Understanding Past Oceans, One Carbon Isotope at a Time

I learned about the water cycle (evaporation, condensation, precipitation – sung to the tune of Clementine) in first grade. The carbon cycle, with its wood, burnt into carbon dioxide (CO₂), breathed by plants, came soon after. Biogeochemistry takes these concepts to a whole new level as we trace elements like carbon, phosphorus, nitrogen, and more through biological, geological, and chemical forms. In the oceans, that might include chemical carbon (CO₂ gas) fixed into biological carbon (organic sugars moving through the food web), organic and inorganic carbon drifting down the water column (as any chemical oceanographer knows), and geological carbon (in the form of carbon-rich sediment piling up on the seafloor, perhaps in a chalky deposit of calcium carbonate).

Carbon Fixation is the conversion of carbon dioxide gas into cell biomass (‘fixed carbon’). Fixed carbon can move through the food web (cells to algae to fish, for example), or is sequestered out of sight.

In this way, the oceans are global superstars, snatching CO₂ – a greenhouse gas – out of the atmosphere. After passing it around among living things within the food web, it can be sent to the abyss for long-term, geological storage. This is called carbon sequestration.

Carbon Sequestration is the storage of carbon out of the atmosphere. Dead things floating down to the ocean floor are one form of carbon sequestration. Forests – trees essentially convert CO₂ into biomass – also do the trick.

Lots of scientists want to better understand marine plankton’s role in carbon fixation. We want to be able to look at not just current processes, but how plankton and the atmosphere have interacted over time. To do this, we can look at carbon isotopes and how their concentrations change from the environment into a cell.

Let’s start with a chemistry review.

How to Track Atoms in an Ecosystem

In the oceans, carbon exists in multiple “species.” Sure, it can be fixed as organic carbon – living things. It can also exist in inorganic forms.

The most obvious on is carbon dioxide. Henry’s Law explains why CO₂ enters seawater; what happens when it does?

Henry’s Law shows that increasing the partial pressure of a gas increases its solubility. For example, increase CO₂ in the atmosphere, and you increase how much CO₂ gets dissolved in the ocean.

Well, it can remain that – dissolved CO₂ molecules. It can also react with water, H₂O, to produce various new carbonate forms. CO₂ and H₂O react to make H₂CO₃, called carbonic acid. Any acid can lose a hydrogen or two (in this case, it can donate two hydrogen atoms) so H₂CO₃ quickly breaks down in seawater to produce HCO₃^– (bicarbonate) and ^­CO₃^2- (carbonate). Each of these carbon species has its own properties in seawater, so we know that the two most common forms of carbon our phytoplankton will be working with are bicarbonate and carbon dioxide. Cells can take up both carbon species, convert them to CO₂ if necessary, and then fix them via photosynthesis.

Now, we can measure how much carbon dioxide and bicarbonate is in seawater. And we can measure carbon dioxide and bicarbonate content inside of cell cultures. What else can we measure? We can determine how many of each kind of carbon isotope exists among the molecules.

An atom has protons, electrons, and neutrons. The number of protons determines what kind of atom it is. The number of electrons determines what the electric charge is. The number of neutrons determines how heavy it is.

Carbon atoms have 6 protons. There are three carbon isotopes, depending on how many neutrons they have. C-14 is the heaviest isotope. It has 6 protons and 8 neutrons, and the nucleus isn’t stable with that many neutrons bound up. So it’s radioactive. We won’t worry about it here; it’s a very small percentage of carbon isotopes, and we are only concerned with stable, non-radioactive isotopes. The next carbon isotope is C-13, with 6 protons and 7 neutrons. It’s slightly heavier than usual, but still stable, and also quite rare. Most carbon is C-12, the well-balanced, 6 proton, 6 neutron isotope.

It turns out that many processes tend to prefer lighter isotopes to heavier ones. When water evaporates, for example, most of the water evaporating off the oceans is H₂O of the nice and light O-16 isotope form. This means that heaver water molecules, with heavier oxygen isotopes, get left behind, with lighter water molecules caught up into clouds and falling as rainfall.

Living things have their preferences too. A phytoplankton prefers light carbon-based molecules to heavier ones. So when photosynthesis occurs, the cell is selecting for lighter carbon isotopes, preferably CO₂ with a good and light C-12. Heavier C-13-based carbon dioxide gets left behind, and biomass is made up of more C-12-based carbon compounds than there are in the environment.

