Animal isotopic labelling experiments are precious and scientifically necessary. These strategies improve our
understanding of metabolism. Experimental protocols can be different: bolus of isotopic tracer, prolonged
administration. Consequently, isotopic tracer incorporation differs in nature and quantity according to the
metabolic pathways targeted. For high enrichment, mass spectrometry can be sufficient to detect labelled
compounds. But, for low enrichment experiments, Isotop-Ratio Mass Spectrometry (IRMS) is completely suitable. This technic allows to measure the relative abundance of isotope in a sample with different configurations. First, samples can be analyzed directly in EA-IRMS (Elemental Analyser) which allows to determine the
global enrichment. Secondly, enrichment can be measured for specific metabolites by gas chromatography
(GC) coupled to IRMS detector.
In this presentation, we focus on the incorporation of 13C lactate through 2 injections into mice. Livers and
inguinal adipose tissues were studied. While labelled tissues or fluids are precious and to have a maximum of
information on a single sample, we developed a dual extraction. This extraction permits to extract at the same
time energetic metabolites and amino acids (in aqueous phase) and lipids (in organic phase). Neutral lipids,
phospholipids and fatty acids of the lipids were separated by Thin Layer Chromatography (TLC) to be analyzed
specifically. Different fractions sample (native tissue, organic and aqueous phase, and lipids TLC fraction)
were analyzed by EA-IRMS. Amino acids and fatty acids were profiled by GC-C-IRMS and GC-MS. Enrichment
have been observed on different fractions and metabolites.
This preliminary work allows the comparison of different techniques to follow and quantified the incorporation of labelled metabolites in polar and apolar fraction extracted from different organs of a mouse.