Temperature-Dependent Metabolic Interactions Between Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus in Milk Fermentation: Insights from GC-IMS Metabolomics

Abstract

Streptococcus (S.) thermophilus and Lactobacillus (L.) delbrueckii ssp. bulgaricus are widely used as a combined starter culture for milk fermentation, often at temperatures of 37°C and 42°C. To investigate the metabolic interplay between these 2 species during the fermentation process, this study examined the growth and fermentation characteristics of different S. thermophilus strains cocultured with L. delbrueckii ssp. bulgaricus ND02 at these 2 temperature conditions. Gas chromatography-ion mobility spectrometry (GC-IMS) metabolomics was employed to analyze changes in the milk metabolome during 3 key fermentation stages: initiation (F0, pH 6.50 ± 0.02), curdling (F1, pH 5.20 ± 0.02), and endpoint (F2, pH 4.50 ± 0.02). The results showed that 42°C fermentation promoted rapid bacterial growth, with significantly reduced fermentation time compared with 37°C. Interestingly, 37°C fermentation favored the enrichment of volatile fatty acids like 2-methylpropanoic acid, 3-methylbutanoic acid, and ethyl acetate. In contrast, 42°C fermentation led to increased levels of ketones such as acetone, 2-hexanone, 2-pentanone, and 2-heptanone. Sensory evaluation indicated that the 42°C fermented milk had higher overall scores. Discriminatory flavor metabolites were more abundant during the later fermentation stage (F1 to F2), while the underlying metabolic pathways became increasingly active. These findings provide insights into the dynamic changes in volatile metabolite profiles of fermented milk produced under different temperature and time conditions using varied starter culture combinations. The results are valuable for optimizing dairy fermentation processes and product quality.