Online volatilome dynamics of Lactiplantibacillus plantarum WCFS1 during growth in MRS, and comparative insights with other wine-associated strains

Lactiplantibacillus plantarum is a metabolically versatile and ecologically adapted lactic acid bacterium widely used in food fermentations. L. plantarum WCFS1, originally isolated from human saliva, represents one of the best-characterised model organisms in lactic acid bacteria research. However, knowledge of its volatilome dynamics during bacterial growth in microbiological media remains limited, although these conditions are relevant for studies that combine microbial biology with the evaluation of food-application potential. In the present study, proton-transfer-reaction time-of-flight mass spectrometry (PTR-ToF-MS) was applied to monitor, across eight fermentation cycles, the online evolution of the volatilome of L. plantarum WCFS1 grown in MRS medium. To also provide a comparative ecological perspective under standardised conditions, five wine-associated L. plantarum strains previously isolated from oenological products were included for comparative analysis. Gas chromatography–mass spectrometry (GC-MS) was employed as a complementary approach to support tentative metabolite annotation. Overall, 142 mass peaks differed significantly from the uninoculated medium, of which 133 were associated with a chemical formula, and 99 were tentatively identified. Multivariate analyses showed that fermentation time was the main source of volatilome variation, with the first four cycles corresponding to the most dynamic metabolic phase and later cycles reflecting stabilisation during stationary growth. Four main kinetic VOC profiles were identified: volatile depletion, transient accumulation, progressive accumulation, and late-stage recovery. Shared metabolites across strains included sulfur compounds, organic acids, branched-chain aldehydes/alcohols, and pyruvate-derived metabolites associated with carbohydrate and amino acid metabolism. WCFS1 showed a distinctive late-stage volatilome, with higher accumulation of pyruvate-, sulfur-, and lipid-related VOCs than wine-associated strains. These results support PTR-ToF-MS as an online sensor for microbial volatilomics and highlight strain-dependent volatile metabolism in L. plantarum.