STUDENT GUIDE
ACTIVITY #9: Investigation Enzyme Action
Most of the chemical reactions that occur in living things are carried out by
protein catalysts called enzymes. Enzymes lower the activation energy required
for reactions to proceed. Enzymes are reusable and work best within certain
ranges of temperature and pH. Since enzymes are specific and can catalyze
only one kind of reaction, it takes thousands different enzymes to conduct life
processes. Enzymes are used by the biotechnology industry in the manufacture
of many food and household products.



Why are enzymes important?
How do enzymes really work?
What are some factors which affect enzyme action?
GOALS: In this lab activity, you will…
 Observe the enzyme catalase at work in
organisms from different kingdoms.
 Learn about the specificity of enzyme catalyzed
reactions.
 Understand the importance of the practical
applications of enzyme catalyzed reactions.
 Determine what factors might affect an enzymecatalyzed reaction.
 Understand what happens to enzymes after
they catalyze a reaction.
MAIN IDEAS: The important concepts and skills covered in this activity are…
 Nearly all metabolic reactions involve enzymes.
 Enzymes are protein catalysts that greatly speed up chemical reactions by
lowering the activation energy required to make reactions proceed within
the narrow range of temperatures in which living organisms function.
 There are many different types of enzymes. Each catalyzes only one or a
few specific reactions. Enzyme specificity is due to the structure of the
enzyme’s active site which must match the shape of the substrate
molecule it acts upon. The interaction of the active site with the substrate
lowers activation energy.
 The rate of enzyme activity is affected by temperature, pH and enzyme
concentration.
 In enzyme reactions, as in all chemical reactions, matter and energy are
conserved.
 In living things, enzymes are not used up – once they catalyze a particular
reaction, they may be reused.
 Enzymes are added to or used in the manufacture of many food and
household products.
ACTIVITY OVERVIEW: A synopsis of this lesson is as follows:
Engage: This short engagement activity is designed to stimulate your
interest in the catalase/H2O2 reaction by having students observe the
oxygen gas bubbles that are produced during the catalytic process. By
testing the reaction in yeast, potato, and beef liver, you will learn that the
enzyme reaction is common to organisms representing the fungi, plant and
animal kingdoms.
Explore: In this exploration activity you will read about enzymes while
working in teams. Since each of the four sets of readings contains
information about different aspects of enzyme action, you must share your
information with the whole class in order to learn all the key concepts about
enzymes.
Explain: In this lab activity you will use a computer, a gas pressure sensor,
and Logger Pro software to measure the rate of the catalase reaction in a
yeast suspension. It is during this activity that you will acquire skills in using
the lab equipment that are necessary for conducting the inquiry activity in
the “Student Designed Experiment” activity which follows.
Elaborate: Students design and carry out their own experiments
investigating some aspect of enzyme activity.
Evaluate: Students answer summary questions.
CONTEXT: The concepts that we have been developing and how this activity
serves as the ‘next step’ can be explained as follows…
In the activity “Observing Mechanical and Chemical Digestion”, students
learned that humans and many other heterotrophs utilize chemical digestion
by way of enzymes to break down complex carbohydrates, lipids and
proteins as they pass from the mouth to the stomach and on to the small
intestines of their digestive systems. If it were not for the presence of these
specific enzymes, the food we eat would never be reduced into small
enough molecules, and would therefore never be able to be utilized by our
bodies at the cellular level for growth and repair. However, enzymes
catalyze more than just digestion reactions. They catalyze many of the
chemical reactions which occur in living things. Students learn about
enzyme action and factors which affect their efficiency throughout these
sets of activities. This activity is the final activity in the Digestion and
Assimilation subunit of the Chemical Biology unit, and ties together chemical
digestion, enzymes and substrates, and the roles of biomolecules in body
function.
CONNECTIONS
Scientific Content –
 Chemical transformations allow a conversion of energy from one form to
another, the breakdown of molecules into smaller units, and the building of
larger molecules from smaller ones.
 Most of these transformations are made possible by protein catalysts called
enzymes.
 Enzymes are protein catalysts that greatly speed up chemical reactions by
lowering the activation energy required to make reactions proceed within
the narrow range of temperatures in which living organisms function.
 There are many different types of enzymes. Each catalyzes only one or a few
specific reactions.
 The rate of enzyme activity is affected by temperature, pH and enzyme
concentration.
