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The Effect of Hydrogen Peroxide on Catalase with Varied Temperature

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The Effect of Hydrogen Peroxide on Catalase with Varied Temperature

Effect of temperature of the action of the Enzyme Catalase.


Background Knowledge

An enzyme is a biological catalyst, it alter the rate of reaction
without being changed itself. Enzymes are proteins; they have a very
precise three-dimensional shape, which forms a one specific active
site on the enzyme. Each enzyme can only convert one kind of substrate
molecule in to one kind of product molecule. These are specific.

What affects Enzymes?

· Temperature- Enzymes stop working if the temperature rises above
40ºC. Increasing the temperature alters the 3D shape and so the enzyme
can no longer fit the substrate.

· pH- They work best in neutral conditions neither acidic nor

What affect does catalase have?

Catalase is a very fast reacting enzyme, it is found in many living
cells, it breaks down hydrogen peroxide to water and oxygen. In fact
one molecule of it can deal with six million molecules of hydrogen
peroxide in 1 minute. Hydrogen peroxide is toxic so needs to be
changed into harmless substances.

Hydrogen peroxide water + oxygen
2H2O2 2H2O + O2

References to practicals referring to enzymes

· Biology for You Pg 30 - Experiment 3.1

From looking at this I found out that catalase reacts with hydrogen
peroxide to give out water and oxygen. Oxygen bubbles produce froth on
the surface of the solution. In my forthcoming experiment I will
expect to see froth being produced.

· Biology- Nelson Science Pg 25 - Picture 4

From looking at this graph, see below. I have learnt that the affect
of temperature does in fact change the rate of reaction. From the
graph the reaction reaches 40ºC but then denatures and the rate of the
reaction decreases. The rate falls rapidly suggesting denaturing.
Taking this information into account I would expect the enzyme
catalase to show a similar pattern with respect to the temperature.

In order to observe the effect of temperature on catalase we will be
maintaining in the amount of oxygen released. The oxygen produces a
froth which we will then measure in mm and the volume of oxygen given
off which will be measure in cm³

Method- measuring the height of froth and volume of oxygen

1. Put work shirt on and goggles on. Carry out the rest of safety
2. Gather equipment as shown on diagram1.
3. Using a cork borer make 5 cylinders from the large potato.
4. Cut them into all the same length (6cm)
5. Using a pestle and mortar mash up each cylinder separately.
6. Measure 25ml of hydrogen peroxide using a measuring cylinder.
7. Select the temperature you are going to study

0ºC- iced water
25ºC-no extra equipment
37ºC-water bath required
55ºC-water bath required
100ºC-beaker of boiling water

8. Place on mashed cylinder into a boiling tube add the measured
hydrogen peroxide and attach the rubber bung connected to the
measuring syringe.
9. Start stop watch and record volume of gas collected every 30
seconds. At the same time measure the amount of froth produced at 30
seconds intervals


· 5 beakers
· 5 test tubes
· Thermometers
· Cork borer
· Potato
· Ruler
· Knife
· Tile
· Measuring syringe
· Heat proof mat
· Bunsen burner
· Tri-pod
· Wire gauze
· Pestle and mortar
· Hydrogen peroxide
· Matches
· Spills
· Ice cubes
· Water bath
· Goggles
· Spatula
· Stopwatch
· Measuring cylinder

Fair test

In this investigation I will keep constant the following

· The surface area of the potato. I will use the mashed up form as it
will be a faster reaction as there is more area to react on, as we
have to consider the time span.
· The same volume of hydrogen peroxide in each part of the
· The same size equipment e.g. boiling tubes as the readings for the
results will be wrong if this is not constant.
· Use the same method for each experiment so that there won´t be any
major differences. Only alter the temperature.
· Keep the amount of potato the same amount.
· Measure the temperature with a thermometer.


In order to make my investigation go to plan I will be as accurate as
I can be so I will measure to the correct measuring size.

· Measure the volume in cm³ and amount of potato in grams to make sure
that they are exactly the same mass before using them in the
· Do the experiment three times to ensure that there isn´t an odd
result. Three is a good number to use as you can see if there is one
odd one where if you just done the experiment twice then you wouldn´t
know which one odd and which isn´t.
· Also to average out the results.

