Enzyme Catalyst Lab

Purpose:
The purpose of this lab to analyze how enzyme affects the conversion of hydrogen peroxide to water and oxygen gas. In other word, how does the enzyme affect the rate of hydrogen peroxide breaking down into water and oxygen. We wanted to test how much enzyme we need to really start affecting the solution to change. The independent variable was how much of the enzyme solution we put into the hydrogen peroxide solution. The dependent variable was how soon the hydrogen peroxide changed to pink/brown and how much enzyme is needed to do so.

Introduction:

This experiment is all about enzymes and how they react to different situations. So what exactly is an enzyme? An enzyme is a substance produced by a living organism that acts as a catalyst to bring about a specific biochemical reaction. A catalyst is something that affects the rate at which the chemical reaction takes place. In this experiment, we will deal with enzyme-catalyzed reactions which means that the substance to be acted upon, the substrate represented in an equation as S, binds to an active site of the enzyme, represented as E. One advantage of enzyme-catalyzed reactions is that it makes the reaction require less activation energy so that products (P) of the reaction form. This is expressed with the following equation:

E + S -> ES -> E + P

It is important to know that each enzyme is specific for a particular reaction because of the amino acid sequence, which has an effect on its structure. Active sites in an enzyme interact with the substrate so that any substrate that changes the shape or blocks the active site will have a completely different purpose/shape than it did before. How can this happen? Well, salt concentration, pH, temperature, and activations & Inhibitors are four ways that enzyme activity can be affected. Depending on the amount of salt concentration, the proteins in the enzyme will denature and in turn become inactive such as with low concentration. On the other hand, concentrations that are too high will cause blockage. Denaturing of proteins also applies to pH and temperature. When the pH & temperature is too high or too low, it causes enzymes to become inactive, since most enzymes perform optimally at about neutral pH and temperature. Lastly, activations and inhibitors regulate how fast the enzyme acts.

In this lab, the enzyme used was catalase. It takes part in some of the many oxidation reactions that occur in cells, but the primary reaction catalyzed by catalase is decomposing H2O2 to form water and oxygen as shown in the following equation:

2 H2O2 -> 2 H2O + O2 (GAS)

Without catalase, a reaction would still in fact occur, but at an obviously slower rate. 

Methods: 

We started out this experiment by putting 10 ml of H2O2  into 7 separate beakers. Each beaker was labeled with a number ranging from 10 to 360 seconds (depending on how much time we would have to swirl it). Then, we added 1 ml of the catalyst. Now we actually got to "mix" the H2O2 by slightly swirling it. We also set a timer to keep track of how long we needed to mix it. Once the timer hit zero, we quickly added 10 ml of H2SO4. Then we put it under the burette and added one drop at a time of the KMnO4. We waited until the the mixture turned permanently pink/brown . This allowed us to see how well the catalyst and enzymes work and how it varies depending on the amount if time it has been in the H2O2 mixture.

Data:

Graph:

Discussion:

For the lab we filled out a chart with our results. The charts explain how long it took our base line. The intial reading was what the tube was filled at before we used it and the final reading was what it was after the reading. We took the base line and added the enzyme and kept on stirring for 10, 30, 60, 90, 180, and 360 seconds. After the time is down we would check and check how much KMnO4 it takes for it to give it a pinkish color. Our results are valid because we compared our results with the class and our data is very close to other groups. In the graph the blue line shows the H2O2 used and the red line shows the KMnO4 consumed.The longer we waited on the H2O2 the more of it was being used in the chemical process and the longer we waited on the KMnO4, less of it was being released because all the enzymes in the chemical reaction are being used.

Initially, we anticipated that, with more time passing between our addition of catalase into our solution of H2O2, the quicker we would be able to see a presence of color indicating the efficiency of the reacting enzyme that was catalase. This proved to be correct as our data began to show a decrease of used catalase from our burette as we began mixing it with our beakers that had been swirled with catalase for a longer period of time.

For example, the beaker that we added and swirled catalase with for a period a six minutes only required .5 mL of KMnO4 to be added until a pink coloration was seen to indicate a reaction whereas our beaker that was swirled for a shortened period of only 10 seconds required a larger quantity of KMnO4 to be added, a difference of 3 mL to our six minute beaker's .5 mL, in order for a pink color hue was seen in the solution. This showed that the more time that catalase was given with the H2O2 solution, the quicker a visual reaction would occur with KMnO4.

Additionally, our collected data showed an inverse relationship with the amount of KMnO4 consumed versus the amount of H2O2 used. Our data showed that when looking at the amount of KMnO4 used (here we are going to describe the amounts in terms of first, the beakers that had a shorter period of time to mix with catalase on to those who had longer time) began high at 3 mL for our 10 second beaker and then dropped down to a smaller amount of .5 mL used, as seen with our 6 minute beaker. However, the amount of H2O2 that was used in our 10 second beaker, the one which had used 3 mL of KMnO4, only .5 mL of H2O2 was used. This inverse relationship between KMnO4 and H2O2 became more evident as we observed the dropping amount of KMnO4 used in regards to the raising amount of H2O2 used as we increased the the time given for catalase to react with our beakers. The 6 minute beaker of H2O2 ended up using .5 mL of KMnO4 to a higher 3 mL of H2O2 used. These two beakers were our extremes and more clearly showed the contrast in use of the two kinds of solutions used in this lab.

We have to acknowledge that for some of our beakers, the time we spent swirling in catalase might have differed by some few seconds or so by either some means of loss of time or distraction of some kind and that may have altered results. For future tests, we should keep close attention to the time spent swirling the catalase with our various beakers for that specified time the lab calls for.

Conclusion: 

Once all data was accumulated from the previous procedure, the results showed an inverse relationship with the amount of KMnO4 consumed and the amount of H2O2 used. As the time that passed increased, KMnO4 increased from its initial 3mL down to a final .5 mL whereas H2O2 decreased from its initial .5mL up to 3mL. This shows that the increased time spent adding H2O2 lowered the amount of substrate needed to catalyze a reaction when in use of KMnO4. The trials exemplified the very aspect of how a present enzyme of catalase sped up the reaction of our KMnO4 solution. As the time increased where our catalase was mixed in with our solution supported the anticipated result that the presence of color in the initial colorless solution would occur more quickly, which shows that less KMnO4 is need as the amount of H2O2 is used. Because the addition of KMnO4 was done by hand, the precise amount of minimal substance that could be added until a color change was seen could be off by a, minute but still present, amount.