Biochemistry - Effect of Temp. on Catalase Activity free essay sample

Qualitative Observations:8 2. 2 Processing Data9 Statistical Processing Calculations9 2. 3 Presenting Processed Data10 Results Table10 3 Conclusion and Evaluation11 3. 1 Conclusion11 Conclusion Statement11 Conclusion Explanation11 3. 2 Evaluation Procedures:11 Reliability11 Errors/Limitations in Experimental procedure11 Significance12 3. 3 Improving the Investigations12 Suggestions for Improvements12 Appendix13 References13 Journal14 1 Design 1. 1 Defining the problem Research Question This lab will be driven by the research question; do changes in temperature (from 0? C, 7? C, 21? C, 37? C, and 90? C) have an effect on the activity of the enzyme catalase (found in beef liver) in the breakdown of hydrogen peroxide? Hypothesis If liver is placed in different temperatures of 0? C, 7? C, 21? C, 37? C, and 90? C and hydrogen peroxide is added to each piece of liver, then the liver placed in 37? C will have the largest reaction height. The liver in 90? C will have the smallest reaction height, followed by the liver in 0? C, then 7? C, and then 21? C. Background Information Pearson Baccalaureate: Standard Level Biology Developed Specifically for the IB Diploma defines enzymes as â€Å"protein molecules which act as catalysts for reactions. As catalysts, the real function of enzymes is to lower the activation energy of the reactions that they catalyze† (166). Enzymes are proteins; therefore the liver has a particularly high concentration of catalase. When hydrogen peroxide (H2O2) is added to liver, catalase catalyzes a reaction in which the hydrogen peroxide is broken down into oxygen gas (O2) and liquid water (H2O). Hydrogen peroxide is a toxic chemical that is produced as a byproduct of many normal cellular reactions, so it is crucial that catalase in the liver breaks the hydrogen peroxide down into the two harmless substances of oxygen gas and liquid water. The hydrogen peroxide must be quickly degraded or converted, and catalase accomplishes this task because one molecule of catalase can deal with six million molecules of hydrogen peroxide in one minute. Enzymes and the temperature of their environment are particularly important to the human body because â€Å"Many of the reactions which represent the digestive process would need far higher temperatures than we are able to maintain safely if enzymes were not involved† (166). The prediction in the hypothesis is based on the concept of denaturation. As Pearson Baccalaureate: Standard Level Biology Developed Specifically for the IB Diploma states, â€Å"Reactions which use enzymes do have an upper limit. That limit is based on the temperature at which the enzyme (as a protein) begins to lose its three-dimensional shape due to intramolecular bonds being stressed and broken. When an enzyme loses it shape, including the shape of the active site, it is said to be denatured† (75). Due to denaturation, the liver placed in 100? C will have the smallest reaction height because at this temperature catalase will begin to denature. Because â€Å"reactions with or without enzymes will increase their reaction rate as temperature (and thus molecular motion) increases†, the liver placed in 0? C, 7? C, and 21? C will have small reaction heights, but the heights will increase as temperature increases (75). The liver placed in 37? C will have the greatest reaction height because â€Å"human catalase works at an optimum temperature of 37? C, which is approximately the temperature of the human body†. Although this lab is using beef liver instead of a human liver, the optimum temperature for beef liver should be similar to that of humans. The predictive graph on the previous page represents how as the temperature of beef liver originally increases, the height of the reaction increases as well due to an increase in molecular collisions. At a certain temperature, the catalase will reach its optimum temperature and have the greatest height of reaction. However, as the temperature continues to increase the enzyme will begin to lose its shape and denature, so the height of the reaction will decrease. This lab will be using beef liver, which contains the specific enzyme catalase, and by placing the liver into different temperatures it will be assessed how catalase performs under certain conditions. After the liver has been placed in different temperatures, hydrogen peroxide will be added to each piece of liver, and by measuring the height of the chemical reaction it will be determined which temperatures catalase performs the best in. Reactions with a high height will represent catalase quickly and efficiently breaking down the hydrogen peroxide, and reactions with a low height will represent catalase slowly and inefficiently breaking down the hydrogen peroxide. This lab will serve as a model for the role of enzymes in the human body, and will outline the importance of enzymes for the human body. Investigation Variables Independent Variables The independent variable is the temperature of the liver, and it will be measured in ? C. The different temperatures used will be 0? C, 7? C, 21? C, 37? C, and 90? C. Dependent Variables The dependent variable is the height of the reaction, and the bubbles of gas produced will be measured in millimetres (mm). 