The concentration of the substrate molecule, or hydrogen peroxide will affect the enzyme rate of activity because if the hydrogen peroxide is more concentrated, there will be more molecules to collide with the enzyme and the reaction will produce more product molecules quicker. To control this we will use the same concentration of hydrogen peroxide for all our experiments.
Enzymes are biological catalysts. They are proteins and aid metabolism within the body by catalysing metabolic reactions. They speed up reactions without themselves being affected by the reaction. They take part in both anabolic reactions, which build up large molecules from smaller ones, and catabolic reactions, which breakdown larger molecules into smaller ones, the breakdown of hydrogen peroxide catalysed by catalase is an example of catabolism. Enzymes take their action because of their specific shape, much work with only one kind of substrate, such as catalase with hydrogen peroxide. They work by giving the substrate molecules a surface to stick to where they can collide with other molecules if they need to, this increases the number of collisions and rate of reactions as without this surface to stick to the molecules would not collide as frequently. Enzyme specificness can be explained by the ‘Lock and Key’ hypothesis.
‘The Lock and Key Hypothesis’
Enzymes are proteins folded into a complex, three-dimensional shape. They have an active site that allows them to act as a catalyst. The active site is a certain shape and only one molecule of substrate, of the same shape will fit into the active site of the enzyme. The molecule of substrate will now react to form a molecule of product, which will leave the active site. The enzyme lowers the energy needed for the reaction, and the reaction is then much more likely to take place. In our investigation, the molecule of substrate is hydrogen peroxide, the enzyme is catalase and the molecules of product are oxygen and water. This hypothesis can be affected by temperature and pH because if the enzyme becomes denatured by either a temperature that is to high, or if the pH is to acidic or alkali and the active site becomes the wrong shape for the specific substrate molecule to fit. Enzymes can be used over and over again though as long as they do not become denatured.
‘The Collision Theory’
‘The Collision Theory’ states that the rate of reaction depends on how often and how hard the reacting particles collide with each other. That the particles have to collide with each other to react, and they have to collide hard enough. So more collisions increases the rate of reaction
This is relevant for the temperature affecting the activity of an enzyme because when the temperature is increased the particles all move quicker. If they’re moving quicker, they’re going to have more collisions. For this investigation it means that when the temperature is increased the hydrogen peroxide molecules and the catalase molecules move quicker, so there will be more collisions of the hydrogen peroxide with the active site of the catalase, increasing the rate of reaction, until the catalase becomes denatured.
This is relevant for the surface area affecting the activity of an enzyme because if one of the reactants is a solid, in this investigation, the potato, which is the source of catalase, breaking it up into smaller pieces, will increase its surface. This means the particles around it in the solution, hydrogen peroxide, will have more area to work on so there’ll be more collisions between the two particles.
SMALL SURFACE AREA LARGE SURFACE AREA
(broken up)
This is relevant for the concentration of the potato, enzyme or hydrogen peroxide affecting the activity of an enzyme because if they are made more concentrated it means that there are more particles that makes collisions more likely.
LOW CONCENTRATION HIGH CONCENTRATION
In our investigation all of the factors that will affect the enzyme activity rate are variables. Temperature is an independent variable, a variable that is changed by the experimenter to see what happens, it is also a continuous spectrum because it can have any value over a continuous range.
The volume of oxygen released is a dependant variable, one which changes as a result of what happens. It is also a continuous spectrum too.
All the other variables are controlled variables. Variables that would affect the result, but are fixed throughout the experiment. These are the pH, surface area of potato and concentrations of potato, enzyme and hydrogen peroxide, these are continuous spectrums. The use of a potato is a categoric variable because other sources of the enzyme could be used, such as liver.
To make sure that we can be sure of cause and effect with the enzyme and that it is a control, we must make sure that temp alone doesn’t cause the hydrogen peroxide to degrade. To test this we will put a boiling tube of just hydrogen peroxide in the water bath and set it up to collect any oxygen evolved, and another boiling tube set up with the catalase in the potato and the hydrogen peroxide. If oxygen is collected with the enzyme present and not without, then the enzyme is a control. But if is oxygen is collected with and without the enzyme present, then the enzyme is not a control.
We must also make sure that hydrogen peroxide doesn’t break itself down with the passage of time. We will set the hydrogen peroxide up in a boiling tube to collect oxygen and see if any is produced.
These controlled experiments make sure that the investigation is a fair test.
We must also make sure that the passage of time doesn’t affect the enzyme. To make sure of this we will test different aged potatoes.
To make sure that the results we will obtain are as precise as possible we will put the boiling tube with the hydrogen peroxide and enzyme in a water bath to keep the conditions at a constant temperature while a certain temperature is being investigated. This is because the water in the water bath will buffer temperature change, so if the temperature changes in the room, the temperature in the water bath will stay the same and the temperature that the hydrogen peroxide is reacting at will stay the same. Water has a high heat capacity; this means that a lot of heat energy is needed for breaking bonds because the bonds between the hydrogen and oxygen atoms in water are very tight so the temperature won’t change until the bonds have been broken.
