How does the change in temperature affect the rate of catalase reaction?

Matthew Smith 11 Brown 23.10.2002

Aim

To investigate the effect of temperature on the enzyme catalase.

Hypothesis

An enzyme is a protein molecule that speeds up chemical reactions in all living things. Without enzymes, these reactions would occur too slowly or not at all, and no life would be possible. All living cells make enzymes, but enzymes are not alive. Enzyme molecules function by altering other molecules. Enzymes combine with the altered molecules to form a complex molecular structure in which chemical reactions take place. The enzyme, which remains unchanged, then separates from the product of the reaction. Therefore, an enzyme is a sort of biological catalyst.

The molecule of hydrogen peroxide approaches the catalyse enzyme.

The hydrogen peroxide then enters the enzyme.

The hydrogen peroxide is turned into oxygen and water by the enzyme.

Enzymes are classified into several broad categories, such as hydrolytic, oxidising, and reducing, depending on the type of reaction they control. Hydrolytic enzymes accelerate reactions in which a substance is broken down into simpler compounds through reaction with water molecules. Oxidising enzymes, known as oxidises, accelerate oxidation reactions; reducing enzymes speed up reduction reactions, in which oxygen is removed.

Catalase is present in the peroxisomes of nearly all aerobic cells. It serves to protect the cell from the toxic effects of hydrogen peroxide by catalysing its decomposition into molecular oxygen and water without the production of free radicals (An atom or a group of atoms with an unpaired electron. Radicals are unusually reactive and are capable of causing a wide range of biological damage).

Hydrogen peroxide + catalase = oxygen + water

Using this scientific knowledge, I predict that increasing the temperature to 35-40ºC will create the fastest reaction. I predict this because catalase are used to speed up biochemical reactions in the body, so they are used to working at body temperatures, and 35-40ºC are typical temperatures. However the cataylse not react quicker than this at temperatures over 40ºC, because these temperatures will cause the catalase to denature, and therefore the reactions will be slower. This is shown in the standard graph below.

Method

The apparatus needed for this experiment is conical flask, hydrogen peroxide, yeast, 10cm3 measuring cylinder, scales, gas syringe, clamp stand, boss head, delivery tube, bung, spatula, measuring boat, stop clock, kettle, ice and a thermometer. The apparatus was used for the following; conical flask to hold the hydrogen peroxide and yeast, 10cm3 measuring cylinder to measure out the volume of hydrogen peroxide, yeast to act as the catalase, hydrogen peroxide to react with the yeast, scales to weigh out the correct mass of yeast, gas syringe to collect the reacted gas, clamp stand and boss head to hold the gas syringe in place, delivery tube to attach the conical flask to the gas syringe, bung to stop any gas escaping the conical flask, except for the gas that goes into the delivery tube, measuring boat to store the yeast, spatula to transfer yeast from tin to measuring boat, stop-clock to time the readings, a kettle to raise the temperature of the hydrogen peroxide, and ice to lower the temperature and the thermometer to check the starting temperature.

Measure out the required mass of yeast using scales, spatula, and measuring boat. Then measure out the required amount of hydrogen peroxide using the 10cm3 measuring cylinder. Attach the gas syringe to the clamp stand using the boss head. Then attach the delivery tube to the end of the gas syringe. Put the hydrogen peroxide into the conical flask and measure the temperature, to raise and lower the temperature of the hydrogen peroxide use a kettle and ice. Add the yeast to the hydrogen peroxide and then quickly but the bung with the delivery tube through it onto the end ...

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The equipment was set up as shown below,

Trial Experiment

The purpose of the trail experiment was to determine the mass of yeast, the volume of hydrogen peroxide, the time between each reading, and the number of readings that were to be taken. The temperature used for my trail experiment is always 25ºC,

because this was room temperature, and therefore it is the easiest temperature to obtain. The results of the first trail experiment are shown below,

Mass of yeast – 2 grams

Volume of Hydrogen Peroxide – 15 cm3

The reason that not all of the results are shown is because even at 25ºC, the reaction was so great that the hydrogen peroxide and yeast started to rise up the delivery tube, and into the gas syringe. Therefore my next trial experiment only used 1 gram of yeast, and 10 cm3 of hydrogen peroxide.

Mass of yeast – 1 gram

Volume of Hydrogen Peroxide – 10 cm3

The same problem occurred with this mass of yeast, the hydrogen peroxide and yeast reacted so vigorously that the started to rise up the delivery tube and into the gas syringe. Because of this I changed my trial experiment again, this time only using 0.5 grams of yeast, but still using 10 cm3 of hydrogen peroxide.

Mass of yeast – 0.5 grams

Volume of Hydrogen Peroxide – 10 cm3

Even though the hydrogen peroxide and yeast did not rise up the delivery tube, there were still problems with this trial experiment. The hydrogen peroxide and yeast were still reacting after 30 seconds, and therefore not all of the results were noted. However, I kept the time going, and the reaction stopped after about 35 seconds.

Using my trial experiment, I decided that for my real experiment, I will use 10 cm3 of hydrogen peroxide, 0.5g of yeast, and time for up to 40 seconds, with 5 second intervals.

