Assessing and avoiding C isotopic contamination artefacts in mesocosm-scale 13CO2/12CO2 labelling systems: from biomass components to purified carbohydrates and dark respiration

Quantitative understanding of plant carbon (C) metabolism by 13CO2/12CO2-labelling studies requires absence (or knowledge) of C-isotopic contamination artefacts during tracer application and sample processing. Surprisingly, this concern has not been addressed systematically and comprehensively yet is especially crucial in experiments at different atmospheric CO2 concentrations ([CO2]), when experimental protocols require frequent access to the labelling chambers. Here, we used a plant growth chamber-based 13CO2/12CO2 gas exchange-facility to address this topic. The facility comprised four independent units, with two chambers routinely operated in parallel under identical conditions except for the isotopic composition of CO2 supplied to them (δ13CCO2 −43.5‰ versus −5.6‰). In this setup, dδ13CX (the measurements-based δ13C-difference between matching samples X collected from the parallel chambers) is expected to equal dδ13CRef (the predictable, non-contaminated δ13C-difference ), if sample-C is completely derived from the contrasting CO2 sources. Accordingly, contamination (fcontam) was determined as fcontam = 1– dδ13CX/dδ13CRef in this experimental setup. Determinations were made for biomass fractions, water-soluble carbohydrate (WSC) components and dark respiration of Lolium perenne (perennial ryegrass) stands following growth for ∼9 weeks at 200, 400 or 800 µmol mol− 1 CO2, with a terminal two weeks-long period of extensive experimental disturbance of the chambers.