A new measurement technique reveals temporal variation in dO of leaf-respired CO2

The oxygen isotope composition of CO2 respired by Ricinus communis leaves (d OR) was measured under non-steady-state conditions with a temporal resolution of 3 min using a tunable diode laser (TDL) absorption spectrometer coupled to a portable gas exchange system. The SD of d 18O measurement by the TDL was 0.2‰ and close to that of traditional mass spectrometers. Further, d OR values at isotopic steady state were comparable to those obtained using traditional flask sampling and mass spectrometric techniques for R. communis grown and measured in similar environmental conditions. As well as higher temporal resolution, the online TDL method described here has a number of advantages over mass spectrometric techniques. At isotopic steady state among plants grown at high light, the ‘one-way flux’ model was required to accurately predict d OR. A comparison of measurements and the model suggests that plants grown under low-light conditions have either a lower proportion of chloroplast CO2 that isotopically equilibrates with chloroplast water, or more enriched d 18O of CO2 in the chloroplast that has not equilibrated with local water. The high temporal resolution of isotopic measurements allowed the first measurements of d OR when stomatal conductance was rapidly changing. Under non-steady-state conditions, d OR varied between 50 and 220‰ for leaves of plants grown under different light and water environments, and varied by as much as 100‰ within 10 min for a single leaf. Stomatal conductance ranged from 0.001 to 1.586 mol m-2 s-1, and had an important influence on d OR under non-steady-state conditions not only via effects on leaf water H2O enrichment, but also via effects on the rate of the one-way fluxes of CO2 into and out of the leaf. Key-words: leaf respiration; leaf water enrichment; oxygen isotope; tunable diode laser. INTRODUCTION Interpretation of variation in the oxygen isotope composition of atmospheric CO2 may provide a valuable method of verification of carbon cycle models at both global (Francey & Tans 1987; Farquhar et al. 1993; Ciais, Denning & Tans 1997) and ecosystem scales (Yakir & Wang 1996; Bowling et al. 2003a; Ogée et al. 2004; Ometto et al. 2005). Both respiration and photosynthesis affect the oxygen isotope composition of atmospheric CO2, and under some conditions by an equally large amount (Cernusak et al. 2004; Seibt et al. 2006; Seibt, Wingate & Berry, in press). CO2 respired by ecosystem components (e.g. leaves, stems and soil) reflects isotopic exchange between oxygen in CO2 and water within components. Water within one ecosystem component may differ isotopically from water in another. For example, leaf water is often significantly more enriched in O than soil water, so that CO2 respired by leaves is expected to be more enriched than soil-respired CO2 (Flanagan et al. 1997; Flanagan, Kubien & Ehleringer 1999). Measurements of the isotope composition of pools of water within ecosystems suggest that leaf water, in particular, is highly dynamic in time and space (Lai et al. 2006; Seibt et al. 2006). Even within relatively simple ecosystems, it is not possible to measure the oxygen isotope composition of leaf-respired CO2 (dOR) at relevant temporal and spatial resolutions, particularly when ecosystem measurements are made at half-hourly temporal resolution using new tunable diode laser (TDL) absorption spectrometric techniques (e.g. Griffis et al. 2005). Hence, accurate models of dOR are required to interpret ecosystem isoflux measurements. A model that describes the environmental and physiological influences on dOR, and accounts for the one-way fluxes of CO2 into and out of the leaf, has recently been presented by Cernusak et al. (2004). The inclusion of the one-way fluxes creates a model analogous to the CO2 invasion effect in soils described by Tans (1998; also see Miller et al. 1999; Stern, Amundson & Baisden 2001), and to the original treatment of isotopic effects during CO2 assimilation (Farquhar et al. 1993). Cernusak et al. (2004) tested their model with leaves in controlled-environment gas exchange chambers under isotopic steady state and were Correspondence: Margaret M. Barbour. Fax: +64 3 325 2418; e-mail: [email protected] Plant, Cell and Environment (2007) 30, 456–468 doi: 10.1111/j.1365-3040.2007.01633.x © 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd 456 able to demonstrate significant departures of dOR from values predicted by simpler models considering just the net respiratory flux (e.g. Flanagan et al. 1997, 1999; Bowling et al. 2003a). dOR predicted by the one-way flux model at isotopic steady state deviates from that predicted by a net flux model when the difference in isotopic composition of CO2 between that in the chloroplast (dOc) and that in the ambient air (dOa) is large, and when stomatal conductance is high. Both these conditions were met in the model evaluation experiment conducted by Cernusak et al. (2004). However, it is not clear if the one-way flux model adequately predicts dOR when stomatal conductance is very low (but significantly greater than cuticular conductance alone; Donovan, Richards & Linton 2003; Barbour et al. 2005), as would be typical for most species at night in natural conditions. We have three objectives in this paper. Firstly, we demonstrate the application of TDL absorption spectroscopy coupled to a portable gas exchange system for leaf-level measurements of dOR and compare with those reported by Cernusak et al. (2004) for plants under similar growth and measurements conditions but using traditional mass spectrometric techniques. Secondly, we quantify nonsteady-state changes in dOR in the first 50 min after the plants were moved from the light into the dark. Finally, we tested theoretical models at isotopic steady state over a wide range in measured stomatal conductance, including very low values (0.001 mol m-2 s-1).