I will use the temperature range 5-60ºC, as this will prove my hypothesis by showing that the rate of reaction is slow at lower temperatures, and slow at high temperatures, but around 35-40ºC it is quickest, because these are body temperatures. I will take results for 40 seconds, in intervals of 5 seconds. This is because in my trial experiment, the reaction was not complete after 30 seconds.

To make sure that this experiment is a fair test, the only thing that will be changed is the temperature. The factors that will be kept the same are; volume of hydrogen peroxide, mass of yeast, time between measurements, continuous shaking of the conical flask. The reason form this is because if one of these factors were changed, then the results from the experiment would be inaccurate. For example, if the mass of yeast is changed, then there would be a larger amount of oxygen created in a quicker time, and therefore the results would all be different. This is because of the nature of enzymes. The enzymes work on a lock and key theory, and if there is more yeast, there would be more enzymes, and therefore more hydrogen peroxide would be broken down, and more oxygen would be produced. If I took readings at different time intervals, then I would not be able to compare the results at the end of the experiment, and therefore would not be able to draw a graph.

To make sure that my results are accurate, my measurements have to also be accurate. The mass of yeast will be correct to 0.01g, because this is the accuracy of the scales that I will use to weigh out the yeast. The volume of hydrogen peroxide will b correct to 1cm3 because the markings on the side of the measuring cylinder go up in 1cm3. The time between each measurement will be correct to 1 second, because even though the stop clock is correct to 1 hundredth of a second, the clock is continuously running, so it is very difficult to be accurate. The volume of oxygen in the gas syringe will be correct to 1cm3 because this is the accuracy of the markings on the side of the syringe.

This method is a good way of carrying out this experiment because it allows me to easily record the rate of reaction between the yeast and the hydrogen peroxide by measuring the volume of oxygen released. This method is such that it is very easy to make repeats of the experiment, to show any anomalous results, and it is very easy to make the initial readings, because the gas syringe has markings on the side.

Results

The values in this table are shown, apart from the volume of oxygen collected in the gas syringe, however this is measured in cm3.

To calculate the rate of reaction I took the volume of oxygen produced after 15 seconds, and divided it by 15 to give the rate of reaction of oxygen in cm3/sec.

Conclusion

My results show that the rate of reaction between hydrogen peroxide and yeast increases as the temperature is increased, but then when the temperature exceeds 40ºC, the reaction slows down. The reaction is quickest at the beginning, and slows down gradually to a stop at around 30 seconds. The results that I obtained agreed mainly with my hypothesis. At 35ºC and 40ºC the reaction was faster than at higher and lower temperatures. However there is one temperature that does not agree with my hypothesis. This is 5ºC. As you can see from my graph and my table; the reaction at 5ºC was quicker than all the other reactions. Also, all of the temperatures except for 5ºC stop reacting when about 82 or 83cm3 of oxygen had been produced. However at 5ºC 97cm3 of oxygen was produced.

The reason why I obtained these results is because of what a catalase is used for in the body. Catalase is present in the peroxisomes of nearly all aerobic cells. It serves to protect the cell from the toxic effects of hydrogen peroxide by catalysing its decomposition into molecular oxygen and water without the production of free radicals. (An atom or a group of atoms with an unpaired electron. Radicals are unusually reactive and are capable of causing a wide range of biological damage). Therefore, when the yeast, the catalase, was added to the hydrogen peroxide, a reaction occurred to produce oxygen, and water. I measured the amount of oxygen produced to show the speed of the reaction. As yeast is a catalase, and a catalase would normally work in the body, the reaction occurred fastest at body temperatures, 35ºC and 40ºC. However at temperatures above 40ºC, the catalase began to denature, and therefore the reaction became slower.

Evaluation

This experiment was an effective way of meeting my aim, and proving my hypothesis. This is because the experiment allowed me to make multiple readings at the same temperature, and to easily change the temperature and take more readings. My results were also reliable, masses correct to 0.01g, volumes correct to 1cm3, and time correct to 1 second. The range of temperatures used was also large enough, allowing me to experiment with low temperatures, body temperatures, and high temperatures, and others in between, in 5ºC intervals. Therefore my results are reliable enough to make my conclusion.

Not all of my results fit in with my conclusion. The temperature where the results did not agree with my hypothesis is 5ºC. At 5ºC the reaction occurred very quickly, quicker than all the other temperatures. The anomalous result, at 5ºC, was cause because of dissociation. This process involves a liquid touching a very cold surface. This causes to molecules in that liquid to split into smaller atoms. So in this case the hydrogen peroxide touching the cold conical flask caused the hydrogen and oxygen to separate, and therefore there is more oxygen produced, as the yeast still reacts with the hydrogen peroxide.

To improve this experiment I would find a way to control the amount of shaking that occurs, because as I have shown, it is very difficult to control. Apart from this, I believe that my method was a good way of carrying out this experiment because it allowed me to collect readings at each temperature, every 5 seconds, and it was easy to control the variable, and the constants of the experiment, apart from the shaking of the conical flask.

To provide further proof of my hypothesis, I would repeat the experiment, but use a smaller range of temperatures, increasing in single ºC. By doing this is would find the exact temperature that the enzymes denature, and the reaction begins to slow down. Also I would like to find the highest temperature at which dissociation occurs.