Study On The Structural Characteristics And Pharmacological Activity Of Polysaccharides From Dictamnus Dasycarpus Turcz

Objective To study the extraction,purification,structure analysis,activity screening and its anti-psoriatic activity of Dictamnus dasycarpus polysaccharide?DDP?,and provide experimental basis for further research on the efficacy and molecular mechanism of anti-psoriatic.DDP.It lays the foundation for the research and development of DDP for the treatment of psoriasis and other immune diseases.Methods?1?The polysaccharide was extracted by water extraction alcohol precipitation,and the content of sugar was determined by phenol-sulfuric acid method.The content of aluronic acid was determined by the sulfuric acid carbazole method,while the content of protein was determined by Kaumas brilliant blue method,then Trichloroacetic acid method was used to remove protein and the activated carbon method was used to remove pigment.The purified polysaccharide was divided into 7 segments with different molecular weights by membrane separation technique.?2?The MTT colorimetric assay was used to determine the immune activity of DDP in vitro,and the antioxidant activity of DDP was determined by using the ability of removing DPPH free radical?DDPH?,superoxide anion radical?·O₂^-?and hydroxyl radical?·OH?in vitro to screen active polysaccharide.?3?We establishe the model of psoriasis induced by Imiquimod Cream in mice and lavage the DDP-D,DDP-E and DDP-F,avd it investigated the anti-psoriatic activity of the three polysaccharides by the expression of IL-17and IL-23 in the serum of mice,skin PASI score,and pathological examination of the back skin.?4?DEAE Cellulose-52 Column chromatography was used to isolate and purify the anti-psoriatic activity of DDP-D and DDP-E.The purity and molecular weights of the polysaccharides were determined by gel permeation chromatography.The composition of the monosaccharide composition of the uniform polysaccharide was analyzed by HPLC method.The structure characteristics of homogeneous of Dictamnus dasycarpus Turcz were analyzed by using periodate oxidation,Smith degradation,methylation analysis and infrared spectroscopy.Results?1?The extraction rate of DDP was 20.25%,the content of polysaccharide was14.1%,the content of aluronic acid was 1.04%,protein content was 1.67%,the removal rate of protein by three chloroacetic acid was 90.5%,the retention rate of polysaccharide was 72.7%,and the 7 segments of different molecular weights were respectively DDP-A?<6 kDa?,DDP-B?10?6 kDa?,DDP-C?20?10 kDa?,DDP-D?50?20 kDa?,DDP-E?100?50 kDa?,DDP-F?150?100 kDa?,DDP-G?>150 kDa?.?2?After screening in vitro,it was found that DDP-A^G could inhibit the proliferation of spleen cells.The best immunosuppressive activity was DDP-C,DDP-D,DDP-E and DDP-F.The antioxidant activity was screened in vitro.It was found that DDP-A^G had the ability to scavenge free radicals,and the most ideal antioxidant activity was DDP-D,DDP-E,DDP-F and DDP-G;the two kinds of compounds were integrated in vitro.Activity screening showed that DDP-D,DDP-E and DDP-F were antioxidative and immunosuppressive.?3?According to the scores of mouse PASI lesions,DDP-E and DDP-D mice skin lesions had significantly improved after 10 days;the skin lesions of DDP-F group were not significantly improved.Pathological observations revealed that the epidermis thickened was significantly of mice in the model group.Compared with the model group,DDP-D group and DDP-E high-dose group had a smooth and thin skin lesion;the DDP-F group had hyperkeratosis of the stratum corneum,and the epidermis thickened significantly.The DDP-D group and DDP-E high dose group had significantly lower serum levels of IL-17 and IL-23 than the model group,comprehensive PASI scores,skin morphology observations,and cytokine results,which indicated that DDP-D,DDP-E had better treatment of psoriasis mice effect.?4?DDP-D1 with a molecular weight of 3.47×10⁴ Da,DDP-D2 with 4.23×10⁴ Da,DDP-E1 with5.01×10⁴ Da,and DDP-E2 with 9.88×10⁴ Da were obtained after separation and purification of DDP-D and DDP-E by DEAE Cellulose-52,and HPLC and GC-MS analysis shows that DDP-D1 is composed of Man,GlcA,GalA,Glc,Gal in the molar ratio of 13.14:1.91:1.00:1.44:25.24,the main chain consists of Man and Glc with the main bond types are 1?3,1?3,6,1?2;DDP-D2 is composed of Man,Rha,Glc,Gal,and Xyl in the molar ratio of 2.62:1.41:1.00:6.2:3.59,the main chain consists of Xyl,Man and Rha with main bond types are 1?3,1?2,6,1?3,4;DDP-E1 is composed of Man,Rha,GlcA,Glc,Gal in the molar ratio of 22.44:17.28:1.73:1.00:11.40,the main chain consists of Man and Rha with the main bond types are 1?3,1?3,4,1?3,6;DDP-E2 is consists of Man,Rha,GalA,Glc,and Gal in the molar ratio of14.19:1.00:1.29:1.68:1.68,the main chain is composed of Man with the main bond types are 1?2,3,1?3,6.Conclusion The polysaccharide of Dictamnus dasycarpus Turcz is an important material basis with anti-oxidation and immunosuppressive effects,DDP-D1 with a molecular weight of3.47×10⁴ Da,DDP-D2 with 4.23×10⁴ Da,DDP-E1 with 5.01×10⁴ Da,and DDP-E2 with9.88×10⁴ Da are ideal anti-psoriasis substances,and its structural characteristics of homogeneous polysaccharides are clear.It is worth to further research and develop,and it has great medicinal value.