Lab 6: Cell Respiration

Part 1 Procedures:

1. Set up the water baths. The water bath will buffer the respirometers against temperature change during the experiment.
   • Place a sheet of paper in the bottom of the water bath. It will make the graduated pipet easier to read.
   • Place a thermometer in each tray. Observe the thermometer to make sure the temperatures of the water bath is stable.
2. Prepare the peas and beads.
   • Respirometer 1: Put 25 mL of water in a graduated tube. Drop in 25 germinating peas. Determine the volume of the water that has been displaced (equivalent to the volume of peas). Record the volume. Remove the peas and place them on a paper towel.
   • Respirometer 2: Refill the graduated tube with 25mL of water. Drop in 25 dry (dormant) peas. Add enough beads to equal the volume of the germinating peas from respirometer 1. Remove the peas and beads and place them on a paper towel.
   • Respirometer 3: Refill the graduated tube with 25 mL of water. Add enough beads to equal the volume of the germinating peas. Remove the beads and place on a paper towel.

3. Prepare samples.

• Place absorbent cotton ball at the bottom of each respirator.
• Use a pipet to saturate the cotton with 2 mL of 15% KOH.
• Place a small wad of dry, nonabsorbent cotton on top of the KOH-soaked cotton. This will prevent the KOH from contacting the peas. The amount of cotton and KOH need to be the same for all respirometers.
• Place 25 germinating peas in respirometer 1.
• Place 25 dry peas and beads in respirometer 2.
• Place the equivalent volume of beads in respirometer 3.
• Insert a stopper fitted with a calibrated pipet into each respirometer vial. The stopper must fit tightly. If the respirometers leak during the experiment, you will have to start over.

4. Place the set of respirometers in the water bath with the pipet tips resting on a piece of tape. Wait 5 minutes before proceeding to allow the respirometers to each thermal equilibrium with the water. If a respirometer fills with water, there is a leak and you need to start over.

5. After the equilibrium period, immerse all the respirometers (including pipet tips) in the water bath. Position the respirometers so that you can read the scale on each pipet. Do NOT put anything into or take anything out of the water bath until you have taken all measurements.

6. Allow the respirometers to equilibrate for another 5 minutes.

7. Observe the initial volume reading on the scale to the nearest 0.01 mL. Record the data for time 0. Observe and record the temperature of the water. Repeat for all 3 samples.

8. Every 5 minutes for 20 minutes, record the temperature of the water and take readings of the volume of air in each of the pipets. Record data and perform calculations.