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The effect of pH on the rate of which Catylase breaks down Hydrogen Peroxide

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stephen turban

on 17 September 2012

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Transcript of The effect of pH on the rate of which Catylase breaks down Hydrogen Peroxide

By Stephen Turban The effect of pH on the rate of which Catylase breaks down Hydrogen Peroxide Contributors - Aniqa and Vikrum Head Body Body Body Body Body Body Body Body Body Body Body Body Introduction The Original Most Interesting Man in the World What is the effect of pH levels on the rate of which catalase breaks down hydrogen peroxide? Question Context Hypothesis: When pH is seven the rate catalase breaks down h202 will be the greatest compared to both pH4 and pH 10 We have already observed that Enzymes have “optimal conditions”. We know that if a pH is too low, or too acidic, then there be a lot more hydrogen than the enzyme normally would exist in. This will cause the tertiary structure to be disrupted, and for the enzyme to denature (Campbell)– making it unable to regulate chemical reaction. Likewise the enzyme will denature if conditions are too basic. The question we are asking is “at what pH is optimal conditions for the enzyme catalase?” as well “what levels of pH are least optimal for the enzyme catalase". The implications of the experiment are huge. Catalase is an enzyme essential to life, especially in reproductive reactions. In order to have this enzyme work most optimally we must first know at what pH the enzyme works best. From that knowledge we can then work to maintain that pH levels in our own body (something generally already done by our body) as well as in experiments involving catalase. How does this fit with what is already known? We will combine catalase with hydrogen peroxide under three different pH levels, 4, 7, and 10 and measure the percent oxygen that is released. This will allow us to compare the reaction rate of the catalase in three different pH levels and ultimately let us see which of the environments is most suitable to catalase. Remember: the higher the percent oxygen released/created the faster the catalase is reacting with hydrogen peroxide.
Experimental Set- Up Why we do the experiment: The body holds of thousands of different types of enzymes that regulate the chemical reactions in our body that we need to live. Without the enzymes working at top capacity we would die; chemical reactions would occur too slowly to support homeostasis. As such, it is important we understand under what conditions enzymes work effectively, and perhaps more significantly, under what conditions enzymes work less effectively.

