Purification of Bacteriocins Produced by Lactic Acid Bacteria.

SAAVEDRA LUCILA, CASTELLANO, PATRICIA; SESMA, FERNANDO

Public Health Microbiology: Methods and Protocols

Humana Press

Año: 2004; p. 331 - 336

Bacteriocins are antibacterial substances of a proteinaceous nature that are produced by different bacterial species. Lactic acid bacteria (LAB) produce biologically active peptides or protein complexes that display a bactericidal mode of action almost exclusively toward Gram-positive bacteria and particularly toward closely related species. Generally they are active against food spoilage and foodborne pathogenic microorganisms including Bacillus cereus, Clostridium perfringens, Staphylococcus aureus, and Listeria monocytogenes (1,2).Bacillus cereus, Clostridium perfringens, Staphylococcus aureus, and Listeria monocytogenes (1,2).Listeria monocytogenes (1,2). There is an increased tendency to use natural occurring metabolites to prevent the growth of undesirable flora in foodstuffs. These metabolites could replace the use of chemical additives such as sorbic acid, sulfur dioxide, nitrite, nitrate, and others. For instance, bacteriocins produced by LAB may be promising for use as biopreservaties (3,4).,4).,4). Bacteriocins of lactic acid bacteria are typically cysteine-rich, cationic, hydrophobic peptides and differ widely in many characteristics including molecular weight, presence of particular groups of amino acids, pI, net positive charge, and post-translational modifications of certain amino acids. This heterogeneity within the LAB bacteriocins may explain the different procedures for isolation and purification developed so far. The methods most frequently used for isolation, concentration, and purification involve salt precipitation of bacteriocins from culture supernatants, followed by various combinations of gel filtration, ion-exchange chromatography, and reverse-phase high-performance liquid chromatography (RP-HPLC). In this chapter, a protocol is described that combines several methods used in our laboratory for the purification of two cationic bacteriocins, Lactocin 705AL and Enterocin CRL10, produced by Lactobacillus casei CRL705 and Enterococcus mundtiiLactobacillus casei CRL705 and Enterococcus mundtii CRL10, respectively