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Amino Acid Analysis in Physiological Samples by GC-MS with Propyl Chloroformate Derivatization and iTRAQ-LC-MS/MS.
Journal article

Amino Acid Analysis in Physiological Samples by GC-MS with Propyl Chloroformate Derivatization and iTRAQ-LC-MS/MS.

  • Dettmer K Institute of Functional Genomics, University of Regensburg, Regensburg, Germany. katja.dettmer@ukr.de.
  • Stevens AP Institute of Functional Genomics, University of Regensburg, Regensburg, Germany.
  • Fagerer SR Laboratory of Organic Chemistry, Department of Chemistry and Applied Biosciences, ETH Zürich, Zürich, Switzerland.
  • Kaspar H Institute of Functional Genomics, University of Regensburg, Regensburg, Germany.
  • Oefner PJ Institute of Functional Genomics, University of Regensburg, Regensburg, Germany.
  • 2019-07-27
Published in:
  • Methods in molecular biology (Clifton, N.J.). - 2019
Amino acid analysis
Amino acids
Derivatization
Gas chromatography
Liquid chromatography-tandem mass spectrometry
Mass spectrometry
Propyl chloroformate
Stable isotope dilution
iTRAQ
Amino Acids
Carbon Isotopes
Cells, Cultured
Chromatography, High Pressure Liquid
Chromatography, Reverse-Phase
Culture Media
Formates
Gas Chromatography-Mass Spectrometry
Humans
Limit of Detection
Oxygen Isotopes
Reagent Kits, Diagnostic
Tandem Mass Spectrometry
English Two mass spectrometry-based methods for the quantitative analysis of free amino acids are described. The first method uses propyl chloroformate/propanol derivatization and gas chromatography-quadrupole mass spectrometry (GC-qMS) analysis in a single-ion monitoring mode. Derivatization is carried out directly in the aqueous samples, thereby allowing automation of the entire procedure, including addition of reagents, extraction, and injection into the GC-MS. The method delivers the quantification of 26 amino acids. The iTRAQ method employs the labeling of amino acids with isobaric iTRAQ tags. The tags contain two different cleavable reporter ions, one for the sample and one for the standard, which are detected by fragmentation in a tandem mass spectrometer (MS/MS). Reversed-phase liquid chromatography (RP-LC) of the labeled amino acids is performed prior to mass spectrometric analysis to separate isobaric amino acids. The commercial iTRAQ kit allows for the analysis of 42 physiological amino acids with a respective isotope-labeled standard for each of these 42 amino acids.
Language
  • English
Open access status
closed
Identifiers
Persistent URL
https://folia.unifr.ch/global/documents/136757
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