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Since milk whey is an abundant dairy by-product and a significant threat to the environment, its utilization is of great interest. The study compares valorization of lactose and lactates—the main carbon sources of whey—by fermentation—an environmentally friendly process. Antimicrobials released during fermentation by food-grade bacteria can help increase the microbiological safety of food. Propionic acid—a strong antimicrobial—is obtained mainly by the petrochemical route, yet there is increasing interest in its synthesis in biotechnological pathway. Five strains of propionic acid bacteria (Acidipropionibacterium acidipropionici, Propionibacterium cyclohexanicum, Propionibacterium freudenreichii, Acidipropionibacterium jensenii and Acidipropionibacterium thoenii) were investigated for their ability to produce organic acids and biomass using Na lactate or lactose as carbon sources. Selected fermentates were investigated for their antimicrobial efficacy during in vitro studies with foodborne pathogens: Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Bacillus subtilis and Staphylococcus aureus. Results confirm that the production of acids and biomass is considerably influenced by the added carbon source. The tested fermentates have strong and specific antimicrobial activity against Pseudomonas aeruginosa, Bacillus subtilis and Staphylococcus aureus. In addition, inhibition of Staphylococcus aureus and Klebsiella pneumonia depends on the activity of produced bacteriocins. The article also discusses the possibility of increasing the antimicrobial activity of fermentates by acidification. [ABSTRACT FROM AUTHOR]