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Liquid and gas chromatography coupled to isotope ratio mass spectrometry for the determination of 13C-valine isotopic ratios in complex biological samples

Breuillé, Denis
Obled, Christiane
Papet, Isabelle
Schierbeek, Henk
Fay, Laurent-Bernard
Published in Journal of mass spectrometry. 2008, vol. 43, no. 10, p. 1334-1343
Abstract On-line gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) is commonly used to measure isotopic ratios at natural abundance as well as for tracer studies in nutritional and medical research. However, high-precision (13)C isotopic enrichment can also be measured by liquid chromatography-isotope ratio mass spectrometry (LC-IRMS). Indeed, LC-IRMS can be used, as shown by the new method reported here, to obtain a baseline separation and to measure (13)C isotopic enrichment of underivatised amino acids (Asp, Thr-Ser, Glu, Pro, Gly, Ala, Cys and Val). In case of Val, at natural abundance, the SD(delta(13)C) reported with this method was found to be below 1 per thousand. Another key feature of the new LC-IRMS method reported in this paper is the comparison of the LC-IRMS approach with the conventional GC-C-IRMS determination. To perform this comparative study, isotopic enrichments were measured from underivatised Val and its N(O, S)-ethoxycarbonyl ethyl ester derivative. Between 0.0 and 1.0 molar percent excess (MPE) (delta(13)C= -12.3 to 150.8 per thousand), the calculated root-mean-square (rms) of SD was 0.38 and 0.46 per thousand and the calculated rms of accuracy was 0.023 and 0.005 MPE, respectively, for GC-C-IRMS and LC-IRMS. Both systems measured accurately low isotopic enrichments (0.002 atom percent excess (APE)) with an SD (APE) of 0.0004. To correlate the relative (delta(13)C) and absolute (atom%, APE and MPE) isotopic enrichment of Val measured by the GC-C-IRMS and LC-IRMS devices, mathematical equations showing the slope and intercept of the curves were established and validated with experimental data between 0.0 to 2.3 MPE. Finally, both GC-C-IRMS and LC-IRMS instruments were also used to assess isotopic enrichment of protein-bound (13)C-Val in tibial epiphysis in a tracer study performed in rats. Isotopic enrichments measured by LC-IRMS and GC-C-IRMS were not statistically different (p>0.05). The results of this work indicate that the LC-IRMS was successful for high-precision (13)C isotopic measurements in tracer studies giving (13)C isotopic enrichment similar to the GC-C-IRMS but without the step of GC derivatisation. Therefore, for clinical studies requiring high-precision isotopic measurement, the LC-IRMS is the method of choice to measure the isotopic ratio.
Keywords AlgorithmsAnimalsBone and Bones/chemistryBuffersChromatography, GasChromatography, High Pressure LiquidData Interpretation, StatisticalMass SpectrometryRatsValine/analysis/metabolism
PMID: 18383273
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GODIN, Jean-Philippe et al. Liquid and gas chromatography coupled to isotope ratio mass spectrometry for the determination of 13C-valine isotopic ratios in complex biological samples. In: Journal of mass spectrometry, 2008, vol. 43, n° 10, p. 1334-1343. doi: 10.1002/jms.1406 https://archive-ouverte.unige.ch/unige:2413

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Deposited on : 2009-08-11

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