Study On The Secondary Metabolites Of Two Plant Endophytes

Microbial natural products have good biological activity and low toxicity,and have incomparable advantage over chemsynthetic drugs in potential druggability.With further study on secondary metabolites of microorganisms,the probability of discovering new active substances decreased gradually because of the repeatablity of microbial resources.Therefore,it has become an important research field to develop new microbial resources and find active compounds with novel structure from microorganism in special ecological environment.Symbiotic microbes derived from plant and animal could produce secondary metabolites with novelty structure and diversity activity,which are important sources for drug lead compoundsResearch on secondary metabolites of two endophytic fungi Alternaria sp.W-1 and Chaetomium globosum IFB-WQ has been carried out in this dissertation.Alternaria sp.W-1 was an endophytic fungus isolated from Laminaria japonica(collected in Weihai seaarea,China)W-1 was cultured by liquid fermentation and the fermentation broth was extracted with ethyl acetate to provide 20 g of crude extract.C.globosum IFB-WQ was an endophytic fungus isolated from a traditional medicinal plant Imperata cylindrical.IFB-WQ was then fermented in solid medium and the crude extract of fermentation broth was extracted with N-butanol to provide 17 g of crude extract.The obtained crude extracts of these two endophyte strains were then fractionated by silica gel column chromatography and gave several polarity fractions Guided by 1H-NMR spectrum and TLC,further separation and purification was performed through silica gel column chromatography,gel filtration over Sephadex LH-20,reversed phase chromatography and HPLC.Five compounds(1-5)were obtained from Alternaria sp.W-1.and five compounds(3 and 6-9)were obtained from C.globosum IFB-WQ.Structures of the obtained compounds were elucidated by modern spectroscopic analysis(MS,1D and 2D NMR)and comparison with literatures.Aforementioned compounds were identified as 6-iso-tricycloalterarene 6a(1),tricycloalterarene 6a(2),uracil(3),5-methyluracil(4),lumichrome(5),Chaetoglobosin E(6)and Chaetoglobosin Vb(7),respectively.Compounds 1 and 2 are two tricycloalterarene compounds and compound 1 was a new compound that had never been reported yet.Compounds 6 and 7 are two cytochalasin compounds.Compound 3 was isolated from both Alternaria sp.W-1 and C.globosum IFB-WQ.Strucures of compounds 8 and 9 need to be further determined.In vitro cytotoxicity of compound 2 against human hepatocarcinama cancer cell line SMMC-7721 was assessed by MTT method.The result showed that compound 2 inhibited the growth of SMMC-7721 cells with the inhibition rate of 44.95%at the concentration of 100?g/mL,while IC50 value of the positive control cisplatin was 8.64?g/mL.Cytochalasins is one kind of fungal secondary metabolites with varied structural types and bioactivity.Our previous experiments found that cytochalasin compounds have in vitro cytotoxicity against human liver cancer cell line SMMC-7721,human gastric cancer cell line SGC-7901 and human cervical cancer cell line Hela.Cytochalasin compounds were also found to have remarkable in vitro neuroprotective activity against Parkinson's disease model cells(MPP+-damaged PC 12 cells)and neonatal hypoxie-ischemic encephalopathy models cells(OGD-induced rat primary astrocytes).Cytochalasins might be one kind of neuroprotective reagent and have the potential in developing anti-PD and anti-NHIE drugs.Tricycloalterarene compounds are also a class of fungal secondary metabolites with multiple structural types and they showed obvious in vitro cytotoxicity to SMMC-7721 cells and SGC-7901 cells in our previous experiment.Among them,in vitro anticancer activity of TCA 3a was related to induced cell cycle arrest in G1 phase and induced apoptosis in SMMC-7721 cells.There are few pharmacological researches on tricycloalterarene compounds,which needs further study.Therefore,continues researches on isolation and purification,structural determination,and pharmacology activity of these two kindsO of fungal secondary metabolites could discover compounds with novel structures and outstanding activity and provide important experimental basis for the development of new drugs,which possess wide prospects in application.