Safety precautions

· Wear goggles
· Tuck tie in skirt
· Wear work shirt
· Handle the hydrogen peroxide with care as it is corrosive and an

Predictions and Reasons

From my research I think that the enzymes will denature after 40ºC and
any other temperature above that. Reason being that enzymes are
proteins and their structure is three-dimensional. Increasing the
temperature disturbs the intra molecular bonds that hold the 3D shape.
Because of this the shape is altered. Enzymes have an active site.
This fits into the substrate molecular (see diagram2-lock and key). If
the active site is altered the substrate will no longer fit in and so
the enzyme doesn´t work properly.
The rise of reaction rate is also due to the increase in temperature,
relating to the kinetic theory. The higher the temperature, the faster
they move. This happens but only to an optimum of 40ºC. The curve
leading up to the optimum point is gradual but as it is reached it
falls dramatically. The reason being that the active site is destroyed
therefore no reaction can take place as there is only one specific
active site per substrate.


Below are my table of results which show the height of froth produced
in cm and the volume of oxygen in cm³ for each of the three tests at
each of the five temperatures studied.

0.5 3 3 2.4 9 2 4
1 3.7 6 3 10 3 8
1.5 4.2 8 3.3 11 4.3 12
2 4.8 10 3.5 12 5.4 12
2.5 5.3 11 3.9 13 6 12
3 5.7 12 4 13 6.2 13
3.5 6.5 12 4.2 13 7.4 13
4 6.8 13 4.4 13 8 14
4.5 7.5 13 4.4 13 8 14
5 8.2 13 4.4 13 8 14



0.5 3 9 4 5 3 8
1 5 14 6 10 4.9 12
1.5 6 18 6.5 14 5.8 15
2 7.5 20 7 18 7.6 19
2.5 9 20 8 20 8.2 20
3 10 20 9 21 9.1 21
3.5 10 20 9 21 10 22
4 10 20 9 21 10 22
4.5 10 20 9 21 10 22
5 10 20 9 21 10 22



0.5 4 7 5 12 4.5 10
1 5.5 14 8 20 6 16
1.5 7 19 10 26 8 22
2 9 22 11 28 10 26
2.5 10 28 12 30 11 28
3 10 28 12 30 11 28
3.5 10 28 12 30 11 28
4 10 28 12 30 11 28
4.5 10 28 12 30 11 28
5 10 28 12 30 11 28


0.5 4 12 5 14 6 15
1 6 18 6 19 7 20
1.5 7 22 6.5 22 8 22
2 8 24 8 24 8 24
2.5 8 25 8 25 8 25
3 8 26 8 25 8 26
3.5 8 26 8 26 8 26
4 8 26 8 26 8 26
4.5 8 26 8 26 8 26
5 8 26 8 26 8 26


0.5 0.1 0.5 0.1 1 0.1 1
1 0.1 0.5 0.1 1 0.1 1
1.5 0.1 0.5 0.1 1 0.1 1
2 0.1 0.5 0.1 1 0.1 1
2.5 0.1 0.5 0.1 1 0.1 1
3 0.1 0.5 0.1 1 0.1 1
3.5 0.1 0.5 0.1 1 0.1 1
4 0.1 0.5 0.1 1 0.1 1
4.5 0.1 0.5 0.1 1 0.1 1
5 0.1 0.5 0.1 1 0.1 1

Table of averages from each of the above temperatures


0.5 2.5 5.0
1 3.2 8.0
1.5 3.9 10.3
2 4.6 11.3
2.5 5.1 12.0
3 5.3 12.7
3.5 6.0 12.7
4 6.4 13.3
4.5 6.6 13.3
5 6.9 13.3


0.5 3.3 7.3
1 5.3 12.0
1.5 6.1 15.7
2 7.4 19.0
2.5 8.4 20.0
3 9.4 20.6
3.5 9.7 21.0
4 9.7 21.0
4.5 9.7 21.0
5 9.7 21.0


0.5 4.5 9.7
1 6.5 16.6
1.5 8.5 22.3
2 10 25.3
2.5 10 28.7
3 10 28.7
3.5 10 28.7
4 10 28.7
4.5 10 28.7
5 10 28.7


