1. Chromatography of Food Colors
Vanderbilt Student Volunteers for Science
Spring, 2007
Training Presentation

2. Important!!!
Please use this resource to reinforce your understanding of the lesson! Make sure you have read and understand the entire lesson prior to picking up the kit!
We recommend that you work through the kit with your team prior to going into the classroom.
This presentation does not contain the entire lesson—only selected experiments that may be difficult to visualize and/or understand.

3. Pre-lab
Use the scissors to cut the top off one Kool-Aid package of each color.
Add one scoop of blue to the blue cups, one scoop of grape to the grape cups, and two scoops of orange to the orange cups.
After you finish, roll the top of the package up and fasten with a piece of scotch tape.
Take the brown food color from the #2 bag and add 2 to 4 drops to the 1-oz cup labeled brown.

4. I. Introduction (p.2)
Ask: What is a mixture? a compound? How do you tell the difference?
Both a mixture and a compound are combinations of substances.
The difference between them is that a mixture can be separated by physical means and a compound can’t.
In a mixture each substance keeps its own properties.
Write the formula for water on the board. (H2O) Explain that water is the combination of two elements: hydrogen and oxygen.
Does water have the properties of oxygen and hydrogen? (no)
Can you separate water into hydrogen and oxygen?
Not easily. It takes special equipment to separate water. Water is a compound, not a mixture.
Ask: What substances make up a salad? How would you separate the mixture?
Salads are mixtures that are easy to separate. You simply take out the lettuce, tomatoes, and carrots. You can physically separate the different substances.

5. II. Check: Identifying mixtures and compounds (p.2)
Substance Mixture or compound? How to separate physically
raisin bread mixture pick out the raisins and nuts with your hand
sugar compound can’t separate (C12H22O11)
salt water mixture filter the salt out of the water or evaporate the water
salt compound can’t separate - NaCl
hamburger mixture buns, lettuce, tomatoes, scrape off catsup, remove meat
rubbing alcohol compound can’t separate (C3H7OH)

6. III. Demo: Separating colors (p.3)
Ask: Is green a pure color or a mixture of colors? If so, how can we physically separate it?
Record their ideas.
Use a 16 oz. plastic cup, chromatography paper strip and a green marker.
Pour a small amount of water in the bottom of the cup.
Place a wooden stick across the top of the cup.
Draw a line with the green marker just above the pencil mark on the chromatography strip.
Fold the paper and hang it from the wooden stick as shown on the next page. Be sure the color line is above the water.
In about 2-5 minutes the green should be separated into blue and yellow.
Take the chromatography paper out of the water as soon as the separate colors can be seen.

7. Explanation (p.3)
As the water travels to the top of the paper, it dissolves the dyes in the pen.
Each dye travels at a different speed depending on how strongly it is absorbed on the paper and how soluble it is in water.
In this case the blue travels faster than the yellow.
The retention factor (RF) of the dye is defined as the distance the dye moves divided by the distance traveled by the water.
The retention factor of any dye is characteristic for a given combination of paper and solvent.
For the same type of paper and solvent, and in the same time allowed, the blue and yellow dyes will always travel to the same position on the paper.
We can use retention factors in experiments to help us identify unknown dyes.
RF = distance dye moved /distance water moved

8. IV. Chromatography of food colors--demo (p.4)
Demonstrate the procedure first!
Show each student where to place the well plates on the worksheet so they can tell what materials will be going into the wells.
Add one scoop of each Kool Aid flavor to its own well and then add 2-4 drops of water to each well containing the Kool Aid.
Use the dropper bottles of food colors to add 2 to 4 drops of food color to the appropriate well.
Pick up the chromatography sheet they will be using and explain that they will be using a different toothpick to draw a line of each food color on one chromatography sheet and Kool-aid colors and brown food color on another chromatography sheet following the color and Kool-aid notations at the top of the two sheets.
Using a different half toothpick each time, draw a line for each of the food colors on the one sheet by using a different toothpick for each color and drawing a line on the pencil line, following the color notation at the top of the chromatography sheet.

9. IV. Chromatography of food colors (cont.)
Gently wave the chromatography sheet to allow the colors to dry.
Show the laminated chromatography sheet for both food colors and Kool-aid colors already drawn on the lines as an example
Refold the chromatography sheet like an accordion so it stands in the cup.
Remove the chromatography sheet and add the contents from the small bottle of water.
Then place the chromatography sheet back in the cup, making sure the water is below the color lines.

10. IV. Chromatography of food colors (cont.)
Distribute the 24-well plates to the students and have each student add one scoop of Kool Aid to the appropriate well.
Take the dropper bottles of food colors around to the student in each pair that has the food colors worksheet, and add 2 to 4 drops of food colors to the appropriate well in the well plate (blue, green, red, yellow for one student and brown for the other student.)
While this is being done, tell students working with the Kool-Aid to add a few drops of water to each well containing Kool-Aid, using their water dropping bottle and stir with the toothpick.
Use the toothpicks to draw a line of color on the pencil line on the student chromatography sheet.
Use a different toothpick for each Kool-Aid solution.
Dip the toothpick in the solution, then draw a line of that color on the pencil line on the chromatography sheet.

11. IV. Chromatography of food colors (cont.)
Gently wave the chromatography sheet to allow the colors to dry.
Refold the chromatography sheet like an accordion so it stands in the cup. Remove the sheets from the cups.
Take out the 2 small bottles of water and add one bottle to each of the 16-oz cups.
Put each chromatography sheet in a cup. It doesn’t matter if it rests against the edge.
Make sure the water in the cup is below all the color lines.
The chromatography sheets should be left to develop (about 20 minutes).
While colors are separating, do the chromatography skit and discuss retention factors and the different food dyes.
After the solvent front reaches the line drawn on the chromatography sheet, remove the chromatography sheet from the cup and place each one on a sheet of paper towel to dry.

12. IV. Chromatography of food colors (cont.)
Look at their food color chromatography sheet and identify which of the food colors listed to the right are present.
Refer to the Kool-aid ingredient sheets and show how to check for what food colors are present.
Ask students to determine what colors are present in each of the Kool-aid chromatograms. These should be compared to the Kool-aid ingredient sheet.
Finally, students should determine what food colors are present in the brown food color.
Question: If you are allergic to yellow food colors, which of the Kool-aid flavors you’ve studied today might cause you to have an allergic reaction?
Answer: Orange
Each student gets to keep their chromatography sheet.
Food Dyes.
The seven food colors approved by the Food and Drug Administration are:
Red #3
Red #40
Yellow #5
Yellow #6
Blue # 1
Blue #2
Green #3.
Only Red #3, Red #40, Yellow #5, Blue #1, and Green #3 are available in food color sets in grocery stores.
Green #3, Yellow #6, and Blue #2 are not used very much in coloring food.
Since Yellow #5 causes allergic reactions in some people, the FDA now requires that it be listed on food labels when it is used.