Separation and Purification of Modified Casein Hydrolysates Using Plastein Reaction and Their Antioxidant Activities
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Abstract:
Casein was hydrolyzed with papain at pH 6.5 for two hours, and the 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging capacity, superoxide anion scavenging capacity, and reducing power of the hydrolysis product (CH) were measured and found to be 35.89%±0.13%, 21.39%±0.33%, and 53.00%±2.00%, respectively. Subsequently, phenylalanine (PHE), tyrosine (TYR), and tryptophan (TRY) were used to modify the casein hydrolysate, respectively. The results showed that the antioxidant activities of the three products (CHPHE, CHTYR, and CHTRY) were highest when the plastein reaction time was six hours. Among them, the highest antioxidant activity was found in CHTRY, whose DPPH radical scavenging capacity, superoxide anion scavenging capacity, and reducing power were 46.84%±1.16%, 34.10%±0.32%, and 70.00%±2%, respectively (p<0.05). CHTRY was separated and purified using a Sephadex G-15 column, and the optimal elution result was obtained when deionized water was used as the elution solution, the concentration of loaded sample was 20 mg/mL, and the flow rate was 0.5 mL/min. Three peaks (P1, P2, and P3) were obtained, and their antioxidant activities were determined. The results showed that P2 had the highest antioxidant activity and its DPPH radical scavenging capacity, superoxide anion scavenging capacity, and reducing power were 58.46% ±0.57%, 38.42%±0.47%, and 80.00%±0.02% (p<0.05), respectively. The P2 product was further separated and identified by liquid chromatography, and a single peak was obtained, confirming a relatively high purity of the product. Finally, antioxidant peptides with a high purity were obtained.
