Alkaloid And Huperzine A Production From Endophytic Fungi Isolated From Huperziaceae

One hundred and twenty-seven endophytic fungal strains were obtained from health tissues of Phlegmariurus fordii, Phlegmariurus petiolatus and Huperzia serrata. Preliminary studies indicated that from one hundred and twenty-seven strains, twenty-eight strains were found to produce alkaloids using fermentation and alkaloid screening condition. Through the screening of acetylcholinesterase inhibitors, ten strains of endophytic fungi producing acetylcholinesterase inhibitors were selected from the twenty-eight strains which produced alkaloids. Based on acid dye colorimetry, total alkaloid production of ten endophytic fungal strains were investigated. The results showed that the highest absorbance was obtained from fermination broth of strain YS-02, and strain YS-13 was the second. The amount of Huperzine A (HupA) produced by strains SY-02 and YS-13 were quantified by HPLC, which showed that they produced 26.4μg/L and 21.0μg/L of HupA, respectively. And further confirmative analyses were done by mass spectroscopy contrast with authentic HupA. Morphology and ITS1-5.8S-ITS2 sequence analysis showed that strains YS-02 and YS-13 were Xylariales and Paecilonmyces tennis, respectively. It was the first report that Xylariales and P. tennis produced HupA. Study on the expansion of cultivation of P. tennis, YS-13 was done by automatic bio-fermentation tank. The fermentation product analysis showed that the endophytic fungal strain P. tennis, YS-13 generate HupA in vitro, producing 20.0μg HupA per liter liquid medium, on the basis of HPLC assays. And the relevant parameters of growth curves were obtained.