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https://doi.org/10.5194/egusphere-2026-1369
© Author(s) 2026. This work is distributed under
the Creative Commons Attribution 4.0 License.
the Creative Commons Attribution 4.0 License.
https://doi.org/10.5194/egusphere-2026-1369
© Author(s) 2026. This work is distributed under
the Creative Commons Attribution 4.0 License.
the Creative Commons Attribution 4.0 License.
08 Apr 2026
| 08 Apr 2026
An intermediary scale setup to measure O2 fractionation factors of aquatic biosphere
Abstract. Earth atmospheric O2 is mainly produced by biosphere photosynthesis, and biosphere respiration is also one of the main consumers of this gas. The evolution of the elemental and isotopic composition of atmospheric O2 is thus linked to global biosphere productivity. Quantitative interpretation of the isotopic composition of O2 in the past as archived in polar ice cores relies on robust estimates of oxygen fractionation factors associated with the relevant biological processes: photosynthesis and respiration. In the past decades, some determinations of these biological fractionation factors were performed in uncontrolled large-scale environments or at the scale of the micro-organisms in conditions very different from the natural environment. This leads to uncertainties in the applicability of these determinations for the interpretation of isotopic composition of atmospheric O2. In order to come up with coherent estimates of oxygen biological fractionation factors applicable to the scale of plants or ecosystems, we developed a set-up of 4 closed biological chambers as a scaled-down replica of aquatic biosphere, with controlled environment parameters, and measured the evolution of O2 concentration and of its isotopic composition. We present here 3 measurement series using this set-up run with the freshwater species Chlorella vulgaris and lasting between 2 and 9 months. These measurement series enabled us to validate and optimise our newly developed system for aquatic closed biological chambers. We also determined the isotopic discrimination associated with 18O/16O of O2 during respiration as -30 ‰ which is in the upper part of the distribution of the previously published values.
How to cite. Bienville, N., Landais, A., Fiorini, S., Piel, C., Sauze, J., Lemaire, B., Geyskens, N., Prie, F., Jossoud, O., Paul, C., Chaillot, J., Lauwers, T., Chollet, S., Abiven, S., and Dapoigny, A.: An intermediary scale setup to measure O2 fractionation factors of aquatic biosphere, EGUsphere [preprint], https://doi.org/10.5194/egusphere-2026-1369, 2026.
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- RC1: 'Comment on egusphere-2026-1369', Anonymous Referee #1, 01 May 2026
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RC2: 'Comment on egusphere-2026-1369', Anonymous Referee #2, 10 Jul 2026
The authors conducted a series of experiments to determine the oxygen isotope fractionation associated with photosynthetic aquatic organisms using a mesocosm system. In contrast to previous studies that analysed dissolved gases, the present study determined the isotope composition of O₂ from gas samples collected from the chamber headspace. This approach is entirely appropriate provided that the experimental system is adequately controlled. However, I have several concerns regarding the experimental design, analytical procedures, and interpretation of the results, as outlined below.１) The manuscript describes repeated gas sampling using 5 mL flasks, but it is not clear how the cumulative removal of gas affected the composition and pressure of the closed atmosphere over experiments lasting several months. Please clarify how the withdrawn gas volume was compensated, estimate the cumulative sampled volume relative to the chamber headspace, and discuss whether repeated sampling could influence the calculated O₂ concentration and isotope ratios.2) The manuscript interprets the observed O₂ and CO₂ dynamics in terms of photosynthesis and respiration, but no quantitative information is provided on the biomass of Chlorella vulgaris during the experiments. Since biomass is expected to change substantially over the several-month incubation, it is difficult to assess whether the observed O₂ fluxes are consistent with the physiological activity of the algal culture. Please clarify whether biomass (e.g., cell density, chlorophyll a, dry weight, or particulate carbon) was monitored during the experiments, or discuss this limitation more explicitly.3) A striking result is that essentially identical photosynthetic discrimination factors are obtained under two biologically very different assumptions: (i) no respiration during the light period and (ii) respiration equal to the dark rate. Since the first assumption is physiologically unrealistic, the weak sensitivity of the calculated discrimination factor to these contrasting assumptions raises concerns about the identifiability of the model. It would be helpful if the authors could explain why the estimated ε¹⁸photo is almost insensitive to the assumed respiratory flux, and discuss whether this reflects limited information content in the measurements or an inherent limitation of the experimental design and modelling framework.4) It has been reported that changes in the O₂/N₂ and O₂/Ar ratios can introduce apparent isotope fractionation in measurements