| According to the records,morchella is a kind of precious fungus that widely be used in edible and medicinal purpose for its taste and function.Modern studies show that Morchella is rich in polysaccharides,fatty acids,amino acids,vitamins,minerals,sterols,organic acids and other chemical components,with anti-tumor,immune regulation,lowering blood lipid,liver protection,anti-oxidation and other pharmacological effects.In recent years,with the development of cultivation technology,many of its varieties can be cultivated artificially,but the yield is not high for the expensive cultivation cost,it is also difficult to achieve large-scale cultivation.In order to provide a theoretical basis for the development and utilization of Morchella,structure analysis and biological activity of the fermentation products were studied by liquid fermentation.A Morchella esculenta strain was used as material,and fermented in potato glucose medium and potato glucose bran bean sprouts medium for 18 L,respectively.The fermentation was divided into three batches,the first batch and the second batch were fermented for 20 days,and the third batch were fermented for 187 days.After removing the mycelium from each batch of fermentation broth,the organic crude extracts were prepared by extraction with ethyl acetate in equal proportion.Each batch of crude extract was retained20 mg for the analysis of secondary metabolites,and the remaining extracts were combined with a total of 8.8031 g for the separation and purification of the compounds.Twelve compounds,JHL01,JHL02,JHL03,JHL04,JHL05,JHL06,JHL07,JHL08,JHL09,JHL10,JHL11 and JHL12,were isolated by silica gel column,silica gel column and Sephadex LH-20 column chromatography.According to NMR spectra,the structures of compounds JHL02,JHL03,JHL04,JHL05,JHL07,JHL09 and JHL10 were respectively analyzed as follows:4,5,7-trihydroxy-3-methoxyisochroman-1-one,N,N'-(pentane-1,5-diyl)diacetamide,Methyl 3,5-dihydroxy-2-(1-hydroxy-2-oxoethyl)benzoate,(3a S,8a R)-1-hydroxy-3,3a,8,8a-tetrahydro-1H-cyclobuta[4,5]cyclohepta[1,2-c]furan-4,7-dione,3,5-dihydroxybenzoic acid,1H-Pyrrole-2-carboxylic acid,7-methoxy-4-methylpyrido[2,1-c][1,4]oxazine-3,6(1H,4H)-dione.Among them,compounds JHL02 is a chromone derivative,JHL04 is a benzoic acid derivative,JHL05 is a furanone substance,and JHL10 is a ketone derivative.After searching,compounds JHL02,JHL04,JHL05 and JHL10 are new structural compounds.HPLC analysis of secondary metabolites showed that the yield of compound JHL01 in PDP medium decreased gradually with the prolongation of fermentation time,and the yield of compound JHL01 was always lower than that of compound JHL02 On the contrary,the yield of compound JHL01 was higher than that of compound JHL02 in PD medium even if the fermentation time was prolonged.The compound JHL10 appeared only at 187 d of fermentation in both PD and PDP media.The results indicated that different medium and fermentation time would obviously affect the liquid fermentation products of Morchella esculenta.The results showed that JHL01,JHL05 and JHL07 had strong antioxidant activity,and JHL02,JHL04,JHL08,JHL09 and JHL10 also had slight antioxidant activity when the concentration of compounds was 0.04 mg.In addition,at the final concentration of 0.2mg/mL,JHL10 showed antibacterial effect against Escherichia coli and Staphylococcus aureus,and the inhibitory rate of JHL05 against acetylcholinesterase was 10.79%.The results of this study showed that the secondary metabolites with various structures and certain biological activity were produced by liquid fermentation of Morchella esculenta,and the liquid fermentation products were significantly affected by different media and fermentation time.The results of this study provide a new way for the development and utilization of the chemical resources of Morchella esculenta. |
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