The stable isotope composition of transpired water and the rate of change in leaf water enrichment in response to variable environments

Previous research has shown that during daylight hours the isotope composition of leaf water is generally well approximated by steady-state leaf water isotope enrichment models. However, there is little direct confirmation of isotopic steady state (ISS) transpiration. Here we use a novel method to evaluate the frequency (or infrequency) of ISS transpiration and the rate of change in leaf water enrichment when leaves are exposed to a variable environment. Specifically, our study had three goals. First, we wanted to develop a new method to measure the isotope fluxes of transpiration that relies on isotope ratio infrared spectroscopy (IRIS) and highlight how an IRIS instrument can be coupled to plant gas exchange systems. In doing so, we also developed a method for controlling the absolute humidity entering the gas exchange cuvettes across a wide range of concentrations (approximately 4000 ppmv to 22000 ppmv) without changing the isotope composition of water vapour entering the cuvette. Second, we quantified variation in the isotope composition of transpired water vapor and the rate of change in leaf water enrichment that can occur as a result of changes in relative humidity, leaf surface conductance to water vapour, leaf temperature and the isotope composition of atmospheric water vapor. Third, we examine the differences between steady state and non-steady state model predictions of leaf water enrichment at the site of evaporation. In our measurements the isotopic compositions of transpired water were neither stable nor equal to source water until leaves had been maintained at physiological steady state for at least 40 minutes. Additionally when transpiration was not at ISS, the steady state model predictions of leaf water enrichment at the site of evaporation exceeded non steady-state model predictions by up to 8 per mil. Further, the rate of change in leaf water enrichment was highly sensitive to variation in leaf water content. Our results suggest that a variable environment is likely to preclude isotopic steady-state transpiration and that this effect would be exacerbated by lengthy leaf water turnover times.