暗黑链霉菌PY-1抑菌活性产物的结构鉴定及其对葡萄霜霉病菌的抑制作用

 从暗黑链霉菌(Streptomyces atratus)PY-1菌株发酵液中分离鉴定抑菌活性产物,评价其对葡萄霜霉病菌(Plasmopara viticola)的抑制作用,为该菌株及其产生的抑菌活性物质的应用提供依据。PY-1菌株发酵液经二氯甲烷萃取,减压浓缩获得抑菌粗提物,稳定性测定结果显示,避光条件下,15℃~65℃、pH 6~10、4℃或常温保存6个月,抑菌活性稳定。采用硅胶柱层析、Sephadex LH-2柱层析、薄层层析和HPLC等技术,对抑菌粗提物进一步分离纯化,得到2种抑菌活性组分;采用ESI-MS、¹H NMR、¹³C NMR等波谱分析技术,对活性组分进行结构鉴定,2种活性组分分别鉴定为5-乙酸环已基酰亚胺(5-acetoxycycloheximide)和环己酰亚胺(cycloheximide)。采用离体叶片法测定2种组分的抑菌活性,不同浓度(10⁴ ng·mL^-1、10² ng·mL^-1、1 ng·mL^-1)的5-乙酸环已基酰亚胺和环己酰亚胺对葡萄霜霉病菌的抑制作用分别为92.59%、86.30%、64.81%和97.04%、91.85%、84.07%。

Structural identification of active compounds from Streptomyces atratus PY-1 and their inhibitory activities against Plasmopara viticola

Separation and identification of the bioactive product from the fermentation medium of Streptomyces atratus strain PY-1 with following the evaluation of inhibition on Plasmopara viticola are essential to further application of the strain itself and the products to control grapevine downy mildew. The crude products with inhibition activity were extracted from the fermentation medium by methylene dichloride with subsequent vacuum condensation, which exhibited stable inhibitory activity against P. viticola under the condition of dark, at 15℃-65℃, pH 6-10, and 6 months storage at 4℃ or room temperature. After the further purification conducted by silica gel column chromatography, Sephadex LH-2, thin-layer chromatography and HPLC methods, we got two components with more stronger inhibitory activity. The two metabolites were identified as 5-acetoxycycloheximide and cycloheximide by means of ESI-MS, ¹H NMR, ¹³C NMR and spectra analysis. The antagonistic activities were then determined by detached-leaf bioassay. The inhibitions of Plasmorpara viticola were calculated at 92.59%, 86.30%, and 64.81%, respectively, when 5-acetoxycycloheximide was diluted to 10⁴ ng·mL^-1, 10² ng·mL^-1 and 1 ng·mL^-1, while cycloheximide had inhibited effects at 97.04%, 91.85% and 84.07% with the same dilution.