SE231:/S081/M01

Sample Set Information

ID	S231
Title	LC-MS based untargeted metabolome analysis of soybean
Description	Untargeted metabolome analysis in leaves of 79 accessions of soybean provided by the World Soybean Core Collection (The Research Center of Genetic Resources, National Agriculture and Food Research Organization, NARO) was conducted using liquid chromatography-mass spectrometry (LC-MS).
Authors	Nozomu Sakurai (National Institute of Genetics, Kazusa DNA Research Institute, Sakura Scientific, email: sakurai (at) kazusa.or.jp)
Reference
Comment

Sample Information

ID	S081
Title	Soybean (GmWMC113, BARITOU 3 A, JP 43384) / Leaf / Indonesia (Landrace) / ダイズ (GmWMC113, BARITOU 3 A [バリ島３Ａ], JP 43384) / 葉 / インドネシア (在来)
Organism - Scientific Name	Glycine max
Organism - ID	NCBI taxonomy: 3847
Compound - ID
Compound - Source
Preparation	The sample was harvested outdoors (July or August, 2021, Japan).
Sample Preparation Details ID	SS1
Comment

Sample Preparation Details Information

ID	SS1
Title	Homogenated and stored at -80 C
Description	After harvesting, the leaves were dried using a ventilation dryer (TTM-435S, TOHMEI Tech Co., Ltd., Osaka, Japan) at 60 ºC for 4 hours. The dried leaves were ground into a fine powder using a mortar and pestle at room temperature, weighed, and stored at 4C until use.
Comment_of_details

Analytical Method Information

ID	M01
Title	LC-QTOF-MS, ESI, Positive
Method Details ID	MS01
Sample Amount	0.5 mg dry weight/ 2 ul injection
Comment

Analytical Method Details Information

ID	MS01
Title	LC-Q-Tof-MS, ESI, Positive
Instrument	Nexera X2 (Shimadzu), Compact (Bruker Daltonics)
Instrument Type	LC-QTOF-MS
Ionization	ESI
Ion Mode	Positive
Description	- Compound Extraction Approximately 10 mg of dried fine powder of soybean leaves was accurately weighed in a 2 mL tube, and a 40 times volume of 75% methanol, containing 1 μM of 7-hydroxy-5-methylflavone as an internal standard, was added. Then, the metabolite extraction and the metabolome analysis were performed according to previously described procedures using LC (Nexera X2, Shimadzu), time-of-flight MS (Compact, Bruker), and C18 column (InertSustain AQ-C18, 3 μm x 2.1 mm x 150 mm, GL Sciences) in both ESI positive and negative modes (Sakurai et al., Nucleic Acids Res 51(D1): D660-677, 2023). - Liquid chromatography (LC)-mass spectrometry (MS) analysis LC-MS anaysis was conducted as previously described (Sakurai et al., Nucleic Acids Res 51(D1): D660-677, 2023) using Nexera X2 system (Shimadzu Corporation) and Compact system (Bruker Japan K.K.). An aliquot (2 uL) of the methanol extract was applied to an InertSustain AQ-C18 column (3 um x 2.1 mm x 150 mm, GL Sciences) connected after a guard column (InertSustain AQ-C18 Cartridge Guard Column E, GL Sciences), and separated by water containing 0.1%(v/v) formic acid (Solvent A) and acetonitrile (Solvent B). The gradient program was as follows: 2% B (0 min), 2% B (3 min), 98% B (30 min), 98% B (35 min), 2% B (35.01 min), and 2% B (42 min). The flow rate was set at 0.2 mL/min. The column oven temperature was set to 40 degree C. The compounds separated by the LC were detected using the mass spectrometer under the conditions below: Ionization, Electrospray Ionization (ESI); Polarity, Positive; Scan rate, 1 Hz; Mass scan range, 50-1200; End plate offset, 500 V; Capillary voltage, 4000 V; Nebulizer gas (N2) pressure, 2.5 bar; Dry gas (N2) flow, 8.0 L/min; Dry gas temperature, 200 degree C; Transfer Funnel1 RF, 200.0 Vpp; Funnel2 RF, 200.0 Vpp; In-source CID Energy, 0.0 eV; Hexapole RF, 50.0 Vpp; Quadrupole Ion Energy, 3.0 eV; Low mass m/z, 55.00; Collision energy, 10.0 eV; Collision RF, 450.0 Vpp; Transfer time, 80.0 us; and PrePulse storage, 3.0 us. The data-dependent MS/MS spectra were obtained with the conditions below: Isolation width, 3-15 Da; Collision energy 35 eV; Precursor ion number, 5; Active Exclusion, on; Exclude, after 3 spectra; Release, after 0.2 min; Reconsider precursor, on; and if current intens. / previous intens., 2.0. For mass calibration, 1 mM sodium formate in 50% (v/v) 2-propanol was injected directly into the MS at 38.50–40.50 min of LC separation with a flow rate 0.1 mL/min. The eluent at 0-3 min was wasted. The raw data were obtained by Hystar software (ver.3.2 SR4, Bruker Daltonik, GmbH).
Comment_of_details	[column] InertSustain AQ-C18 (2.1 x 150 mm, 3 micrometer; GL Sciences) [gradient] Solvent A: water containing 0.1% v/v formic acid; Solvent B: acetonitrile; Gradient 2% B (0 min), 2% B (3 min), 98% B (30 min), 98% B (35 min), 2% B (35.01 min), and 2% B (42 min) [total separation time] 42 min