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 * Catalase Lab Report** Lab Group 5 //Michael Motier, Carrie Reinking, Joey Cover//

Enzymes are an integral part of the metabolic process, they are responsible for regulating the chemical reaction between molecules by acting as catalysts (1). Enzymes are substrate specific; meaning they can only be activated by a specific molecule or molecular compound (1). The enzyme contains an active site where in most cases digestion or combination takes place (1). Enzymes aid to speed the chemical reaction process but are not consumed in the digestion or combination phase (1).  Catalase is an enzyme that is site specific to peroxide (H2O2). It works by accepting the peroxide and breaking it in to water (H2O) and releasing oxygen (O2) in the form of bubbles (2). Peroxide on its own is dangerous to most organisms because it is an instable molecule (2) (free-radical).  We intend to test the effects of potato catalase when peroxide is applied in various experiments. Our tests will include the effects of temperature as it applies to the reaction between catalase and peroxide, the effects of increased, neutral and low PH levels and how the concentration of catalase affects the chemical reaction with the peroxide. We hypothesize that when the PH levels are changed it will have a denaturing effect on the catalase, thus directly affecting the enzyme function. We believe that by increasing or decreasing the temperature less of a reaction will occur between catalase and peroxide because temperature has a direct effect on enzyme function (1). We predict that by increasing the concentration of catalase present during the test we will see an increase in activity; the more catalase present, the longer the reaction will be sustained. We expect to offer conclusive evidence that under certain circumstances the effects of catalase can be measured and to an extent predicted. In an attempt to better describe enzyme function we will first start by examining the effects of temperature on the catalase enzyme. Before starting the temperature portion of this experiment we had to first label and mark 3 test tubes to measurements of 1 cm and 5 cm. Next, each tube was filled to the 1 cm mark with catalase buffered to a pH of 7. The temperature of the buffered catalase was then adjusted to specific temperatures, tube 1 was cooled to a temperature of 7° C, tube 2 was placed in an incubator until the temperature was 37°C, tube 3 was placed in boiling water until the temperature reached 100°C, and as a control, tube 4 was adjusted to room temperature (20°C). Once the tubes were within the accurate temperature ranges, hydrogen peroxide (H2O2) was added until the volume of each tube reached the 5 cm mark. We then observed the reaction for 20 seconds after which we obtained and recorded the results by measuring the height of the bubble column from the 5 cm mark. The incubator (37°C) produced the highest bubble column at 3mm with the lowest column produced by the boiling water (100°C) at 0mm. We continue our tests by examining the effects of concentration on enzyme activity. Before starting the experiment, we had to label three test tubes at various measurements. Next we filled test tube 1 to 1 cm with buffered catalase, test tube 2 to 2 cm buffered catalase, and test tube 3 to 3 cm buffered catalase. Then we filled test tube 1 to 5 cm with hydrogen peroxide, test tube 2 to 6 cm with hydrogen peroxide, and test tube 3 to 7 cm hydrogen peroxide. After waiting 20 seconds, we recorded the height of the bubble columns to determine the rate of reaction in each test tube. The measurements were taken starting from the highest mark on each test tube (5 cm, 6 cm, 7 cm). We later noted that over more time, the bubble columns in each tube reached equal heights Our final test was to examine the effects of pH on enzyme activity. Before starting the testing portion of the experiment, we had to label each test tube at the 1 cm, 3 cm, and 7 cm marks. We filled each test tube to 1 cm with nonbuffered catalase. Tube 1 was filled to the 3 cm mark with water that was adjusted adjusted to an acidic pH of 3 by the addition of HCl. Tube 2 was filled it to the 3 cm mark with water adjusted to a neutral pH of 7.Tube 3 was filled it to the 3 cm mark with water adjusted to a basic pH 11 by the addition of NaOH. Next, we filled the three test tubes to the 7 cm mark with hydrogen peroxide. We had to allow the mixtures to sit for one minute, and then stir the mixtures, then wait for another 20 seconds. After the 20 seconds has passed, we measured the reaction in the tubes in the form of bubbles. The bubble column heights were measured from the 7 cm marks on the test tubes.
 * **Tube** || **Temperature °C**  || **Bubble Column Height (mm)**  ||
 * 1 Refrigerator || 7°C || 1 mm ||
 * 2 Incubator || 37°C || 3 mm ||
 * 3 Boiling water || <span style="font-family: Times New Roman,serif;">100°C || <span style="font-family: Times New Roman,serif;">0 mm ||
 * <span style="font-family: Times New Roman,serif;">4 Room Temperature || <span style="font-family: Times New Roman,serif;">20°C || <span style="font-family: Times New Roman,serif;">.5 mm ||
 * <span style="font-family: Times New Roman,serif;">**Tube** || <span style="font-family: Times New Roman,serif;">**Amount of Catalase**  || <span style="font-family: Times New Roman,serif;">**Bubble Column Height (mm)**  ||
 * <span style="font-family: Times New Roman,serif;">1 || <span style="font-family: Times New Roman,serif;">1 cm || <span style="font-family: Times New Roman,serif;">1 mm ||
 * <span style="font-family: Times New Roman,serif;">2 || <span style="font-family: Times New Roman,serif;">2 cm || <span style="font-family: Times New Roman,serif;">1 mm ||
 * <span style="font-family: Times New Roman,serif;">3 || <span style="font-family: Times New Roman,serif;">3 cm || <span style="font-family: Times New Roman,serif;">.5 mm ||
 * <span style="font-family: Times New Roman,serif;">**Tube** || <span style="font-family: Times New Roman,serif;">**PH**  || <span style="font-family: Times New Roman,serif;">**Bubble Column Height (mm)**  ||
 * <span style="font-family: Times New Roman,serif;">1 || <span style="font-family: Times New Roman,serif;">3 || <span style="font-family: Times New Roman,serif;">0 mm ||
 * <span style="font-family: Times New Roman,serif;">2 || <span style="font-family: Times New Roman,serif;">7 || <span style="font-family: Times New Roman,serif;">.5 mm ||
 * <span style="font-family: Times New Roman,serif;">3 || <span style="font-family: Times New Roman,serif;">11 || <span style="font-family: Times New Roman,serif;">0 mm ||

