Research On The Interaction And Mechanism Between Flavonoids And α-amylase

Flavonoids are a kind of polyphenolic compounds, which widely existing in nature. Asflavonoids possess lots of biological activity, including antioxidant, inhibiting lipidperoxidation, prevention of cardiovascular disease and cancer, flavonoids are extensivelyresearched, developed and utilized in food and pharmaceutical industry. There are some activephenolic hydroxyls in flavonoid molecule, it may interact with digestive enzyme, such assalivary amylase, pancreatic amylase, and influence catalytic activity of enzyme after took inbody. Therefore, six kinds of representative flavonoid (luteolin, diosmetin, narigenin,dihydromyricetin, kaempferol, quericetin) in structure are selected in this article. In vitrosimulating biological environment in vivo, we have researched the changes of theircharacteristic respectively after six flavonoid interacted with pig pancreatic α-amylase, anddiscussed the main binding force and structure-activity relationship between them.In the process of researching six kinds of flavonoids on pig pancreatic α-amylase catalyticactivity, it was concluded that six kinds of flavonoids (luteolin, diosmetin, narigenin,dihydromyricetin, kaempferol, quericetin) have an effect on pig pancreatic α-amylasecatalytic activity. When pig pancreatic α-amylase was0.616U/mL, the flavonoidconcentration was1mg/mL, the inhibitory effect of luteolin on catalytic activity of pigpancreatic α-amylase was maximu and the inhibition rate reached81.24%. The inhibitoryeffect of quericetin on catalytic activity of pig pancreatic α-amylase was minimum and theinhibition rate was only9.82%. The inhibition rate of six kinds of flavonoids on the catalyticactivity ofpig pancreatic α-amylase was luteolin> diosmetin> narigenin> dihydromyricetin>kaempferol> quericetin. Besides, through the preliminary study of the kinetics of pigpancreatic α-amylase, we knew that the inhibitory effect of these kinds of flavonoids on pigpancreatic α-amylase belong to noncompetitive.In the process of researching six kinds of flavonoids on pig pancreatic α-amylasefluorescence spectra, we found that there are different degrees quenching effect of six kinds offlavonoids on pig pancreatic α-amylase fluorescence characteristic. With the increasing offlavonoids concentration, the endogenous fluorescence intensity of pig pancreatic α-amylasedecreased regularly. Fluorescence quenching constants of pig pancreatic α-amylase interacting with six kinds of flavonoids were greater than the maximum dynamic quenchingconstant(2×1010L/mol·s) of all kinds of quencher on biological macromolecules, and thefluorescence quenching type was confirmed as static quenching. What’s more, we could alsoobtain only one binding site and greater binding constant for the interaction between six kindsof flavonoid and pig pancreatic α-amylase, with the analysis and calculation of bindingconstant and the number of binding site. According to the magnitude of the binding constant,reflecting the bonding strength of the interaction between six kinds of flavonoid and pigpancreatic α-amylase, we can make a conclusion that the strength order was luteolin>diosmetin> narigenin> dihydromyricetin> kaempferol> quericetin. Finally, the interactiontypes between six kinds of flavonoid and pig pancreatic α-amylase were dominated byhydrophobic force through thermodynamic study.In the process of researching the effects of pig pancreatic α-amylase on antioxidantcapacity of six kinds of flavonoid, the inhibition effects of pig pancreatic α-amylase onoxygen radical absorbance capacity, scavenging DDPH free radical and reducing power ofsix kind of flavonoid were all not obvious, the maximum of inhibition rate were19.88%、18.91%、17.04%respectively. However, the inhibition rate of pig pancreatic α-amylase onresisting oxidation of linoleic acid was44.21%to the maximum. For the above four kinds ofantioxidant capacity evaluation method, the strength orders of inhibition effect of pigpancreatic α-amylase on antioxidant capacity of six kinds of flavonoid were all luteolin>diosmetin> narigenin> dihydromyricetin> kaempferol> quericetin.Through analyzing the structure-activity relationship after six different structure offlavonoids interacted with pig pancreatic α-amylase, we could make a conclusion that the totalnumber of hydroxyl groups and the hydroxyl group on ring B had no significatantly effects onthe binding force between flavonoids and pig pancreatic α-amylase; the C-3hydroxyl groupon ring C made against to the binding interaction between flavonoids and pig pancreaticα-amylase; The methylation of the hydroxyl group in ring B of flavonoid could also weakenthe binding force between flavonoids and pig pancreatic α-amylase.