Adam Jon Sher 10BL1
Investigating the rate of reaction of the enzyme catalase in a potato
Planning
It is the purpose of our investigation to discover whether the temperature of the enzyme catalase affects its’ turnover rate. The experiment that we will carry out to investigate this hypothesis will involve using the catalase in the cells of a potato (a Wilja potato to be exact) and reacting it with a test tube of hydrogen peroxide.
When catalase meets hydrogen peroxide, it turns the hydrogen peroxide into water and oxygen. The full equation is 2H2O2 2H20 + O2. This is the equation on which we are basing our investigation. We are going to drop pieces of cut potato into hydrogen peroxide, because the potato is cut then catalase will be present on the cut surfaces of the potato, and will therefore make contact with the hydrogen peroxide. The oxygen bubbles released will buoy the piece of potato up to the surface, and we will time how long this takes to happen. We think that the temperature will affect the turnover rate of the catalase (i.e. how many hydrogen peroxide molecules that it can break down in one second), and that oxygen bubbles will therefore be released quicker and the potato will rise to the surface quicker at a certain temperature. One of the things that we will find in this investigation is the temperature at which catalase works best, which will be the temperature at which the potato rises fastest.
We are going to measure out 5cm of hydrogen peroxide in a test tube. The reason that we are measuring vertical height rather than volume is because we are going to time how long it takes for the piece of potato to rise back up to the surface of the hydrogen peroxide by the oxygen bubbles released in the reaction, this means that the depth that the potato has to fall through is more crucial than the amount of hydrogen peroxide that surrounds it. All the test tubes used must be of the same height, so that in each experiment we will be dropping the potato from the same height.
We will use a cork borer to take out a section of potato, and then we will slice that into pieces 3mm thick. These will be the pieces of potato that we will use in all our experiments. If we use the same potato each time, then it is more likely that the same amount of catalase will be found on each piece, whereas if we used two different potatoes, the likelihood is that the slices from one potato would have more catalase than the slices of another.When not in use, these pieces of potato will be folded in a moist paper towel to stop them from drying out.
We will be varying the temperature of the hydrogen peroxide as well as the temperature of the potato, and therefore the temperature of the catalase. This will be done by heating up a beaker of water using a Bunsen burner to the desired temperature, and placing two test tubes in it. The first will contain the hydrogen peroxide, and the second will contain the piece of potato, which will be wrapped in a moist paper towel inside the test tube so as not to evaporate the chemicals on the surface of the potato. We will leave the beaker under the Bunsen burner, and we will take the temperature of the water every so often until the correct temperature is reached. At this point we will turn off the Bunsen burner, and drop the piece of potato into the hydrogen peroxide. We will use tweezers so that the oil on our fingers does not get onto the surface of the potato and block the catalase from the hydrogen peroxide. We will time using a stopwatch from the point that we drop the potato until the point that the potato hits the surface of the hydrogen peroxide on its’ way back up.
We will record the result and we will repeat each temperature value five times for accuracy. If there is a case of a clear anomalous result then we will repeat the experiment again for that value. The temperatures that we will use will be 0°C, 10°C, 20°C, 30°C, 40°C, 50°C and 60°C. We will calculate the mean of our results and we will call this value the correct value that we will plot in the graph. This is a suitable range with which to draw a conclusion, because preliminary work showed that the enzyme will denature at 60°C. We should get results that are accurate enough with these temperatures without having to measure every 5°C, but if we wanted to make our results more accurate, then this would be a good way to start.
We will ensure that we carry out a fair investigation by keeping the variables down to a minimum. The only thing that we are changing is going to be the temperature of the experiment. The variables that we must try to keep constant are the depth and volume of the hydrogen peroxide, and the weight and surface area (and therefore the amount of catalase) of the potato. We hope to achieve the same amount of catalase on each piece of potato by keeping the surface area the same because the catalase is found inside the cells, and therefore only on the cut surfaces of the potato. As we are using a cork borer to extract the section of the potato that we will be using, the circumference of each piece of potato will be identical, and hopefully each 3mm slice of potato will be the same weight, as a heavier slice of potato would need more oxygen bubbles to raise it to the surface. We will ensure that the hydrogen peroxide is taken from the same bottle to ensure that it is equally diluted in all the experiments. We will take all the potato cuttings from the same potato to ensure that they contain similar amounts of catalase, and we will conduct all the experiments on the same day, to ensure that room temperature and pressure remain constant throughout. We will use the same thermometer (0-100°C, because preliminary research showed that temperatures would exceed 50°C, and that we would therefore need a thermometer with a larger range than the 0-50°C thermometer) to measure the temperature of the water, and the same ruler (0-015cm) to measure the depth of the hydrogen peroxide, again for consistency in measurement because any error in the measuring apparatus will be consistent throughout all the readings.
