植物内生阿维链霉菌I06A-01716次生代谢产物的研究

Study on metabolites produced by Streptomyces avermitilis strain I06A-01716

目的分离鉴定阿维链霉菌I06A-01716发酵液中抗植物病原真菌活性成分,并进行抗真菌、抗肿瘤和免疫抑制活性研究。方法在抗真菌活性指导下,采用硅胶柱层析、凝胶柱层析、HPLC等分离手段对活性组分进行分离;通过高分辨质谱,1D-NMR和2D-NMR对其结构进行鉴定;采用琼脂平板纸片法,进行抗植物病原真菌的最低抑制浓度测定;采用MTT法检测其对肿瘤细胞生长的抑制作用;采用流式细胞仪测定其对细胞周期的影响。结果分离得到2个活性组分:1716-1和1716-2。1716-1和1716-2对人肝癌细胞HepG2的IC50值分别为2.42μg/mL和3.42μg/mL;对人口腔上皮癌细胞KB的IC50值分别为1.64μg/mL和6.73μg/mL。细胞周期研究表明,随着1716-2的浓度增加,在KB细胞中,G2/M期比例由对照组的15.23%逐步升高到36.53%;在HepG2细胞中,G2/M期比例由对照组的4.39%升高到35.78%。结论1716-1和1716-2结构分别与寡霉素B和寡霉素A一致。1716-1和1716-2有抗多种植物病原真菌活性。1716-1和1716-2在较低浓度下能够抑制人肝癌细胞HepG2和人口腔上皮癌细胞KB生长。1716-2使HepG2细胞和KB细胞阻滞在G2/M期。

Objective To isolate, purify and identify the antifungal components from the fermentation broth ofStreptomyces avermitilis strain I06A-01716, and to study their biological activities. Methods Different column chromatography methods were used to isolate and purify bioactive components under guidance of antifungal assay; HR-ESI-MS and, 1D-NMR, 2D-NMR were analyzed to elucidate the structures of the active components; Antifungal test against plant pathogenic fungi were tested by agar diffusion method. The cell growth inhibitory effects of 1716-1and 1716-2 on KB cell and HepG2 cell lines were detected by MTT assay; Effects of 1716-2 on cell cycle of KB cell and HepG2 cell were examined by flow cytometry. Results Tow bioactive components： 1716-1 and 1716-2 were purified. The IC50 of HepG2 and KB to 1716-1 or 1716-2 respectively were 2.42μg/mL, 1.64μg/mL or 3.42μg/mL, 6.73μg/mL. As the concentration increasing of 1716-2, in KB cells, the proportion of G2/M phase rise to 36.53% from 15.23%; in HepG2 cells, the proportion of G2/ M phase go up to 35.78% from 4.39%. Conclusion 1716-1 and 1716-2 were identified as Oligomycin B and Oligomycin A. 1716-1 and 1716-2 significantly inhibited the growth of several plant pathogenic fungi. 1716-1 and 1716-2 can inhibit the growth of both HepG2 and KB cells in low concentrations. The arrest at G2/M phase was detected by flow cytometry in both HepG2 and KB cells after 1716-2 treatment with different concentrations.