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13 protocols using 2 ethylfuran

1

Analytical Determination of Aldehydes and Ketones

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Propanal, pentanal, hexanal, heptanal, octanal, nonanal, trans-2-butenal, trans-2-pentenal, trans-2-hexenal, trans-2-heptenal, trans-2-octenal, trans-2-nonenal, trans-2-decenal, trans-2-undecenal (≥95%), trans,trans-2,4-heptadienal, trans,trans-2,4-decadienal, 1-penten-3-ol, 2-heptanone, 2-octanone, 2-pentylfuran, 2-ethylfuran, 2-butyl-2-octenal, 2-methyl-2-pentenal, propanoic acid, caproic acid, iron(II) chloride (tetrahydrate), iron(III) nitrate (nonahydrate), and iron(II) sulphate (heptahydrate) were obtained from Merck KGaA, Darmstadt, Germany. Acetic acid, chloroform, potassium iodide and acetonitrile were purchased from Carl Roth GmbH + Co. KG, Karlsruhe, Germany. Sodium thiosulphate (0.1 N) was purchased from Bernd Kraft GmbH, Duisburg, Germany. 1,2-Dipalmitoyl-sn-glycero-3-phosphoethanolamine (PE, >96%), and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (PC) were obtained from TCI Chemicals Europe N.V., Zwijndrecht, Belgium. Paraffin oil (Pfeiffer® P3) was purchased from MasCom Technologies GmbH, Bremen, Germany. All of the chemicals were of analytical grade, if not mentioned otherwise.
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2

Aromatic Compound Quantification Protocol

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Ethyl hexanoate, ethyl decanoate, 1-octen-3-one, (E)-2-nonenal, (E)-2-octenal, hexanal, heptanal, methional, β-damascenone, ethyl 3-methylbutyrate, ethyl isobutyrate, 3-methylbutyl acetate, 3-methyl-1-butanol, 1,1-diethoxyethane, 3-methylbutanal, 2-phenylethanol, 2-phenylethyl acetate, 2-pentylfuran, and 2-ethylfuran were purchased from Merck, Ireland (Arklow, Co., Wicklow, Ireland) and ethanol absolute (Scharlab, S.L., Barcelona, Spain).
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3

Volatile Organic Compound Identification

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Identities of all VOCs considered as potential marker substances were confirmed through analysis of pure reference substances.
Acetone, 2-butanone, hexanal, nonanal, isoprene and benzene were acquired from Ionimed Analytik GmbH (Innsbruck, Austria). Butane, pentane and hexane were bought from Supelco (Bellefonte, USA). Methyl-isobutyl-ketone, 1-propanol, styrene, 3-octanone, 2-heptanone, furan, heptane, methylacetate, 2-methylfuran, 2-ethylfuran, 2-pentylfuran, 2-pentanone, 2-hexanone, 3-hexanone, 3-methyl-2-butanone, 3-methyl-2-pentanone, 2-propanethiol and dimethylsulfide were purchased from Fluka/Sigma-Aldrich (Steinheim, Germany). 2-Methyl-butanal and dimethyldisulfide were bought from Abbott GmbH & Co.KG (Wiesbaden, Germany). Ethylbenzene was acquired from Merck (Darmstadt, Germany). 3-methylfuran and 3-methylbutanal was purchased from TCI Europe N.V. (Zwijndrecht, Belgium).
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4

Furan Derivatives Procurement and Analysis

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The furan standard was procured from Alfa Aesar Co. (Ward hill, MA, USA), while 3-methylfuran was from Acros Co., (Carlsbad, CA, USA), and 2-pentylfuran from Tokyo Chemical Industry Co. (Tokyo, Japan). The other furan derivative standards, including 2-methylfuran, 2,3-dimethylfuran, 2,5-dimethylfuran, 2-ethylfuran, furfural, furfuryl alcohol, 2-butylfuran, and 2-acetylfuran, were from Sigma-Aldrich (St. Louis, MO, USA). The internal standard d4-furan was from Dr. Ehrenstorfer GmbH Co. (Augsburg, Germany). The HPLC-grade methanol was from Merck (Darmstadt, Germany), and the sodium chloride was from Uni Region Bio-Tech (Los Altos, CA, USA). The sodium chloride was purchased from UNI-ONWARD CORP (New Taipei City, Taiwan).
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5

Analytical Preparation of Organic Compounds

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Water used in the experiments was purified by a Milli-Q purification system (Millipore Sigma, Burlington, MA, USA), and solvents used were of HPLC-isocratic grade or higher. The chemicals used as standards in the experiments were purchased from several different vendors. From Acros Organics (Antwerp, Belgium), we purchased 2-hexenal. From Chem Service Inc. (West Chester, PA, USA), we purchased acetophenone, fructose, glucose, and sucrose. From Fluka (Steinheim, Switzerland), we purchased chlorogenic acid and eucalyptol. From Merck (Darmstadt, Germany), we purchased quinic acid. From Polyphenols AS (Sandnes, Norway), we purchased cyanidin-3-O-galactoside. From Sigma-Aldrich (St. Louis, MO, USA), we purchased benzaldehyde, catechin hydrate, citric acid, o-cymene, p-cymene, 2-ethyl furan, ethyl octanoate, (E,E)-2,4-heptadienal, (E,E)-2,4-hexadienal, hexanal, hexanoic acid, 1-hexanol, hexyl acetate, linalool, d-limonene, malic acid, methylbutanal, 3-methyl-1-butanol acetate, 6-methyl-5-hepten-2-one, 4-Methyl-2-pentanol, n-alkane mixture (C7–C30), neryl acetate, 1-octen-3-ol, 1-pentanol, quercetin-3-O-rutinoside, 2-α-pinene, shikimic acid, and γ-terpinene.
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6

