Relationship between antigenicity and allergenic epitopes of extensive hydrolyzed whey protein: LC-MS/MS-based analysis

Whey protein is the main protein in milk that causes allergic reactions in infants and children. In this work, Alcalase and Neutrase were combined to hydrolyze whey protein concentrate (WPC). The degree of hydrolysis (DH) and antigenicity reduction rate of WPC were 26.45% and 59.41% after hydrolyzed at the mass ratio of Alcalase and Neutrase of 1:1 and the enzyme to substrate ratio of 6000 U g−1 at pH 8.5, 50 °C for 3 h. HPLC results showed that when DH was about 26%, the molecular weight distribution of WPC hydrolysate less than 3000 Da exceeded 95%, and 18.33% of WPC hydrolysate was less than 180 Da (free amino acids). LC-MS/MS was used to detect peptides with 8–25 amino acid residues, the analysis indicated that most of the allergenic epitopes of β-lactoglobulin were destroyed, especially the major allergenic peptide fragments at positions 41–60 and 102–124. In conclusion, the combined hydrolysis of Alcalase and Neutrase significantly reduced the antigenicity of whey protein. This study could provide a potential value for the of WPC hydrolysates as a hypoallergenic infant formula.