Proteolytic Activity and Volatile Compound Formation by Staphylococcus warneri, Rothia uropygioeca, and Enterococcus lactis in Pea Protein Fermentations

The transition toward clean label, plant-based foods has intensified the search for natural fermentation strategies to reduce off-flavors, while improving taste and nutritional quality of plant proteins. This study evaluated 142 microorganisms isolated from animal-derived ripened foods, selecting four proteolytic Gram-positive cocci (Staphylococcus warneri GC 13 and 28, Rothia uropygioeca GC 15, and Enterococcus lactis GC 40) for fermenting pea protein isolate and textured pea protein as plant-based model substrates. Fermentations at 25 °C for 48 h significantly modified microbiological and physicochemical parameters in a matrix-dependent manner, with textured pea protein supporting higher lactic acid bacteria growth and stronger acidification than pea protein isolate. Free amino acid profiling demonstrated strain- and matrix-dependent proteolytic activity, with GC 13 and 15 promoting the highest total free amino acids in pea protein isolate, while GC 15, 28, and 40 markedly increased multiple amino acids in textured pea protein. Biogenic amine analysis revealed that tyramine was the predominant amine, reaching 162–212 ng/g in inoculated textured pea protein and 348 ng/g in uninoculated controls, whereas only trace levels were detected in the pea protein isolate. Volatile organic compound analysis grouped compounds into three clusters, evidencing a reduction in native beany aldehydes and the formation of alcohols, ketones, and acids, particularly in textured protein, indicating substantial reshaping of the aroma profile. Overall, the results showed that the evaluated strains can be exploited as clean-label tools to enhance proteolysis, attenuate off-flavors, and maintain acceptable biogenic amine levels in pea protein matrices, with R. uropygioeca 15 emerging as a promising candidate for plant protein fermentation.