 In enzyme reactions, as in all chemical reactions, matter and energy are
conserved.
Scientific Process –
 There is quantitative data collected in this activity.
 Students are asked in the “Elaborate” (or Part B) section to design their own
experiments as an extension of Part A, identify the components of their
experiments (variables, control, constants) and to analyze the data from
their experiments.
Math/Graphing –
 There is graphing associated with Part A and Part B. Students will
appropriately graph their data on graphs provided.
ENGAGE:
ENZYME ACTION
Observing Reactions
Name _________________________ Class _______________
Date ________
INTRODUCTION:
You will begin your study of enzymes by observing and describing the reaction
that occurs when hydrogen peroxide (H2O2) comes in contact with yeast, potato,
and/or beef liver cells.
MATERIALS:
You may be assigned to a lab station that has one of the following sets of
materials:
Station A
Yeast suspension
1.5% H2O2
Petri dish
Dropper pipette
Station B
Slice of potato
1.5% H2O2
Petri dish
Station C
Piece of beef liver
1.5 % H2O2
Petri dish
Forceps
PROCEDURE:
1. Obtain and wear chemical splash goggles.
2. If you are at Station A, use a dropper pipette to place 10 drops of yeast
suspension in the Petri dish. If you are at Station B or C, place a slice of
potato or a piece of liver (use forceps) in the Petri dish.
3. Using the dropper in the H2O2 bottle, add 20 drops of 1.5% H2O2 directly
on the material (yeast, potato, or liver) you placed in the Petri dish.
4. In the space below, describe the reaction that occurs in the dish.
DATA
5. Clean-up by emptying the materials in the Petri dish in a waste container
designated by your teacher. Wash the Petri dish and return it to your lab
station.
6. In a class discussion, share your description of the reaction.
7. As a class, brainstorm what might be causing the bubbles and about the
gas that might be in the bubbles. List any other observations that might
help in explaining the reaction. Summarize these ideas in the space
below. (Use on the back of this page if necessary.)
SUMMARY
EXPLORE:
SET A
What are Peroxisomes?
ENZYME ACTION
Exploring Key Concepts
Peroxisomes are cell organelles found in many kinds of cells. They contain at
least 50 different kinds of enzymes that are involved in a variety of metabolic
reactions. During some important reactions in peroxisomes, hydrogen peroxide,
a molecule toxic to cells, is produced a by-product. Peroxisomes also contain
the enzyme catalase which converts the toxic hydrogen peroxide molecule into
oxygen and water.
Peroxisomes denoted with a P, Image courtesy of Rick Rachubinski.
http://biology.about.com/science/biology/library/weekly/aa051100a.htm
ENZYME ACTION
SET A
Exploring Key Concepts
Question
Why does hydrogen peroxide foam when you put it on a cut? And why
doesn't it foam in the bottle or when I put it on my skin?
Answer
Hydrogen Peroxide (H2O2) is something you can buy at the drugstore. What you
are buying is a 3% solution, meaning the bottle contains 97% water and 3%
hydrogen peroxide. Most people use it as an antiseptic. It turns out it is not very
good as an antiseptic, but it is not bad for washing cuts and scrapes and the
foaming looks cool.
The reason why it foams is because blood and cells contain an enzyme called
catalase. Since a cut or scrape contains both blood and damaged cells, there is
lots of catalase floating around. When the catalase comes in contact with
hydrogen peroxide, it turns the H2O2 into water (H2O) and oxygen gas (O2).
Catalase does this extremely efficiently - up to 200,000 reactions per second.
The bubbles you see in the foam are pure oxygen bubbles being created by the
catalase. Try putting a little hydrogen peroxide on a cut potato and it will do the
same thing for the same reason - catalase in the damaged potato cells reacts
with the hydrogen peroxide.
Hydrogen Peroxide does not foam in the bottle or on your skin because there is
no catalase to help the reaction to occur. Hydrogen peroxide is stable at room
temperature.
http://www.howstuffworks.com/question115.htm
SET A
More juice
from apples
NCBE
Enzymatic juice extraction from apples was introduced about 25 years ago and
today some 3-5 million tons of apples are processed into juice annually
throughout the world.
The commercial production process is as follows;



After they have been crushed, apples are usually left for 20-30 minutes so
that enzyme inhibitors in the pulp are oxidized.
