Effect of Enzyme Concentration on Enzyme Activity
Xochilt Garcia
BIOL 1730.862
October 24, 2013
Caroline Vela
Thursday 11-1:50
Abstract
This experiment examined the effect of the enzyme concentration on the activity of amylase. To
test this, iodine was added to a starch solution in which amylase had been introduced. Starch
solution was placed in the wells of two spot plates and a four different concentrations of amylase
were introduced. At five minute intervals, iodine was added to determine if the amylase had
broken down the starch. The results of this experiment were inconclusive.
Introduction
The activity of enzymes are affected by three factors: temperature, pH, and enzyme
concentration. To test the effect of concentration, amylase is tested to see if it has acted. Iodine is
added to a starch solution. If the amylase acted, the solution will stay a light brown as an
indicator that it broke the starch down into maltose. If the starch solutions becomes black or
blue-black, the enzyme has not acted (Wachtmeister). As the enzyme concentration drops, the
enzyme takes longer to act.
Materials and Methods
Eight test tubes (labeled 1-8) were filled with different solutions according to Table 1. Two
porcelain spot plates were arranged and labeled as shown in Table 2. Each of the 24 wells were
filled with 5 drops of the starch solution and two drops of the pH 7 buffer. Mix the solution by
stirring each well with a toothpick. Then as quickly as possible, one drop of the different amylase
concentrations were added consecutively; the undiluted amylase solution was placed in Row A;
the enzymes from tube six were placed in Row B; tube 7 in Row C; and tube 8 into row D. For
Time 0, one drop of iodine was placed immediately in column 1 and stirred and the results were
recorded in Table 3. If the solution was stained black or blue-black, a 1 was recorded. If the
solution remained unchanged, a 2 was recorded. At five minute intervals, iodine was introduced
and stirred into every well of a column.
Table 1: Test Tubes 1-8 and Materials for Each
1
4ml enzyme/amylase
2
3ml pH 7 Buffer
3
3ml distilled water
4
3ml starch
5
3ml iodine
6
1:2 enzyme dilution 4ml
7
1:4 enzyme dilution 4ml
8
1:8 enzyme dilution 4ml
Table 1: Test tubes and the materials placed in each tube
Table 2:
Porcelain Plate Arrangement and Labeling
1
A
B
2
3
4
5
6
C
D
Table 2: The correct positioning and labeling of the porcelain tables and wells.
Table 3
Results of Enzyme Concentration Test
Enzyme/
Time 0
Time 5
Time 10
Time 15
Time 20
Time 25
Test Tube
1
6
7
8
Table 3: Table for recording observations and results of expirement.
Results
Amylase did not break down starch and the wells were not stained. Most wells remained
unstained. This differs from other groups in that more wells in their experiments were stained by
the iodine. As shown in Table 4, the findings seem to be inconsistent and do not follow the
expected pattern.
Table 4
Results of Enzyme Concentration Test
Enzyme/
Time 0
Time 5
Time 10
Time 15
Time 20
Time 25
1
2
2
2
2
2
2
6
1
2
2
2
2
2
7
1
2
1
2
2
2
8
2
1
2
2
2
2
Test Tube
Table 4: Results of Enzyme Concentration Test
Discussion
Due to the inconsistent findings, the hypothesis is rejected. The concentrations did not behave in
the way they were expected. There are several possible human errors that occurred that may have
skewed the results. During the first trial, solutions were placed in incorrect wells. The porcelain
plates were then rinsed but not properly dried. Excess water could have affected the results of
this test by slightly altering the enzyme concentrations. Additionally, it is possible too much of
any of the materials could have introduced or time wasn’t kept adequately. This experiment does
not accurately portray the effects of enzyme concentration on the enzyme activity.
References
Wachtmeister, H. F. E., & Scott, L. J. (2006). Encounters with life: General biology laboratory
manual. (Seventh ed., pp. 44-45). Morton Publishing Company.
Aragon, J. J., & Sols, A. (1991). Regulation of enzyme activity in the cell: effect of enzyme
concentration. The FASEB Journal, 5(14), 2945-2950.