Structural Elucidation And The Biological Activity Of Polysaccharides From Saussurea Laniceps

Snow lotus is a famous precious medicinal material in China,which is widely distributed in Xinjiang,Tibet,Qinghai,Gansu and other high attitude regions.It has attracted much attention because of its good biological activity.The content of polysaccharides in Snow lotus is very rich,but the current papers on snow lotus polysaccharides are very scarce.In order to enrich the research content of Snow lotus,improve its application value and avoid the waste of this medicinal resource,the structure analysis and biological activity of polysaccharides from Saussurea laniceps in Tibet were studied in this paper,and the main results are as follows:(1)The crude polysaccharides of the torus and petal of Saussurea laniceps were obtained by water extraction and alcohol precipitation,and two polysaccharides,SLT-3 and SLP-4,were isolated and purified by DEAE-Sepharose Fast Flow chromatography column and Sephadex G-150 chromatography column.The purity of polysaccharides was 96.89% and 96.56%,respectively.The chemical structure showed that the molecular weights of SLT-3 and SLP-4 were 10113 Da and 19681 Da;SLT-3 was composed of mannose,rhamnose,glucuronic acid,galacturonic acid,glucose,galactose,xylose and arabinose in a molar ratio of 0.25:0.53:0.19:15.35:0.51:1.10:0.63:1.73,whereas SLP-4 consisted of mannose,rhamnose,galacturonic acid,glucose,galactose,xylose and arabinose in a molar ratio of 0.83:2.03:15.00:0.84:8.26:4.04:6.01.The results of FT-IR spectroscopy and Congo red test showed that SLT-3 and SLP-4 were acidic polysaccharides with a triple helix structure;SLP-4 showed a honeycomb-like structure with massive cavities existed inside it,while the surface of SLT-3 was smooth with a small amount of cavities.The results of methylation analysis and NMR analysis suggested that both SLT-3 and SLP-4 were a kind of pectin polysaccharide;SLP-4 was composed of HG,RG I and AG II with a branching structure,while SLT-3 was composed of composed of HG with a straight-chain structure.(2)The antioxidant activities of SLT-3 and SLP-4 were studied by chemical antioxidant test,human hepatocellular carcinoma cell model and human erythrocyte oxidative hemolysis model.The results showed that both SLT-3 and SLP-4 could scavenge DPPH radical,ABTS radical,hydroxyl radical and superoxide anion radical,and had excellent reducing power.ORAC results showed that SLT-3 and SLP-4 had moderate antioxidant activity.In addition,SLT-3 and SLP-4 could protect Hep G2 cells and human erythrocytes from the damage of free radical induced by AAPH.Further studies suggested that SLT-3 and SLP-4 could effectively inhibit the production of ROS,reduce the content of MDA and repair the AAPH-induced erythrocyte oxidative damage by regulating the balance of GSH-GSSG and the activities of CAT,GSH-Px and SOD(maintaining the balance between non-enzymatic and enzymatic antioxidant defenses),and as a result,the normal physiological activity in erythrocytes was maintained.(3)The immune activities of SLT-3 and SLP4 were preliminarily evaluated by using mouse macrophage RAW264.7 cells as the experimental model.The results showed that both SLT-3 and SLP-4 could promote the secretion of cytokines like,NO,TNF-? and IL-6.The authors mainly focused on the investigation of SLP-4 for it showing stronger immunomodulatory effect,and the results were as follows: SLP-4 could significantly enhance the pinocytosis and phagocytosis of macrophages.In addition,SLP-4 could induce M1 polarization of original macrophages and convert M2 macrophages into M1 phenotype,although SLP-4 could not promote the conversion of M1 macrophages to M2 phenotype,it could still inhibit the inflammation induced by LPS.Further studies showed that SLP-4 could specifically react with the surface membrane receptors,like TLR2 and TLR4 of RAW264.7 cells,and then,the immune response was activated through a series of possible signal transduction pathways,such as MAPK and NF-?B.(4)The anti-hepatitis B virus(HBV)activity of SLT-3 and SLP-4 was studied by using Hep G2.2.15 cells as the experimental model.Results showed that both SLT-3 and SLP-4 had good inhibitory effect on the hepatitis B surface antigen(HBs Ag)and hepatitis B e antigen(HBe Ag)in Hep G2.2.15,and the inhibitory effect of SLP-4 was stronger.Different from the common polysaccharides with anti-HBV activity or anti-HBV drugs,the inhibition rate of SLT-3 and SLP-4 on HBe Ag was higher than that on HBs Ag.Through the study of the mechanisms,it was found that the anti-HBV activity of SLT3 and SLP-4 was not achieved by inhibiting hepatitis B virus DNA(HBV-DNA),activating the immune system of the machine or inhibiting the virus invasion.Further studies indicated that there was an interaction between SLT-3 and HBs Ag(or HBe Ag),SLP-4 and HBs Ag(or HBe Ag),which might block the ELISA detection of HBs Ag and HBe Ag,resulting in a decrease in absorbance and thus,making SLT-3 and SLP-4 have the anti-HBV activity.(5)Oxidized low-density lipoprotein(ox-LDL)was used to induce the foaming of RAW264.7 cells,and the lipid-lowering activity of SLT-3 and SLP-4 was studied.It was found that the intracellular cholesterol content of RAW264.7 cells decreased most significantly after the co-incubation with SLT-3 or SLP-4 and ox-LDL,therefore,the author chose this modeling method for the following experiments.The results o showed that both SLT-3 and SLP-4 could reduce the lipid accumulation,the content of ROS and the accumulation of MDA and the secretion of inflammatory cytokines(TNF-?,IL-1 ? and IL-6)in foam cells.Additionally,there was an interaction between SLT-3 and ox-LDL,SLP-4 and ox-LDL,which might prevent some ox-LDL from entering into the RAW264.7 cells.Therefore,the lipid-lowering effect of SLT-3 and SLP-4 might be achieved through the joint action.