1. 2 Controlling Variables Control Variables Table 1: Control variables and their treatments Variable | Effect | Control | Size of liver | Could increase or decrease the height of the reaction. A larger piece of liver means more catalase, which could break down the H2O2 at a more efficient rate than smaller pieces of liver. | Cut every piece of liver the same size by weighing each piece on an electric beam balance. Each piece of liver is approximately 1. 4g. pH | pH has an effect on enzymes and each enzyme has an optimal pH. By making the liver more acidic, basic, or neutral the height of the reaction could increase or decrease as it deviates from its optimal pH. | Do not change the pH of any of the solutions. This lab is only investigating the effect of temperature on enzyme activity, not the effect of pH on enzyme activity, so nothing should be added to the li ver that would increase or decrease the pH. | Time | If some pieces of liver are kept in their specific temperatures longer than others, the height of the reaction could be greatly affected. By keeping the liver in its temperature for a shorter period of time, the liver itself has less time to change temperature which could greatly alter the results. | Keep all of the pieces of liver in their specified temperatures for five minutes. | Size of test tube| Different sized test tubes will hold different volumes of O2 and will give incorrect measurements with the ruler| Keep all the test tubes the same size – with volume of 30cm3| Uncontrolled Variables The variables that will be difficult to control include air pressure in the lab and the volume of the liver. Measures will be taken to reduce the effect of these on the experiment by keeping air conditioning at 21? C and cutting the liver close to the same shape and size. Control Experiment The control experiment for this investigation will be the experimental setup at 0? C. All the steps in the method will be followed at this temperature. At 0? C the experiment should not proceed as the energy needed for enzyme action is unavailable. 1. 3 Experimental Method Materials and Equipment 10 Test Tubes 1 Package of Beef Liver 2 250mL Beakers 2 Test Tube Racks 1Forceps Hot Plate 1LIce 1 Plastic Bin 20 mlHydrogen Peroxide 1 100 mL Beaker 1 10mL Graduated Cylinder 5 Thermometers 1 Sharpie 1 Electronic Beam Balance 1 Knife Method 1. Prepare an ice bath by placing ice into a container, and place a thermometer into the ice bath. Wait until the temperature has reached 0? C. 2. Prepare two more water baths like this at 7? C and 21? C. (You can replace these two temperatures with that of the fridge and ro om) 3. Prepare a hot water bath by placing a 250mL beaker filled with water on a hot plate, and place a thermometer into the water. Wait until the temperature has reached 90? C. You can use and electric water bath if one is available. 4. Prepare a warm water bath by placing a 250mL beaker filled with water on a hot plate, and place a thermometer into the water. Wait until the temperature has reached 37? C. You can use and electric water bath if one is available. 5. Obtain beef liver and cut the liver into 10 slices which are approximately the same size. Weigh each slice on an electronic beam balance to ensure they are the same weight. 6. Obtain 10 test tubes and place them into a test tube rack. Label 5 test tubes with a number from 1-5, and repeat for the remaining five test tubes. 7. Obtain 15mL of hydrogen peroxide and a graduated cylinder. 8. With forceps, place a piece of liver into each of the test tubes. 9. After the liver has been placed in the test tubes labelled 3, place a thermometer into the test tube rack and wait for 5 minutes. After 5 minutes, pour 2mL of hydrogen peroxide into each of the test tubes, observe the reaction, and label the height of the reaction after 1 minute with a Sharpie. (This is the experiment at room temperature. If you have prepared a water bath at 21? C then use it instead). 10. Remove both of the test tubes labelled 1 from the test tube rack, place them in the ice bath, and wait for 5 minutes. After 5 minutes, remove the test tubes from the ice bath, add 2mL of hydrogen peroxide into each of the test tubes, observe the reaction, and label the height of the reaction after 1 minute. 