Managing the Nitrogen Cycle

[intelius]

This year I would like to do a project on the following engineering grand challenge (these questions summarize the nitrogen cycle challenge on the engineering challenges website http://www.engineeringchallenges.org/ch ... rogen.aspx):

Managing the Nitrogen Cycle (ie. how can we increase denitrification? how do we control the release of nitrogen oxides produced in high-temperature burning of fuels? how can we improve the efficiency of various human activities related to nitrogen?)

I would like to answer one of these questions through my science project. I'm not exactly sure how to go about it. Any brainstorming would definitely help.

[SciB]

Hi Intelius and welcome to Scibuddies. Your potential science project is in a great environmentally important area. We have only one planet to live on so far, and if we mess it up we may go the way of the dinosaurs.

The application of huge amounts of nitrogen fertilizers has allowed farmers in the U.S. to increase crop yields per acre but not all of that fertilizer is used by plants. The excess may end up in waterways and areas such as the northern Gulf of Mexico where it can stimulate algae blooms that decompose and deplete the oxygen in the water killing fish and other marine creatures (http://water.epa.gov/type/watersheds/na ... n/zone.cfm). There is a natural nitrogen cycle in which decomposers and nitrogen-fixing bacteria produce ammonium or nitrate ions that plants can use and the excess is converted by denitrifying bacteria back to a gas—nitrous oxide or nitrogen (https://en.wikipedia.org/wiki/Denitrification). By overuse of nitrogen fertilizers, humans have overloaded the system and created an imbalance that needs to be fixed before any more damage is done. That is the challenge referred to in your link.

There are many possible projects within this challenge. You mentioned denitrification which is a biochemical process performed by certain types of soil microbe that removes nitrate by converting it to another form such as nitrous oxide, N2O, or ultimately to molecular nitrogen, N2 (https://en.wikipedia.org/wiki/Denitrifying_bacteria). If the runoff from nitrogen fertilizer applied to crop lands could be treated by denitrifying bacteria then there would be less to pollute the rivers. Is this a project you might be interested in?

Working with bacteria, especially the ones that need to be grown without oxygen can be difficult. The complexity of this kind of project would require that you have access to a microbiology lab with apparatus for growing and monitoring specific kinds of bacteria. What resources do you have access to for your project?

Denitrifying bacteria occur naturally in certain environments where the amount of oxygen is low such as boggy soils or the muck at the bottom of a pond. You might be able to create a habitat that promotes the growth and activity of denitrifiers and use it to remove excess nitrate in runoff. Measuring the relative changes in nitrate in an environmental situation as opposed to a lab can be very difficult, however. On the other hand, an environmental solution might be more likely to succeed in actually reducing the nitrate excess in a real-world situation.

Let us know more specifically what your interests and resources are and we can better help you to pose a good hypothesis and design experiments to test it.

Good luck!

Sybee

[intelius]

You might be able to create a habitat that promotes the growth and activity of denitrifiers and use it to remove excess nitrate in runoff

Sybee,

I am actually quite interested in this idea, but I do not have access to a lab. Is it possible to do a project of this complexity at home? What would I need?

[SciB]

Hi Intelius,

I have never tried to experiment with bacteria that live in the absence of oxygen. I do know that they can be found a few centimeters down in the mud at the bottom of a pond. I think you could use this mud as your source of denitrifying bacteria. I don't know if this will work or not, but you can find out by doing the experiment. Hopefully one of the other experts who has some experience working with anoxic bacteria can offer more suggestions.

Get six 2-liter bottles and in three put pond water with about 4 cm of mud in the bottom and in the other three put only pond water. Let the bottles sit undisturbed for 24 hours in a darkened area. These are your test 'ponds'.

Your hypothesis might be: Runoff from farms or feedlots can be treated by anaerobic bacteria to decompose the nitrate. To test the hypothesis you add a small amount of fertilizer containing nitrate to the ponds, measure the nitrate level in each one at time zero then measure it every day until the nitrate level stops changing.

The bacteria in the mud should be able to decompose the nitrate while little decomposition will take place in the bottles without mud. You can buy nitrate test strips or solutions at pet stores or aquarium suppliers. Here's one source, but you can find many more if you do a google search: http://www.petco.com/product/14733/API- ... riums.aspx

Do some more reading about the kinds of bacteria that grow in pond muck. Here are some possible links:
http://water.me.vccs.edu/courses/env149/lesson5.htm
http://www.dairyingfortomorrow.com/uplo ... 0ponds.pdf
https://en.wikipedia.org/wiki/Anaerobic_lagoon

I'm sure you will have more questions so feel free to post again and we will try to get your project on track for success.

Good luck!

Sybee

[intelius]

Runoff from farms or feedlots can be treated by anaerobic bacteria to decompose the nitrate.

