Peroxidase Temperature Lab Report

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:

The data in proves that our hypothesis was correct. When we increased the temperature to 35°C, the the enzyme activity increased because kinetic energy increased, increasing the collisions between the substrate and the enzyme, and thus creating a higher chance of reaction. When we increased the temperature to 45°C, the enzyme activity decreased as the enzyme became denatured,because the atoms in the enzyme had enough energy to overcome the hydrogen bonds between the R groups that give the enzyme its shape From our data, we could conclude that the optimal temperature of turnip peroxidase is around 35°C and around 45°C, it will start to denature.

Peroxidase is an enzyme found in potatoes that catalyzes the breakdown of hydrogen peroxide, H2O2, into O2 gas and water. We examined the different pH environments that can affect the enzyme activity during the breakdown of H2O2. In order to do this, we added different levels of pH, low, medium, and high, into different test tubes with the enzyme and H2O2, and we then inverted the tube. The amount of O2 gas produced was then measured and recorded. The result was that the higher pH produced more gas, followed by medium pH, then low pH. The enzymes were more active in the pH of about 10. It increased

Turnips and horse radish roots are rich source of this enzyme. In this experiment, we would carry out a reaction between hydrogen peroxide and guaiacol which is colorless dye, using peroxidase as a catalyst, to produce water and an oxidized form of guaiacol which is brown. The formation of brown color would serve as an indicator that the breakdown of Hydrogen Peroxide took place. The enzyme activity would be directly proportional to the brown color intensity. The color intensity would be measured using a spectrophotometer and standardized to find the corresponding concentration for each absorbance unit.

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.

I also hypothesized that when peroxidase enzymes go over a temperature of 60°C or go under 30°C they will denature. My results show the temperature graph had a spike at 37°C, and its reaction rate was almost 2 times greater than any other temperature at this temperature. This concludes that the optimal temperature of the peroxidase enzyme is 37°C. Again, the enzymes did not stop functioning completely when they were not in their optimal temperature zone, but they became less active and efficient. My last hypothesis was that when peroxidase enzymes have a concentration of 2 mL or more, they will denature.

Discussion: When solutions are increased in temperature, there is more kinetic energy, and the molecules speed up as they transition from liquids to gas. This increased movement of molecules, increases the collision of molecules which, increases the rate of reactions (1). It was hypothesized that as the temperature of the hydrogen peroxide increased that rate of reaction with the potato enzyme would increase until the temperature caused the enzyme to

In this study, the effects of temperature and pH were measured on the catalytic ability of the enzyme catechol oxidase (also known as tyrosinase, diphenol oxidase, or polyphenal oxidase). Enzymes are defined as catalysts in biological systems that lower that energy of activation or Ea of a reaction. When a substrate bonds to a the active site of an enzyme, this forms an enzyme-substrate complex. An enzyme substrate complex consists of one or more substrates bonded to the enzymes active site, which changes shape slightly when bonded to, so as to appropriately fit the substrate(s). This slight change in enzyme shape is called induced fit.

The role of an enzyme is to catalyse reactions within a cell. The enzyme present in a potato (Solanum Tuberosum) is catechol oxidase. In this experiment, the enzyme activity was tested under different temperature and pH conditions. The objective of this experiment was to determine the ideal conditions under which catechol oxidase catalyses reactions. In order to do this, catechol was catalyzed by catechol oxidase into benzoquinone at diverse temperatures and pH values. The enzyme was exposed to its new environment for 5 minutes before the absorbance of the catechol oxidase was measured at 420 nm using a spectrophotometer. The use of a spectrophotometer was crucial for the collection of data in this experiment. When exposed to hot and cold temperatures, some enzymes were found to denature causing the activity to decrease. Similarly, when the pH was too high or low, then the catechol oxidase enzyme experienced a significant decrease in activity. It can be concluded after completing this experiment that the optimal pH for catechol oxidase is 7 and that the prime temperature is 20º C. Due to the fact that the catechol oxidase was only tested under several different temperatures and pH values, it is always possible to get a more precise result by decreasing the increments between the test values. However, our experiment was able to produce accurate results as to the

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.