11. Remove both of the test tubes labelled 2 from the test tube rack, place them in a different test tube rack and place this rack into the fridge. Place a thermometer into the fridge as well. Wait for 5 minutes, and then read the temperature of the fridge and remove the test tubes. Pour 2mL of hydrogen peroxide into each of the test tubes, observe the reaction, and label the height of the reaction after 1 minute. (If you have prepared a water bath at 7? C then use it instead). 12. Remove both of the test tubes labelled 4 from the test tube rack, place them in the warm water bath (37? C), and wait for 5 minutes. After 5 minutes, remove the test tubes from the warm water bath, add 2mL of hydrogen peroxide into each of the test tubes, observe the reaction, and label the height of the reaction after 1 minute. 3. Remove both of the test tubes labelled 5 from the test tube rack, place them in the hot water bath (90? C), and wait for 5 minutes. After 5 minutes, remove the test tubes from the hot water bath, add 2mL of hydrogen peroxide into each of the test tubes, observe the reaction, and label the height of the reaction after 1 minute. 14. With a ruler, measure from the bottom of the test tube to the mark which labels the maximum height of the reaction after 1 minute. Repeat this for all 10 test tubes, and record the measurements in your data table. 15. Pour the liver from each test tube into a waste beaker, clean each of the test tubes out, and put all materials away. 16. Combine the data that your own group obtained with the data from two other groups. This will allow for six trials worth of data, and once you obtain this sufficient amount of data, calculate the average height of the reaction for each of the five temperatures over the six trials, and then calculate the standard deviation for each of the five temperatures over the six trials as well. Variable | Unit of Precision | Error/Uncertainty | Temperature | ? C | +/- 0. 5? C | Height | mm | +/- 0. 5mm | In the procedure, the maximum height of the reaction will be determined by marking the position of the tallest bubble that results from the reaction. Practical Safety and Risk Assessment All care will be taken when dealing with hot plate and hot water to prevent burning and scalding. The knife and glassware will be used carefully and hand gloves will be worn when handling hydrogen peroxide. Note the following hazards with hydrogen peroxide: Contact with eyes can cause serious long term damage. The solution is corrosive and can cause skin burns. For eye contact, immediately flush eyes with plenty of water. For skin contact, wash off with plenty of water. Setup Diagram Liver | 2 ml of Hydrogen peroxide | Liver | 2 ml of Hydrogen peroxide | 2 Data collection and processing 2. 1 Recording Raw Data Sufficiency of Data This lab will be investigating the effect of temperature on the activity of the enzyme catalase. In this lab, five different temperatures will be investigated (0? C, 7? C, 21? C, 37? C, 90? C). Each temperature will have six trials, and this will ensure the reliability of the data. With the sufficient data, the standard deviation and mean will be calculated for each temperature. The rate of reaction at the different temperatures will also be calculated. Results – Raw Data Table 2: height of reaction of hydrogen peroxide on liquid beef liver in different temperatures | Height of Reaction ( ±0. 5mm)| Temperature ( ±0. 5C °) | Trial 1| Trial 2| Trial 3| Trial 4| Trial 5| Trial 6| 0. 0| 115. 0| 123. 0| 134. 0| 132. 0| 102. 0| 108. 0| 7. | 125. 0| 134. 0| 128. 0| 118. 0| 96. 0| 90. 0| 21. 0| 127. 0| 124. 0| 134. 0| 129. 0| 110. 0| 114. 0| 37. 0| 147. 0| 139. 0| 149. 0| 137. 0| 128. 0| 106. 0| 90. 0| 0. 0| 0. 0| 0. 0| 0. 0| 0. 0| 0. 0| Note: The highlighted data is the data obtained by me. The rest of the data has been collected from Brendon Song and Skye Rodgers. Qualitative Observations: In this photo, both test tubes were put into the freezer and kept at a tempera ture of 0 degrees. 2mL of hydrogen peroxide were added into the liquid beef liver and the bubbles were formed. However, it is evident that there is a significant height difference between the two trials and the test tube on the right has a large gap at the bottom. Also, the left test tube’s bubbles seem to be much more concentrated than the right tube, this could cause inaccuracy in the data due to the gap increased the height of reaction but the amount of bubbles produced wasn’t actually significantly higher. This test tube was placed in 90 degrees hot water for five minutes, and the colour of the liquid beef liver has changed from a dark pink colour to a brownish colour as shown in the picture. This can possibly suggest that the catalase in beef liver was denatured in the high temperature which resulted in no reaction at all when hydrogen peroxide was added in. 2. 