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7 protocols using 2 phenylethyl acetate

1

Volatile Organic Compounds for Behavior Tests

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The compounds used in behaviour tests namely 3-methylbut-1-yl propionate (≥99% chemical purity), 3-methylbutan-1-ol (≥99% chemical purity), ethyl octanoate (≥99% chemical purity), 2-phenylethyl acetate (≥99% chemical purity) and 2-phenyl ethanol (≥99% chemical purity) were purchased from Sigma-Aldrich (St. Louis, MO, USA); ethyl propionate (99% chemical purity), 3-methylbut-1-yl acetate (99% chemical purity), ethyl hexanoate (99% chemical purity) were purchased from Alfa Aesar (Ward Hill, MA, USA); ethyl acetate (≥99% chemical purity), hexane as a solvent (≥99% chemical purity) was obtained from Carl Roth (Karsruhe, Germany).
D (+)-Glucose monohydrate (obtained from Merck Millipore, Darmstadt, Germany), yeast extract, peptone and agar (purchased from Liofilchem, Rosetodegli Abruzzi, Italy) were used for YPD-agar medium preparation and yeast cultivation. To avoid bacterial contamination, YPD medium was supplemented with chloramphenicol (purchased from Applichem, Darmstadt, Germany).
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2

Analysis of Volatile Organic Compounds

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The following compounds were purchased from Sigma–Aldrich (St. Louis, MO, USA): hexanal (98% chemical purity), (E)-2-hexenal (98% chemical purity), octanal (99% chemical purity), nonanal (95% chemical purity), decanal (≥98% chemical purity), (E)-2-nonenal (97% chemical purity), benzaldehyde (≥99% chemical purity), 6-methyl-5-hepten-2-one (99% chemical purity), 1-hexanol (≥99% chemical purity), (Z)-3-hexen-1-ol (98% chemical purity), 1-octen-3-ol (98% chemical purity), propyl 3-methylbutanoate (≥98% chemical purity), 3-methylbutyl propionate (≥98% chemical purity), 3-methylbutyl 3-methylbutanoate (≥98% chemical purity), hexyl acetate (99% chemical purity), ethyl octanoate (≥99% chemical purity), (Z)-3-hexenyl acetate, (Z)-3-hexenyl 3-methylbutanoate (≥97% chemical purity), α-pinene (98% chemical purity), α-phellandrene (≥95% chemical purity), limonene (≥98% chemical purity), (E)-β-ocimene (≥90% chemical purity), p-cymene (99% chemical purity), linalool (97% chemical purity), α-terpinyl acetate, 2-phenylethyl acetate (99% chemical purity), (E)-geranyl acetone (96% chemical purity), hexane (≥99% chemical purity). Ethyl hexanoate (99% chemical purity), 3-methylbutyl acetate (98% chemical purity) were purchased from Alfa Aesar (Ward Hill, MA, USA), δ-3-cerene from Carl Roth (Karlsruhe, Germany), α-farnesene from Bedoukian Research Inc. (Danbury, CT, USA).
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3

Analytical Standards for Aroma Compounds

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Dichloromethane, ethyl acetate, absolute ethanol (99%), and anhydrous sodium sulfate were purchased from VWR International (Leuven, Belgium). Standards of ethyl benzoate, methyl anthranilate, phenylacetaldehyde, 2-methylbutanal, 3-methylbutanal, heptan-2-one, heptan-2-ol, 2-methylbutyl acetate, trans-2-nonenal, methional, isobutyl acetate, benzaldehyde, myrcene, linalool, ethyl heptanoate, ethyl octanoate, 2-phenylethyl acetate, ethyl acetate, ethyl butanoate, isoamyl propionate, acetophenone, and 2-acetylthiophene were purchased from Sigma-Aldrich (Bornem, Belgium). Standard of nonan-2-one was purchased from Janssen Chimica (Geel, Belgium). Standards of dimethyltrisulfide and 1-octen-3-ol were purchased from Acros Organics (Geel, Belgium). Standard of 3-methylbutan-1-ol was purchased from Merck Millipore (Burlington, NJ, USA). Standard of acetoin was purchased from Supelco (Bellefonte, PA, USA).
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4

