Colin’s first diatom growth experiment is complete. A day or so ago, I talked about the final stages of the experiment at which point the diatoms in Diatom Alley were growing successfully. Once they have consumed around 50% of the nitrate available to them, it’s time to filter the water in which they have been living. Last night, Colin and Ivia carried this out for two of the carboys. Today, I observed Colin’s filtration process and had a hands-on experience with Ivia as she filtered the remaining water from the third carboy.
Prior to filtering, Colin and Ivia divide up the water from the carboy. Note how greenish-brown the water is now, indicating the presence of phytoplankton.
Colin uses a GF/F (0.7 micron glass fiber filter) and a vacuum pump to filter out the organic bulk (any material that isn’t dissolved in the water; in other words, the particles).
The filter is thickly coated with diatoms and other particles.
The filter is then packaged, labelled, and frozen for transport back to the lab in Rhode Island. There it will be combusted in a furnace at 1000°C to generate carbon and nitrogen gases. All the nitrogen atoms will be converted to N2 gas and carbon will be converted to CO2. Gas chromatography will separate the gases based on how much they “stick” to the column. Then the N2 and CO2 will be measured in a machine called an isotope ratio mass spectrometer to determine the isotopic composition of N (d15N) and C (d13C). Some of the particle-free liquid that passed through the GF/F is bottled and frozen and will undergo analysis back in the lab, too.
Ivia continues to fill each filtering device as its water level lowers.
Meanwhile, Ivia has taken the bulk of the water from the carboys to the biology lab and is filtering out the particles using a vacuum pump. This is a slower process, as there’s a lot of water and considerable material in the water. Some of it is desirable—like the diatoms Colin has grown—while some of the phytoplankton slow the filtration process. This cannot be avoided because everyone is happy to grow when there is ample nutrients and suitable conditions such as Colin has provided. The organism gumming up the works’ name is Phaeocystis, and it’s a very common phytoplankton species. Phaeocystis individuals bunch together in the sticky mucous they produce, and together they can clog up the filters and slow down the filtering process.
All told, about 18-19L of water from each carboy are filtered through 5 micron polycarbonate filters (many are required because of how much Phaeocystis clogs the filters!), twelve at a time. An aspirator pump provides the pressure to pull water through. When the water has been filtered through, the filters are collected and frozen for cleaning and measurement of the nitrogen bound up in diatom frustules and the silicon of the frustules. Because only a small fraction of the total nitrogen in a diatom is bound in the frustule, and we clean all the other nitrogen out of the sample before analysis, we require much more material for this part of the experiment. In fact, we are filtering about 200 times more water for the diatom-bound isotope analysis than we did for the bulk analysis (19L vs 100mL)!
With a remaining liter of the water, Ivia filters out biogenic silica—silica that the diatoms have incorporated into their frustules (their glass skeletons). She uses a very fine filter (0.06 µm) for this process, but unlike Colin, she must use one made of polycarbonate plastic. If she used a GF/F (which is made of glass), even a very small and insignificant amount of glass would drastically change the proportion of the different isotopes we have in the natural environment and bias our interpretation of the isotopic signal.
Once filtration is complete, she folds it up and places it in a tiny test tube, which she will then cook in an oven at 60 degrees Celsius to drive out all the water. I (Marlo) took a turn at placing the filter on the filtration device and folding it up once filtration was complete, and it’s a fiddly business requiring great care and dexterity.
Back in the lab, and through a complex chemical process, Ivia will concentrate and purify the silicon on these filters and in the remaining water. She will send this purified, concentrated silicon through a machine called a mass spectrometer to measure the silicon isotopes present.
We look forward to seeing how the next growth experiment proceeds after Colin starts it again when we reach 61°S. When it’s complete, this filtration process will start all over again.
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