The Function of Enzymes in Extracting Catalase from Raw Potato Tissue

Introduction

Enzymes work as catalysts, which are substances that speed up chemical reactions. Enzymes are proteins and their shape determines the specificity of the enzyme, as well as plays an important part in its catalytic activity. (Catalase Lab, May 1, 2011). This means that there are hundreds of different enzymes that need to catalyze the various reactions that take place in the human body each day. (Belk, C. & Maier, V. Borden, 2012).Enzyme activity is often influenced by many factors. One of these factors is going to be examined in this lab and that’s ion concentration on catalase activity. In this lab I will be using the enzyme, catalase which is mainly found in the cells and tissues of many if not all living organisms. The function of catalase is to help breakdown toxic substances that arise from normal cellular metabolism. (UNT Laboratory Staff, 2012 Fall Semester).

I will be extracting the catalase from the raw potato tissue. With the Hydrogen peroxide (H2O2) being a reactant as well as a waste product that comes from metabolic processes in the cell, its toxicity will cause the cell to be damaged if not removed quickly. (UNT Laboratory Staff, 2012 Fall Semester). Thus the catalase will rapidly convert the hydrogen peroxide to oxygen and water (Worthington, K. & Worthington, V., 2011). The purpose of this experiment is to observe how ion (salt) concentration effects catalase activity as it involves enzymes. Which leads to my hypothesis, when more NaCl concentration is added, the height of the foam (bubbles) which indicates catalase activity, will increase in the solution. The reasoning used to develop my hypothesis was that I believed when you have more reactants it results in a greater amount of solution.

Materials

• Catalase
• 3% H2O2
• NaCl solutions (0%, 0.9%, 5%, 10%, 20%)
• 1 Cork borers/straws (~ 10mm)
• 5 Test tubes
• 1 Test tube rack
• 1 Mortar and pestle
• 1 Small beaker of water
• 1 Sharp blade
• 2 Plastic pipettes
• 1 Metal Spatula (narrow)
• 1 Small cutting board
• 1 Timer/clock
• 1 Wax pencil
• 1 Metric ruler

Procedure

First of all get all your materials unless they are laid out for you already. When all of this is done you will need to think up a hypothesis with your group. Get your raw potato out. Remove a solid cylinder of the raw potato tissue out by using the cork borer with a diameter of about 8 mm. You will only need one solid cylinder of raw potato tissue, but if you mess up then you have a whole potato to get it right.

Next use the sharp single-edged razor blade to cut that cylinder into pieces that are 1 cm long. You will need 5 pieces in total for this experiment. Once done if not using the potato cylinders that have been just cut you will need to put them into the small beaker of water, so that they don’t dry out. Get out your test tubes and label them with your wax pencil. Label the tubes respectively 0%, 0.9%, 5%, 10%, and 20%.

You will now need to use the metric ruler to measure 1 cm and 2 cm on each tube from the bottom, and make sure you use the wax pencil to mark the tube. Next you will use the mortar and pestle to deeply mash in a potato tissue cylinder piece, and then use the metal spatula to scrape all of the contents into the tube labeled “0%”. Then add “0% NaCl” to the 1 cm mark in the tube by using the plastic pipette, and make sure it is as close to the 1 cm mark as possible. Repeat this step for all of the other tubes labeled 0.9%, 5%, 10%, and 20%, making sure that the amount of NaCl solution is correctly added.

Have the timer ready. In repeating this step take out your test tube rack, so then you can let the tubes stand for at least 10 minutes. Use your timer now, to be accurate (It is okay if it goes over 10 minutes but it has to be at least 10 minutes) before going on to the next thing. Once done waiting for the tubes to stand, you can now go to the refrigerator and take the H2O2 out. It will be in a container, but still handle with care.

You will need to add this H2O2 to each of the test tubes to the 2 cm mark. After 2 minutes, you will measure the height of the foam (bubbles).You have to make sure you measure the test tube from the very bottom to the very top of the bubbles. Record this and any other observations on a Table and use this data to make graph. Once done empty all the contents of the tubes into the waste bucket(s) provided. Place these used test tubes in the tub that will be by the sink. Make sure all your equipment and solutions are returned to their original places.

In this experiment the independent variable is the sodium chloride (NaCl) solution, dependent variable is the hydrogen peroxide (H2O2) and height of foam (mm) and the standardized variable is the fresh potato tissue. The kind of data collected was the height of the foam (mm).The method of gathering data was making a table and from that table making a graph to further understand the results. The kind of measurements/observations that were made in order to quantify the data was noting the color of the solution, and how the potato dispersed in the tube.

Results/Observations

Table 1 The effect of ion concentration (NaCl) on catalase activity

Concentration of NaCl Height of foam (mm) Other observations

0% 2.5 Potato some at bottom, some at top

Color: white

0.9% 2.8 Potato is mostly at top Color: yellow

5% 2.4 Potato mainly at top Color: semi-yellowish

10% 2.3 Potato all around (middle, top & bottom) Color: white

20% 2.2 Potato all at top none at bottom Color: white

Figure 1

Table 1 shows in the first column the different percentages of NaCl concentration per test tube that was added, the second column shows the height of foam that was measured once the hydrogen peroxide was added to each test tube in millimeters and lastly the third column shows the other observations of where the potato was in the test tube and what the color of the solution became once the hydrogen peroxide was added. Figure 1 shows the data from Table 1, but on a graph, it’s another visual to see what the effect of ion concentration (NaCl) had on the catalase activity.

Discussion/Conclusions

The purpose of this experiment is to observe how ion (salt) concentration effects catalase activity as it involves enzymes. Which leads to my hypothesis, when more NaCl concentration is added, the height of the foam (bubbles) which indicates catalase activity, will increase in the solution. Unfortunately my results don’t support my hypothesis. Table 1 is the first instance where my hypothesis is rejected. In Table 1 with the first row, first column dealing with 0% of NaCl concentration, the results were 2.5mm for the height of the foam. Knowing that information the height of the foam had to increase with more NaCl concentration and it did with the second row, first column 0.9% concentration of NaCl. It’s height of foam was 2.8mm, so far validating my hypothesis.

I then did the test tube involving 5% of NaCl concentration third row, first column and the results were baffling to me the height of the foam was 2.4 mm. It dropped drastically and thus proving my hypothesis invalid, but maybe this is a fluke, so I did the 10% concentration of NaCl fourth row, first column and it dropped again by 0.1 mm to 2.3mm. Officially my hypothesis has been rejected thus making it invalid. Figure 1 helps to further Table 1 by showing it as a visual.

I wanted to know why and how the hypothesis was invalid and thus rejected. I researched later that when you have a higher ion concentration the ion comes in contact with the bonds between the side groups of the amino acids that make up the enzyme. This in turn causes the enzyme to lose its shape, thus causing it to no longer accept, or react the way it’s supposed to. The rate of the enzyme (catalase) activity decreases instead of increases in the solution. (Enzymes, Science of Everyday Things, 2002).

Some factors that might have affected the results would be the temperature of the hydrogen peroxide if it was warm or hot instead of cold, and if you shook the test tube when it was supposed to stand still for the allotted time. Some errors include: if you didn’t measure all the substances correctly, didn’t time the experiment where needed and label the test tubes with their names, thus causing a mix up.

Some applications/usefulness this experiment might have for the real world include: metabolic enzymes, enzymes that deal with the body from breathing to thinking, your diet as in the raw foods you eat that aid in digesting those foods, and lastly in digestion, involving your digestion glands secreting juices containing enzymes (Enzymes, Science of Everyday Things, 2002).