0.5 5.0 13.7
1 6.3 19.0
1.5 7.1 22.0
2 8.0 24.0
2.5 8.0 25.0
3 8.0 25.7
3.5 8.0 26.0
4 8.0 26.0
4.5 8.0 26.0
5 8.0 26.0


0.5 0.1 0.83
1 0.1 0.83
1.5 0.1 0.83
2 0.1 0.83
2.5 0.1 0.83
3 0.1 0.83
3.5 0.1 0.83
4 0.1 0.83
4.5 0.1 0.83
5 0.1 0.83

These two tables show the average measurement that we recorded for
each temperature.


Time (mins)
Temperature (ºC) 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 5
10ºC 0 2.5 3.2 3.9 4.6 5.0 5.3 6.0 6.4 6.6 6.9
25ºC 0 3.3 5.3 6.1 7.4 8.4 9.4 9.4 9.4 9.4 9.4
37ºC 0 4.5 6.5 8.5 10 10 10 10 10 10 10
55ºC 0 5.0 6.3 7.1 8 8 8 8 8 8 8
100ºC 0 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1 0.1


Time (mins)
Temperature (ºC) 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 5
10ºC 0 5.3 8 10.3 11.3 12 12.7 12.7 13.3 13.3 13.3
25ºC 0 7.3 12 26.7 19 20 20.6 21 21 21 21
37ºC 0 9.7 16.6 22.3 25.3 26.7 26.7 26.7 26.7 26.7 26.7
55ºC 0 13.7 19 22 24 25 26.7 26 26 26 26
100ºC 0 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8

Analysing results and Conclusion

From my results it appears that catalase works best at 37ºC, and it is
virtually denatured at boiling point.
Looking at the initial part of the reaction (see graph 1) it is clear
that the gradient at the beginning gets steeper when looking at the
temperatures between 10ºC-55ºC. At each temperature the line levels
off towards the end of five minutes. Looking at graph 2, there is a
steady rise in height of froth up to 37ºC and then a gradual fall up
to 100ºC.
Looking at my background knowledge and prior experiments using enzymes
I can explain my results as follows.
Kinetic theory states that particles, which gain heat energy, move
more quickly. In our case the reacting particles are the substrate
(hydrogen peroxide) and the enzyme catalase. As the temperature is
increased the particles of hydrogen peroxide have more energy
therefore they collide with the potato more frequently and so
increasing the rate at which the product is formed. However at a
certain temperature this is no longer the case. This is because
enzymes are proteins and proteins can be denatured at high
temperatures. This is because proteins have a 3D shape. In our case
the catalase has a certain shape that the substrate fits into. At high
temperatures the active site on the enzyme is altered, see diagram

(Diagram showing active site on the enzyme is altered therefore
stopping products being formed)

This stops the substrate from 'fitting´ and so no product is formed.
My results do not totally support or undermine my original prediction.
The reason being that on graph 1, my results suit my prediction. It
shows that the temperature, 37ºC was the fastest and 100ºC is when the
enzyme denatures. But in graph 2, my results undermine my original
prediction as at 55ºC the reaction still takes place where as in my
prediction I stated that enzymes would denature at 40ºC approximately,
I didn´t expect this is happen.


