SE189:/MS1

Sample Set Information

ID	SE189
Title	Clarification of pathway-specific inhibition by Fourier transform ion cyclotron resonance/mass spectrometry-based metabolic phenotyping studies.
Description	We have developed a metabolic profiling scheme based on direct-infusion Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR/MS). The scheme consists of: (1) reproducible data collection under optimized FT-ICR/MS analytical conditions; (2) automatic mass-error correction and multivariate analyses for metabolome characterization using a newly developed metabolomics tool (DMASS software); (3) identification of marker metabolite candidates by searching a species-metabolite relationship database, KNApSAcK; and (4) structural analyses by an MS/MS method. The scheme was applied to metabolic phenotyping of Arabidopsis (Arabidopsis thaliana) seedlings treated with different herbicidal chemical classes for pathway-specific inhibitions. Arabidopsis extracts were directly infused into an electrospray ionization source on an FT-ICR/MS system. Acquired metabolomics data were comprised of mass-to-charge ratio values with ion intensity information subjected to principal component analysis, and metabolic phenotypes from the herbicide treatments were clearly differentiated from those of the herbicide-free treatment. From each herbicide treatment, candidate metabolites representing such metabolic phenotypes were found through the KNApSAcK database search. The database search and MS/MS analyses suggested dose-dependent accumulation patterns of specific metabolites including several flavonoid glycosides. The metabolic phenotyping scheme on the basis of FT-ICR/MS coupled with the DMASS program is discussed as a general tool for high throughput metabolic phenotyping studies.
Authors	Oikawa, A., Nakamura, Y., Ogura, T., Kimura, A., Suzuki, H., Sakurai, N., Shinbo, Y., Shibata, D., Kanaya, S. and Ohta, D.
Reference	Plant Physiol. 2006 Oct;142(2):398-413
Comment

PubMed

Analytical Method Details Information

ID	MS1
Title	FT-ICR/MS
Instrument	IonSpec Explorer FT-ICR/MS (IonSpec)
Instrument Type
Ionization	ESI
Ion Mode	Positive and Negative
Description	Sample Preparation Thirty 1-week-old plants were frozen in liquid N2 and ground to powder, which was used for methanol extraction. The extracts were filtered through disposable membrane filter units (DISMIC-13JP, ADVANTEC), evaporated under N2 atmosphere, and stored at −80°C until use. Upon FT-ICR/MS analysis, the extracts were dissolved in 50% (v/v) acetonitrile/water. IMCs for the positive ion mode were lidocaine ([M + H]+ = 235.18049), prochloraz ([M + H]+ = 376.03809), reserpine ([M + H]+ = 609.28066), and bombesin ([M + 2H]2+ = 810.41481). A set of 2,4-dichlorophenoxy acetic acid ([M−H]− = 218.96212), ampicillin ([M−H]− = 348.10235), 3-[(3-cholamidopropyl) dimethylammonio] propanesulfonic acid ([M−H]− = 613.38920), and tetra-N-acetylchitotetraose ([M−H]− = 829.32078) were used as the IMCs in the negative ion mode analysis. FT-ICR/MS Conditions Mass analysis was done using an IonSpec Explorer FT-ICR/MS (IonSpec) equipped with a 7-tesla actively shielded superconducting magnet. Ions were generated from an ESI source with a fused silica needle of 0.005-inch i.d. Samples were infused using a Harvard syringe pump model 22 at a flow rate of 0.5 to 1.0 μL min−1 through a 100-μL Hamilton syringe. All the experimental events were controlled using Omega8 software (IonSpec). Briefly, the potentials on the electrospray emitters were set to 3.0 kV and −3.0 kV for the positive and the negative electrosprays, respectively. The base pressure in the source region was approximately 5 × 10−5 torr (1 torr = 133.3 Pa). For the positive and negative electrosprays, sample solutions were prepared in 50% (v/v) acetonitrile/water with 0.1% (v/v) of formic acid and ammonium hydroxide, respectively. Ionized metabolites were accumulated for a period of 2,500 to 5,000 ms in a hexapole ion trap/guide and transferred through a radiofrequency-only quadrupole into the FT-ICR cell in the superconducting magnetic field, where they were again trapped. The direct current potentials in the positive and negative ion mode analyses were −2 V and 2 V during the ion accumulation and 2 V and −2 V for the ion transfer into the FT-ICR cell, respectively. These ions trapped in the hexapole were extracted for the transfer into the FT-ICR cell. In the positive and negative ion modes, the potentials on the extraction plate were 12 V and −12 V during the ion trapping and were reversed to −2 V and 2 V for the extraction. The base pressure in the analyzer region was set to the level of approximately 4 × 10−10 torr. ESI-MS spectra were acquired over the m/z range 55 to 1,000 from 1,024,000 independent data points. MS/MS analyses were done using the sustained off-resonance irradiation SORI-CID methods (Gauthier et al., 1991; Laskin and Futrell, 2005). SORI Rf was set at 0.5 to 1.5 V, and the N2 collision gas was used with 400-ms pulse.
Comment_of_details