Practical 1. Investigating the effect of substrate concentration on the rate of an enzyme
controlled reaction.
Apparatus and techniques covered:
ATa
ATc
use appropriate apparatus to record a range of quantitative measurements (to
include mass, time, volume, temperature, length and pH).
use laboratory glassware apparatus for a variety of experimental techniques to
include serial dilutions.
Introduction.
A convenient system to investigate the effect of substrate concentration on the rate of
enzyme controlled reaction is provided by the hydrolysis of hydrogen peroxide to water and
oxygen that is catalysed by the enzyme catalase:
You will investigate the effect of hydrogen peroxide (substrate) concentration on the rate of
reaction by indirectly measuring the rate of oxygen liberation.
Method.
Safety Note: 6 % hydrogen peroxide is an Irritant. Eye protection must be worn at all times during
this practical and skin contact should be avoided. See CLEAPPS Student Hazcard 57.
Safety Note: Dilute catalase is low risk. See CLEAPPS Student Hazcard 2.
1. Draw lines 4 cm from the bottom of 5 boiling tubes.
2. Make a series of hydrogen peroxide concentrations in these boiling tubes following the
table below.
Concentration of hydrogen
peroxide solution / %
0.75
1.50
3.00
4.50
6.00
3.
4.
5.
6.
7.
8.
9.
10.
11.
Volume of 6% hydrogen
peroxide / cm3
2.5
5.0
10.0
15.0
20.0
Volume of water / cm3
17.5
15.0
10.0
5.0
0.0
Label the tubes appropriately.
Place 1 mL of catalase solution in a watch glass.
Using forceps add three paper discs to the solution and allow to soak for 30 seconds.
Using forceps transfer one disc to one of your boiling tubes of hydrogen peroxide.
Using a glass rod push the disc to the bottom of the tube.
Start a stop clock and release the disc.
Record the time taken for the disc to reach the 4 cm line.
Repeat steps 6 to 9 for the remaining two discs in the same tube of hydrogen peroxide.
Repeat steps 5 to 10 for the remaining concentrations of hydrogen peroxide.
Report.
1. Write a short introduction including the aim, the reaction you are studying, and your
hypothesis.
2. Refer to this document as the Method noting any modifications you made.
3. Include a statement of the risks involved and how you minimised them.
4. Describe how you will use your data to calculate a rate of enzyme action (Hint: rate=time
taken for a measurable event).
5. Using your raw data calculate the rate for each disc, and a mean rate for each
concentration.
6. Include these values with correct units and your raw data in a ‘neat’ table.
7. Plot a graph of mean rate against hydrogen peroxide concentration; include range bars to
represent variation from the mean rates.
8. Write a short paragraph describing your results. (Hint: pick out major trends and refer to
your data as you describe them.)
Discussion.
Under the heading ‘Discussion’ write a brief analysis of your data and compare it to data you
can find online considering the following points:
1. Did the experiment meet the aim and was your hypothesis correct?
2. What does your data show about the effect of substrate concentration on the rate of an
enzyme controlled reaction?
3. How does the trend you have observed compare to information you can find on the
internet?
Remember to cite the website you use including the date you accessed the website.
Evaluation.
Under the heading ‘Evaluation’ write a brief analysis of the practical considering:
1.
2.
3.
4.
Which variables did you control in this experiment?
How accurate were your results?
Based on the replicates how reliable were your results?
What were the limitations of this experiment and how you could improve it?