LABELED AMINO-ACID INFUSION STUDIES OF INVIVO PROTEIN-SYNTHESIS WITH STABLE ISOTOPE TRACERS AND GAS-CHROMATOGRAPHY MASS-SPECTROMETRY

Infusion of amino acids labeled with stable isotopes and monitoring of the enrichment in individual proteins allows the in vivo determination of the fractional rates of synthesis of human proteins. Interpretation of such studies requires that the specific activity of the tracer at the site of protein synthesis be known. To avoid having to measure amino acyl-ribonucleic acid (t-RNA), methods are described for the isolation, and subsequent evaluation of the enrichment of four amino-acid metabolites in human plasma or urine. The degree to which each metabolite allows intracellular amino acyl-t-RNA enrichment to be estimated by measuring the steady-state protein enrichment during long-term tracer infusion is assessed. Three healthy subjects were used. Four amino acid tracers were infused simultaneously for 16 h and the enrichment in amino acids isolated from apolipoprotein B (apoB) and in the corresponding amino acid metabolites was measured. The pairs of compounds used were [N-15]glycine and [N-15]hippurate, [1-C-13]leucine and [1-C-13]ketoisocaproate, [1-C-13]lysine and [1-C-13]aminoadipate, and [ring-H-2(5)]phenylalanine and [H-2(5)]phenylacetyl-glutamine. Enrichment in apoB reached a plateau at the end of the study. Hippurate, ketoisocaproate, and aminoadipate appear to be candidates as precursor equivalents in stable-isotope studies of hepatic protein turnover.