| Summary: | Proteins and protein-derived materials, such as enzymatic hydrolysates, have been shown to improve the oxidative stability of food lipids. However, the mechanisms of protein antioxidant action are complex, particularly in heterogeneous, dynamic environments such as food systems. The goal of this research is to provide a fundamental understanding of the various mechanisms by which proteins inhibit oxidative reactions in food lipid dispersions. The presence of low concentrations of the whey protein β-lactoglobulin (β-Lg) present in the continuous phase of oil-in-water emulsions was observed to inhibit lipid oxidation. Furthermore, the antioxidant activity of continuous phase β-Lg was significantly improved by thermal denaturation (95°C; 15 min) at pH 7.0, despite having lower iron binding values and concentration of reactive sulfhydryls than untreated, native β-Lg. The observed enhancement in antioxidant activity of heated β-Lg seems to result from improved solvent accessibility of radical scavenging amino acids, and/or changes in β-Lg's surface activity, leading to increase interactions with lipophilic oxidants. Cysteine and tryptophan residues in native β-Lg dispersed in the continuous phase of emulsions were observed to oxidize prior to emulsified lipid, suggesting that these amino acids possess antioxidant activity. However, methionine residues in β-Lg did not appear to participate in free radical scavenging in this system, despite its reported oxidative lability. This may be due to the nature of the endogenous oxidant (i.e. lipid-derived radical species) or to the fact that β-Lg's methionine residues have low solvent accessibility when the protein is in its native conformation. Finally, it was observed that β-Lg's antioxidant activity is enhanced with enzymatic (e.g. chymotrypsin) hydrolysis. Increased peroxyl radical scavenging activity and iron chelation seem to be responsible for the increased antioxidant activity of hydrolyzed β-Lg. Liquid chromatography - mass spectrometry (LC-MS) was used to assess the oxidative modification of three specific amino acid residues in the selected chymotrypic hydrolysates of β-Lg during the period preceding the formation of lipid oxidation products. Significant oxidation of tyrosine and methionine were observed during this period, suggesting that these residues are capable of free radical scavenging, thereby conferring improved oxidative stability to oil-in-water food emulsions.
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