Fractionation is a purification process. In this case, we are looking at how carbon molecules are purified to a mixture richer in C-12 isotopes than before.

We call this selective process fractionation. It’s basically a purification process, since it distills molecules by their weight. We can measure which isotopes are present in cells. This can give us a glimpse into past environments. Fractionation rates are influenced by nutrient and light availability, so give us insight into what’s going on in cell physiology. Apply this to the fossil record, and we can use it to piece together what our atmosphere looked like in the past.

If we can track changes in carbon isotope fractionation according to an algal cell’s physiology, we have a tool for piecing together past atmospheres.

Designing a Model

Cell modeling can help us here. An algal cell is a highly complicated system, and of course different algal species might have very different mechanisms for shuffling carbon around their various compartments. In A general model for carbon isotopes in red-lineage phytoplankton: Interplay between unidirectional processes and fractionation by RubisCO (Wilkes & Pearson , 2019), researchers set about to do just that. Now, rubisco is the enzyme that actually fixes CO₂ into biomass. There are several membranes carbon must get through, however, before reaching that rubisco.

In an algae cell, rubisco is found inside the pyrenoid. That means that carbon must pass through the cell membrane to enter the cytosol. Then it must pass through another membrane to enter the chloroplast. Then it must enter the pyrenoid – and it can either pass directly through the pyrenoid membrane, or enter through thylakoid membranes, busy with the Light Reactions (carbon fixation is a dark step of photosynthesis, no light required). Each of these membrane passages results in isotope fractionation. Each passage selects for C-12, and distills the carbon pool into a richer and richer source of C-12 based carbon compounds.

To further complicate things, there are two carbon species being used here. Both dissolved CO₂ and soluble bicarbonate can be taken up by the algae. CO₂ passes through a membrane faster than HCO₃^– (diffusion is harder for ions than it is for chargeless molecules), but the cell wants to maintain a gradient of CO₂.

Diffusion is the passive transport of molecules across a membrane. Molecules will passively move across a membrane to regions of a lower concentration.

The cell wants lots of CO₂, but if it builds up too much of it, diffusion will stop working in the cell’s favor. By keeping CO₂ concentrations low, the cell will have a constant influx of CO₂ from the surrounding seawater. So algae cells actually convert CO₂ to bicarbonate, maintaining that gradient to keep the carbon coming. Finally, within the pyrenoid, any bicarbonate is converted into CO₂ so that the rubisco enzyme is saturated with it. That keeps rubisco running happily, fixing carbon via the Calvin cycle

All of these chemical reactions are summed up in Fig. 2 of the paper. And that provides a nice and clean little picture of everything going on – what do we do with it?

It turns out that this diagram, with its chemical reactions and carbon flux and fractionation across each step, can be converted into a system of equations. These equations can be used to solve for different physiological factors in the cell. So data of, say, the carbon isotope fractionation of carbon dioxide in each pool (for example) would inform a calculation of the physiology of rubisco.

In other words, experimentally derived physiological data can be used to determine model-derived physiological estimates.

After developing a model like this, then, you want data to test it with. Wilkes & Pearson did a literature review to gather cell data they could use with their model. Their literature review provided information like growth rates, CO₂ and bicarbonate concentrations in seawater, and fixed carbon in the cell. After testing their model, they ended up with some really good estimations of how selective rubisco is when picking C-12 instead of C-13. And guess what? It’s selectivity appears to depend on how much light and nutrients the cells are getting.

What This Means

Photosynthesis, environmental carbon, nutrient availability, and cell physiology (like growth rate) are all related to each other in algal cells. Their relationship to each other can be summed up in a model that tracks carbon isotope fractionation throughout the cell.

It’s typically very simple to measure carbon isotope fractionation values. That means that this model empowers a simple experimental measurement the potential to provide lots more information. With this model, we can relate fractionation to various states of algal physiology, what’s going on with the rubisco enzyme, and – even climate conditions throughout the algal fossil record.

Marine phytoplankton are, and have long been, major players in earth’s climate. They fix carbon, feed food webs, produce half of global oxygen, and protect the environment with carbon sequestration. With this new model, we have an exciting new tool to better understand them and their role.

References

Wilkes, E. B., & Pearson, A. (2019). A general model for carbon isotopes in red-lineage phytoplankton: Interplay between unidirectional processes and fractionation by RubisCO. Geochimica et Cosmochimica Acta, 265, 163–181. https://doi.org/10.1016/j.gca.2019.08.043