In my investigation I was pleased with my achievements.
In my method, keeping the temperature constant throughout the
investigation was hard to maintain, as the temperature of the contents
of the tube would change quite quickly and therefore the hydrogen
peroxide wouldn´t be at the temperature required. To overcome this
problem I could keep the test tubes in a hot water bath for all the
temperatures making sure that the water bath was the suitable depth.
This would ensure constant temperature throughout the whole 5 mins.
Also another problem that I encountered was to keep the height of the
froth fair. I measured the height of the froth with a 30cm ruler
against the test tube rack, with the support of my hand. As I was
measuring, my hand would move from time to time and therefore didn´t
know where I should place my ruler afterwards. To over come this I
should attach the ruler onto the test tube rack with cello tape, as it
is transparent or maybe use a pointer.
With respect to I measured the height of froth in cm, but to be more
precise I should have measured it in mm. To over come this I should
use a ruler with mm readings. Also another problem that I observed on
accuracy was that I didn´t allow the temperature to equilibrate to the
right temperature. In this case I wasn´t using the correct temperature
that I wanted, this could have led to some anomalous results. Ideally
I should have brought the temperature of the hydrogen peroxide up to
the needed temperature before adding to the potato.
Looking back at my results I found some anomalous results in my
findings. When averaging I used these results, which could of made the
average either lower or higher than it should be. To improve this I
should have missed these results. Not including some sets of results
when making averages may have led to better values.
My results are in line with those I predicted. Graphs indicate rise in
temperature up a point leads to an increase in oxygen production. This
is in line with kinetic theory. However it is very clear that after a
certain temperature is reached the enzyme actually virtually stops.
This supports my theory of lock and key fit.
However optimum activity of enzyme is at about 37ºC this is as we
expected. But at 55ºC the enzyme is still not denatured according to
my results. This is a higher temperature than I would expect. Possible
not allowing solutions to reach temperatures selected has led to an
inaccuracy. It may be that in fact that many temperatures of solutions
were lower than we stated.
Overall, due to reliable repeats and in general predictions being
confirmed I feel my results are reliable enough to make a conclusion.
The obvious thing I would improve about the measurements I made would
be to increase the range of temperatures used. Especially between
55ºC-100ºC. In this way it may be clearer at the temperature which
denaturing took place, and would possibly give a graph that resembled
the graph in background knowledge.
Another way of improving this investigation is to change the method. I
measure the volume of oxygen that was produced. In order to get pure
oxygen without any other gases that are in the air I would use the
same equipment but make sure that the gap between the rubber bung and
solution was free from any other gases.

The Effect Or Varying Enzyme Concentration On The Breakdown Of
Hydrogen Peroxide In The Presence of

Hypothesis - Hydrogen peroxide will breakdown to oxygen and water in
the presence of Catalase. The reaction will increase with increasing
enzyme concentration when molecules of hydrogen peroxide are freely
available. However, when molecules of the substrate are in short
supply, the increase in rate of reaction is limited and will have
little effect.

Variables - In this investigation, the variables that affect the
activity of the enzyme, Catalase, were considered and controlled so
that they would not disrupt the success of the experiment.

i) Temperature - As temperature increases, molecules move faster
(kinetic theory). In an enzyme catalysed reaction, such as the
decomposition of hydrogen peroxide, this increases the rate at which
the enzyme and substrate molecules meet and therefore the rate at
which the products are formed. As the temperature continues to rise,
however, the hydrogen and ionic bonds, which hold the enzyme molecules
in shape, are broken. If the molecular structure is disrupted, the
enzyme ceases to function as the active site no longer accommodates
the substrate. The enzyme is denatured.

To control this variable, the temperature was maintained at a fairly
constant level that allowed the enzyme to work effectively (room
temperature, approximately 23°C). This was achieved by using a test
tube rack and tongs to handle the apparatus so that the heat from my
hands did not affect the Catalase.

ii) pH - Any change in pH affects the ionic and hydrogen bonding in an
enzyme and so alters it shape. Each enzyme has an optimum pH at which
its active site best fits the substrate. Variation either side of pH
results in denaturation of the enzyme and a slower rate of reaction.

In this experiment, the pH was kept constant using a pH 7 buffer,
selected to maintain a pH level suited to the enzyme by being equal to
the natural environment of the enzyme (potato tissue).

iii) Substrate Concentration - When there is an excess of enzyme
molecules, an increase in the substrate concentration, produces a
corresponding increase in the rate of reaction. If there are
sufficient substrate molecules to occupy all of the enzymes' active
sites, the rate of reaction is unaffected by further increases in
substrate concentration as the enzymes are unable to break down the
greater quantity of substrate.

