Study On Extraction,Isolation And Antioxidant Structure-Activity Relationship Of Polyphenols In Lotus Leaf

Lotus leaf belongs to the lotus leaf of Nymphaeaceae.It is distributed in the Yangtze River and Yellow River basins in China.It has been used as food and medicine since ancient times,and it has the advantages of safety and non-toxic side effects.The lotus leaf is rich in many biologically active ingredients,such as polyphenols,flavonoids,alkaloids and anthocyanins,among which polyphenols are the main active substances.Polyphenols have a variety of biological activities,such as anti-oxidation,hypolipidemic,antibacterial and enhance immunity.At present,foods,medicines,etc.developed from nature,safe and harmless plants are increasingly being sought by the general public.However,resources of lotus leaf have not been fully utilized in our country,and a large number of lotus leaves have been abandoned in lakes.Extracting natural functional elements with excellent biologicalactivity from lotus leaf can effectively improve the economic value.This paper mainly cover four parts: extraction,purification,anti-oxidation and anti-oxidation structure-activity relationship study of lotus leaf polyphenols,providing references for the development and utilization of lotus leaf resources.The main tests and results are as follows:(1)The polyphenols in the lotus leaf were extracted and the response surface optimization experiment was performed on the basis of the single factor experimental results.After the Box-Behnken experiment design,the optimal conditions were simulated and predicted,and the optimal extraction conditions were: The extraction temperature was 78?,the extraction time was 95 min,the p H value was 4.7,and the ratio of solid to liquid was 1:30.The verification experiment showed that the extraction rate of the leaf polyphenols was 26.49 mg GAE/g,which was in good agreement with the predicted value.(2)The primary purification of lotus leaf polyphenols was carried out by ion precipitation method and AB-8 macroporous resin.The optimal conditions for ion precipitation and purification of lotus leaf polyphenols were determined by ion precipitation in the purification experiment of lotus leaf polyphenols: Under the condition of 25?,the mass ratio of precipitant ZnCl2 was 1:5,and the optimal precipitation p H was 6.Under this condition,the precipitation rate of polyphenols from lotus leaf reached 84.5%.The best purification conditions of lotus leafpolyphenols were determined by AB-8 macroporous resin purification of lotus leaf polyphenols: loading concentration 1.50 mg/m L,loading rate 2BV/h;elution conditions 70% ethanol solution eluted at a flow rate of 1BV/h,and the macroporous resin had a resolution of 82.6%.(3)Separation and purification of polyphenols from the leaf polyphenols by HSCCC,and the isolation conditions for isolation and purification of polyphenols from lotus leaf were determined using a solvent system of n-hexane:ethyl acetate:methanol:water(1:6:1:6,v/v).Two kinds of polyphenols were obtained;Adopted two-phase solvent system of n-butanol:methanol:water(5:1:4,v/v)was used to separate and obtain a polyphenol.(4)The three target component(Peak1,Peak2,Peak3)obtained was separated standards detected by HPLC analysis of a control experiment,three kinds of the target component is very likely to give a structure glycosides are Violetin,isoquercitrin and quercetin.(5)The antioxidant activity of three target components of lotus leaf polyphenols isolated and purified by HSCCC under the three systems of scavenging hydroxyl radical,DPPH radical and ABTS radical was determined.The three target components were tested to have a certain scavenging effect on all three kinds of free radicals,and all had a concentration-dependent effect.The antioxidant activity in the hydroxyl radical,DPPH radical,and ABTS radical systems was in the followingorder: quercetin > isoquercitrin> astragalin.(6)Quantum chemical calculations were performed on three target components using the Gaussian software package.The results show that C3-OH contributes to the coplanarity of the molecules.When C3 is due to glycosides,it affects the conjugation between B-ring and C-ring,making The conformation of the molecule is reversed,which affects the antioxidant activity;the BDE values of the three target components are calculated and the anti-oxidation active sites are mainly located on the B-ring;Using the theory of frontier orbits to calculate the components to get the theoretical scavenging capacity of free radicals has the following sequence: quercetin> isoquercitrin> astragalin,consistent with the results of antioxidant experiments.