Comparative Analysis of Antimicrobial and α-Amylase Inhibitory Activities and GC-MS-based Metabolic Profiling of Cow Milk Fermented by Lactic Acid Bacteria.

Lactic acid bacteria (LAB) play a key role in milk fermentation by producing a wide range of metabolites that influence the nutritional, functional, and health-promoting properties of fermented dairy products. The present study aimed to characterize the volatile metabolite profile of LAB-fermented cow milk obtained from different cow breeds using gas chromatography-mass spectrometry (GC-MS) and to identify metabolites potentially associated with its biological activities. Additionally, the antimicrobial activity of fermented milk was evaluated against Bacillus subtilis, Escherichia coli, Pseudomonas fluorescens, and Micrococcus luteus using the agar well diffusion method, while α-amylase inhibitory activity was determined using an in vitro assay. The antibacterial and α-amylase inhibitory activities of fermented milk samples were significantly higher (p < 0.05) than unfermented milk samples. Following ultrafiltration using 3 and 10 kDa membranes, the 3 kDa retentate from the Sahiwal cow milk (SCM) fermented milk sample exhibited the highest antimicrobial activity (inhibition zone: 1.78 cm), whereas the 10 kDa filtrate from the Holstein Friesian cow (HFCM) fermented milk sample showed the highest α-amylase inhibitory activity (51.90%). GC-MS analysis revealed a diverse array of fermentation-derived metabolites, including alcohols, phenols, fatty acids, esters, and heterocyclic compounds. Most of the compounds are known to exhibit antimicrobial, antioxidant, anti-inflammatory, antidiabetic, and anticancer properties. Overall, this study provides valuable insights into the metabolite composition of LAB-fermented cow milk and the influence of cow breed and LAB fermentation on metabolite production, highlighting the potential of fermented milk as a functional food with enhanced health-promoting properties.