A new high-quality set of singly ((2) H) and doubly ((2) H and (18) O) stable isotope labeled reference waters for biomedical and other isotope-labeled research.

RATIONALE: Research using water with enriched levels of the rare stable isotopes of hydrogen and/or oxygen requires well-characterized enriched reference waters. The International Atomic Energy Agency (IAEA) did have such reference waters available, but these are now exhausted. New reference waters thus had to be produced in sufficient quantity, and higher characterization quality was desired. METHODS: The reference waters have been prepared gravimetrically from three parent waters: natural water, pure (2) H water and highly (18) O-enriched water. These parent waters have been thoroughly assessed for their full isotopic compositions. To ensure the integrity and correctness of the gravimetric procedure, validation measurements have been carried out on the isotopic composition of the produced reference waters by two of our laboratories. These measurements corroborate the values obtained on the basis of gravimetric data. RESULTS: Two new sets of three reference waters enriched in the stable isotopes have been produced and certified: one set of singly labeled waters, only enriched in (2) H, and another set of Doubly Labeled Waters, enriched in both (2) H and (18) O. They cover δ(2) H and δ(18) O values in the range of 800-16000 ‰ and 100-2000 ‰, respectively. The process has led to highly accurate isotopic values for these waters. CONCLUSIONS: These reference waters are now available (called IAEA-604 to IAEA-609). They will be valuable as reference materials for all fields using isotope labeling of water, most prominently, but not exclusively, biomedical research (body composition analyses, metabolic rate measurements). The two waters with the lowest enrichments will also be useful as anchor values for isotope measurements around the natural range.

A thoroughly validated spreadsheet for calculating isotopic abundances (2H, 17O, 18O) for mixtures of waters with different isotopic compositions.

RATIONALE: Oxygen and hydrogen stable isotopes are widely used tracers for studies on naturally occurring and laboratory mixtures of isotopically different waters. Although the mixing calculations are straightforward to perform, there are ample possibilities to make mistakes, especially when dealing with a large number of mixed fluids. To facilitate isotope mixing calculations and to avoid computational mistakes, a flexible tool to carry out these calculations is in demand. METHODS: We developed, in three independent efforts, spreadsheets to carry out the mixing calculations for a combination of waters with different isotopic compositions using the isotope mass balance equation. We validated our calculations by comparison of the results of the three spreadsheets for a large number of test calculations. For all the cases, we obtained identical results down to the 12(th) to 14(th) significant digit. RESULTS: We present a user-friendly, thoroughly validated spreadsheet for calculating (2) H, (17) O and (18) O stable isotopic abundances and respective isotope delta values for mixtures of waters with arbitrary isotopic compositions. The spreadsheet allows the mixing of up to 10 different waters, of which up to five can be specified using their isotopic abundances and up to five others using their isotope delta values. The spreadsheet is implemented in Microsoft Excel and is freely available from our research groups' websites. CONCLUSIONS: The present tool will be applicable in the production and characterization of singly and doubly labeled water (DLW) mother solutions, the analysis of isotope dilution measurements, the deduction of unknown isotope values of constituents for mixtures of natural waters, and many other applications.