In Vitro Antioxidant Activity Screening Of Genistein Structure Modification

Objective: From the nine genistein structure transformation to extract derivatives which are antioxidant activity better than genistein, for the study of the structure-activity relationship and Isoflavones drug research to provide theoretical guidance.Methods: 1. The Fenton reaction-Phen-Fe2+ photometric method: ·OH with H2O2 as the substrate, Determination of genistein structure modification to remove ·OH ability;2. Using the FRAP method: under acid condition, Fe3+-TPTZ reducing substances in the samples can be reduced into Fe2+-TPTZ form, and present a clear blue, in 593 nm place has the largest light absorption, to reflect the sample total reducing power;3. Tiefa thiocyanate(FTC) : linoleic acid oxidation in the process of automatically generating peroxide, use quantitative method to investigate antioxidant ability to resist lipid peroxides;4. In vitro with H2O2(100 ~ 900 n M) dealing with human umbilical vein endothelial cells(HUVEC), and uses the CCK-8 method to detect the cell vitality, build the HUVEC oxidative stress injury induced by H2O2(IC50)model;5. CCK-8 method is used to detect genistein and preliminary screening out 4, 9 structure modification on the oxidative stress induced HUVEC toxicity of protection;6. Observe the genistein and preliminary screening out of 4, 9 structure modification.It influence on endothelial cell number and form;7. Through preliminary screening out in 4, 9 structure modification,in drug concentration(20 n M, 100 n M, 100 n M) under the pretreatment of HUVEC, MDA, LDH, GSH and SOD and CAT enzyme activity as an index, respectively at different concentrations of its protective effect on HUVEC oxidative stress injury.Results: 1. Genistein structure modification can remove system of ·OH, 2-9 samples in 0 ~ 80 u M with the increase of the concentration of its clearance ability enhancement, concentrations in 80 u M, ·OH clearance for 9 > VC > genistein > 8> 7 > 6> 4, other did not see obvious difference;2. Genistein structure modification are reducing power, at 0 ~ 400 u M its reducing power as the rise of the concentration increased, 9 FRAP values greater than genistein significantly(p < 0.05), other did not see obvious difference compared with genistein;3. Genistein structure modification has the ability to resist lipid peroxides, when measuring the sample concentration is 400 u M, 4 days system response, the determination results for 4, 9 resisting lipid peroxide significantly stronger than genistein, 4 which is significantly higher than genistein(p < 0.05);4. In vitro built 750 u M HUVEC oxidative stress injury induced by H2O2(IC50) model;5. In concentration(20n M ~ 500 n M) within the scope of visible genistein, 4 and 9 protection is concentration dependent, therefore its decline on H2O2 induced endothelial cell vitality has significant inhibitory effect, and has a 9 maternal stronger inhibition;6. The sample cell shape of each dose group were different degrees of improvement, most cells short a polygon or fusiform, intercellular space smaller, showed that genistein, 4, 9 of HUVEC form has good protection;7. Genistein, 4, 9(20 n M ~ 500 n M) concentration dependence of raising the level of SOD, CAT, GSH, and inhibition of oxidative stress induced the decrease of SOD, CAT, GSH levels among them 9 increase SOD, CAT, GSH levels significantly better than that of genistein(p < 0.01); Genistein, 4, 9(20n M ~ 500 n M) for lower concentration dependent on the level of LDH and MDA, and inhibit oxidative stress induced increased LDH leakage and MDA levels rising, 4 which cut level of LDH and MDA was significantly better than that of genistein(p < 0.01).Conclusion: Through the in vitro antioxidant experiment Preliminary screen the 4, 9 genistein derivatives.