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

Abstract

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 delta 13C abundance measured at -34.01 +/- 0.60 and -23.62 +/- 0.95 delta per mil (parts per thousand) (/1000), 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 1.54/1000, 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 (530/1000). 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 +/- 0.5/1000, enriched sample +268.61 +/- 8.0/1000. 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.