After a certain point, the rate of reaction begins to fall drastically – at about 60°C, when you might think things should really be moving, there is no reaction at all. Why not? The answer lies in the fact that all enzymes are proteins. As with other proteins an enzyme has a very specific shape, and is held in this shape by strong disulphide bridges and much weaker hydrogen bonds, ionic interactions, etc. Heating the enzymes above a certain level will provide enough energy to break these weaker bonds and unfold the amino acid chain. When this happens the enzyme is said to be ‘denatured’. The enzyme is effectively destroyed, and usually will not work again even if the temperature falls.
The graph shows that there Is an optimum temperature for enzyme activity, i.e. warm enough to make the reaction proceed rapidly, but not hot enough to denature the enzyme. For most of our enzymes, this optimum lies in the range of 37-43°C. It Is no coincidence that the body temperature of humans is about 37°C. In fact, endothermic animals (those organisms such as birds and mammals that can maintain a body temperature Independent of that of their surroundings) have a number of specialised features to keep their temperature at the optimum point. Even ectotherms (those animals such as lizards whose body temperature fluctuates with that of their environment) make use of sun and shade to maintain their temperature within this range.
I predict that my graph will look like this: -
I predict that my graph will look like this because the rate of reaction will slowly increase as the concentration of amylase increases. I believe that when the concentration of the amylase will reach a certain gradient the rate of reaction will stay constant from there on. By this I mean that when the concentration of amylase reaches a maximum, for example 100%, the rate of reaction cannot increase any further due to the fact that the number of enzymes will be the same as the number of substrates. I carry on increasing the number of enzymes; only the number of enzymes equal to the number of substrates will react causing a maximum reaction rate. This is known as the V-max.
To explain my predicted graph further I have decided to split it up into sections: -
At point ‘a’, I predict that the rate of reaction will increase slowerr than proportional to the concentration of amylase. This is because the amylase would have to work quicker to break down all the starch. At point ‘b’, I predict that the rate of reaction will increase in proportional to the concentration of amylase. This is because the concentration of amylase would be at a point where there might be 5 starch molecules to every amylase molecule. At point ‘c’, I predict that the rate of reaction will increase quicker than proportional to the concentration of amylase. This is because at this point there might be two starch molecules to every amylase molecule, gradually becoming one starch molecule to one amylase molecule. At point ‘d’, I predict that the rate of reaction will stay the same even though the concentration of amylase still increases. This is because at this point there will be one starch molecule to one amylase molecule. Now if I carry on increasing the number of amylase molecules, the number of starch molecules will stay the same, and therefore the number of substrates needed to be broken down will also stay the same. This means that part ‘d’ of my predicted graph will not show on my graph of results, because it contains a concentration of more than 100% and my graph will only go up to 100%. Therefore I now predict that my graph will look like this: -
If there were 50 starch molecules and 5 amylase molecules then each amylase molecules would have to break down 10 starch molecules, this would take a long time. But if there were 50 starch molecules and 25 amylase molecules then the reaction would be 5 times quicker that 50 starch and 5 amylase. However if there were 50 starch molecules and 50 amylase molecules then the reaction would be at its quickest because each amylase molecule would have only to break down one starch molecule. Now if there were 50 starch molecules and 100 amylase molecules the reaction time would be as quick as 50 starch molecules and 50 amylase molecules. This is because on 50 amylase molecules are needed to break down the 50 starch molecules, meaning that the other 50-amylase molecules do nothing. This is called the V-Max.
Fair Test
I will change the concentration of amylase by diluting it with water. The variables that I need keep the same are: -
- Starch Concentration
- Temperature
- The number of drops of iodine
- Overall Variable
The independent variable is the concentration of amylase and the dependent variable is the amount of time taken for the starch to break down into glucose. To measure the concentration of amylase I will mix the amylase with the amount of water that will give me the correct concentration gradient I would need for that specific result. Then take 1 ml from that concentration and add it to the starch.
