The Effect of pH on the Activity of Catalase

The Effect of pH on the Activity of Catalase
Secondary resources: from my research I carried out, I have learnt that: Catalyse is an enzyme that is found in foods such as potato and liver. Catalyse speeds up the decomposition of hydrogen peroxide into water because the shape of the hydrogen peroxide molecule. This type of where a molecule is broken down into smaller pieces is called an anabolic reaction. This enzyme catalysis is the breakdown of hydrogen peroxide and it forms oxygen gas and water. Enzymes are very large and complex organic molecules that are synthesized by the cell to perform very specific functions. These biological catalysts are important because they speed up the rate of the reaction they catalyze that would otherwise be too slow to support life.
Catalyse is an enzyme present in the cells of plants, animals and aerobic bacteria. It promotes the conversion of hydrogen peroxide, a powerful and potentially harmful oxidizing agent, to water and molecular oxygen. Catalase is located in a cell organelle called the peroxisome. Peroxisomes in animal cells are involved in the oxidation of fatty acids, and the synthesis of cholesterol and bile acids. Hydrogen peroxide is a by-product of fatty acid oxidation. White blood cells produce hydrogen peroxide to kill bacteria. In both cases catalase prevents the hydrogen peroxide from harming the cell itself.Peroxisomes in plant cells are involved in photorespiration and symbiotic nitrogen fixation (the breaking apart of the nitrogen molecule N2 to reactive nitrogen atoms). Hydrogen peroxide is produced as an intermediate during these chemical processes and must be removed to prevent damage to cellular machinery. Aerobic (oxygen requiring) bacteria produce hydrogen peroxide as a by-product of metabolism. This fact is used when identifying bacteria. If hydrogen peroxide is added to a bacterial colony and bubbles are produced, this is evidence of oxygen production and confirms that the colony is aerobic. The activity of catalase can be measured by finding the rate at which the oxygen gas is released from the decomposition of hydrogen peroxide. The buffer solutions that I am using are pH ? 4.4, 5.2, 6.5, 7.5 and 8.4. Hydrogen peroxide = 2H2O2 Hydrogen peroxide + catalase → oxygen + water The books and resources I used were: Energy and chemistry ? p229 Gareth Williams, biology for you pages 29-31 www.google.com- enzymes, catalase, hydrogen peroxide ocr staged assessment phase 1 module by Mary Jones Pre-test:
In the pre-test I done it by putting my pH and hydrogen peroxide
mixed solution in the syringe
Then put the potato in the conical flask
Put the bung on
Put water in the measuring cylinder and put it upside down without
the water coming out
I then put the delivery tube in the measuring cylinder
I got the stop watch
Started the time when I pushed the mixed solution in the conical
flask with the potato
Stopped the time when the gas was collected 10cm3
Recorded my result in a table
Repeat that experiment another 2 more time
Do the other pH solutions
Weight of the Potato (g) Volume of pH Trial 1 ? time taken for the gas to reach 10cm3 (s) Trial 2 Trial 3 Average 5 4.4 32.73 35.04 33.96 79.09 5 6.5 48.11 29.84 36.45 90.1 5 8.4 25.8 18.87 18.85 21.17 The second row in my table has got the anomalous result. It can?t have been 90.1 because that is too much to be the average set of results. This happened because I let some gas out as I was holding the cylinder, I didn?t realise the tube came out and then I thought why is it taking so long. In my pre-test I had some mistakes, they were firstly I didn?t use hydrogen peroxide with my pH, so I had to do the whole experiment again. The second mistake was, the gas I was measuring I wasn?t too sure if I was doing 20 or 15 so then I decided to do 10. [image] My graph that I have made from the results of my pre-test isn?t exactly right because looking at my results it doesn?t seem right. This happened because may be some of the gas escaped from the side of the cylinder, which is why it took time for my gas to be collected. Introduction: Enzymes are biological catalysts; this basically means that they speed up the chemical reactions in living things. Without enzymes, our guts would take at least weeks and weeks to digest out food, our muscles, nerves and bones would not work properly and so on, so we would not be living. A catalyst is any substance, which makes a chemical reaction go faster, without is self being changed. A catalyst can be used over and over again in a chemical reaction but I wont get used up. Enzymes are very much the same except that they are easily denatured by heat. Our enzymes work best at body temperature. Our enzymes also have to have the correct pH. All enzymes