Bonds breaking
Ammonia
Water
Rate
Heating up
Heating up
Heat
This graph shows how the heat capacity of water is much more than of ammonia because the ammonia has a steeper gradient than the water. This means that ammonia will heat up much more easily than water.
We will deliberately not vary the pH, surface area of potato, concentration of potato, enzyme and hydrogen peroxide throughout the investigation to make sure that it is a fait test and because no potato is the same because of their age, type or crop, we will use the same source of enzyme, the same potato, to make the investigation a fair test.
Prediction
In this investigation, I predict that, as the temperature rises, the rate of enzyme activity will increase until the high temperatures denature the enzyme and the rate of enzyme activity will decrease. The enzyme will denature because when the temperature increases the atoms within the enzyme molecule will begin to vibrate, at first the rate will be low because the enzyme will not be vibrating much so it will not have much energy for catalysing the reaction. The more the temperature increases the more the atoms will vibrate which will begin to knock the enzyme out of shape and denature it until the active site of the enzyme will no longer match up with its specific substrate molecule.
The collision theory also states that as the temperature increases, the rate of reaction increases because the particles move about faster.
The collision theory states that the rate of reaction depends on how often and how hard the reacting particles collide with each other. That the particles have to collide with each other to react, and they have to collide hard enough. So more collisions increases the rate of reaction
When the temperature is increased the particles all move quicker. If they’re moving quicker, they’re going to have more collisions. This means that when the temperature is increased the hydrogen peroxide molecules and the catalase molecules move quicker, so there will be more collisions of the hydrogen peroxide with the active site of the catalase, increasing the rate of reaction, until the catalase becomes denatured.
This is a sketch graph showing what I predict should happen. It shows the rate gradually increasing as the temperature rises until the enzyme reaches its optimum and then as the enzyme denatures the rate drops off.
Rate
Temperature
In the investigation I am going to investigate the five temperatures of;
Use hydrogen peroxide of concentration 25%, and measure how much oxygen is evolved within a time period of 5 minutes. These were decided with the use of preliminary experiments.
We used preliminary experiments to find out the best concentration and temperatures to work with because, if the concentration of the hydrogen peroxide was to high, then too much oxygen would be produced to quickly to collect and therefore it would become hard to gain accurate results, or if it was to low, then not enough oxygen would be produced to record.
If the temperatures were to high or low then not enough oxygen would be produced to record. The preliminary experiments gave us a rough idea of where the optimum temperature for the enzyme was so that we could focus our main experiment within that temperature region.
We will repeat each experiment three times, this is to reduce experimental error and increase precision and accuracy of the results by working out the average of the three results. We will also use the same apparatus for each experiment to make it a fair test.
If any anomalous points occur during the experiments I will have a sketch graph available to identify any as they occur so that they can be repeated.
Method
- Set the apparatus up as shown.
Delivery Tube
Boiling Tube
Thermometer
Measuring Cylinder filled
with water being displaced
by oxygen bubbles
Water Bath
Bowl
Water
Potato
Hydrogen Peroxide
Stopwatch
-
Measure out 25cm of hydrogen peroxide in a measuring cylinder and pour it into the boiling tube. Turn on the water bath to 20°C on the thermometer and put the boiling tube in so that the hydrogen peroxide can settle at 20°C.
- Bore 3 pieces of potato, remove the skin and then cut them down to a length of 4 cm. Then cut the potato down the middle once and across six times so that there is a total of twelve pieces for each sample of potato.
- Put the potato sample into the hydrogen peroxide and as soon as the last piece enters start the stopwatch and put the delivery tube onto the top of the boiling tube (make sure that the measuring cylinder for collecting the oxygen is full of water so that it can be displaced by any oxygen evolved).
- Check the temperature on the thermometer in water bath every minute to make sure that it doesn’t change.
- After 5 minutes record the total volume of oxygen evolved in the measuring cylinder and reset the stopwatch.
- Repeat this process two more times with the remaining two samples of potato.
- Repeat steps 1 to 7 with the other temperatures;
Preliminary Experiment
Temperature: - 30°C
The temperature produced a good volume of oxygen so we will work with temperatures around 30°C. The concentration at 25% also gives the most constant results so we will work with that concentration of hydrogen peroxide.
This preliminary experiment helped me to decide what concentration to work with and what temperature to work with as before I would have just had to do a broad range of temperatures from 0°C to 100°C and therefore the results would not be very precise as we would not focus around where the optimum temperature may be.
It helped me to decide to use a thinner measuring cylinder for collecting the oxygen for more precise results, as it would have a smaller, more precise gradient for measuring the volume.
Results
Tally Chart Showing Temperature checks every minute for 5minutes
Sketch Graph Used During Experiment
Analysis
The line of best fit on the graph of results shows a positive correlation up to 30°C and then a sudden negative correlation that starts off with a steep gradient, which levels out towards 80°C.
This is because, as the temperature rises, the activity of the enzyme increases as the atoms within the enzyme begin to vibrate more vigorously, but at 30°C the vibrating atoms begin to knock the enzyme out of shape and it becomes denatured and its active site will no longer combine with the hydrogen peroxide so the volume of oxygen being produced, and rate of enzyme activity decreases and the correlation of the graph comes down. The graph and results suggest that the optimum temperature at which catalase works is 30°C.
