Stable-isotope method for determining the gastrointestinal handling of [1-13C]palmitic acid

The 13C enrichment in individual fatty acids extracted from human feces following the oral administration of [1‐13C]palmitic acid has been determined using a novel approach based upon gas chromatography‐isotope ratio mass spectrometry. The method was established and tested for precision and repeatability. Analytical precision was determined from 10 repeated injections of a sample containing 16∶0 and 18∶0 with levels of δ13C abundance measured at −34.01±0.60 and −23.62±0.95 delta per mil (parts per thousand) (‰), respectively (mean±SD). For the repeatability study, measurement of enrichment of the same mixture of unlabeled fatty acid methyl ester (FAME) standards (13∶0, 14∶0, 16∶0, and 18∶0) was found to have standard deviations (0.45, 0.56, 1.46 and 1.54‰ respectively). When labeled [1‐13C]palmitic acid was serially diluted with naturally enriched palmitic acid, a linear relationship was obtained to a dilution of 10% enriched compound (530‰). FAME were prepared from two fecal samples from a normal healthy adult; the first, a baseline specimen, containing no added label and the second, followed a single oral dose of [1‐13C]palmitic acid and was enriched. Enrichment in 13C was confined to the solvent‐soluble fraction following lipid extraction, and was only identified with prior acidification. The enrichments were measured in triplicate, baseline sample −32.66±0.5‰ enriched sample +268.61±8.0‰ Enrichment was restricted to the labeled species consumed, 16∶0. The methodology described here allows for the separation of compounds prior to the determination of enrichment and can be utilized to contribute to a more complete description of the gastrointestinal handling of labeled substrates than previously obtained.