Study On The Stability Of Flavonoids From Tartarty Buckwheat And The Interaction With Trypsin

Buckwheat has abundant flavonoid compounds, known as medicinal and edible plants. Buckwheat flavonoids include rutin, and rutin can be hydrolysised into isoquercetin and quercetin. Flavonoids have efficiency of hypoglycemic hypolipidemic, cancer prevention, enhance immunity, antiseptic, antioxidant, free radical scavenging. Trypsin can rapidly activate trypsinogen, and many other proenzyme, plays a vital role in the digestive process. The occurrence of many diseases are associated with trypsin, therefore we Study the stability of flavonoids from Tartarty Buckwheat and the interaction with trypsin.In this study, spectrophotometry is used to evaluate the influences of different conditions on flavonoids extracted from tartary buckwheat brans by two types of method, designated Extract#1and Extract#2. Conditions include varied preserving time, light, heat, pH, food additives, oxidizer, reducing agents and metal ions. The results showed:in reaction to preserving time, light, heat, acid, food additives, antioxidants, metal ions K+, Na+, Ca2+, Mg2+, both extracts had better stability; in reaction to alkaline, reducing agents, metal ions Cu2+, Fe2+, A13+, Fe3+, stabiliies were vulnerable. Under all conditions, Extract#1is slightly more stable than Extract#2. And Extract#1, Extract#2, Extract#3were extracted from buckwheat bran by ethanol and water. The effects of3kinds of extracts on trypsin were studied. Results showed Extract#1and Extract#2effectively inhibits the function of trypsin with Extract#2having an obvious advantage over Exact#1. Extract#3only showed inhibitory activity in higher concentration. The50%inhibiting concentration of3kinds of extracts were0.634g·L-1,0.166g·L-6.859g·L-1, respectively.In our work, the interactions between3flavonoids (Que, Rut, Iso) and trypsin were researched and analyzed by fluorescence spectroscopy. A number of related data and experimental results were made available. During the experiment, strong static fluorescence quenching and red shifts of light were showed with interactions between all3flavonoids (Que, Iso, Rut) and trypsin, meaning the binding effects have changed the microenvironment of tryptophan redidues in trypsin. The binding constants and the number of binding sites were calculated respectively. Based on thermodynamic principles, reactions between the3flavonoids and trypsin were spontaneous, entropy-increasing and energy-generating. The main forces were hydrophobic interaction and hydrogen bonds. Under the same concentration (with the molar ratio1:1), flavonoids absorption spectra and trypsin fluorescence spectra overlapped at300to500nm, which meant a nonradiative energy transferred between the3flavonoids (Que, Iso, Rut), and Trp residues on trypsin, thus resulted in fluorescence quenching.At last, in order to analyse the interaction between three flavonoids and trypsin better, UV spectrophotometry and related principle of enzyme kinetics applied, we studied the influence of3flavonoids compounds (Que, Rut, Iso) on trypsin activity. The results showed:under the concentrations set for this study, Que has a strong inhibitory effect on trypsin, while Iso and Rut had no significant inhibition on trypsin activity. The50%inhibiting concentration of Que is0.115mol·L-1. When the calculated value of Km for trypsin was3.97X10-4mol·L-1, the inhibition constant (Ki) of Que on trypsin is14.439x10-8mol·L-1. It can be seen that Que has a strong affinity with trypsin, compared with Iso and Rut, which makes it easier to combine with trypsin. From the experimental results in this chapter, we conclude that deep processing technology of Tartary buckwheat should be developed, leading to a wide array of industrialization development of Tartary buckwheat food.