The Effect of a Catalase on the Breakdown of Hydrogen Peroxide

The Effect of a Catalase on the Breakdown of Hydrogen Peroxide

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The Effect of a Catalase on the Breakdown of Hydrogen Peroxide

Aim

To follow the progress of a catalysed reaction by measuring the volume
of gas produced as the reaction proceeds. Using the initial rates of a
series of experiments I will be able to find the orders of the
reaction with respect to enzyme and substrate. Also to find out if
concentration has an effect on the reaction when an enzyme is used to
accelerate the breakdown of hydrogen peroxide.

Introduction / Background Information.

This is an experiment to examine how the concentration of the
substrate Hydrogen Peroxide (H2O2) affects the rate of reaction of the
enzyme Catalase. In this experiment I will be using yeast as a source
of catalase.

Enzymes are catalysts which speed up specific reactions. Enzymes such
as catalase are protein molecules, which speed up a specific reaction
within the cell. They are all very specific as each enzyme just
performs one particular reaction.

Catalase is an enzyme found in food such as potato and liver, (in this
case I will be using yeast as my source) It is used for removing
hydrogen peroxide from cells. This need to be done as hydrogen
peroxide is the poisonous by product of the metabolism. Catalase
speeds up the decomposition of hydrogen peroxide into water and oxygen
as shown in the equation below.

2H2O(aq) à 2H2O(l) + O2 (g)

It is able to speed up the decomposition of hydrogen peroxide because
the shape of its active site matches the shape of the hydrogen
peroxide molecule. This type of reaction were a molecule is broken
down into smaller pieces is called an anabolic reaction.

I will be studying the effect of concentration of the catalase in this
reaction.

Hypothesis

Hydrogen peroxide will breakdown to oxygen in water in the presence of
catalase. The reaction will increase with the increasing enzyme
concentration when the molecules of hydrogen peroxide are freely
available. The more concentrated the catalase the more chance of the

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Related Searches

molecules colliding with the hydrogen peroxide molecules resulting in
a faster reaction.

Variables

In this investigation I will only be changing one variable
(concentration), the variables that effect the activity of the enzyme,
catalase , will be considered and controlled so that they will not
disrupt the success of the investigation.

* Temperature - this will speed up the reaction (kinetic theory) so
to control this I will be using tongues and to handle the boiling
tube to keep the solution at room temperature (approx 20 degrees
Celsius)

* pH - Any change in pH will alter or denature the shape of the
substrate, so to control this I will attempt to use a ph buffer to
keep a constant pH of 7 (if available)

Foreign objects or chemicals - all equipment and glass were must be
clean so no other factors disrupt the investigation.

Safety Considerations / COSSH.

Hydrogen Peroxide - corrosive - irritant to skin and eyes.

A lab coat must be warn (to prevent spillages on skin and clothing)

Safety glasses (to prevent splashes into eyes)

Gloves (to prevent contact onto skin)

All spillages must be reported and cleaned up immediately.

All breakages must be reported immediately and reported.

Equipment.

* Burette (50cm3)

* Trough or bowl

* Boiling tube with bung and delivery tube

* Graduated pipette (5cm3) and safety filler

* Measuring cylinder (10cm3)

* Hydrogen peroxide solution, 5 vol (25cm3)

* Yeast suspension (20cm3), made from 2g dried yeast in 160cm3 water
aerated for several hours.

* Stopwatch

Method.

1. - Fill a burette with warm water and invert it into a trough of
water. Hold it in place with a clamp and check that the burette is
not leaking. Make sure you leave enough room in the trough for
water which will be displaced from the burette.

2. - Place 2.5cm3 of well stirred yeast suspension in a boiling tube
and set up the apparatus as in fig 1. Carefully open the tap on
the burette until the meniscus reads the 50cm3 mark. (it will be
zero in this experiment as the burette is upside down)

3. - Measure out 5cm3 of hydrogen peroxide in a 10cm3 measuring
cylinder. Record the readings of volume at 10 second intervals for
4 minutes.

4. - Add 5cm3 of hydrogen peroxide to the yeast suspension and
quickly replace the bung. Zero time is counted as the time of the
first bubble appears in the burette. Take a reading of the volume
of gas in the burette every 10 seconds for 4 minutes.

5. - Wash out the boiling tube, refill the burette, and repeat steps
1-4 four more times using:

4cm3 H2O2 + 1cm3 distilled H2O

3cm3 H2O2 + 2cm3 distilled H2O

2cm3 H2O2 + 3cm3 distilled H2O

1cm3 H2O2 + 4cm3 distilled H2O

6. - Plot the results on a graph. Finding the initial rate of the
reaction, measure the gradient of each tangent.

Diagram.

See attached sheet. (fig 1)

Results.

Exp 1

4cm3 H2O2 +

1cm3 distilled H2O

Exp 2

3cm3 H2O2 +

2cm3 distilled H2O

Exp 3

2cm3 H2O2 +

3cm3 distilled H2O

Exp 4

1cm3 H2O2 +

4cm3 distilled H2O

Time (secs)

Vol O2

(cm3)

Time

(secs)

Vol O2

(cm3)

Time

(secs)

Vol O2

(cm3)

Time

(secs)

Vol O2

(cm3)

10

20

10

10

10

5

10

5

20

35

20

20

20

10

20

5

30

47

30

30

30

15

30

6

40

60

40

35

40

20

40

6

50

60

50

46

50

22

50

7

1min

60

70

60

50

60

25

60

7

70

80

70

52

70

28

70

8

80

85

80

54

80

30

80

10

90

90

90

55

90

31

90

14

100

95

100

56

100

32

100

15

110

100

110

57

110

33

110

16

2min

120

105

120

58

120

34

120

17

130

110

130

59

130

35

130

18

140

120

140

60

140

36

140

20

150

125

150

62

150

37

150

21

160

130

160

65

160

38

160

22

170

135

170

70

170

39

170

23

3min

180

135

180

71

180

39

180

24

190

137

190

72

190

40

190

24

200

139

200

73

200

40

200

25

210

141

210

74

210

40

210

25

220

141

220

76

220

41

220

26

230

141

230

77

230

41

230

27

4min

240

141

240

78

240

41

240

28

Graphs

See attached sheets - fig 2 - fig 3.

Evaluation of Results

By looking at the results it shows that my hypothesis agreed with the
results from my experiment. The shape of the graph showed that the
amount of oxygen produced decreased as the concentration lessened. An
explanation for this is the higher the concentration the more chance
of a successful collision rate, for low concentrations the rate of
reaction is therefore directly proportional to the concentration of
hydrogen peroxide. As the concentrations of a substrate increases not
al the collisions will be successful because some active sites will be
saturated.

Critique / Evaluation / Conclusion

Overall I found this experiment satisfactory; I had problems when
plotting graphs as some of the results may have not been totally
accurate. I would need to ask for a second person to read the burette
whist I noted down the results. This was a problem on my own because
the reaction happened fast and my readings were almost certain to be
inaccurate. I would also attempt to change variables to see if this
affected my investigation, giving me a greater understanding of enzyme
action.

Bibliography.

www

www.chemguide.co.uk

www.austin.ac/chemreview

Books

General chemistry - Stuart Marton - Penguin - 1991
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