Fermentation in Coculture with Limosilactobacillus fermentum (LBF 433) and Lacticaseibacillus casei (LBC 237) as a Strategy to Diversify Peptides in Milk

This study investigated the impact of coculture fermentation using Limosilactobacillus fermentum (LBF 433) and Lacticaseibacillus casei (LBC 237) on the generation of bioactive peptides from whole milk. Fermentations with individual strains and coculture were performed, followed by peptide identification using nanoLC-MS/MS and de novo sequencing via PEAKS XPRO. Coculture increased the number of peptides, with L. fermentum predominantly contributing to proteolysis. Most peptides in coculture overlapped with the LBF 433 monoculture. Physicochemical analysis showed peptides with near-neutral charge, moderate hydrophobicity, and high aliphatic residue content─traits linked to improved solubility and bioactivity. Principal component analysis revealed two main axes: hydrophobicity related to aliphatic residues and isoelectric point negatively correlated with the acid–base profile. These features affect membrane interaction and bioavailability. Overall, coculture fermentation is an effective strategy to modulate peptide profiles and obtain compounds with desirable structural and functional properties for use in functional foods and nutraceuticals.