Novel insights into the biochemical drivers shaping hydrogen isotope values of sugar and cellulose within a plants’ leaf

Recent methodological achievements in determining the non-exchangeable hydrogen isotopic composition (δ²H_ne) of non-structural carbohydrates such as sugars allow to disentangle of so far hidden hydrogen isotope (²H) fractionation processes influencing δ²H_ne of plant carbohydrates. We conducted two climate chamber experiments to have a closer look at the basic biochemical drivers of the photosynthetic ²H fractionation between water and sugar and the post-photosynthetic ²H fractionation between sugars and cellulose in leaves: First, we studied the impact of the different biochemical reactions in 10 species with C₃, 7 species with C₄, and 8 species with CAM carbon fixation pathways, and their response to changes in temperature and vapor pressure deficit (VPD). Second, we investigated the impact of a temperature increase from 10 to 40°C in 5°C steps under a constant VPD on leaf level photosynthesis and metabolic functioning of 7 plant species. The first experiment revealed distinct differences in the photosynthetic ²H fractionation between C₃, C₄, and CAM plants. In addition, the observed intensity and direction of the shifts in δ²H_ne in response to changes in temperature and VPD in C₃ plants was species specific, absent in C₄ plants, and again species-specific in CAM plants. However, post-photosynthetic ²H fractionation was very similar among the three types of carbon fixation. We demonstrate that, in contrary to widespread believes, the ²H enrichment during post-photosynthetic ²H fractionation is driven by the carbohydrate metabolism, and not by an isotopic exchange with surrounding water. The results of the second experiment identified a plants’ metabolic activity, and its response to changes in temperature, as a major driver of the post-photosynthetic ²H fractionation of leaf sugars in C₃ species. Our results clearly demonstrate that δ²H_ne of plant carbohydrates are driven by plants metabolism and its response to the environment, which are species-specific. This will help to improve our current ability to interpret δ²H_ne chronologies in tree rings and other plant archives, and to use ²H fractionation in carbohydrates as a novel proxy to study a plants’ metabolic properties.