节点文献
海绵共附生系状菌中生物活性物质的研究
Studies on the Bioactive Metabolites of Sponge-associated Fungi
【作者】 盖悦;
【导师】 胡昌奇;
【作者基本信息】 复旦大学 , 药物化学, 2007, 博士
【摘要】 本文以稻瘟霉(Pyricularia oryzae)分生孢子和菌丝形态变异或生长抑制为指标,从海绵共附生系状菌的发酵产物中筛选具有抗真菌和抗肿瘤等生物活性的物质。通过对100株海绵来源的系状真菌的发酵液进行抗稻瘟霉活性筛选,得到9株具有良好抗稻瘟霉活性且稳定的菌株。采用代谢产物的提取分离和活性检测同步进行的方法,对活性最好的菌株,即05JANF163/165、05JANF151、05JANF154、05ABR145、05ABR116、98F152和S1-1,进行了大量发酵,从其发酵液中共分离得到20个化合物,根据理化性质和各种波谱学等方法鉴定了19个单体化合物的结构,分别为Fusarielin A(1)、Fusarielin B(2)、Fusarielin E(3)、(-)-Oxysporidinone(4)、Roridin A(5)、Roridin H(6)、Roridin J(7)、Verrucarin A(8)、Verrucarin B(9)、Verrucarin L Acetate(10)、Verrucarin J(11)、Deoxyfusapyrone(12)、5′-Hydroxyzearalenol(13)、Zearalenol (14)、Zearalenone(15)、8′-Hydroxyzearalenone(16)、Zeaenone(17)、α,β-Dehydrocurvularin(18)、MycophenolicAcid(19)。其中化合物(3)和(13)为新的天然产物,并首次得到了化合物(6)和(7)的X-射线单晶衍射的立体结构。化合物(1)和(3)能使稻瘟霉分生孢子发生严重变形或使菌丝生长发生弯曲变形,最小抑制浓度均为3.1μg/mL;化合物(5)、(6)、(9)和(11)能使稻瘟霉菌丝发生弱卷曲变形,它们的最小抑制浓度分别为12.5μg/mL、3.1μg/mL、0.2μg/mL和0.2μg/mL;化合物(12)能使稻瘟霉菌丝发生弱卷曲强串珠状变形,其最小抑制浓度为3.1μg/mL;化合物(4)、(7)、(8)、(10)、(15)、(17)、(18)和(19)抑制稻瘟霉菌丝生长的最小抑制浓度分别为25μg/mL、6.2μg/mL、1.6μg/mL、0.2μg/mL、3.1μg/mL、0.1μg/mL、25μg/mL和25μg/mL。利用稻瘟霉生物活性模型筛选出的这些生物活性化合物同文献中报道的有关它们的其它生物活性试验结果有很好的一致性,这也说明该生物模型能够有效的指导海洋生物活性物质的分离纯化,是一个适用的初筛体系。
【Abstract】 Bioactive substances that interfere with microtubule function can generally inhibit the conidia growth of Pyricularia oryzae (a plant pathogenic fungus) and induce morphological deformation of conidia or mycelia of Pyricularia oryzae by swelling, curling, beads formation and hyper-divergence effect. We apply the method using conidia of Pyricularia oryzae as a primary screening assay for compounds such as antifungal antibiotics and anticancer substances from secondary metabolites of sponge-associated fungi. From 100 fungi samples isolated from sponges, we obtained 9 strains possessing better and stable activity. From the fermentation broth of strain 05JANF163/165, 05JANF151, 05JANF154, 05ABR145, 05ABR116, 98F152 and S1-1 , twenty active compounds were isolated and purified by activity-guided fractionation. Structures of nineteen compounds were established on the basis of various NMR spectroscopic analyses, including 1D-and 2D-NMR spectroscopic analyses (1H-1H COSY, ROESY, HMQC, and HMBC spectra), and MS (FAB, EI and ESI). The nineteen compounds were Fusarielin A (1) , Fusarielin B (2) , Fusarielin E (3) , (-)-Oxysporidinone (4) , Roridin A (5) , Roridin H (6) , Roridin J (7) , Verrucarin A (8) , Verrucarin B (9) , Verrucarin L Acetate (10) , Verrucarin J (11) , Deoxyfusapyrone (12) , 5’-Hydroxyzearalenol (13) , Zearalenol (14) , 8’-Hydroxyzearalenone (15) , Zearalenone(16) , Zeaenone (17) ,α,β-Dehydrocurvularin (18) and Mycophenolic Acid (19) . Compounds (3) and (13) were new nature compounds, and compounds (6) and (7) were first elucidated their spectroscopic structures by X-ray single-crystal analysis.Compounds (1) and (3) inhibited the conidia growth of Pyricularia oryzae by swelling effect and induced morphological changes of the mycelia by curling effect with the minimum inhibitory concentration (MIC) 3.1μg/mL. Compounds (5) , (6) , (9) and (11) induced morphological changes of the mycelia by weak curling effect with MIC 12.5μg/mL, 3.1 μg/mL, 0.2μg/mL and 0.2μg/mL, respectively. Compound (12) induced morphological changes of the mycelia by weak curling and strong beads formation with MIC 3.1μg/mL. Compounds (4) , (7) , (8) , (10) , (15) , (17) , (18) and (19) inhibited the mycelia growth with MIC 25μg/mL, 6.2μg/mL, 1.6μg/mL, 0.2μg/mL, 3.1μg/mL, 0.1μg/mL, 25μg/mL and 25μg/mL, respectively.The activity results of our study accorded with the relevant reports and it showed that the bioassay method using conidia of Pyricularia oryzae could efficiently guide the isolation of the bioactive metabolites of marine-derived fungi and could be a primary screening for bioactive substances.