Introduction Enzymes are proteins that act as a catalyst, this allows a process to undergo more smoothly than without. Enzymes cause a chemical reaction to happen without hesitation. We tested the hypothesis: if peroxidase is exposed to a pH of seven, then it should have a high initial reaction velocity (IRV). Materials and Methods

Every enzyme has a temperature range of optimum activity from 30°C to 37°C since the rate of reaction is highest (Meihua Zhaoa, Guangming Zenga, & Danlian Huanga, 2014). The temperature ranges outside the optimum temperature the enzyme peroxidase is rendered inactive and inhibited. It occurs because as the temperature changes (increases and decreases) would supply enough energy to break some of the intramolecular attractions between polar groups (dipole-dipole attractions and Hydrogen bonding) as well as the hydrophobic forces between non-polar groups within the protein structure (DaasAmiour & LeilaHambaba, 2015). The forces are changed and disturbed thus causes a change in the secondary and tertiary levels of protein structure (Pierre Bauduin & Touraud, 2006). The active site is altered in its conformation beyond its ability to accommodate the substrate molecules it was intended to catalyze. Most enzymes within the human cells will shut down at a body temperature below a certain value such below 4°C which

Temperature can affect the reaction of catechol oxidase by speeding up or slowing down the reaction. I was able to see what happened to the absorbance after changing the temperature of the catechol oxidase solution. I did this by heating and cooling the solutions to measure the absorbances in hot, cold, warm, and room temperature. Then the data was compared to see how the temperature effected the solution. The catechol oxidase solutions reacted best in room temperature (twenty-three degrees Celsius) and the worst in the cold (zero degrees Celsius). I concluded that temperature really does affect the way catechol oxidase reacts.

Heat effects the enzyme activity by speeding up the reaction and/ or completely denatures the enzyme. When the yeast and hydrogen peroxide mixture was put into the 80 degree C, the amount of O2 mL evolved constantly stayed at 0 ml for the complete 10 minutes contrary to the room temperature water in which the amount of O2 evolved increased by about an average of 7 mL for about 6 minutes and then increase by about 1-2 mL. An enzyme denatures with high temperature because heat changes the shape of the active site permanently which causes the enzyme to cease function. On the other hand, a colder temperature will slow the down the enzyme reaction. In 2.4 degree C solution, the amount of 02 evolved by about 1-3 mL every 30 seconds and by 6 min

The purpose of this lab is to explore the effects of enzyme concentration, substrate concentration, temperature, and inhibitors on reaction rate, respectively. To test each of these factors, four activities were completed. The enzyme from Turnip Extract and the substrate Peroxide were tested. The turnip extract was tested at the following concentrations: .5ml, 1.0ml, and 2.0ml. Peroxide was tested at the following concentrations: 0.1ml, 0.2ml, and 0.4ml. In order to understand the effect of temperature on reaction rate the following temperatures were tested: 4C, 23C, 37C, and 60C. To achieve the desired temperatures, an ice bath and water baths were utilized. Lastly, the effect of the inhibitor Hydroxylamine was tested in the following amounts: 0 drops, 1 drop, and 5 drops. In the experiment, Guaiacol was used to determine the rate of reaction by the absorbance measured. When Guaiacol is oxidized it has a brownish color, the presence of the brownish color indicates Peroxide (substrate of peroxidase) has undergone the reaction and has been reduced to water. By this, the reaction rate can be determined by the color change or absorbance over time. For each activity, a spectrophotometer was used to measure the absorbance. After that, the data for each activity was translated to graph where linear regression was used to analyze the effects. The results of the enzyme concentrations on reaction rate are the following: low enzyme concentration (0.5 ml) had a reaction rate of