2 Processing Data Statistical Processing Calculations Table 3: Statistical Processing – The following sample calculations will be done for the raw data Statistical Analysis| Formulae| Meaning of Symbols| Sample Calculation| Mean| | The mean or averagesum of sample measurements number of samples | = 115+113+134+132+102+108 6 ? 17. 3| Standard Deviation| | S = standard deviationx = each individual valueX = mean of all measurements= deviation from mean = degrees of freedom| Calculated in Microsoft Excel| | Height of Reaction ( ±0. 5mm)| | Temperature( ±0. 5 °C) | Trial 1 | Trial 2 | Trial 3 | Trial 4 | Trial 5 | Trial 6 | Average Height of Reaction | Standard Deviation (mm +/- 0. 5mm) | 0. 0| 115. 0| 113. 0| 134. 0| 132. 0| 102. 0| 108. 0| 117. 3| 12. 9| 7. 0| 125. 0| 134. 0| 128. 0| 118. 0| 96. 0| 90. 0| 115. 1| 18. 0| 21. 0| 127. 0| 124. | 134. 0| 129. 0| 110. 0| 114. 0| 123. 0| 9. 2| 37. 0| 147. 0| 139. 0| 149. 0| 137. 0| 128. 0| 106. 0| 134. 3| 15. 8| 90. 0| 0. 0| 0. 0| 0. 0| 0. 0| 0. 0| 0. 0| 0. 0| 0. 0| Table 4: Average height of reaction and standard deviation of hydrogen peroxide on liquid beef liver in different temperatures 2. 3 Presenting Processed Data Results Table Graph 1: Temperature vs. average height of reaction 3 Conclusion and Evaluation 3. 1 Conclusion Conclusion Statement The results obtained from this lab support my hypothesis. My hypothesis was: If liver is placed in different temperatures of 0? C, 7? C, 21? C, 37? C, and 90? C and hydrogen peroxide is added to each piece of liver, then the liver placed in 37? C will have the largest reaction height. The liver in 90? C will have the smallest reaction height, followed by the liver in 0? C, then 7? C, and then 21? C. By viewing the average height of the reaction, these results are supported. The liver placed in 37 °C had an average reaction height of 134. 3mm, 0? C has a height of 117. 3mm, 7? C has a height of 115. mm, 21? C has a height of 123. 0mm and 90? Chas a height of 0. 0mm. This data coincides with the predictions stated in the hypothesis, where I stated â€Å"The liver in 90? C will have the smallest reaction height. † By observing the constructed graph in Introduction, it is clear that the average height of the reaction initially increased as the temperature increased, but when temperatures increased past optimum (37 °C), the rate of re action starts decreasing until the temperature reaches to 90 °C and the reaction completely stopped. Graph 1 constructed from the obtained data shows that rate of reaction continuously to rise to 37 degrees and starts dropping until 90 degrees. These results outline the effect of temperature on enzyme activity, and it is apparent that changes in temperature do have an effect on the enzyme catalase. Conclusion Explanation The results obtained give increased insight into the role of enzymes in the human body. The beef liver closely modelled the human liver, and it is clear that catalase works best at an optimum temperature of 37 degrees. This is important because humans maintain a stable body temperature of 36 to 37 degrees, and with the aid of enzymes this temperature provides enough activation energy for metabolic reactions, in this case the breakdown of hydrogen peroxide into oxygen gas and liquid water. The liquid liver catalase didn’t react well in 100 degrees temperature because the high temperature that the liver was exposed to cause the protein enzyme to become denatured resulting in a loss of 3-D structure and hence stop enzyme activity when hydrogen peroxide it added in. 3. 2 Evaluation Procedures: Reliability While the results obtained from this lab clearly support my hypothesis, the examination of the calculated standard deviation for each temperature is 12. 9mm, 18. 0mm, 9. 2mm, 15. 8mm and 0. 0mm. This range in standard deviation is due to errors that may have occurred throughout the lab. The liver placed in 0? C had a standard deviation of 12. 9mm, the liver placed in 12. 9mm for 0 degrees, 18. 0mm for 7 degrees. 9. 2mm for 21 degrees, 15. 8mm for 37 degree and 0. 0mm for 90 degrees. The larger the standard deviation, the less reliable the data is. The error bars are the graphical representation of the variability of data it either shows range of data or standard deviation. The smaller the errors bars are the more reliable the data is. Errors bars can vary greatly depending on the number of trials in an experiment, the more trials means higher accuracy hence smaller error bars. Errors/Limitations in Experimental procedure The first limitation in this experiment was that the temperature wasn’t very accurate and stable throughout the conduct of this lab due to the fridge and freezer’s door was frequently opened by other students who were doing the same experiment. The second limitation is that the