Measuring Fungal VOC Emission Rates

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A mixture of fungal VOCs containing fusel alcohols and acetates: 3-methyl-1-butanol (Sigma- Aldrich, 95%), 2-methyl-1-butanol (Sigma-Aldrich, 9%), 3-methyl-1-butyl acetate (Sigma-Aldrich, 95%), 2-phenylethanol Sigma-Aldrich, 99%), 2-methyl-1-butyl acetate (Sigma-Aldrich, 99%), and 2-phenylethyl acetate (Sigma-Aldrich, 98%) was prepared at a ratio of 1:1:1:1:1:1 (v/v). The actual release rates of the dispensers were determined in the laboratory under ambient conditions (25°C, 0.5 m/s airflow) using the gravimetric method (Table 1). Three replicates of each dispenser type were weighed for 10 days at regular intervals, and weight loss was recorded to the nearest 0.0001 g using a fine balance. The average release rate for each dispenser type was calculated based on the average weight loss over 24 h. Additionally, the average release rate of dispensers, under natural field conditions, was determined using the gravimetric method by periodically weighing the dispensers used in the field.
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5

Cultivation and Flavor Analysis of Aspergillus oryzae

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Soybeans, raw wheat flour and edible salt (NaCl) were purchased from a local supermarket (Zhenjiang, China, 2021). 3-Methylbutanal, ethanol, 3-methyl-1-butanol, ethyl propanoate, ethyl acetate, ethyl isovalerate, 2-phenylethanol, benzeneacetaldehyde, 3-(methylthio) propyl acetate, ethyl 3-(methylthio) propionate, methyl benzoate, (E)-β-damascenone, 2-phenylethyl acetate, 2-furanmethanol, 3-methylthio-1-propanol, 3-methylthio propanal, 2,6-dimethyl pyrazine, trimethyl pyrazine, 2-acetyl pyrroline, 4-hydroxy-2,5-dimethyl-3(2H)-furanone (HDMF) and 4-hydroxy-2(or 5)-ethyl-5(or 2)-methyl-3(2H)-furanone (HEMF) were all ordered from Sigma-Aldrich (Shanghai, China, 2021). 3-Furaldehyde, 4-ethylphenol, 4-ethylguaiacol, 4-vinylguaiacol, 3-methylbutanoic acid, acetic acid, 2-methylbutanoic acid, dimethyl trisulfide, 1-octen-3-ol and C6-C33 n-alkanes were provided by Aladdin Holdings Group (Shanghai, China, 2021). Other analytically pure reagents were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China, 2021). A. oryzae 3.042 were obtained from the Guangdong Institute of Microbiology (Guangzhou, China, 2021).
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6

Volatile Compound Extraction and Analysis

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Furfural, ethyl isovalerate, hexanol, isoamyl acetate, γ-butyrolactone, benzaldehyde, hexanoic acid, ethyl hexanoate, limonene, ethyl 2-furoate, 1-octanol, 2-nonanone, 2-nonanol, 2-phenylethanol, diethyl succinate, ethyl octanoate, octanoic acid, β-cyclocitral, 2-phenylethyl acetate, nonanoic acid, ethyl nonanoate, decanoic acid, β-damascenone, ethyl decanoate, tetradecane, isoamyl octanoate, lilial, dodecanoic acid, ethyl dodecanoate, hexadecane, tetradecanoic acid, ethyl pentadecanoate, methyl pentadecanoate and ethyl hexadecanoate were purchased from Sigma-Aldrich-Fluka (St. Louis, Missouri, USA). Aliphatic hydrocarbons (retention index mixture for GC (KI) with purity above 95%) were also supplied by Sigma-Aldrich.
A 2 cm long Divinylbenzene/Carboxen/Polydimethylsiloxane (DVB/CAR/PDMS) fibre was supplied by Supelco (Bellefonte, PA, USA). Before extraction the fibre was conditioned according to the manufacturer's recommendations.
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7

HPLC Analysis of Anthocyanins and Terpenes

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Solvents of HPLC-gradient grade and all other chemicals of analytical-reagent grade were purchased from Sigma-Aldrich (Milan, Italy). The solutions were prepared in deionized water produced by a Milli-Q system (Merck Millipore, Darmstadt, DE) . Chemical standards of delphinidin-3-O-glucoside chloride, cyanidin-3-O-glucoside chloride, petunidin chloride, peonidin-3-O-glucoside chloride, malvidin-3-O-glucoside chloride, and cyanidin chloride were supplied by Extrasynthèse (Genay, France), whereas those of (+)-catechin, linalool oxide (mixture of isomers), linalool, α-terpineol, citral, citronellol, nerol, geraniol, rose oxide (mixture of isomers), geranyl acetate, geranic acid, benzyl alcohol, 2-phenylethanol, and 2-phenylethyl acetate were purchased from Sigma-Aldrich.
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