Experiment to Investigate how enzyme concentration can affect the initial rate of

reaction

Aim:
To investigate how enzyme concentration can affect the initial rate of reaction by
using hydrogen peroxide and potato disks.
Hypothesis:
I predict that there will be more oxygen and a higher initial rate of reaction when a
larger amount of potato disks will be used to catalyse the hydrogen peroxide. This will
mean that the Catalase found in potatoes will react with the hydrogen peroxide,
causing it to breakdown into water and oxygen, shown by the following equation:

2H2O2

2H2O + O2

Method:

Mount a test tube that will serve as the reaction tube, in a water bath using a
clamp. Place a rubber bung with a delivery tube connected to it at the tip of the
reaction tube to plug it. Secondly, clamp a beaker outside the water bath with
distilled water in it. Place a syringe or collecting tube into the beaker upside
down so it is submerged in the water, and completely filled up with it.
Remember to mount this correctly, ensuring that no air bubbles, or an air gap
is left at the top of the colleting tube. Finally, place the delivery tube into the
inverted tube, to allow bubbles of oxygen to later fill the collecting tube up.

Using a scalpel, cut 10 discs of potato, each 0.2mm thick, use a cork borer to
ensure that the disks are the same size. Place these in the reaction tube along
with 5cm3 of buffer solution.

Using a syringe add 5cm3 of hydrogen peroxide solution to the potato discs.
After that, immediately place the rubber bung and delivery tube firmly into the
reaction tube. Make sure the open end of the delivery tube is firmly placed in
the collecting tube.

Start a timer as soon as the first bubble of oxygen leaves the delivery tube into
the collecting tube. Collect the gas produced for 3 minutes. Shake the solution
to ensure the contents has fully mixed and that the reaction occurs efficiently
through the 3-minute period. If no oxygen bubble is produced within three
minutes of adding the hydrogen peroxide record a zero value for the given test.

Jan Stankiewicz 12PH

At the end of the 3-minute period, you can disconnect the rubber bung to stop
any more oxygen entering it, and then measure the volume of oxygen
produced in the 3-minute period. Remember to place the collecting tube in
such a way that when you read off the volume, you do not get any parallax
error.

Wash out the reaction tube thoroughly, ensuring all the hydrogen peroxide has
been emptied.

Repeat the steps 2-6 using a range of different number of potato discs e.g.
10,15,20... , ensuring that other conditions are unchanged. Dismount the
collecting tube, empty the tube of gas and water and repeat the water filling
process.

Results:
Number of potato
discs

Volume of oxygen
collected/ cm3

10

2.0

15

4.7

20

7.8

25

8.5

30

9.7

Jan Stankiewicz 12PH

Logarithm graph showing the volume of oxygen produced with the

number of potatoes

Conclusion:
Catalase is a protein molecule, which is found in living cells, such as blood cells. It is
used to speed up specific reactions within the cell. One of its functions, is to remove a
by-product of metabolism called hydrogen peroxide. This strong oxidizing agent can
be found in our cells, and if not removed, can be poisonous. Outside the human body,
the enzyme Catalase can be found in potatoes. If these potatoes are mixed with the
hydrogen peroxide, Catalase from the potatoes will catalyse the reaction of the
breakdown of hydrogen peroxide, 2H 2O2
2H 2O + O2. As shown in the
reaction, the decomposition of the compound forms water and oxygen. Catalase
works in such a way that the shape of its active site exactly matches the shape of the
substrate molecule (hydrogen peroxide), which forms temporary bonds between the
hydrogen peroxide and the Catalase. As the bonds are forms, the hydrogen peroxide
molecules change into a slightly different shape than the original shape of the
molecule, making it easier for the bonds to be broken. These bonds then break and
result in a product being formed (oxygen and water). From the results obtained, we
can see that there is a general trend throughout the results. There is an directly
proportional increase in the results, showing that the more potato discs used, the more
oxygen is produced from the reaction. This is because the more potato discs added, a
higher concentration of Catalase is then used, causing a higher rate of reaction and
more oxygen to be formed form the decomposition of hydrogen peroxide. A buffer
solution of pH 7 was used to keep the pH levels the same in all the reactions. All the
reactions were also placed in a water bath with the same temperature of 30 oC
Jan Stankiewicz 12PH

throughout. Looking at the logarithmic scale we can see that there is a big increase in
the volume obtained until the reaction where 20 discs of potato are used. During the
20, 25 and 30 disc reactions, there is not that much of an increase compared to the
reactions using 10 and 15 discs of potato. This is because the hydrogen peroxide in
the reaction is the limiting reagent, and the more potato discs use, the more is used up
constantly, so it will react will all the Catalase within the potato discs, eventually
slowing the reaction down. From these speculations and results, we can confirm my
hypothesis that predicted what happened in the experiment.
Evaluation:
The experiment has proven to have been successful, however there are a few
modifications that can be made to improve the accuracy of the results and reliability
of them. Firstly, we can use a more accurate measuring system for the collection of
oxygen. The accuracy of the collecting tube is only relevant to .1mm 3. This can
provide inaccuracies as small amounts of oxygen can be produced that will not be
noted in the results as accurately. This can be solved by using a thinner collecting tube
that has a much more spread out, and more accurate scale. Another problem is how
the oxygen volume is read from the collection tube. Moving up and down can cause a
parallax error that will alter the results, therefore the collecting tube has to be put
horizontally against a medium with a marking point to avoid this error. Another error
could be the plugging of the boiling tube (reaction tube) with the rubber bung after
adding the potato discs. This can let some oxygen immediately escape, making results
unreliable and less valid.

Jan Stankiewicz 12PH