The pulp is then heated to 30 °C before pectinases are added (this
compares with a temperature of 50-60 °C which is needed if enzymes are
not used). Enzyme treatment takes anything from 15 minutes to 2 hours
depending upon the exact nature of the enzyme, the dosage rate, the
reaction temperature and the variety of apple used. Some varieties, like
Golden Delicious, are notoriously difficult to break down. During
incubation, the pectinases degrade soluble pectin in the pulp, making the
juice flow more freely.
Next the apples are pressed. Yields of juice may be increased by up to
20% by enzyme treatment, depending upon the age and variety of apple
used and whether pre-oxidation is employed. Pectinase treatment is
particularly effective with mature apples and those from cold storage.
Significant increases in yield are not usually achieved from fresh, early
season fruit.
http://www.eibe.reading.ac.uk/ncbe/PROTOCOLS/PRACBK/juice.html
http://dgl.microsoft.com/
Set A
1. What are peroxisomes?
2. What chemical reactions occur in peroxisomes?
3. How is hydrogen peroxide used around the home?
4. What is the name of the enzyme found in blood and cells that causes
hydrogen peroxide to foam?
5. What substances is hydrogen peroxide broken into by catalase?
6. What gas is in the foam or bubbles produced by the catalase reaction?
7. How efficient (or fast) is catalase in breaking down hydrogen peroxide?
8. What enzyme is used to get more juice from apples? How does it work?
ENZYME ACTION
SET B
Exploring Key Concepts
The Enzyme-Substrate Reaction
Enzymes catalyze reactions by lowering the activation energy necessary for a
reaction to occur. The molecule that an enzyme acts on is called the substrate. In
an enzyme reaction, substrate molecules are changed, and product is formed.
The enzyme molecule remains unchanged after the reaction, and it can continue
to catalyze the same type of reaction over and over. Figure 1 illustrates the
reaction between the enzyme catalase with the substrate hydrogen peroxide to
form oxygen and water as products.
SUBSTRATE
ENZYME
PRODUCT
catalase
2H2O2
(hydrogen peroxide)
O2 +
(oxygen)
2H2O
(water)
Figure 1
A lab test for detecting the presence of catalase is illustrated in Figure 2.
Figure 2
http://www.labbench.com/sample/concepts.html
SET B
ENZYME ACTION
Exploring Key Concepts
Enzyme Structure
Enzymes are globular proteins. Their folded structure creates an
area known as the active site. The nature and arrangement of
amino acids in the active site make it specific for only one type of
substrate.
Binding Specificity
Even when different substrate molecules are
present, only those that have the specific
shape complementary to the active site are
able to bind with the enzyme's active site.
http://www.labbench.com/sample/active.html
SET B
http://www.lawrysfoods.com/davemayer/adallnatorun1.html
Meat tenderizer, the ant bite miracle
Every year, approximately 5 million Americans are stung and over 25,000
people seek medical attention for painful ant bites. Fire ant bites occur
more frequently than wasp, bee, or hornet stings and are often the most
serious bites if not treated properly.
When fire ant bites occur, they are painful and can cause serious
problems, especially for young children. Although the bite is not life
threatening, a secondary infection caused from broken blisters, known
as pustules, could be.
If applied immediately to an ant bite, Meat Tenderizer will prevent
further infections, swelling, and pustules. Why does Meat Tenderizer
alleviate the sting and swelling; furthermore, preventing infections? A
common ingredient in most Meat Tenderizers includes an enzyme called
Papain. Papain is taken from the famous Papaya fruit. This particular
enzyme breaks down the venom proteins injected by the insect,
rendering their poison harmless.
Meat Tenderizer may be found in any grocery store and is certainly
proven to be an effective household item. Although Meat Tenderizer may
not be a complete cure for ant bites, it is a definite early preventative
for more serious infections.
http://ohoh.essortment.com/whatpapainenzy_runv.htm
Set B
1. Write an equation that summarizes the hydrogen peroxide/catalase
reaction.
2. What gas is found in the bubbles produced during the reaction?
3. Define the term substrate and name the substrate in the hydrogen
peroxide/catalase reaction.
