Mousy Off-Flavor of Wine:  Precursors and Biosynthesis of the Causative N-Heterocycles 2-Ethyltetrahydropyridine, 2-Acetyltetrahydropyridine, and 2-Acetyl-1-pyrroline by Lactobacillus hilgardii DSM 20176

Abstract

The N-heterocyclic bases, 2-ethyltetrahydropyridine (1), 2-acetyl-1-pyrroline (2), and 2-acetyltetrahydropyridine (3) are associated with the occurrence of mousy off-flavor in wine. The biosynthesis of these N-heterocycles by the wine lactic acid bacterium, Lactobacillus hilgardii DSM 20176, was studied by high-cell-density incubation in combination with a minimal chemically defined N-heterocycle assay medium. The key components of the defined N-heterocycle assay medium included D-fructose, ethanol, L-lysine, L-ornithine, and mineral salts. N-heterocycle formation was quantitatively determined by gas chromatography-mass spectrometry. The formation of 2 and 3 required the concomitant availability of a fermentable carbohydrate (D-fructose), ethanol, and iron (Fe(2+)). In addition, L-ornithine stimulated the formation of 2 and repressed 3 formation, whereas L-lysine stimulated the formation of 3 and repressed 2 formation. Incorporation of d(6)-ethanol into the acetyl side chain of 2 and 3, and of d(4)-acetaldehyde into the acetyl side chain of 3, confirmed that ethanol and acetaldehyde could serve as major side chain precursors. A pathway for the formation of 2 and 3 by heterofermentative lactic acid bacteria is proposed involving the interaction of accumulated C-2 intermediates from the heterolactic pathway and N-heterocyclic intermediates derived from the metabolism of L-ornithine and L-lysine.

Malolactic Fermentation: The ABC’s of MLF

“…The availability of the precursor's lysine and ornithine has a significant impact on the ability of LAB to produce these compounds (Costello & Henschke, 2002). Costello & Henschke (2002) also found that the lack of ethanol drastically reduces the ability of L. hilgardii to produce Nheterocycles, whereas elevated concentrations of acetaldehyde sti mulates formation. Lactic acid bacteria differ in their preference for the formation of the different nitrogen heterocyclic compounds.…”

Lactobacillus: the Next Generation of Malolactic Fermentation Starter Cultures—an Overview

“…The odour thresholds of ACPY and ACTPY have been determined in water as 0.1 and 1.6 μg/L, respectively. Wines that are found to be spoiled by the mousy taint have been found to contain one or more of the three main compounds: ETPY levels of 2.7-18.7 μg/L, ACPY concentrations up to 7.8 μg/L and ACTPY at concentrations of 4.8-106 μg/L (Costello and Henschke 2002). Costello and Henschke (2002) investigated the biosynthesis of ETPY, ACPY and ACTPY by L. hilgardii DSM 20176.…”

“…It was found that ornithine stimulates the formation of ACPY while lysine stimulates the formation ACTPY. Conversely, ornithine represses the formation of ACTPY while lysine represses the formation of ACPY (Costello and Henschke 2002). Costello et al (2001) found that LAB differ in their ability to produce these compounds.…”

Mechanism of 2-acetyl-1-pyrroline biosynthesis in Bassia latifolia Roxb. flowers

“…Trace experiments with 15 N-proline, 15 N-glycine, and proline-1-13 C indicated that the nitrogen source of 2AP was proline, whereas the carbon source of the acetyl group was not its carboxyl group (Yoshihashi et al 2002). Costello and Henschke (2002) suggested role of ornithine as nitrogen source of 2AP via Putrescine pathway. Intermediate 1-pyrroline could then be subjected to acylation of C-2 position of acyl-CoA derivatives, thus leading to synthesis of 2AP.…”