Strain-Dependent Lactic Fermentation Primes Milk Proteins for Gastrointestinal Digestion Under Age-Specific Physiological Conditions

Lactic fermentation modifies milk proteins through the proteolytic activity of lactic acid bacteria, potentially altering their susceptibility to gastrointestinal digestion. This study evaluated how fermentation by Lactobacillus delbrueckii and Lactobacillus helveticus, individually and in co-culture, affects the structure of bovine milk proteins, their subsequent gastrointestinal digestion, and residual immunoreactivity. Milk was fermented with L. delbrueckii, L. helveticus, or their co-culture. Residual immunoreactivity of caseins (CNs) and β-lactoglobulin (βLG) was determined by ELISA, and protein profiles were evaluated by SDS-PAGE before and after in vitro gastrointestinal digestion using age-specific physiological models representative of 1-, 3-, and 6-month-old infants and adults. Fermentation with L. delbrueckii and the co-culture reduced the ELISA-detectable CN fraction by 7.42% and 8.85%, respectively, while L. delbrueckii was the only treatment to significantly decrease residual βLG immunoreactivity (2.17%). The effects of fermentation were most evident in the infant digestion models. Fermentation by L. delbrueckii was associated with more pronounced reductions in bands within the molecular weight regions expected for caseins and whey proteins during gastric digestion in the 1- and 3-month-old infant models, whereas these differences progressively diminished with digestive maturity. Throughout all digestion models, bands within the molecular weight range expected for βLG remained among the most persistent protein bands during gastric digestion. In silico analyses predicted preferential hydrolysis of caseins, the release of peptides with potential biological activities, and the differential persistence of linear allergenic epitope sequences among milk proteins. Strain-dependent lactic fermentation partially remodeled milk proteins before digestion, influencing their subsequent gastrointestinal behavior. The results indicate that the effects of fermentation depend not only on strain-specific protein modifications but also on the physiological conditions under which digestion occurs, providing new insights into the interaction between fermentation-induced protein remodeling and digestive maturity.