Accounting for mesophyll conductance substantially improves ¹³ C-based estimates of intrinsic water-use efficiency

Carbon isotope discrimination (Δ) has been used widely to infer intrinsic water-use efficiency (iWUE) of C₃ plants, a key parameter linking carbon and water fluxes. Despite the essential role of mesophyll conductance (g_m ) in photosynthesis and Δ, its effect on Δ-based predictions of iWUE has generally been neglected. Here, we derive a mathematical expression of iWUE as a function of Δ that includes g_m (iWUE_mes ) and exploits the g_m -stomatal conductance (g_sc ) relationship across drought-stress levels and plant functional groups (deciduous or semideciduous woody, evergreen woody and herbaceous species) in a global database. iWUE_mes was further validated with an independent dataset of online-Δ and CO₂ and H₂ O gas exchange measurements with seven species. Drought stress reduced g_sc and g_m by nearly one-half across all plant functional groups, but had no significant effect on the g_sc  : g_m ratio, with a well supported value of 0.79 ± 0.07 (95% CI, n = 198). g_m was negatively correlated to iWUE. Incorporating the g_sc  : g_m ratio greatly improved estimates of iWUE, compared with calculations that assumed infinite g_m . The inclusion of the g_sc  : g_m ratio, fixed at 0.79 when g_m was unknown, proved desirable to eliminate significant errors in estimating iWUE from Δ across various C₃ vegetation types.