Phytoremediation

[Elan_Wilson1]

Hello, I am a student who is entering the ninth grade and I really want to win at science fair. I'm thinking about conducting a project comparing two commonly used chelators, EDTA and EDDS, in terms of phytoremediation. I really need help with coming up with a scientific procedure and an idea of necessary materials.

[SciB]

Hi Elan,

Your project is very timely as pollution with heavy metals such as cadmium and lead is a worldwide health problem. Using plants to concentrate the metals and then harvesting the plants is a great green solution to this problem and one that could be done in poorer countries where the need is greatest. Metal chelators can be used to enhance the uptake of the metals by certain plants.

I know about EDTA but EDDS was unfamiliar, so I did some reading and I see that EDDS is being considered because it is broken down in the environment and does not persist and cause other problems as EDTA can.

I noticed in your description of your project that you want to use phytoremediation with chelators to decrease heavy metal pollution in bodies of water. Removing metals from a water supply is definitely important, but I am wondering how you were planning to do the comparison of EDTA with EDDS. The phytoremediation projects I am familiar with all target soil pollution using plants such as sunflowers that grow on land.

You could use a water plant like water hyacinths since they are very easy to grow and easy to harvest. If you live in a southern state as I do, water hyacinths can easily be found in any lake or pond. Water lettuce would be another easily found and grown choice.

Which metals were you going to use for comparing your chelators and how were you going to detect them in the plant? I would not recommend cadmium as it is very toxic. Lead is also toxic but not as much. Do you have access to a chemistry lab at a nearby college or university? They could help you with the method for detection.
In terms of materials, I would think you could use plastic bins as growth containers. You need to keep the lighting, temperature, volume of water, number and size of plants, and the concentration of the metal ions in the water all constant so that the only variable is the chelator you add.

I found a paper about comparing EDTA and EDDS in doing phytoremediation and attached it. It’s from a scientific journal called “Chemosphere” and is very technical so if you get stuck reading it please post back and I will help.

Good luck! I really like this project and I know you can bring it off successfully with some careful planning.

Best wishes,

SciB

[Elan_Wilson1]

Thank you........as an answer to your question....i will have access to a chemistry lab and I plan to use lead concentrations in my water samples, but I am not very familiar with phytoremediation itself and need to know a precise process in which I could conduct my experiment.Also, I was wondering if there was any place where I could possibly purchase the chelators for my project.

[SciB]

Hi Elan,
Since you have permission to work in a chemistry lab, you could ask the people there if they have the chemicals you need and if you could use them. Most labs have EDTA, but they may have to order EDDS. I usually order chemicals from Sigma-Aldrich, http://www.sigmaaldrich.com/united-states.html. I checked and they do have EDDS: http://www.sigmaaldrich.com/catalog/sea ... partialmax

Also on the Sigma page for EDDS there was an abstract from a paper on using EDDS with sunflowers for phytoremediation of lead in soil:

Effects of indole-3-acetic acid (IAA) on sunflower growth and heavy metal uptake in combination with ethylene diamine disuccinic acid (EDDS)
Erika E Fässler, Michael W MW Evangelou, Brett H BH Robinson, Rainer R Schulin

Abstract
The use of plants for phytoextraction of heavy metals from contaminated soil is limited by the ability of the plants to grow on these soils and take up the target metals, as well as by the availability of the metals for plant uptake in the soil solution. The hypotheses of this study were that the growth-promoting phytohormone auxin (indole-3-acetic acid, IAA) can alleviate toxic effects of metals on plants and increase metal phytoextraction in combination with the biodegradable chelating agent ethylene diamine disuccinic acid (EDDS). To test these hypotheses we performed two sets of experiments with sunflowers (Helianthusannuus L.) in hydroponic solution. In the first set of experiments, five IAA concentrations (0, 10^-^1^2, 10^-^1^1, 10^-^1^0, 10^-^9M) were applied in combination with Pb (2.5@mM) or Zn (15@mM). In the second set of experiments we applied combinations of IAA (0 or 10^-^1^0M) and EDDS (0 or 500@mM) to Pb or Zn-stressed sunflowers. Root and shoot growth of metal-stressed plants were most effectively increased with 10^-^1^0M IAA, and also the extraction of both metals was significantly increased at this treatment level. IAA reduced the negative metal effects, such as reduced shoot and root dry weight, root length, root volume and root surface area. EDDS significantly decreased metal uptake by the plants, thus reducing metal stress and promoting plant growth. The combined application of IAA with EDDS significantly increased Zn uptake in comparison to EDDS only treated plants. The experiments indicate that IAA can alleviate toxic effects of Pb and Zn on plant root and shoot growth and can in combination with chelants such as EDDS increase the phytoextraction potential of these plants.

