CCEA How Temperature affects amylase

Volume of the starch amylase sample

Time

Time taken between the samples

Independent Variable

I have chosen to investigate the temperature of the solutions, and I shall record it in oC.

I have decided to investigate the following temperatures 20 oC, 25 oC, 30 oC, 35 oC, 40 oC and 45 oC

Dependant Variables

Time taken for amylase to break starch down into maltose

I will record this time in seconds

For each temperature I will repeat the experiment three times and I will find the average time it took for each temperature

Controlled Variables

The factors which must be kept the same each time I do the experiment are:

Volume of amylase used-2cm3

Concentration of amylase used

Volume of iodine used – 2 drops to each of the dimples

Concentration of iodine used

Volume of starch used- 8cm3

Concentration of starch used

Volume of the starch / amylase sample – 2 drops

Time taken between the samples – 30 seconds

I will also control the apparatus I will use, e.g. I will be using two 75ml beakers and three100ml beakers etc.

Prediction

I predict that with an increase in temperature there will also be a decrease in the time it takes amylase to break down starch, i.e. the rate of reaction will increase. I also predict that the rate of reaction will peak at around 40oC, and as the temperature increases past 40oC the rate of reaction will slow until it abruptly stops at around 60oC.

I predict the above as at a low temperature the molecules of starch and the active sites of the enzymes have few collisions yet as the temperature increase the molecules gain energy and there are more collisions between the molecules and the active sites and the reaction occurs faster. At 40oC the maximum rate is reached as this is the maximum temperature the enzyme can react in, after this temperature the enzymes active sites become denatured where the enzymes start to become misshapen and at 60oC all the enzymes are so denatured that no starch molecule can fit into the amylase’s active sites; so from the temperature of 40oC the reaction slows until 60oC where no reactions can occur.

Apparatus

5 Water Baths

2 filter funnels

5 thermometers

2 burettes

30 test tubes

2 teat pipettes

2 dimple tiles

1 pair of safety goggles

3 100ml beakers

2 75ml beakers

        

Plan

1. I shall collect my apparatus and place it in the middle of my bench.
2. I shall set up my water baths at the following temperatures: 25oC, 30 oC, 35 oC, 40oC and 45 oC which I will test with my thermometers.
3. I shall set up my burettes and place filter funnels at the top of each. I will also place a 100ml beaker below each of them.
4. I will then use a 75ml beaker to add amylase to a burette, zeroing the burette into the 100ml beaker below it.
5. Repeat step 4. except with starch.
6. I will then pour 2ml of amylase into a test tube and repeat it for the other two test tubes going into that water bath.
7. Next I will pour 8ml of starch and place into the other two test tubes in the same water bath as the amylase I added in step 6.
8. I shall then repeat steps 6. and 7. with the other four water baths.
9. Then using a teat pipette I will add two drops of iodine to all the dimples in the dimple squares.
10. Using a thermometer I will check the temperature of the first water bath and record it in a prepared table.
11. I then shall mix the starch from one test tube with the amylase of another (both taken from  the water bath I just recorded the temperature from). At the same time I will start the stop clock. Every thirty seconds I shall take a small sample from the test tube and add two drops of this to a dimple. I shall repeat until when I add the sample the iodine stays orange. I will then record the time it took for this to happen off the stop clock.
12. I will reset the stop clock and repeat step 11. twice for the other two test tubes in the 25oC beaker still recording the time taken.
13. I will then repeat steps 9. - 11. with the for the water baths of temperatures 30 oC, 35 oC, 40 oC and 45 oC.

Safety Precautions

Make sure there are no obstructions on the floor, i.e. school bags

Make sure my tie is out of the way

Make sure the apparatus is far enough away from the end of the bench

Handle all chemicals carefully

Wear safety goggles

Strategy for dealing with results

1. Firstly I shall calculate the average time it took the amylase to break down the starch for each temperature. I will use the following equation:

      I shall record these times in a table

2. I intend to calculate the rate of reaction for amylase to break down starch for each      temperature. This is done with the following equation:

3. I will also plot a graph showing time against temperature. Time in seconds will go on the y-axis and temperature will go on the x-axis. I expect this graph to look similar to the following:

4. Finally I plan to plot a graph for the rate at which amylase breaks down the starch against the temperature. I will have Rate of Reaction on the y-axis and temperature on the x axis. I expect the graph to look similar to the graph below:

Obtaining Evidence

Table of results showing the actual temperature the samples were taken at, the time recorded in seconds, the average time and the rate of reaction

Calculations

Average time=

Rate of Reaction =

Interpreting and Evaluation

Interpreting

My graphs can be seen at the end of this section.