are made of protein, this is why they are sensitive to heat, pH and heavy metal ions. Unlike ordinary catalysts, they are specific to one chemical reaction, but an enzyme only works for one specific reaction. Enzymes must have the correct shape to do their job. They are made of proteins, and are very easily affected by heat, pH and heavy metal ions. Most people say an enzyme works like a key in a lock. If the key has been twisted by heat, or dissolved in acid or stuck up with chewing gum it will not work. Enzymes change their shape if the temperature of pH changes. Aim: The aim of my experiment is to see the effect of pH on the activity of catalase Apparatus: I have decided to use the following equipment in order to carry out my experiment:
1 Stopwatch
1 Beehive shelf
1 Trough
1 50ml Measuring cylinder
3 beakers 100mm
1 potato
Hydrogen peroxide (H2O2)
pH solutions
Conical flask with bung
Goggles
Measuring scale
1 20ml syringes
Delivery tube
Petri dish
Method:
1st I got all of my apparatus
Then grated my potato
Get the hydrogen peroxide in a beaker
Get the pH solution in a beaker
Measure both of them 10ml in the syringe and put them in the other
beaker
Get a trough with half filled with water
Put the measuring cylinder in the trough and fill it with water
and turn it up side down in the water
Put the delivery tube in the cylinder and hold the cylinder from
the top so that the cylinder doesn?t tip over
Measure the potato 5g in the Petri dish
Once I have measured the potato I put that in the conical flask
Get the mixed solution of hydrogen peroxide and pH solution they
should both add up to 20ml in the beaker
Make sure the other end of the tube is blocked with the syringe
Get the stopwatch
Put the solution of the hydrogen peroxide and pH in the flask and
put the bung on
As soon as you put the bung on start the watch
I timed it until the gas reached 10cm3
I recorded the time in a table
I repeated this 3 more times so I got a set of results
I then done it with the different types of pH
Fair test: To make it a fair test I will need to have not let any gas escape, if I do my test or time will take longer which won?t give me accurate results. I have to make sure that when I am measuring the pH solutions and the hydrogen peroxide I need to ensure that I am using the same amount for all of the experiments that I do for my actual experiment and the pre-test. I need to also make certain that the amount of potato that I am going to use, when I measure it, I need to use the same amount every time, which were 5g. I also need to make sure that when the oxygen has reached 10cm3, I need to stop the watch straight away and record the time. I am going to keep all of these things constant. I also require that I should need to check that I am using the same equipment through out the whole experiment. The only thing I am changing is going to be the buffer solutions, every time I do the 1 pH I made sure I repeated it a couple more time preferably 3 times until I got a set of results that are not more than 2 seconds more than my 1st result. If all of these factors are controlled it will make my test a fair one and also lead me to gain a reliable results. Safety: To ensure that I carry out a safe experiment, I will be careful when using the hydrogen peroxide because it tends to bleach clothes and damage the skin, I will also be careful with hydrogen peroxide by always closing the lid because the hydrogen peroxide can decompose if it is not bottled in. I will wear goggles so that my eyes are protected from the substrate. I will also wear protective footwear to protect my feet from any spills. Prediction: I predict that as soon as I pour the pH solution and hydrogen peroxide, I will have oxygen gas formed in my measuring cylinder. This happens because the potato and the solution react fast to give me my products. I also predict that the pH 4.4 would be less reactive than the other pH?s. I think this because as the pH value gets lower the reaction time will go faster and the rate of reaction will increase so when it gets lower from 7 to 6.5 and 4.4 the reaction time will increase. This is because the collision theory says that chemical reactions occur when particles of the reactants collide. They must collide with a certain minimum energy, called the activation energy. 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 type of biological catalyst. Those enzymes identified now number more than 700. 