Alkane Standard Solutions for GC-MS

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Alkane standard solutions from C8 to C20 (40 mg/L each in hexane) and C21 to C40 (40 mg/L each in toluene) were purchased from Sigma-Aldrich. A standard solution from C5 to C10 was prepared using pure substances in a ratio resulting in narrow and symmetric peak shapes as described by Weingart, et al.35 (link).
Pure VOCs were selected according to the SPME/GC-MS results, such as Benzenethanol, β-caryophyllene, trans-2-pentenal, 2-ethylfuran and β-cyclocitral (Sigma-Aldrich); cadinene (a mixture of ɣ-cadinene and δ-cadinene; (BOC Sciences); β-selinene and ledol (Xiamen Freede Industry). Pure VOCs were used in functional assays and for identity confirmation with HS-SPME/GC-MS analysis (Supplementary Fig. S2)
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7

Analytical Standards for Amino Acids, Sugars, and Furan Compounds

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Analytical standards for twenty-two amino acids (list in Table 2, norvaline and sarcosine), six sugars (D-fructose, D-glucose, sucrose, Dmaltose, β-lactose and phenyl-b-D-glucopyranoside), furan and five alkylfurans (2-methylfuran, 3-methylfuran, 2-ethylfuran, 2,5-dimethylfuran and 2,3-dimethylfuran) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Thiophene and 2-methylthiophene were bought from VWR (Radnor, PA, USA) and acrylamide from Chiron (Trondheim, Norway). For labelled standards analogous, furan and alkyls (furan-d 4 , 2-methylfuran-d 6 , 3-methylfuran-d 3 and 2,5-dimethylfuran-d 3 ) were bought at Toronto Research Chemicals (Toronto, ON, Canada), while acrylamide (acrylamide-d 3 ) comes from Chiron.
Methanol HPLC grade, boric acid, trifluoroacetic acid (TFA), hexamethyldisilazane (HMDS) and Carrez Solution II (30% ZnSO4) were
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8

Quantification of Furan Derivatives

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Furan standard was purchased from Alfa Aesar Co. (Ward hill, MA, USA), while 3-methylfuran was from Acros Co., (Carlsbad, CA, USA), and 2-pentylfuran from Tokyo Chemical Industry Co. (Tokyo, Japan). The other furan derivative standards including 2-methylfuran, 2,3-dimethylfuran, 2,5-dimethylfuran, 2-ethylfuran, furfural, furfuryl alcohol, 2-butylfuran and 2-acetylfuran were from Sigma–Aldrich (St. Louis, MO, USA). Internal standard d4-furan was from Dr. Ehrenstorfer GmbH Co. (Augsburg, Germany). The HPLC-grade solvents methanol and petroleum ether were from Merck (Darmstadt, Germany). Deionized water was produced by a Milli-Q water purification system (Bedford, MA, USA). Sodium chloride, ninhydrin, 3,5-dinitrosalicylic acid, d-dextrose, potassium sodium tartrate 4-hydrate, and glycine was purchased from Sigma–Aldrich (St. Louis, MO, USA).
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9

Quantitative Analysis of Furan Compounds

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Water for chromatography (HPLC grade), methanol gradient grade for liquid chromatography and sodium chloride were supplied by Merck (Darmstadt, Germany). Furan (chemical purity ≥ 98 %), 2methylFuran (chemical purity ≥ 99 %), 3-methylFuran (chemical purity ≥ 98 %), 2-ethylFuran (chemical purity ≥ 99 %), 2,5-dimethylFuran (chemical purity ≥ 99 %) and 2-pentylFuran (chemical purity ≥ 97 %) were purchased at Sigma Aldrich (St. Louis, USA). 2,5-DimethylFuran-d 6 (chemical purity ≥ 98 %, isotopic purity ≥ 99 %) was purchased at AlfaAesar (Haverhill, Massachusetts, USA). Furan-d 4 (chemical purity ≥ 99 %, isotopic purity ≥ 99 %) was purchased at HPC Standards GmbH (Cunnersdorf, Germany). All the other internal standards (IS) were from Toronto Research Chemicals (Toronto, Canada) with the following labelling and purity : 2-methylFuran-d 6 (chemical purity ≥ 98 %, isotopic purity ≥ 98 %), 2ethylFuran-d 5 (chemical purity ≥ 90 %, isotopic purity ≥ 94 %) and 2-pentylFuran-d 11 (chemical purity ≥ 96 %, isotopic purity ≥ 98 %).
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10

Flavor Compound Identification and Quantification

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Acetone, trimethylamine, 1-propanol, dimethyl sulfide, 2-methyl-1-propanol, 1-penten-3-ol, (z)-2-penten-1-ol, 3-pentanone, 3-methyl-butanal, 2-methyl-1-butanol, 1-pentanol, 3-pentanol, phenol, methyl-pyrazine, 2-ethyl-furan, 2-furanmethanol, propyl acetate, 1-hexanol, benzyl alcohol, 2,3-dimethyl-pyrazine, 2,5-dimethyl-pyrazine, 1heptanol, 2-ethyl-5-methyl-pyrazine, phenylethyl alcohol were purchased from Sigma-Aldrich (Shanghai, China). Mequinol, 2, 2, acetic acid, propanoic acid, acetoin, styrene, China) . The purity of these flavor standards was more than 99%. The n-alkanes (C6 to C30) were acquired from Sigma-Aldrich (Shanghai, China). The isotope internal standards (n-nonan-d20, n-dodecane-d26, n-nonadecane-d40, n-tridecane-d28, n-hexadecane-d34) were purchased from Cambridge Isotope Laboratories, Inc.
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