Reliability
To make my results more reliable I will repeat each concentration gradient 2 more times so that I have a wide range of results. From this I will calculate their averages and from the averages I will calculate the rate of reaction.
Accuracy
To make my results more accurate, I will measure them will extreme care and will be precise. I will make sure that I measure all the liquids at the lower meniscus and not at the upper meniscus. I will make sure that I start the stopwatch as soon as I pour all the substances into the test tube. Then I will stop the stopwatch as soon as the substances in the test tube turn colourless.
Safety
In order to conduct a safe experiment, I will follow all the lab rules of the classroom and will behave in a respective and mature manner. I will not do anything dangerous that would put any fellow classmates at risk. I will wear safety goggles and will clear my work area of any unnecessary equipment.
Apparatus
I will make this experiment safe by following the lab instructions of the classroom.
Method
- Pour 5 ml of starch into the measuring cylinder.
- Use one Pipette to make sure that you have measured exactly 5 ml.
- Pour that into a test tube.
- Put the test tube into the test tube rack.
- Heat water in a kettle.
- Pour 100 ml of boiling water into a beaker to create a water bath.
- Put the thermometer in the water bath.
- Add a sufficient amount of cold water into the water bath so that the temperature of the water bath is at 40°C.
REMEMBER: DO NOT FORGET TO MAINTAIN THE TEMPERATURE OF THE WATER BATH, THE TEMPERATURE MUST REMAIN AT 40°C AT ALL TIMES!
- Measure 5 ml of amylase into the small beaker.
- Use the other Pipette to make sure that you have measured exactly 5 ml.
- Use the Pipette to take 1 ml of amylase from that.
- Put the test tube into the water bath.
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Take 3 drops of iodine and add them into the test tube of starch at the same time as the amylase. [THE SOLUTION SHOULD TURN BLACK]
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Start timing immediately using the stopwatch.
- Stop timing when the solution turns clear.
- Record your results in your results table.
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Repeat 1 – 16 two more times.
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Repeat 1 –17, however this time use 4 ml of amylase and 1 ml of water, next time use 3 ml of amylase and 2 ml of water, now use 2 ml of amylase and 3 ml of water, finally use 1 ml of amylase and add 4 ml of water.
- Calculate the average time by adding up all your results and then dividing the answer by the number of results you have for that particular concentration gradient.
- Calculate the reaction rate by dividing 1 by the average time.
- After recording all your results tidy up the equipment.
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Finally draw a graph of your results. Label the y-axis rate of reaction and label the x-axis concentration of amylase.
Empty Table of Results
I have decided to use these values on my table based on the results of the preliminary work that I carried out prior to this experiment. I will go up every 20% as I feel that is a sufficient gradient that will give me a reliable graph.
Empty Graph
I have decided to use these values on my graph based on the results of the preliminary work that I carried out prior to this experiment. My preliminary work consisted of short experiments know as ‘the spotting tile experiments’. I had to put drops of amylase and starch along with iodine solution onto a spotting tile. The amylase has to be at a certain temperature. I would then time how long the reaction took and then would record that onto a table of results. I would then divide 1 by the time to obtain the rate of reaction. I obtained 0.018 t-1 as the highest reading for the rate of reaction; hence this is the highest reading on my graph. The highest value on the x-axis, concentration of amylase is 100% as there is no value higher.
This is a diagram of my preliminary work: -
Table of Results
I will now draw another table in which I will show the rate of reaction and the points to be plotted on my graph.
Draw the graph yourself!
