Yeast Peroxidase Lab

Then the disk should be placed into the hydrogen peroxide solution and times for a reaction. Then another disk is placed into a beaker labeled potato extract for 5 seconds, then placed into the hydrogen peroxide solution and timed for a reaction. The hypothesis for the pH test were correct because the reactions did slow down as the pH got higher. In the data, the average rate of reaction for the highest pH level was 180 seconds or higher. The lowest pH level had the fastest reaction of them all at about 61 seconds.

This lab activity had students to conduct various tests on enzymes in different environments to determine how the rate of reaction in decomposing hydrogen peroxide (H2O2) to water and oxygen gas was dependent on environmental adjustments. Controls are essential in a lab to help rule out any unnecessary results.

The purpose of this experiment was to simply measure oxygen production rates released from decomposed hydrogen peroxide under different conditions (concentration of enzymes, temperature, and PH level).

Figure 1: Amount of O2 gas curves to the time at which it was measured according to low, medium, and high pH.

The purpose of this laboratory experiment is to explore the effects of pH has on a reaction rate. The reaction studied was the breakdown of hydrogen peroxide catalyzed by enzyme peroxidase. Peroxidase is a large protein containing heme co factors in its active site. Four trials were ran at pH levels of 3, 5, 7, and 9. I hypothesized that the reaction would run very quickly at a pH 7, since that is the normal condition of cells where peroxidase is found.

The purpose of this experiment is to learn the effects of a certain enzyme (Peroxidase) concentration, to figure out the temperature and pH effects on Peroxidase activity and the effect of an inhibitor. The procedure includes using pH5, H202, Enzyme Extract, and Guaiacol and calibrating a spectrophotometer to determine the effect of enzyme concentration. As the experiment continues, the same reagents are used with the spectrophotometer to determine the temperature and pH effects on Peroxidase activity. Lastly, to determine the effect of an inhibitor on Peroxidase, an inhibitor is added to the extract. It was found that an increase in enzyme concentration also caused an increase in the reaction rate. The reaction rate of peroxidase increases at 40oC. Peroxidase performed the best under pH5 and declined as it became more basic. The inhibitor (Hydroxy-lamine) caused a decline in the reaction rate. The significance of this experiment is to find the optimal living conditions for Peroxidase. This enzyme is vital because it gets rid of hydrogen peroxide, which is toxic to living environments.

In one tube went 0.1 ml guaiacol, 0.2 ml H202¬ and 4.7 ml dH20 for a total of 5 ml. In the other test tube 1.0 ml of peroxidase and 4.0 ml dH20 was combined for a total of 5 ml. The second part of this test was to observe the reaction rate between the peroxidase enzyme and the hydrogen peroxide substrate with guaiacol as the reducing agent every 20 seconds for 10 minutes. The contents of the two test tubes were mixed together and then transferred some of the mixture into a cuvette that could fit into the spectrophotometer. The liquids were combined, poured into the cuvette, put into the spectrophotometer and its absorption rates were recorded every 20 seconds for 10 minutes.

The mixture at 60 °C had values of zero for both the at temperature mixtures and the recovered mixture, indicating that the high heat completely denatured the enzyme. This leads to the conclusion that peroxidase has an optimal temperature near 22°C and anything lower will have low yield, while anything higher will denature. This proves the importance of temperature on enzyme shape and thus

The research and observations of this lab primarily focused on the enzyme activity of the enzyme Peroxidase. Peroxidase is a large protein and is composed of more than three hundred amino acids. The enzyme was selected as it is easy to experiment with and effectively showcases the effects of varying independent variables, such as pH and temperature. Peroxidase catalyzes the decomposition reaction of the chemical Hydrogen Peroxide ( H2 O2 ) into water and an electron donating molecule, which stands for R in the written chemical equation. ( The equation is displayed below:

Each of the five tables shows the amount of oxygen produced by five varying pH levels, over a certain amount of time ranging from fifteen seconds to thirty seconds. In Table 1.0 the volume of oxygen produced by combining the catalase with pH level 3 solutions displayed. Correspondingly,

Hypothesis: If the pH is around seven, then the rate of the reaction for the enzyme peroxidase will be the highest. This is because the optimal pH range for the enzyme peroxidase is around seven and at its optimal pH an enzyme functions the best.

pH levels effect on Enzyme Activity and Concentration Abstract: pH level may affect enzyme activity and the performance of the enzyme which is crucial in order for the enzyme to properly function in our biological systems. We investigated the relationship between pH level and rate of enzyme function at that certain pH level. The enzyme concentration was recorded at varying levels of pH using the enzyme Lactase, substrate ONPG, and phosphate buffer in a spectrophotometer set at wavelength 405nm. As pH levels became extremely high or low, enzyme concentration decreased.

This experiment is designed to analyze how the enzyme catalase activity is affected by the pH levels. The experiment has also been designed to outline all of the directions and the ways by which the observation can be made clearly and accurately. Yeast, will be used as the enzyme and hydrogen peroxide will be used as a substrate. This experiment will be used to determine the effects of the concentration of the hydrogen peroxide versus the rate of reaction of the enzyme catalase.

The results from my preliminary experiment show that 100 cm3 of oxygen has been produced in the first 30 seconds.. This reaction is far too quick and will prevent me from analysing the effects different substrate concentrations have on enzymes if I decide to continue. I will therefore lower my yeast concentration to 1%. I will also measure the volume of oxygen produced every 15 seconds, instead of every 30 seconds.

Catalase is an enzyme found in mammalian tissues and catalyzes the reaction by which hydrogen peroxide (H2O2) is decomposed into oxygen (O2) and water (H2O).(4) Enzyme activity and efficiency is affected by the conditions that the reaction occurs in. Changes in conditions can result in altering of the enzyme’s three dimensional shape and affect the enzyme’s function.(1) The conditions that influence the activity of an enzyme include pH, temperature, substrate concentration(s), ionic strength, and nature of salts present.(2) At a constant set enzyme concentration, the reaction rate increases with increasing substrate concentration until all available enzyme in bound to the substrates, at that point adding more substrate will not further reaction rate.(3)