4. What is an enzyme’s “active site”?
5. Can an enzyme bind to any kind of substrate? ___________ Explain.
6. Are enzymes changed during the reactions they catalyze?
7. Are enzymes used up during reactions they catalyze?
8. Explain why meat tenderizers are good for treating ant bites.
SET C
ENZYME ACTION
Exploring Key Concepts
Beef Liver Catalase
Active Site (Heme group and
Several amino acids)
s ds
Amino Acid Chains
Figure 2
http://www.clunet.edu/BioDev/omm/catalase/cat1.htm
http://madsci.wustl.edu/posts/archives/dec96/847573455.Bc.r.html
http://tidepool.st.usm.edu/crswr/catalysis.html
SET C
ENZYME ACTION
Exploring Key Concepts
How does the enzyme catalase break down hydrogen peroxide?
The enzyme catalase carries out the following reaction: H2O2 + H2O2 -> 2 H2O + O2
Catalase takes two molecules of hydrogen peroxide and converts them to two water
molecules plus a molecule of oxygen gas. A common test for the presence of catalase in
material such as cells or tissues involves suspending the material in a dilute solution of
hydrogen peroxide. If the material produces catalase, bubbles form in the solution from
the release of oxygen.
An answer of 'how' catalase carries out this reaction is complicated, and enters the
realm of biochemistry. As with all enzymes, catalase is a protein, meaning that it is
synthesized within the cell from building blocks called amino acids (kind of like pieces
from a complicated leggo set..). In addition to the amino acids that make up the protein,
catalase carries around a heme group and may contain NADP. An iron atom sits in the
middle of the heme group. (see Figure 2 on the next page). You may have heard of
heme before, particularly as hemoglobin - the stuff that carries oxygen in blood and
makes it red. The heme group and some nearby amino acids make up the active site in
the catalase molecule. The active site is the location where bonds in hydrogen peroxide
are broken converting it into water and oxygen.
Enzymes and other catalysts lower the amount of energy necessary for chemical
reactions to occur. This energy is called activation energy. By lowering activation
energy, enzymes make it possible for many reactions to occur in a short period of time.
One catalase active site can breakdown 200,000 hydrogen peroxide molecules per
second. Low activation energy also makes it possible for chemical reactions in living
things to occur at low temperatures. This is important because high temperatures would
cause death or severe damage to living things. Figure 1 illustrates the influence of a
catalyst (such as an enzyme) on the progress of a chemical reaction. Note that the effect
of the catalyst is to lower the activation energy required to initiate the reaction.
Figure 1
SET C
Question: How is cheese
made?
Answer:
Cheese is a solid or semisolid food product prepared from the milk of cows,
ewes, goats or other mammals. The first step in cheese-making is to separate
the milk into curds and whey. Curds are milk solids that consist of casein(a milk
protein), fat, water, and some carbohydrates. Whey is a liquid that contains
water, milk sugar, and whey protein. Bacterial cultures and enzymes are added
to reduce the pH and to prepare the milk for the addition of rennet (Endeavor).
Rennet is added after the cultures and enzymes are added. Rennet is the dried
extract of rennin, the enzyme that is added to help speed up the separation
process. Traditionally, the rennet is derived from the fourth stomach of a suckling
mammal, usually a calf or a lamb(j chem). The next step is to salt the cheese to
add flavor and to aid in curing. Then the cheese is pressed to shape it and to
eliminate more whey. The cheese is now ready to be cured.
There are substitutions for animal derived rennet. Some vegetable juices and
extracts are used as rennin but this is a very expensive alternative so it is seldom
used. Modern advances in science have enabled microorganisms to produce
enzymes that mimic animal rennet. This procedure is less expensive than
vegetable rennet. Common microorganisms include fungi and bacteria. So rennet
can come from animals, vegetables or microorganisms and still taste just as
good.
http://crystal.biol.csufresno.edu:8080/projects/56.html
ENZYME ACTION
SET D
Exploring Key Concepts
Some Factors That Affect Enzyme Action
The shape of an enzyme is maintained by interactions between the various amino acids
that compose it, and this shape is sensitive to changes in the enzyme's environment.
Two important influences are pH and temperature. When an enzyme's shape is
significantly altered because of pH or temperature variation, the enzyme may no longer
catalyze reactions. An enzyme is said to be denatured when it loses its functional shape.
pH and Enzyme Function
Each enzyme functions best within a certain pH range. For example, the enzyme pepsin,
which digests proteins in your stomach, functions best in a strongly acidic environment.
Trypsin, a protein digesting enzyme found in your small intestine, works best in a basic
environment.