of O₂ isotopes (Abe and Yoshida, 2003; Barkan and Luz, 2003). These studies demonstrated that variations in O₂/N₂ and O₂/Ar affect not only Δ¹⁷O but also δ¹⁸O measurements. Since substantial changes in O₂ concentration are expected in the present incubation experiments, it is important to clarify whether such analytical artefacts were evaluated and corrected. If no correction was applied to the δ¹⁸O data, the authors should explain why this was considered unnecessary and discuss the potential impact on the reported fractionation factors.5) I strongly recommend incorporating continuous monitoring of key aquatic parameters, such as dissolved oxygen concentration and chlorophyll a, those are able to monitor by sensors. Such measurements would provide valuable information on changes in biomass and physiological status throughout the experiments, and would greatly strengthen the interpretation of the observed O₂ fluxes and isotope fractionation.Citation: https://doi.org/
10.5194/egusphere-2026-1369-RC2
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Nicolas Bienville
CORRESPONDING AUTHOR
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Amaelle Landais
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Sarah Fiorini
CEREEP Ecotron-IDF, CNRS, UMS3194, 77140 Saint-Pierre-lès-Nemours, France
Clément Piel
Ecotron Européen de Montpellier (UAR 3248), CNRS, Université de Montpellier, 34980 Montferrier-sur-Lez, France
Joana Sauze
Ecotron Européen de Montpellier (UAR 3248), CNRS, Université de Montpellier, 34980 Montferrier-sur-Lez, France
Benoit Lemaire
INSU Division Technique, CNRS, 91191 Gif-sur-Yvette, France
Nicolas Geyskens
INSU Division Technique, CNRS, 91191 Gif-sur-Yvette, France
Frederic Prie
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Olivier Jossoud
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Clémence Paul
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Justin Chaillot
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Thomas Lauwers
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Simon Chollet
CEREEP Ecotron-IDF, CNRS, UMS3194, 77140 Saint-Pierre-lès-Nemours, France
Samuel Abiven
CEREEP Ecotron-IDF, CNRS, UMS3194, 77140 Saint-Pierre-lès-Nemours, France
Laboratoire de Géologie de L’ENS, ENS-PSL, CNRS, IPSL, 75005, Paris, France
Arnaud Dapoigny
Laboratoire des Sciences du Climat et de l’Environnement, LSCE – IPSL, CNRS, CEA, UVSQ, Université Paris-Saclay, 91191 Gif-sur-Yvette, France
Short summary
This study describes an instrumented system of 4 closed aquatic biological chambers that allows determination of the oxygen isotopic fractionation factors of aquatic biosphere in controlled realistic conditions. We validated our system with a freshwater algae species and determined its ¹⁸O/¹⁶O isotopic discrimination during respiration to be −30 ‰, in the upper range of previously published values.
This study describes an instrumented system of 4 closed aquatic biological chambers that allows...
The reviewed manuscript presents measurements of an important parameter: The discrimination against 18O of O2 during respiration. The authors here choose to use a mesocosm approach. The average values reported here are at the top of the range of previously reported values.
While these results are interesting and important, I feel a lot is lacking. A large part of the previous work they compare to is based on experimental work in the ocean, which should reflect discrimination in natural ecosystems. As the authors note, the (dark respiration) discrimination in their mesocosm experiment is at least partly a function of the temperature, nutrient status, and experiment temperature. Thus, it is unclear how the results obtained in the experiments here could be extrapolated to Earth’s oceans.
The authors acknowledge this limitation and suggest that further experiments should test for the effects of the temperature, species, etc. To me, the advantage of a controlled lab experiment is that one can set up these parameters and understand how they control the isotopic discrimination. Moreover, in such lab experiments, the question is whether this strong discrimination is cyanide-resistant respiration-related, which could be answered by the use of inhibitors. But as a minimum, it seems to me that the temperature effect on the results presented here should be tested before the manuscript is resubmitted.
In addition, some of the reported discrimination values reported here are extreme, about -40 permil. This is well beyond what is known for respiration processes. How is this explained?
As for discrimination during photosynthesis, the authors acknowledge that their results are too uncertain to shed any new light on this question. So I wonder if including these results and discussing them adds much to the paper.
In general, the manuscript is well written and easy to follow.
Minor comments:
Title: Instead of “intermediary scale setup”, maybe a “mesocosm” will be a shorter and more often used term?
Line 70: In this paragraph, the following papers should be cited and added to the discussion :
Severinghaus, J. P., Beaudette, R., Headly, M. A., Taylor, K., & Brook, E. J. (2009). Oxygen-18 of O2 records the impact of abrupt climate change on the terrestrial biosphere. Science, 324(5933), 1431-1434.
Tables and figures: Please report epsilon in permil (i.e., -29 instead of -0.029), as in the text.