Catalase Lab

Abstract In this laboratory exercise we studied enzyme catalase, which accelerates the breakdown of hydrogen peroxide into water and oxygen. The purpose was to isolate catalase and measure the rate of activity under different conditions. The laboratory was also conducted in association with a second laboratory that measured the effects of an inhibitor on the enzymes. Changes in temperature and pH along with Substrate Concentration and Enzyme Concentration were the conditions tested in the experiment.
Each lab was assigned to a group A, B, C and D in our class performed this experiment. The data presented in this report will reflect the average rates of change in O2 ml/min of our class results. In the later discussion sections, it will become
obvious that human error was the deciding factor in the data collection.
Introduction
In each individual cell of a human there are many chemical reactions taking place, performing necessary functions for being a large, complex, multicellular organism. This is pretty easy to understand. How do these reactions happen is not so easy to understand. Chemical reactions involve the breaking and reforming of chemical bonds between molecules (substrate(s) of the reaction), which are transformed into different molecules (product(s) of the reaction). Chemical reactions can occur spontaneously (without added energy or intervention), and a lot of the chemical reactions necessary for life processes are spontaneous; some however, are not. Metabolic pathways are processes which involve many chemical reactions that occur in a specific order.
For example, to get energy out of a molecule of glucose, a series of reactions must take place in a specific order to break the bonds between the carbons of the glucose molecule. In addition, we have to rely on a series of chemical reactions that break down stored glycogen into glucose molecules to have glucose molecules in the first place. If we had to rely on these reactions to take place spontaneously, we would have to wait a very long time we probably wouldn't be here. Enzymes catalyze chemical reactions so that they occur in a timely and sequential manner to produce a product.
Enzymes are biological catalysts. They help to increase the rate of chemical reactions. Enzymes are most often proteins and their three-dimensional shape is important to their catalytic activity. Because of their 3-D shape, enzymes are highly specific for the substrates that they will act upon. So any one "function", such as getting energy from a glucose molecule, actually involves many reactions, each with a specific enzyme.
Enzyme activity is influenced by many factors. We (the class) examined some of the major factors, which influence the activity of an enzyme called catalase. Catalase is an enzyme that is found in many cells. The highest levels of catalase are in the liver because the liver often functions to break down toxins present in the blood. Catalase catalyzes the breakdown of hydrogen peroxide: Peroxides can form in the body during respiration, and are chemically reactive, which means that they can chemically modify (and thus render useless) other biological molecules. In this lab we studied; the influence of catalase concentration on enzyme activity, the influence of Substrate concentration on enzyme activity, the influence of pH on catalase activity and the influence of temperature on catalase activity.

Methods This lab experiment was done in four different procedures labeled A, B, C and D we worked as a group of 4 with our lab partners at our tables to perform the experiments. Each group was assigned one of the specific experiments in the lab. We recorded our results on the on the board so we were able to get the data for each experiment performed. Each group had to analyze all 4 parts (A-D) of the experiments from the lab's pooled data even though each group only performed one part.
The following “general” procedures were used to obtain oxygen production and were to be repeated for all sections A-D.
A. The influence of Catalase Concentration on Enzyme Activity
B. The influence of Substrate Concentration on Enzyme Activity
C. The influence of pH on Catalase Activity
D. The influence of Temperature on Catalase Activity
Each person in our group was assigned a particular task and that task was repeated by the same person each time we repeated a step to test another level of the influence of Catalase Concentration on Enzyme Activity (this should have been uniform procedure for each lab group).
Jenny - Loaded the soaked filter disks into the reaction chamber and added the hydrogen peroxide.
Shelby – Monitored the timing of the experiment
Ariel – measured the amount of oxygen produced, recorded the data and performed the calculations
I (Karee) – Prepared the apparatus for O2 collection
General procedures (for experiments A-D) continued
Part 1: Preparation and Collection of Equipment
1. Collected the kit with the material needed to conduct the lab experiment. Which included: 50ml graduated cylinder, clamp, and a large glass bowl.
2. Filled the glass bowl with tap water
3. Filled the graduated cylinder with 50 ml of tap water
4. Turned the graduated cylinder upside down in the glass bowl filled with water and leave the top half under water
5. Suspended the inverted cylinder with the clamp and stopped the top of the cylinder 3 cm above the bottom of the bowl. We tightened the clap to help keep the cylinder in place.
Part 2: Preparation of the Reaction Chamber
1. Loaded the reaction chamber with 4 liver extract soaked disks
2. Added 10ml of hydrogen peroxide into the reaction chamber without allowing it to touch the disks
3. Sealed the chamber with a rubber stopper that has a hose attached to it
4. Laid the reaction chamber on it’s the side with no disks
5. Inserted the hose into the graduated cylinder Recorded the initial water level then proceeded to flip over the reaction chamber to initiate the reaction
6. Let the reaction continue for 1 minute
7. After 1 minute read the final water level and recorded the value in the data table
8. Calculated the difference between the final reading and the initial reading and recorded it in the table.
9. Broke down the reaction chamber; discarded the tested disk(s) and solution, cleaned and dried the reaction chamber & repeat the experiment using the variation indicated for the group assigned
Group A Procedure: The influence of Catalase Concentration on Enzyme Activity (my group performed this procedure).

