| Phlebopus portentosus is a new rare and precious edible and medicinal fungus in China,which is favored by consumers all over the world.The mycelium of Phlebopus portentosus is very easy to senescence,which seriously restricts the development of it’s industry.At present,there are relatively few studies on mycelial senescence of Phlebopus portentosus domestic and abroad,and the characteristics and physiological changes of mycelial aging are not clear.In this study,the medium optimization,vitality evaluation and aging cause analysis of its mycelium were carried out,in order to provide data reference for the strain maintenance of Phlebopus portentosus and the senescence mechanism.The main results are as follows:1.Through molecular identification,PH-21 was identified as Phlebopus portentosus.The optimum growth condition of mycelium was30℃,p H 6.5.There were significant differences in polysaccharides,triterpenoids,soluble protein content,total enzyme activity and specific activity of SOD between wild strains and commercial strains.The SOD of three different sources of Phlebopus portentosus were sensitive to H2O2 and remained relatively stable to"ethanol chloroform"solution.The highest activity condition of PH-21 SOD was 35℃,p H 5.5.The SOD activity stored at 4℃for a short time was relatively stable,with the extension of storage time,the SOD activity decreased rapidly.2.Through response surface analysis,the best addition amounts of inorganic salts are Fe SO4 1.28 mg/m L,Ca SO4 1.10 mg/m L and Mn SO40.81 mg/m L,the optimum amount of vitamin is VB1 128.5μg/m L,VB295.1μg/m L,VB3 94.8μg/m L.Compared with PDA medium,the contents of mycelial polysaccharides and triterpenoids in the optimized compound medium increased by 61.4%and 53.5%respectively,the free radical scavenging rates of DPPH and ABTS were increased by 89.2%and44.6%,respectively.The content of MDA in cells decreased significantly,the activities of SOD,POD and CAT decreased significantly.3.PH-21 aging mycelium showed significant browning,creeping,secretion and other properties.Microscopic examination showed that the cells had local swelling,melanin deposition on the cell wall and hyphal tangle.The secretion contains cell membrane and nucleic acid decomposition products,membrane lipid peroxide intermediates and other cell senescence related substances.The lipophilic substances in PH-21 secretion and fermentation broth had certain antibacterial effects on Staphylococcus aureus and Bacillus subtilis.UV-vis scanning showed that melanin had the maximum absorption peak at 210 nm.The liquid strain with low melanin concentration grew better.4.UPLC non targeted metabolomics analysis of PH-21 showed that the contents of lipids,phenols and amino acids in metabolites were high.There were 76 overlapping and different metabolites in mycelium of each mycelium stage,43 metabolites were down regulated in the whole stages,and 24 metabolites were up regulation and down regulation continued to change,mainly lipid and amino acid precursors,9 metabolites were up-regulated at all stages,including intermediates of cell aging markers such as methyl malonic acid,and substances related to cell antioxidant and stress functions such as guanidine acetic acid.The number of down regulated metabolites in mycelium was positively correlated with mycelium age.At 70 d and 42 d,the enrichment degree of many metabolic pathways changed,among which propionic acid metabolism and glutathione metabolism were significantly different from those of cellular antioxidant and functional protein synthesis. |
