filling a terrarium with carbon dioxide

[Lorenzo192]

For my experiment, I am trying to show that cyanobacteria can change the atmosphere in a tank filled with carbon dioxide. This is supposed to show that Mars could be terraformed with cyanobacteria. However, I am having real problems getting the tank filled with carbon dioxide. I can get a lot in there, but it seems to disappear and won't stay above a certain level. We have oxygen and CO2 sensors, so we know oxygen is coming in. We have tried all sorts of things to seal up the tank, but there are holes (taped up and with sealed with vacuum grease) where the probes are. We can get it to less than 1% oxygen, but then it goes up to 10-12% after a few hours. How can I show that the oxygen increase is due to the cyanobacteria if I can't be sure it isn't just leaking in? How low should we be able to get the oxygen level? And is there a better way to do this than with dry ice (that is also affordable)?

Thanks for any help you can give. One thought: would we need to do another control, like have another tank that starts out at the same level? That's the only thing I can think of, but it's not something I really want to do! I thought it we got it stabilized at a certain level, that would be enough.

Lorenzo Montoya

[SciB]

Hi Lorenzo,

What I great idea for a science project! I’m hoping the Mars mission gets underway soon so I can see the planet converted into an earthlike home for humans—and maybe get to go there as an astrotourist!

Since I can’t see what your ‘terrarium’ looks like I’m going to suggest that you might want to try just using a large jar with a screw lid that has a silicone gasket. You could drill holes in the lid for the sensors and pass them through using small rubber stoppers with holes in them of the appropriate size. An oxygen molecule is not very big and you will always get some leakage, especially when atmospheric pressure changes.

You mentioned that you did not want to do the control minus the cyanobacteria—not a good idea. That is exactly the control you do need to prove that the O2 is coming from the bacteria and not the air. You would not have to buy another set of sensors. Just make holes in the lid as you did for the other jar and put in solid rubber stoppers instead of the ones with holes. When you want to measure the gas all you have to do is switch the stoppers.

Dry ice is a good, cheap source of CO2 and I would stick with that.

Temperature has a fairly large effect on the volume of a gas and bacteria generally have an optimal temperature that they like to grow at. I did a quick search and this article gives 29.2C as the optimum temp for cyanobacteria: http://onlinelibrary.wiley.com/doi/10.1 ... x/abstract
29C is warmer than most people keep their homes especially in winter so you might want to rig up an incubator using a lamp with an incandescent light bulb in a big cardboard or styrofoam box. The cyanos do need light to grow, right? So with an incubator you can provide them light and heat at the same time. Here’s one plan for building a diy incubator but there are lots more on the web: http://skeptacles.blogspot.com/2012/09/ ... ogurt.html

I hope this gives you some ideas for solving your leak problem. Please let us know how it is going and we’ll try to help you if you get stuck again.

Good luck!

Sybee

[Lorenzo192]

I tested a jar before setting up the experiment and it held the level of oxygen pretty well. Now the experiment is set up with the control and the Cyanobacteria. Both jars have sand (regolith) and some water. Now the levels of oxygen are increasing. Is this caused by a reaction with the carbon dioxide and the water?
Thanks for your help again.
Lorenzo

[SciB]

You did not say whether both the control and the cyanos were showing increased O2. How much of a change in O2 are you seeing? Where did the water come from that you added? Water can contain some dissolved O2 that could be released. I wouldn't say it was caused by a reaction of CO2 with water. CO2 will dissolve in water to form carbonic acid but this does not produce O2.

I think you may have a problem with too much CO2 in the jar as this could acidify the water so much that the cyanos will die or at least stop growing and photosynthesizing which would defeat your terraforming process. You could add a buffer compound to the water to maintain the pH. Have you measured the pH of the water in the jars?