To control the substrate concentration, identical quantities of the
substrate were used for each reading. To ensure that this was measured
precisely, 5ml syringes were used to accurately gauge to exact

iv) Inhibition - Inhibitors compete with the substrate for the active
sites of the enzyme (competitive inhibitors) or attach themselves to
the enzyme, altering the shape of the active site so that the
substrate is unable to occupy it and the enzyme cannot function
(non-competitive inhibitors). Inhibitors therefore slow the rate of
reaction. They should not have affected this investigation, however,
as none were added.

v) Enzyme cofactors - cofactors are none protein substances which
influence the functioning of enzymes. They include activators that are
essential for the activation of some enzymes. Coenzymes also influence
the functioning of enzymes although are not bonded to the enzyme.

Unless enzyme cofactors were present in the potato tissue containing
the Catalase, they were not included in this investigation and
therefore would not have affected the rate of reaction and the results
of this experiment.

vi) Enzyme Concentration - Provided there is an excess substrate, an
increase in enzyme concentration will lead to a corresponding increase
in rate of reaction. Where the substrate is in short supply (i.e. it
is limiting) an increase in enzyme concentration has no effect.

I varied the enzyme concentration by altering the number of equal
sized discs of potato that contain the Catalase, in the reaction. The
greater the number of discs, the greater the enzyme concentration.

Apparatus -

i) A manometer

ii) 30ml hydrogen peroxide

iii) Manometer fluid

iv) 6 boiling tubes

v) Tongs

vi) A test tube rack

vii) A potato

viii) A petri dish

ix) A cork borer

x) Distilled water

xi) A razor blade

xii) A stop watch

xiii) A ruler

xiv) Rubber tubing

xv) A marker pen

xvi) A clamp

xvii) A stop watch

xviii) 2 5ml syringes

xix) pH 7 buffer

xx) A bung

Procedures - Three tubes, 10mm in diameter were bored from a potato
using a cork borer. Using a razor blade and a ruler, 122 discs, 1mm
thick, were cut from the tubes and placed under distilled water in a
petri dish. This prevented the potato from being contaminated or

5ml of hydrogen peroxide and 5ml of a pH 7 buffer were then measured
and added to each of six boiling tubes using a syringe. Care was taken
to view the syringes from the side to ensure the bottom of the
meniscus was lined up properly with the gradations and there were no
air bubbles in the syringe. A pH buffer was added to the boiling tubes
to maintain the pH at a constant level so that changes in pH as a
result of the reaction would not affect the activity of the enzyme and
disrupt the results. pH 7 buffer was selected to match the natural pH
of the potato tissue and therefore suit the enzyme so that it could
work efficiently.

One of the boiling tubes was then connected to a manometer containing
manometer fluid using a bung (see diagram below). Holding the
manometer level by the bung to ensure that the fluid was at its lowest
level, a mark was drawn to indicate this point using a marker pen. A
further mark was then drawn 5cm above the original, measured using a

15 pieces of potato were placed in to the boiling tube using a pair of
tweezers to prevent contamination. A clamp was then placed over the
rubber tubing on the bung to ensure that all of the oxygen gas
released will travel up the manometer tube and not escape. Once the
clamp was closed, the stopwatch was started to record the time taken
for the manometer fluid to travel to the second mark. When this had
been achieved the time was noted in a results table and the clamp
opened to allow the gas to be released and the manometer fluid to
return to its original level. Once the apparatus had been reset and
any air bubbles in the manometer fluid removed, a second and later
third reading was taken by re-closing the clamp and measuring the time
taken for sufficient gas to be released from the reaction to force the
manometer fluid back up to the top mark. By taking several readings
for each enzyme concentration, it enabled me to average the results to
minimise the extent of any inaccuracies. The experiment was then
repeated for different quantities of potato discs (enzyme
concentration) by using different boiling tubes containing hydrogen
peroxide (see table below for quantities). I began with an enzyme
concentration of 15 potato discs rather than a lower quantity to
ensure that the apparatus was working correctly.

Vol. pH buffer (ml)

Vol. Hydrogen peroxide (ml)

No.Potato discs






















Observations and Measurements - In the boiling tubes it was clear that
a reaction was taking place by the observation of bubbles of oxygen
gas being released creating a 'fizzing' in the boiling tubes.

