| The resource of animal's blood is rich in China.It would be of great waste of protein and mind cause pollution to the environment if the blood was not rationally tapped and directly discharged into the environment.So it's of great significance to industrialize the production of animal-blood products and protect the environment by using modern high-tech to develop high value-added products and further improve blood's utilization. The aim of this study was procine hemoglobin.Used the protease to hydrolyze it and gained products of enzyme hydrolysis with anti-oxidation activity.For further exploration of the feasibility of extracting antioxidative peptide from animal blood hemoglobin,the anti-oxidation activity of the products of enzyme hydrolysis were tested in some measurements of anti-oxidation activity.1.Single factor and orthogonal experiments were desigend to study the enzyme hydrolysis function of neutral protease,papain,trypsinase single enzyme system on blood hemoglobin.The degree of reducing power of different enzyme with the best enzyme hydrolysis process was determined by assaying the degree of hydrolysis(DH) and reducing power.According to the results of DH and reducing power,the relationship between degree of DH and reducing power with anti-oxidation activity of the products of enzyme hydrolysis was not that the greater the DH,the stronger the reducing power.In the condition of a certain time and through the orthogonal experiments,the fixed conditions for the neutral protease enzymatic hydrolysis with the best reducing power were temperature 40℃,pH7.0,hydrolysis time4 hours,the substrate concentration 8%,enzyme6000U/g,and the DH and reducing power can reach 24.35%,0.432 respectively. The fixed conditions for the papain enzymatic hydrolysis with the best reducing power were temperature 53℃,pH7.5,hydrolysis time4 hours,the substrate concentration 6%,enzyme7500U/g,and the DH and reducing power can reach 18.21%,0.302 respectively. The fixed conditions for the trypsinase enzymatic hydrolysis with the best reducing power were temperature 43℃,pH8.0,hydrolysis time 3hours,the substrate concentration 9%,enzyme6000U/g,and the DH and reducing power can reach 21.66%,0.342 respectively. The experimental results showed that the anti-oxidation activity of the enzyme hydrolysis products of neutral protease were both superior to trypsinase and papain.Compare the clearance rate on·OH and O2- between pig's blood hemoglobin and the enzymolysis solution of pig's blood hemoglobin hydrolysed by 3 enzymes at the optimum conditions.The results showed that all three enzymolysis products presented good ability to clean up·OH and O2-,obviously higher than that of original pig's blood hemoglobin. The enzymolysis componets of neutral protease had the greatest clearance capacity,the clearance rate on·OH and O2- were 56.23%and 45.61%respectively.2.The enzymolysis solution of procine hemoglobin was decolorizated by the activated carbon and the CMC at the temperature 50℃,pH4.0,decolorization time 20min,the amount of activated carbon and CMC were 0.20%and 0.30%.The results showed that the decolorization ability of the CMC was better than that of the activated carbon,and 10.85%protein lost.The anti-oxidation activity of the neutral protease's enzymolysis solution of procine hemoglobin was compared before and after decolorization.After decoloration,the reducing power decreased by 21.74%,the clearance rate of the hydroxyl radical decreased by 25.15%,and the clearance rate of superoxide anion decreased by 30.93%.3.The enzymolysis solution of procine hemoglobin was filtrated respectively by ultra filtration tube with molecular weight 100KDa,10KDa,5KDa.Then studied the anti-oxidation activity of the separated components of different molecular weigh.The results showed that the strongest reducing power,the strongest capacity of getting rid of superoxide anion and the strongest capacity of scavenging hydroxyl radical of the separated components were all the molecular weight less than 5KDa.At a concentration of 50ug/mL condition,the reducing power achieved 0.37,the clearance rate of superoxide anion was39.41%,and the removal rate of hydroxyl radical was 50.32%.4.selected the concentration of neutral protease of enzyme solution 0.00%,0.05%,0.10%,0.20%,0.50%and studied the anti-oxidation activity in lard and vegetable oil separately.Compared the anti-oxidation activity with Vc and BHT.The results showed that the enzyme soIution can inhibit the formation of peroxides in the oil system.When the concentration of enzyme0.05%the anti-oxidation activity in lard and vegetable oil were6.78%,6.41%,When the concentration of enzyme0.5%the anti-oxidation activity in lard and vegetable oil were 9.35%,10.42%.At the same concentration of enzyme solution the antioxidant capacity on the lard and vegetable oil was higher than VC but lower than BHT.5.According to rat liver tissue homogenate and liver mitochondria on bath temperature in vitro test,studied the enzymatic hydrolysis of procine hemoglobin on the inhibition of MDA.The antioxidation activity of the enzymatic hydrolysi about procine hemoglobin was evaluated by the inhibitory effect.The results showed that when the enzyme solution reached the concentration of 10mg/mL,the clearance capacity on MDA in rat liver tissue homogenate achieved a significant level(P＜0.05),the clearance capacity on MDA in liver tissue mitochondria got a significant level(P＜0.05) as well.
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