| This work was funded by the Key Projects in the National Science and Technology PillarProgram during the Twelfth Five-Year Plan Period (2012BAD33B03). Alkaline protease, trypsinand papain were used to hydrolyze the corn gluten meal, the results showed that the hydrolysatestreated by alkaline protease had better antioxidant activity and high degree of hydrolysis. Theselected alkaline protease and flavourzyme were combined to prepare the corn antioxidant peptide.And two enzymatic hydrolysis models were established by multiple linear regression design andresponse surface design, the obtained models fit well. Through analyzing the models, optimalenzymatic process parameters were obtained as follows: alkaline protease: pH9.5, the amount ofenzyme was8%, the ratio of solid to liquid was1:15and the hydrolysis time was75min; Alkalineprotease-flavourzyme: pH7, the amount of enzyme was4.2%and the hydrolysis time was66min.Experiments were done at the optimal hydrolysis parameters, the DPPH scavenging activity andFe2+chelating activity of the obtained hydrolysates were26.3%and46.3%respectively. Thehydrolysates were separated by ultrafiltration to obtain the hydrolysates with different molecularweight distribution, and their antioxidant activities were determined. The antioxidant activities ofthe hydrolysates or peptides were evaluated by free radical scavenging capacity, metal ionchelating activity and lipid peroxidation inhibitory capacity. The results indicated that those withmolecular weight <10kDa exhibited highest antioxidant activity in all relevant assays. Thehydrolysates were subsequently purified by gel filtration chromatography, and fraction F3showedthe highest antioxidant activity. Three peptides were identified from fraction F3usingLC-ESI-Q-TOF MS/MS as Leu-Pro-Phe (375.46Da), Leu-Leu-Pro-Phe (488.64Da) andPhe-Leu-Pro-Phe (522.64Da). These peptides exhibited good free radical scavenging activity andlipid peroxidation inhibitory effect.Zein protein was extracted from the by-product corn gluten meal. The obtained zein protein was first hydrolyzed by four different proteases. Among hydrolysates produced, alkaline proteasehydrolysates exhibited the highest antioxidant activity. One regression model was established byuniform deign to optimize the hydrolysis conditions. The optimal hydrolysis conditions were: pHof8.5, an E/S ratio of12%and a hydrolysis time of90min. The hydrolysates were then separatedby ultrafiltration, and hydrolysates with molecular weight <3kDa exhibited highest antioxidantactivity in all relevant assays. The hydrolysates with molecular weight <3kDa were subsequentlypurified by gel filtration chromatography, and fraction F3showed the highest antioxidantactivities. Two peptides were identified from fraction F3using LC-ESI-Q-TOF as Pro-Phe(263.13Da) and Leu-Pro-Phe (375.46Da). Peptide Pro-Phe exhibited good antioxidant activity.Effects of temperature, pH, food ingredients, metal ions and gastrointestinal proteases onDPPH/ABTS radical scavenging activity of corn antioxidant peptide Leu-Pro-Phe weredetermined. The results showed that peptide Leu-Pro-Phe exhibited strong thermal stability, it stillshowed strong free radical scavenging activities when it was heated to100℃for5hours. Thestructure of Leu-Pro-Phe can not be broken in alkaline and acid environments. Furthermore, atalkaline pH, peptide Leu-Pro-Phe was more effective to scavenge ABTS radical. The foodingredients significantly affected the antioxidant activities of Leu-Pro-Phe. The DPPH radicalscavenging activity significantly increased (p<0.01) when sucrose, NaCl and citric acid wereadded. The DPPPH radical scavenging activity of peptide Leu-Pro-Phe was significantlyinfluenced (p<0.05) by K+, Ca2+, Mg2+, Zn2+and Cu2+ions. However, the ABTS radicalscavenging activity of Leu-Pro-Phe was only affected by Cu2+ions, the activity was significantlydecreased (p<0.01) when high concentrations of Cu2+ions were added. The peptide Leu-Pro-Phestill exhibited strong free radical scavenging activities after digested by pepsin and trypsin. |
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