Effect of substrate pH on yeast catalase and hydrogen peroxide reaction...need h

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Useful info The Enzyme Catalase is a protein molecule which is found in living cells. It is used to speed up reactions in the cells. It is a very specific enzyme and just performs one particular reaction. Catalase is an enzyme found in cells in potatoes and liver and is used for removing Hydrogen Peroxide from the cells. Hydrogen Peroxide is the poison produced during metabolism. Catalase aids the decomposition of Hydrogen Peroxide the by products of which are water and oxygen as shown in the equations below. Formula: Catalase Hydrogen Peroxide-------------->Water + Oxygen Catalase 2H2O2-------------->2H2O+O2 It is able to speed up the decomposition of Hydrogen Peroxide because the shape of it's active site is the same as that of the Hydrogen Peroxide molecule. This is the type of reaction where a molecule is broken down into smaller pieces and is called an anabolic reaction. Theory Enzymes are catalysts which means they speed up reactions and allow them to take place more easily using less energy. They do this because they each have a specialized active site which is designed to fit a specific molecule. (Pepsin= Protease which catalyses reactions involving proteins) etc Every type of enzyme has a different shaped active site and likes to work in different conditions some like more neutral conditions others like acidic conditions the conditions vary from enzyme to enzyme. The Apparatus I will Need For the tests. Gas Syringe Metal Stand Yeast Catalase Hydrogen Peroxide Test Tubes Beakers Test Tube Rack Stop Watch Pipette Pipette Filler Tap Water Plan 1. Put 2cm3 of yeast into one test tube. And 4cm3 of hydrogen peroxide solution at a concentration of 20% to the other test tube. Use a pipette to measure the volumes. It is very important to measure the amounts of Hydrogen Peroxide , Yeast and water accurately , to ensure it is a fair test. 2. Pour the hydrogen peroxide into the test tube that contains the yeast and immediately put the gas syringe bung on the end of the test tube immediately , at the same time start the stopwatch. 3. Bubbles should start to rise up the tube and the gas syringe will move outwards, when the volume of gas inside the gas syringe reaches the 30cm3 mark stop the clock and note the time to the nearest 1/10th of a second. 4. Repeat the experiment with hydrogen peroxide concentrations of 16%, 12%, 10%, 8% and 4%. The table below shows what amounts of Hydrogen Peroxide and water are needed to make the solutions. Concentration Of Hydrogen Peroxide Volume Of Hydrogen Peroxide (cm3) Volume Of Water (cm3) 20% 4 0 16% 3.2 0.8 12% 2.4 1.6 10% 2 2 8% 1.6 2.4 5. Repeat the tests at least three times so that you can work out an average. Repeating the experiments several times will produce better , more accurate results as inaccuracies in one experiment should be made up for by the other experiments. I am going to use yeast catalase as opposed to catalase from apples, potatoes or liver . To ensure this is a fair test all the variables except for the concentration of the Hydrogen Peroxide must be kept the same for all of the experiments. Variables that must not be altered include:- The temperature of yeast, the temperature of Hydrogen peroxide, the yeast concentration, the batch of yeast, the volume of yeast and the volume of hydrogen peroxide. When measuring out the volumes of Hydrogen Peroxide, Yeast and Water the measurement should be taken by looking at the scale at an angle of 90 degrees to it to avoid any errors while measuring . My Prediction I predict that as the substrate concentration increases, the rate of reaction will increase at a rate that is directly proportional to the rate to the increase in the substrate concentration until the solution is saturated with hydrogen peroxide. When this point is reached, adding extra substrate will make no difference to the rate of the reaction. Once the number of substrate molecules added is more than the number of active sites which are unfilled then the rate of reaction will no longer go up. This is because the maximum number of reactions are taking place at once so any more substrate molecules have to wait until some active sites are no longer in use. Results When I carried out the experiment I got these results. Hydrogen Peroxide Concentration 0% 4% 8% 10% 12% 16% 20% Time Taken (Test 1) (secs) 47.3 18.4 17.3 14.5 10.6 9.7 Time Taken (Test 2) (secs) 43.3 19 16.7 14.9 11.2 10 Time Taken (Test 3) (secs) 52.2 17.2 18.5 11.2 8.6 7.8 Average of the Tests (secs) 47.6 18.2 17.5 13.5 10.1 9.2 Rate=30/Average (Cm3/second) 0 0.63 1.65 1.71 2.22 2.97 3.26 The average results are written down to one decimal place because it is impossible to get accurate times to two decimal places , because our reaction times are not fast enough to stop the clock that accurately . From these results I was able to plot a graph of the rate of reaction against concentration of Hydrogen Peroxide. Graph To show the volume of oxygen produced per second. Volume Of Oxygen Produced per second CM3 [IMAGE] Substrate concentration Analysis of results When the Hydrogen Peroxide concentration is increased, the rate of reaction increases at a directly proportional rate until the concentration of Hydrogen Peroxide reaches about 16%. When the concentration doubled from 8-16% the rate of reaction went up from 1.65-2.97 Cm3 Oxygen produced per second, which is an increase of 1.8 . I expected the rate of reaction to increase twice when the Hydrogen Peroxide concentration was increased twice because there were twice as many substrate molecules which could join onto the active sites of the enzymes . The reason that the number is less than double could be because at 16% the Enzyme's active sites may already have been close to being saturated with Hydrogen Peroxide. There may also have been an experimental error which caused inaccuracies. After 16% the increase in the rate of reaction slowed down. This is shown by the curve of the graph starting to level out . At this point almost all the active sites were being used so the active sites were said to be saturated with Hydrogen Peroxide. Increasing the Hydrogen Peroxide Concentration after the point of saturation had been reached would not have caused the rate of reaction to go any faster . All the active sites were being used so any more Hydrogen Peroxide molecules would have had to wait until an active site became vacant. The optimum rate of reaction ,theoretically, is when all the active sites are being used but in reality this is never reached because not all the active sites are being used at the time. Factors that Limited the experiments accuracy To help make my experiment more accurate, I repeated each test three times and then averaged all the results to plot the graph. I tried to keep the variables except the concentration of Hydrogen Peroxide the same for all of the experiments. There are a few factors which may have made the experiment less than perfectly accurate such as : The miniscule delay between mixing the Hydrogen Peroxide and the yeast , putting the bung on top of the beaker and starting the stopwatch. This will effect all the results but I carried out all the three steps in the same way every time so it should not make a noticeable difference to my results. Anomalous results The plotted points on the graph give a straight line of best fit at beginning which then curves off steadily as the reaction slows . The o anomalous results are the ones at 8% and 10%. The one at 8% is slightly above the line of best fit and the 10% result is slightly below it. This is probably due to an error involving the factor I mentioned.

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