New insights into the mechanisms of post-rubisco isotope fractionation from combined analysis of intramolecular 13C and deuterium abundances in Pinus nigra tree-ring glucose

Understanding isotope fractionation mechanisms is fundamental for analyses of plant ecophysiology and paleoclimate based on tree-ring isotope data. To gain new insights into isotope fractionation, we analysed intramolecular 13C discrimination in tree-ring glucose ({Delta}i, i = C-1 to C-6) and metabolic deuterium fractionation at H1 and H2 ({varepsilon}met) combinedly. This dual-isotope approach was used for isotope-signal deconvolution. We found evidence for metabolic processes affecting{Delta} 1 and{Delta} 3 which respond to air vapour pressure deficit (VPD), and processes affecting{Delta} 1,{Delta} 2, and{varepsilon} met which respond to precipitation but not VPD. These relationships exhibit change points dividing a period of homeostasis (1961-1980) from a period of metabolic adjustment (1983-1995). Homeostasis may result from sufficient groundwater availability. Additionally, we found{Delta} 5 and{Delta} 6 relationships with radiation and temperature which are temporally stable and consistent with previously proposed isotope fractionation mechanisms. Based on the multitude of climate covariables, intramolecular carbon isotope analysis has a remarkable potential for climate reconstruction. While isotope fractionation beyond leaves is currently considered to be constant, we propose significant parts of the carbon and hydrogen isotope variation in tree-ring glucose originate in stems (precipitation-dependent signals). As basis for follow-up studies, we propose mechanisms introducing{Delta} 1,{Delta} 2,{Delta} 3, and{varepsilon} met variability.