The landmark experiments of Schoenheimer and Rittenberg in the 1930s provided the scientific foundation for the ensuing development and application of stable isotope techniques in clinical diagnosis and research.1

Stable isotopes are non-radioactive atoms of the same chemical element, which differ only in their number of neutrons.2 Many elements also have radioactive (non-stable) isotopes. Aspects of macronutrient metabolism have been investigated employing molecules labelled with ²H (D or deuterium), ¹³C,¹⁵N, and ¹⁸O. Extensive literature is available detailing the use of ²⁵Mg, ²⁶Mg,⁴²Ca, ⁴⁶Ca, ⁴⁸Ca, ⁵⁷Fe,⁵⁸Fe, ⁶⁷Zn, and ⁷⁰Zn isotopes in studies of mineral metabolism. The most commonly used radioactive isotopes are ¹⁴C and ³H (tritium).2 More than 6000 stable isotope labelled compounds (tracers) are commercially available for use in metabolic studies. Examples of some of these tracers are [1-¹³C] leucine, [1-¹³C, ¹⁵N] leucine, [ring-D₅] phenylalanine, and [6,6]-D₂glucose. It is currently accepted that these compounds have negligible biological side effects, which renders them ethically acceptable for use in children.3

Following intravascular or oral administration, the tracer is metabolically indistinguishable from the equivalent unlabelled compound of interest (tracee). The metabolic fate of the compound can be assessed qualitatively and quantitatively by measuring the relative abundance of tracer and tracee and/or their respective metabolites with time. The detectable mass difference of tracer and tracee allows the analysis of compounds, extracted from plasma, by gas chromatography–mass spectrometry (GC–MS, picogram sample size, analytical range 0.1–100 mole %, precision ±0.2 mole %).4 ,5 The detection limit is considerably less than 0.1 mole %, when tracers with multiple stable isotope labels (for example ring-D₅ phenylalanine) are used.6Stable isotopes in breath (¹²CO_{2 …}