6. Cell Energy - Colorful Respiration

Blow through a straw into bluish liquid and watch it turn green then yellow before your eyes. Put some plants into the yellow liquid, leave it in a sunny window, come back the next day and the liquid is green. What if you leave the plants in the dark? What if you put some pond snails in? What if you put both pond snails and plants? What’s going on?

The liquid is bromthymol blue (BTB) a non-toxic acid-base indicator that can be used to indirectly measure levels of dissolved carbon dioxide (CO2). The amount of CO2 in a solution changes the pH. An increase in CO2 makes a solution more acidic (the pH gets lower). A decrease in CO2 makes a solution more basic (the pH gets higher). The reason for this is that carbon dioxide that is dissolved in water is in equilibrium with carbonic acid (H2CO3).

CO2 + H2O ↔ H2CO3

In any solution, while the majority of CO2 stays as CO2, some of it is converted to H2CO3, turning the solution slightly acidic. If CO2 is added to the water, the level of H2CO3 will rise and the solution will become more acidic. If CO2 is removed from the water, the amount of H2CO3 falls and the solution becomes more basic. Thus, acid-base indicators such as BTB can indirectly measure the amount of CO2 in a solution.

For more than you ever wanted to know about carbonic acid, see the Wikipedia article on carbonic acid. For the example lesson plans developed by Bob Culler through Access Excellence at the National Health Museum. For a great time lapse video showing BTB color changes using elodea and snails, see Activity C13 from Addison-Wesley’s Science 10 curriculum.


  • Bromthymol blue (BTB can be ordered from any science supply company such as Flinn Scientific $9 for 1 liter 0.04% BTB solution).
  • Several 2 liter soda bottles
  • Test tubes
  • 500 ml beakers or disposable plastic or paper cups
  • Water (since the pH of tap water varies, you may wish to use distilled water for your master BTB solution)
  • Drinking straws
  • Plastic wrap
  • Elodea
  • Pond snails


  1. Before the lesson, the teacher should mix a master BTB solution in one or more 2 liter soda bottles. For each 2 liter bottle, mix 120 ml 0.04% BTB with 1800 ml water. The end result should be a medium blue master BTB solution, dilute enough to be safe for plants and snails but dark enough to see the color changes.
  2. Pour 200 ml diluted BTB in a beaker or cup.
  3. Take a deep breath then blow bubbles in the BTB solution through a drinking straw. What happened? Why?
  4. Set up a test tube rack with 3 tubes. In tube #1 put unbubbled BTB solution (blue). In tube #2 put bubbled BTB solution (yellow). Tubes #1 and #2 will be your comparison tubes. In tube #3 you have a choice of what to do. Choose one option from each of the following columns:
    BTB solution Living things
    Light conditions
    bubbled BTB (yellow) spring of Elodea Sunny window/bright light
    unbubbled BTB (blue) 5 pond snails Dark closet/drak heavy cloth
    both Elodea and 5 pond snails  
  5. Make a hypothesis about what will happen to your tube.
  6. After 24 hours, check the color of your tube. What happened? Why?

Going Further

  1. Investigate the effect of exercise. Compare blowing bubbles in BTB for 3 seconds before and after vigorous exercise (such as doing 2 minutes of jumping jacks).
  2. Investigate the effect of holding your breath. Compare blowing bubbles in BTB for 3 seconds before and after holding your breath for as long as possible (without passing out).
  3. Try this lesson developed by NASA. It describes how to capture various gases (room air, human exhalation, car exhaust, or carbon dioxide from a chemical reaction) in a balloon and use BTB to measure the carbon dioxide content.

Attachment Size
bromthymol_blue.doc 20.5 KB
colorful_respiration.doc 38.5 KB