When the pH changes, the shape of the active site progressively distorts and affects
enzyme function. When an enzyme is subjected to a pH far from its optimum range, it
may cease to function.
substrate
enzyme
Normal shape at optimum pH
enzyme
Altered shape at pH beyond optimum
Temperature and Enzyme Function
Chemical reactions speed up as temperature is increased, so, in general, enzyme action
will increase at higher temperatures. However, each enzyme has a temperature
optimum, and beyond this point the enzyme's functional shape is lost. Boiling
temperatures will denature or destroy most enzymes.
http://www.labbench.com/sample/active.html
ENZYME ACTION
SET D
Exploring Key Concepts
How and why does temperature, pH level, and enzyme
concentration affect an enzyme's reaction rate?
All chemical reactions are affected by temperature. According to the laws of
thermodynamics, molecular motion increases as temperature increases. The increase
in molecular motion which occurs at high temperatures, such as boiling, causes
enzymes to denature. When an enzyme is denatured, it looses its 3-dimensional shape
and can no longer perform its function. Freezing temperatures also damage enzymes.
100
%
maximum
activity
Figure 1
Enzyme activity as function
of temperature for an
“average” enzyme
0
0
20
40
Temperature (degrees Celsius)
60
pH effects are more complex because they involve chemical reactions with ions. Certain
concentrations of ions in the environment of an enzyme can alter the shape of the
enzyme causing it to loose its ability to catalyze reactions. Because of this, enzymes
can operate only within a specific pH range. Ions can also affect enzyme action by
changing the substrates that enzymes act upon.
Figure 2
Increasing
activity
trypsin
pepsin
0
2
4
Enzyme activity as
a function of pH.
Pepsin and trypsin
are both enzymes
that digest protein.
6
8
10
12
14
pH
The effect of enzyme concentration is easier to understand. Keep in mind that enzymes
are not destroyed or used up during reactions they catalyze. In general, increasing the
amount an enzyme will increase the reaction rate. However, the reaction rate will
eventually become slower when the amount substrate (the material on which the
enzyme is acting) decreases as a result of the action of the enzyme.
Set C
1. Write an equation that summarizes the reaction between hydrogen
peroxide and catalase.
2. What type of molecule is catalase? (protein, fat, carbohydrate, or nucleic
acid)
3. Catalase is made of subunits of _______________________which are
arranged in long chains.
4. What structures make up the active site in catalase?
5. What happens at the active site in catalase?
6. What is activation energy?
9. What do enzymes do to amount of activation energy necessary for
chemical reactions and how is this important to living things?
10. What enzymes are used in the manufacture of cheese? Why are they
used?
Boston® One Step
Liquid Enzymatic
Cleaner is an easy
weekly method for
removing stubborn
protein deposits
from RGP (rigid gas
permeable) lenses
and maintaining
contact lens
wearing comfort.
Convenient liquid
works right in your
lens case as you
disinfect your lenses
so it's easy to use
and saves time.
Most practitioners
recommend using
an enzymatic
cleaner at least
once a week.
Remove protein
deposits while you
disinfect — in one
easy step
Simplifies lens
care. No tablets.
No vials.
Unique SmartDrop
dispenser assures
easy and accurate
dispensing
Clear, odorless
formula
SET D
SET D
The Triple Threat to Stains!
Era Max is formulated with three types of active enzymes
to help fight many stains that active families encounter.
For great stain removal and color-safe whitening, we've
added an advanced stain-fighting enzyme.
Sizes
• 100 oz.
• 200 oz.
Powerful Stain Removal and Pretreating!
Era continues to be the power tool for stain removal and
pretreating. Like Era Max, Era contains advanced enzymes
to fight your family's tough stains and help get your whole
wash clean.
Sizes
• 50 oz.
• 100 oz.
• 150 oz.
• 200 oz.
• 300 oz.
EraserBall to the Rescue!
Era EraserBall is both a pretreating and measuring device
that helps make doing your laundry easy. The EraserBall,
specifically designed for use with Era detergent, is safe for
your washer and clothes.
Included with the following formulas/sizes
• Era Max 200
oz.
• Era 200 oz.
• Era 300 oz.
© 2001 Procter & Gamble. / Valid only in the U.S. Terms & Conditions . Privacy Statement
SET D
1. List three factors that affect the rate of enzyme reactions.
2. How does temperature affect the rate of enzyme reactions?
3. What do you think is the optimum temperature for human catalase?
________ Explain.
4. How does pH affect the rate of enzyme reactions?
5. What do think is the optimum pH for human catalase? ___________
Explain.
