Studies On Adjuvant Acivity Of Solanum Lyratum Sponin Fractions

Solanum Lyratum Thunb, a kind of solanun plant,which has been used as a valued chinese traditional medicine over twenty centures. According to traditional chinese medicine, The pharmacolegical nature of this herb is little cold and bitter. It has the function of Qingrejiedu and wind-expelling phlegm which usually used to treat many kinds of tumors and cancers.In the present research, Total sponins were obtained from Solanum Lyratum Thunb by traditional exrtacting method,From which the different saponin fractions were isolated initially . With the aim of selecting pure sponin mononer with high adjuvant activity. the activity of each fraction was studied by immune responses of ICR mice against ovalbumin (OVA), recombinant hepatitis B surface antigen (rHBsAg) . The thesis mainly contains three parts:1 Isolation and identification of total sponin and its fractions from Solanum Lyratum Thunb.The dry Solanum Lyratum Thunb were extracted with 80% EtOH, and then concentrated in a small volume. EtOH extract was suspended in water and defatted with ether. followed by partitioning with n-BuOH. The combined BuOH layers were concentrated to obtain a total sponin. The total sponin was subjected to D101, washed with H2O, and eluted with EtOH of four kinds of concentrations to get four sponin fractions A,B,C and D. Identfied by Molish reaction as well as other methods,There are sponins in each fractions.2 Adjuvant effect of Solanum Lyratum Thunb saponin fractions on the immune responses to OVA-immunized mice .Detection of specific antibody (IgG, IgG1and IgG2a)levels in ICR mice against OVA by the method of ELISA,The result showed each Solanum Lyratum Thunb sponin fraction could significantlly enhance the levels of IgG, IgG1and IgG2a(P < 0.05orP < 0.01).Among them, C group has the most significant adjuvant activity(P < 0.01).The result also proved Solanum Lyratum Thunb saponin could enhance cellular and humoral immune responses of ICR mice against OVA. But the result showed that the adjuvant functions of Solanum Lyratum Thunb saponin fractions seemed having no correlation with their polarity.According to the studied result,the polarity of sponins have a certain relation to their sugar chains,But the adjuvant effect of saponins is mainly connected with the kind of sugar residues shared by them.3 Adjuvant activity of Solanum Lyratum Thunb saponin fractions on recombinant hepatitis B surface antigen (rHBsAg) vaccine.By the same mathod above to detect specific antibody (IgG, IgG1and IgG2a)levels in ICR mice against recombinant hepatitis B surface antigen (rHBsAg) vaccine,The result showed,Compared to rHBsAg control group ,Solanum Lyratum Thunb saponin fractions could significantly not only enhance rHBsAg-specific IgG and IgG1 antibody levels, but also increase IgG2a antibody levels in rHBsAg-immunized mice (P < 0.05orP < 0.01). Alum also increased the rHBsAg-specific IgG and IgG1 antibody responses.But there were no significant differences in IgG2a antibody levels between the rHBsAg group and Alum/rHBsAg group. In IgG2a antibody levels, Solanum Lyratum Thunb saponin fractions groups were significantly higher than Alum/rHBsAg(P < 0.05orP < 0.01), Especially C group (P < 0.01) . All suggest that Alum induces almost exclusively a Th2 response, while Solanum Lyratum Thunb saponins elicits both Th1 and Th2 response.The detecting result of MTT showed Solanum Lyratum Thunb saponins could significantly enhance Con A, LPS and rHBsAg - induced splenocyte proliferation in the mice against rHBsAg vaccine( P < 0.05or P < 0.01),C fraction group is the highest,which was significantly higher than Alum group( P < 0.01).The effects of Solanum Lyratum Thunb saponins on natural killer (NK) cell and rHBsAg-specific CTL activity from splenocytes in rHBsAg-immunized mice were measured.The results show,Compared to Alum group., Lyratum Thunb saponins groups could significantly improve NK and CTL activity(P<0.05orP<0.01).The adjuvant activity of C group is the highest( P < 0.01).suggesting that the usage of Solanum Lyratum Thunb saponins in rHBsAg vaccine can help to improve specific and non-specific ability to kill target cell.