DETERMINATION OF THE ISOTOPE ENRICHMENT OF ONE OR A MIXTURE OF 2 STABLE LABELED TRACERS OF THE SAME COMPOUND USING THE COMPLETE ISOTOPOMER DISTRIBUTION OF AN ION FRAGMENT, THEORY AND APPLICATION TO IN-VIVO HUMAN TRACER STUDIES

Calculations of flux rates for stable isotope tracer studies are based upon enrichment values of an infused tracer. We propose the determina tion of enrichment values by gas chromatography/mass spectrometry, whi ch is based on tracer mole fraction and mass spectrometer signals, nor malized over the total signal of an ion fragment isotopomer distributi on. The method accounts for overlap of the signals of one or two trace rs and the tracee, high tracer mole fraction and incomplete labelling of the (infused) tracer. For the single and multiple tracer case a lin ear relationship between tracer mole fraction (from zero to one) and a ll normalized mass spectrometer signals is derived. This linearity ove r the entire range is demonstrated with a single (1-C-13)glucose trace r and for mixtures of (1-C-13)- and (3,3-H-2(2))tyrosine tracers. The linearity allows determination of the tracer mole fraction for two tra cers, using multiple linear regression. The corresponding calibration can rely on measurements of the pure tracer and tracee compound, witho ut weighing or check for chemical purity. This is compared with a cali bration based on tracer/tracee mixtures. Estimates for the tracer mole fraction are slightly better if based on a calibration, using standar d mixtures. In all cases the tracer mole fraction can be determined wi th high precision (coefficient of variation smaller than 5%) and high accuracy. For tyrosine it is demonstrated that the measurement of seve n channels rather than three, for the main isotopomers, does not reduc e the precision in the prediction of the tracer mole fraction. Equatio ns are also derived to use the tracer mole fraction to estimate the en dogenous production of the tracee under study conditions, assuming a s teady state of the host metabolism.