Your idea of using water plants to remove lead from a pond or stream is good because most of the studies i see are about using plants to concentrate and remove heavy metals from polluted soil. You asked for more info on phytoremediation, so i attached a paper that describes how it works and what it can do. Here's a link to a site that explains the difference between using plants for extracting Pb from soil vs water: http://www.webapps.cee.vt.edu/ewr/envir ... index.html

Let me see if i understand your project correctly. You want to see how well a water plant such as water hyacinth, water lettuce or duckweed can take up and hold lead from water to remove this pollutant from the environment. To test this you would set up several containers of water containing various concentrations of lead from zero to some higher value that would reflect actual pollution. If you get water plants from a pond, i would also get the water from the pond to use in your experiments because you know the plants will grow in it.

Each container would contain the same volume of water and get the same number, size and type of plants and these would be allowed to grow for a certain length of time in the sun. The plants would then be collected, dried and analyzed for lead to see how much they were able to remove. So, your dependent variable would be the amount of lead per gram of dried plant material and your independent variables could be how much lead you added to the water, how long you grew the plants or how many plants you started with.

Is that what you were thinking of doing? Please post back so i can give you more specific help. This project will take a lot of work and planning but i think it could be a winner and provide some useful information about cleaning up the environment naturally.

Best wishes,

SciB

[Elan_Wilson1]

Yes....your interpretation of my project idea was very accurate. However, instead of selecting my plants from the environment, I plan to actually purchase them, mature them, then place them in a chamber with contaminated water(with lead or other heavy metals).I was also planning to( instead of collecting lead concentrations from the plants themselves) measure the remaining concentration of lead in the water, but I am not sure how to do that.Finally, I was also unsure how to set up the plants in a way that they do not die from the water, I was wondering how I should specifically apply the chelators to the experiment(I know what they are....but I am unsure of their actual application), and I was wondering how exactly I would detect pollution levels in the water.

[SciB]

Hi Elan,
The reason I suggested using wild plants is that you would have greater control over choosing the size and number appropriate for your experiments. Which plant are you thinking of using? Some plants are better than others at concentrating metals from water.
Your idea to measure the change in concentration of metal ions in the water over time should work although deciding on the volume of water to use for a certain amount of plant material might be a little tricky. Did you plan to weigh the plants and try to get exactly the same amount in each test chamber? This might be easier with a small plant like duckweed (Lemna minor) whose total weight could be more easily adjusted than that of a large plant like a water lily or hyacinth. I have attached a reference for the use of duckweed to remove lead, zinc and copper from ponds. Don’t worry about understanding all the technical details. There is some useful general information and I want you to see how scientists are using plants to do experiments like what you are thinking of.
Lead and zinc are two fairly safe metals when used properly in the lab, but do handle them as you would any toxic substance—wear gloves, goggles and lab apron. Since you will be working in a lab, there won’t be any problems with disposal. You don’t want to pour your solutions down the drain so they end up in the environment!
You will need to set up your experiments to test a range of metal concentrations and you will have to check the scientific literature to decide what concentrations to use. You should have a zero concentration as a control, then increase it stepwise until you reach a level that would be considered very polluted. I don’t know how sensitive a plant such as duckweed is to metal levels in the water, but the references should give you an idea.
Finally, you need a way to measure lead and zinc with sufficient sensitivity to detect the changes in concentration in the water caused by metal uptake by the plants. If you have access to a chemistry lab, they should have an instrument called an atomic absorption spectrometer that is the most common way to measure metal ion concentrations in solution. Here’s some links that will give you more information on how this is done:
http://enterprise.astm.org/filtrexx40.c ... /D3559.htm
http://enterprise.astm.org/filtrexx40.c ... /D1691.htm
I think I have given you an awful lot to think about, so I will stop here. Get back to us when you have more questions and we will help some more.
Best wishes,
SciB

