What Color Are the Leaves Really Turning?

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Difficulty
Time Required Very Short (≤ 1 day)
Prerequisites None
Material Availability For your convenience a kit is available from our partner Home Science Tools.
Cost Low ($20 - $50)
Safety Adult supervisions is required while using the stove and hot water. Alcohol is flammable and toxic. Adult supervision is recommended while working with the isopropyl alcohol.

Abstract

Everyone loves the beautiful colors of fall, but where do they come from and how does the change in colors happen? In this project, you will uncover the hidden colors of fall by separating plant pigments with paper chromatography. What colors will you see?

Objective

In this project, you will uncover the changing colors of fall leaves by separating plant pigments with paper chromatography.

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Credits

Sara Agee, PhD, Science Buddies
Edited by Svenja Lohner, PhD, Science Buddies

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General citation information is provided here. Be sure to check the formatting, including capitalization, for the method you are using and update your citation, as needed.

MLA Style

Science Buddies Staff. "What Color Are the Leaves Really Turning?" Science Buddies, 23 Jan. 2019, https://www.sciencebuddies.org/science-fair-projects/project-ideas/PlantBio_p032/plant-biology/leaves-turn-color-pigments. Accessed 25 June 2019.

APA Style

Science Buddies Staff. (2019, January 23). What Color Are the Leaves Really Turning? Retrieved from https://www.sciencebuddies.org/science-fair-projects/project-ideas/PlantBio_p032/plant-biology/leaves-turn-color-pigments


Last edit date: 2019-01-23

Introduction

Everyone loves the beautiful colors of fall, but where do they come from? What happens to make the leaves turn from green to yellow, orange, and red? Part of the answer is that tree leaves have pigments or colorful molecules, inside them. These pigments make food for the tree during warm, sunny months through the process of photosynthesis. Photosynthesis is when the energy of the sun is absorbed by the plant pigments and turned into glucose, or food energy, which keeps the leaves green. But winter brings colder, shorter days and without as much sunlight, the tree shuts down the food-making leaves for the winter. So what you see in the changing autumn colors is the gradual change of these pigments.

  • Chlorophyll (green): Chlorophyll is necessary for photosynthesis, which is the chemical reaction that enables plants to use sunlight to manufacture sugars for their food. Trees in the temperate zones store these sugars for their winter dormant period and so do not need to actively make food during the winter. Tree leaves must constantly make chlorophyll when they need it for photosynthesis, so once they stop, the green goes away.
  • Xanthophylls (yellow) and Carotenoids (orange): These pigments are also used for photosynthesis, and are there all summer long, you just don't see them because they are usually masked by the strong color of the green chlorophyll. These pigments do not break down as fast as chlorophyll, so they hang around longer during the fall. These pigments are also commonly found in such things as corn, carrots, and daffodils, as well as rutabagas, buttercups, and bananas.
  • Anthocyanins (red): The anthocyanins are different, because they begin to appear after the plant stops making chlorophyll. So these pigments are not there during the summer, and they appear latest in the fall succession of color. Anthocyanins are very intense color molecules, and they also give color to such familiar things as cranberries, red apples, concord grapes, blueberries, cherries, strawberries, and plums. They are water soluble and appear in the watery liquid of leaf cells.

When you look at a leaf, you see the result of all of these colors mixing together. But if you separate the pigment molecules, you can see each individual pigment color on its own. One method for separating mixtures of different components is called paper chromatography. In this method, a mixture (such as your pigment mixture) is applied onto a chromatography paper. The paper strip is dipped into a liquid, called the solvent or mobile phase. The liquid will start traveling up the paper strip and carry all the components within the mixture (such as your different color pigments) along through the chromatography paper. While traveling up the paper, each component interacts with the paper and the solvent differently depending on its chemical properties. Some of them are more attracted to the paper whereas others prefer to stay in the mobile phase. As a result, each individual component travels along the paper at a different speed, as shown in Figure 1. This is how with paper chromatography a colorful mixture of pigment molecules can be separated into each individual pigment component.

Chromatography apparatus
Figure 1. Paper chromatography. Molecules are separated from each other, depending on how fast they migrate with the solvent up the chromatography paper. (Wikipedia, 2008.)

