Period5.C2


 * **Independent Variable ** || We will be manipulating the pH ||
 * **Dependent Variable(s) ** || The height of the bubbles created in the reaction, and the change in pressure ||
 * **Controlled Variables ** || Temperature, amount of liver, amount of either acidic or base solution, time that solution is allowed to mix with liver, time that hydrogen peroxide is allowed to react with enzymes ||
 * **How will you change your independent variable? (be specific) ** || First we will test the reaction of the enzymes without any addition of base or acid, and then add either a solution of a base or an acid to compare the reactions of the enzymes with the hydrogen peroxide ||
 * **How will you measure your dependent variable? ** || We will take quantitative measurements of the changing height in the bubbles after the reactions. Also we will measure the change in pressure after the reaction in the beaker. ||
 * **How will you set up a control? ** || Just make sure the temperature is stable, and nothing contaminates the liver besides the acid and base solutions. Also the amount of solutions that are added will be equal in volume and concentration. ||
 * **Hypothesis: ** || The reaction will be less efficient between the enzymes and the hydrogen peroxide if the pH of the enzymes is changed from its original state. ||

**Procedure for Enzymatic Activity Lab ** Jacqueline Disch, Kaelynn Maloney, <span style="font-family: 'Times New Roman','serif'; font-size: 12pt; msoansilanguage: EN-US; msobidilanguage: AR-SA; msofareastfontfamily: 'Times New Roman'; msofareastlanguage: EN-US;">Ian Randall


 * <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-size: 12.0pt; mso-hansi-theme-font: major-latin;">Introduction: **

<span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-size: 12.0pt; mso-hansi-theme-font: major-latin;">Throughout the different roles in life on earth, enzymes have always been most important to sustenance of life. Enzymes are mostly protein based molecules that act as catalysts. A catalyst is a chemical compound or substance that initiates or accelerates a chemical reaction without itself being consumed or affected during the reaction. This process is more formally known as catalysis. Catalytic reaction has a lower rate-limiting [|free energy change] to the transition state than the corresponding uncatalyzed reaction, resulting in a larger reaction rate at the same temperature, meaning an enzyme catalyzes reactions by lowering the activation energy necessary for a reaction to occur. The molecule that the enzyme acts on is called the substrate, and <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: Verdana; mso-bidi-font-size: 12.0pt; mso-hansi-theme-font: major-latin;">each enzyme is specific for the reaction it will catalyze. Naturally enzymes are highly specific towards the substrate, the temperature, and the pH levels for which it must react with. <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-size: 12.0pt; mso-hansi-theme-font: major-latin;">In this lab we studied the enzyme Catalase, produced by the liver.

<span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: Helvetica; mso-bidi-font-size: 12.0pt; mso-bidi-font-weight: bold; mso-hansi-theme-font: major-latin;">Catalase <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: Helvetica; mso-bidi-font-size: 12.0pt; mso-hansi-theme-font: major-latin;"> is a common enzyme found in nearly all living organisms which are exposed to oxygen, where it functions to catalyze the decomposition of hydrogen peroxide to water and oxygen. This is the chemical formula of the reaction. <span style="display: block; font-family: 'Calibri','sans-serif'; line-height: 115%; msoasciithemefont: major-latin; msobidifontfamily: Verdana; msobidifontsize: 12.0pt; msohansithemefont: major-latin; text-align: center;">H2O2 -> H2O + O2 (Catalase) <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: Verdana; mso-bidi-font-size: 12.0pt; mso-hansi-theme-font: major-latin;">As the experiment in this lab, we tested the functionality of Catalase in environments with different pH levels. We will do this by adding hydrochloric acid (HCL), a naturally acidic substance with a pH level of 2, and we will add separately sodium hydroxide (NaOH) a naturally basic substance that can have ph level ranging from 12- 14. Lastly we will be testing it with water, which has a pH level of 7 as the control. This is because water exactly in the middle between the acidity of hydrochloric acid and the baseness of sodium hydroxide.

