Study On The Key Technology Optimization And Biological Activity Of AFG Purification

In this paper,a new separation and purification process of AFG was established and optimized by single factor test and orthogonal test.The experimental results show that the best process condition is A₃B₃C₂D₃,that is,the ratio of ethanol and AFG synthetic liquid is 4:1,the alcohol is precipitated three times,and the AFG crude product obtained after each stirring for 45 minutes passes through the polyacrylamide column four times.In this paper,the protective effect of AFG on CCl₄induced acute liver injury in mice was studied,and the mechanism of its treatment was discussed.They were divided into the control group,the model group and the administration group(50,100,200mg·kg^-1).The model group and the administration group were injected with peanut oil solution of CCl₄intraperitoneally for 7 days after continuous gavage,and the blank group was injected with peanut oil intraperitoneally.The indexes of liver and spleen were measured 16 hours later,and the levels of ALT and AST in serum were measured by kit method.The levels of SOD and MDA in liver were measured by colorimetry.The liver tissue was taken for histopathological observation.The expression levels of TNF-?,IL-1?and IL-6 were detected by RT-PCR.Western blot was used to detect the protein expression of TNF-?,IL-1?and IL-6 in liver tissue.The results showed that the administration group could reduce the index of liver and spleen,the activity of ALT and AST in serum,the content of MDA in liver and the level of SOD in liver tissue.The pathological phenomenon of histological section was improved.The expression of TNF-?,IL-1?and IL-6 m RNA and protein in liver tissue were decreased.It is suggested that AFG has protective effect on the liver of CCl₄induced acute liver injury mice,and its mechanism may be related to the gene and protein expression of TNF-?,IL-1?and IL-6.In this paper,the inhibitory effect of AFG on proliferation and apoptosis of mouse melanoma B16 cells was studied.CCK-8 method was used to detect the toxicity of AFG to B16 cells,and the best concentration was selected.The LDH content of B16 cells was detected by colorimetry.Hoechst 33258 fluorescent staining was used to observe the effect of AFG on apoptosis of B16 cells.RT-PCR was used to detect the expression of Bcl-2,Bax and Caspase-3 in B16 cells.Western blot was used to detect the expression of Bcl-2,Bax and Caspase-3 in B16 cells.The results showed that the proliferation of B16 cells was inhibited by AFG at 50,75 and 100?mol/L.Under the microscope,the number of B16 cells apoptosis increased significantly in each group.The m RNA and protein levels of Bcl-2 and Bax and Caspase-3 in B16cells were significantly decreased,while the m RNA and protein levels of Bax and caspase-3 were significantly increased.AFG can inhibit the proliferation and promote the apoptosis of B16 cells,which may be related to the gene and protein expression of Bcl-2,Bax and caspase-3.