What species of cyanobacteria are you using? Were you going to add a growth medium for them? Here's a long list of media that are used for cyanos and I have no idea which would be appropriate for your bacteria: http://www-cyanosite.bio.purdue.edu/med ... media.html
If the medium contains a buffer then that would prevent the pH from dropping too low from the CO2. Also, the extra nutrients will make the cyanos grow better. You do want them to photosynthesize, however, so they make O2. Do you have lights next to the jars to mimic sunlight?

Let us know when you check the pH and if the O2 levels are rising with the cyanos as they grow and photosynthesize. Be sure to take photos at various times to show any changes in the color of the medium as they grow.

Good luck!

Sybee

[Lorenzo192]

Thank you for your help. I wanted to do this project because I was interested in terraforming Mars, but I didn’t have much direction and now I’m in a jam. The bacteria I used are nostoc and oscillatoria. The nostoc is a very dark green, almost black, and oscillatoria is a light green. It is impossible to tell what color they are now because the Martian soil made the water a reddish brown color and I put them both in the test bottle. I went from using the fish tank to using two canning jars with one jar being the control as you suggested. I tested the jars before hand and found that they could keep out oxygen pretty well with the percent going from 0% to a little over 1% after a few days. However, when I put the soil and water in the jars and the bacteria in the test jar, the oxygen level went from 0 to 2.95% (test) and 3.80% (control). There was more water in the control jar and that made me think it had something to do with the water. I have not tested the Ph yet because I need to get a Ph test kit.
When I put the CO2 in I found that it worked best when I crumbled the dry ice into small pieces (I used insulated gloves). This got the oxygen level down faster and further than putting small chunks of dry ice in a cup of water and letting the CO2 flow into the jar. When I did this some small dry ice chunks got into the water. This was probably bad for the bacteria as I was not thinking about carbonate acid which may have killed the bacteria. Fortunately, I did not use all of the bacteria and I have more than half left.
What I propose doing is starting over by emptying out the jars, put equal amounts of soil and water in each, put in most of the bacteria in the test, and put in the CO2 by the gas flow in from a container of dry ice and water or use a strainer with chunks of dry ice in it holding it over the jar. I would only let the oxygen level get down to somewhere around 8%. What are your suggestions? Do I need to put a Ph buffer in the test jar and what kind?

[SciB]

Thanks for the details! That's what we really need to answer your questions in the best way since we can't be there in person to see what you are doing.

Now i understand your problems a lot better.

So basically you are just using some kind of soil and water, right? I hope you are not planning to use chlorinated tapwater as this might kill the cyanos or at least slow them down a lot. The soil could provide some nutrients but if you want the bacteria to grow best they will need a source of nitrate and the water will have to be buffered to keep the pH around 8 which is what they prefer.

I tried to find the culture medium for oscillatoria and i found it mentioned in this paper: http://www.google.com/url?sa=t&rct=j&q= ... 8268,d.eXY

However, when i looked at the methods section i found that they did not give the recipe for the culture medium which is called Z8! Instead, they had referenced two other papers, neither of which i could get on PubMed. If you do a search for 'Z8 culture medium' you may be able to find it eventually. I know that it contains nitrate and orthophosphate but i don't know the concentrations. If you can't find Z8, I would just do a search for a generic minimal salts bacterial medium which will probably work ok--certainly better than water alone.

What are you using as a light source? The cyanos need artificial light with a spectrum that is close to sunlight if you have them inside which I would recommend since they need a temperature of at least 20C to grow. If you have a sunny south window that would be good for algae.

I think your idea to only reduce the O2 level to about 8% is good and flowing the CO2 into the two jars is definitely better than putting bits of dry ice into the water. You are going to have to use a buffer in the water because otherwise the dissolved CO2 is going to make it too acidic for the cyanos. The minimal salts medium uses phosphate as the buffer usually and when you find the recipe for Z8 it should tell you how to adjust the pH to about 8.0 which is optimal for oscillatoria and nostoc. It would really help if you could work in a lab with basic chemicals available and a pH meter.

This is really an interesting project and I hope you can make it all work. We'll be here to help.

Good luck!

Sybee