融合ESI FT-ICR/MS法分析拟南芥代谢组

Reiko Motohashi; Masakazu Satou; Fumiyoshi Myouga; Akira Oikawa; Daisaku Ohta

doi:10.21769/BioProtoc.1463

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Peer-reviewed

Arabidopsis Metabolome Analysis Using Infusion ESI FT-ICR/MS

融合ESI FT-ICR/MS法分析拟南芥代谢组

Reiko Motohashi email

发布: 2015年05月05日第5卷第9期 DOI: 10.21769/BioProtoc.1463 浏览次数: 8296

评审: Arsalan DaudiSekhar KambakamAnonymous reviewer(s)

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Cover of Plant Molecular Biology, featuring study using the protocol.

Jul 2014

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Abstract

We made the method for Arabidopsis metabolome analysis based on direct-infusion Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR/MS) (IonSpec). This method was sufficiently applied to metabolic phenotyping of Arabidopsis. This method is simple in that after homogenizing samples, powdered samples are dissolved in extraction solvents (acetone and methanol) to 20% fresh weight/volume. Extracted sample solutions are dried and dissolved in 50% (v/v) acetonitrile. Mass analysis using FT-ICR/MS (IonSpec) is performed in positive and negative ionization operation modes. Mass spectra are acquired over the 100-1,000 m/z range and accumulated to improve the S/N ratio.

Keywords: Metabolome (代谢组学)

Arabidopsis (拟南芥)

Albino (白化)

ESI FT-ICR/MS (ESI FT-ICR质谱)

Materials and Reagents

3-week-old Arabidopsis plants
Liquid nitrogen
Acetone (HPLC grade) (Wako Chemicals USA, catalog number: 014-08681 )
Methanol (HPLC grade) (Wako Chemicals USA, catalog number: 134-14523 )
Nitrogen gas
EN1-16 (TAITEC, catalog number: 0076417-000 )
Formic acid (HPLC grade) (Wako Chemicals USA, catalog number: 063-04192 )
Lidocaine (anaesthetic) m/z 235.18104 (Wako Chemicals USA, catalog number: 120-02691 )
Prochloraz (agricultural chemical) m/z 376.03863 (Wako Chemicals USA, catalog number: 164-25131 )
Reserpine (alkaloid sedative drug) m/z 609.28121 (Wako Chemicals USA, catalog number: 184-00691 )
Bombesin (peptide) m/z 810.41479 (Wako Chemicals USA, catalog number: 339-40861 )
28~30% ammonia solution (Wako Chemicals USA, catalog number: 016-03146 )
Negative mode internal standards: 2.4-D (plant hormone) m/z 218.96157 (Wako Chemicals USA, catalog number: 040-18532 )
Ampicillin (antibiotic) m/z 348.10180 (Wako Chemicals USA, catalog number: 017-20531 , CHAPS 8 detergent) m/z 613.388865 (Wako Chemicals USA, catalog number: 341-04721 )
Tetra-N-acetylchitotetraose [(GluNAc)₄] m/z 829.32023 (Tokyo Chemical Industry, catalog number: T2910 )
50% (v/v) acetonitrile (HPLC grade) (Wako Chemicals USA, catalog number: 018-19853 ) [use distilled water to dilute acetonitrile to 50%(v/v)]
Acetic acid (Wako Chemicals USA, catalog number: 014-20063 )
28~30% ammonia solution (Wako Chemicals USA, catalog number: 016-03146 )
Ultrapure water (recommended but not required)

Equipment

An IonSpec Explorer FT-ICR/MS equipped with a 7-tesla actively shielded superconducting magnet (Agilent, Ionspec)
Glass vial and teflon cap (GL Biochem, catalog number: 1030-46716 )
Glass pipette
0.45 µm filters (PTFE) (ADVANTEC^®, catalog number: 13HP045AN )
Heat block

Procedure

English

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Motohashi, R., Satou, M., Myouga, F., Oikawa, A. and Ohta, D. (2015). Arabidopsis Metabolome Analysis Using Infusion ESI FT-ICR/MS. Bio-protocol 5(9): e1463. DOI: 10.21769/BioProtoc.1463.