Materials: 3 200mL graduated cylinders (to put three liver samples in) 3 cuts of liver (approximately 1" by 1") 5 mL of 0.1M HCl 5 mL of 0.1 M NaOH 1 micropipette 1 gas pressure sensor to hook to the computer 30 mL of hydrogen peroxide (10 mL for each liver sample) 1 computer (to hook gas pressure sensor to) Tongs to handle liver 3 pairs of goggles 3 pairs of rubber gloves

Procedure: 1. Gather materials 2. Put on rubber gloves and goggles 3. Using a knife cut three pieces of liver into 1 inch by 1 inch pieces 4. Connect the pressure sensor to the computer 5. Using tongs put one of the pieces of liver into a 200 mL beaker 6. Place pressure sensor in the same beaker with the piece of liver 7. Measure out 10mL of hydrogen peroxide using a graduated cylinder 8. Using a pipette add 10mL of hydrogen peroxide to the beaker 9. Allow computer to record the pressure results 10. Measure the maximum height of the bubbles in the reaction with a ruler and record results 11. Using tongs put the second piece of liver in the second beaker 12. Measure 5mL of O.1M HCI using a graduated cylinder 13. Using a pipette add 5mL of 0.1M HCI to the beaker directly on top of the piece of liver 14. Repeat steps 7-9 15. Using tongs put the third piece of liver into the third beaker 16. Measure 5mL of 0.1mL NaOH using a graduated cylinder 17. Using a pipette add 5mL of 0.1M NaOH to the beaker directly on top of the piece of liver 18. Repeat steps 7-9 19. Observe results and graph of the pressure changes with different pH levels on the computer

Data: <span style="font-family: 'Times New Roman','serif'; font-size: 12pt; mso-ansi-language: EN-US; mso-bidi-language: AR-SA; mso-fareast-font-family: 'Times New Roman'; mso-fareast-language: EN-US;">

<span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-fareast-font-family: 'Times New Roman'; mso-hansi-theme-font: major-latin;"> 1.) What do your results tell you about how your variable affected catalase activity? Include specific data from your experiment to help you answer this question
 * <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-fareast-font-family: 'Times New Roman'; mso-hansi-theme-font: major-latin;">Liver Lab Analysis Questions: **

The results have shown that when the liver becomes more basic or acidic, catalase activity doesn’t occur because the enzymes are deactivated. Therefore the catalase enzymes do not break down the hydrogen peroxide. Since the hydrogen peroxide isn’t broken down, no bubbles are created form the reaction and the pressure and the height of the bubbles remains constant. This is demonstrated through are more basic and more acidic pieces of liver. The piece of liver that had base, NaOH, stayed around 99 kPa, while the piece of liver that had become more acidic with HCI stayed around 101 kPa.

2.) What were some sources of error and how might they have affected your data? What changes would you make to your procedure –or—what changes did your peer editor suggest that you make?

One source of error could have been that the stopper wasn’t completely secure on the top of the flask, letting of some of the pressure. This could have cause a bit of difference between the pressures in the lab and the pressures we should have gotten in our results. Also any of the measurements of HCI, NaOH, or hydrogen peroxide could have been, creating a greater increase or decrease in the reaction of catalase and hydrogen peroxide, manipulating the pressure. Also before the experiment, after peer editing a couple of sources of error in the procedure were corrected before the actual lab. One, in the original lab procedure, the liver was to be places in a beaker with its corresponding ingredients, but we changed it to a flask so we could better record the pressure. Next, we included adding distilled water to the control. Also the original called for 5 mL of acid, base or distilled water but was altered to 10 mL, because 5ml wouldn’t of cause a big enough reaction to clearly observe the results.

3.) Catalase is located in a cell organelle called the peroxisome. What do you think is the function of the peroxisome? Explain your reasoning.

Catalase is an enzyme that breaks down hydrogen peroxide, into oxygen and water. Based on its function, the peroxisome must deal with breaking down and regulating other chemicals, in addition to hydrogen peroxide, in the body. The high hydrogen perioxide levels damage serotonergic terminals, destroying neurons and forever effecting part of your brian. This effects your short term memory and verbal skills. This is expecailly dangerous at a party because loss of short term memory and trouble communicating could lead to some bad desisions.

4.) Researchers studying the effects of ecstasy (methylenedioxymethamphetamine or MDMA) on mice found that ecstasy decreased catalase activity in the liver. They also noticed that ecstasy’s effects on catalase were increased as the temperature of the mouse’s environment increased. Based on this information, can you draw any conclusions about what might be going on inside a person’s body when he/she takes ecstasy at a party? What risks are involved?