The collision theory also means that the rate of enzyme activity and volume of oxygen evolved increases because of how often and how hard the reacting particles collided with each other. The particles have to collide with each other to react, and they have to collide hard enough. So more collisions has increased the rate of reaction
When the temperature was increased the particles all moved quicker, because they’re moving quicker, they have more collisions. So when the temperature increased the hydrogen peroxide molecules and the catalase molecules moved quicker, so there was more collisions between the hydrogen peroxide and the active site of the catalase, increasing the rate of reaction, until the catalase became denatured.
The results turned out as I predicted they would. My prediction said that in this investigation, as the temperature rises, the rate of enzyme activity will increase until the high temperatures denature the enzyme and then the rate of enzyme activity will decrease. The enzyme will denature because when the temperature increases the atoms within the enzyme molecule will begin to vibrate, at first the rate will be low because the enzyme will not be vibrating much so it will not have much energy for catalysing the reaction. The more the temperature increases the more the atoms will vibrate which will begin to knock the enzyme out of shape and denature it until the active site of the enzyme will no longer match up with its specific substrate molecule.
The collision theory states that as the temperature increases, the rate of reaction increases because the particles move about faster.
It states that the rate of reaction depends on how often and how hard the reacting particles collide with each other. That the particles have to collide with each other to react, and they have to collide hard enough. So more collisions increases the rate of reaction
When the temperature is increased the particles all move quicker. If they’re moving quicker, they’re going to have more collisions. This means that when the temperature is increased the hydrogen peroxide molecules and the catalase molecules move quicker, so there will be more collisions of the hydrogen peroxide with the active site of the catalase, increasing the rate of reaction, until the catalase becomes denatured.
This sketch graph shows what I predicted should happen. It shows the rate gradually increasing as the temperature rises until the enzyme reaches its optimum and then as the enzyme denatures the rate drops off. It matches the results graph almost perfectly.
Rate
Temperature
The results for each experiment on the graph are close together. This gives me more confidence in the results that they are reliable. The only results that aren’t close together are the results that were taken at 36°C. One of the results is far apart from the other two, which are close together. This makes this set of results slightly less reliable but because all the other results are reliable this anomalous point can be excused because of experimental error.
Evaluation
I believe that my investigation was a success because my results and evidence are close together, making them reliable and giving me confidence in my conclusions and they show my prediction to be correct. I think my results were accurate enough for the task and they showed that the optimum temperature that catalase works at is 30°C.
There was one anomalous result in my results, which was the result from experiment 3 for the temperature of 36°C. This is most likely to be there because of experimental error.
To improve my experiment, I could have finished it off before I had to leave and put the potato in a bath of water. This is because leaving the potato in water for the prolonged period of time may have allowed it to be affected by osmosis. When we came back to complete the investigation the potato had swollen up in size. The action of osmosis, the involuntary diffusion of water between a semi-permeable surface to balance out the concentration of water between the two surfaces, may have resulted in the concentration of the catalase being lowered so this may have affected the activity rate of the enzyme. According to the collision theory, if the concentration of he enzyme is less concentrated, it means that there are less particles to collide reducing the likelihood of collisions.
HIGH CONCENTRATION LOW CONCENTRATION
Other ways in which I could have improved the investigation to try and get even more accurate evidence is;
- We could have used alternative equipment in the same conditions and seen whether the results are the same.
- Use a manometer instead of a measuring cylinder to collect the oxygen and compare the results.
- Use a thinner measuring cylinder to collect oxygen because it would be more accurate.
- Use a more precise, digital thermometer because it would give us more accurate temperatures to more decimal places.
- Take more results around where the optimum is to have more confidence about the results.
The method, which I used, is reliable and can be counted on to give accurate results. The use of a sketch graph during the experiments helped me with this because it allowed me to identify if a large amount of anomalous results appeared and then they could be repeated. Also, there are no evident random errors, errors that have aroused by fault of myself or any systematic errors, an error that has aroused because of fault with the apparatus.
I think I have enough evidence to draw a conclusion, as most of my results are close to the line of best fit, which gives me, confidence that my results are correct. Also, all the data agrees with the hypotheses, theories and predictions making it seem accurate, correct and reliable.
To obtain more evidence to support my conclusion I could compare my results with others to identify any systematic errors and if the results were the same it would give me even more confidence in them. You could use other methods of measuring enzyme activity; use a different source of catalase, such as liver. Or use just catalase, extracted from the source and if the same results were always produced then that would support my conclusion solidly.
CONCLUSION
The optimum temperature at which catalase catalyses a reaction at the quickest rate is 30°C. As temperature increases during an enzyme catalysed reaction, the enzyme activity rate increases as well until the enzyme reaches its optimum and is denatured by the vibrations of its atoms becoming too vigorous and knocking it out of shape because the temperature rise has caused them to gain more energy. Then, as the temperature rises, the enzyme activity rate drops because the enzyme is no longer the right shape to catalyse the reaction.
