Spectroscopy for stable isotope measurements in metabolic physiology

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Stable isotopes are heavier elements due to an additional neutron (or neutrons) over the normal isotope, but they are non-radioactive. These isotopes are used to tag molecules that we wish to measure the metabolism of in experiments. There are very few laboratories that use stable isotope-labelled tracers to study human physiology. Traditionally, the method of a primed infusion of stable isotope-labelled biomolecules has been used. Infused tracers are expensive, require sterile and pyrogen-free preparation, and only yield short-term (hours) results as participants are confined to a bed/chair while being infused. Recently, we have begun to use oral ingestion of heavy water (heavy hydrogen - 2H- or deuterium D-labelled), which rapidly equilibrates and, via normal metabolism, labels biomolecules such as non-essential amino acids, which then become the tracer. We are now perfecting methods to measure the D-labelling of other molecules, including lipids, RNA, and DNA. The oral dosing of heavy water allows us to make highly sensitive flux measurements of several biomolecules, simultaneously saving time and money and rapidly advancing our discovery work. Compared to conventional infused tracers, ingestion of heavy water is remarkably cost-effective, lower risk as it is ingested rather than injected, and has greater biomolecule labelling potential, and thus has several methodological advantages over conventional infused tracers. We also wish to incorporate heavy oxygen (18O) analysis to add greater utility to this stable isotope methodology. The instrument we seek is a cavity ring-down spectroscopy (CRDS) unit that will measure both D and 18O in biological samples to support integrative and innovative studies of metabolic physiology and energy expenditure in humans. The CRDS unit is more affordable than, and yet as accurate as, the traditional (high-maintenance cost) isotope-ratio mass spectrometer. Acquisition of a CRDS unit would provide a unique training opportunity for HQP. The CRDS unit will result in a rapid advance of planned discovery research by alleviating a substantial bottleneck for sample analysis, greatly reducing analytical costs, and expanding the use of lower-cost isotope-labelled water for evaluating multiple metabolic pathways. The CRDS can analyze both 18O and D and allowing estimation of free-living energy expenditure and whole-body water content, which are key research applications for each of the applicants on this proposal. This equipment will be used by three senior and two junior NSERC-supported researchers to advance their use of stable isotopes in physiological research. The equipment will be used by ~20 HQP at various levels of training from undergraduate to postdoctoral fellows. Because of its ease of use, we will use the CRDS to run samples from various applications. Acquisition of the CRDS unit would elevate our research productivity and us to forge new national and international interdisciplinary collaborations.