Bubble The Trouble Away
(Teacher Notes)
Standard-Objective-Eligible Content: II-4, a (See pages B-2 - B-10.)
Lab Time: 50-60 minutes
Background:
Chemical reactions make life possible. If these reactions proceed too slowly, the ordinary
activities of life would come to a halt. A substance that can speed up that rate is called a
catalyst. Catalysts work by lowering the “start up” or activation energy of a reaction and are not
changed or used up themselves.
Living organisms contain their own special catalysts, which are proteins known as enzymes. An
enzyme may accelerate a reaction by a factor of 10 to the 10th power, so that a reaction that
might take 1500 years can be completed in a cell in just five seconds!
Enzymes speed up a reaction by binding to the substances that enter the reaction. These
substrates bind to the enzymes at a region known as the active site. The way the reaction is
catalyzed may occur because the enzyme holds two substances together in positions in which
they can react with each other; or an enzyme can twist a substrate so that a chemical bond is
broken, producing two smaller molecules. An enzyme’s shape is so specific for that substrate
that it can be compared to a lock and key. In fact, simple cells have as many as 2000 different
enzymes, each to catalyze a different reaction!
Materials/Equipment: See student handout.
The 3% hydrogen peroxide and liver (chicken or beef) can be purchased at a grocery store. For
investigating temperature influence, provide thermometers, ice, and boiling water. Provide
lemon juice, ammonia, and wide-range pH paper to investigate pH as a factor. Other plant and
animal tissues, such as fresh potato, apple, carrot, chicken meat, could be purchased to test the
occurrences of catalase in various living tissues.
Pre-Activity: (15-20 minutes)
1. Divide into work groups and instruct teams to propose an answer for the question, “What
would happen to your cells if they made a poisonous chemical?” List responses on the
board.
2. Explain the role of enzymes in speeding up the breakdown of toxins by modeling enzyme
activity with jigsaw-shaped pieces to represent the enzyme catalase and its substrate
hydrogen peroxide. Display the “lock and key” nature. Write on the back of the jigsaw
pieces: enzyme, substrate, and the area of active site.
3. Write the balanced equation for this reaction: H2O -H2O + O2 . The word catalase
belongs over the arrow in this reaction. Have students identify the substrate, the enzyme,
and the products of this reaction.
Activity: (45 minutes)
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1. Allow one team to gather materials for the activity while the other groups read the
Background and Procedures.
2. Instruct the demo team to follow procedure in Steps 1-4 and make observations aloud for the
class. Ask the class to devise a way to standardize their estimates of the bubbling activity
(such as a scale of 0-5 with 5=greatest and 0=none), and to explain why such a scale would
be important.
3. Refer to procedure Steps 3 through 4 in the Student Handout to discuss the concepts that
enzymes are not altered by the reaction they participate in, i.e., are not “used up,” and that
catalase activity is exothermic in nature.
4. Guide groups to answer the questions.
Designing the experiment: (2 approaches)
1. Ask groups to propose factors that would influence the rate of enzyme activity. Use guided
questions to select one factor that the class will examine. As they decide how to approach
the problem, assess their understanding of dependent and independent variables and a need
for a control. Remind them to organize their observed data into comparison charts and
graphs using the same rating scale as before.
OR
2. Choose from the suggested activities below to test influencing factors.
A. What is the Effect of Temperature on Catalase Activity?
1. Place a small piece of liver, covered with distilled water, in a boiling water bath
(100C) for five minutes. Predict the effects on the enzyme.
2. CAUTION: Carefully remove the test tube from hot water using tongs, allow it to
cool, and pour off the water. Add 2 mL of 3% hydrogen peroxide and record the rate
of reaction. Explain the results.
3. Place equal quantities of liver into each of three test tubes and 2 mL of hydrogen
peroxide into three others. Place a test tube of liver and one of hydrogen peroxide
into each of the following: 0C (ice) water bath, 22C (room temperature), and 37C
(warm water bath).
4. After three minutes, pour the tube of hydrogen peroxide into the corresponding tube
of liver for each temperature. Record the reaction rates for each.
5. Make a graph of the reaction rates compared to temperature. What is the best
(optimum) temperature for catalase activity?
B. What is the Effect of pH on Catalase Activity?
1. Measure and add 2 mL of hydrogen peroxide to each of three clean test tubes. Then
to:
Tube 1--add 10 drops of lemon juice (or 1N HCl) using stirring rod to mix.
Tube 2--add 10 drops of ammonia (or 1N NaOH) using stirring rod to mix.
Tube 3--add 2 drops of ammonia and 1 drop of lemon juice using stirring rod to mix.
2. Determine pH with wide range pH paper or sensor. Record the pH number value for
each tube.
3. Add a small piece of liver to each test tube. Estimate and record the rates of
reaction.
