Ultrafiltration of einkorn protein hydrolysates enhances antioxidant and techno-functional properties

Einkorn is an ancient cultivated wheat, characterized by high protein content, yet it remains an underutilized grain. This study explored the production of bioactive peptides from einkorn (Triticum monococcum) proteins using crude protease from Bacillus mojavensis sp. EBTA7, offering a potentially cost-effective alternative to purified enzymes due to the absence of downstream purification steps. Hydrolysis was performed under optimal conditions (pH 9, 60 °C), and hydrolysates were fractionated by ultrafiltration into four molecular weight (MW) groups (≤1 kDa, 1-5 kDa, 5-10 kDa, and > 10 kDa). The degree of hydrolysis ranged from 19.9% to 24.3%, with the hydrolysate produced at 20,600 U/g protein selected for subsequent ultrafiltration and characterization. Antioxidant assays revealed that the >10 kDa fraction exhibited the strongest activity, with the lowest IC₅₀ values of 15.0 mg/mL (DPPH) and 3.9 mg/mL (ABTS), along with the highest reducing power, while low-MW fractions showed superior Fe²⁺ chelation. Functional analyses indicated that high-MW peptides had enhanced oil holding capacity, emulsifying activity, and foaming stability, whereas smaller peptides improved emulsion stability. Structural characterization (hydrophobicity, FTIR, zeta potential, amino acid profiling) confirmed the role of hydrophobic and aromatic residues in bioactivities and techno-functional properties. In conclusion, mild hydrolysis combined with ultrafiltration effectively generated functional peptides from einkorn proteins. The >10 kDa fraction demonstrated promising antioxidant and technological properties, suggesting potential food applications, although further validation is needed. This research provides value-added opportunities for einkorn utilization and demonstrates the efficiency of crude protease-assisted hydrolysis coupled with ultrafiltration in producing functional peptide fractions.