Novel method to extract large amounts of bacteriocins from lactic acid bacteria

Abstract

Antimicrobial peptides, bacteriocins, produced by lactic acid bacteria were adsorbed on the cells of producing strains and other gram-positive bacteria. pH was a crucial factor in determining the degree of adsorption of these peptides onto cell surfaces. In general, between 93 and 100% of the bacteriocin molecules were adsorbed at pHs near 6.0, and the lowest (c5%) adsorption took place at pH 1.5 to 2.0. On the basis of this property, a novel isolation method was developed for bacteriocins from four genera of lactic acid bacteria. By using this method we made preparations of pediocin AcH, nisin, sakacin A, and leuconocin Lcml that were potent and concentrated. This method produced a higher yield than isolation procedures, which rely on precipitation of the bacteriocins from the cell-free culture liquor. It is simple and can be used to produce large quantities of bacteriocins from lactic acid bacteria to be used as food biopreservatives.

Characterization of an antibacterial substance produced by Pediococcus pentosaceus Iz3.13 isolated from Japanese fermented marine food

“…The molecular mass of pediocin AcH was reported to be 4628 Da 35 Further, pediocin Iz3.13 did not adsorb to the producer cells when the pH of the medium was raised. This is contrary to the findings of Yang et al 19 who reports that pediocin AcH produced by Ped. acidilactici LB 42-923 could be completely recovered from growth media by cell adsorption and desorption.…”

“…Similarly, high recovery rates were reported by other researchers. 19 There is a need to determine whether the differences between pediocin Iz3.13 and pediocin AcH are due to differences in methods of anlaysis, genetic variability of indicator strains, or novelty of the pediocin Iz3.13. At this stage, results indicate that pediocin Iz3.13 produced by Ped.…”

“…Purification of the bacteriocin was done by a modification of the cell adsorption-desorption method. 19 Briefly, cells from 400 mL of a 22 h culture (2% peptone, 2% glucose, 2% yeast extract, pH 6.0, at 30°C) of the producing strain were harvested by centrifugation (15 000 ¥g, 2 min). Extraction of bacteriocin from the cells was done by re-suspension in 10 mL of 100 mM NaCl (pH 2.0) with stirring at room temperature for 4 h. The supernatant containing the extracted bacteriocin was then harvested and filter-sterilized using a 0.2 mm-pore-size filter (Minisart; Sartorius, Göttingen, Germany).…”

“…Suspension in 100 mM NaCl was to prevent clumping of bacteriocin molecules. 19 Throughout the experiment, the optical density (660 nm) was observed, and periodically 0.1 mL samples were serially diluted for determination of colony counts by spread plate method on TSBYE agar. For the bacteriocin-free control, an equal amount of 100 mM NaCl at pH 2.5 was added.…”

Production kinetics of acidophilin 801, a bacteriocin produced by Lactobacillus acidophilus IBB 801

“…However, constant pH values optimal for growth decrease bioavailable bacteriocin activity when cells enter the stationary phase. This loss of bacteriocin activity can be attributed to proteolytic degradation by either speci¢c or non-speci¢c proteases [7,15], a limited immunity of the producer cells if one suggests that immunity is based either on the complex formation between the mature bacteriocin molecules and the immunity peptides or on the interaction of immunity peptides and (limited) cell receptor sites [17], or to the adsorption of the inhibitory compound on the surface of the producer cells [8,18]. The latter would be the main reason for the apparent loss of bacteriocin activity [8,19].…”