4. Make a graph of estimated reaction rates compared to pH. At what pH does there
appear to be a best reaction? What is the effect of low or high pH on enzyme
activity?
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Sample Data and Calculations:
Generally the temperature effect will show greatest activity around 30-37C then start dropping
until there is no activity at 100C (protein enzymes are denatured and shape is changed at those
settings).
The optimum pH for catalase activity is between pH 7 and pH 10 (slightly basic) with the lowest
being in the acid range of pH 2-4. (High levels of hydrogen ion concentration -low pH- also tend
to denature the protein conformation or alter the polarity of the molecules.)
At higher concentrations of hydrogen peroxide, there is a greater chance that an enzyme
molecule will collide with this substrate.
1. Measure and place 2mL of the 3% hydrogen peroxide into a clean test tube or cup.
2. Using the forceps, add a small piece of liver to the test tube pushing it into the peroxide with
the stirring rod.
3. Pour off that liquid into another test tube.
4. Add another 2 mL of the 3% hydrogen peroxide to the liver remaining in the test tube from
Step 2.
5. What factors would influence the rate of this enzyme activity? Design an experiment to test
that influence. Be sure to include materials, procedure, data table, and conclusions.
Student Questions and Answers:
1. Describe the peroxide. Is it bubbling? colorless liquid, no bubbles
2. What do you observe? Which product of this reaction is being released? rapidly rising
froth of bubbles, oxygen
3. The liquid is now composed of what? What would happen if more liver were added to this
liquid? Why? water; nothing, the substrate has already been catalyzed
4. Is there any reaction? Predict what would happen if this liquid were poured off and more
hydrogen peroxide added to the liver again. yes, more bubbles, predict same reaction every
time since enzymes are not altered or used up in a reaction
5. What factors would influence the rate of this enzyme activity? Class discussion will vary
depending on available materials and time frame.
Additional Questions:
1. What other way could the products of this enzyme be identified? (glowing splint for oxygen
gas)
2. What other way could the products of this enzyme be measured? (pressure sensor for gas)
3. What happens to the heat produced when this reaction occurs in living cells? (generates body
heat)
4. Predict the effect of prolonged high body temperatures (fever).
Extensions:
Hydroxylamine is a competitive inhibitor of catalase that attaches to catalase and interferes with
its normal binding with hydrogen peroxide. To test its effects, five drops of 5% hydroxylamine
can be added to 2 mL of hydrogen peroxide before the piece of liver is introduced. Other
extensions could include research into conditions and diseases associated with enzyme absence
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or malfunctions.
Reading Comprehension Connection: I-2, I-3 (See page B-11.)
Resources:
Internet:
Community College of Baltimore County, MD - Dr. Gary E. Kaiser-BIO 141 Microbiology Lab
Manual
http://www.cat.cc.md.us/courses/bio141/labmanua/lab8/index.html
Access Excellence High School Biology - Computer Interfacing Experiments
http://outcast.gene.com/ae/21st/TE/PW/ciexp.html
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Bubble The Trouble Away
(Student Handout)
Purpose: To examine the activity of an enzyme in living tissues
Background:
By the time a student can count down 5...4...3...2...1 seconds, an enzyme can complete a
chemical reaction that normally might take 1500 years! In this activity, you will study one such
enzyme called catalase. It is found in many living cells and can speed up the reaction that
breaks down poisonous hydrogen peroxide into two harmless substances: water and oxygen.
Since hydrogen peroxide is a by-product of so many normal cell activities, it must be quickly
broken down or those cells would die. Beef- or chicken-liver cells will be used to demonstrate
the activity of catalase. Even though these cells are actually no longer alive, their enzymes
remain active for several weeks.
Materials/Equipment:
2mL 3% hydrogen peroxide
10 mL graduated cylinder (or small calibrated measure)
2 test tubes with rack (or small, clear containers)
Stirring rod
Forceps
Pea-sized piece of liver (chicken or beef)
Safety Considerations: Always wear goggles in lab. Use test tube holder for hot test tubes.
Handle carefully. Be sure to clean stirring rod each time.
Procedure:
What is Normal Catalase Activity?
1. Measure and place 2mL of the 3% hydrogen peroxide into a clean test tube or cup. Describe
the peroxide. Is it bubbling?
2. Using the forceps, add a small piece of liver to the test tube pushing it into the peroxide with
the stirring rod. What do you observe? Which product of this reaction is being released?
3. Pour off that liquid into another test tube. What do you think the liquid is now composed
of? What do you think would happen if you added more liver to this liquid? Why?
4. Add another 2 mL of the 3% hydrogen peroxide to the liver remaining in the test tube from
Step 2. Do you observe any reaction? Predict what would happen if you poured off this
liquid and added more hydrogen peroxide to the liver again.
5. What factors do you think would influence the rate of this enzyme activity? Design an
experiment to test that influence. Be sure to include materials, procedure, data table, and
conclusions.
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