Download The Affect of Enzymes on a Chemical Reaction

Running head: REACTION RATES OVER TIME
The Reaction Rates of Enzyme-Catalyzed Reactions Over Time
Trevor M. Dopp
Madison High School
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Abstract
Enzymes are proteins that catalyze chemical reactions. Catalase catalyzes the process
of breaking down hydrogen peroxide into water and oxygen. Enzyme-catalyzed reactions
were created in order to measure the reaction rates over time. Catalase was added to hydrogen
peroxide solutions and bubbles were formed showing that a reaction was occurring. Each
solution was allowed to react for a different amount of time before being stopped by adding
sulfuric acid. Each solution was then titrated using potassium promaganate to measure the
amount of hydrogen peroxide used up in each solution. Using the equation
y 2  y1
, the
x 2  x1
reaction rates over each time interval were calculated. The data showed that the reaction rate
of the catalyzed reaction was nearly constant at the beginning of the reaction but slowed down
to almost a stop as the substrate was used up. Further study could be done to test the affects of
temperature on enzyme-catalyzed reactions to show how body temp could inhibit the function
of enzymes in the body.
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Introduction
Enzymes are proteins that catalyze chemical reactions. They have active sites where
substrates attach to the enzyme and a chemical reaction occurs in which a product is
produced. Although a chemical reaction occurs, the enzyme remains unchanged. Salt
concentration, pH, temperature, and competitive and noncompetitive inhibitors can inhibit the
effectiveness of an enzyme (Ap biology lab, 2001). If these factors are not at their optimal
levels the enzyme will become ineffective and may even denature. A change in pH,
temperature, or salt concentration will cause the enzyme to denature. Competitive inhibitors
will mimic the structure of the substrate and trick the enzyme into thinking that it is the
substrate. Noncompetitive inhibitors will attach to the enzyme and alter the shape of its active
site, not allowing for an induced fit with the substrate. “Biological systems utilize free energy
and molecular building blocks to grow, to reproduce, and to maintain dynamic homeostasis”
through the use of enzymes (Campbell, Reece, Urry, Cain, Wasserman, Minorsky & Jackson,
2008). Enzymes are proteins, which are macromolecules that have many functions including
speeding up chemical reaction in the form of enzymes. An organism uses enzymes to catalyze
chemical reactions in the body so the reactions can occur quickly in order to maintain
homeostasis.
This experiment was done in order to measure the reaction rates of enzyme-catalyzed
reactions that occurred for different time lengths (Ap biology lab, 2001). It should show the
change in reaction rate of an enzyme-catalyzed reaction over a specific period of time, which
should be almost constant at first but slow down to almost zero over time (Ap biology lab,
2001). It should also show the amount of hydrogen peroxide that is converted to water and
oxygen over specific periods of time due to enzyme catalysis.
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If an enzyme is added to its corresponding substrate solution then the reaction rate of
the solution will increase at first but the reaction will slow down over time as the substrate is
used up, because enzymes catalyze chemical reactions. Catalase, the enzyme used in this lab,
contains four polypeptide chains each made up of over 500 amino acids (Ap biology lab,
2001). Catalase regulates hydrogen peroxide levels in the body, so adding it to a solution
composed of its corresponding substrate, hydrogen peroxide, will speed up the chemical
reaction of breaking down hydrogen peroxide into water and oxygen. The enzyme and
substrate have an induced fit at the active site where they react. In order to stop the reaction
after the desired period of time, sulfuric acid will be added to the solution, which will
denature the enzyme and stop the reaction.
Materials and Methods
In order to conduct this lab I used 11 Dixie cups, a glass beaker, a burette, two
syringes, a stopwatch, potassium promaganate, catalase, sulfuric acid, hydrogen peroxide
latex gloves, goggles, a marker, tape, and my lab notebook. First I used the tape and marker to
label each Dixie cup as follows: 0 sec, 10 sec, 30 sec, 60 sec, 90 sec, 120 sec, 180 sec, 360
sec, H2O2, catalase, and titration. I filled the H2O2 Dixie cup with hydrogen peroxide, the
catalase Dixie cup with catalase, and the beaker with sulfuric acid. I then used a syringe to put
10mL of H2O2 in each of the Dixie cups labeled 0 sec, 10 sec, 30 sec, 60 sec, 90 sec, 120 sec,
180 sec, and 360 sec. I cleaned out the syringe I used for the hydrogen peroxide and filled it
with 1mL of catalase and filled the other syringe with 10mL of sulfuric acid. I started with the
Dixie cup labeled 10 sec and started the stopwatch at the same time I put the 1mL of catalase
in the solution and stirred the solution for the designated time labeled on the Dixie cup. After
the designated time I put the 10mL of sulfuric acid in the solution to stop the reaction. I
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repeated these steps with the other Dixie cups labeled with a specific time on them. When
doing the 0 sec Dixie cup I did not add any catalase, but I did add the 10mL of sulfuric acid
(this solution will be used to find the base line). In order to find the base line I filled my
burette with potassium promaganate and recorded the initial reading of the burette in my lab
notebook. 5 mL of the 0 sec solution was placed in the Dixie cup labeled “titration” using a
clean syringe. After placing the solution under the burette I added the potassium promaganate
to the solution drop by drop until the solution changed color. I recorded the final reading of
the burette in my lab notebook. This is the base line assay. I repeated the previous steps for
each of the solutions I created in order to find the amount of hydrogen peroxide used in each
reaction. Because I thought there may have been an error in titrating the 90sec solution, I
redid the titration for that solution.
