Tuesday, November 5, 2013

Enzyme Catalysis Lab

Lab 2A:

Purpose:    The purpose of lab 2a was to see the effects of temperature on enzymatic activity and to see the presents of catalase in living tissue. We know that enzymes may act differently when it isn't performing at its optimal pH and that enzymatic activity can be amplified by the presents of a substrate. This experiment demonstrates these things. The independent variable in this experiment was the substances being added ( the temperature change or the potato/liver). The dependent variable is the product that comes from the reaction.

Introduction:      As I mentioned, enzymes activity can be affected by a few different things. These things are temperature, pH, and chemicals that influence the enzyme. Enzymes function at an optimal pH and temperature. Once the enzyme leaves that optimal value a few things could happen. In the case of temperature, once the enzyme goes past it's optimal temperature it denatures, meaning that it loses its shape due to disruption of bonds. Another way that the enzyme may be effected positively is by something we call cooperativity. When a substrate binds to the active site of an enzyme in its active form it increases the chances of other substrates being able to bind and produces more product. 

Methods:     At the start of the experiment we wanted to observe the basic reaction that we would be study. To do so, we put 10 mL of a 1.5 % solution of hydrogen peroxide into a beaker. We then added 1 mL of the catalase solution. After completing this and seeing the reaction, we then began the test of temperature.  First, we took 5 mL of the catalase and put it in a test tube. We then placed the test tube in a boiling water bath for 5 minutes. After the catalase had cooled, we out 10 mL of the 1.5% solution in a beaker. Then we added the cooled catalase and observed the reaction. Our last test was to demonstrate the presence of catalase in living tissue. What we did to show that was, we first cut a 1 cm3 piece of potato and once again set up our 10 mL of the 1.5% solution in a beaker. 
We then placed the potato in and observed the reaction one last time. 

Graphs and Charts:    Since the lab was not actually performed but us, leaving us with no data or charts to fill in, here is a graph from our book. This graph shows how the temperature affects the enzymes.


Discussion:      What we noticed in our first part of the experiment were that bubbles appeared. These bubbles were O2 (also our product). These bubbles are formed because of the H2O2 being broken down. This was just a normal reaction and it did what it was supposed to do. If the enzyme and the substrate are operating in its optimal conditions, then the reaction should occur and eventually slow to a halt. We then wanted to see the affects of temperature. What we noticed was that no bubbles were formed during this reaction. That's because the catalase was boiled therefore leaving its optimal temperature and because of that it denatured. What it means to denature is that it's "a process in which a protein loses its native shape due to the disruption of weak chemical bonds and interactions, thereby becoming biologically inactive". All that is saying is that when the heat is raised too high, the bonds are broken and the enzyme can't perform work anymore. We didn't see the bubbles because the catalase had been boiled to a point where it changed the active site and the substrate couldn't bond to the enzyme and then couldn't produce the O2. Then in our final test, we wanted to see if catalase was present in living tissue. When we placed the potato in, the reaction once again occurred. We saw this because there was in fact catalase in the potato. A question posed by the lab was, "what if the potato had been boiled before we placed it in the beaker?". The answer to that is, that we wouldn't see a reaction. The heat would have also denatured the catalase in the potato producing no reaction. 

Conclusion:    As this lab demonstrates, enzymes are very specific molecules that can't always work in whatever they are thrown into. What makes this all relevant to us is that we have enzymes inside us! What happens to us when we have a fever? And our enzymes begin to go past their optimal temperature? This is why people can die of high fevers. Humans enzymes can't function that high either and denature and we die. This is why it's so important that we learn about this things, because they can apply to you and me in our every day life. 

References:  Reece, Jane B. Campbell Biology. San Francisco: Pearson Benjamin Cummings, 2011. Print

Lab 2. Enzyme Catalysis. College Board. 

http://www.biologyjunction.com/ap_sample4lab2.htm


Lab 2B:

Purpose:
The purpose of this experiment was to find an initial base line. There was no independent or dependent variable because this was the control itself this was the dependent variable.

Introduction:
Enzymes are macromolecules that act as a catalyst. They speed up the metabolic reactions by lowering the energy barriers. They speed up the reaction without consuming the reaction itself.

Methods:
We added 1.5% hydrogen peroxide, water and sulfuric acid. When this was combined we titrated it so we could then get a baseline to use for the next experiment.

Data:

Discussion:
With finding the base line we figured out how much hydrogen peroxide is used in the experiment we conducted after. Our baseline ended up being 3.7. 