In order to decide how varying the enzyme concentration affected the
decomposition of hydrogen peroxide, the rate of reaction was measured.
To do this accurately, the time taken for a specific quantity of
oxygen gas (a product of the reaction) to be released was determined.
This was achieved by observing the time taken for the manometer fluid
to travel between the two marked fixed points as it was forced through
the manometer by the rising gas. This was an accurate measure of how
the enzyme concentration influenced the breakdown of hydrogen
peroxide, as the quantity and speed of gas produced is dependant on
the rate of reaction. The marked points remained the same distance
apart for each reading for different enzyme concentrations so that
they could be accurately compared and the trend observed.

All measurements were taken so that the stopwatch was started once the
rubber tubing was sealed and the stopwatch stopped once the manometer
fluid had reached the base of the highest marked point. To judge
accurately, the point at which the fluid reached the marked line, it
was examined at eye level and the measurement taken when the bottom of
the meniscus was lined up to the mark. This was the same for every

Data handling - The data obtained from this investigation has been
recorded in a table showing the time, enzyme concentration and rate of
reaction. This means that the results of the experiment are presented
in a clear and orderly fashion that allows patterns in the results to
become more obvious.

The rate of reaction was calculated by dividing 1000 by the time taken
for the quantity of gas to be produced from the reaction. By
calculating the rate of reaction instead of merely using the time
readings, the quicker reactions will be represented as a greater value
for the rate of reaction rather than a small time value. This makes
the graph more clear and easier to analyse.

Patterns within the results collected from the experiment, are best
shown on a graph. This is because overall trends between the enzyme
concentration and rate of reaction can be portrayed more effectively
and become more obvious.

Limitations and Precautions - In this investigation, I measured the
rate of reaction with enzyme concentrations of between 0 and 35 units
(potato discs). At 0, there should be no reaction as there will be no
substrate, however, I included it to act as a control. This will show
that it is the variable, enzyme concentration that is being measured.

I decided to vary the enzyme concentration by varying the number of
potato discs. However, although the enzyme, Catalase, occurs in the
potato tissue, I did not know the exact quantity and certain discs
might have more Catalase than others. This could be a major limitation
in this investigation. I have tried to compensate for this, however,
by taking multiple readings for each enzyme concentration so that
inaccuracies are minimised once averaged.

As a precaution, I have limited my contact with the boiling tubes, as
my body heat will raise the temperature, increasing the rate of
reaction or expanding the gas inside the test tube moving the
manometer fluid.

I also monitored the temperature using a thermometer to ensure that it
remained constant and not disrupt the results of the experiment by
affecting the activity of the Catalase.

A pH buffer was used to maintain a consistent pH level in the boiling
tubes. This way there was no variation in pH that might have resulted
in an increase or decrease in the rate of reaction.

A major limitation of this investigation was the time. It meant that
only 8different enzyme concentrations could be measured at intervals
of 5 units or potato discs. This means that only very general, overall
trends can be identified across the results. Patterns between these
values can only be approximated and are not necessarily accurate.

Safety - Laboratory coats were worn during the investigation to
prevent chemicals from spoiling clothes. Care was also taken whilst
handling the chemicals as hydrogen peroxide is corrosive and the
manometer fluid is permanently staining. Whilst using the razor
blades, care was also taken to hold them by the handle and not the
blade to prevent an accident occurring.

Results - The rate at which hydrogen peroxide was broken down to water
and oxygen in the presence of Catalase:

The graph "The decomposition of hydrogen peroxide in the presence of
potato catalase Chart 2" shows the rate of reaction up to an enzyme
concentration of 25. Up to this point the line of best fit is a
straight line through the origin. This shows that without the enzyme,
catalase, present no reaction takes place. It also indicates that the
enzyme concentration is directly proportional to the rate of reaction
for the decomposition of hydrogen peroxide in the presence of catalase
(the rate of reaction increases with increasing enzyme concentration).

The other graph, "The activity of potato catalase with differing
enzyme concentrations Chart 1", shows how the rate of reaction varies
with differing enzyme concentrations over the whole range that I
experimented with. After an enzyme concentration of 25 potato discs,
the line of best fit is no longer a straight line and begins to level
off. The enzyme concentration is no longer proportional to the rate of
reaction, and the increases in the rate of reaction reduce

Conclusion - The reaction was fastest at an enzyme concentration of 35
potato discs. At this enzyme concentration there were the greatest
number of free active sites available to the substrate molecules so
that they could be broken down.