[Elan_Wilson1]

Thank you for the sources, they are all very in depth and they actually helped me begin the foundation of my project.Right now I am about 70% through my proper research paper, and the only main subjects that I need to cover are the procedures and materials.I have recently discovered a link of another project that would give you a good idea on how I plan to organize my procedure and experimentation. I plan to construct a similar device as shown, but I am also unsure if I should focus on extracting the lead content from a controlled source of water, or a controlled source of soil( I also need an idea concerning what unit of measurement I should use to describe the lead content).I just want to get the application right to impress my judges.I also have decided to use Indian mustard plants due to their heavy application in extracting lead content from water and soil, and I plan to apply EDTA and EDDS to the soil of my plants in steady increasing intervals over time(ex/ in the first experiment I will apply 0.5g of EDTA and EDDS while in the second I will apply 1g and so forth).I also need a professional opinion if this project is capable of winning or not, and some helpful tips to make it above and beyond.

https://www.googlesciencefair.com/en/pr ... XpldHVEQQw

[SciB]

Hi Elan,

Congratulations! You have your project well on the way to success--but i see what you are looking for is that extra tweak that makes it a prize winner.

My feeling is that removing lead from a volume of water using your mustard plants growing on some kind of support in the water is the best way to do the test. How to remove heavy metal contaminants from waste water is a common problem and i think it would be much easier to measure lead in a water sample than to try and extract it from soil.

The rate of decrease in lead ions in your water container is the dependent variable, but what will you use as your independent variable? The number or size of plants per volume of water?

Companies or municipal governments are looking for ways that are cheap and effective to produce clean water and phytoremediation seems like a great solution since a plant like mustard can easily and quickly be grown from seed. I was thinking, if there is some way that you could get actual water from a local water treatment plant it would bring your project to life. It would have to be water that has already been treated so that there are no coliforms or other potentially harmful agents--like 'gray water' that people use to water their lawns but don't drink. Some cities have connections to this source for lawn watering and if you could get that water, you could spike it with lead (assuming that it doesn't already have high lead content) and use that for your test.

Let me know your thoughts about this and we can try to help you create a really outstanding project.

Best wishes,

SciB

[Elan_Wilson1]

To elaborate, as an independant variable, I plan to use the quantity of lead in the water as an independant variable and I was thinking of creating some flotation device that suspeded the plants above the water while not letting them be completely submerged. To collect data, I plan to take samples from the water and inspect them with a specrtometer since lead tends to give off some sort of light(inference). I also really like the idea of getting the water from a water treatment plant, and I was wondering, from your opinion, what you think should be my controls in the experiment and what would be a reasonable amount of time to grow the plants before they are exposed to the contaminated water.

[SciB]

Hi Elan,
I don’t know how hard it would be to get wastewater from a water treatment plant in your city, but it would make your project more relevant to real life. As I understand water treatment, it involves several stages—settling, aeration, filtering, etc—and your project to remove heavy metals through the use of plants would be one of those stages.

Why don’t you call the nearest water treatment plant to where you live and explain to them that you are a student who is doing a science fair project to develop a phytoremediation method for reducing heavy metal contamination in wastewater. I’m betting they will help you and be just as excited as you are about this project. What you want is a source of water that has been treated but is not yet drinkable so you can use your mustard plants-plus-chelator to clean up the metals that are still in it before it goes to the consumer.

Did you state your hypothesis? I don’t remember seeing it. Also, are you planning to use the number of plants for a given water volume as one of the independent variables? Thinking about this as a water engineer, one of the first questions they would naturally ask is how many plants are needed to clean up a certain volume of water. Addition of a chelator is supposed to increase the rate of heavy metal removal, but I don’t know if it also increases the amount of metal removed by a single plant.

