两株共生菌的次生代谢产物研究

发布时间：2018-04-18 17:35

本文选题：共生菌 + 次生代谢产物　；参考：《扬州大学》2014年硕士论文

【摘要】：共生菌在与宿主长期协同进化的过程中,往往拥有比较特殊的代谢途径,能够产生丰富的结构多样性和生物学活性多样性的次生代谢产物。共生菌还具有可再生性,且生长周期短、代谢易于控制、菌种易于选育等诸多优势,因此,共生菌来源的天然产物研究越来越受到人们的重视。菌株IFB-WQ是南京大学功能生物分子研究所从白茅(Imperata cylindrical)中分离获得的一株内生真菌,根据形态学特征,鉴定为球毛壳菌(Chaetomiun globosum)。本论文课题对该菌株固体发酵产物的化学成分进行了研究。球毛壳菌IFB-WQ固体发酵产物经氯仿／甲醇(v/v1：1)混合溶剂浸提数次,减压去除溶剂,除油后得粗浸膏26g。粗浸膏经硅胶柱粗分得各极性组分；以1H-NMR谱和TLC示踪,组合使用硅胶柱层析、凝胶柱层析、高效液相色谱等方法对各组分进行分离纯化获得6个化合物。采用高分辨质谱、1D和2D核磁共振谱等现代波谱技术及文献比对的方法对化合物进行结构鉴定,上述化合物分别被鉴定为Chaetoglobosin F (1)、Chaetoglobosin Fex (2)、Chaetoglobosin E (3)、Cytoglobosins A (4)、Penochalasin C (5)和isochaetoglobosin D (6),它们皆为细胞松弛素类化合物。菌株IFB-WQ2是南京大学功能生物分子研究所从海螺(Busycon canaliculatu)中分离获得的一株放线菌,根据形态学特征,鉴定为链霉菌(Streptomyces sp.)。本研究对该菌株固体发酵产物的化学成分进行了分离和鉴定。放线菌IFB-WQ2固体发酵产物经氯仿／甲醇(v/v1：1)混合溶剂浸提数次,减压去除溶剂,除油后得粗浸膏35g。粗浸膏经硅胶柱粗分得各极性组分,以1H-NMR谱和TLC示踪,组合使用硅胶柱层析、凝胶柱层析、高效液相色谱等方法对各组分进行分离纯化获得4个化合物。采用高分辨质谱、1D和2D核磁共振谱等现代波谱技术及文献比对的方法对化合物进行结构鉴定,它们分别被鉴定为己六醇(7)、胡萝卜苷(8)、2-oxo-3-Pyrrolidinecarboxylic acid (9)和麦角甾醇(10)。采用MTT法测定了化合物1、3和5对人肝癌细胞株SMMC-7721和人宫颈癌细胞株HeLa的体外细胞毒活性,结果发现：化合物1、3和5对SMMC-7721细胞均具有一定的细胞毒活性,其IC50值分别为49.2μg/ml (0.092μmol/ml)、51.7μg/ml (0.097μmol/ml)和35.4μg/ml (0.069μmol/ml);化合物1、3和5对HeLa细胞也均具有一定的细胞毒活性,其IC50值分别为52.7μg/ml (0.099μmol/ml)、12.6μg/ml (0.024μmol/ml)和43.9μg/ml (0.086μmol/ml)。此外,还开展了化合物1-6的体外清除DPPH抗氧化应激活性研究,结果显示：化合物1和6具有很强的抗氧化活性,化合物1的IC50值为0.002μmol/ml,化合物6的IC5o值为0.002μmol/ml,化合物2-4具有弱的抗氧化活性,化合物5无抗氧化活性。天然产物是药物先导化合物的丰富来源,本研究从白茅内生菌C. globosum IFB-WQ中分离获得的细胞松弛素类化合物既具有体外抗肿瘤活性,又具有显著的体外抗氧化活性,值得深入研究。
[Abstract]:In the process of long-term co-evolution with the host, symbiotic bacteria often have a special metabolic pathway, which can produce abundant secondary metabolites of structural diversity and biological activity diversity.Symbiotic bacteria also have many advantages such as reproducibility, short growth cycle, easy control of metabolism, easy selection and so on. Therefore, people pay more and more attention to the study of natural products from symbiotic bacteria.Strain IFB-WQ is an endophytic fungus isolated from Imperata Cylindricalus from the Institute of functional Biomolecular, Nanjing University. According to its morphological characteristics, it was identified as Chaetomiun globosumsuma.In this paper, the chemical constituents of the solid fermentation products of the strain were studied.The solid fermentation product of IFB-WQ was extracted with chloroform / methanol v / v 1: 1 mixed solvent for several times, then the solvent was removed by decompression, and the crude extract was obtained after removing oil.Six compounds were isolated and purified by silica gel column chromatography, gel column chromatography and high performance liquid chromatography with 1H-NMR spectrum and TLC tracer.The structures of the compounds were identified by means of modern spectral techniques such as high resolution mass spectrometry 1D and 2D nuclear magnetic resonance spectroscopy and literature comparison.The above compounds were identified as Chaetoglobosin F ~ (1) and isochaetoglobosin D ~ (6), which are cytochalasins.The strain IFB-WQ2 was isolated from Busycon canaliculatu by the Institute of functional Biomolecular, Nanjing University. It was identified as Streptomyces sp.In this study, the chemical constituents of the solid fermentation product of the strain were isolated and identified.The solid fermentation product of actinomycetes IFB-WQ2 was extracted with chloroform / methanol v / v 1: 1) mixed solvent several times, the solvent was removed by decompression, the crude extract 35 g was obtained after removing oil.Four compounds were isolated and purified by silica gel column chromatography, gel column chromatography and high performance liquid chromatography with 1H-NMR spectrum and TLC tracer.The structures of the compounds were identified by modern spectroscopic techniques such as 1D and 2D NMR spectra and literature comparison. They were identified as hexadecanol 7, daucoside 2 oxo-3-Pyrrolidinecarboxylic acid 9) and ergosterol 1010, respectively.The cytotoxic activities of compounds 1t3 and 5 on human hepatoma cell line SMMC-7721 and human cervical cancer cell line HeLa in vitro were determined by MTT assay. The results showed that compounds 1t3 and 5 had cytotoxic activity to SMMC-7721 cells.Their IC50 values were 49.2 渭 g/ml (0.092 渭 mol / ml) and 35.4 渭 g/ml (0.069 渭 mol / ml), respectively, and their IC50 values were 52.7 渭 g/ml 0.099 渭 mol / ml 12.6 渭 g/ml 0.024 渭 mol / ml and 43.9 渭 g/ml 0.086 渭 mol / ml, respectively.In addition, the anti-oxidative stress activity of compounds 1-6 was also studied in vitro. The results showed that compounds 1 and 6 had strong antioxidant activity.The IC50 value of compound 1 was 0.002 渭 mol / ml, and the IC5o value of compound 6 was 0.002 渭 mol / ml. Compound 2-4 had weak antioxidant activity, while compound 5 had no antioxidant activity.Natural products are rich sources of drug leading compounds. The cytochalasins isolated from C. globosum IFB-WQ in this study not only have antitumor activity in vitro, but also have obvious antioxidant activity in vitro, which is worthy of further study.
【学位授予单位】：扬州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：Q936

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