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Metabolites 2019, 9(4), 63; https://doi.org/10.3390/metabo9040063

HILIC-Enabled 13C Metabolomics Strategies: Comparing Quantitative Precision and Spectral Accuracy of QTOF High- and QQQ Low-Resolution Mass Spectrometry

Institute of Biochemical Engineering, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany
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Equal first authors.
Received: 29 January 2019 / Revised: 21 March 2019 / Accepted: 28 March 2019 / Published: 2 April 2019
(This article belongs to the Special Issue Mass Spectrometry-Based Metabolomics and Its Applications)
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Abstract

Dynamic 13C-tracer-based flux analyses of in vivo reaction networks still require a continuous development of advanced quantification methods applying state-of-the-art mass spectrometry platforms. Utilizing alkaline HILIC chromatography, we adapt strategies for a systematic quantification study in non- and 13C-labeled multicomponent endogenous Corynebacterium glutamicum extracts by LC-QTOF high resolution (HRMS) and LC-QQQ tandem mass spectrometry (MS/MS). Without prior derivatization, a representative cross-section of 17 central carbon and anabolic key intermediates were analyzed with high selectivity and sensitivity under optimized ESI-MS settings. In column detection limits for the absolute quantification range were between 6.8–304.7 (QQQ) and 28.7–881.5 fmol (QTOF) with comparable linearities (3–5 orders of magnitude) and enhanced precision using QQQ-MRM detection. Tailor-made preparations of uniformly (U)13C-labeled cultivation extracts for isotope dilution mass spectrometry enabled the accurate quantification in complex sample matrices and extended linearities without effect on method parameters. Furthermore, evaluation of metabolite-specific m+1-to-m+0 ratios (ISR1:0) in non-labeled extracts exhibited sufficient methodical spectral accuracies with mean deviations of 3.89 ± 3.54% (QTOF) and 4.01 ± 3.01% (QQQ). Based on the excellent HILIC performance, conformity analysis of time-resolved isotopic enrichments in 13C-tracer experiments revealed sufficient spectral accuracy for QQQ-SIM detection. However, only QTOF-HRMS ensures determination of the full isotopologue space in complex matrices without mass interferences. View Full-Text
Keywords: LC-QTOF-HRMS; LC-QQQ-MS/MS; HILIC; Quantitative metabolomics; IDMS; Isotopic distribution; Spectral accuracy; 13C-tracer studies; Corynebacterium glutamicum LC-QTOF-HRMS; LC-QQQ-MS/MS; HILIC; Quantitative metabolomics; IDMS; Isotopic distribution; Spectral accuracy; 13C-tracer studies; Corynebacterium glutamicum
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Feith, A.; Teleki, A.; Graf, M.; Favilli, L.; Takors, R. HILIC-Enabled 13C Metabolomics Strategies: Comparing Quantitative Precision and Spectral Accuracy of QTOF High- and QQQ Low-Resolution Mass Spectrometry. Metabolites 2019, 9, 63.

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