The control in this lab is the 0 sec Dixie cup solution because no catalase was added to
it. The independent variable is the time each enzyme-catalyzed reaction occurred for, while
the dependent variable is the amount of hydrogen peroxide that is used up in the reaction. In
order to calculate the amount of potassium promaganate used in each solution the following
formula was used: final reading of the burette minus the initial reading of the burette. The
answer to the previous formula was then subtracted from the base line assay to get the amount
of hydrogen peroxide that was used up in each reaction. These formulas should show that the
longer the enzyme-catalyzed reaction occurred, the more hydrogen peroxide was used. The
amount of hydrogen peroxide used should always be less than the base line assay as well. The
reaction rate is found by using the equation
y 2  y1
.
x 2  x1
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Results
When titrating each sample of the hydrogen peroxide solution, a logistic trend was
found. The longer the catalase reacted with the hydrogen peroxide solution, the more
hydrogen peroxide was used up. It was found that the rate of reaction of the catalase and
hydrogen peroxide was almost constant at first, but slowed down over time. While the 10sec
solution consumed 2.5mL of potassium promaganate, the 360sec solution only consumed
0.01mL of potassium promaganate.
Titration of H2O2
Table 1
Reaction Rate of Enzyme-Catalyzed Reaction
Table 2
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Figure 1
Discussion
Enzymes have an induced fit with their corresponding substrate that allows them to
efficiently react with the substrate. If an enzyme is added to its corresponding substrate
solution then the reaction rate of the solution will increase at first but slow down over time as
the substrate is used up, because enzymes catalyze chemical reactions. The data collected
supports this hypothesis. The enzyme used was catalase and its corresponding substrate is
hydrogen peroxide. It was observed that when 1mL of catalase was added to 10mL of
hydrogen peroxide, bubbles formed showing that a chemical reaction was occurring. It was
expected that the reaction rate of the enzyme-catalyzed reaction would slow down to close to
zero as more hydrogen peroxide was used up. This was also supported with the data. The
titration showed that the reaction rate from time 0 seconds to 10 seconds was 0.05 mL/sec. As
more hydrogen peroxide was used up over time, the reaction rate slowed to 0.0005 mL/sec
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from time 180 seconds to 360 seconds. An error that could have occurred with the titration of
the solutions was a time constraint. Doing the titration a day after catalyzing the hydrogen
peroxide solution could affect the results because the hydrogen peroxide may have broken
down even more within a day.
It would be beneficial to test the affects of temperature on the enzyme-catalyzed
solution in order to see how body temperature affects enzyme function and show the affects
that long-term fevers can have on enzymes. This could be done by repeating the same
procedure done in this lab but making a few changes. Instead of catalyzing each hydrogen
peroxide solution for different amounts of time, each solution would be held at a different
temperature. Each solution could be catalyzed for 10 seconds but the temperature of each
solution would vary. For example the solutions could be set at 0oC, 20 oC, 40 oC, 60 oC, 80 oC,
and 100 oC using a hot plate and ice bath. The titration should show that the enzyme works
best at or around body temperature, which is 37oC. If the enzyme is too cold, it will not
function and when the enzyme becomes too hot, it will denature and become ineffective.
Conclusion
This lab was conducted to measure the reaction rate of an enzyme-catalyzed reaction
over time. The results of the lab show that enzyme-catalyzed reactions are almost constant at
first, but slow down over time as the substrate is used up. The idea that sulfuric acid denatures
enzymes and stops chemical reactions was also supported by the observations. It was also
shown that catalase does indeed catalyze the process of breaking down hydrogen peroxide
into water and oxygen.
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References
(2001). Ap biology lab manual for students. (lab 2 enzyme catalysis). College Entrance
Examination Board.
Campbell, N., Reece, J., Urry, L., Cain, M., Wasserman, S., Minorsky, P., & Jackson, R.
(2008). Ap edition biology. (Eighth ed.). San Francisco, CA: Benjamin Cummings.