Вы находитесь на странице: 1из 7

Enzyme Lab Report Robert Becerra Period 4 10/25/10 Introduction: Chemical reactions are necessary for sustentation of a living

cell, which must be carried out swiftly and efficiently. A cell cant depend on random phenomena to rely on these reactions. Therefore the cell creates enzymes to maneuver the process. An enzyme can be utilized to cause a particular molecule to split or to come together. Each enzyme must be specific to do work on a particular molecule. It must fit over the molecule in order to do this. The portion of the enzyme that fits over the molecule is called the active site. The substrate is the molecule that the active site is working with. When the enzyme and the substrate are reacting, they form a complex. Enzymes are catalysts; in which they allow the reactions to occur more quickly. They lower the activation energy that is needed to start the reaction. Enzymes are affected by their environments which need a specific set of conditions in order to function best. Raising the temperature of a substance will cause a rise in its kinetic energy allowing the heat to be expressed in motion. Most enzymes require a neutral pH; however, some will reach their greatest potential at a very acidic or basic pH. Concentration of the actual enzyme will also affect the way the enzyme works on the molecule. Coenzymes are chemicals required by a number of enzymes for proper functioning and those molecules that bond to the active site and compete with the substrate are called competitive inhibitors. Objectives: Observe the reaction of catalase and hydrogen peroxide Demonstrate the effects of extreme temperatures on catalase activity Learn how to establish a baseline for the amount of peroxide in a 1.5% solution Use titration techniques to determine the rate of hydrogen peroxide decomposition by enzyme catalysis Investigate spontaneous decomposition of hydrogen peroxide to oxygen and water

Hypothesis: If more time is added to the hydrogen peroxide and catalase solution per turn; then less hydrogen peroxide will be used as it is being absorbed by the catalase. Materials: A. Testing Enzyme Activity Hydrogen peroxide 1.5%, 30 ml Catalase working solution, 1 ml Boiled catalase working solution, 1 ml

1 Syringe, 10 ml 1 Pipet, 1 ml 1 Glass rod 1 Knife or Scalpel Potato or beef liver 2 plastic cups, 2 oz. 1 Beaker, 250 ml B. Establishing a Baseline Determining the Amount of Hydrogen Peroxide in a 1.5% solution Hydrogen Peroxide, 1.5%, 10 ml Catalase working solution, 1 ml Sulfuric acid, 1 M, 10 ml Potassium permanganate, 2% 1 Pipet, 1 ml 1 Syringe, 10 ml 1 Titration syringe 2 plastic cups, 2 oz. C. Rate of Hydrogen Peroxide Spontaneous Decomposition Hydrogen peroxide, 1.5%, 25 ml Catalase working solution, 1ml Sulfuric acid, 1 M, 10 ml Potassium permanganate, 2% 1 pipet, 1 ml 1 Syringe, 10 ml 1 Syringe, 5 ml 1 Titration syringe 3 plastic cups, 2 oz. D. Rate of Hydrogen Peroxide Decomposition by Enzyme Catalysis Hydrogen peroxide 1.5%, 60 ml Catalase working solution, 6 ml Sulfuric acid, 1 M, 60 ml Potassium permanganate, 2% 1 pipet, 1 ml 1 syringe, 10 ml 1 titration syringe 6 plastic cups 1 stopwatch 2 plastic cups, 2 oz. Procedure: A. Testing Enzyme Activity

1. 2. 3. 4.

Obtain a 10 ml syringe, remove tip, and label syringe H for hydrogen peroxide (H2O2) Using the syringe, add 10 ml of hydrogen peroxide to 2 oz. plastic cup. Using a pipet, add 1 ml of catalase solution to cup. Mix the contents by swirling and observe for 30-60 secs. Record observations in Table 1.