<span style="font-family: Times New Roman,serif;">Overall, our results were very conlusive as to how the catalase reacted under certain conditions. Our hypothesis for the effect of temperature on catalase reaction was supported by our data. We found that the incubator, which had the most natural temperature, caused the fastest reaction causing the bubble column to grow taller than the others in the time allotted for the experiment. We believe this occurred because enzymes denature when heat is applied, causing no reaction, and when temperature is decreased, molecules slow down and therefore cannot react as quickly as normal. With these results and continued experimenting, one could determine there is an optimal temperature range at which catalase will react with peroxide. Our hypothesis on how the concentration of catalase affected the reaction was incorrect. We expected more of a reaction to occur with a higher concentration of catalase but the results showed the opposite, with a higher catalase concentration delaying the reaction. Our explanation for this was because the peroxide was becoming diluted through the catalase, therefore taking longer to react. As previously stated, we later noted the bubble columns reached equal heights with more time, showing that the concentration doesn't limit the amount of reaction it only slows it down.We hypohesized that by changing the pH of the catalase, either increasing or decreasing, the enzyme would denature and less of a reaction would occur. This prediction was supported by our results where no reaction at all ocurred with the addition of an acid or base. When the pH was kept neutral, the most reaction ocurred with a .5 mm bubble column. The enzyme catalase, being a protein reacted as we expected with the change in temperature and pH causing denaturing. Although our prediction for the concentration of catalase was incorrect we now understand the reasoning behind the results of our testing.

//<span style="font-family: Times New Roman,serif;">Works Cited // //﻿// <span style="font-family: Times New Roman,serif;">﻿(1) Campbell, et al. Biology. 8th ed. San Francisco: Pearson Benjamin Cummings, 2008. Print.

<span style="font-family: Times New Roman,serif;">(2) Mader, Sylvia S. Biology Laboratory Manual. Ed. Lynn Nagel. 10th ed. Boston: McGraw-Hill Learning Solutions, 2010. Print.