Understanding the potential of pressurized CO₂ to assist enzymatic hydrolysis of egg yolk waste for the production of hydrolysates rich in protein, peptides and amino acids with antioxidant activity

Egg yolk waste from the poultry processing industry was valorized to produce protein hydrolysates using enzymatic hydrolysis in pressurized CO₂ media. Hydrolysis parameters such as pepsin concentration (1.0–2.0%, w /w), CO₂ pressure (60–300 bar) and temperature (37 and 50 °C) were evaluated. First, lipids were removed (92.7% recovery) using supercritical CO₂ (SC-CO₂) + ethanol (10%, w /w) at 300 bar/50 °C, enhancing the protein content in the defatted residue to 61.7%. Then, defatted residues were enzymatically hydrolyzed in pressurized CO₂ media (37 °C/300 bar/3 h) to produce hydrolysates rich in soluble protein, low-molecular weight peptides, free amino acids with higher degree of hydrolysis (20.3%) and antioxidant power compared to the control. The FTIR and SEM analyses confirmed that CO₂-induced structural changes, protein unfolding, and facilitated enzyme accessibility. Overall, integrated SC-CO₂ extraction and pressurized CO₂ hydrolysis provided a sustainable approach to obtain protein hydrolysates from egg yolk waste with potential applications in functional foods and nutraceuticals.