Here is a diagram of the experiment that we will carry out:
In order to predict how catalase is affected by temperature first we must understand how this enzyme works. Enzymes do not work equally well at all temperatures, if the temperature is too low or too high the enzyme can be denatured, which means that the protein strings that make up the enzyme begin to unravel and fall apart. We must remember that this enzyme is inside a potato, so it will logically function best at the temperature at which the potato would grow in its’ natural habitat. Potatoes originally come from the Peruvian-Bolivian Andes, and the average summer temperature there is around 21°C. Plant enzymes also work over a larger temperature range than mammalian enzymes because unlike mammals, they have no way of controlling their ownliving temperature.
This is what the enzyme catalase looks like:
Enzymes and catabolic reactions can be easily explained using the lock and key hypothesis. This states that each substance is like a lock, and only the correct enzyme, which is the “key” to the lock, can catabolise it. The helps to explain exactly what the catalase is doing, which is being a catabolic “key” to the “lock” which in this case is hydrogen peroxide. If an enzyme gets too hot or too cold and denatures, then the shape of the enzyme is changed. This means that the shape of the “key” is changed and will no longer fit the “lock”, which means that the catalase can no longer break down hydrogen peroxide because it has distorted out of shape.
From the above information, I can make several informed predictions about the results of this experiment. If a graph were drawn that plotted temperature against the rate of reaction, I predict that a gradual curve would be plotted that showed the rate of reaction to be proportional to the temperature, similar to the one in the graph shown below. This is because when an object is cold, there is less kinetic energy present and the particles would vibrate slowly, but it there were more kinetic energy present, then the particles would vibrate harder and faster, which means that the reactivity (rate of reaction in this case) would increase because more collisions would occur every second. This can also be explained as the fact that heat acts as a catalyst, something that speeds up reactions, so one would expect heat to act as a catalyst in proportion to how much heat there is.
I expect the experiment to peak (at which point the rate of reaction would cease to be proportional to the temperature) at about21°Cbecause this is the average summer temperature in the Peruvian-Bolivian Andes where potatoes originated. I also think that the catalase would work over a wide range of temperatures, because plants have no way of controlling their own living temperature, so their enzymes must be able to withstand quite large fluctuations in the temperature.Also, potatoes are grown here in England, so we can assume that they can also deal with the colder climates of England as well as the warmer climates of places like France or Peru where potatoes are also grown. I think that the rate of reaction will read 0 at about 60°C, because in my preliminary work, no oxygen bubbles were released at 60°C, which must mean that the enzyme has been denatured.
On the y axis of the graph we will plot 1/t because we are measuring the rate of the reaction, not the time. On the x axis we will plot the temperature that the catalase was at when the experiment took place, this would be measured in °C.
This graph would show the catalase working on catabolising the hydrogen peroxide at low temperatures, but the turnover rate increasing in proportion as the temperature rose. This would peak at the optimum temperature and then descend as the temperature got so hot that the catalase started to distort and work less efficiently in its’ distorted state. When the temperature got even hotter the enzyme would denature and stop working, which would be shown by a sudden drop on the graph, ending at zero.
There are a few safety issues that we need to take into account during this experiment. Firstly, we must wear goggles as hydrogen peroxide is corrosive and it is extremely dangerous if it gets into your eyes.Secondly, we must take care when boiling water, as apparatus can get hot and boiling water can scald the skin.
Obtaining Evidence
Here is the results table that I will enter the results into:
Temperature(°C) / Time (1)
(s) / Time (2)
(s) / Time (3)
(s) / Time (4)
(s) / Time (5)
(s)
0
10
20
30
40
50
60
In case of anomalous results, repeat readings here:
Temperature(°C) / Time(s)Calculations and final data:
Temperature(°C) / Mean Time (s) / Rate of Reaction (1/t)
0
10
20
30
40
50
60
Analysing Evidence
If we look at the affixed graph, and at the results table, we can see that catalase is most efficient at or around 40°C. We can also see that it will still work anywhere between 0°C and 59.9°C. The enzyme denatures at 60°C, that is it completely breaks apart and will not work at all, but from 50°C the rate of reaction drops rapidly, showing that the enzyme’s turnover rate is being severely reduced.
We can see that there were quite a number of anomalous results that we discarded when we drew the graph, for reasons as to why these anomalous results may have occurred, please see the evaluation section of this investigation.
