Temperature-controlled ultrasonication improves protein extraction from soybean okara by various energy-dependent mechanisms

The food industry by-product soybean okara is a sustainable source of soy proteins, but its proteins have low extractability, hindering efficient protein isolation. This study investigated how temperature-controlled ultrasound treatments (<30 °C) at varying energy inputs improve okara protein isolation. In three independent okara batches, the extractability of about 50% of the proteins was restricted by non-covalent and covalent aggregation, while the other 50% could not be extracted even in the presence of denaturing and reducing agents, suggesting physical inaccessibility. Ultrasound treatments at pH 9.0 with low applied energy (2,000 J/g fresh weight [FW] okara) enabled extraction of physically accessible aggregated proteins (extraction yield 49%). Increasing the applied energy (10,000-40,000 J/g FW okara) further improved the extraction yield, reaching up to 90%, through additional particle and cell disruption. Therefore, temperature-controlled ultrasonication enhances okara protein extraction via two mechanisms: aggregate dispersion at low applied energy and additional matrix disruption at higher applied energies. After precipitation at pH 4.5, protein isolates produced using ultrasound-assisted processes had purities of 43-47 g/100 g dry weight due to pronounced co-isolation of non-protein constituents like lipids. Using higher ultrasound energy levels during extraction improved the foamability of the produced okara protein isolates, likely due to a less extensively aggregated state of proteins in these isolates. However, overall solubilities and foaming performances remained sub-par, requiring further investigation towards improvement. This study provides new insights into how temperature-controlled ultrasonication facilitates the extraction and functionalization of (aggregated) okara proteins.