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The subject of the study is a strain of Bacillus subtilis PSF-19 isolated by us from vegetable raw materials (Passiflora preparation), which is capable of producing antimicrobial substance (AMV) suppressing pathogenic bacteria. The article discusses the method and conditions of extraction from the culture fluid fraction of AMV, active against Listeria monocytogenes – one of the dangerous intestinal pathogens that infect food. Using biochemical methods and mass spectroscopy, the molecular weight and the peptide nature of the active fraction of AMV were determined. A fraction of AMV with a molecular weight of 3,4–3,6 kDa has bactericidal activity, which is destroyed by treatment with proteolytic enzymes, which allowed it to be attributed to the group of low-molecular antimicrobial peptides – bacteriocins. The studies allow to consider the strain Bacillus subtilis PSF-19 as a producer of bacteriocin, especially effective against pathogens of intestinal listeriosis. The establishment of the fact of destructibility of proteolytic enzymes, finding the conditions of microbiological synthesis of AMV, its isolation and accumulation for research, provide opportunities for practical use as a means for decontamination of the environment instead of traditional antibiotics.