In this science project, you will use paper chromatography to separate the colored pigment molecules from fall leaves. By collecting leaves at different stages of turning, you will be able to capture all of the colors of fall. Will you be able to uncover the hidden colors, and tell the full fall story?

Terms and Concepts

To do this type of experiment you should know what the following terms mean. Have an adult help you search the Internet or take you to your local library to find out more.

  • Pigment
  • Photosynthesis
  • Chlorophyll
  • Xanthophyll
  • Carotenoid
  • Anthocyanin
  • Paper chromatography
  • Solvent
Questions
  • What makes a leaf look so colorful?
  • Can I extract the pigment from a leaf?
  • Why do leaves turn fall colors?
  • Are plant pigments involved in leaves turning fall colors?

Bibliography

These resources will give you more information about chromatography and teach you about the types of chromatography used in research labs today:

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Materials and Equipment Product Kit Available

These specialty items can be purchased from our partner Home Science Tools:

  • Candy Chromatography Science Kit (1). You will need these items from the kit:
    • Chromatography paper strips; at least 5. The kit comes with 20 strips; additional chromatography paper can be purchased separately from our partner Home Science Tools.
    • 100 mL beaker
    • 90% isopropyl alcohol
    • Mini binder clips (2)
    • Wooden splints
    • Pipette
  • Note: This kit contains additional items to do other chromatography science projects. See the kit instructions page for details. Downloadable materials safety data sheets (MSDS) are available for all kit chemicals if required by your science fair.

You will also need to gather these items:

  • Leaves at different stages of turning colors (30-40, 10 per color group)
  • Scissors
  • Heat resistant, strong glasses (3-4)
  • Stove
  • Pot
  • Water
  • Wooden spoon
  • Fork
  • Ruler
  • Pencil
  • Plate

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What Color Are the Leaves Really Turning?

www.sciencebuddies.org/science-fair-projects/project-ideas/PlantBio_p032/plant-biology/leaves-turn-color-pigments

Experimental Procedure

  1. Go on a nice walk with an adult and collect some fresh leaves from different stages of color change during fall. It is best for all of your leaves to come from the same tree, so look for a tree with a variety of leaves at different stages. Figure 2 shows an example of some collected leaves:

    Fall leaves collected to do a leaf paper chromatography science experiment.
    Figure 2. Red, green, and yellow leaves collected from a tree in the neighborhood.

  2. Separate and group the leaves into color groups, with ten good leaves in each group (unless you are using a tree with small leaves, like aspen or birch, then you should use a higher number of leaves). Try to form groups from colors that are as different as possible. For example, make a green group, a yellow group, and a red group, as shown in Figure 3. In each group, chose leaves in the deepest colors possible:

    Fall leaves sorted in to piles of green, yellow, and red in preparation for leaf paper chromatography.
    Figure 3. All leaves are separated into a pile of yellow, red, and green leaves.

  3. Cut the leaves into very small pieces with your scissors (all pieces should be smaller than about ¾ inch long) and put each group into the bottom of a heat resistant, strong glass (Figure 4):

    Chopped fall leaves sorted by color into drinking glasses.
    Figure 4. Each pile of leaves is cut into small pieces and placed into a different heat resistant strong glass.

