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MB Sample ID: SA211192

Local Sample ID:Col-0_W+N+_P3ID33_c
Subject ID:SU002287
Subject Type:Plant
Subject Species:Arabidopsis thaliana
Taxonomy ID:3702

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Combined analysis:

Analysis ID AN003604
Analysis type MS
Chromatography type GC
Chromatography system Agilent 7890B
Column Restek Rxi-5Sil (30m x 0.25mm,0.25m) with 10m precolumn
MS Type EI
MS instrument type Single quadrupole
MS instrument name Agilent 5977B
Ion Mode POSITIVE
Units µg/mg FW and arbitrary/mg FW

MS:

MS ID:MS003359
Analysis ID:AN003604
Instrument Name:Agilent 5977B
Instrument Type:Single quadrupole
MS Type:EI
MS Comments:The instrument was an Agilent 7890B gas chromatograph coupled to an Agilent 5977B mass spectrometer. The column was an Rxi-5SilMS from Restek (30 m with 10 m integraguard column). The liner (Restek # 20994) was changed before the analysis. Oven temperature ramp was 70 °C for 7 min then 10 °C/min to 330 °C for 5 min (run length 38 min). Helium constant flow was 0.7 mL/min. Temperatures were the following: injector: 250°C, transfer line: 290°C, source: 250 °C and quadripole 150 °C. 5 scans per second were acquired spanning a 50 to 600 Da range. Instrument was tuned with PFTBA with the 69 m/z and 219 m/z of equal intensities. 5 scans per second were acquired. The split mode conditions were: 70°C for 2 min then 30°C per min to 330 °C for 5 min. Helium constant flow 1 mL/min.Data processing: Raw Agilent datafiles were converted in NetCDF format and analyzed with AMDIS http://chemdata.nist.gov/mass-spc/amdis/. An home retention indices/ mass spectra library built from the NIST, Golm , http://gmd.mpimp-golm.mpg.de/ and Fiehn databases and standard compounds was used for metabolites identification. Peak areas were also determined with the Targetlynx software (Waters) after conversion of the NetCDF file in masslynx format. AMDIS, Target Lynx in splitless and split 30 modes data were compiled in one single Excel File for comparison. After blank mean substraction peak areas were normalized to Ribitol and Fresh Weight. Statistical analysis was made with TMEV http://www.tm4.org/mev.html : univariate analysis by permutation (1way-anova and 2-way anova) were firstly used to select the significant metabolites (P-value < 0.01). Multivariate analysis (hierarchical clustering an principal component analysis) were then made on them. Mapman http://www.gabipd.org/projects/MapMan/ was used for graphical representation of the metabolic changes after Log2 transformation of the mean of the 3 replicates. Absolute quantification: A response coefficient was determined for 4 ng each of a set of 103 métabolites, respectively to the same amount of ribitol. This factor was used to give an estimation of the absolute concentration of the metabolite in what we may call a “one point calibration”. Metabolites rich in nitrogen (basic aminoacids and polyamines) gave several analytes (up to 5 for glutamine and asparagine). The peak area as TIC equivalent of these analytes were summed to express the contents of these metabolites. They are referred to “sum” in the tables.
Ion Mode:POSITIVE
Helium Flow:0.7 ml/min
Ion Source Temperature:250
Ionization:EI
Source Temperature:250
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