temperature could not be kept at the same degree, the temperature will rise once the test tubes are taken out of the freezer/fridge this can cause some inaccuracy when recording the data. Another limitation which may cause the unreliability of the data is that the hydrogen peroxide acid couldn’t be added to the test tubes at the same time, this could cause a small temperature difference between the two trials which may result in slightly different height of reaction. The forth limitation in this lab is the measurement of the height of reaction weren’t recorded at the exact same time due to it takes time to measure each sample’s reaction height so some bubbles may evaporate during this process and therefore decrease the height of reaction. The last limitation is that the measurement of the amount of hydrogen peroxide added into the liquid liver couldn’t be measured at exactly 2mL because of human errors when using measuring cylinder to obtain the acid, this can result in slight differences between each trial. Significance The five limitations stated above were mainly temperature errors and measuring errors. All these limitations have a certain effect the accuracy of the data; however, the uncertainties of each limitation were very small to have significant effect on the final result. The temperature difference for the first limitation stated above could be as small as 1 degree which won’t make any large difference for the data. The measuring limitations also nly have a small effect on overall result because six trials’ of height of reaction for each temperature was recorded so the measuring inaccuracy can be ignored. Overall, the uncertainties for this experiment are fairly low therefore the results obtained are quite reliable for school study purpose. 3. 3 Improving the Investigations Suggestions for Improvements For the first limitation stated above, the inaccuracy was caused by frequently opening the fridge’s doors so the temperature can’t stay at same which resulted in some differences when recording the data. This limitation could be avoided if all the students place their test tubes in the fridge at the same time and leave the fridge closed for a full five minutes before opening again. By doing this, the temperature of the liquid liver catalase can be assured to be at the designated degrees after 5 minutes and the data collected will be more accurate. The second limitation can be avoided only if the hydrogen peroxide acid was added in right after the samples get taken out from the fridge. This can make sure the temperature is remained at the same degrees when the solute is added in hence the data will be a bit more accurate. However, this improvement is hard to carry out because it takes some time to walk back to the working station and add in the hydrogen peroxide, during this process, the temperature may increase due to room temperature is higher. The third limitation could possibly be avoided if two measuring cylinders were available for each student to use and the conductor of this lab use both hands to put the hydrogen peroxide acid in at the exact same time. The results’ accuracy will improve by doing this but it won’t make any big difference in the final result. The forth limitation can be avoided if another student helps to measure the second test tube while the conductor of this experiment measure another one at the same time so that the maximum height of reaction is accurately recorded without any delay. The last limitation can be avoided using a pipette when obtaining the hydrogen peroxide acid, the uncertainty will be greatly reduced by doing this and thereby making the results more reliable. Appendix References Damon, Alan, Randy McGonegal, Patricia Tosto, and William Ward. Pearson Baccalaureate: Standard Level Biology for the IB Diploma (Pearson International Baccalaureate Diploma: International Editions). n/a: Imprint Unknown, 2008. Print. â€Å"Effect of Catalase on Hydrogen Peroxide. † http://www. sciencegeek. net/Biology/biopdfs/Lab_Catalase. pdf. (25 Jan. 2013). Journal Name: Jerry Li Research Journal Project title: Investigation of a Factor Affecting Enzyme Action Total hours research: 2 hours Date| Research/Activity/Action/Outcome| Time Used| 1/02/2013| Task sheet and instructions were given in class today | 30 mins| 09/02/2013| Researched factors that affect enzyme activities and wrote down some important notes for the experiment| 90 mins| 13/02/2013| The experiment was done in class today, raw data were collected and put into the raw data table| 60 mins| 18/02/2013| Data from other students were collected and put into the table of raw data| 30 mins| 25/02/2013| C ompleted writing Part 2 (Data collecting and Processing) of the IA| 90 mins| 02/03/2013| Full lab report completed and fixed up a few errors| 120 mins |