Disentangling direct effects of temperature and vapour pressure deficit on leaf gas exchange: mechanistic insights from online stable isotope techniques

Strong covariation between temperature and vapour pressure deficit (VPD) in nature creates challenges for understanding the direct effects of the two on leaf gas exchange. Measurements of stable isotope discrimination in CO₂ and H₂O provide additional insights into physiological and biochemical processes during leaf gas exchange. We investigated mechanistic causes of variations in net photosynthesis rate (A_n) and stomatal conductance (g_s) with increasing temperature at constant VPD and increasing VPD at constant temperatures.

We conducted combined leaf gas exchange and online isotope discrimination measurements on four common European tree species (Fagus sylvatica, Picea abies, Quercus petraea, and Tilia cordata) (1) across a temperature range of 5–40°C, while maintaining a constant VPD (~0.8 kPa) and (2) across a VPD range of 1–4 kPa, while maintaining a constant temperature (~30°C). The experiments were conducted without soil water limitation. The whole plant along with the whole instrumental setup were heated to prevent condensation when the dew point temperature within the leaf cuvette was higher than the room temperature.

Above the optimum temperature for photosynthesis (~30°C), we observed a decoupling of g_s and A_n across all tested species, with g_s increasing but A_n decreasing. Measurements of carbon and oxygen isotope discrimination indicated that during this decoupling, mesophyll conductance to the chloroplast decreased consistently and significantly among species; however, this reduction did not lead to reductions in CO₂ concentration at the chloroplast surface or the chloroplast stroma. Both g_s and A_n decreased, while the transpiration rate increased with increasing VPD. The relative humidity inside the leaf, derived from the oxygen isotope discrimination measurements, decreased from 100% to around 70% with increasing VPD, suggesting a progressive unsaturation of vapour pressure in the substomatal cavity. Accounting for the unsaturation, we found decreased CO₂ concentration in the intercellular air spaces and at the chloroplast stroma with increasing VPD; however, mesophyll conductance and CO₂ concentration at the chloroplast surface remained relatively stable.

We conclude that the effects of temperature and VPD on leaf gas exchange are distinctly different. The reduction in A_n at higher temperatures, unlike that at higher VPD, was not associated with stomatal closure and thus a restricted supply of CO₂ to the chloroplasts. Instead, it was more likely caused by Rubisco deactivation and/or a reduction of the electron transport rate. The unsaturation of vapour pressure inside leaves must not be ignored at VPD higher than 1 kPa, as it is vital for accurate estimations of g_s and the CO₂ concentration in the internal air spaces of leaves. Under non limiting soil water supply, the increases in leaf water loss due to increased leaf transpiration at higher temperature and VPD are important for plants to strategically cope with severe heat and dry conditions.