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Proposing a validation scheme for 13C metabolite tracer studies in high-resolution mass spectrometry
- Source :
- Analytical and Bioanalytical Chemistry
- Publication Year :
- 2019
- Publisher :
- Springer Berlin Heidelberg, 2019.
-
Abstract
- 13C metabolite tracer and metabolic flux analyses require upfront experimental planning and validation tools. Here, we present a validation scheme including a comparison of different LC methods that allow for customization of analytical strategies for tracer studies with regard to the targeted metabolites. As the measurement of significant changes in labeling patterns depends on the spectral accuracy, we investigate this aspect comprehensively for high-resolution orbitrap mass spectrometry combined with reversed-phase chromatography, hydrophilic interaction liquid chromatography, or anion-exchange chromatography. Moreover, we propose a quality control protocol based on (1) a metabolite containing selenium to assess the instrument performance and on (2) in vivo synthesized isotopically enriched Pichia pastoris to validate the accuracy of carbon isotopologue distributions (CIDs), in this case considering each isotopologue of a targeted metabolite panel. Finally, validation involved a thorough assessment of procedural blanks and matrix interferences. We compared the analytical figures of merit regarding CID determination for over 40 metabolites between the three methods. Excellent precisions of less than 1% and trueness bias as small as 0.01–1% were found for the majority of compounds, whereas the CID determination of a small fraction was affected by contaminants. For most compounds, changes of labeling pattern as low as 1% could be measured. Graphical abstract Electronic supplementary material The online version of this article (10.1007/s00216-019-01773-7) contains supplementary material, which is available to authorized users.
- Subjects :
- Metabolite
02 engineering and technology
Validation Studies as Topic
Mass spectrometry
Orbitrap
01 natural sciences
Biochemistry
Pichia
Analytical Chemistry
law.invention
Matrix (chemical analysis)
chemistry.chemical_compound
Selenium
Metabolomics
law
Stable isotope tracer experiments
TRACER
Carbon isotopologue distribution
Isotopologue
Stable isotope labeling experiments
Anion Exchange Resins
Carbon Isotopes
Chromatography, Reverse-Phase
Chromatography
Chemistry
Hydrophilic interaction chromatography
010401 analytical chemistry
Reference Standards
021001 nanoscience & nanotechnology
Chromatography, Ion Exchange
0104 chemical sciences
13C labeling
0210 nano-technology
Hydrophobic and Hydrophilic Interactions
Research Paper
Subjects
Details
- Language :
- English
- ISSN :
- 16182650 and 16182642
- Volume :
- 411
- Issue :
- 14
- Database :
- OpenAIRE
- Journal :
- Analytical and Bioanalytical Chemistry
- Accession number :
- edsair.doi.dedup.....f560a343b41ecb2e2db250203fd45fee