Abstract

Objective

The goal of this project is to determine if sediment from parking lots contain compounds that are harmful for aquatic organisms.

Introduction

As human population centers grow to cover more and more of the planet, watersheds are increasingly affected by the presence of buildings, roadways, and parking lots. A watershed is "the area of land that catches rain and snow and drains or seeps into a marsh, stream, river, lake or groundwater." (EPA, 2006b.) This project asks the question: "Can run-off from parking lots be toxic to organisms in nearby ponds and streams?" You will learn how to conduct a bioassay to determine if sediments from a parking lot are toxic for aquatic organisms.

As the name suggests, a bioassay uses living organisms as the "detector" for an experimental procedure. When doing environmental testing for toxins, the bioassay is typically a viability assay. You count how many organisms are present at the beginning of the experiment, expose the organisms to different concentrations of the suspsected toxin, and count the number of organisms that remain viable. It is critically important to maintain a control population of the organisms, so that you don't mistake the naturally-occuring death rate of the organisms as the effect of toxins.

The organism that you will use for testing water toxicity is Daphnia magna. Daphnia magna (common name "water fleas") are tiny freshwater crustaceans. They are filter feeders, and can survive in culture by eating algae, bacteria, or yeast.

Figure 1. Photomicrograph of Daphnia, the common water flea.

If you make a graph of the percentage of Daphnia that remain viable (y-axis) vs. the concentration of the compound tested (x-axis), you can determine the LC₅₀ value for the compound. This is the concentration at which 50% of the population remains viable. It is one useful measure for comparing the relative toxicities of compounds.

Terms, Concepts, and Questions to Start Background Research

To do this project, you should do research that enables you to understand the following terms and concepts:

• bioassay,
• watershed,
• Daphnia magna,
• toxicity,
• LC₅₀,
• environmental samples.

Questions

• What is the LC₅₀ for a compound and how is it determined?

Bibliography

• Environmental Inquiry, 2006a. "Bioassays Using Daphnia," Cornell University and Penn State University [accessed October 9, 2006] http://ei.cornell.edu/toxicology/bioassays/daphnia/.
• Information on analyzing Daphnia viability assays:
  Environmental Inquiry, 2006b. "Analyzing Daphnia Data," Cornell University and Penn State University [accessed October 9, 2006] http://ei.cornell.edu/toxicology/bioassays/daphnia/analyze.asp.
• Information on testing environmental samples:
  Environmental Inquiry, 2006c. "Testing Environmental Samples," Cornell University and Penn State University [accessed October 9, 2006] http://ei.cornell.edu/toxicology/bioassays/daphnia/environmnt.asp.
• These two sites have information on culturing Daphnia:
  
  • Environmental Inquiry, 2006d. "Culturing Daphnia," Cornell University and Penn State University [accessed October 9, 2006] http://ei.cornell.edu/toxicology/bioassays/daphnia/culture.asp;
  • Clare, J., 2002. "Daphnia: An Aquarist's Guide," A Caudata.org website [accessed October 9, 2006] http://www.caudata.org/daphnia/.
• For information on toxicity of specific compounds, you can search the following databases:
  
  • EPA, 2006a. "ECOTOX Database," U.S. Environmental Protection Agency [accessed October 19, 2006] http://cfpub.epa.gov/ecotox/.
  • NLM, 2006. "TOXNET Toxicology Data Network," U.S. National Library of Medicine [accessed October 19, 2006] http://toxnet.nlm.nih.gov/f.
• For information on watersheds, see: EPA, 2006b. "Surf Your Watershed," U.S. Environmental Protection Agency [accessed October 19, 2006] http://www.epa.gov/surf/.

Materials and Equipment

To do this experiment you will need the following materials and equipment:

• Daphnia magna:
  • you may be able to purchase these at a local tropical fish store, or
  • a culture kit is available from Carolina Biological Supply Company (part number 14-2326),
• plastic transfer pipettes (to move individual Daphnia from one container to another),
• small fish net (for catching and moving larger numbers of Daphnia),
• broom and dustpan (for collecting sediment),
• electronic balance (for weighing collected sediment samples), such as the Fast Weigh MS-500-BLK Digital Pocket Scale, 500 by 0.1 G, available from Amazon.com
• pure spring water (or dechlorinated tap water),
• small containers (for holding your experimental and control groups before testing).

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Experimental Procedure

1. Do your background research so that you are knowledgeable about the terms, concepts, and questions, above.
2. You will need to plan ahead for this experiment, because it takes time to order and receive the Daphnia magna, and to establish a stable culture of the organisms before testing can begin.
3. Follow the instructions for culturing Daphnia that come with the culture kit. It's a good idea to keep two separate containers of Daphnia. If something goes wrong with one of them, you'll have a back-up.
4. Do not use plain tap water for your Daphnia cultures or test conditions. Most tap water contains chlorine and/or chloramine as a disinfectant. These compounds are lethal for Daphnia. Use spring water or dechlorinated tap water.
5. Once your Daphnia culture is well-established, you are ready to conduct your bioassay. Collect sediment samples from a local parking lot. Take one of your parents along to look out for cars and keep you safe. You should also take notes on where each sample was collected (maybe take a photo), and label each sample.
6. Weigh the sediment samples.
7. Dilute them with spring water at different concentrations, to simulate rainwater run-off after a storm. Label each container with concentration (g/L) of sediment in the water.
8. Transfer 30–50 Daphnia into each container. Keep a population of Daphnia in sediment-free water as a control. Keep all containers in the same environmental conditions (same temperature, light level, feeding, etc.).
9. Count and record the number of Daphnia in each container at 8, 16, 24, and 48 hours after the start of the experiment.
10. Calculate the percentage of viable Daphnia in each container for each time point.
11. Graph the percentage of viable Daphnia (y-axis) vs. sediment concentration (x-axis), for each time point.
12. Does the sediment have any effect on Daphnia viability?
13. If there is a decrease in viability, can you estimate the LC₅₀ for the sediment?

Variations

• Are sediments from other locations (e.g., a playground, or a sidewalk) toxic to Daphnia?
• You could also use this bioassay technique to test water from a local pond or stream. Another idea would be to collect run-off water from your lawn at different time points after application of fertilizer. To collect the run-off water, dig a hole so that you can put a plastic collection container with its opening just below the soil surface level. Sample water at one day, three days and one week after application of fertilizer.
• Since Daphnia are transparent, it is possible to monitor their heart rate under a microscope. This makes pharmacological bioassays a possibility. See the Science Buddies project Caffeine and Heart Rate: A Pharmacological Study Using Daphnia magna.
• Here are some additional Science Buddies projects that involve testing environmental samples with aquatic bioassays: There's Something Fishy About That Fertilizer, Acid Rain and Aquatic Life, and Heavy Metals and Aquatic Environments.

• For more science project ideas in this area of science, see Environmental Science Project Ideas.

Credits

Andrew Olson, Ph.D., Science Buddies

Sources

This project is based on:

• Environmental Inquiry, 2006b. "Analyzing Daphnia Data," Cornell University and Penn State University [accessed October 9, 2006] http://ei.cornell.edu/toxicology/bioassays/daphnia/analyze.asp, and
• Environmental Inquiry, 2006c. "Testing Environmental Samples," Cornell University and Penn State University [accessed October 9, 2006] http://ei.cornell.edu/toxicology/bioassays/daphnia/environmnt.asp.

Last edit date: 2011-11-18 12:00:00