Conclusion
Both my graphs have show that my predictions were correct. I had predicted that the points on my graph would slowly increase until they reached a certain point where they would stay the same/stable this is called the V-Max. I had predicted that my graph would look like this:-
This has happened because the rate of reaction slowly increases as the concentration of amylase increases. For example if there are 50 starch molecules and 5 amylase molecules then each amylase molecules would have to break down 10 starch molecules, this would take a long time. But if there were 50 starch molecules and 25 amylase molecules then the reaction would be 5 times quicker that 50 starch and 5 amylase. However if there were 50 starch molecules and 50 amylase molecules then the reaction would be at its quickest because each amylase molecule would have only to break down one starch molecule. Now if there were 50 starch molecules and 100 amylase molecules the reaction time would be as quick as 50 starch molecules and 50 amylase molecules. This is because on 50 amylase molecules are needed to break down the 50 starch molecules, meaning that the other 50-amylase molecules do nothing. This is called the V-Max.
To explain the trend of my graph better, I will split it up into three sections: -
At point ‘a’, the rate of reaction is increasing slower than proportion to the concentration of amylase. This is because the amylase would have to work quicker to break down all the starch. At point ‘b’, the rate of reaction is increasing in proportion to the concentration of amylase. This is because the concentration of amylase would be at a point where there might be 5 starch molecules to every amylase molecule. At point ‘c’, the rate of reaction is increasing quicker than proportional to the concentration of amylase. This is because at this point there might be two starch molecules to every amylase molecule, gradually becoming one starch molecule to one amylase molecule. Hence my graph is a curve.
My results support my original predictions because I had predicted correctly using the scientific knowledge I have gained from various sources and also from the preliminary work I had done prior to this experiment. Evidence of my predictions being correct is shown in my graphs.
Evaluation
This experiment was a fair test because I only changed one variable and left all other variable the same where possible. For example I could not change the room temperature because I do not have control over it. I only changed the concentration of amylase.
To make sure that I measured accurately, I poured the liquids slowly and looked very carefully into the measuring cylinder to see where I was meant to stop pouring. When timing with the stopwatch I tried to stop the stopwatch as soon as I saw the solution go clear, however there area for human error here and what might look clear to me might not look clear to a classmate of mine. To see how much human error myself, could cause, I tested my reactions, the test consisted of starting and stopping a stopwatch, the lower down the time the quicker your reactions. On average I got 0.16 seconds, but the most I got was 0.07 seconds, which is quite quick. So you could estimate that I could be + 0.16 seconds on all the times I have measured because that was my average reaction time.
I believe that the procedures or method I used to carry out this experiment are not the most accurate procedures that could be used to carry out this investigation. Another method you could use would be the use of a computerised system, you could use a computer appliance to drop the iodine and concentration of amylase into the starch and start timing as soon as the iodine and amylase mix with the starch. The results that could be obtained from this experiment would be more reliable because there would be very little area for human error.
I believe that my results are quite reliable because I repeated the experiment twice to give me three sets of results from which I calculated averages. My results in the end followed the trend that I said would occur in my prediction, so this means that my prediction was correct. Most of my results were accurate, however some were not because they were anomalous results and did not follow the trend.
If I could do the experiment again, I would improve it by trying to cut out human error. By this I mean that would be more careful whist measuring and would also be quicker in starting the stopwatch. If I were to do the experiment again I would prefer to use a different method such as the method I suggested earlier, a computerised method rather than using this same method again because the computerised method is more accurate and has less of a margin of human error. However the method I used can produce very inaccurate results if I was not careful in measuring and also has a larger margin for human error.
Another alternative method is to use a colorimeter to measure the colour of the amylase, starch and iodine solution, to see when the solution has turned colourless. By using this method you are able to cut out all human error when it comes to measuring the colour of the solution. I could also use a syringe to measure the volume of the liquids before pouring them into the test tube instead of a measuring cylinder. A syringe has smaller scale divisions, of which the thickness of the line is even very small. A measuring cylinders division are large and the thickness of the line on it is quite thick. This would enable be to be more accurate.
Results Reliability
I believe that my results are reliable and sufficient because all the points are on a straight line, the tests covered enough range to show the full shape of the graph, as a result I am certain that I have got enough results to be sure about my conclusion as being correct, acceptable.
Here is a graph to show the reliability of my results: -
The error bars in red on the y-axis show the percentage error of 10 for the time taken for each concentration gradient.