Catalase is responsible to taking hydrogen peroxide and turning it into water and oxygen. If a person was to take ecstasy, which slows catalase activity, the concentration of hydrogen peroxide would build up. <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: Calibri; mso-bidi-font-size: 12.0pt; mso-bidi-theme-font: major-latin; mso-fareast-font-family: Calibri; mso-fareast-theme-font: major-latin; mso-hansi-theme-font: major-latin; msoasciithemefont: major-latin; msobidifontfamily: Calibri; msobidifontsize: 12.0pt; msobidithemefont: major-latin; msofareastfontfamily: Calibri; msofareastthemefont: major-latin; msohansithemefont: major-latin; msolist: Ignore;"> 1. <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-fareast-font-family: 'Times New Roman'; mso-hansi-theme-font: major-latin;">5.) Catalase is used commercially whenever hydrogen peroxide is used as a germicide. For example, it breaks down the H2O2 that was used to pasteurize milk prior to cheese-making (Chu //et al//. 1975).

a. As a cheese-maker, what conditions would you want to have in your cheese factory to ensure that catalase was working best? (use the class findings to help you answer this question) As a cheese maker you would have to have to perfect environment for the enzymes to work, so they aren’t deactivated. According to our results, the pH of the cheese would have to be at 7, the middle of the pH scale, to work the best. Being too acidic or basic would harm or even cease catalase activity. According to other groups in the class, temperature would have to have temperature be right as well.

b.) why would you need to make sure that the temperature of the catalase never went above a certain temperature? You would need to make sure the temperature of the catalase never went to high or sales the catalase enzymes would be deactivated. Without it there would be too high of a hydrogen peroxide lever in the cheese, which would cause a reaction, and possibly create too many bubbles in the cheese. Conclusion: ** <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-fareast-font-family: 'Times New Roman'; mso-hansi-theme-font: major-latin;"> The enzymatic activity lab allowed students to design experiments to demonstrate how enzyme activity was regulated. Also, by changing different variables one could observe to what extent enzymes regulated the rate chemical reactions. During this specific procedure, an acid, base, or water was introduced to the liver sample before hydrogen peroxide was added to observe whether or not a change in the pH level of the liver would change the rate at which the enzymes worked. The data proved that when the pH was manipulated by the means of <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: Verdana; mso-bidi-font-size: 12.0pt; mso-hansi-theme-font: major-latin;">hydrochloric acid and sodium hydroxide solutions, the enzymes did not function properly. When hydrogen peroxide was added to the liver after the acid or the base, there was hardly a reaction, meaning that no oxygen was produced. However, during the test of the liver that was accompanied by the distilled water, an abundance of oxygen was produced by the enzyme catalase. This reaction took place because the distilled water was utilized as a variable, therefore it did not alter the natural pH of the liver. Subsequently, this lab procedure proves that the enzyme catalase functions best under the condition of its natural pH level. Any minor changes in the pH level result in the enzymes not functioning properly. One may draw a parallel to the human body to examine this point further. If the pH levels in the body were to change, then the preprogrammed enzymes would not be able to carry out their functions as instructed because they would not know how to operate under foreign conditions.
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The results of the liver lab prove that the hypothesis was correct in that any change in the pH level of the liver would result in the enzyme catalase not functioning properly, however there may have been minor flaws in the procedure. Although these flaws did not drastically affect the results of the lab, they must be acknowledged in order to improve next time. One mistake made, was that the liver samples were not allowed to soak in the HCL, NaOH, and distilled water for the same amounts of time. The time was supposed to be observed, but there was some discrepancy in this area. Finally, the liver samples that were subjected to the testing were not exactly <span style="font-family: 'Calibri','sans-serif'; line-height: 115%; mso-ascii-theme-font: major-latin; mso-bidi-font-family: 'Times New Roman'; mso-bidi-font-size: 12.0pt; mso-fareast-font-family: 'Times New Roman'; mso-hansi-theme-font: major-latin;">the same size. They were similar in mass and size; however more accurate measurements would have yielded more precise results.

In conclusion, the liver lab allowed the students to take independent control of their procedure, therefore allowing them to interact more closely with the measurements and steps that needed to be taken in order to produce accurate results. Although the procedure may have exhibited minor flaws the results clearly reveal that a change in the pH of the liver inhibits the enzymes to function properly.