The Study Of Isolation And Purification Of Rice Peptide And Its Antioxidant Activity

Using protease to separate the rice peptide and the starch had some advantages: the reacting conditions were mild; the technology was simple and the most important one was that the properties of the rice starch were excellent. As the outgrowth of the rice starch, rice peptide had some activities, such as hypocholesterolemic and antioxidant activities. But in the hydrolysates, there were still some kinds of sugar, which would influence the quality of the peptide. In this study, protease was used to hydrolyze rice to separate rice protein and rice starch at the same time. The emphases are the eluting of sugar and the hypocholesterolemic activities and the free radical scavenging abilities of the rice peptides. Amino acid sequences of the single peptides with high activity were identified finally.Based on the the sugar in the hydrolysates came from the step of hydrolysis, we analyzed the component and the molecular weight of the peptide and the sugar to confirm that D201resin could separate the sugar and the peptide effectively. Optimum conditions of ion-exchange are: initial buffer pH 7 with 0.01 mol / L ion strength, the molar ratio of borax and sugar 1: 8, and the ratio of sample and resin 1: 1. The elution we have chosen was water first (1v) and then 0.1 mol/L HCl (2v). The peptide sample was achieved after rotation evaporation and freeze-drying. The protein content (based on dry) of the peptide was 80.0% and the sugar content (based on dry) of the peptide was 9.0%.Jing mi and nuo mi as raw, we studied the hypocholesterolemic activity and the radical scavenging abilities of the peptides hydrolyzed by Protease N and Alcalase. The results showed that the jing mi peptide hydrolyzed by Alcalase and Protease N had a little high activity of hypocholesterolemic(38%when the peptide concentration was 7.5mg/mL), but lower than soy peptide(44-63%when the peptide concentration was 0.2mg/mL); Jing mi peptide hydrolyzed by Protease N had the highest ability of radical scavenging: the IC50 of DPPH was 7.5mg/mL;the IC50 of OH was 12.6mg/mL;the IC50 of O2- was 24.7mg/mL.RP-HPLC was used to purify the Jing mi peptide hydrolyzed by Protease N.14 fractions from the separation of the antioxidant peptides were obtained with a preparative RP-HPLC column. The most active DPPH radical scavenging fraction was fraction 6(IC50 was 32.6ug/mL); the most active hydroxyl radical scavenging fraction was fraction 2 (66.2% when the peptide concentration was35ug/mL) and the most active superoxygen radical scavenging fraction was fraction 4(39.5% when the peptide concentration was35ug/mL). The structures of the three kinds of fractions were determined by LC-MS. The possible amino acid sequences were: VAK/AKV,AAAA,VN/VGG,VDM/VFV of fraction 2, CCIIK/CCLIK/CCIIQ/CCLIQ,MFM and WDP/WLV/WIV of fraction 4 and EYDP/EYLV/EYIV,PGHV/GHVP and IPV/PVI of fraction 6.