千层塔中产石杉碱甲内生真菌的研究

Study on Huperzine A-Producing Endophytic Fungi Isolated from Huperzia Serrata

从千层塔中分离产石杉碱甲内生真菌。采用平板分离法分离千层塔内生真菌,通过抑制乙酰胆碱酯酶活性和抗氧化活性筛选,结合薄层层析和HPLC测定活性菌株中的石杉碱甲含量,通过形态学ITS-rDNA序列分析鉴定内生真菌。共分离得到94株内生真菌,筛选到S29、L44、S94共3株乙酰胆碱酯酶抑制活性和清除DPPH·自由基活性都在50%以上,筛选到一株S29能产石杉碱甲的菌株。摇瓶发酵产量为每克干菌体的石杉碱甲含量为50.64μg。形态学与ITS-rDNA序列分析法鉴定该内生真菌为Podospora sp.属。药用植物千层塔体内Podospora sp.属的内生真菌可产石杉碱甲。

In this study,huperzine A-producing endophytic fungi were isolated and purified from Huperzia serrate using plate separation method. The endophytic fungi were fermented in liquid culture medium. The DPPH · scavenging activity and AchE inhibitory activity of the fermented product were determined. In addition, the fermented products were analyzed by TLC and HPLC for the presence of huperzine A （HupA）. The morphological characters and ribosomal DNA （rDNA） sequence of the fungi were also analyzed. The results showed that 94 endophytic fungi were isolated from H. serrate, in which strains S29, L44, S94 owned the significant AchE-inhihitory and DPPH. scavenging activities. Strain S29 was found to produce HupA with a yield of 50. 64μg in one gram dry mycelium. Morphology and ITS sequence analysis showed that strain S29 was a Podospora species.