| A polysaccharides component(LP-2)was isolated and purified from longan pulp.The structure of LP-2 was analyzed using gel chromatography-laser light scattering(GPC-LLS),ion chromatography,gas phase mass spectrometry(GC-MS),infrared spectroscopy(IR)and nuclear magnetic resonance(NMR).The effects of continuous digestion of saliva,gastric juice,and intestinal juice on the structure of LP-2 and activation of DC cells were analyzed by simulated digestion model.And the main results are as follows:1.LP-2 was composed of galactose and glucose in a relative molar ratio of 1.45%:95.50%,with a weight average molecular weight MW of 4.67×10~7(±3.43%),a dispersion coefficient of 1.31(Mw/Mn),and a radius of rotation RZ of 116 nm.The results of methylation showed that the LP-2 glycosidic linkages were mainly 70.32%of 1,6-Glcp,10.76%of t-Glcp and 10.13%of 3,6-Glcp.NMR results indicate that LP-2 is a polysaccharide with 1,6-?-D-Glcp as the main chain,1,3,6-?-D-Glcp as the branch,and T-?-D-Glcp as the terminal polysaccharide..2.The digestion model in vitro was used to studied its effect on the structure of LP-2.The results showed that LP-2 had no significant changes in molecular weight and reducing sugar content during saliva digestion,continuous saliva-gastric digestion and continuous saliva-gastric-intestinal digestion,and no free monosaccharides in the final digestive fluid.The simulated digested polysaccharide(DLP-2)was composed of galactose and glucose in a relative molar ratio of 3.15%:91.15%,and its glycosidic linkage were 63.93%of 6-Glcp,9.14%of 3,6-Glcp and 8.53%of t-Glcp,and there is no significant difference between the infrared spectra of LP-2 and DLP-2.The results showed that saliva,gastric juice,and intestinal fluid did not cause significant changes in the structure of LP-2.3.Flow cytometry and other methods were used to evaluate the effect of in vitro simulated digestion on the activation of dendritic cells(DC)by LP-2.The results showed that there was no significant difference between 50-200?g/mL of LP-2 and PMB-treated LP2 in phagocytic rate of DC2.4 cells and IL-12 secretion by DC2.4cells(p<0.05),indicated that the activation of LP2 on DC cells was not caused by endotoxin.When the concentrations of LP-2 and DLP-2 were in the range of 50-200?g/mL,the relative proliferation rate of DC2.4 cells was above 95%,which was not significantly different from the blank control group(p<0.05).The phagocytic rates of 200?g/mL LP-2 and DLP-2 on DC2.4 cells were 6.82±0.69%and 6.33±0.61%respectively,which were 73.89%and 68.58%of blank control group;the secretion of IL-10 and IL-12 that are stimulated by LP-2 were 200.26±21.72 pg/mL and 38.92±2.59 pg/mL respectively,which were 121.29%and 127.11%of the blank control group;the secretion of IL-10and IL-12 that are stimulated by DLP-2 were 237.31±41.95 pg/mL and 38.35±4.54pg/mL respectively,which were 143.73%and 125.24%of the blank control group.The same concentration of DLP-2 and LP-2 had no significant difference in proliferation of DC2.4,phagocytosis of fluorescent microspheres and the secretion of IL-10 and IL-12(p<0.05),indicating that the simulated digestion process in vitro has no significant effect on the immunomodulatory activity of longan polysaccharides. |
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