Science Fair Project Title: Which Plants can Resist Walnut Tree Allelopathy?

Allelopathy is a process by which a plant releases chemicals that can either inhibit or benefit other plants. Juglone is the allelopathic substance produced by the Black Walnut tree that works like a herbicide to prevent the germination and growth of plants near the tree.

Project Investigation Protocol:
Two fast growing vegetable plants, red radish and tomato will be treated with two different dosages of juglone extracted from the leaves of the Black Walnut over the course of 7 days. Observations will be made of any adverse affects seen aligned to specific growth criteria. Based on the results of these observations a data table will be prepared.

Important Vocabulary Terms to Start Background Research:
Allelopathy, Juglone, allelopathic effect, autotoxicity, herbicide, walnut toxicity, Black Walnut Tree, juglone toxicity

Research Questions
  • What is Allelopathy and Juglone?
  • How did the juglone solution react with the tomato plants?
  • How did the juglone solution react with the radish plants?
  • How many days did it take for the plant that received 30 ml of juglone solution to die?
  • How long did it take for the plant that received 60 ml of juglone solution to die?
  • Which plant died first?
  • Which of the two plants seemed tolerant of Juglone?

  • Required Materials:
    Radish seeds
    Young tomato plants
    Distilled water
    Black Walnut leaves (bark or roots)*
    Metric measuring cup
    Plastic storage containers
    Large cooking pot
    Potting soil

    Project Experimental Procedure:
    1. Gather 3 young tomato plants and place each in separate flower pots or containers.
    2. Radishes are very well suited to growing in containers. Before planting the seeds soak them in water for not more than ten minutes.
    3. Place 3 radish seeds into 3 separated small flower pots or containers. Make sure the soil is loose, with no rocks or other obstructions. This allows the roots to grow to their proper size without any barriers.
    4. Radish seeds usually germinate within a week. Begin the next part of the investigation when all of the radish plants are at about the same height as the young tomato plants.
    5. Place 20 black walnut leaves (bark or roots) in a pot and fill the pot half full with distilled water.
    6. Bring the leaf-water mixture to a boil. Boil the leaves and water for 10 minutes. Use the stopwatch to keep track of time.
    7. After 10 minutes, use a large spoon to remove the leaves from the water.
    8. Continue to boil the now extracted juglone solution for 20 minutes.
    9. After 20 minutes, allow the liquid to cool completely. Once the liquid has cooled, transfer it to one of the plastic storage containers.
    10. Place all of the plants in an area where they will receive equal amounts of sunlight.
    11. Using a metric measuring cup measure 60 ml (about 2 oz) of juglone solution and add this amount to the first radish plant. Again measure 60 ml of Juglone solution and add to the tomato container.
    12. Label the two plants containers “60 ml of Juglone.”
    13. Measure 30 ml (about 1 oz.) of juglone solution and add 30 ml of distilled water. Pour this amount into the 2nd radish container. Repeat the same procedure with the tomato.
    14. Label the plant containers 30ml Juglone + 30ml water.
    15. Thoroughly rinse out the measuring cup. Water the third radish plant with 60 ml of distilled water only. Label the container “Distilled Water Control.” Repeat the procedure using with the 3rd tomato plant.
    16. Pour Juglone solution on each of the plants at the same time every day. Make sure that the soil is moist, but not wet.
    17. If the Juglone solution does not moisten the soil, add additional distilled water to the plant so that it is moist.
    18. Keep track of the amount of juglone solution made and make more, as needed, following steps 5 through 9.
    19. Repeat step 16 every day for at least seven days. Water the plants at the same time every day.
    20. Observe the plants noting any changes seen. Record all of the observations in a table similar to the one shown.
    21. Record the data in the table. Use the phrases like “no visible change,” “change in leaf color,” “withering of steam,” “stunting of growth,” "wilting of plant,” etc when describing the plants appearance.
    Radish Plant                 Observations per Day
    Solution Dosage  
    1       2      
    6       7      
    60 ml Juglone

    30 ml Juglone

    Distilled water

    Tomato Plant                   Observations per Day
    Solution Dosage  
    1       2      
    6       7      
    60 ml Juglone

    30 ml Juglone

    Distilled water

    Safety Issues:
    Care should be exercised when handling the juglone solution some people and animals have allergic reactions (irritations to the skin, eyes, nose, throat and lungs) to products from walnut trees. Asthma may also be stimulated by exposure to juglone. Disposable plastic gloves or garden gloves should warn when conducting this science fair project, which should be carried-out in a well ventilated area.

    Although juglone has low water solubility and does not move far in the soil, small amounts may be injurious to sensitive houseplants.  Try not to permit juglone, or soil mixed with juglone, and any of the Black walnut parts (bark, leaves, roots, etc) to come in contact with household plants.   

    For more background information on Allelopathy in general and Black Walnut Tree Allelopathy in particular, along with other science fair projects on this subject, visit the websites listed below:

    1) Allelopathy
    2) The Biodegradability of an Allelopathic Chemical in Soil
    3) Black Walnut Toxicity
    4) Black Walnut Trees and Juglone
    5) Which Part of the Walnut Tree Contains the Greatest Concentration of Allelopathic Substances?

    Preparing juglone from Black Walnut tree tissue roots can be quite messy Science In a Bag has designed a kit that includes everything needed (including juglone solution) to conduct this and other allelopathy science fair projects.

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