Isolation,identification And Their Anti-tumor Activities Of Endophytic Fungi From Ginger And Other Plants

With the development of society and economy,environmental pollution is getting worse and worse,the incidence of cancer is increasing at an alarming rate which has become the second killer to human.Endophytes can produce the same or similar metabolites as their host during long-term cooperative coevolution and widely distributed in plants,so isolating endophytes whose secondary metabolite could show the similar inhibitory activity has become an important strategy on the research of anti-tumor drugs.In the microecosystem of medical plants,the endophytic fungi established a long-term harmonious and symbiotic relationship with its host.Its secondary metabolite are abundant,and it shows many biological activities including antibacterial,anti-tumor activities and so on,and it also can promote plant growth.Endophytes from medical plant have potential appliance value,and it is one of efficient ways to screen ingredients or compounds with biological activities.The object of the study is four kinds of medicinal and edible plants.The endophytic fungi are isolated from plants by plate method and purified by the streak plate method.The endophytic fungi which have anti-tumor activity are screened out,and some strains that behave stonger anti-tumor activity were identified by morphology and molecular biology.The summary is as follows:1.Isolation and purification of the endophytic fungi:58 strains of endophytic fungi was isolated and purified from ginger,lilium,yam and momordica charantia L..2.Morphological identification of endophtic fungi:46 strains of endophytic fungi was identified by morphology primarily.It turns out that they respectively belong to Penicillium sp.,Saccharomyces sp.,Fusarium sp.,Aspergillus sp.,Trichoderma sp.and so on.The species resources of these endophtic fungi are abundant,and BH-F strains belong to Trichoderma sp..There are no spore of BH-A observed on the medium,so that we can't identify the strain by morphological identification.3.The fermentation of endophytic fungi and the treatment of its fermented products:58strains of endophytic fungi were fermented in potato dextrose liquid medium,the fermentation products of each strains were extracted by EtOAc and the mycelium was extracted by ethanol.At last,there are 60 crude extract samples were obtained.4.Screening strains with the anti-tumor activity:5 tumor cells including A549,Hela,HepG2,SGC7901,SGC7901/ADR were selected as subject cell.Firstly,using MTT method to screen out the active strains with anti-tumor activity from the 60 samples.It turns out that18 samples had shown cytotoxic activity against at least one tumor cell lines,which accounts for 30%of the total sample and belong to 14 kinds of strains.Then 14 kinds of active strains were selected again,and IC₅₀0 of every strain to every tumor cell are calculated.The results are that BH-A strains and BH-F strains show stronger anti-tumor activity than others.5.Molecular biology identification of active strains:BH-A and BH-F were identified by molecular biological methods.Their ITS fragment was amplified by PCR.After blasting through the GenBank data base.Ultimately,the strain BH-A and strain BH-F was identified as Trametes versicolor and Trichoderma sp.