Introduction

The fundamental building block of biological individuality in all (known) living systems is the protein. The protein is a biomacromolecular structure built from a specific combinatorial linear sequence of at most 20 unique Lego-like amino acids. Back a few billion years ago during the Paleoproterozoic era (2.5 bya - 1.6 bya) proteins were grown at a staggeringly slow rate - not to mention their reaction rates were slow as well. The metabolic action of early anaerobes produced O₂ or unbound oxygen. Although useful for some species, this change apparently killed many organisms right into extinction. As soon as photosynthesis showed up on the block there was a significant amount of free Oxygen (O) in the atmosphere on Earth, so suddenly metabolism kicked into high gear (an estimated 1500% increase). This is known as the Oxygen Catastrophe in the Siderian period.

That was all well and dandy for the most part, except for the fact that, somehow, the nonbacterial metabolic processes that survived continued to produce a deadly byproduct toxic compound, H₂O₂ (hydrogen peroxide). They had been doing that for many millions of years but now the process was significantly increased. The peroxisomes in animal cells are involved in the oxidation of fatty acids for digestion and H₂O₂ is a product of that fatty-acid oxidation (metabolic) process. In turn, the enzyme catalase increases the reaction rate of the natural decomposition of H₂O₂ assisting in the maintained livelihood of the remaining cellular systems despite the increased metabolic reaction rates.

Molecular enzymic action is not stable in all possible environments. Indeed, it is not even assured that catalase can break down the unfortunate byproducts of fatty acid oxidation quickly enough. In this lab experiment, there are times when the environmental conditions that may prohibit the statistically normal functioning of catalase. This experiment gave a broad overview of ways of measuring and changing the reaction rates of enzyme-catalyzed reactions.

A number of beakers are required to perform the experiments, and I counted at a minimum, 7, and that would be with extensive washing of that seventh cup. Roughly 20 mL of catalase is required. A 50 mL beaker (not one of the first seven) would be useful to keep some distilled water nearby. Also, at least 50 mL of KMnO₄ (potassium permanganate), and another 50 mL of H₂O₂ (hydrogen peroxide), and yet again another 50 mL beaker full of H₂SO₄ (sulfuric acid). Use at least one unique tool for measuring and attaining a portion of each chemical compound so as to not mix chemicals (this is drastically important), plus at least one extra for transfer of a concoction that will be explained later—more if one does not wish to clean so very frequently.

In general, the basic procedure involves adding catalase to hydrogen peroxide and then proceeding to stop the reaction at some established moment by the addition of sulfuric acid, which lower the pH level in the beaker. By adding the KMnO₄ it is possible to ascertain the amount of remaining hydrogen peroxide, which tells of how much has been catalyzed with respect to time by the catalase. The hypothesis behind this is that if sulfuric acid is used to stop the catalytic reaction then it is possible to ascertain the reaction rate of catalase relative to the amount of remaining hydrogen peroxide.

The average baseline for the September 7^th experiments was 11.2 mL. The data was collected in terms of 10 seconds, 30 seconds, 60 seconds, 120 seconds, 180 seconds, and 360 seconds. Running the experiments showed that roughly 10 mL, 2 mL, 1 mL, 1 mL, 1 mL, and 1 mL of hydrogen peroxide was decomposed (respectively) whereas the sample set of data suggests that is more closely 2.5 mL, 1.6 mL, 1 mL, 0.3 mL, 0.1 mL, and 1 drop (respective to the times in their given order).

The graph shows the reaction rate of catalase and its substrate hydrogen peroxide with respect to time. The reaction rate decreases with a positive increase in time. It is possible to graph a curve to this set of data, attain a formula, and then find the slope at a given point to determine the reaction rate of catalase in the environments that were used throughout this experiment. The data shows that the rate of reaction is not constant and instead is changing with respect to time. The fact that more and more substrate is being catalyzed implies that there is a decelerating reaction rate (there is more “clutter” and less available substrates to be catalyzed).

The catalysis of hydrogen peroxide decomposition is greater than hydrogen peroxide on its own. The reaction rate is much greater during the first few seconds of reaction and slows down as the number of un-catalyzed substrates increases and “blocks” the paths for the catalase to reach the hydrogen peroxide, neglecting even the increased distance between catalase and one molecule of hydrogen peroxide. Intriguingly, apparently catalase can be extracted from liver by mashing up the large chunks of tissue and thus smashing individual cells to the point of producing the inner liquids of the cell. At first, one might think that a number of other materials are also being expelled from the cells, but, once remembering that the liver was prepackaged, one can tell that catalase is able to survive in whatever the conditions of the packaged chicken liver and not many other proteins and thus why we do not have any other peculiar reactions beyond the catalysis of hydrogen peroxide in the experiment.

There are a number of areas where error could have easily crept into the numbers taken in this experiment. The environmental influences of the room and the greater Kyle-Buda area could easily have changed our results, even in a very tiny sense. The cleanliness of the lab equipment and lab environment itself is clearly an issue. Some preliminary results before the actual data was collected showed the importance of maintaining clean laboratory equipment such that results are closer to what others gather.

The sample data shows that there is a drop in the ability to measure how little is required of the KMnO₄to turn the solution a brownish color and thus suggesting that my values for the 120 second, 180 second, and 360 second trails (in terms of the remaining hydrogen peroxide) is likely incorrect and largely refutable.

4.5, 11, 7.5, 15.5, - and that 15.5 we have to kind of eliminate, since it's so extreme to the other averages. In the first 10 to 30 seconds it's going to be doing “the most damage”. That third number is going to be less because there are less numbers to catalyze.

Look at the above data. Any inferences? Yes, it is inconsistent, what makes it inconsistent? Everybody did it a little differently. Ground to one group may not be so to another. Inconsistencies involve the standards of swishing, amount of time, cleanliness, color recognition, etc.

How much of the substrate (the hydrogen peroxide) was catalyzed by the enzyme? If we take all of the data collectively even with the errors we can come up with some conclusions.

Sample set of data: 2.5, 1.6, 1, .3, .1, 1 drop … and the baseline for the sample was 3. The `3' represents the concentration of the hydrogen peroxide (H₂O₂). Our concentration of peroxide was probably over 1.5%, thus the baseline being so high. If you consider the sample data is a 3 baseline and our data is an 11.2 baseline, then you see, then, it starts to make a little more sense. It's going to use it most at 10 seconds (the sample), and then drop off - this is consistent with Bryan's set of data (10, 2, 1, 1, 1, 1).

The standard curve. We will get our average data with our top 3. Graph the average data, and also graph the sample data - use two separate lines so that you see what it's supposed to look. We will see that the longer the enzyme is affected in the solution, the lower the rate of consumption. There are a number of reasons for this. Giving a high concentration of peroxide and catalase, the more peroxide that is broken down, the less molecules of catalase are engaged, so the rate slows down. At 6 minutes there should have been a very small rate of consumption - everything is already done at that point.

Graph the sample data so that you understand what it should look like. Draw conclusions from the sample data, since that's the “perfect” lab exercise. The AP book (online) has a similar lab like the one we have done so that you can see what they're thinking about this.