A quantitative measurement of the substrate fluxes that determine fasting or postprandial glucose concentrations is central to the determination of the pathophysiology of disease states and of measuring the effect of therapeutic interventions on the disease state. During the fasting state, glucose concentrations are dependent on the rate of endogenous glucose production (EGP) relative to the rate of glucose disappearance; measurement using tracer techniques is quite straightforward. The situation is more complex in the postprandial state, where gastric emptying, glucose absorption, the net sum of splanchnic extraction of ingested glucose (and therefore the rate of systemic appearance of ingested glucose), EGP, and glucose disappearance determine postprandial glucose concentrations (1, 2). The tracer-based methodologies used to measure these fluxes, and their potential pitfalls, will be discussed. Tracers: stable vs. radioisotopes. The use of a substance (tracer) that can be used to follow a naturally occurring substrate (tracee) requires that such a substance experience the identical metabolic fate as the tracee. The ideal tracer can be detected with such precision as to require administration in trace amounts, thereby avoiding any alteration in the metabolism of the tracee (3). Tracers used for the purpose of metabolic research in humans are usually identical to the tracee, except that one or more atoms differ from the more abundant naturally occurring form of that atom. Such isotopes are radioactive when they spontaneously disintegrate to form another element, releasing radiation as a by-product of the decay. For example, 3H, which has one proton and two neutrons, emits an electron when decaying to 2He (two protons and one neutron). The energy emitted by such disintegrations is detectable by a scintillation counter. The rate of decay of such isotopes may determine their utility or otherwise in the study of human metabolism; in practice, isotopes with a short half-life are typically not used in metabolic studies except to image metabolic events in specific tissues. Stable isotopes also differ in the number of neutrons present but do not decay spontaneously and therefore do not emit radiation. Gas chromatography–mass spectrometry is necessary to distinguish such isotopes. To be of use in metabolic research, the natural abundance of a given isotope (and the presence of multiple other isotopes) needs to be low. Other than the absence of radiation exposure associated with the use of stable isotopes, there is another important key distinction from radioisotopes. The presence of naturally occurring stable isotopes within the body prior to tracer infusion means that the tracer (merely) enriches the amount of this isotope. Knowledge of the background abundance of the isotope is necessary to determine the degree of enrichment required. While scintillation counting is sufficiently sensitive to detect small quantities of radioisotope, permitting infusion of “massless” amounts of tracer, this is not the case with mass spectrometry. The use of stable isotopes generally requires infusion of amounts of labeled compound large enough that the mass of tracer infused is significant and must be accounted for when calculating a metabolic rate using such isotopes.

The assumption that tracers are treated in identical fashion to their naturally occurring counterparts underpins all tracer methodology. However, the presence of radioactive impurities can introduce significant errors into calculations (4, 5). This is because a portion of the radioactive material infused is not behaving as a tracer but as a “bystander” playing no role in the reaction rate being measured and with a clearance rate …