Study On The Interaction Mechanism Between Three Citrus Flavonoids And Digestive Enzymes

In recent years,the incidence of type II diabetes and obesity are increasing rapidly,which seriously threatens human health.With?-glucosidase,?-amylase and pancreatic lipase as the main targets,exploring natural plant components with low toxicity and low side effects has become one of the hot topics in the field of type II diabetes and obesity prevention.Citrus flavonoids are found in citrus peel residue and have anti-oxidation,anti-cancer and anti-virus effects.Exploring the interaction mechanism of citrus flavonoids and digestive related enzymes is of great significance for the prevention and treatment of type II diabetes and obesity and the effective use of cholesterol by-product resources.In this paper,the quenching types of interactions of eriocitrin,eriodictyol,sinensetin with?-glucosidase,?-amylase and pancreatic lipase were studied by fluorescence spectroscopy.Binding constant,number of binding sites and thermodynamic parameters were used to explore the main force types of citrus flavonoids-enzyme complex stability.Synchronous fluorescence spectroscopy,three-dimensional fluorescence spectroscopy,circular dichroism spectroscopy and Fourier transform infrared spectroscopy were used to study the interaction of eriocitrin,eriodictyol,sinensetin binding with?-glucosidase,?-amylase and pancreatic lipase with the microenvironment and secondary structure of amino acids of enzymes.Molecular docking experiments were performed to explore the binding characteristics of eriocitrin,eriodictyol,sinensetin with?-glucosidase,?-amylase and pancreatic lipase,respectively.The main research contents and experimental results of this article are as follows:1.The fluorescence intensity of?-glucosidase decreased gradually with the increasing concentration of eriocitrin,eriodictyol and sinensetin.The binding processes were predominately driven by hydrophobic interaction and only a binding site and the binding constants were?7.02±0.22?×10⁴,?4.57±0.16?×10⁴ and?5.70±0.12?×10⁴ L·mol^-1?298 K?.The microenvironment around the tyrosine and tryptophan were changed after?-glucosidase interacted with the three citrus flavonoids.The three citrus flavonoids were combined with?-glucosidase resulted in changes in the secondary structure of the enzyme molecule.The results of molecular docking experiments showed that the interactions were found between three citrus flavonoids and amino acids residues in the?-glucosidase.2.The fluorescence intensity of?-amylase decreased gradually with the increasing concentration of eriocitrin,eriodictyol and sinensetin.The binding processes were predominately driven by hydrophobic interaction and only a binding site and the binding constants were?1.26±0.12?×10⁵,?4.90±0.08?×10⁴ and?1.51±0.12?×105 L·mol-1?298 K?.The microenvironment around the tyrosine and tryptophan were changed after?-amylase interacted with the three citrus flavonoids.The results of molecular docking experiments showed that the interactions were found between three citrus flavonoids and amino acids residues in the?-amylase.3.The fluorescence intensity of pancreatic lipase decreased gradually with the increasing concentration of eriocitrin,eriodictyol and sinensetin.The binding processes were predominately driven by hydrophobic interaction and only a binding site and the binding constants were?2.66±0.03?×10⁵,?1.58±0.04?×10⁵ and?7.94±0.06?×105 L·mol-1?298 K?.The microenvironment around the tyrosine and tryptophan were changed after pancreatic lipase interacted with the three citrus flavonoids.The results of molecular docking experiments showed that the interactions were found between three citrus flavonoids and amino acids residues in the pancreatic lipase.The results of this paper proved that the fluorescence intensity of?-glucosidase,?-amylase and pancreatic lipase decreased gradually with the increasing concentration of eriocitrin,eriodictyol and sinensetin by a static quenching mechanism and in this way the microenvironment and protein secondary structure of the three enzymes have also changed.This study will provide experimental basis for the application of eriocitrin,eriodictyol and sinensetin in the prevention and treatment of type II diabetes and obesity as"multi-target"natural products that can simultaneously combine with?-glucosidase,?-amylase and pancreatic lipase.