Your other independent variable would be the amount of EDTA or EDDS you add to the water and you will have to do some checking in the scientific literature to determine an appropriate concentration. Also, you need to decide what the concentration of lead should be to represent a real-world situation. Again, you will have to google it or use pubmed to find out what concentrations of lead are considered minor, moderate and severe pollution. Be sure to ask your mentor in the lab what is the lowest concentration of lead ions that can be detected by the instrument that you will be using. You want to be sure to be able to detect the changes in lead concentration in the water.

Can you explain to me how you plan to get the mustard plants accustomed to growing in water before you put them in the experimental container? I’m assuming you are going to buy plants, wash the soil off the roots and support them on a styrofoam raft in a hydroponic solution to get them used to growing that way. I can’t say how long it will take before they are ready to use in your experiment, but be sure to treat all of the plants the same way. You don’t have to vary the number of plants per volume of water as one of your variables—I just thought that would be a good way to see if there is a linear relationship—twice the number of plants equals twice the amount of lead removed, three times the number of plants removes three times the lead, and so on.

As for controls, an obvious one is NO plants. You will be comparing the amount of lead removed by plants with and without a chelator, so the minus chelator condition will be your control there. There is one more control that you would do if you were publishing these results and that would be to compare the growth of the plants in wastewater with and without lead and in distilled water with hydroponic nutrients. Maybe the wastewater contains some chemicals or ions that will inhibit the growth of the plants or even kill them. You wouldn’t know that unless you compared their growth in a hydroponic solution to growth in wastewater. If you are able to get water from the treatment plant, you might want to try growing the plants in it first before you start the experiments just to make sure they will survive.

How long you grow the plants before putting them in the water with the lead is something I don’t know, as is also how long you let the plants grow in the wastewater before analyzing the water for lead. If you decide not to vary the number of plants per volume of water, then what you could do is take samples of the water at 2, 4, 8 days, 2 ,3, 4 weeks, etc., and measure the lead content. This will show you how fast the plants take up the lead and you can compare the rate of lead absorption with and without chelator over time.

Whew! That was a lot of writing, and I hope I haven’t made things more confusing by including too much information. Think about your hypothesis and which variables will give you the most interesting and useful data to present. I will continue to help you with the experimental details and really look forward to your results.

Best wishes,

SciB

[Elan_Wilson1]

I have now gathered all of my materials and have composed my experimental procedure completely, but I am having difficulty finding Indian mustard seeds or plants.

[deleted-140482]

You can buy Brassica rapa (Wisconsin fast plant, or one kind of mustard plant) at Carolina Biological (http://www.carolina.com/wfp-mutant-seed ... /158804.pr). I assume these are the plants you are looking for. I'm not sure of somewhere you could buy the plants partially grown, but they do grow and mature in a very short period of time, so hopefully the seeds will be good enough. The seeds are also available to Amazon, but I can't vouch for their quality.

[SciB]

Hi Elan,

Happy to hear from you again. I wondered how you made out with your project since July.

Indian mustard, Brassica juncea, is a good choice for phytoremediation because the plants are known to accumulate heavy metals such as lead and cadmium, more-so than other plants. As JMP suggests, I would raise your own plants from seed because they germinate and grow fast. I did a search for Brassica juncea for sprouting and found one company that sells seed: http://www.johnnyseeds.com/p-8581-orien ... ng-og.aspx

The seeds are cheap and 1 oz will give you more than enough for all your experiments.

Keep me posted on your experimental set up. How are you going to control for the amount of plant material in each test sample? You need to have equal amounts if you are going to compare them. My hypothesis would be: The more plants, the more lead will be removed from a given volume of water. That's why I suggested before that you make the number of plants per volume of water your independent variable. Do you have an accurate digital scale that is sensitive to tenths of a gram? I think you should carefully weigh and record the amount of plant material you put into each test container and at the end of the test period, you should weigh it again. Knowing the weights you will be able to normalize your changes in lead concentration to a specific weight of mustard plant. Comparing the plant's weight before and after the test period will tell you if the plant's growth is inhibited by the lead in the water.

How did you plan on suspending the plants in the water? What volume of water will you use? How many plants per test? There are a lot of details to work out!

Let us know when you have more questions. By the way, when is the due date for your project?

Best wishes,

Sybee