Hmm...I feel like this type of project has been done already as we already know that the denitrifying bacteria will remove nitrogen from water. (see:
http://www.scientificamerican.com/artic ... s-streams/
"Denitrification is the only process that we know for sure removes nitrogen from water,")

However, I think your idea from earlier about creating habitats that would promote the growth and activity of the denitrifiers would make a great science fair project. We could add on to the method you had suggested earlier by using mud as our source of denitrifiers and the bottles as the habitats. We would have yet another bottle with some mechanism to increase the growth of the denitrifying bacteria I suppose? I'm just a bit lost. I hope you can guide me through this. I just want to add on to your idea to make it more original. If we do successfully denitrify the water in the bottles, how would we implement this to dentrify runoff in waterways? I think if we could figure out a way to promote the growth of the denitrifiers it would be best if we focused this project on implementing these methods (please do give feedback!).

I also had a few other questions that I know you said you weren't quite sure about:
1. Where can this dentrifying bacteria be found? Is it surely found in deep mud from a pond? Is there a way to test the presence of denitrifying bacteria?
2. What methods can we use to promote the growth and activity of the denitrifiers once we get a sample of them?

[intelius]

After re-reading my last response I see that it was a bit confusing. Basically this is what I would like to know:
1. How do we promote the growth and activity of denitrifiers? (I understand that I will have to do some research on this but I am not sure where to start)
2. Once we find a way to promote the growth of the denitrifiers how do we implement this method to remove excess nitrate from runoff in waterways?

For question #2 I don't know if this should be part of the project or if I should just solely focus on promoting the growth of the denitrifiers. Could you give me some pointers on this? (haha this seems to be turning into 2 different projects: a science project and an engineering project )

[SciB]

I know what you mean. The best projects and research involve multidisciplinary attacks involving biologists, engineers, chemists, physicists, statisticians, etc.

You are right that a lot of science projects have already been done and the answers are already known, so they aren't that interesting. So, how can you make your project more interesting? I think your idea about boosting the denitrifiers is a good one but I'm not sure how to do that.

Bacteria in general require three nutrients--nitrogen (nitrate in this case), phosphorus (from the fertilizer) and a carbohydrate. The last item may be your best shot for increasing the denitrifying activity. When I grow bacteria in liquid medium I use glucose as the carbon source. You need to see if you can find a recipe for growth medium for denitrifying bacteria so you can see what they use as carbon source and how much per liter. Adding it to the pond water will stimulate growth of the bacteria and hopefully also of their denitrifying activity.

You can always test for the presence and activity of denitrifiers by simply adding nitrate to the pond water, measuring the concentration with the nitrate test strips, then measuring again after a day or so. If there are active denitrifying bacteria [lets abbreviate them to DNB] then the nitrate level will decrease with time.

Does this help? I wish some of the others on this forum would chime in with suggestions. I'm no expert in working with anaerobic bacteria and I do want your experiments to succeed. Do some more reading about DNB and maybe you will find some other ideas for making your project unique. Hey..lots of scientists got started in their garage or basement with just an idea and minimal supplies. The challenge is what makes it fun!

Good luck!

Sybee

[intelius]

You have definitely helped Sybee! I was doing some research and I ran across this science project which uses denitrifying bacteria to reduce nitrate levels in water and it uses Nitrobacter and Nitrosomonas bacteria to do this. Aren't these two nitrifying bacteria? I was just curious about this (here's the link: http://www.all-science-fair-projects.co ... 122_2.html) I'm not sure what the difference is between denitrifying and nitrifying bactera

I will see if I can obtain the bacteria in some form at a pond supply store.

I also found some articles which explored optimal conditions for the growth of denitrying bacteria and I will see if I can implement these somehow:
-http://www.rpi.edu/dept/chem-eng/Biotec ... ation.html
"Substrate Dependence: Denitrifying bacteria are highly sensitive to the substrate available for their use. Easily degraded materials such as methanol and acetic acid give the highest rates of denitrification. Raw wastewater does not yield such a high degradation rate, but it is significantly higher than what is observed with non organic carbon sources.
Temperature Dependence: Rates of denitrification reactions increase with temperature in a manor similar to what rates in nitrification reactions. They are not as limited in range though, as some bacteria can continue to carry out the processes at temperatures up to 60°C. These bacteria are relatively rare, and as a result reactions are not usually carried out at such high temperatures despite the expected increases in reaction rate. Click Here to see the effect temperate has on reaction rates for several different substrates.

