The purpose of this lab is to illustrate the importance of monitoring the water quality of effluent (wastewater outflow) and its effects on aquatic organisms.
|Annie McEntyre||Physical Science
EEn.2.3.2 Explain how ground water and surface water interact.
EEn.2.4.1 Evaluate human influences on freshwater availability.
EEn.2.4.2 Evaluate human influences on water quality in North Carolina’s river basins, wetlands and tidal environments.
- How will soluble zinc affect an aquatic ecosystem?
- Will all of the organisms be affect similarly or different?
- Why do industries need to ensure they are meeting regulatory requirements when working with potentially toxic chemicals?
- Science and Engineering Practices
- Analyzing and interpreting data
- Construction explanations and designing solutions
- Obtaining, evaluating, and communicating information
Rationale: “In order to comply with the Clean Water Act, Befesa Zinc Metal’s water discharge permit requires quarterly Whole Effluent Toxicity (WET) testing. Samples of AZP’s effluent are sent to ETT Lab in Greer, SC for WET test analysis, which measures its effects on specific test organisms’ ability to survive, grow and reproduce. The purpose of this lab is to illustrate the importance of monitoring the quality of the effluent and its effects on aquatic organisms.”
Objective: One way to test for the presence of toxic compounds in a water sample is a bioassay. In a bioassay, a living organism serves as a detector for toxins—the same way canaries were used in coal mines to detect invisible toxic gases. In this project, water fleas (Daphnia magna), a freshwater crustacean, are used in a bioassay to monitor water quality.
The water flea (Daphnia magna) was used as a sensitive indicator for assessing the toxicity due to synthetic detergents. Acute and chronic toxicity of detergents to the water flea was studied under laboratory conditions by following the median tolerance limit (TLM) at 48 hr and the rate of survival.
You might know that lead can be toxic and that you can get lead poisoning from eating or inhaling old paint dust. Lead is called heavy metal, and other sources of heavy metals can be toxic, too. Silver, copper, mercury, nickel, cadmium, arsenic, and chromium are all heavy metals that can be toxic in certain environments and at certain concentrations. In this experiment, find out if one common heavy metal, copper, can be toxic to an aquatic environment.
In this experiment, students will test the effects of the heavy metal copper (Cu) on an aquatic environment containing Daphnia, snails, and plants.
You might know that lead can be toxic and that you can get lead poisoning from eating or inhaling old paint dust. Lead is a heavy metal, and other sources of heavy metals can be toxic, too. Silver, copper, mercury, nickel, cadmium, arsenic, and chromium are all heavy metals that can be toxic in certain environments.
“Toxic metals, including “heavy metals,” are individual metals and metal compounds that have been shown to negatively affect people’s health. In very small amounts many of these metals are necessary to support life. However, in larger amounts, they become toxic. They may build up in biological systems and become a significant health hazard.” (OSHA, 2004)
In this project, you will determine if one common heavy metal, copper (Cu), can be toxic to an aquatic environment. You will use copper because it is one of the heavy metals that is easy to find and is not very toxic to humans. You will use copper sulfate as a source of copper that is soluble, meaning it will dissolve, or turn into a solution when mixed with water.
The amount of an ion in solution is often measured in parts per million (ppm). This means that if there is 1 ppm ion in the solution, then there is 1 milligram (mg) of the ion present in each liter (L) of solution. Heavy metals release free ions in solutions and can cause an effect at very low doses.
In this project, a range of copper concentrations should be tested from 0ppm (no copper sulfate added) to 0.4ppm (with increments of 0.1ppm). These are very low concentrations, so it takes some careful planning to make the solutions. We will show you here how to make a copper ion solution with the lowest ion concentration you will be testing: 0.1ppm.
