Measurement of Precursor Enrichment for Calculating Intramuscular Triglyceride Fractional Synthesis Rate

Increased intramuscular (IM) triglyceride (TG) is associated with insulin resistance. Although stable isotope methods may provide insight; their application is hindered by measurement of the precursor. We have established the methods of measuring IM palmitoyl‐CoA (PalCoA) and Palmitoyl‐carnitine (PalCn) enrichment using LC/MS. However, the measured PalCoA enrichment is often lower than PalCn. The aim of this study was to assess the validity of these methods in the measurement of precursor enrichment. Methods Infusion of U‐ 13 C 16 ‐palmitate in 5 rabbits was performed. Leg muscle was collected before and after infusion. Samples were rapidly washed in ice‐cold saline and frozen in liquid nitrogen. On processing, samples were kept cold by means of dry ice and ice‐water bath. After extraction and TLC isolation, muscle free palmitate enrichment was measured by GC/MS. IM PalCoA and PalCn were extracted and measured using LC/MS. Results Enrichment of IM free palmitate was the lowest and PalCn was the highest, and PalCoA was at the middle (Fig. 1). The sum of PalCoA and PalCn was 2.08 ± 0.36 nmol/g, which is very small in relation to IM lipid pool (5800 nmol/g). Conclusion The findings support the notion that any lipid breakdown during muscle sampling and processing would promptly dilute the IM precursor enrichment at the sequence of free palmitate to PalCoA to PalCn. Because PalCn is the least to be affected by potential tracee dilution from in vitro lipid breakdown, it may better represent precursor enrichment in the measurement of IM TG FSR .