Development of an accurate and precise water-vapor equilibration method for hydrogen isotope analysis of plant non-structural carbohydrates and first applications

Philipp Schuler, Jobin Joseph, Valentina Vitali, Matthias Saurer and Marco M. Lehmann, Forest Dynamics, Swiss Federal Institute for Forest Snow and Landscape Research WSL, Birmensdorf, Zürich, Switzerland, Marc-André Cormier, Department of Earth Sciences, University of Oxford, Oxford, United Kingdom, Roland A. Werner, Nina Buchmann and Arthur Gessler, Eidgenössische Technische Hochschule Zürich ETH, Zürich, Switzerland

Background/Question/Methods
Recent studies show that the hydrogen isotopic composition (δ2H) of plant carbohydrates can be used as a proxy for metabolic processes and carbon dynamics in plants, which could be of similar importance than carbon and oxygen isotopes in ecophysiological research. Today, the analysis of carbon-bound non-exchangeable hydrogen isotopes in non-structural carbohydrates (NSC; i.e. sugars and starch) are limited due to a bias related to H-exchange between oxygen-bound hydrogen in NSC with surrounding water and vapor. However, precise and accurate δ2H measurements of NSC are urgently needed to constrain isotope fractionation processes between leaf water, primary assimilates, and structural components such as cellulose. Thus, we developed a new high-throughput method to measure non-exchangeable δ2H in NSC using a water-vapor equilibration technique. We applied the method on NSC standard material, as well as on plant NSC derived from climate chamber experiments with various species of different photosynthetic pathways (C3, C4 and CAM), growing under controlled environmental conditions (i.e. two temperature and two VPD levels).
Results/Conclusions
Our new methods enables the analysis of δ2H values in sugars and starches with high accuracy (3.0 and 4.7 ‰ compared to their reference standards) and precision (standard deviation of 3.1‰ and 1.4‰, respectively). First applications indicate that δ2H values of sugars strongly differ between C3, C4 and CAM plants at high temperature (30°C) and high VPD (2.5 kPa). δ2H values of C3 plant sugars were the lowest (~-100‰), intermediate of C4 plant sugars (~-65‰) and the highest (~-26‰) of CAM plant sugars. Further ongoing δ2H analyses of leaf water, starch and cellulose of more species under different climate settings will illuminate the variability of 2H-fractionation between species of different photosynthetic modes and the species-specific variability within these photosynthetic modes in response to climate conditions. These results will fill the gaps that are needed to improve current 2H-fractionation models, which might find widespread application in ecophysiology and dendrochronology, as well as on other paleobotanical archives for the reconstruction of past climate and plant physiological responses.