The rate increased steadily from 0 up to a concentration of 25 and
slowed beyond this point to give a "maximum level". It appears that at
this "maximum level", increasing the enzyme concentration had little
effect and other factors such as substrate concentration were limiting
the reaction and prevented any further increases in the rate of

Discussion - The results of this investigation are as I predicted in
the hypothesis: "The reaction will increase with increasing enzyme
concentration when molecules of hydrogen peroxide are freely
available. However, when molecules of the substrate are in short
supply, the increase in rate of reaction is limited and will have
little effect". The reasons for this are that there are number of
variables that influence the decomposition of hydrogen peroxide in the
presence of Catalase. Some of which can be classified as limiting
factors i.e. the reaction is dependant or "limited" by their
availability, to be able to function effectively; these include enzyme
concentration, temperature and substrate concentration. All of these
factors are required for an efficient reaction to take place, even
when one is freely available the reaction can still be limited by the
availability of the others. When I increased the enzyme concentration,
it meant that there were more free active sites for the substrate
molecules so that a greater quantity of substrate molecules could be
broken down into products. However past a certain point, which in my
investigation was at an enzyme concentration of 25 potato discs, there
were many free active sites but insufficient substrate molecules to
occupy them. Increasing the enzyme concentration further without
increasing the substrate concentration has no effect on the rate of
reaction which eventually will remain constant.

From the line of best fit on the graph "Chart 1", it is clear that
some of the points do not exactly fit. They are anomalies. Although
they have only slight inaccuracies, they are an indicator of possible
errors in the investigation. These may have occurred in either the
measurement of the quantities of the enzyme and substrate or the
measurement of the time taken for the manometer fluid to rise five
centimetres up the manometer tube. Another possibility was that
fluctuations in temperature caused the rate of reaction to increase or
the gas inside the boiling tube to expand, forcing the fluid to rise
up the manometer tube. Although minimal contact was made with the
apparatus during the investigation, slight undetected variations in
the room temperature may have led to inaccuracies.

The precision of this experiment, generally, was very limited since
insufficient readings were taken. Although the range of enzyme
concentrations taken was large, the difference in enzyme concentration
between each reading was too great to distinguish a value between
them. For example, the rate of reaction at an enzyme concentration of
15 potato discs was 35 + or - 4. This results in an error of
uncertainty of 11%

The shape of the graph is as I predicted showing that as enzyme
concentration increases so does the rate of reaction. This is because
at a greater enzyme concentration, there are more free active sites
available for the substrate and so more products can be made in a
shorter length of time. However, it is not possible to take precise
readings from the graph between the plotted points since insufficient
readings were taken. To be able to do this, intermediate enzyme
concentrations would have to be measured so that the shape of the
graph would be more exact.

Suggestions and Improvements - To create a more accurate experiment in
the future, several precautions or alterations can be made:

· Instead of using potato discs that have slight variations in size,
and volume of catalase, as a source for the enzyme, a 1 molar solution
of the enzyme could have been diluted to create different
concentrations. This way the concentrations can be measured far more
accurately reducing the chances of errors in the investigation.

· In this experiment 8 enzyme concentrations were considered. However,
although there was a large range, insufficient intermediate
measurements were made creating gaps between the measurements where
guess work is needed to predict the rate of reaction at these points
e.g. point A on graph "Chart 2". In a future investigation, a far
greater number of enzyme concentrations between those already recorded
should be tested reducing the extent of any anomalies on a graph where
the line of best fit must be drawn.

· In this investigation each reading was repeated so that an average
rate of reaction for each enzyme concentration could be calculated.
This could be improved by repeating the reading more frequently thus
reducing the extent of any anomalies further, once averaged.

The Rate Of Catalyse Activity

Catalyse is an enzyme that breaks down hydrogen peroxide into water
and oxygen.

