Preparation,Characterization,Antioxidant And Antiinflammatory Effect Of The Chelate Of Flammulina Velutipes Po Lysaccharide With Zn

Flammulina velutipes,one of the most popular edible mushrooms,is currently artificially cultivated on large-scale in China and Japan,where it has gained popularity due to its high nutritional value and delicious taste.Most studies have paid much attention on the active constituents and pharmacological properties of Flammulinavelutipes.Polysaccharides,one of the most important components of mushrooms,have exhibited extensive and powerful benefits applicable in developing novel drugs and functional foods,which are correlated to their immune-modulating,anti-oxidation,antitumor and nontoxic effects.Several polysaccharides have been isolated from F.velutipes fruiting bodies or mycelia,and their antioxidation,anti-tumor and immuno-reglatory effects studied.Nowadays,there are many studies related to the antioxidant activities and anti-inflammatory of polysaccharides,however,few investigations had been carried out on the chelation of Zn with polysaccharides from edible mushroom.Therefore,Zn-Flammulina velutipes polysaccharide(Zn-FVP)was prepared and the anti-inflammatory activity of the polysaccharides before and after chelation were comparatively analyzed in this study.Moreover,Structural characteristics of Flammulina velutipes purified polysaccharide(FVPp)and Zn-FVP were investigated.It provides a theoretical basis for the development of new functional products of Zn-FVP.The main research contents are as follows:1.Crude polysaccharide(FVPc)was extracted from Flammulina velutipes.Then,optimal conditions for preparing Zn-Flammulina velutipes polysaccharide(Zn-FVP)chelate were studied by using L9(34)orthogonal test.The results showed that the optimized conditions for the chelating reaction were as follows:initial concentration of Zn(?),6 g/L;ratio of purified polysaccharide(FVPp)to Zn(?),5:1;chelating time,8 h;and pH,5.Chelating rate achieved 98.11%under the optimal conditions.2.The structure of FVPp and Zn-FVP were characterized by Fourier transform infrared spectroscopy(FTIR)and powder X-ray diffraction anaysis(XRD).The FTIR spectroscopy results may be used to elucidate the configuration of the polysaccharide and it was a furanose for there being two intense absorption bands between 1100 cm-1 and 1010 cm-1.Characteristic bands for OH stretching at 3000-3500 cm-1 and bands between 1000-1100 cm-1 corresponding to C-H variable angle vibration,showed some obvious differences for chelation of polysaccharides and zinc.At the same time,the absorption peak of N-H at ca.1654 cm-1 shifted to the 1616 cm-1 due to the redshift phenomenon,and the shift of the absorption bands groups may contributed to the linkage of N-H and zinc ions.The absorption band at 620 cm-1 was related to the sulfate group which indicated that physisorption played an important role in the reaction apart from chelation.The qualitative spectrum of Zn-FVP indicated the presence of ZnSO4.This result was also supported by FTIR Analysis.New peaks was attributed to the changes of structure in Zn-FVP compared with FVPp.This result indicated that the abundant SO42-and Zn(?)groups were widely present in the chelated samples.Compared to chelated samples,Zn signal was not observed in FVPp.XRD showed the existence of new functional groups as evidenced by the appearance of new peaks.3.The antioxidant activities of the polysaccharides before and after chelation were evaluated.Zn-FVP showed stronger scavenging activities on DPPH,O2-· and ·OH than FVP.FVP and Zn-FVP exhibited almost the same antioxidant activities in H2O2.To summarise,the antioxidant activity evaluation were all improved after the chelation with Zn2+.The study demonstrates that the increase of the content of zinc can enhance the antioxidant activity of Flammulina velutipes purified polysaccharide.4.The anti-inflammatory effect of FVPp and Zn-FVP on Lipopolysaccharide(LPS)-stimulated RAW 264.7 cells was evaluated.In the range of 5-50?g/mL,Zn-FVP didn't exhibit obvious inhibition of the survival rate of cells.Zn-FVP in noncytotoxic levels showed strong suppressive effect on the inflammatory cytokines(IL-6,TNF-a,INF-y and NO)and might be exploited as an effective therapeutic drug.5.The antitumor effect of Flammulina velutipes purified polysaccharide before and after chelation were evaluated.To investigate the antitumor effect of the polysaccharide/Zn2+Chelate on L929 tumor cell,the methlthiazolyl tetrazolium(MTT)experiment was carried out and the shape of L929 cells was observed with different concentration of polysaccharide solution.The results showed that at the concentration of 5-50 ?g/mL,crude polysaccharide and purified polysaccharide had little effect on the growth of L929 cells while the polysaccharide/Zn2+ Chelate played an important role in inhibitory effect.Furthermore,the cultural supernatant of macrophages cocultured with Flammulina velutipes polysaccharides could kill L929 cells.