DataEntryID 63
    General information
      Manuscript title: Metabolomics-driven quantitative analysis of ammonia assimilation in E. coli.
      PubMed ID: http://www.ncbi.nlm.nih.gov/pubmed/19690571
      Journal: Molecular Systems Biology
      Year: 2009
      Authors: Jie Yuan, Christopher D Doucette, William U Fowler, Xiao-Jiang Feng, Matthew Piazza, Herschel A Rabitz, Ned S Wingreen and Joshua D Rabinowitz
      Affiliations: Lewis Sigler Institute for Integrative Genomics, Princeton University, Princeton, NJ, USA and Department of Chemistry, Princeton University, Princeton, NJ, USA
      Keywords: E. coli, ammonia assimilation, nitrogen upshift
      Full text article: https://kimosys.org/rails/active_storage/blobs/eyJfcmFpbHMiOnsibWVzc2FnZSI6IkJBaHBBb0FFIiwiZXhwIjpudWxsLCJwdXIiOiJibG9iX2lkIn19--16cf1ea9d90cd424c31a238c3ce7798ef422f3ce/Yuan_2009.pdf
      Project name: not specified

    Experiment description
      Organism: Escherichia coli
      Strain: NCM 3722 and mutants
      Data type: time-series data of metabolites
      Data units: mM
      Execution date: not specified

    Experimental details
      Temperature (°C): 37.0
      pH: ≈ 7.0
      Carbon source: glucose
      Culture mode: batch
      Process condition: anaerobic
      Dilution rate (h⁻¹): —
      Working volume: not specified L
      Biomass concentration (g/L): not specified
      Medium composition: Minimal salts media with 10 mM NH4Cl and 0.4 % glucose.
      General protocol information: Sampling Method: filter cultures were prepared by passing 5 ml of exponentially growing liquid batch culture through membrane filters.; Quenching: filters were submerged directly into -75 ºC methanol or -20 ºC 40:40:20 acetonitrile:methanol:water with 0.1 % formic acid.; Extraction list: methanol; Analysis list: LC-ESI-MS;
      Methods description: Detailed protocols for preparing filter cultures and extracting metabolites have been published [1,2].
Metabolite measurement - Cell extracts were analyzed by LC-electrospray ionization (ESI)-MS/MS on a Thermo Quantum triple quadrupole mass spectrometer operating in selected reaction monitoring (SRM) mode. All samples were analyzed within 24 h of their preparation. Separate LC runs were conducted for positive and negative ionization modes, using hydrophilic interaction chromatography and ion-pairing reversed phase chromatography, respectively [3].
Amino acids were derivatized before their quantitation by LC-ESIMS/MS. Cell extract (200 µl) was mixed with triethylamine (5 µl) and benzyl chloral formate (1 µl) to convert amines to N-benzylcarbamate (Cbz) derivatives [4].
As internal standard signals did not vary substantially or systematically between samples within a given time course, the reported metabolite concentration changes are between-sample ratios of the peak heights of the SRM chromatograms without correction for the internal standard response, except for glutamate and glutamine, in which correction for internal standards was performed to maximize quantitative accuracy.

-----------References-------------
[1] Bennett B, Yuan J, Kimball E, Rabinowitz J (2008) Absolute quantitation of intracellular metabolite concentrations by an isotope ratio-based approach. Nat Protoc 3: 1299–1311. http://doi.org/dx4wxn
[2] Yuan J, Bennett B, Rabinowitz J (2008) Kinetic flux profiling for quantitation of cellular metabolic fluxes. Nature Protoc 3: 1328–1340. http://doi.org/cprs6m
[3] Lu W, Bennett BD, Rabinowitz JD (2008). Analytical strategies for LC-MS-based targeted metabolomics. J Chromatogr B Analyt Technol Biomed Life Sci 871: 236–242. http://doi.org/bc7n2m
[4] Kraml CM, Zhou D, Byrne N, McConnell O (2005) Enhanced chromatographic resolution of amine enantiomers as carbobenzyloxy derivatives in high-performance liquid chromatography and supercritical fluid chromatography. J Chromatogr A 1100: 108–115. http://doi.org/fwnt9q 
      Data file: http://kimosys.org/repository/63/download?parameter=1151; 
      Alternative formats: http://kimosys.org/repository/63/attached_files/60/download; http://kimosys.org/repository/63/attached_files/59/download; http://kimosys.org/repository/63/attached_files/58/download; http://kimosys.org/repository/63/attached_files/57/download; http://kimosys.org/repository/63/attached_files/56/download; http://kimosys.org/repository/63/attached_files/55/download; http://kimosys.org/repository/63/attached_files/54/download; http://kimosys.org/repository/63/attached_files/53/download; 

    Submission and curation
      Entered by: Administrator KiMoSys
      Created: 2013-06-12 13:00:46 UTC
      Updated: 2020-04-24 16:10:33 UTC
      Version: 0
      Status: (reviewed) 2013-12-06 17:17:38 UTC
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