EXPERIMENT:

Studying the effects of enzyme

concentration on the activity of
salivary amylase
Problem Statement:
 What are the effects of enzyme
concentration on the activity of
salivary amylase?
Hypothesis
 The rate of enzymatic reaction
increases with the increase in
enzyme concentration as long as
there are no other factors limiting
the rate of reaction.
Variables:
 Manipulated variable:
Concentration of enzyme
 Responding variable:
Time taken for hydrolysis of starch
to be complete
 Fixed variable:
Substrate concentration,
temperature and pH of medium.
Materials:
 1% starch suspension, 0.8%
amylase or saliva suspension,
iodine solution and distilled water.
Apparatus:
 5ml syringes, 1ml syringes, test
tubes, glass rods, a stopwatch, a
white tile with grooves, measuring
cylinder and droppers.
Procedure:
1. Six test tubes, labelled A to F are
prepared.
2. The test tubes contains the
following mixtures.
A: 0.5ml of 0.8% amylase + 2.5ml of distilled water

B: 1.0ml of 0.8% amylase + 2.0ml of distilled water

C: 1.5ml of 0.8% amylase + 1.5ml of distilled water

D: 2.0ml of 0.8% amylase + 1.0ml of distilled water

E: 2.5ml of 0.8% amylase + 0.5ml of distilled water

F: 3.0ml of 0.8% amylase

3. Test tubes A to F are immersed in
a water bath set at 37°C.
4. Meanwhile, drops of iodine
solution are added separately
onto the grooves of the white tile.
5. 4ml of 1% starch suspension is
added to test tube A using a
syringe. Immediately the
stopwatch is started and the time
is recorded as 0 minute.
6. The mixture in the test tube is
stirred using a glass rod. A small
amount of the mixture is removed
and tested with the iodine solution
on the tile.
7. The iodine test is repeated at
30~seconds intervals until the
mixture does not turn blue~black
in colour when tested with iodine
solution. The time taken for the
hydrolysis of starch to be
completed is recorded
8. Steps 4 to 7 are repeated with
the contents of test tubes of B, C,
D, E and F.
9. All results are recorded and
tabulated in a table.
10. The rate of enzymatic reaction
(l/time) is then calculated. A
graph of the rate of enzymatic
reaction, l/t against the enzyme
concentration is plotted.
 Results (Table):
Test Concentration Time taken for the Rate of
Tube of amylase (%) hydrolysis of enzymatic
starch to be reaction
completed
(l/t)
seconds minutes
A 0.17 330 5.5 0.18
B 0.33 150 2.5 0.40
C 0.50 90 1.5 0.67
D 0.67 60 1.0 1.00
E 0.83 60 1.0 1.00
F 1.00 60 1.0 1.00
 Results (Graph):
Rate of Reaction
1.2

0.8

0.6

0.4

0.2

0
0.17 0.33 0.5 0.67 0.83 1
Concentration of amylase (%)
Discussion:
 The time taken for amylase to
hydrolyse starch decreases as the
enzyme concentration increases.
 A higher concentration of the
enzyme contains more enzyme
molecules to hydrolyse the starch
molecules. Hence, it takes less
time for the hydrolysis of starch to
be completed.
 From the graph, it can be seen that
the rate of reaction increases when
enzyme concentration increases
until a certain concentration, when
the further increase in enzyme
concentration will not increase the
rate of reaction. This is because
the substrate concentration has
become the limiting factor.
Conclusion:
 The rate of reaction increases with
the increase in enzyme
concentration until a certain
concentration of enzyme is
achieved.