| This article is mainly study the effect of Rhizoma gastrodiae(R.gastrodiae)ethanol extract on mycelia β-glucan and composition of fermentation broth by submerged culture of Grifola frondose(G.frondsa).The method of extraction of mycelium β-glucan from G.frondsa was optimized by orthogonal design.R.gastrodiae ethanol extracts of different concentration were added into the fermentation system of G.frondsa to study the effect of additives on mycelia biomass,mycelia polysaccharides and β-glucan from G.frondsa,the content of gastrodin(GA),hydroxybenzaldehyde(HBA)and hydroxybenzyl alcohol(HA)in R.gastrodiae ethanol extracts was determined by High Performance Liquid Chromatography(HPLC),and the main ingredients of R.gastrodiae which one is the most responsible to accelerate biomass,mycelia polysaccharides and β-glucan productions and optimum supplemental levels were researched.Hydroxybenzaldehyde were added into the fermentation system of G.frondsa to study the effect of additives on composition of fermentation broth by submerged culture of G.frondsa.The effects of G.frondsa fermentation broth on immune function in immunosuppressed mice was studide.The main results as follows:The β-glucan content of two strains of G.frondsa(strain 1 and 2)was tested under the same culture and extraction conditions,and the results showed that the β-glucan content of strain 2 was higher than that of strain 1,therefore,we choose the strain 2 for further study of strain.The strain 2 as the object of study,the extraction method of β-glucan from G.frondsa was compared through four methods: hot water extraction,hot water-enzyme extraction,ultrasonic,and ultrasonic combined with enzyme hydrolysis method,and the results showed that the extraction rate of β-glucan by ultrasonic combined with enzyme hydrolysis method is higher than that of the other three methods.On the basis of single factor test,with the extraction rate ofβ-glucan as the evaluation index,orthogonal test was used to optimize the extraction conditions of ultrasonic combined with enzyme hydrolysis method,and the results showed that the optimum extraction conditions were ultrasonic power 300 W,ultrasonic time 15 min,compound enzyme additive amount 1.5 %,the temperature of the enzyme hydrolysis 40 ℃,the extraction yield of β-glucan from G.frondsa mycelium reached 2.80 mg/g under the conditions.R.gastrodiae ethanol extract of different concentrations were supplemented into the fermentation system of G.frondsa to study the effects of R.gastrodiae ethanol extract on mycelia biomass,mycelia polysaccharides and β-glucan of G.frondsa by submerged fermentation.The contents of gastrodin(GA),p-hydroxybenzyl alcohol(HA)and p-hydroxylbenzaldehyde(HBA)in R.gastrodiae ethanol extract were determined by the method of High Performance Liquid Chromatography(HPLC).The results showed that 7 g/L R.gastrodiae ethanol extract gave the best results,the contents of characteristic constituents of R.gastrodiae ethanol extract were gastrodin(GA)5.8375 mg/g,p-hydroxybenzyl alcohol(HA)1.1072 mg/g,p-hydroxylbenzaldehyde(HBA)0.6601 mg/g.Three kinds of characteristic components of R.gastrodiae ethanol extract of different concentrations were supplemented into the fermentation system of G.frondsa to study the key characteristic constituents which one is the most responsible to accelerate biomass,mycelia polysaccharides and β-glucan productions and optimum supplemental levels.The results showed that HBA is the most responsible ingredients to promote the biomass,mycelia polysaccharides and β-glucan productions in submerged culture of G.frondosa which optimum supplemental levels were 100 mg/L,250 mg/L and 150 mg/L,respectively.In order to study the effects of HBA on composition of fermentation broth by submerged culture of G.frondosa,change rule of the amount of mycelium biomass,reducing sugar,p H value,extracellular polysaccharide,extracellular protein,and the content of polyphenol flavonoids in the fermentation broth extract were investigated.The results showed that HBA can promote the biosynthesis of polysaccharides,proteins and polyphenol flavonoids in G.frondosa fermentation broth to some extent.In order to study the effect of G.frondosa fermentation broth on immunity in mice,we used G.frondosa fermentation broth as test substance and immunocompromised mice as subjects.The results showed that high dose blankfermentation liquid and three doses of R.gastrodiae fermentation broth can significantly improve the spleen gland index of immunocompromised mice.R.gastrodiae fermentation broth can significantly improve the ear swelling of immunosuppressed mice and stimulate delayed allergy.High dose of blank fermentation broth and high dose of R.gastrodiae fermentation broth can significantly improve the content of serum Ig G in mice.High and medium dose of blank fermentation broth and high and medium dose of R.gastrodiae fermentation broth can significantly improve the content of serum Ig M in mice. |
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