SE198:/S93/M011/D1

Sample Set Information

ID	SE198
Title	Untargeted metabolome analysis of 15N- or 34S-labeled plants (lettuce) / 15Nまたは34S全標識した植物のLC-MSメタボローム解析（レタス）
Description	Untargeted metabolome analyses of lettuce were performed using liquid chromatography – mass spectrometry (LC-MS). This data acquisition aims at an improvement of peak annotation by using plant samples that were fully labeled with stable isotope 15N or 34S. Two lettuce varieties, Leaf Lettuce Green and King Crown were used. The information of the peaks detected in Leaf Lettuce Green was available at the Plant Metabolome Repository website (http://metabolites.in/plants). 15Nおよび34Sで全標識した植物試料を用いることで、液体クロマトグラフィー-質量分析（LC）で検出される化合物ピークの推定を向上させることを目的として、ノンターゲットメタボローム解析を行った。 植物試料として、二種類のレタス（リーフレタスグリーンおよびキングクラウン）を用いた。 リーフレタスグリーンで検出されたピークは、植物メタボロームレポジトリ (http://metabolites.in/plants) から公開されている。
Authors	Sakurai N1,2, Suda K1, Akimoto N1, Hoshi K1, Osawa S1, Ikeda C1, Ozawa K1, Yamada M1, Muneto R1, Shibata D1 (1 Kazusa DNA Research Institute, 2 National Institute of Genetics), Contact: sakurai AT nig.ac.jp (replace AT with @) 櫻井望1,2、須田邦裕1、秋元奈弓1、星久美1、大澤祥子1、池田千晶1、小澤馨史1、山田学1、宗藤玲子1、柴田大輔1（1 かずさDNA研究所, 2 国立遺伝学研究所）
Reference
Comment

Sample Information

ID	S93
Title	Mock (without plant sample)
Organism - Scientific Name
Organism - ID
Compound - ID
Compound - Source
Preparation
Sample Preparation Details ID
Comment

Analytical Method Information

ID	M011
Title	ESI Positive (Method 1)
Method Details ID	MS01
Sample Amount	20 uL injection
Comment

Analytical Method Details Information

ID	MS01
Title	LC-FT-MS, ESI, Positive
Instrument	Agilent1100 HPLC (Agilent), LTQ-FT (Thermo Fisher Scientific)
Instrument Type	LC-FTICR-MS
Ionization	ESI
Ion Mode	Positive
Description	Metabolite extraction 300 mg of powdered sample was mixed with 900 uL of 100% methanol solution, and 18 mg freeze-dried sample was mixed with 1200 uL of 75% methanol solution in 2 mL tube. Both 100% and 75% methanol solution contain 25 uM of 7-hydroxy-5-methylflavone as internal standard (IS). The sample was homogenized using Mixer Mill MM 300 (QIAGEN) at 25 Hz, for 2 min, twice. The homogenate was centrifuged under 17,400 x g, 5 min at 4 C. A supernatant was passed through a Polytetrafluoroethylene (PTFE) filter (pore size 0.2 um, Millipore), and the filtrate was applied to a C18 silica column (MonoSpin C18, GL Science) to remove highly-hydrophobic contaminants. The filtrate passed through the C18 column was used for LC-MS analyzes. The same extraction procedure with 75% methanol was performed without a sample to prepare mock samples and used as negative controls. Liquid chromatography (LC)-mass spectrometry (MS) analysis Agilent 1100 system (Agilent) and Finnigan LTQ-FT (Thermo Fisher Scientific) were used. Aliquots (5 uL) of the methanol extract were applied to a TSK-gel ODS-100V column (4.6 x 250 mm, 5 um, TOSO), and separated by water containing 0.1%(v/v) formic acid (Solvent A) and acetonitrile containing 0.1%(v/v) formic acid (Solvent B). The gradient program was as follows: 3% B (0 min), 97% B (90 min), 97% B (100 min), 3% B (100.1 min), and 3% B (107 min). The flow rate was set at 0.25 mL/min for 0-100 min, and 0.5 mL/min for 100.1-107 min. The conditions of mass spectrometry were as below: - Method 1: This method is used for measuring the accurate mass of the peaks and obtaining MS/MS spectra. [Full scan] resolution, 100,000 by FT; m/z range, 100-1500 [MS2 scan] data-dependent scan by IT for the most intense 5 ions in the full scan. [Dynamic exclusion setting for MS2 scan] repeat count, 3; repeat duration, 30 s; exclusion list size, 500; margin, 10 ppm; exclusion duration, 20 s. - Method 2: Same as Method 1 except below: [Dynamic exclusion setting for MS2 scan] exclusion duration, 30 s. - Method 3: Same as Method 1 except below: [Dynamic exclusion setting for MS2 scan] repeat count, 2; exclusion duration, 30 s. - Method 4: Same as Method 1 except below: [Dynamic exclusion setting for MS2 scan] off. - Method 5: This method is used for obtaining MS3 spectra. [Full scan] resolution, 12,500 by FT; m/z range, 100-1500. [MS2 scan] data-dependent scan by IT for the most intense 5 ions in the full scan. [Dynamic exclusion setting for MS2 scan] same as Method 1. [MS3 scan] data-dependent scan by IT for the most intense 2 ions in the MS2 scan - Method 6: This method is used for obtaining MS2 spectra in high-resolution: [Full scan] resolution, 12,500 by FT; m/z range, 100-1500. [MS2 scan] data-dependent scan by FT for the most intense 5 ions in the full scan. resolution, 12,500. [Dynamic exclusion setting for MS2 scan] repeat count, 1; repeat duration, 30 s; exclusion list size, 500; margin, 10 ppm; exclusion duration, 40 s. - Method 7: Same as Method 5 except below: [Dynamic exclusion setting for MS2 scan] same as Method 2 ESI: electrospray ionization, FT: Fourier transformation ion cyclotron resonance type mass spectrometry, IT: ion trap type mass spectrometry. The mass analyses with electrospray ionization (ESI) in positive or negative modes were performed. The combinations of the Methods 1-7 and ionization polarity are different between the samples, but, at least one method for accurate mass measurement (Method 1-4) was applied for each sample. The raw data were acquired using Xcalibur software (Thermo Fisher Scientific).
Comment_of_details	[column] TSK-gel ODS-100V (4.6 x 250 mm, 5 micrometer; TOSOH) [gradient] Solvent A: water containing 0.1% v/v formic acid; Solvent B: acetonitrile containing 0.1% v/v formic acid; Gradient: 3% B (0 min), 97% B (90 min), 97% B (100 min), 3% B (100.1 min), and 3% B (107 min) [total separation time] 107 min

Data Analysis Information

ID	D1
Title	Peak detection (Method 1)
Data Analysis Details ID	DS1
Recommended decimal places of m/z	default
Comment

Data Analysis Details Information

ID	DS1
Title	Peak detection using PowerGetBatch
Description	The PowerGetBach software (version 0.5.4, http://www.kazusa.or.jp/komics/software/PowerGetBatch) was used for peak detection. In the case of the sample with more than three biological or technical replications, a single parameter setting for high resolution (Method 1-4) was applied for peak detection. In the case of the sample without replications, a single raw datum was processed using three different parameter settings for high resolution (Method 1-4). The data obtained in low-resolution settings (Method 5 and 7) was processed with a single parameter setting. The detailed peak detection parameters for PowerGetBatch are available at the Plant Metabolome Repository website (http://metabolites.in/plants/data/PGB_params.zip).
Comment_of_details