Making a copper sulfate solution with a concentration of 0.1ppm means that you will need 0.1mg of the copper sulfate in one liter of water. However, it is very difficult to measure 0.1mg. To give some perspective, a new U.S. dollar bill weighs 1g. If a dollar bill were cut into 1,000 pieces, each piece would weigh 1mg. If each of those pieces were further cut into ten pieces, cutting the dollar bill into a total of 10,000 pieces, then each piece would weigh 0.1mg. This is the amount of copper sulfate needed in one liter of water to make a 0.1ppm solution. It should now be clear why it would be hard to accurately weigh out something this small!
This is why we need another way to make a copper solution with a concentration of 0.1ppm. Specifically, you will be making a solution that is at a much higher concentration than you will be testing, called a stock solution, and then you will be diluting the stock solution to create the concentrations you want to test.
First, 1 gram (g) of copper sulfate will be weighed out and mixed with 2 liters (L) of water. This can be written as
1g copper sulfate
This means that 0.5g (or 500mg) of copper sulfate will be dissolved into 1L of water, creating a copper solution with a concentration of 500ppm. How are we going to decrease the concentration even more? We can take one drop of this solution and dilute it with more water. There are about 20 drops of water in 1 milliliter (mL). Knowing this, we can figure out how much copper sulfate will be in one drop of water using the following calculation:
1g copper sulfate x 1L water x 1mL water = 0.000025g copper sulfate
2L water 1000mL water 20mL water 1 drop water
This tells us that there are 0.000025 grams (which equals 0.025mg) of copper sulfate in one drop of water. This means that four drops of water have 0.1mg of copper sulfate. Because a 0.1ppm solution is 0.1mg dissolved in 1L of water, if we mix four drops of the 500 ppm copper sulfate solution (0.1mg) with 1L of water, we will have a copper sulfate solution with a concentration of 0.1ppm.
The 0.1ppm copper sulfate solution will be the lowest concentration of copper that you will test in your experimental aquatic environments. You will add increasing amounts of copper (by adding additional drops) to the experimental environments to test the effects of increased concentrations of copper. After each experimental environment has had copper added to it, you will add a number of aquatic organisms, specifically duckweed and snails, to each environment. How will adding copper affect the organisms in each environment?
- How will soluble copper affect an aquatic ecosystem?
- Will all of the organisms be affected similarly or differently?
- How much copper will cause an effect?
- Why do industries need to ensure they are meeting regulatory requirements when working with potentially toxic chemicals?
These items can be purchased from Carolina Biological Supply Company
- Copper sulfate pentahydrate (1 gram)
- Safety goggles
- Small pond snails (at least 15).
- Duckweed (at least 50)
- Daphnia Culture
- Large, reusable plastic containers with lids, 6 cups / 48 oz volume (e.g. Glad or Zip-lock) (5)
- Empty recycled milk container with lid, 1 gallon (cleaned and rinsed thoroughly)
- Permanent marker
- Disposable gloves (at least 1 pair)
- Piece of scratch paper
- Kitchen scale
- Bottled water (7 L)
- Medicine dropper or pipet
- Lab notebook
Activity: Part 1: Set up
Rinse each container thoroughly with water. Do not use soap because it can coat the plastic container and may be harmful to the organisms in your experiment. Prepare the stock copper solution.
A stock solution is a solution that is at a higher concentration than you will use in your tests, and it is used to make solutions with lower concentrations (the concentrations you want to test). This is made by the teacher
Label the 1-gallon milk container “Copper sulfate solution (500ppm)” using the permanent marker. Add 2L of bottled water to the 1-gallon milk container.
Put on safety goggles and gloves. Take the copper sulfate, and the 1-gallon milk container with the water, and scale to a well-ventilated area.
Lay a piece of scratch paper on the scale. Be careful not to let the copper sulfate touch your bare skin or the scale (it can touch the paper), measure out 1 gram of copper sulfate.
Carefully create a funnel using scratch paper to pour the copper sulfate into the 1-gallon milk container with the water. Secure the lid on the container and carefully shake it until the copper sulfate dissolves.
The copper sulfate may take a couple of minutes to dissolve in the water.
