Methionol

Commercially available food additive products were used in the human sensory test. Food-grade NaCl was purchased from Naikaisyoji (Tokyo, Japan), and monosodium glutamate (MSG) was purchased from Ajinomoto Co., Inc. (Tokyo, Japan). All flavor compounds (of food additive grade), including propionaldehyde, butanal, isobutylaldehyde, 2-methylbutylaldehyde, pentanal, IVA, hexanal, heptanal, octanal, propionic acid, 2-methylbutyric acid, isovaleric acid, hexanoic acid, heptanoic acid, propyl alcohol, 2-methyl-1-butanol, hexanol, heptanol, methional, methionol, and 3-(methiylthiol) propionic acid, were purchased from Sigma-Aldrich (St. Louis. MO, USA). Deionized water for test solutions and mouth rinsing was supplied by a Mill-Q water purification system (Millipore, Bedford, MA, USA). γ-Glu-Val-Gly was obtained from Ajinomoto Co., Inc. (Tokyo, Japan). NPS-2143, a CaSR inhibitor, was synthesized, as described by Rybczynska et al. [41 (link)]. Chemicals for cell based-assays, such as CaCl₂, ethylenediaminetetraacetic acid (EDTA), and probenecid, were purchased from Sigma-Aldrich (St. Louis. MO, USA).

All chemicals, unless otherwise indicated, were purchased from Sigma-Aldrich (Germany). Analytical-grade anhydrous ethanol was purchased from Ajax Finechem (Taren Point, NSW, Australia), and D-glucose was purchased from Merck (Kenilworth, NJ, USA). The internal standard dimethyl-d₆ sulfide (d₆-DMS), along with methionol, methanethiol, and dimethyl disulfide, were purchased from Sigma-Aldrich (Darmstadt, Germany). Liquid YPD or YPD agar plates were used for routine maintenance of standard yeast cultures, while chemically defined SGM mimicking grape juice (21°Brix, pH 3.2, yeast assimilable nitrogen 300 mg L⁻¹) was used for fermentative growth, as described in previous reports (Kinzurik et al. 2015 (link), Deed et al. 2019 (link)). To cater for the methionine auxotrophy of the BY4741 strain, 10-fold concentrations of L-methionine were supplemented in the final SGM (0.3 mM). (E)-hex-2-enal (Sigma-Aldrich, Darmstadt, Germany) stock solution was freshly prepared and aliquoted by dissolving in anhydrous ethanol at 1.5 mg mL⁻¹ and supplemented into the medium preparation at a final concentration of 1.5 mg L⁻¹ just before yeast seeding. YNB medium (yeast nitrogen base without amino acids or ammonium sulfate 1.7 g L⁻¹, ammonium chloride 0.3 g L⁻¹, L-malic acid 3 g L⁻¹, citric acid 0.2 g L⁻¹, sucrose 200 g L⁻¹, pH = 3.2) was used for early precursor kinetics.

The following chemical standards were used in the aroma base and were obtained from Sigma-Aldrich Co. (St. Louis, MO, USA): ethanol (≥99%), 2,3-butanedione (97%), hexan-1-ol (98%), methionol (≥98%), butanoic acid (≥99%), decanoic acid (≥98%), 2-methylpropanoic acid (99%), 2-methylbutanoic acid (98%), 3-methylbutanoic acid (99%), 2-methylpropyl ethanoate (≥97%), ethyl phenylacetate (≥99%), ethyl 3-methylbutanoate (98%), ethyl 2-methylpropanoate ((≥98%), ethyl 2-methylbutyrate (99%), ethyl butanoate (≥98%), ethyl decanoate (≥99%), ethyl hexanoate (≥99%), ethyl octanoate (≥98%), hexanoic acid (99%), octanoic acid (≥99%), 2-methylpropan-1-ol (≥99%), ethyl acetate (≥99%) and isoamyl acetate (≥99%). Acetic acid (≥99%) was obtained from VWR International (Radnor, PA, USA). The terpenes compound added to the model wine for the treatments were: (S)-(−)-limonene (96%), (R)-(+)-limonene (97%), linalool (≥97%) and α-terpineol, which were purchased from Sigma-Aldrich Co. (St. Louis, MO, USA).