Conclusion:
This experiment helped the future ones we did because without it we would not have a control for the other parts of lab two.  

References:
Robert B. "Chapter 8." Campbell Biology Ninth Edition. N.p.: n.p., n.d. N. pag. Print.

Lab 2C:

Purpose: the purpose was to determine the rate of spontaneous conversion of hydrogen peroxide to water and oxygen.

Introduction: An enzyme is a chemical that catalyzes, or speeds up, a reaction and is not consumed in the process. A substrate binds to its active site, and the enzyme speeds up an otherwise slow reaction and releases the product. In the case of the enzyme in this experiment, catalase, it speeds up the breakdown of hydrogen peroxide into water and oxygen. This is essential in the human body, because a buildup of hydrogen peroxide is toxic to humans, but catalase converts it into substances usable by the human body. 

Method: In this lab we took 10 mL of hydrogen peroxide that sat overnight, 1mL of water and 10 mL of sulfuric acid. After it was mixed we took 5 mL of the solution and titrated it. The amount of KMnO4 used was 4mL.

Data:


Discussion:    In this lab, we added Potassium permanganate to our 1.5% solution to see the rate of spontaneous conversion from H2O2 to H2O and O2 with an uncatalyzed reaction. What the potassium permanganate does is it shows how much hydrogen peroxide is left in the solution after it had been allowed to sit. A problem we have with our data is that we found a negative number for our amount of hydrogen peroxide that was spontaneously decomposed. That doesn't make sense, because once the reaction has occurred, it can't return to its original form since the oxygen has already escaped into the air. This impossible number is most likely due to an area we had in calculations. We also found that the percent of the hydrogen peroxide that was spontaneously decomposed in 24 hours was only 25%. That number also should have a been higher. This is probably once again due to an error in the set up of the lab or the calculations. 

Conclusion: the rate of conversion of hydrogen peroxide to water and oxygen is about 25% per day uncatalyzed  this data will be helpful to compare to in later reactiins wiht catalase added.

Citations: Campbell Biology Ninth Edition, College Board Enzyme Catalysis Lab.


Lab 2D:

Purpose: the purpose of this experiment is to determine how much hydrogen peroxide can by catalyzed by the enzyme catalase within various windows of time. The independent variable of this experiment was the amount of time the catalase was allowed to react, and the dependent variable was the amount of hydrogen peroxide consumed. 

Introduction: An enzyme is a chemical that catalyzes, or speeds up, a reaction and is not consumed in the process. A substrate binds to its active site, and the enzyme speeds up an otherwise slow reaction and releases the product. In the case of the enzyme in this experiment, catalase, it speeds up the breakdown of hydrogen peroxide into water and oxygen. This is essential in the human body, because a buildup of hydrogen peroxide is toxic to humans, but catalase converts it into substances usable by the human body. 

Methods: First we put 10 mL of 1.5% hydrogen peroxide into a cup. We added one mL of catalase extract and let the substance react for 10 seconds, and repeated the process for increasing amounts of time before stopping the reaction by denaturing the enzyme with sulfuric acid. We titrated potassium permanganate into the solution to determine how much hydrogen peroxide was reacted.

Data:



Graphs and Charts:





Discussion: The most obvious trend in our data is that as the enzyme was allowed to react for more time, we had to titrate less and less potassium permanganate, meaning increasing amounts of hydrogen peroxide were consumed. This is because the more time catalase was allowed to react, the more hydrogen peroxide it was able to break down, because the enzymes had more time to move on to more substrate after breaking one down. This data did fluctuate, most notably at the 60 and 120 second trials, but this is almost certainly an experimental error. We were probably impatient in our titration, and added more than necessary, giving us an inaccurate reading. But, the overall trend was that more hydrogen peroxide was consumed when the catalase was given more time to work, which would make sense, because giving catalase more time gave it more time to break down hydrogen peroxide. 

Conclusion: The more time catalase is given to work on hydrogen peroxide, the more will be broken down. This trend is supported by our data with a couple errors, but comparing our data to that of other groups showed that these data fluctuations were just errors. Overall, we can conclude enzymes can break down more substrate given more time to react. 

References: Campbell Biology Ninth Edition
Lab 2: Enzyme Catalysis, College Board
http://www.princeton.edu/~achaney/tmwiki100k/docs/Catalase.html

1 comment:

  1. Next time use a line of best fit - you would be able to see the curve better at the longer reaction rates.

    ReplyDelete