From 0°C to 40°C, we can see a steady rise as we are nearing the optimum temperature that catalase operates at, but the catalase is still working at 0°C, which means that potatoes can be grown in quite cold climates, but not necessarily at 0°C due to other factors such as water freezing and therefore not flowing to where the plant is situated.
We can also see that the increase from 0°C to 40°C was roughly proportional to the temperature of the catalase, as at 10°C the rate of reaction was 0.05, whereas at 30°C, which is thrice as hot, the rate of reaction was thrice as much at 0.15. These diagrams may help to explain why this occurs:
The conclusion that I have drawn from this evidence is that the enzyme catalase can work at a range of temperatures, which it must be able to do because plants cannot control their own living temperature, and in places like Peru where potatoes originated, the temperature can be as low as 9°C and as high as 40°C in one year. The potato must be able to survive these conditions, otherwise it would die out during a hot summer or a cold winter. The enzyme catalase is most efficient (i.e. it has the highest turnover rate) at 40°C and it denatures (will not break down hydrogen peroxide at all) at 60°C. This was shown in the experiment because as we conducted the experiment at 60°C, the potato sank to the bottom and no oxygen bubbles were produced.I can finally conclude that the rate of reaction of catalase is directly proportional to the temperature at which it is acting between 0°C and 40°C.
My conclusion does not accurately support my prediction because I stated that 21°C was the optimum temperature for the enzyme catalase, as that is the average temperature of a potato in its’ native Peru during the growing season.After conducting the experiment, we have found that in fact 40°C is the optimum temperature at which catalase works. For reasons as to why there is such a large gap between my prediction and my conclusion, please see the evaluation section of this infestation.
Nevertheless, my conclusion supports my prediction about the rate of reaction being proportional to the temperature until the rate of reaction peaks, and also about the catalase becoming denatured at 60°C, which was shown on the results table as the catalase failed to react with the hydrogen peroxide at all, and consequently no oxygen bubbles were given off and the potato did not rise to the surface.
Evaluation
My procedure was good enough to enable me to collect valid evidence, but it was far from perfect, as the high number of anomalous results showed. The evidence obtained was accurate once it was processed,because it could be explained using scientific knowledge, but there were eight anomalous readings that were recorded. These readings were:
Temperature (°C) / Reading (s)0 / 15
0 / 18
0 / 14
10 / 4
10 / 6
20 / 4
30 / 17
30 / 42
The procedure was simple to carry out, and it gave us good results, but it could be improved greatly. There were too many variables that we could not control, such as room temperature, because although my plan stated to conduct all experiments on the same day, there simply was not enough time to do so, and the data had to be collected over a number of days. We could alsonever be certain that each slice of potato contained the same amount of catalase because even if they had the same surface area, some pieces might have had more catalase on that surface area than others.Also, the hydrogen peroxide would expand whilst heated, affecting the depth, and our experiment failed to take this into account meaning that our results at higher temperatures could be inaccurate as the potato had to rise through a greater depth of water than the potatoes that were placed in the lower temperature hydrogen peroxide.
One idea that occurred to me was that the way of calculating how much oxygen given off would have been much simpler if we had simply collected it in a gas syringe rather than attempting to make it buoy pieces of potato up to the surface. If we collected it in a gas syringe, and the room temperature and pressure was kept constant throughout all the readings, then this would have been a more accurate way to gauge the reactivity of thecatalase because the amount of oxygen liberated could have been simply read off the side of the gas syringe.
If we wanted to make the results more accurate but without changing the method too much, we could have simply repeated the experiment several times, or we could have measured every 5°C instead of every 10°C, which would have meant that we could see if the line of best fit on the graph that we have already drawn is accurate, or if it needs adjusting.
I think that the evidence represented here is reliable, because I expected my results to plot a very similar graph to the one that was plotted. It definitely supports the conclusion, but I think that further work on different varieties of potato is needed if I am able to state that catalase will always react in this way. I therefore think that the evidence is not sufficient to support the conclusion that catalase will always react in this way, but I was able to back up the evidence with scientific knowledge, so it seems likely that catalase would indeed act in this way for other types of potato.
The eight anomalous results could have been caused by any number of variables. There were two bottles of hydrogen peroxide that we used, and one may have been diluted more than the other. We might have let some of the pieces of potato on the side longer than others, giving them a chance to dry out before we could conduct our experiment. The anomalous results could also have been caused by the temperature of the experiment dropping or rising whilst we were picking up the potato with the tweezers, which would have given us inaccurate results.