6. How do you think adding more catalase to the reaction will affect the rate
of the reaction? ____________________ Explain.
7. Why are enzymes added to contact lens cleaners and laundry detergents?
Let’s Investigate Part A:
Procedure Part A:
Enzyme Action:
Testing Catalase Activity
Many organisms can decompose hydrogen peroxide (H2O2) enzymatically.
Enzymes are globular proteins, responsible for most of the chemical activities of
living organisms. They act as catalysts, as substances that speed up chemical
reactions without being destroyed or altered during the process. Enzymes are
extremely efficient and may be used over and over again. One enzyme may
catalyze thousands of reactions every second. Both the temperature and the pH
at which enzymes function are extremely important. Most organisms have a
preferred temperature range in which they survive, and their enzymes most likely
function best within that temperature range. If the environment of the enzyme is
too acidic or too basic, the enzyme may irreversibly denature, or unravel, until it
no longer has the shape necessary for proper functioning.
H2O2 is toxic to most living organisms. Many organisms are capable of
enzymatically destroying the H2O2 before it can do much damage. H2O2 can be
converted to oxygen and water, as follows:
2 H2O2 → 2 H2O + O2
Although this reaction occurs spontaneously, enzymes increase the rate
considerably. At least two different enzymes are known to catalyze this reaction:
catalase, found in animals and protists, and peroxidase, found in plants. A great
deal can be learned about enzymes by studying the rates of enzyme-catalyzed
reactions.
In Part A of this investigation, you will measure the rate of enzyme activity under
different concentrations of enzyme. In Part B you will design and conduct an
investigation of your own choosing to study a factor such as temperature and pH
which may affect enzyme activity.
It is possible to measure the pressure of oxygen gas formed as H 2O2 is
destroyed. If a plot is made, it may appear similar to that of Figure 1.
rate
Pressure
Time
Figure 1
At the start of the reaction, there is no product, and the pressure is the same as
the atmospheric pressure. After a short time, oxygen accumulates at a rather
constant rate. The slope of the curve at this initial time is constant and is called
the initial rate. As the peroxide is destroyed, less of it is available to react and the
O2 is produced at lower rates. When no more peroxide is left, O2 is no longer
produced.
OBJECTIVES
In this experiment, you will



use a computer and pressure sensor to measure the destruction of
hydrogen peroxide by the enzyme catalase or peroxidase at various
enzyme concentrations.
measure and compare the initial rates of reaction for this enzyme when
different concentrations of enzyme react with H2O2.
design and conduct an investigation to study a factor such as temperature
or pH which affects the destruction of hydrogen peroxide by catalase or
peroxidase.
Pressure Sensor
Figure 2
MATERIALS
Macintosh or IBM-compatible computer
Vernier Interface
Vernier Biology Gas Pressure Sensor
Logger Pro
1-hole rubber stopper assembly
enzyme suspension
four 18 X 150 mm test tubes
3% H2O2
150-mL beaker of water
test tube rack
10-mL graduated cylinder
600-mL beaker
three dropper pipettes
Part A – How Does Enzyme Concentration Affect the
Rate of Enzyme Action
PROCEDURE
1. Obtain and wear goggles.
2. Prepare the computer for data collection by opening “EXP06” from the
Biology with Computers experiment files of Logger Pro. The vertical axis has
pressure scaled from 0.9 to 1.4 atmospheres. The horizontal axis has time
scaled from 0 to 3 minutes. The data rate is set to 50 points/minute.
3. Assemble the apparatus shown in Figure 2.
4. Leave the side valve of the pressure sensor open
when you
start the experiment. Align the blue handle with the
arm
that leads to the pressure sensor as shown in Figure
3.
Figure 3
5. Place four test tubes in a rack and label them 1, 2, 3, and 4. Partially fill a
beaker with
tap water for use in Step 6.
6. Add 3 mL of water and 3 mL of 3% H2O2 to each test tube.
7. Using a clean dropper pipette, add 1 drop of enzyme suspension to Test
Tube 1. Note: Be sure not to let the enzyme fall against the side of the test
tube.
Table 1
Test tube label
Volume of 3% H2O2 (mL)
Volume of water (mL)
1
3
3
2
3
3
3
3
3
4
3
3
8. Insert the 1-hole stopper assembly into the test tube. Note: Firmly twist the
stopper for
an airtight fit.
9. Shake the tube to swirl and thoroughly mix its contents. The reaction should
begin. The next two steps should be completed as rapidly as possible.