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 (micro body organelles that house various oxidation reactions in which toxic peroxides are generated as side products) 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). The lower the concentration the less gas particles, the higher the concentration the more gas particles collide. From my research I should see my graph the same as the one that I have from my results table. If I don?t, then there is probably something wrong with the pH or my results table wasn?t as accurate. Diagram of experiment: Observation-results: Potato (g) Volume of pH Trial 1 ? time taken for the gas to reach 10cm3 (s) Time taken for the gas to reach 10cm3 (s) Trial 2 Time taken for the gas to reach 10cm3 (s) Trial 3 Average 5 4.4 16.93 18.85 15.82 17.2 5 5.2 14.63 16.89 15.67 15.73 5 6.5 10.5 12.28 14.66 12.48 5 7.5 9.45 10.62 8.86 9.64 5 8.4 6.45 8.92 7.25 7.54 Conclusion: From my results I can see that the experiment I carried out and my prediction was correct because I predicted that pH 4.4 would be the less reactive one as it took more time for the oxygen to be collected 10cm3. So pH 7.5 was the optimum pH at which the enzymes works best. As the pH increases or decreases, the enzyme becomes less effective. The higher the enzyme concentration was, the more oxygen was produced, or in the case of my experiment, more bubbles were produced. This is because the higher the enzyme concentration, the more active sites there are where a collision between an enzyme and the substrate can take place. As you can see in the graph, the amount of bubbles increases as the amount of enzyme concentration decreases. This indicates that it was reaching the optimum state of catalyse activity. These set of results tell me that the best pH that effects the catalyse to collect the oxygen using the hydrogen peroxide is 8.4 because it has the less amount of time in seconds to collect the oxygen and it reacts really fast. [image]Graph: My graph looks good because the time is going from high to low. This is because the pH 4.4 is the least reactive, as I done my experiment with the catalyse. According to the graph, there are no visible outstanding results. However, if I had done the experiment more times and got a more precise average, then there may have been a few outstanding results. (By outstanding results I mean results that do not fit in with the other results pre-test graph, for example they may be too high). According to the graph the optimum pH, which is the most reactive, is 8.4 because it took the less time for it to react with the catalyse. Evaluation: The data that I collected was reasonable for the accuracy of my experiment, it was fairly reliable meaning that the results we collected were mostly expected but again with human error and contamination there were a few different results in my pre-test, which stood out quite a lot. A major cause for concern on accuracy was the reading of the overall measurement of the oxygen produced but also the measurements of hydrogen peroxide and the pH used for the experiment. So if these miss-readings were carried out through the whole investigation then my results will be quite a long way out. Looking back on experiment I believe that there are many ways to improve it. One way is the accuracy of the measuring of the potato this can be done by measuring accurately on the scale because it did have 2 more decimal numbers which could have been just five. Measuring the oxygen produced is a little harder, to measure this it may include equipment that is not available to me and so cannot be carried out. All that is best is that the same person reads it off just in case someone else?s vision is not as good. Also it was quiet harder because I wasn?t using a beehive shelf and the measuring cylinder was a bit tilted because obviously I had to see and reading and to collect the gas I needed the delivery tube to be there as well. My graph is very basic and one way to improve this will be to increase my range off tests. This will give me more results and so when I plot my graph again there will be more points producing a greater accuracy and a better curve with which I will be able to pull stronger conclusions from. I think the timing was as accurate and reliable as it could be, but only a second or third experiment would back that up fully. The equipment I used was reasonable. I think contamination was minor so that did not cause a problem. One problem with our first experiments was that I had to change the Hydrogen peroxide because I felt that it was old, and was probably not as concentrated. This could of seriously effected my main experiment with using one chemical but out of several bottles, to eliminate this I must get one bottle big enough for the whole experiment and use that one only; so that there is no change in the quality of the H2O2. My experiment has gone quite well, but I could do certain things in order to get more accurate results. Firstly, if I had time, I would certainly do the experiment more times, as this would give me a more precise average.

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