Effect of Extreme Temperature on Enzyme Activity 1. Using your syringe, dispense 10 ml of hydrogen peroxide solution into a 2 oz cup. 2. Your instructor has already prepared a sample of boiled catalase for the class. Add 1 ml of boiled catalase to your cup. 3. Mix contents by swirling and observe for 30-60 secs. Record any observations in Table 1. Presence of Catalase in Living Tissue 1. Use a knife or scalpel to carefully cur a 1 cm cube of potato or beef liver. 2. Macerate the potato or liver piece with a mortar and pestle. Place the tissue in the 50 ml beaker. 3. Add 10 ml of hydrogen peroxide to the beaker containing the macerated tissue. Observe any reaction that takes place. 4. Record your observations in Table 1. Suggest what might happen in the potato or liver was boiled before it was added to the hydrogen peroxide. B. Establishing a Baseline- Determining the Amount of Hydrogen Peroxide in a 1.5 % solution 1. Obtain two more 10 ml syringes, remove tips, and label one of the syringes S for sulfuric acid. Label the other syringe T for transfer. 2. Dispense 10 ml of hydrogen peroxide into a 2 oz. cup using the properly labeled syringe. 3. Add 1 ml of distilled water to the cup using a pipet. 4. Using the syringe labeled S, carefully add 10 ml of sulfuric acid to the cup. Mix the contents by gently swirling. 5. Using the syringe labeled T, transfer 10 ml of the mixture into a new 2 oz. plastic cup. 6. Fill a titration syringe to the 10 ml marking with potassium permanganate (KMnO4). Note the initial reading in Table 2 in the Analysis section of the lab. 7. Slowly add one drop of potassium permanganate and swirl the solution to mix. Continue to add potassium permanganate, on drop at a time and swirl after each addition, until the solution permanently turns pink or brown. The amount of KMnO4 added is proportional to the amount of H2O2 that was present in the solution. 8. Record the final volume in the titration syringe in Table 2. C. Rate of Hydrogen Peroxide Spontaneous Decomposition 1. Dispense 25 ml of hydrogen peroxide in a 2 oz. plastic cup. Let the beaker sit, uncovered, for 24 hours at room temperature. 2. After 24 hours, dispense 10 ml hydrogen peroxide into a new 2 oz. cup using the properly labeled syringe. 3. Add 1 ml of distilled water to the cup using a pipet.

4. Using the syringe labeled S, carefully add 10 ml of sulfuric acid to the cup. Mix the contents by gently swirling. 5. Using the syringe labeled T, transfer 10 ml of the mixture into a new 2 oz. plastic cup. 6. Fill a titration syringe to the 10 ml marking with potassium permanganate. Note the initial reading in Table 3 in the Analysis section of the lab. 7. Slowly add one drop of potassium permanganate and swirl the solution to mix. Continue to add potassium permanganate, one drop at a time and swirling after each addition, until the solution permanently turns pink or brown. The amount of KMnO4 added is proportional to the amount of H2O2 that was present in the solution. 8. Record the final volume in the titration syringe in Table 3. D. Rate of Hydrogen Peroxide Decomposition by Enzyme Catalysis 1. Dispense 10 ml of hydrogen peroxide in a 2 oz. plastic cup using the 10 ml syringe labeled H. 2. Using a pipet, add 1 ml of catalase solution and swirl gently for 10 secs. To mix. 3. Using the syringe labeled S, add 10 ml of sulfuric acid to stop the reaction. 4. Using the syringe labeled T, transfer 10 ml of the mixture into a new 2 oz. plastic cup. 5. Fill a titration syringe to the 10 ml marking with potassium permanganate. Note the initial reading in Table 4. 6. Slowly add one drop of potassium permanganate and swirl the solution to mix. Continue to add potassium permanganate, one drop at a time and swirling after each addition, until the solution permanently turns pink or brown. The amount of KMnO4 added is proportional to the amount of H2O2 that was present in the solution. 7. Record the final volume in the titration syringe in Table 4. 8. Repeat the procedure for 30, 60, 120, and 180 seconds. 9. Graph your results in the Analysis section of the lab. Plot the amount of hydrogen peroxide used on the y-axis and the time, in seconds, on the X-axis. Analysis: Table 1: Enzyme Activity Activity Enzyme Activity Effect of Extreme Temperature Presence of Catalase in Living Tissue Observations Substance fizzes with swirl Substance does not fizz with swirl Fizzes extremely

