Yeast Fermentation Lab

Introduction:

During anaerobic conditions high levels of NADH develop, leaving a shortage of NAD⁺. Low levels of NAD⁺ slow the rate of glycolysis. Fermentation restores NAD⁺ levels while producing alcohol and CO₂.

During aerobic respiration glucose is broken down into water and carbon dioxide.
C₆H₁₂O₆ + O₂ =====> 6H₂O + 6CO₂
Under ideal conditions most eukaryotic cells produce 36 ATP molecules from one molecule of glucose.

During fermentation baker's yeast breaks glucose into ethyl alcohol and carbon dioxide.
C₆H₁₂O₆ =====> 2C₂H₅OH + 2CO₂
This process only yields 2 ATP per glucose molecule.

Materials:

15 mL plastic centrifuge tubes with caps

7% yeast solution (4 packages yeast and 400 mL water)

5% glucose solution (20 grams glucose in 400 mL water)

TES-TAPE (available at Wal-Mart and pharmacies)

large beakers (250 to 500 mL)

water bath at 40 degrees C

permanent fine point lab markers

Prelab preparation:

1. Poke 3 to 4 small holes in the centrifuge tube caps using a pin or thumbtack.
2. Prepare the yeast solution by mixing 4 packages active dry yeast in 400 mL tap water.
3. Prepare the sugar solution by mixing 20 grams glucose in 400 mL tap water.
4. Preheat the water bath and the solutions to 40 degrees C.
5. Fill beakers with water and place in the water bath to preheat.
6. Fill beakers with water and leave at room temperature.

Procedure:

1. Fill each tube halfway with sugar solution
2. Fill the rest of each tube with yeast solution, extending the fluid level above the top of the tube
3. Take a small piece of TES-TAPE and measure the amount of glucose
4. Screw the cap on the tubes (a few drops will spurt out the holes)
5. Check to make sure there are no bubbles visible in the tube
6. Invert the tubes and mark two tubes L and the other two tubes C
7. Place one L tube and one C tube in each beaker
8. Return the 40 degree C beaker to the water bath
9. After 5, 10, 15, and 20 minutes...
   • take the L tubes out of the water
   • dry the tops with a paper towel
   • mark the level of gas bubbles (include any foam as part of the bubble
   • return the L tubes to the beakers
   • take the C tubes out of the water
   • turn the tubes upright and remove the caps
   • take two small pieces of TES-TAPE and measure the amount of glucose
   • return the tubes are return to the beakers
10. After 20 minutes, empty the contents of all tubes into the waste beaker
11. Record the mL of CO₂ at each mark
12. Wash and rinse out your tubes
13. Graph the change in glucose and CO₂ over time in your journal
14. Answer the following questions in your journal
    • Why did you have two tubes at each each temperature?
    • What was the variable in this experiment?
    • What was the gas that formed in the tubes?
    • Describe an experiment that you could use to test if a different type of sugar would give the same results.

Reference:

"Fermentation, Respiration, and Enzyme Specificity: A Simple Device and Key Experiments with Yeast," by L. Reinking, J. Reinking, and K. Miller, The American Biology Teacher, Vol. 56, March 1994, pp. 164-168.

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