Abstract
A new method for accurately analyzing octanoate enrichment in plasma was developed and validated. Samples were derivatized directly in plasma by transesterification with isobutanol and were analyzed by gas chromatography–mass spectrometry (GC-MS). This method was developed to analyze the precursor enrichment in a stable isotope tracer protocol. Glyceryl tri[1,2,3,4-13C4] octanoate, a stable isotope-labeled medium-chain triglyceride (MCT), was orally administered in combination with (1) exclusively MCT or (2) a combination of protein, carbohydrates, and MCT to investigate the metabolic route of oral MCT under various conditions. Accurate analysis of octanoate enrichment in plasma at concentrations as low as 0.43 μM (lower limit of quantification, LLOQ) was performed. This is an improvement of about twenty times for the LLOQ for analysis of the enrichment of octanoate when compared with the gold-standard method for fatty acid analysis (methyl esterification). Moreover, we found that‚ with this gold-standard method, study samples were easily contaminated with (unlabeled) octanoate from other sources, leading to biased, incorrect results. The precision and linearity obtained using the new method were good (coefficient of variation intraday < 9.1%, interday < 9.3%, R2 of the calibration curve > 0.99). The sensitivity was sufficient for analyzing samples obtained using the stable isotope protocol. This new method is more sensitive than methyl esterification and it minimizes the risk of contamination.Graphical abstract
Highlights
One of the clinical applications of medium-chain triglycerides (MCT) is as a dietary supplement for patients with a disorder involving mitochondrial long-chain fatty acid oxidation [1]
To measure the precursor enrichment in plasma samples obtained during the stable isotope protocol, we aimed to develop and validate a gas chromatograph (GC)-MS method to determine the enrichment of the 1,2,3,4-13C4-octanoate tracer at low concentrations in plasma
We developed a new gas chromatography–mass spectrometry (GC-MS) method for plasma octanoate enrichment analysis using isobutylation
Summary
One of the clinical applications of medium-chain triglycerides (MCT) is as a dietary supplement for patients with a disorder involving mitochondrial long-chain fatty acid oxidation [1]. These patients are unable to oxidize long-chain triglycerides (LCT), but are capable of oxidation of MCT. Both are fats, MCT and LCT are processed differently within the body. MCT oxidation circumvents per definition the defective pathway of the patients with a disorder involving mitochondrial long-chain fatty acid oxidation. MCT is able to compensate for the insufficient long-chain fatty acid oxidation and serves as an van Harskamp D. et al
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