Photosynthetic C18OO fractionation is related to within- and between-species variations in the ratio of intercellular to atmospheric CO2.

The C18OO fractionation during photosynthesis (Δ18OA), a powerful isotopic proxy, is strongly influenced by the 18O exchange between CO2 and leaf water. Δ18OA is believed to relate to physiological parameters such as the evaporative 18O enrichment of leaf water (Δe), CO2 influx and efflux; however, the key mechanism remains unclear. Here, we investigated the response of Δ18OA to short-term changes in CO2 levels in 3 C3 species (Helianthus annuus, Vigna unguiculata and Triticum aestivum) and one C4 species (Cleistogenes squarrosa) grown under different levels of vapor pressure deficit (VPD) and nitrogen supply. We found a significant CO2 effect on Δ18OA for C. squarrosa, but not for the C3 species. Δ18OA was not significantly correlated with Δe in the C3 species, and Δ18OA of the C4 species was not sensitive to changes in Δe driven by VPD. The gross CO2 efflux and Δ18OA were significantly correlated for both C3 and C4 species (r2 = 0.92), demonstrating its role in regulating Δ18OA. We also found that the C3 species had significantly higher Δ18OA than the C4 species, due to the lower ratio of intercellular to atmospheric CO2 (Ci/Ca) in the latter. Δ18OA was significantly (r2 = 0.94) correlated with Ci/Ca, supported by a compiled literature dataset that included 31 species. Our study reveals the distinct difference in Δ18OA between C3 and C4 species and the remarkable relationship between Δ18OA and Ci/Ca, providing new insights into how Δ18OA can be used to infer carbon cycle processes from leaf to ecosystem scales.