Studies On Bioactive Compounds Production By Submerged Fermentation Of Phellinus Baumii

Phellinus baumii, a highly-valued medicinal mushroom, which is always being named Sang huang Gu, Sang chen,Sang er in Chinese. More recent research has shown Phellinus sp. inhibited cancer cell growth, antioxidant, anti-hepatic fibrosis function and possessed immunomodulatory properties.At present, many researches were focused on the chemical composition, structure, pharmacology of polysaccharides and proteoglycan of fruiting bodies. Some reports also showed the activity components of the mycelia extracts were similar to the fruit bodies. The cycle culture time of fermentation is shorter than that of planting and the cost of fermentation is cheaper than that of planting, furthermore, the original material is easy to be obtained. Therefore, it is the better choice to produce bioactive Compounds Production by Submerged Fermentation. In order to offer technique support, the identification and screening of Phellinus sp. strains, the optimum of submerged fermentation parameters, and the activity of extracts were carried out.There are three parts in this dissertation. Study on the identification and screening of strains, the optimum of submerged fermentation parameters, and the activity of extracts were discussed in this dissertation.1 Studies on screening of strainsThe four strains, PB-10, PI-12, MYSH and NFSH belong to Phellinus genus by ITS sequence analysis. Based on the ITS sequence analysis and Antagonistic effect, the four strains are denified as Phellinus baumii, Phellinus baumii, Phellinus linteus, Phellinus badius. Taking the mycelia growth rate, biomass, content and yield of total flavones, content and yield of polysaccharide, as detecting indexes, the four strains were screened. The results showed that the four detecting indexes of PB-10 were 5.10mm/d, 1.18g/100mL, 2.1%, 55.83mg/100mL; which were superior or equal to other strains. The anti-oxidant activity, proliferation effects of crude polysaccharide on lymphocyte of mice, inhibitory effects of ethanol extracts on L1210 tumor cells, all these were compared. The results are as follows: At the concentration of 250μg/mL, the inhibition rate of tumor cells L1210 by the ethanol extracts of PB-10 is 82%.The scavenging activity of superoxide anion is related to the content of flavones. The value of IC₅₀ on scavenging superoxide anion of ethanol extract of PB-10 is 2.3μg/mL, which is much less than other strains. Therefore, PB-10 strain, an appropriate strain which can produce more flavonoids was screened out from different strains of Phellinus.2 Studies on the optimum of submerged fermentation parametersThe best fermentation condition of producing flavones or intracellular polysaccharide were conformed by various media and conditions, including single factor test and L₉(3⁴)orthogonal test.The optimal culture of producing flavones was: glucose 10g/L, corn flour 40g/L, Soybean cake powder 5g/L, KH₂PO₄ 1.5g/L,MgSO₄ 1g/L; culture temperature 28℃, rotation speed 160r/min, pH normal,inoculation volume8～10%,the amount of liquid 100mL(250mL canonical flask),and fermentation time 144hrs. The yield of total flavones can be reached 26.25mg/100mL.The optimal culture of producing intracellular polysaccharide was: glucose 5g/L, corn flour 40g/L, Soybean cake powder 20g/L, KH₂PO₄ 1.5g/L,MgSO₄ 1g/L; culture temperature 28℃, rotation speed 140r/min, pH normal,inoculation volume 10%,the amount of liquid 100mL(250mL canonical flask),and fermentation time 132 hrs. The yield of intracellular polysaccharide is reach 118.5 mg/100mL.Under the condition: glucose 5～10g/L, corn flour 40g/L, Soybean cake powder 5～20g/L, KH₂PO₄ 1.5～3g/L, MgSO₄ 1～1.5g/L; culture temperature 28℃, rotation speed 140～160r/min, pH normal,inoculation volume 8～10%, the amount of liquid 100mL(250mL canonical flask),and fermentation time 132hrs～144hrs, amount of flavones and polysaccharide could be obtained in the same condition.3 Studies on the activity of mycelia extractsThe ethanol extracts of mycelia in different phases and the fraction of extraction were all active. The sequence of scavenging of active oxygen is n-butanol, ethyl acetate, ethanol, chloroform; the scavenging of active oxygen is dose-dependent on the content of total flavones. The inhibitory effects of different fraction from ethanol extractions on L1210 tumor cells were obvious. All these results indicate it is feasibility and valuable to produce bioactive components of Phellinus baumii by liquid submerged fermentation.