With that in mind, we decided to test the effect of the pH of the environment on the rate of which catalase (the enzyme we chose) breaks down Hydrogen Peroxide. What this truly measures is the effectiveness of the enzyme under different environments. If, for example, under a more acidic environment catalase breaks down more Hydrogen peroxide then at a neutral or basic pH we know that the most congenial environment for catalase is more acidic. Context Materials and Methods Materials (this is per a group, if you want the total materials for all three groups, then multiply all of these materials by three)
15ml of buffers of 4, 7, 10 respectively
45 ml hydrogen peroxide 3%
One graduated cylinder (50ml)
One test tube
One Oxygen Sensor
One Nalgene Bottle
One dropper
One Graphing Calculator Materials - Set up Oxygen Sensor (refer to enzyme lab “setting up oxygen sensors”
- Measure 5ml 3% Hydrogen Peroxide in graudated cylinder and add to test tube
- Measure 5ml of the Buffer pH of 7 and add to test tube (This is the control of pH of 7, later on we will retry the experiment with a pH of 10 and 4. This Buffer level is the IV of the experiment)
- Add 10 drops of Catalase to test tube and shake, add catalase solution into Nalgene bottle and cap and immediately cap with 0xygen sensor.
- Take oxygen measurement after 30 seconds, wait till the oxygen sensor is finished and record reaction rate - the percent oxygen released is the DV for the experiment. How high percent of oxygen is created is a product of how well catalase is reacting(for all specifics involving how to use the calculator/sensor please refer to Measuring Oxygen Production hand out)
- Repeat all steps (except for the oxygen sensor) two more times (for a total of three trials)
- Repeat all steps (except for the oxygen sensor) again, but with buffers of pH of 4 and 10 Procedure Results Over all, the results indicated that, on average, when the pH of the environment was four Catalase broke down Hydrogen peroxide and created oxygen most quickly. If only average is looked at, at 10pH Catalase broke down Hydrogen Peroxide at the second highest rate, and 7pH the worst. However, the median indicates that at 7pH Catalase had a higher catalyzation rate than at 10pH - indicating a pattern of Catalase being more effective the more acidic the environment.
Results Results The most significant results we got, by far, was how much more oxygen we measured was produced at a pH of 4 over a pH of 7 and 10 based upon our class averages. (Around 1000 times more).
As well, our class averages indicated that at a pH of 10 the second highest rate of oxygen was produced.
Looking at my group individually, however, different results are found. The highest pH is still a pH 4. But the oxygen production at 4.708(10^-4) percent oxygen produced per a second is still relatively similar to the production of pH 7 (1.028(10^-4) percent oxygen produced per a second. As well, the pH of 10 is not too far behind at 8.70(10^-5 )percent oxygen produced per a second . This shows a marked contrast between the nearly three fold difference found between pH4 and pH of 7 and 10 in the class averages. Significant Results The original question asked was “at what pH conditions does the enzyme catalase catalyze hydrogen peroxide most effectively”. The results that we found indicate that on average at a pH of 4 Catalase is working best. At a pH of 10 it is working second best, and at a pH of 7 it is working the worst. These results show that different pH levels will influence the speed in which an enzyme can catalyze. How Do These Results Relate to the Original Question The data we collected for the most part did NOT support our hypothesis. The averages in each of the groups supported the notion that catalase had the highest rate of breaking down hydrogen peroxide at pH 4, second highest at pH 7, and third highest at pH 10. This held true for other measurements of central tendency – The medians for example, also indicated that pH 4 was the best environment for catalase breaking down Hydrogen Peroxide. Data vs. Hypothesis - The data we collected ran mostly opposite towards what other researchers have reported. Most reporters outside of classroom have reported optimal conditions of catalase to be at about 7 pH (1). While we found the opposite to be true, both pH of 4 and 10 allowed for higher 02 production thatn pH 7. Data vs. Other Researchers -The results we received were unexpected. The relatively huge numbers of group 2 (especially for pH of 4) undoubtedly skewed the results in favor of showing pH 4 of being heavily more favorable towards catalase. Potential sources of error could be faulty set up of the oxygen sensors. As well as measurement stretched over two errors differences in set-up might have caused variation in the results. Catalase is mostly found in the liver. If the liver is more acidic than basic, than perhaps these results are more reliable than thought. (however, the liver usually stays at a pH around 7 (Maehly)
Unexpected Results Another way to interpret the results is to change the way of finding the point of central tendency. Instead of using the average, which has produced huge differences due to abnormal results from group two, the use of a median might show more useful information.
Class Medians are
pH 4: 3.37E-5
pH 7: 2.32E-5
pH10: 1.685E-5
Reinterpret the Results: First of all, to answer the question raised by our results a more thorough and extensive experiment would need to take place. Though nine trials were used, it is seen clearly that the missteps of one group can strongly impact the results of a class as a whole. As well, more levels of pH would be useful to see what exactly is the best pH for catalase and its break down. Looking at Catalase in it’s natural environment would also be a useful way to study Catalase’s “optimal conditions”. Further Research The results we arrived at fit into the big picture by showing that pH does in fact affect enzyme productivity/ability to speed up chemical reactions. The faster catalase could break hydrogen peroxide into oxygen and water the faster that it could speed up chemical reactions in the bodies of organisms. The results, at first may seem dry, but they demonstate how crucially important it is we know what the optimal pH is for enzymes like catalase. This knowledge allows us to understand better our bodies, and the chemical reactions inside it, as well as enable us to protect the systems that we depend on to stay alive. Big Picture Conclusion In conclusion, our results showed that the enzyme catalase breaks down H202 most effectively at a pH of 4 –better than at a pH of 7 and 10. These results should be read well salted, however, the data given by group two clearly warped the results. It is doubtful that at a pH of 4 an enzyme found in the human body is over a thousand times more productive then at a pH of 7. Nevertheless, while the data collected itself did directly support my hypothesis, there is still a lot that can be taken away from this experiment. Most significantly, enzyme activity is affected by the pH of the environment. If the pH is too high or too low it will denature the enzyme, not allowing it to regulate the chemical reactions necessary for life. Conclusion: References 2. Maehly A, Chance B (1954). "The assay of catalases and peroxidases". Methods Biochem Anal. Methods of Biochemical Analysis 1: 357–424. 1 Campbell A. Neil and (2005) AP Edition Biology. San Fransisco, Pearson Education p.154-155 Oxygen Production of Catalase in percent Oxygen released per a second pH level of the Environment The Effect of different pH in the Environment on the oxygen released in the breakdown of hydrogen peroxide by Catalase 4 7 10
Group 1 Avg. 0.000471 0.000103 0.0000873
Group 2 Avg. 0.0688 0.0000127 0.004203
Group 3 Avg. 0.0000126 0.0000117 0.0000066
Class Avg. 0.0203 0.0000424 0.000499 Data Table Graph The most important safety concern for this experiment was that the buffers remained separate.This is so that the buffers do not mix and create a chemical reaction. As well, it was important not to shake too violently when “gently shaking” the catalase and the buffer in the same test tube. Safety Concerns
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