Levilactobacillus brevis with High Production of Putrescine Isolated from Blue Cheese and Its Application

Abstract

Polyamine intake has been reported to help extend the lifespan of animals. Fermented foods contain high concentrations of polyamines, produced by fermenting bacteria. Therefore, the bacteria, isolated from fermented foods that produce large amounts of polyamines, are potentially used as a source of polyamines for humans. In this study, the strain Levilactobacillus brevis FB215, which has the ability to accumulate approximately 200 µM of putrescine in the culture supernatant, was isolated from fermented foods, specifically the Blue Stilton cheese. Furthermore, L. brevis FB215 synthesized putrescine from agmatine and ornithine, which are known polyamine precursors. When cultured in the extract of Sakekasu, a byproduct obtained during the brewing of Japanese rice wine containing high levels of both agmatine and ornithine, L. brevis FB215 grew to OD₆₀₀ = 1.7 after 83 h of cultivation and accumulated high concentrations (~1 mM) of putrescine in the culture supernatant. The fermentation product also did not contain histamine or tyramine. The Sakekasu-derived ingredient fermented by the food-derived lactic acid bacteria developed in this study could contribute to increasing polyamine intake in humans.

Keywords: Levilactobacillus brevis; blue cheese; food; polyamine; putrescine.

Figure 1

Putrescine concentration of culture supernatants of bacteria isolated from blue cheese. (A) The putrescine concentration of the culture supernatants of the isolated 38 strains was measured using a simplified quantitative method (PuO-POD-4AA-TOPS method described previously [32]); (B) The culture supernatant of the 14 strains that contained more than 100 µM putrescine in the culture supernatant was quantified by HPLC.

Figure 2

The previously reported putrescine biosynthetic pathways (3157043).

Figure 3

Production of putrescine when Levilactobacillus brevis FB215 is grown in MRS broth supplemented with putrescine precursors. L. brevis FB215 was inoculated in 50 mL of MRS broth supplemented with ornithine or agmatine at a final concentration of 1 mM and incubated under anaerobic conditions for 48 h at 37 °C. At the end of the cultivation period, the polyamine concentration in the culture supernatant was determined using HPLC (n = 3). Two-tailed t-tests were performed.

Figure 4

Putrescine concentration of the culture supernatant of Levilactobacillus brevis FB215 grown in Sakekasu extract. L. brevis FB215, S. epidermidis FB146 [34], and L. curvatus KP3-4 [23] were inoculated in Sakekasu extract and incubated at 37 °C. Polyamine concentrations in culture supernatants at different growth phases were determined. The levels of putrescine in the culture supernatants of L. brevis FB215, S. epidermidis FB146, and L. curvatus KP3-4 are represented by red squares, blue triangles, and green circles, respectively (n = 3).