Study On The Separation And Purification Of Perilla Seed Antioxidant Peptide And Its Hypolipidemic Function

Perilla is a traditional medicinal and edible plant in my country.It has a history of more than two thousand years of medicinal use in China.Perilla is a treasure.Its roots,stems and leaves are rich in protein,fat,vitamins,carbohydrates and other nutrients.It has anti-allergic,hypolipidemic,anti-oxidant,and prevent cell aging and canceration.Perilla seeds are mainly used for the extraction of perilla oil,and the largest flow of by-product perilla seed meal is for livestock and poultry feed.Perilla seed meal has excellent protein odor and rich amino acid composition.It is a complete protein with a complete range of amino acids.This topic uses perilla meal as raw material to prepare perilla protein isolate by ultrasonic extraction and flash extraction,and study its functional properties and antioxidant activity.Further enzymatic hydrolysis was used to prepare perilla hydrolyzed peptides,and the hydrolyzed peptides were purified and identified to study their lipid-lowering and antioxidant activities.The main research contents are as follows:(1)Using ultrasonic and flash extraction assisted alkaline solution acid precipitation method to prepare perilla protein,and optimize the extraction process of perilla protein through response surface based on single factor test.The best process conditions for ultrasonic extraction are:material-liquid ratio1:20(g/m L),alkaline extraction p H 10,alkaline extraction time 20 min,under these conditions,the extraction rate of perilla protein isolate stabilized at 53.85%.The optimal process conditions for flash extraction are:material-to-liquid ratio 1:20(g/m L),flash p H 10,flash extraction time 30 s.Under these optimized conditions,the extraction rate of perilla protein isolate is stable at 46.54%.(2)Ultrasonic extraction of protein isolate(UEOPI)and Flash extraction of protein isolate(FEOPI)functional properties and anti-oxidation studies have shown that UEOPI has better DPPH,ABTS,O₂^-,·OH radical scavenging ability and reducing power;higher solubility(NSI),water holding capacity(WHC),emulsification(EAI),emulsification stability(ES),foaming and foaming stability.Therefore,the follow-up experiment selects the ultrasonic extraction protein isolate(UEOPI)with higher extraction rate,better antioxidant activity,and better functional properties for research.(3)Use alkaline protease to enzymatically hydrolyze UEOPI.After ultrafiltration,four components with molecular masses of<3 k Da,3-5 k Da,5-10 k Da and 10 k Da are obtained,O₂^-,·OH radical scavenging rate and reducing power)The research screened out the strong antioxidant component as molecular weight<3 k Da.DEAE-32 ion exchange chromatography further separates and purifies the components with molecular weight less than 3 k Da to obtain three components D1,D2,and D3,and determine their DPPH,ABTS,O₂^-,·OH radical scavenging rate and reducing power,The results show that D3 has the strongest antioxidant activity.D3 was further separated and purified by Sephadex G-25 gel chromatography to obtain three components of G1,G2,and G3.The DPPH,ABTS,O₂^-,·OH radical scavenging rate and reducing power were measured,and the resistance of component G2 was determined.The oxidation activity is the highest.According to LC-MS-MS,the antioxidant activity peptide G2 is composed of7 amino acids,with a molecular mass of 790.3755 Da,and its amino acid sequence is AADCRAK,that is,alanine(Ala)-alanine(Ala)-Asparagine(Asp)-Cysteine??(Cys)-Arginine(Arg)-Alanine(Ala)-Lysine(Lys).(4)The activity of GSH-Px,SOD,CAT and MDA content in the liver tissues of the tested mice were tested to understand the effect of the active peptide G2 of perilla seed meal on the antioxidant capacity of the mice.The changes in the contents of TC,TG,LDL-C and HDL-C are used to understand the effect of mouse G2 on the blood lipid level of mice.The results show that the mice treated with G2 in the low,medium and high dose groups are higher than those treated by G2 intragastric administration.In the lipid model group,the MDA content in mice decreased significantly or extremely significantly(P<0.05 or P<0.01);the activities of GSH-Px,SOD and CAT in mice increased significantly or extremely significantly(P<0.05 or P<0.01));the content of TC,TG and LDL-C in mice is significantly or very significantly reduced(P<0.05 or P<0.01),and the content of HDL-C is significantly or very significantly increased(P<0.05or P<0.01),indicating Perilla active peptide G2 has the effect of improving the antioxidant capacity of mice,protecting the body from free radical attacks,and regulating the abnormal blood lipid metabolism of hyperlipidemia mice.