From Metabolonote
jump-to-nav Jump to: navigation, search

Sample Set Information

Title Integrated strategy for unknown EI-MS identification using quality control calibration curve, multivariate analysis, EI-MS spectral database, and retention index prediction
Description Compound identification using unknown electron ionization (EI) mass spectra in gas chromatography coupled with mass spectrometry (GC-MS) is challenging in untargeted metabolomics, natural product chemistry, or exposome research. While the total count of EI-MS records included in publicly or commercially available databases is over 900 000, efficient use of this huge database has not been achieved in metabolomics. Therefore, we proposed a "four-step" strategy for the identification of biologically significant metabolites using an integrated cheminformatics approach: (i) quality control calibration curve to reduce background noise, (ii) variable selection by hypothesis testing in principal component analysis for the efficient selection of target peaks, (iii) searching the EI-MS spectral database, and (iv) retention index (RI) filtering in combination with RI predictions. In this study, the new MS-FINDER spectral search engine was developed and utilized for searching EI-MS databases using mass spectral similarity with the evaluation of false discovery rate. Moreover, in silico derivatization software, MetaboloDerivatizer, was developed to calculate the chemical properties of derivative compounds, and all retention indexes in EI-MS databases were predicted using a simple mathematical model. The strategy was showcased in the identification of three novel metabolites (butane-1,2,3-triol, 3-deoxyglucosone, and palatinitol) in Chinese medicine Senkyu for quality assessment, as validated using authentic standard compounds. All tools and curated public EI-MS databases are freely available in the 'Computational MS-based metabolomics' section of the RIKEN PRIMe Web site ( ).
Authors Teruko Matsuo, Hiroshi Tsugawa, Hiromi Miyagawa, Eiichiro Fukusaki
Reference Matsuo et al. Analytical Chemistry (2017) 89(12):6766–6773

Link icon article.png

Link icon database.png Link icon dropmet.png

The raw data files are available at DROP Met web site in PRIMe database of RIKEN.

Sample Information

Title Senkyu
Organism - Scientific Name Cnidium officinate Makino or Ligusticum chuanxiong Hort
Organism - ID Cnidium officianate:NCBI taxonomy 54711  Ligusticum chuanxiong:NCBI taxonomy 49555
Compound - ID
Compound - Source
Preparation Six types of Chinese medicine Senkyu were supplied by Tochimoto Tenkaido Co. Ltd. (Osaka, Japan), and their origins were defined by species (Cnidium officinate Makino or Ligusticum chuanxiong Hort.), cultivation area (Japan or China), and manufacturing process (with or without steaming), as summarized in Supporting Information Table S1.
Sample Preparation Details ID

Table S1. Details of Chinese medicine Senkyu used in this research
Species Cultivation
Process Abbreviation
Cnidium officinale Makino Japan Steaming and dry CJSD
Cnidium officinale Makino Japan Dry CJD
Cnidium officinale Makino China Steaming and dry CCSD
Cnidium officinale Makino China Dry CCD
Ligusticum chuanxiong Hort China Steaming and dry LCSD
Ligusticum chuanxiong Hort China Dry LCD

Analytical Method Information

Title EI and GC-MS (Quadrupole)
Method Details ID MS1
Sample Amount 1 μL

Analytical Method Details Information

Title EI and GC-MS (Quadrupole)
Instrument GCMS-QP2010 Ultra, Shimazu
Instrument Type
Ionization EI
Ion Mode Positive
Description Extraction from sample

A 10 mg sample of each dried root was weighed and homogenized in a 2-mL Eppendorf tube and 1,000 μL of a single-phase mixture of methanol/water/chloroform (5:2:2, v/v/v) containing an internal standard (ribitol, 0.4 mg/mL H2O) was added for metabolite extraction.The extraction was carried out at room temperature using mixer mills (MM 301, Verder Scientific Co. Ltd., Haan, German) at 20 Hz for 5 min.After centrifugation at 16000 ×g for 3 min at 4°C, 800 μL of the supernatant was transferred to a new 1.5-mL Eppendorf tube, to which 400 μL of distilled water was added and vortexed for 10 s. After centrifugation at 16000 ×g for 3 min at 4 °C, 400 μL of the upper layer containing hydrophilic metabolites was transferred to a new 1.5-mL Eppendorf tube, and then freeze-dried (Taitec model VD-800F freeze dryer, Taitec Co. Ltd., Saitama, Japan). For quality control samples, 10 mg of each sample was added into a 15-mL tube (total 360 mg; six biological samples and six biological replicates) and then divided into six 2-mL Eppendorf tubes such that they ontained 10 mg, 20 mg, 30 mg, 40 mg, 50 mg, and 60 mg, respectively. An empty tube was also prepared, labeled as 0 mg QC, for monitoring background effects. Metabolite extraction of these dilution series was performed sing the procedure described above.
For oximation, 50 μL of methoxyamine hydrochloride in pyridine (20 mg/mL) was added and incubated at 30 °C for 90 min. For silylation, 50 μL of N-methyl-N-(trimethylsilyl)trifluoroacetamide (MSTFA) was added and ncubated at 37 °C for 30 min. The mixture was centrifuged at 16000 ×g for 5 min at 20 °C and a 50-μL aliquot of the resultant supernatant was transferred into GC–MS vial. Each sample (1 μL) was injected into a GCMS-QP2010 Ultra instrument (Shimadzu Co., Kyoto, Japan) in split mode (25/1, v/v). Each diluted QC sample was analyzed four times from the same GC–MS vial. Analytical conditions were optimized as described in a previous study.14 The column was a fused silica capillary column (30 m × 0.25 mm i.d.) coated with 0.25 μm CP-SIL 8 CB low bleed/MS (Agilent Technologies Japan, Ltd., California, USA). The front inlet temperaturewas 230 °C, the helium gas flow rate was 1 mL/min, and the column temperature was held at 80 °C for 2 min isothermally, then raised by 15 °C/min to 330 °C, and finally held for 6 min isothermally. The transfer line and ion source temperatures were 250 °C and 200 °C, respectively. Twenty scans per second were recorded over the mass range 85–500 m/z.

Personal tools
View and Edit Metadata