Abstract
Bioactive peptides can be released, during gastrointestinal digestion or food processing, from many plant and animal proteins and represent an important source of potential health-enhancing components. Therefore, the aim of this study is to investigate antioxidant, anticancer and antihypertensive activities of proteins and peptides extracted from egg yolk. Defatted egg yolk (EY) was hydrolysed using pepsin and pancreatin and sequentially fractionated by ultrafiltration, followed by gel filtration (EYGF), resulting in three different fractions of which two (EYGF-23 and EYGF-33) effectively inhibited the oxidation of linoleic acid in a dose-dependent manner. Both fractions (80 mg/ml) exhibited antioxidant activity comparable to trolox and butylated hydroxytoluene (0.2 mg/ml). The antioxidant mechanism involved superoxide anion and hydroxyl radicals scavenging, ferrous chelating and reducing ability. The protective effect of one of the fractions (EYGF-23) was also tested in Caco-2 colon cancer epithelial cells, exposed to 3 mM of tert-butyl hydroperoxide (t-BHP) to induce oxidative stress. Induced cell toxicity by t-BHP was significantly inhibited when Caco-2 cells were treated with 1.0 mg/ml of the fraction for 24 hours. This fraction protected the cells by inhibiting intracellular ROS as well as lipid peroxidation products by 60.0% (p <0.001) and 24.15%, (p <0.05). In addition, total glutathione level and superoxide dismutase activity were significantly elevated to 0.29 μM (p <0.001) and 55.10% (p <0.01), respectively, in comparison to cells treated with t-BHP alone. Amino acids lysine, proline, tyrosine and tryptophan in this fraction may be responsible for antioxidant activity. The main highlight of this study was that cytotoxic effects of the second fraction (EYGF-33) was observed in vitro in cancer colon cells but not in normal colon cells. Apoptosis was mediated by mitochondrial ROS, evidenced by PARP-1 cleavage. Reduced cell viability could be explained by cell cycle arrest in the S-phase which stopped DNA synthesis. Arginine, lysine, leucine, alanine, valine and tryptophan residues, which are mainly presented in the second fraction, may be responsible for the anti-proliferative activity. A third fraction (EYLCF-16) was successfully purified from egg yolk protein by both gel filtration and HPLC. This fraction exhibited high angiotensin converting enzyme (ACE) inhibitory activity (91.8%) at 10 mg/ml and IC50 value (1.53). ACE inhibition exhibited by the fraction was probably due to the positively charged amino acid residues, lysine and arginine, and hydrophobic amino acid residues, leucine and tryptophan. These project findings indicate a substantial potential for producing peptides with antioxidant, anticancer and ACE inhibitory activity from egg yolk.