Affects of Ph on Peroxidase Enzyme

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The Affects of pH on Peroxidase Enzyme

Claire Bell
Biology 1130
Jazmen Myers
Section 36

Abstract
Peroxidase is a turnip enzyme; it is used in the oxidation of hydrogen peroxide to lower activation energy, speeding up the reaction. The activity of peroxidase is highly dependent on its environment and most importantly the pH level. Peroxidase has been the focus of many recent studies and is believed to possibly reduce swelling among other things. We conducted an experiment testing the effect different levels of pH had on the reaction rate of peroxidase. In the experiment we created different solutions all containing hydrogen peroxide, peroxidase, and guaiacol. However each cuvette contained a different pH level, 2,5,7,or 10. Through the experiment we found the activity of peroxidase is highest at its optimal pH level between 6.0 and 7.0 and ceases to react at pH levels 2.5 and lower and between 8.5 and 9.5 and higher. Our data supported the idea that peroxidase is slightly acidic enzyme; however the more acidic pH levels are more effective as inhibitors opposed to the highly basic pH levels.

Introduction
In this experiment we tested the effects of the environmental parameter pH on peroxidase activity. We performed this experiment to observe and analyze the effect pH has on peroxidase activity and find the optimal pH level for peroxidase, as well as the ceasing point in the reaction. Parameters are characteristics that can alter an experiment. The parameter we used in this experiment was pH. The pH scale ranges from levels 0 to 14, 0 being the most acidic, and 14 being the most basic. An enzyme is a biological catalyst made up of proteins. Enzymes speed up reactions by bringing reactants close together and lowering the activation energy. Enzymes are extremely specific to their substrates. A substrate is material where an organism lives or the surface upon which an organism grows or is attached. Gauicol was the substance we used as a reducing agent; it served as an indicator changing color from clear to brown after losing a hydrogen ion reduced it. In this experiment we found the optimal pH level and ceasing point by analyzing the substance’s saturation point. A saturation point is the point where a substance cannot take in more of another substance in a solution. How does changing the pH influence the reaction rate of peroxidase? For our null hypothesis we predicted there would not be an effect when pH is added. Our alternative hypothesis predicted that there would be an effect if pH were added.

Materials and Methods
In this experiment there was a variety of needed materials. Protective equipment consisted of lab goggles, latex gloves, and a fume hood. We also needed micropipettes to measure and transfer our solutes, Para film to protect our solution, and twelve cuvettes to hold our assay solutions. The most important materials were the assay solutes including: peroxidase, guaiacol, hydrogen peroxide, and our pH levels 2,5,7 and 10. Peroxidase is the enzyme solution we used; it consisted of 5 grams of turnip blended in 500 milliliters of water and was filtered. Guaiacol acted as the indicator. Hydrogen peroxide was the substrate solution in the experiment. The Spectrophotometer determined the saturation level of the assay solutions. To prepare the assay solutions we began by labeling the 12 cuvettes. The first four are labeled A, B, C and D and acted as the blank cuvettes. The last eight were our experimentals numbered 1 through 8. When creating the blank cuvettes the pH buffers were added first. Each cuvette contained 2mL of different pH levels either 2,5,7, or 10. Following the pH levels, 1 mL of peroxidase was added. Lastly, since hydrogen peroxide is light sensitive the 0.1 ML of H202 was added to the blank cuvettes individually before placed in the spectrophotometer. There was no Guaiacol added in the blank cuvettes. Before testing the experimental cuvette, the...
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