Potato Catalase lab

Investigating the Effects of pH and Temperature on Enzyme Activity

Background: In this lab exercise we’ll investigate some of the properties of enzymes. Almost all chemical reactions that occur in living things are catalyzed by enzymes. Enzymes are large protein molecules (macromolecules) that catalyze or speed up chemical reactions without being altered themselves. Proteins can be just about any size or shape, which is useful since it’s the shape of an enzyme that determines the reactions it can catalyze. However, enzymes are sensitive to changes in temperature and pH, which alter their shapes and can even destroy catalytic ability (denaturing). Enzymes have evolved to work most efficiently at the temperature and pH found in the part of the organism where they are needed. Many enzymes in the human body function most efficiently at 37^oC and at a pH of 7.4.

The enzyme you will investigate is called catalase. Catalase is found in tissues of many organisms (both plants and animals) because it plays a very important role in protecting cells. Its purpose is to destroy toxic substances which may be introduced into cells. Also, some cells use catalase to destroy cellular debris or worn out organelles. Hydrogen peroxide is a normal by-product of cellular metabolism but, it is also toxic to cells. Under normal conditions organisms produce the enzyme catalase that quickly changes hydrogen peroxide into two harmless substances, oxygen and water. However, the function of the enzyme is affected by changes in the environment. Catalase works to break down hydrogen peroxide by the following chemical reaction:

2 H₂O₂ ------------------>2 H₂O+ O₂

(Catalase)

Purpose: What is the effect of altering the pH on the function of catalase?

Pre Lab Questions (ANSWER IN COMPLETE SENTENCES IN YOUR LAB NOTEBOOK. This answers should be directly below the procedure that you worked on in class).

1. What is catalase and where is it found in living organisms?
2. What reaction does catalase speed up (catalyze)?
3. Write the correct chemical reaction for hydrogen peroxide being catalysed into water and oxygen.
4. What is the substrate in the chemical reaction?
5. What are the products in the chemical reaction when catalase breaks down hydrogen peroxide?
6. Write hypotheses for how you think changes in pH AND temperature will affect the action of the catalase enzyme in the potato.
Hypothesis for change in pH.
Hypothesis for change in temperature:

Materials:

• 0.1 M HCl (hydrochloric acid)
• 0.1 M NaOH (sodium hydroxide)
• 6 large test tubes
• test tube rack
• hot water bath
• ice bath
• 10ml graduated cylinder
• 100ml beaker
• Transfer pipet
• 3.0% H₂O₂ (Hydrogen Peroxide)
• test tube holders (for hot water bath)
• Safety glasses
• Potatoes
• Ruler

Safety: HCl and NaOH are strong acids and bases. If you spill these chemicals on your skin, immediately wash in running water. Wear safety glasses at all times. Be careful of hot water and hot plates.

Part A:

1. Label 3 large test tubes hot, cold and room temp.
2. Cut 3 1.5cm³ pieces of potato with peel removed.
3. Use a mortar and pestle to macerate (mash/grind up) each cube.
4. Place ALL of the macerated potato cubes into each labeled test tube.
5. Add 1ml of distilled water to each test tube containing potato mash.
6. Place the test tube labeled hot in the hot water (100^oC) bath for 3 minutes.
7. Place the test tube labeled cold in the cold water bath (0-4^oC) for 3 minutes.
8. Leave the test tube labeled room temperature in the test tube rack.
9. After 3 minutes, remove the test tube from the baths and allow the hot test tube to cool.
10. Add 5ml of H₂O₂ (3.0% hydrogen peroxide) to each tube.
11. Wait one minute while reaction occurs then measure the height of the bubbles in each tube in cm.
12. Record in Table 1. Measure the radius of the test tube and record in Table 1.

Part B:

1. Label 3 test tubes acid, base and water.
2. Cut 3 1.5cm³ pieces of potato with peel removed.
3. Use a mortar and pestle to macerate (mash/grind up) each cube.
4. Place ALL of the macerated potato cubes into each labeled test tube.
5. Add 1ml of 0.1M HCl (hydrochloric acid) to the tube labeled acid. Gently mix contents with wooden or glass rods. SEE SAFETY NOTE ABOVE.
6. Test the pH of the solution by dropping a small amount of liquid onto a pH strip. Record in Data Table 2.
7. Add 1ml of 0.1M NaOH (sodium hydroxide) to the tube labeled base. Gently mix contents with wooden or glass rod. SEE SAFETY NOTE ABOVE.
8. Test the pH of the solution by dropping a small amount of liquid onto a pH strip. Record in Data Table 2.
9. Add 1ml of water to the tube labeled water. Mix contents by gently swirling. Test the pH of the solution by dropping a small amount of liquid onto a pH strip. Record in Data Table 2.
10. Wait 3 minutes then add 5ml of 3%H₂O₂ to each tube.
11. Wait 1 minute for reaction to occur, then measure and record the height of the bubbles in cm.
12. Record in Table 2. Measure the radius of the test tube and record in Table 2.

This lab is borrowed from EDCC