Oxygen Inhibition: Oxygen inhibits denitrification by providing a better electron acceptor for metabolism. When oxygen is present, the microorganisms will use oxygen instead of nitrate. The concentration of oxygen of importance is the concentration at the cell's surface, which may be significantly different than the bulk concentration.

pH Dependence: The optimum pH for denitrification is in the 7-9 range, with rates falling off sharply outside this region (see figure). At lower pH the generation of N2O and NO is increaced. "

-http://www.wastewaterhandbook.com/docum ... isites.pdf
"The necessary conditions for the denitrification process to develop in an activated
sludge process can be summarised as:
(1) Presence of a facultative bacterial mass;
(2) Presence of nitrate and absence of DO in the mixed liquor (i.e. an anoxic environment);
(3) Suitable environmental conditions for bacterial growth;
(4) Presence of an electron donor (nitrate reductor). "


If I tested these however I still feel that my project wouldn't be original. Is it the best route to go to search for new ways to promote the growth of denitrifying bacteria or somehow implement these methods to reduce the nitrate in runoff? Both of these objectives seem a little far-fetched to me as I don't know how to approach either

[SciB]

Developing a doable original research project is a hard job even for an experienced professional researcher. I've gone down several dead ends that sounded great at the time but didn't work. You are up against a big hurdle and that is lack of lab resources. Your experiments are limited to what you can do at home or in your high school lab .

What do you see as the practical purpose of your project? I was thinking of a pond that a farmer could dig that would catch the runoff from his field or feedlot and eliminate the nitrate pollution. The denitrifying bacteria could be stimulated by the addition of some easily renewed carbon source--but what that would be I don't know.

The other factors that you mentioned like pH, which can be controlled, and temperature which would be difficult, are secondary to making the bacteria efficient enough to eliminate all the nitrate in the runoff. Your experiment could tell you how much nitrate a small sample of a pond could process and you could attempt to calculate from that what size of treatment pond that would be needed for a farm or feedlot of a certain size. That is something that I don't think has been done before. With a calculation like that you could prove that your hypothesis would work when scaled up. That is something a funding agency is always asking--is your process scalable?

You are right that Nitrosomonas is a nitrate-producing bacterium. In nature, these bacteria work with DNB. The nitrifying bacteria convert ammonia, a common pollutant in farm runoff, to nitrite and then nitrate. The DNB then convert the nitrate into nitrous oxide or nitrogen to finish the process.

Do you have a pond or lake in your area where you can get mud that hopefully contains DNB? I would try to do this rather than buy bacteria because pure cultures can cost over a hundred dollars. Get some pond water with mud, buy a nitrate test kit and some cheap fertilizer and see if your 'pond' will eliminate nitrate. If it works then you have your testable model system. If not then you may have to go to plan B. Is there a waste water treatment plant in your area? They have ponds that contain anaerobic bacteria and if you tell them you are a high school student doing a science project that has as its goal the reduction of nitrate runoff they may be willing to give you some 'mud' containing active DNB.

Let us know what you decide and we'll help you with the planning, design and experiments.

Good luck!

Sybee

[intelius]

I was thinking of a pond that a farmer could dig that would catch the runoff from his field or feedlot and eliminate the nitrate pollution

Ohh I now understand what you were getting at earlier,

Your experiment could tell you how much nitrate a small sample of a pond could process and you could attempt to calculate from that what size of treatment pond that would be needed for a farm or feedlot of a certain size.

I really like this idea! It seems more doable since I do not have access to a lab. I would need some help with the calculations though!

Is there a waste water treatment plant in your area? They have ponds that contain anaerobic bacteria and if you tell them you are a high school student doing a science project that has as its goal the reduction of nitrate runoff they may be willing to give you some 'mud' containing active DNB.

There are actually quite a few very close by. I will contact them today and keep you posted.

Once again, thank you for helping me on this I really appreciate it!

[SciB]

You are most welcome!

Read about waste water treatment before you talk to the people at a plant so they see that you are a serious student who has taken the time to understand about denitrifying bacteria. I hope they can help you but they have a lot of legal constraints on what they can do, so they may say they would like to help but can't. They are a good source of information, however, and a personal tour of their plant would be awesome! You can ask if they have public tours.

I still think you should try to use mud from a natural lake or pond if there's one near enough to where you live. in fact, you could try natural mud vs waste water treatment plant 'mud' and see which one is better at eliminating nitrate.

Good luck and keep us updated on your progress.

Sybee

[intelius]

Hmm... I was trying to start my formal report and I couldn't figure out what question this project addresses. Is this really an engineering project? Or is the question, how large of a pond does a farm need to reduce the nitrogen pollution in its run-off?


Intelius

[SciB]

Your project has to do with biology not engineering. Leave that part to the people who dig the pond. What you are doing is figuring out what size pond they need to make. You are doing this by using your test 'ponds ' to measure how much nitrate they can convert. Since you know the size of your mud all you have to do is multiply it by how much nitrate the farmer has in his run-off. Does that sound reasonable?

Let us know if you have more questions.

Good luck!

Sybee