Breath water-based doubly labelled water method for the noninvasive determination of CO₂ production and energy expenditure in mice

ABSTRACT

We explored a novel doubly labelled water (DLW) method based on breath water (BW-DLW) in mice to determine whole body CO₂ production and energy expenditure noninvasively. The BW-DLW method was compared to the DLW based on blood plasma. Mice (n = 11, 43.5 ± 4.6 g body mass (BM)) were administered orally a single bolus of doubly labelled water (1.2 g H₂¹⁸O kg BM⁻¹ and 0.4 g ²H₂O kg BM⁻¹, 99 atom% (AP) ¹⁸O or ²H). To sample breath water, the mice were placed into a respiration vessel. The exhaled water vapour was condensed in a cold-trap. The isotope enrichments of breath water were compared with plasma samples. The ²H/¹H and ¹⁸O/¹⁶O isotope ratios were measured by means of isotope ratio mass spectrometry. The CO₂ production (RCO₂) was calculated from the ²H and ¹⁸O enrichments in breath water and plasma over 5 days. The isotope enrichments of breath water vs. plasma were correlated (R² = 0.89 for ²H and 0.95 for ¹⁸O) linearly. The RCO₂ determined based on breath water and plasma was not different (113.2 ± 12.7 vs. 111.4 ± 11.0 mmol d^–1), respectively. In conclusion, the novel BW-DLW method is appropriate to obtain reliable estimates of RCO₂ avoiding blood sampling.

KEYWORDS:

• Energy expenditure
• doubly labelled water
• hydrogen-2
• in vivo measurements
• mice
• oxygen-18
• stable isotope tracer techniques

Acknowledgements

We thank Ingrid Brüning, Ute Lüdtke, Kirsten Karpati, Axel Fischer and Helmut Scholze for analytical and statistical assistance, Marzena Kucia is gratefully acknowledged for animal management, and Roland Gaeth for help with animal care.

Disclosure statement

No potential conflict of interest was reported by the authors.