Antioxidant properties of caseinophosphopeptides and casein hydrolysates

Chelators are valuable ingredients used to improve the oxidative stability of many processed foods. Caseins and casein peptides have phosphoseryl residues capable of binding prooxidant metals such as iron and may have antioxidant activity. In 5% corn oil-in-water emulsions CPP (25 μM) inhibited the formation of lipid oxidation at both pH 3.0 and 7.0 as determined by lipid hydroperoxides and hexanal. Calcium (0–100 mM) had no influence on the antioxidant activity of CPP. Casein hydrolysates were tested for their antioxidant properties as hydrolysates represent a more economical source of phosphopeptides. Casein hydrolysates were more effective inhibitors of lipid oxidation than CPP at equal phosphorous content (0–310 μM) suggesting that antioxidant properties were not uniquely related to chelating prooxidant metals but also to free radicals scavenging activity. The antioxidant activity of CPP and casein hydrolysates was studied in phosphatidylcholine liposomes. CPP (\u3c1.0 mg/ml) and casein hydrolysates (0.3–1.7 mg/ml) were effective inhibitors of TBARS development when oxidation was promoted by iron. High amounts of CPP (\u3e1.0 mg/ml) were prooxidant while casein hydrolysates were only observed to be antioxidative. In the presence of peroxyl radicals, casein hydrolysates were more effective scavengers than enriched CPP (3–15 mM). When antioxidant activity was evaluated in cooked ground beef, TBARS formation was inhibited 75 and 39% by 0.5% CPP and casein hydrolysates, respectively, after 4 days of storage. The binding interactions of CPP and casein hydrolysates to iron were studied using isothermal titration calorimetry (ITC) with Fe-nitrilotriacetic acid (NTA) as the titrant. The observed binding constants were in the range of 5.8–1.7 × 103 M−1 for CPP and casein hydrolysates. The number of binding sites was below 1.0 suggesting that more than one phosphoseryl group is needed to bind iron or that other amino acids such as glutamic acid might participate in the binding. The interactions seem to be independent of pH since the apparent binding constants were similar between pH 3.0–7.0. These results show that CPP and casein hydrolysates are promising sources of natural antioxidants in foods since they have the potential ability to act as chelators of prooxidant metals and as free radical scavengers