10. Close the air valve on the pressure sensor. Align the
blue
handle with the side stem, as shown in Figure 4.
Figure 4
Start measuring the pressure by clicking on the “Collect” button.
11.
12.
If you do not manually stop the program, data collection will end after 3
minutes. If the pressure exceeds 1.3 atmospheres, the pressure inside the
tube will be too great and the rubber stopper is likely to pop off. If the
pressure exceeds 1.3 atmospheres, stop the data collection by clicking on the
Stop button and open the air valve on the pressure sensor.
13. Find the rate of enzyme activity:



While holding down on the left mouse button, drag the cursor over the
initial part of the curve that you want to analyze. The part of the curve you
selected will appear in a dashed-line box.
Click the “R=” button
Record the value of the slope of the line (m) in Table 2.
Figure 5: Note the highlighted region of
the graph
14. Find the rate of enzyme activity for Test Tubes 2 – 4: Make sure you “store”
the run after each test

Add 2 drops of the enzyme solution to Test Tube 2. Repeat Steps 7 – 13.

Add 3 drops of the enzyme solution to Test Tube 3. Repeat Steps 7 – 13.

Add 4 drops of the enzyme solution to Test Tube 4. Repeat Steps 7 – 13.
15. Draw your results on the graph on the following page. Be sure to label each
line.
DATA
Table 2
Tube #
Number of Drops of
Enzyme Solution
1
1
2
2
3
3
4
4
Slope (m) or Rate in
Atm/min
ANALYZING THE DATA
1. How does changing the concentration affect the rate of decomposition of
H2O2?
2. If you increased the concentration of enzyme to 5 drops, what do you think
would happen to the rate of the reaction? Predict what the rate (slope of the
line) would be for 5 drops.
3. Based on your knowledge of enzyme action, explain why the increased
concentration of enzyme results in an increased reaction rate.
Let’s Investigate Part B:
Procedure Part B:
You have now measured the effects of the enzyme concentration on the rate of
the enzyme activity of catalase. It is now time to try your hand at designing your
own experiment to answer a question you pose about the process of enzyme
catalyzed reactions. You should select a variable to investigate that you think
might influence this rate.
1. Ask a Question
What question might you ask about the rate of
enzyme reactions that could be answered in an experiment that uses the
catalase enzyme?
2. Background Information What background information leads you to
believe that the variable you have identified in your question will affect the
rate of enzyme catalysis?
3. Hypothesis Write a hypothesis that will be useful in guiding your
investigation. Remember, a hypothesis is a tentative solution to your
question. It states the results you expect to get when you conduct the
experiment.
4. Design Your Experiment Tell what steps you will follow in conducting
your experiment. Include a drawing of your apparatus. Be sure to
describe your control.
5. Variables What is your independent variable?
What is your dependent variable?
What are your constants?
6. Data
7. Graph
Draw a data table for recording your data.
Draw your results on the graph below.
8. Analysis and Conclusions Do your data support or refute your
hypothesis? Explain your answer with evidence from your data and
observations.
9. Possible Errors It is possible that you made errors in your experimental
design, in setting up your equipment, or in collecting and recording data.
Please discuss these errors and explain how you would conduct the
experiment if you had to do it over again.
10. Other Questions The answers obtained from an experiment often lead
scientists to ask additional questions about the topic they are studying.
State at least one question about enzyme catalyzed reactions that arises
from your experiment that you would like to investigate if you had the
chance.
Investigating Further…
Yogurt is produced through fermentation catalyzed by enzymes in bacteria. Research
yogurt production and propose the optimal conditions under which the enzymes could
operate in order to produce the greatest quantity/quality of product.
Summary of Activity…
Select a commercial product that is either commercially produced through enzymatic
processes, or one that is a commercially prepared enzyme. Write a description of the
enzyme’s chemical activity (action of enzyme on the substrate); including optimum
conditions for production or activity as well as any other interesting pieces of information.
Applying what you have learned…
Stone washed blue jeans were actually made by washing them with pumice
stones. This process has created a disposal problem, so the biotechnology
industry came up with a solution. Research the application of enzymes to the
“stone-washing” industry and report your findings. Include enzyme(s) used in the
stone-washing process; how they are produced in quantity for those purposes;
what the enzyme(s) actually do in the “stone-washing” process; what would be
the optimal conditions for the enzyme(s) to do their work?