  4. Add 1 Tbsp. of 90% isopropyl alcohol to each glass.
  5. Using the blunt end of a wooden spoon, macerate (soften) the chopped leaves by squashing them into the isopropyl alcohol at the bottom of the cup.
  6. As you squish the leaves, you will notice that the alcohol will start to turn the color of the leaves. This is called extraction, and the isopropyl alcohol is called the solvent.
  7. Continue until the liquid turns a deep shade of the color of the leaves, about 5 minutes per glass.
  8. Let the macerated suspensions sit for 30 minutes in a dark, room-temperature place to allow the color molecules to fully extract.
  9. Using a fork, lift out the bits and pieces of leaf material and set them aside. Take care to remove any liquid by gently pressing the leafy bits against the glass before you remove them. You should be left with a dark suspension of leafy color in isopropyl alcohol at the bottom of your cup.
  10. Pour some water into a pot and heat it up on the stove. Then switch off the stove and place the three heat resistant glasses with your extracts into the hot water. Let them sit in the hot water for about 20–30 minutes, or until the isopropyl alcohol has mostly evaporated. As the alcohol evaporates, your extracts should become thicker when stirred with a fork.
    1. Concentrating the extracts should not require more than 30 minutes of evaporating off the isopropyl alcohol, although the extracts will become more concentrated the longer the alcohol is allowed to evaporate off.
    2. Stir each color thoroughly to blend and loosen any bits of dried up pigment from the side of the bowl. Be sure to use a clean fork for each color so you do not mix them!
  11. In the meantime, prepare your chromatography paper. Cut the chromatography paper into strips approximately 2 centimeters (cm) wide by 6.5 cm long. Prepare a total of 6 chromatography strips this way.
    1. Science Buddies Kit: The kit comes with 20 long strips of chromatography paper; two 6.5 cm strips can be cut from each long strip.
  12. Take one of the chromatography strips and use a ruler and pencil to draw a line across it horizontally 1 cm from the bottom. This is the origin line, see Figure 5 for details. Repeat this step for all 6 of the chromatography strips.
chromatography strip with origin line
Food Science Cooking science project
Figure 5. Draw an origin line on the chromatography strip. The leaf extract to be tested will be spotted in the middle of the origin line.
  1. When your color extracts are concentrated enough, use the pipette to place a small dot of extract at the center of the origin line of one chromatography strip as shown in Figure 5. The drop size should be about the size of a pencil eraser. The more extract you have on your paper towel strip the darker the chromatography will be. Some plant pigments can stain, so you should do this on a plate so that the color won't seep through and stain your work surface. Try to apply the extract as smoothly and evenly as you can along the line. Repeat with 2-3 more strips, using the same color extract, so that you have triplicates for each color pigment.
    1. Use a pencil to label the chromatography strip "red extract, yellow extract, or green extract". Do not use a pen to label the strips: the ink might run when the alcohol passes through the strips.
    2. Allow the spot to dry. If the spot is too faint, you will need to thicken your extract and repeat step 13 on a new strip.
  2. Repeat step 13 with the other colored extracts so that you have three paper strips for each color extract. Be sure to rinse the pipette between each color extract! Allow the strips to dry.
  3. Using your binder clips, clip two of the prepared chromatography strips to a wooden splint, as shown in Figure 6. Make sure the two strips do not touch each other and that the bottoms align.
  4. Rest the wooden splint on top of the 100 mL beaker so that the strips hang into the beaker and do not touch the sides of the beaker, nor the bottom of the beaker. Estimate the distance between the bottom of the beaker and the end of the chromatography strips.
  5. Remove the wooden splint with the chromatography strips and add 90% isopropyl alcohol to the beaker so the alcohol would just reach the end of the chromatography strips if they are placed back.
  6. Place the wooden splint back on top of the beaker. Do both strips touch the alcohol? The goal is to have the strip just touching the surface of the alcohol, as shown in Figure 6. As long as the alcohol touches the strips and the alcohol level is below the extract dots, you are good to go!
    1. If necessary, remove the paper strips from the beaker and add more 90% isopropyl alcohol to reach the desired alcohol level.
experimental setup for paper chromatography of black ink
Food Science Cooking science project
Figure 6. Your setup should look similar to this example. The end of the chromatography strip should just touch the alcohol. Note: This picture does not show chromatography strips with plant extract. The colors on your paper strips should look different.
  1. Set the beaker aside for about 30 to 60 minutes, and watch as the pigments separate along the length of the strip. As soon as one of the colors reaches the top of a strip, remove the strips and allow them to dry.
  2. Repeat steps 15 to 19 with all your remaining paper strips.
  3. Compare the colors found in the different strips. What happened to the colors? Did the different groups of leaves have unique colors, or shared colors, or both? Is each color found in the same place along each strip, or in different places? Are the colors in the same order, or in a different order of separation along the strip?
  4. Tip: If you have a pale chromatography, next time try using more leaves, cutting them up into smaller pieces (in step 3), and/or adding more of your colored extract solution onto the pencil line on the chromatography paper (step 13). Alternatively, you can also try to use the extracts themselves as your solvent. Replace the isopropyl alcohol in your beaker with one of the extracts. Then hang the paper strips directly into the extract. Repeat with the other leaf extracts.

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