Recent advances in understanding the metabolic origin and the temporal dynamics in delta(13)C of dark-respired CO(2) (delta(13)C(res)) have led to an increasing awareness of the importance of plant isotopic fractionation in respiratory processes. Pronounced dynamics in delta(13)C(res) have been observed in a number of species and three main hypotheses have been proposed: first, diurnal changes in delta(13)C of respiratory substrates; second, post-photosynthetic discrimination in respiratory pathways; and third, dynamic decarboxylation of enriched carbon pools during the post-illumination respiration period. Since different functional groups exhibit distinct diurnal patterns in delta(13)C(res) (ranging from 0 to 10 per thousand diurnal increase), we explored these hypotheses for different ecotypes and environmental (i.e. growth light) conditions. Mass balance calculations revealed that the effect of respiratory substrates on diurnal changes in delta(13)C(res) was negligible in all investigated species. Further, rapid post-illumination changes in delta(13)C(res) (30 min), which increased from 2.6 per thousand to 5 per thousand over the course of the day, were examined by positional (13)C-labelling to quantify changes in pyruvate dehydrogenase (PDH) and Krebs cycle (KC) activity. We investigated the origin of these dynamics with Rayleigh mass balance calculations based on theoretical assumptions on fractionation processes. Neither the estimated changes of PDH and KC, nor decarboxylation of a malate pool entirely explained the observed pattern in delta(13)C(res). However, a Rayleigh fractionation of (12)C-discriminating enzymes and/or a rapid decline in the decarboxylation rate of an enriched substrate pool may explain the post-illumination peak in delta(13)C(res). These results are highly relevant since delta(13)C(res) is used in large-scale carbon cycle studies.
2009 John Wiley & Sons, Ltd.