In This Laboratory Exercise The Class Will Determine The Rate Of Catalase Activity Under Various Conditions Such As Di 1 (165.77 KiB) Viewed 10 times
In this laboratory exercise, the class will determine the rate of Catalase activity under various conditions, such as different enzyme concentrations, pH values, and temperatures. Check with your instructor before proceeding! MATERIALS Laptop Computer Vernier computer interface Logger Pro application Vernier O₂ Gas Sensor 400 mL beaker 10 mL graduated cylinder 250 mL Nalgene bottle three dropper pipettes Exercise I: TESTING THE EFFECT OF ENZYME CONCENTRATION PROCEDURE & RESULTS 1. Connect the Oxygen Gas Sensor to the computer interface. Prepare the computer for data collection by opening the file "06A Enzyme (O₂)" from the Biology with Computers folder of Logger Pro. 2. Place four test tubes in a rack and label them 1, 2, 3, & 4. Fill each test tube with 10 ml of 1.5% H₂O2 (use the Re-pipet bottle on your table, if provided). To use the re-pipettor, hold the test tube under the output tube. Pull the plunger top straight up and gently push straight down. When the plunger is down the red line should be set to 10 ml. 3. Initiate the enzyme catalyzed reaction. • 1.5% and 3.0% H₂O2 enzyme suspension five 18 x 150 mm test tubes Ice · pH buffers test tube rack Thermometer . Pour the 10 ml of 1.5% H2O2 into the 250 ml Nalgene bottle. Using the labeled transfer pipette, draw up some of the enzyme suspension.. Add 5 drops of enzyme solution to Nalgene bottle. Quickly, swirl the bottle once and place the O₂ Gas Sensor into the bottle as shown in Figure 1. Gently push the sensor down into the bottle until it stops. The sensor is designed to seal the bottle without the need for unnecessary force. Click Collect on computer to begin data collection. The data collection will automatically stop after 3 minutes. 4. When data collection is finished, remove the O₂ gas sensor from the Nalgene bottle. Rinse the bottle with water and dry the sensor and the neck of the bottle with a paper towel. 5. Move your data to a stored run. To do this, choose Store Latest Run from the Experiment menu. 6. Measure the slope of the generated line plot at different time intervals. Click on "Tangent" button and use the mouse to drag the cursor to each time interval listed in Table 2 and enter the slope values. Table 2. Rate of O2 production within different time intervals (%O2/min) 0.5-1.0 min 1.0-1.5 min 1.5-2.0 min 0-0.5 min 0.404 0.510 1.121' 1.832 5 Drops Question: Which time interval shows the slowest and highest enzymatic rates? Why? Question: Does the rate drop in the time interval after the highest one? If yes, explain why? 7. Collect data for test tubes 2, 3, and 4. Repeat Steps 3-5 using different amounts of enzyme: 10. 20. And 0 drops. € 40 2.0-3.0 min 1.543
8. Determine the maximum slopes, known as "initial" rate of the enzyme activity for each of the line plots. Click on "Tangent" button and use the mouse to drag the cursor to the time interval with the highest slope, which is usually between minutes 1 and 2. These slopes represent Catalase activity under the conditions of each line plot. Enter the values of the highest slopes in Table 3. 9. Table 3. Summary of the Effect of Enzyme concentration on Catalase Activity Test tube label Maximum Slope, or rate (%O₂/min) 5 Drops 10 Drops 20 Drops 0 Drops 1.832 3.551 12. 135 0.060 Check with your instructor, before proceeding, and enter your results into the class results file. QUESTIONS 1. Which enzyme concentration produced is the lowest reaction rate? Explain why. 2. Which enzyme concentration produced is the highest reaction rate? Explain why. 3. What do you think will happen to the rate of the reaction if one increases the concentration of enzyme to twenty-five drops? Predict what the rate would be for 25 drops. 4. How does changing the concentration of enzyme affect the rate of decomposition of H₂O₂? Exercise II: TESTING THE EFFECT OF TEMPERATURE (Refer to your instructor's protocol) Exercise III: TESTING THE EFFECT OF pH (Refer to your instructor's protocol) 41 3
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