Edexcel IGCSE Biology · Spec 2.12
Investigating Enzymes Practical
Required practical: investigating how temperature affects the enzyme amylase, using starch and iodine.
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Investigating Enzymes Practical
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Explained
Timing amylase with iodine
Amylase breaks starch down into maltose. Iodine solution is orange brown normally and blue black when starch is present. Put those two facts together and you have a way of seeing exactly when the starch has gone.
The method
Put drops of iodine solution into the wells of a spotting tile. Warm a tube of starch solution and a tube of amylase separately in a water bath at your chosen temperature, then mix them and start a stopwatch.
Every thirty seconds, take a drop of the mixture and add it to a fresh well of iodine. While starch remains, the iodine turns blue black. When it stays orange brown, the starch has all been digested, and that is your end point.
Repeat the whole thing at a range of temperatures, using water baths at say 10, 20, 30, 40 and 50 degrees Celsius.
Turning time into rate
A shorter time means a faster reaction, so time on its own is a slightly awkward measure. Calculate the rate as one divided by the time taken, and the numbers then rise as the reaction gets faster, which makes the graph easier to read.
What the results show
Rate rises with temperature at first, because the enzyme and substrate molecules have more kinetic energy, move faster and collide more often, so more successful collisions form enzyme substrate complexes.
It peaks at the optimum temperature, around 37 degrees Celsius for a human enzyme.
Above that it falls sharply, and for a different reason. The heat breaks the bonds holding the enzyme's shape, so the active site changes shape and the substrate no longer fits. The enzyme has been denatured, and unlike the rise this change cannot be undone by cooling.
Notice that the two halves of the curve have different explanations. The rise is about collisions; the fall is about shape. Answers that use kinetic energy for both miss what is being tested.
What the mark scheme accepts and rejects
An Edexcel International GCSE Biology mark scheme for a question on temperature and enzyme activity credits the ideas that the enzyme denatures at the higher temperature or that the optimum is below the stated value, and that the enzyme and substrate no longer bind or that the active site has changed shape.
It then adds a fallback worth knowing about. If none of those points is awarded, one alternative mark is available for linking temperature to kinetic energy or to collisions. That is the mark scheme rescuing an answer that has the general idea but not the specific one, and it tells you which is the better answer to give.
Note the wording it uses for denaturing. The active site changes shape, and the substrate no longer fits. The enzyme is not killed, because it was never alive, and it is not melted. Those two words are common and both are wrong.
Making it a fair test
Only the temperature should change. Keep the volume and concentration of the starch the same, the volume and concentration of the amylase the same, and the pH the same by using a buffer.
Two details improve the results. Bring both solutions to the test temperature before mixing them, otherwise the mixture spends the first part of the experiment at the wrong temperature. And keep the sampling interval the same throughout, because the interval limits how precisely you can identify the end point.
Repeat each temperature and take a mean, which reduces the effect of random error and lets you spot an anomalous result.
Spec 2.12
What you need to know
- Investigate how temperature affects an enzyme
- Describe the method using amylase and starch
- Explain the results
Active recall
Quick check
Answer each question before opening the answer.
How can you tell when amylase has fully broken down starch?
Test samples with iodine at intervals; when it no longer turns blue-black, the starch has been digested.
Name two variables you must control in this experiment.
Any two of: pH, temperature, enzyme concentration, substrate concentration, volumes used.
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