The Effect of Temperature on the Reaction Between Amylase and Starch
The Effect of Temperature on the Reaction Between Amylase and Starch
I decided to investigate how a change in temperature would affect the rate of reaction between starch and amylase.
From previous work done on the starch/amylase reaction, I know that amylase is an enzyme which breaks down starch into smaller molecules. It is used inside the body where the temperature is certainly higher the room temperature.
Hypothesis
I predict that the rate of reaction between amylase and starch will increase as the temperature increases. In general the rate of an enzyme catalysed reaction doubles for every 10oC rise in temperature and I would expect amylase to follow a similar trend.
Of course the reaction will stop when the temperature rises too high because, as an increase in temperature increases vibrations ion the molecule, the vibrations become so violent that the bonds holding the structure together are broken and he enzyme denatures.
The enzyme should be most effective at 40oC because this is body temperature, at which is it most used to working. In order for it to function most efficiently in the body, amylase must have an optimum temperature of 40oC.
I would expect my results to look like this on graph. The reasoning behind this is that an increase in temperature will increase the vibration of the molecules, meaning that they will collide more often and more violently, therefore increasing the rate of the reaction.
Using preliminary work, I devised an experiment to test my hypothesis. As I was altering temperature, I would have to keep all my other variables constant as to ensure a fair test and to obtain accurate results.
Variables
Variables How I will Keep them Constant
Concentration of Starch/Amylase/Iodine By using the same concentration throughout my investigation
Volume of Starch/Amylase/Iodine by using a calibrated pipette and by making sure to eliminate parallax errors
As a control test, I will put amylase in one test tube, starch in another, then add iodine to view the ...
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Using preliminary work, I devised an experiment to test my hypothesis. As I was altering temperature, I would have to keep all my other variables constant as to ensure a fair test and to obtain accurate results.
Variables
Variables How I will Keep them Constant
Concentration of Starch/Amylase/Iodine By using the same concentration throughout my investigation
Volume of Starch/Amylase/Iodine by using a calibrated pipette and by making sure to eliminate parallax errors
As a control test, I will put amylase in one test tube, starch in another, then add iodine to view the reaction
Preliminary work
By using a dropping tile and different concentrations of amylase and starch, I found that 0.25% amylase worked well because it did not dissolve the starch too quickly. This turned out to be the biggest problem in my trial experiments.
Different temperatures were explored by placing the starch and amylase in water baths for a certain period of time and then adding the iodine. The mixtures were then viewed in a colorimeter.
The final values I decided on were:
Starch: 8cm3 1%
Amylase: 0.5cm3 0.25%
Iodine: 2cm3
Temperatures: room temperature - 100 oC
I also decided to repeat each experiment at least once to increase the accuracy of my experiment.
Method
. Take a test tube and fill it with the starch
2. Place in the set temperature for 30 seconds
3. Add the amylase and wait for 30 more seconds
4. Add the iodine (this stops the reaction)
5. View the mixture on a colorimeter and record the reading
6. Repeat at least once
7. Repeat steps 1 - 6 for each different temperature
! Take care when handling glass, wear goggles and be careful of the hot water !
Results
Temperature Colorimeter Readings
Celsius Arbitrary Units
Expt1 Expt2 Expt3 Average
25 7 6.8 6.4 6.7
35 6.2 6.2 6 6.1
45 3.5 4.7 3.9 4
55 1.6 2.4 1.8 1.9
60 0.7 1.2 1 1
75 0.3 0.3 0.3 0.3
00 0.1 0.1 0.3 0.2
From these figures I was able to draw a graph to help me analyse the experiment. As the temperature increased, the colorimeter reading decreased, slowly at the start, then more quickly, then slowly again.
The colorimeter reading does not equal the rate of reaction because when the iodine is added, the more starch in the mixture, the darker the colour and the colorimeter reading is higher when the colour is darker. Therefore the higher the reading on the colorimeter, the lower the rate of reaction, so the graph was not immensely helpful. I managed to come to the conclusion that, my experiment confirmed my prediction to an extent. Kinetic theory is the idea that, when a substance is heated, its molecules, having been supplied with energy, move around faster. In this experiment, as the temperature increased, the enzyme and starch molecules collided more frequently (Brownian motion) and with more energy, which caused them to react more efficiently. At low temperatures, the molecules did not collide as frequently and the starch was therefore not broken down as quickly. This is true of any reaction, whether or not it involves catalysts, biological or otherwise. The first half of the results (up to about 50 C) were as expected, but instead of denaturing at higher temperatures, the enzyme seemed to work even faster, creating a few anomalous results. This was probably down to the fact that I did not heat the amylase before using it in my experiment, which meant that it was warming up as the experiment went on. At the higher temperatures it warmed up more, without denaturing. If I had left the amylase in the water bath before I added it to the starch, this would have heated it to the correct temperature before the reaction started.
On the whole I think my results were fairly reliable and accurate apart from the aforementioned anomalous results. More repeats would have helped, but I tried to squeeze as many as I could into the allotted time.
The apparatus used could have been improved in many ways. The water baths used were not all at the exact temperatures required, and each contained a different amount of water. If better quality water baths had been used, and there was time to ensure that each had exactly the same amount of water and was at the exact temperature required, anomalous results could have been eliminated. This could also have been achieved by repeating the experiment for each temperature more than twice, and also by performing the experiment at intervals smaller than 10oC Seeing how amylase worked under colder temperatures may have been interesting also, but in the end I achieved what I set out to do, and I would not change much should I have to do this experiment again.