1. Prepared filter disks
a. Collected and mixed liver extract
b. Pour a small amount of the liver extract into a petri dish
c. Cut out 25 filter disks and soaked them in the liver extract
d. Added distilled water into a different petri dish
e. Cut 4 filters and let them soak in the distilled water

2. Prepared the reaction Chamber
a. Loaded the reaction chamber with 4 extract soaked disks, ran the experiment as explained in the general directions and recorded the results in table 1.
b. Repeated the experiment using one, two, three, five and six extract soaked disks and recorded the results in table 1
c. Repeated the experiment using the four disks soaked in distilled water and recorded the results in table 1.
3. Cleaned up and Washed our hands
4. Recorded and calculated the rate of reaction per minute (see Table 1)
5. Created a line graph to show our data (see Graph 1)
6. Analyzed our data and created a written overview of the results (see results for procedure A)
Group B Procedure: The influence of Substrate Concentration on Enzyme Activity
1. Prepared filter disks
a. Collected and mixed liver extract.
b. Pour a small amount of the liver extract into a petri dish.
c. Cut out 20 filter disks and soaked them in the liver extract.
2. Prepared the reaction chamber
a. This group loaded the reaction chamber with 4 extract soaked disks. They used 3% H2O2 and then ran the experiment they recorded their data in table 2.
b. They then repeated the experiment using 1.5% H2O2, 0.75% H2O2, 0.3% H2O2 and distilled H2O.
3. Cleaned up and Washed their hands
4. Recorded and calculated the rate of reaction per minute (see Table 2)
5. Created a line graph to show their data (see Graph 2)
6. Analyzed their data and created a written overview of the results (see results for procedure B)

Group C Procedure: The influence of pH on Catalase Activity
1. Prepared filter disks
d. Collected and mixed liver extract.
e. Pour a small amount of the liver extract into a petri dish.
f. Cut out 25 filter disks and soaked them in the liver extract.
2. Prepared the reaction chamber
c.
This group loaded the reaction chamber with 4 extract soaked disks. They used 3% H2O2 with a pH of 3 and then ran experiment they recorded their data in Table 3.
d. Then they repeated the experiment using 3% H2O2 with a pH of 5 and recorded the results in Table 3
e. Then they repeated the experiment using 3% H2O2 with a pH of 7 and recorded the results in Table 3
f. Then they repeated the experiment using 3% H2O2 with a pH of 9 and recorded the results in Table 3
g. Then they repeated the experiment using 3% H2O2 with a pH of 11 and recorded the results in Table 3
3. Cleaned up and Washed their hands
4. Recorded and calculated the rate of reaction per minute (see Table 3)
5. Created a line graph to show their data (see Graph 3)
6. Analyzed their data and created a written overview of the results (see results for procedure C)
Group D procedure: The influence of Temperature on Catalase Activity
1. Prepared filter disks
a. Collected and mixed liver extract
b. They put 4 ml of the extract in a test tube and placed it in boiling water for five minutes. Then they put the boiled extract into a petri dish and soaked 4 disks in it
c. Then they pour the rest of the liver extract into a small petri
dish
d. Then they placed 20 filter disks into the liver extract and let them soak
2. Prepared the reaction chamber
a. They loaded the reaction chamber with 4 soaked disks and ran the reaction at 210 C room temperature. They recorded their results in Table 4
b. They repeated step a three more times once at O0C, 370C and once at 1000 C and recorded their results in Table 4
3. Cleaned up and Washed their hands
4. Recorded and calculated the rate of reaction per minute (see Table 4)
5. Created a line graph to show their data (see Graph 4)
6. Analyzed their data and created a written overview of the results (see results for procedure D)