The Effect Of Gastrodia Elata Extract On The Physicochemical Properties, Antioxidant And Immune Activities Of Grifola Frondosa Extracellular Polysaccharide

In this paper,the effects of Rhizoma gastrodiae extract(R.gastrodiae extract)on the exopolysaccharide fractions,physicochemical properties,structural characteristics and biological activities of Grifola frondosa(G.frondosa)were carried out.The researches mainly included that adding the 7%(v/v)R.gastrodiae extract in the submerged culture of G.frondosa to find out the changes of the main components and its contents of R.gastrodiae extract,exploring the exopolysaccharide arrangements and contents of G.frondosa.Taking the main fractions of polysaccharide as research objects(EPS-2 and REPS-2),we have studied to determine the differences of polysaccharide contents,protein contents,uronic acid contents,molecular weights,monosaccharide compositions and glycosidic bonds of EPS-2 and REPS-2,and to explore the effects of R.gastrodiae extract on the primary structure of exopolysaccharide of G.frondosa.In order to explore the antioxidant activities in vitro and the immunological activities in mice as research goals,we have studied to determine the changes of superoxide anion radical scavenging rates,DPPH radical scavenging rates,ABTS radical scavenging rates,mouse immune organ indexs,serum cytokines and immune globulins contents of EPS-2 and REPS-2,exploring the effects of R.gastrodiae extract on the bioactivities of exopolysaccharides from G.frondosa and summaring of the relationships between structural characteristics and differences in biological activities.The research results are as follows:The qualitative and quantitative detection methods of gastrodin,p-hydroxybenzyl alcohol(HA),p-hydroxybenzaldehyde(HBA)and parishin were studied and established.The changes of R.gastrodiae extract main components before and after autoclaving,and the utilization of main components of R.gastrodiae extract by G.frondosa were analyzed.The main components of R.gastrodiae extract after autoclaving was changed,and the content of gastrodin after sterilization increased by66.06%,HA increased by 36.96%,HBA decreased by 37.64% and the parishin decreased by 90.47%.During the fermentation of G.frondosa,the main components of R.gastrodiae extract were changed,which the gastrodin content decreased by24.68%,the HA content increased by 6.50%,the HBA content decreased by 42.59%,and the parishin content increased by 9.85%;The R.gastrodiae extract was added into the fermentation of G.frondosa,the physicochemical properties and structural characteristics of the obtained EPS-2 and REPS-2 were structurally tested.The results showed that the polysaccharide contents of EPS-2 and REPS-2 were 82.68% and 84.12%,the protein contents were 0.31% and0.23%,and neither contained uronic acid.In addition,the molecular weights of EPS-2and REPS-2 were 1.295×104 Da and 2.904×104 Da,respectively.The monosaccharide composition of EPS-2 was mainly composed of glucose,mannose and galactose,which the molar ratio was about 4.11:4.68:0.04.Moreover,the monosaccharide composition of REPS-2 was mainly composed of glucose,mannose,galactose and fucose,and its molar ratio was about 3.02:5.72:0.09:0.16.EPS-2 and REPS-2 had typical polysaccharide absorption peaks.GC-MS methylation indicated that both EPS-2 and REPS-2 were composed of six glycosidic bonds: t-Glcp,t-Manp,1,2-Glcp,1,3-Manp,1,6-Manp and 1,3,6-Manp.EPS-2 was mainly composed of t-Glcp,1,2-Glcp.1,3-Manp,1,6-Manp and 1,3,6-Manp,and the molar ratio was approximately about 13.9:23.5:8.6:26.8:10.2.Meanwhile,REPS-2 was mainly composed of t-Manp,1,2-Glcp,1,3-Manp,1,6-Manp and 1,3,6-Manp,which the molar ratio was approximately about 10.5:19.7:9.2:32.1:14.5;The experiment results of antioxidant activities in vitro showed that EPS-2 and REPS-2 had good antioxidant capacities(superoxide anion radical scavenging rates,DPPH radical scavenging rates and ABTS radical scavenging rates),and the radicals scavenging rates increased with increasing sugar concentration,which were apparented concentration dependence,but were lower than the positive control Vc group.Among them,REPS-2 promoted the DPPH radical scavenging most obviously,followed by the ABTS radical and the superoxide anion radical.The results were significantly higher than EPS-2(P<0.05),and the semi-inhibitory concentrations(ED50)were all less than 4 mg/m L;The experimental results of immunological activities in mice showed that the EPS-2 and REPS-2 with intragastric doses of a middle-dose(100 mg/kg)and ahigh-dose(200 mg/kg)were effectively increased the thymus index,spleen index,serum cytokines and immune globulins in immunodeficient mice induced by cyclophosphamide(CTX)(P<0.05).Among them,at the same dose,REPS-2 was significantly better than EPS-2 in promoting the secretion of serum cytokines(IFN-?and IL-2)and immune globulin(Ig G)(P<0.05).These data indicate that REPS-2,dominated by mannan/glucan,was more effective in regulating immune activities of mice than EPS-2.