg/ml concentration on ethyl acetate layer. The ethyl acetate layer was purified by LH-20 column, high performance liquid chromatography (HPLC) to yield eight compounds. Compound1-4 are separation from liquid state fermentation(5S,8S,9R)-8-benzoyl-9-hydroxy-8-methoxy-3-methyl-2-(5-methylfuran-2-yl)-1-oxa-7-azaspiro[4.4]non-2-ene-4,6-dione(1), (3S,3aR,7aS)-3a,7a-dihydroxy-3-((S)-1'-hydroxy-4'-methylpent-3'-en-1'-yl)-4-methoxy-3-methylhexahydroisobenzofuran-5(3H)-(2), chlovalicin(3), (4S,4aR,5S,8aR)-4,4a,8a-trihydroxy-5-methoxy-4-methyl-3-(3'-methylbut-2'-en-1'-yl)octahydro-6H-isothiochromen-6-one(4), compound 5-8 are separation from solid state fermentation. Destruxin A(5), Destruxin B(6), Destruxin E chlorohydrin(7) and Destruxin E diol(8). 　　All structures were elucidated by NMR spectra techniques, including 1H, 13C, DEPT, COSY, HMQC, HMBC, and NOESY, with physical data(mass spectrometry, optical rotation, IR spectrometry), also compare with the compound data in the literature to establish.Compound 1 is the new compound, compound 1-4 are the first natural compounds isolated from Metarhizium anisopliae.