Hydrogen Peroxide Water + Oxygen
2H2O2 2H2O +O2

The factors that affect enzyme activity are:

· Enzyme concentration (Gareth Williams, Biology For You page 31)
· Substrate concentration (Mary + Geoff Jones, Biology page )
· Temperature (Gareth Williams, Biology For You page 31)
· pH (Gareth Williams, Biology For You page 31)

Background Information

Enzymes such as catalyse are protein molecules, which are found in
living cells. They are used to speed up specific reactions in the
cells. They are all very specific as each enzyme just performs one
particular reaction.

Catalyse is an enzyme found in food such as potato and liver. It is
used for removing Hydrogen Peroxide from the cells. Catalyse speeds up
the decomposition of Hydrogen Peroxide into water and oxygen. It is
able to speed up the decomposition of Hydrogen Peroxide because of the
shape of the Hydrogen Peroxide molecule. This type of reaction where a
molecule is broken down into smaller pieces is called an anabolic

In my investigation I will study the effect of enzyme concentration on
the rate of catalyse activity.


I predict that if I increase the enzyme concentration this will
increase the rate of catalyse activity because there will be more
active sites for reactions, resulting in more chemical reactions
caused by successful collisions between the active site of the enzyme
(catalyse) and the substrate (hydrogen peroxide)
(Gareth Williams, Biology For You pages 29-31)

10cm3 measuring cylinder
Hydrogen Peroxide
Water bath
Pureed potato
Test tube
Delivery tube


I will use the equipment as shown in the diagram to measure the amount
of oxygen that gathers into the measuring cylinder. The amount of
oxygen is directly related to the rate of catalyse activity. The more
oxygen, the more catalyse activity and the less oxygen, the less
catalyse activity. I will be able to measure the oxygen using the
measuring cylinder.

I will be using pureed potato to provide the enzymes for this
experiment. This is the best kind of potato to use because it has more
surface area than whole potatoes and this means more active sites for
the substrate to react with.I will be change the enzyme
concentration(amount of pureed potato) in order to measure its effects
on the rate of catalyse activity. I will perform the experiment using
0cm3, 1cm3, 2cm3, 3cm3, 4cm3, 5cm3 of pureed potato. In order to make
it a fair test I must keep the volume the same for each test. The set
volume will be 5cm3 so I will use water to keep the volume at 5cm3,
this means that for example when I do the experiment with 3cm3 of
pureed potato I will add 2cm3 of water. I will add 5cm3 of hydrogen
peroxide each time in order to keep the test fair. I will use a
measuring cylinder to measure out all these values. I will repeat the
test at each value three times and then take an average to make it a
fair test and I will also keep the temperature of the room the same
throughout the experiment using the thermostatic controls in the
laboratory. The temperature will remain at 20ºC.

These are the measurements I will be using.

TEST TUBE Enzyme(POTATO)(Cm3) WATER(Cm3) Substrate (Hydrogen
peroxide)(Cm3) TOTAL VOLUME(Cm3)
1 0 5 5 10
2 1 4 5 10
3 2 3 5 10
4 3 2 5 10
5 4 1 5 10
6 5 0 4 10


Test Tube Experiment No.1 Experiment No.2 Experiment No.3 Total
1 0 0 0 0 0
2 2.3 2.1 2.4 6.8 2.3
3 4.3 4.5 4.4 13.2 4.4
4 6.8 6.5 7.1 20.4 6.8
5 7.4 7.4 7.1 21.9 7.3
6 7.4 7.5 7.2 22.7 7.4

In conclusion I can see that my prediction was correct. The higher the
enzyme concentration was, the more oxygen was produced. This is
because the with higher enzyme concentration, there are more active
sites where more successful collisions will take place between the
active site of the enzyme and the substrate. The graph shows that as
the enzyme concentration was increasing more and more, the rate of
catalyse activity was increasing less an less which indicates that it
was nearing the optimum rate of catalyse activity.

My experiment worked well although if I were to do it again I could do
certain things differently in order to gain more accurate results. I
would repeat the experiment more times as this would give me a more
accurate average and I would also uses more different values in order
to get a more detailed outlook on how enzyme concentration affects the
rate of catalyse activity. I would also use a measuring syringe to
measure the amount of oxygen because this is a much more accurate way
of doing it.

How to Cite this Page

MLA Citation:
"The Effect of Hydrogen Peroxide on Catalase with Varied Temperature." 18 Apr 2014

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