Synergistic enhancement of physicochemical properties, flavor, and functionality of fermented milk through co-fermentation with Lactiplantibacillus plantarum LP315 and Streptococcus salivarius subsp. thermophilus S10

This study aimed to identify the optimal ratio of Lactiplantibacillus plantarum LP315 (LP315) and Streptococcus salivarius subsp. thermophilus S10 (S10) and to evaluate the effects of co-fermented milk during storage. The physicochemical properties of co-fermented milk produced using different LP315:S10 ratios (1:1, 1:10, 10:1, 1:100, and 1:1000) were systematically assessed. Co-fermentation at a 1:1 ratio yielded fermented milk with superior viable counts, acidity, viscosity, water-holding capacity, texture, and sensory attributes after 1 day of storage. Based on this optimal ratio, the physicochemical properties of co-fermented milk were further compared with those of single-strain fermented milks on days 1, 7, 14, and 21 of storage. The results showed that co-fermented milk showed improved physicochemical characteristics compared with single-strain fermented milks, particularly on day 1. Untargeted metabolomics analysis identified 12 key differential metabolites significantly enriched in co-fermented milk and associated with flavor and functionality, including carbohydrates (D-glucose and lacto-N-tetraose), vitamins (pyridoxal and pantothenic acid), organic acids (L-2-hydroxyhexanoic acid, 3-(3-hydroxyphenyl) propionic acid, and L-lactic acid), and amino acids (phenylalanine, leucine, proline, glutamic acid, and threonine). These findings highlight LP315 as a promising adjunct strain for use with commercial starters in the production of functional fermented dairy products.