Name ______Date ______
Testing Enzyme Activity
Background Information
Like nonliving matter, all organisms are made of chemicals. No single substance in an organism is alive yet together, all the substances that make up an organism are alive.
Organisms are mixtures that contain mostly water. Water has a pH that ranges from 6.5 to 7.5. It dissolves many things and makes many of the chemical activities that occur in organisms possible. Enzymes control the chemical activities of living things. Enzymes reduce the amount of activation energy needed to get chemical reactions started. The ability of enzymes to function is influenced by pH.
In this laboratory exercise, you will examine the influence of pH on the enzyme catalase. Catalase will break down hydrogen peroxide (H2O2) into water (H2O and O2). Liver contains the enzyme catalase. It will be the source of catalase for our lab today.
Problem
How does pH influence the activity of the enzyme catalase?
Materials
5 125 mL test tubes
Test tube rack
Petri dish
3 pipettes
Stiring rod
pH paper
0.1M Sodium Hydroxide (NaOH)
0.1M Hydrochloric Acid (HCl)
3% Hydrogen peroxide
Liver puree (catalase)
Small piece of raw beef liver
Labeling tape
Marker
Safety glasses
Procedure
1. Thoroughly read through the lab and do the Prelab table provided below. Record what will go into each test tube prior to starting the experiment. If this is not completed in advance of the lab, you will not be conducting the laboratory and miss out on all the fun!!
Prelab Chart: For Procedure #4 – 9.
Test Tube / Amount of Water / Amount of HCl / Amount of NaOH / Amount of Liver / Amount of H2O21
2
3
4
5
2. Put a drop of raw liver puree on a petri dish. Using a dropper, put a drop of hydrogen peroxide on the liver. Observe what happens and record it below
3. Clean out the petri dish and now place a piece of boiled liver puree onto the petri dish. Again, put a drop of hydrogen peroxide on the liver. Observe what happens and record it below.
4. Using the marker and labeling tape, label the 5 test tubes 1- 5. With a clean dropper, put 10 drops of water in test tube #1. Using a graduated cylinder, measure out 5mL of hydrochloric acid and add it to the water in test tube #1. (When diluting an acid, always pour the acid into the water). Rinse the dropper and the graduated cylinder in the sink.
5. Prepare the following solutions for the remaining test tubes. Use procedures similar to those used in step 3.
a. Test tube 2: 10 drops of hydrochloric acid added to 5 mL of water.
b. Test tube 3: 5.5 mL of water alone
c. Test tube 4: 10 drops of sodium hydroxide added to 5 mL of water
d. Test tube 5: 5 mL of sodium hydroxide added to 10 drops of water.
6. Using the stirring rod, put a drop of the solution from Test tube 1 onto a piece of pH paper. Wipe off the stirring rod. Record its pH. Repeat the procedure for the solutions in Test tubes 2- 5.
7. Place a paper towel under the test tube rack with the 5 tests tubes in it. Using a clean dropper, drop 5 drops of liver puree to each solution and mix well by swirling the test tube gently.
8. Using the clean graduated cylinder, add 5mL of hydrogen peroxide to each test tube. Get ready to identify the amount of activity in each test tube from 0 –4, as you did in step 6.
9. Your teacher will do this part for you as a demonstration to not waste lab time.
Using a clean graduated cylinder, add 5mL of hydrogen peroxide to each test tube. Look for bubbles. Identify the amount of activity in each flask by assigning it a number from 0-4, with 0 meaning no bubbling and 5 meaning many bubbles.
Observations:
Data Table: Observations of Catalase activity level
Test Tube / Without Liver Puree* / With Liver PureepH / Activity (0-4) / pH / Activity (0-4)
1
2
3
4
5
* Conducted by your teacher
1. What happened when hydrogen peroxide was added to each test tube without the liver puree? Discuss why this happened.
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2. What happened when hydrogen peroxide was added to the test tubes with the liver puree? Discuss why this happened.
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3. In which test tube was the bubbling (reaction) activity the greatest? What pH was that solution?
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Analysis and Conclusions:
1. What was the purpose of the test tubes without the liver puree in them?
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2. At what pH does catalase function best at breaking hydrogen peroxide down into water and oxygen? How do you know this?
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3. What conclusion can you draw therefore concerning the pH in body cells containing catalase?
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4. Predict ways in which pH affects the chemical reactions in living cells.
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5. In the investigation conducted, how could you tell that it was the pH that affected the chemical activity of the enzyme rather than the chemical activity of the hydrogen peroxide?
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6. Water is the major component of cells. What does this tell you about the optimum pH for most enzymes? Explain.
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7. The stomach produces hydrochloric acid to help digest food. How does this affect the pH of the stomach?
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8. What would happen if cells did not have any enzymes at all?
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