6.5. Isotopic tracer techniques
The doubly-labelled water technique is an appropriate method to measure field metabolic rates of free ranging insects (Wolf et al., 1996; Speakman, 1997, 1998). It uses the differential elimination of 2H and 18O isotopes from the body tissues to determine metabolic rate. Note, these are stable isotopes and, thus, not radioactive. The principle is explained by Wolf et al. (1996) as “the fact that isotope concentrations decrease exponentially with time through the natural ‘wash-out’ of CO2 and water. The hydrogen isotope is lost as water only, and the oxygen isotope as both water and CO2. Therefore, the apparent turnover rate of 18O is higher than that of 2H, and the difference between the two apparent turnover rates reflects the CO2 production rate". Wolf et al. (1996) validated this method for bumble bees. They injected small volumes (1 µl) of a mixture containing 2H and 18O. After ~10 minutes, when the mixture had equilibrated with the body water pool, they withdrew 1–2 µl of haemolymph to determine the initial 18O concentration. After 5–7 h, final blood isotope content was analysed by mass spectrometry. This technique actually provides rates of CO2 production, which can be directly used to calculate energy turnover of honey bees, because their RQ = 1 (Rothe and Nachtigall, 1989).