Conclusion

As you can see from my graph of rate of reaction against temperature, at 19oC the rate of reaction is low. With an increase in temperature the rate of reaction increases rapidly, but as it approaches 43 oC the rate of reaction decelerates, and as the temperature increases past 43 oC the rate of reaction drops quite rapidly, until my last reading at 50 oC.

I conclude from my results that temperature does affect the rate at which amylase breaks down starch. I further conclude that the optimum temperature for this reaction is at 43 oC.

My original predication was “that with an increase in temperature there will also be a decrease in the time it takes amylase to break down starch, i.e. the rate of reaction will increase. I also predict that the rate of reaction will peak at around 40oC, and as the temperature increases past 40oC the rate of reaction will slow until it abruptly stops at around 60oC.”

My results were pretty similar to my prediction, except my predication for the optimum temperature slightly differed from  the result I got – I predicted 40 oC would be the optimum temperature, but in fact from my results I can say it is in fact 43 oC. I also predicted that the reaction would stop at 60 oC and in fact this was true, as I didn’t get a result for this temperature.

Explanation of results

The rate of reaction was slow at 19 oC which is because the amylase and starch molecules have little energy which therefore means they are moving slowly. This means that the collisions between the starch molecules and the active sites of the amylase occur less often than they would the temperature was higher. This means the reaction takes longer than it would if the temperature was higher.

The optimum temperature for this experiment was 43 oC. This was so, as at 43 oC the starch molecules are colliding with the active sites of the amylase as fast as they can, and therefore maltose is being produced as fast as possible. As the temperature increase, some of the active sites of the amylase start to deform, which means there are less active sites for the starch molecules to react with so the reaction takes longer. By 60 oC the enzyme is denatured, which means its active sites cannot react with starch anymore, and therefore there is simply no reaction

Evaluation

1. There was one anomalous result on my graph and that was the point plotted for the 32oC when the rate of reaction was 2.2s-1. Most likely one of the two groups who were recording these results, recorded them wrongly, which affected the average time.

2. The graph draw from the results my class gathered is similar to that of the theoretical results. The optimum temperature I got (43 oC ) was slightly different from the one which I predicted (40 oC). However, I believe there is enough evidence to support a firm conclusion and that amylase works best at 43 oC.

3. Evaluation of measuring instruments

Water Bath- Temperature remained pretty much constant, however the actual temperature of the bath differed from the temperature we set using the dial on the water bath.

Teat pipette- There wasn’t much problem with the amount of iodine and of the sample we were adding- i.e. 2 drops, however there was a problem when we were taking the sample and that was too much tended to get sucked up, and by the end we had little sample left.

4. If I had to do this experiment again there are several things I would change. The first being I would use a syringe instead of a teat pipette, which allows an accurate volume of sample to be taken.  I wouldn’t change the burette however as it allowed accurate volumes to be taken, and it done what we needed of it. I also wouldn’t change the stop clock as it did what we required of it.

5. Evaluation of method.

There are several things I would change in my method if I were doing the experiment again. A problem my group had was judging when the iodine was blue-black and when we should sop the clock. Possibly I would use a colour chart if I was to have to do this experiment again or if I could I would use a colorimeter which would make my results far more accurate. Also I think that the time in-between sample was too long, and that a time in-between samples of 10 seconds would make my results more accurate. Also I feel the number of samples we took was slightly too small and maybe if we took sample for every 5 oC from 20 oC to 60 oC. Also if we took several extra samples between 35 oC and 45 oC we may get a more accurate result for optimum temperature.

f)    To conclude this experiment, I set out to investigate how temperature affects the reaction between iodine and starch. After completing the experiment, I concluded that as temperature increases, the rate of reaction also increases, until 43 oC which is the optimum temperature, and when the rate of reaction is largest. The rate of reaction decreases quite rapidly as the temperature increases past 43 oC until the reaction stops at 60 oC as the amylase is denatured, and cannot break down any starch.

Bibliography

New Edition Biology- Mary Jones, Jeff Jones – Published by Cambridge

Class notes taken down from my teacher

www.images.google.com