You will be creating five experimental aquatic environments, and each one will have a different concentration of copper ions in it, as shown in Table 1.
Using the permanent marker, label your containers with the concentration of the copper ion solution you will put in them (in ppm).
Add 1000mL (1 L) of water to each plastic container.
To each plastic container, add drops of the copper sulfate solution (500 ppm) according to Table 1 above. Add the solution slowly, one drop at a time.
Add at least three snails (more would be better as it would provide more robust data; make sure to keep the number constant between test conditions) to each container, making sure that they are right side up at the bottom of the container. This will help you see if they are alive or not.
Put 10 duckweed plants into each container. Make sure that each duckweed plant has a stem (the part sticking down) and exactly two leaves (the part that floats).
You will be counting the number of leaves on each duckweed plant to see if it grows new leaves over time or not.
For each container, record the number of snails in it and the total number of leaves on the duckweed plants in it (this should be 20 for the first day).
Observe the animals every day for 5 days and write down your observations in your lab notebook in data tables like Tables 2 and 3. For each observation, count the number of snails that are still alive and the total number of leaves on the duckweed plants in each container. Observe the Daphnia under a microscope and describe their behavior.
Activity: Part 2: Graphing
Make graphs of your results. On the bottom (x-axis) of the graphs, put the concentration of copper (in ppm), and on the left side (y-axis) of the graphs put the number of snails living (Graph 1) or the total number of leaves on the duckweed plants (Graph 2) at the end of the experiment (Day 5).
Also, make graphs of the snails and leaves over time. Put the time (in days) on the X-axis and either the number of snails living (Graph 3) or the total number of leaves on the duckweed plants (Graph 4) on the Y-axis.
Examine your graphs. What effect did the arsenic have on the snails? What about duckweed? How did the arsenic concentration impact the effect?
Copper pentahydrate sulfate needs to be disposed of responsibly. Dispose of it according to your local guidelines for pesticide disposal.
Activity: Part 3: Data Tables
|Water in Each Plastic Container (mL)||Drops of Copper Sulfate Solution to Add (500 ppm)||Total Copper Ion Concentration in the Plastic Container (ppm)|
|1,000 ml||0||0 ppm|
|1,000 ml||4||0.1 ppm|
|1,000 ml||8||0.2 ppm|
|1,000 ml||12||0.3 ppm|
|1,000 ml||16||0.4 ppm|
|1,000 ml||20||0.5 ppm|
Wrap-Up and Action
Assessing the students will be done informally through individual group and whole group discussions.
Kick Nets to determine water quality by analyzing macroinvertebrate populations.
Daphnia Culture: $11.40
Ramshorn Snails 25 pack $27.25 x 10
Duckweed $10.50 x 4
Cupric Sulfate, Pentahydrate (Lab Grade) $13.75
Shipping Cost Carolina Biological $76.53
Total Cost Carolina Biological $445.31
64 oz deli containers 20 pack $35.71 x 2
Total Amazon $76.41
Optional: Dissolved Oxygen Meter (Amazon) $89.44
Total Project Cost: $521.72 or $611.16 (with optional materials)
About the Author
My name is Annie McEntyre, I am a 2022-2023 Kenan Fellow. I completed my internship with Befesa Zinc Metal. I am currently a physical science teacher in Rutherford County Schools at East Rutherford High School. I have been teaching for the past twelve years. During my teaching career, I have taught elementary through high school mathematics and science classes.
About the Fellowship
I had the privilege of interning with Befesa Zinc Metal. I was able to shadow all aspects of the facility and work with the process engineers at each step of the zinc recycling process. I also was able to shadow the environmental compliance division learning about how Befesa Zinc Metal protects the environment. This included protecting the Broad River. I was able to visit the ETT lab, the lab responsible for testing water samples from Befesa Zinc Metal. Working with the environmental compliance division and visiting the ETT lab inspired the water quality toxicity investigation.
Download a copy of this lesson and supporting student pages here.