Determination of extracellular and intracellular enrichments of [1-13C]-α-ketoisovalerate using enzymatically converted [1-13C]-valine standardization curves and gas chromatography–mass spectrometry

Add time:08/31/2019    Source:sciencedirect.com

We report a validated method for the determination of extra- and intracellular [1-13C]-α-ketoisovalerate ([1-13C]-KIV) enrichments by gas chromatography–mass spectrometry. Standardization curves were prepared by enzymatic oxidation of [1-13C]-valine enriched standards of known composition. Slopes of [1-13C]-valine standardization curves (mean±SD: 0.99±0.02, n=5) and [1-13C]-KIV standardization curves (mean±SD: 0.98±0.01, n=7) were not significantly different. The method was applied for the determination of [1-13C]-KIV enrichments in plasma and tissues during [1-13C]-valine infusion in a piglet. [1-13C]-KIV enrichment could be determined±0.1 MPE (C.V. 1%), and extracellular [1-13C]-KIV enrichment was a reliable estimate of intracellular (skeletal muscle, bone growth plate) [1-13C]-KIV enrichment.

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