Study On Chemical Composition And Antibacterial Activity Of Pholiota Nameko Based On OSMAC Strategy

The natural products with high chemical structure in higher fungi provide a material basis for the discovery and discovery of drug lead compounds.This paper studies the species of Pholiota nameko belonging to the Basidimy coat,Hymenomycetes,Agarieales,and Pholiota.its chemical composition and antibacterial activity were studied.Because the mushroom is sensitive to environmental changes,different structural types of compounds can be produced under different culture conditions.In order to make full use of the mushroom resources and obtain more types of compounds,a strain is used to produce a variety of compounds"(One The strategy of strain-many compounds(OSMAC)is to optimize the fermentation of the strain by changing the medium composition and ratio,culture temperature,culture time,shaking speed,illumination,addition of biosynthetic pathway precursors and key enzyme inhibitors.Conditions to fully exploit the potential of secondary metabolites of microbial strains in order to obtain more compounds with novel structural or bacteriostatic activity from the mushroom.In this paper,the OSMAC strategy was first used to design 14 different fermentation conditions for fermentation of the mushroom.The chemical composition of the fermentation product was analyzed by HPLC.The chemical composition of the same strain under different fermentation conditions was found to be significant.The three metabolites of the three kinds of fermentation conditions are abundant:(1)Fermentation conditions 5: Adding fungal secondary metabolite precursor material caryophyllene to GP medium,constant temperature at 24 ° C,rotation speed 150 r / min,shaking culture for 7 days,adding substrate caryophyllen in shaker culture 7 After 7 days,it was allowed to stand at room temperature for 7 days(2 cycles);(2)Fermentation conditions 6: 75 ml of rice medium and 200 mL of ultrapure water for mushroom culture,standing at room temperature for 60 days;(3)Fermentation conditions 10: The mushroom was cultured in an optimized GP medium,24 ° C,150 r / min,shake flask dark culture for 30 days.Subsequently,the chemical constituents were separated,purified and structurally identified from the ethyl acetate extracts of the three kinds of fermentation conditions.The experiments were carried out by forward and reverse phase column chromatography,Sephadex LH-20 gel column chromatography and analysis,and semi-preparative high performance liquid chromatography.The spectra were analyzed by various spectral methods including nuclear magnetic resonance(1D NMR and 2D).NMR),ultraviolet(UV),mass spectrometry(ESI and HRESI),infrared(IR),optical rotation(ORD),circular dichroism(CD),X-ray single crystal diffraction(X-Ray),etc.Compounds were identified for structure.Results are as follows :1.Six compounds were isolated and identified from the extract of ethyl acetate in the fermentation broth obtained under fermentation condition 6,compounds Pyrophen(2),Bicyclo[1.1.0]butane-4-hydroxyl-2-[Benzeneacetic acid,2’-hydroxy-3’,5’-dimethyl-],methyl ester(4),12,15-dihydroxy-cubenol(6),Donacinol(7),(2R,5R,6S,9R)-6,9-epoxy-2,6,10-trimethylhendeca-1,5,10-triol(8),(22E,24R)-Ergosta-7,22-dien-3β,5α,6β-triol(15),respectively.Among them,two new compounds are compounds 4 and 6,respectively.The absolute configuration of compound 2 is determined by X-Ray single crystal diffraction.The compound type involves benzene ring derivatives,sesquiterpene,sterol,etc.2.Ten compounds were isolated and identified from the extract of ethyl acetate in the fermentation broth obtained under fermentation condition 10.which were compounds(3R,6S,7S,8R,10S)-3,7,14-trihydroxy-1-sterpurene(1),3βH-7βH-3,11-dihydroxyeremophil-1(10)-en-2-one(3),12,13,14,15-tetramethyl-4-hydro-xy-2-en-6-oxo-hexahydro-1H-Indene(5),Butanedioic acid,2-hydroxy-2-(1-methylethyl)-1,4-dimethyl ester(9),1,3-Diethyl ester-2-methyl ester citric acid(10),Diethyl(2R)-2-hydroxy-2-methylsuccinate(11),Succinic acid(12),β-Sitosterol methyl ether(14),β-sitosterol(17),Daucosterol(18).Among them,there are 3 new compounds(1,3,5).The types of compounds involve sterpurane type sesquiterpenes,cadinane type sesquiterpenes,lignin-type sesquiterpenes,acid esters,sitosterol and ergosterol.The absolute configuration of Compound 1 was determined by X-Ray single crystal diffraction,and the absolute configuration of Compound 3 was determined by ECD calculation.3.Two sterol compounds were isolated from the extract of ethyl acetate extracted from fermentation broth 5,respectively.compounds(22E,24R)-Ergosta-7,22-dien-3β-methoxy(13),(22E,24R)-Ergosta-7,22-dien-3β-ol(16).Finally,the preliminary antibacterial activity of a large amount of monomeric compounds isolated from the mushroom was tested by K-B method..The bacteriostatic strains include Grampositive bacteria Staphylococcus aureus,Pseudomonas aeruginosa,Candida albicans,and Gramnegative bacteria Escherichia coli.The results showed that the inhibitory circles of compounds 1,2,5,12,14,15 and 17 were all less than 7 mm in diameter,and had no obvious antibacterial activity against these four pathogenic bacteria.