Table 2: Establishing a Baseline Titration Syringe Initial Reading Final Reading Baseline (Initial Final)

20.0 ml 3.1 ml 16.9 ml

Table 3: Hydrogen Peroxide Spontaneous Decomposition Using a Titration Syringe Initial Reading Final Reading Volume Used After 24 Hours Amount H2O2 Spontaneously Decomposed (baseline volume used after 24 hours) % H2O2 Spontaneously Decomposed (amount H2O2 Spontaneously decomposed / baseline) X 100 20.0 ml 9.7 ml 10.3 ml 6.6 ml 39%

Table 4: Hydrogen Peroxide Decomposition by Enzyme Catalysis Using a Titration Syringe Time (seconds) 10 30 16.9 ml 16.9 ml 20 ml 10 ml 4.2 ml 3.6 ml 15.8 ml 6.4 ml 1.1 ml 10.5 ml

Baseline Volume Initial Volume Final Volume Amount KMnO4 Used (initial final) Amount H2O2 Used (baseline - KMnO4 used)

60 16.9 ml 10 ml 6.0 ml 4.0 ml 12.9 ml

120 16.9 ml 10 ml 7.8 ml 2.2 ml 14.7 ml

180 16.9 ml -----------------------------

Hydrogen Peroxide Decomposititon


(Part D)
20 18 16 14 12 10 8 6 4 2 0 10 20 60 120 Time in Seconds

H2O2 Used

Conclusion: My hypothesis dealt with the portion of the lab, Part D. Part D had us create a solution with hydrogen peroxide and its enzyme, catalase. This chemical reaction was monitored over time with different intervals. Unfortunately however, our group had a hard time with controlling the reaction so we were forced to re-do the reaction several times and because of this we were unable to accomplish our last 180 second session with the reaction because of the loss of time. Using the rest of the data, I analyzed the remaining data and observed the change of time and its effect it had on the hydrogen peroxide decomposed. My hypothesis was proved wrong as time was added to the solution; more and more hydrogen was being used thanks to the catalase. One could see the effects of this by observing the data; with a mere 10 seconds, only 1.1 ml of hydrogen peroxide was used. As time went on, with 120 seconds, 14.7 ml of the hydrogen peroxide were used. Assessment Questions: 1. The function of enzymes is to lower the activation energy to start chemical reactions much faster. 2. Enzymes that work well in acidic environments would occur in the stomach. The stomach must have a very acidic stomach acid necessary to digest food. 3. The liver will break down an even amount of hydrogen peroxide in the body during any season because the temperature in the human body must be kept regulated. If it was hotter outside and that made it hotter inside the body, then the summer would help the liver break down more. 4. The smellulase enzyme would bind to the molecules that cause flatulence to become smelly and it would go away. 5. Insulin is an enzyme that allows diabetics to regulate blood sugar. A regular daily injection allows for proper blood sugar levels. 6. The Amylase activity at 25 C will not work as well at the ideal temperatures of 3540 C. Enzymes require a specific temperature to work properly. 7. Oxygen gas was escaping the solution in Part A. This happened because hydrogen peroxide loses O2 when it is exposed to heat. Your could use a test to see if you were correct. 8. In this lab, not keeping track of time was our biggest blunder. Not paying attention to the chemicals you use and how much can be costly as well. 9. The reaction of hydrogen peroxide and catalase would continue until there was no more hydrogen peroxide. 10. Catalase- is the enzyme that began the reaction in breaking down of hydrogen peroxide. Hydrogen peroxide- the solution that the enzyme acted upon breaking down. Sulfuric Acid- the solution that stops the reaction; it stopped the breakdown of hydrogen peroxide. Potassium permanganate- tested amount of H2O2 present in a solution

11. The baseline acted as a control group so that changes can be observed and recorded. 12. To determine if the amount varies hydrogen peroxide should be added to both subjects and tested periodically and compared. 13. It would probably denaturalize the catalase since an enzyme is a protein, and breaking down protein is the job of the proteases.