Period5.B

How do different amounts of hydrogen peroxide effect its reaction with liver? By: Maddie Bertschmann, Emma McWilliams, Megan Fujiyoshi, and Olivia Turner ** Introduction
 * Enzymatic Activity Lab

The purpose of this lab is to observe and record how much oxygen is released in the chemical reaction between liver and hydrogen peroxide. Catalase is an enzyme (a type of protein) that is found in the liver. The liver helps to break down toxins in the body. The purpose of the enzyme, catalase, is to accelerate a chemical reaction, and it also breaks down hydrogen peroxide. In final, oxygen and water are released. With oxygen and water as harmless byproducts, the liver is able to successfully rid the body of the harmful toxin (hydrogen peroxide). Because it is known that oxygen is in fact released, the amount of oxygen released will be measured using a gas pressure sensor in the experiment. By using different amounts of the hydrogen peroxide, we will measure gas pressure to see if more oxygen production is a result of a greater amount of hydrogen peroxide.

Materials  
 * 1-125 ml flask
 * 1 piece of liver at least 5 cm3 (about 10 grams)
 * 1 knife
 * 8-1 mL pipettes
 * 1 gas pressure sensor
 * 1 LabPro
 * 1 computer
 * 1 pair of tongs
 * At least 20 ml of hydrogen peroxide
 * 1 pair of goggles for each group member (4 total)
 * 1 pair of gloves for each group member (4 total)
 * Procedure**
 * 1) Set up gas pressure sensor probe by plugging in the gas pressure sensor to channel 1 on the LabPro.
 * 2) Prepared the computer for data collection by opening the Experiment 12B folder from the Biology with Computers folder of Logger Pro.
 * 3) <span style="color: black; font-family: 'Verdana','sans-serif'; font-size: 11pt;">Connected the plastic tubing to the valve on the gas pressure sensor.
 * 4) <span style="color: black; font-family: 'Verdana','sans-serif'; font-size: 11pt;">Inserted the single-holed rubber stopper into the flask for an airtight fit. At this time, tubing should not be connected to the stopper.
 * 5) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Cut five 1 cm3 pieces of liver (about 2 grams each)
 * 6) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Using the pair of tongs, placed 1 piece of liver in a 125 ml flask.
 * 7) <span style="color: black; font-family: 'Verdana','sans-serif'; font-size: 11pt;">Immediately connected the plastic tubing to the connector in the rubber stopper.
 * 8) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Observed the gas pressure in the flask for 5 minutes (this trial is the control).
 * 9) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Printed the data generated by the gas pressure sensor and the LabPro.
 * 10) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Emptied and thoroughly cleaned the flask.
 * 11) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Using the pair of tongs, placed 1 piece of liver in a 125 ml flask.
 * 12) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Measured 2 ml of hydrogen peroxide with the micropipette and a sterile tip.
 * 13) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Added the hydrogen peroxide to the 125 ml flask.
 * 14) <span style="color: black; font-family: 'Verdana','sans-serif'; font-size: 11pt;">Immediately connected the plastic tubing to the connector in the rubber stopper.
 * 15) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Observed the gas pressure in the flask for 5 minutes.
 * 16) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Printed the data generated by the gas pressure sensor and the LabPro.
 * 17) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Emptied and thoroughly cleaned the flask.
 * 18) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Repeated steps 12-18 with 4 ml, 6ml, and 8ml of hydrogen peroxide.
 * 19) <span style="font-family: 'Verdana','sans-serif'; font-size: 11pt;">Compared the results of the gas pressure produced by each reaction.

The independent variable is the amount of hydrogen peroxide used in the reaction. || The dependent variable is the amount of gas pressure caused by the release of oxygen from each reaction. ||
 * **<span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">Independent Variable ** || <span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">
 * **<span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">Dependent Variable(s) ** || <span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">
 * **<span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">Controlled Variables ** || <span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">The controlled variables include the amount of liver used, the type of liver, the temperature, the pH, and the gas pressure sensor. ||
 * **<span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">How will you change your independent variable? (be specific) ** || <span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">We will start by creating a control using 0 ml of hydrogen peroxide. In each of the following experiments we will increase the amount of hydrogen peroxide by 2 ml. We will conduct five experiments total, with 0ml, 2ml, 4ml, 6ml, and 8ml of hydrogen peroxide. ||
 * **<span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">How will you measure your dependent variable? ** || <span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">We will measure the release of oxygen from each reaction with a gas pressure sensor. ||
 * **<span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">How will you set up a control? ** || <span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">We will set up a control by doing an experiment without using any hydrogen peroxide. ||
 * **<span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">Hypothesis: ** || <span style="font-family: 'Verdana','sans-serif'; font-size: 10pt;">If there is more hydrogen peroxide added to the flask containing the liver, then the released oxygen will have a greater pressure than if the flask containing the liver has less hydrogen peroxide. ||
 * Hydrogen Peroxide Reaction Data Table **
 * ** Amount of Hydrogen Peroxide (ml) ** || ** Original Air Pressure (kPa) ** || ** Air Pressure After Five Minutes (kPa) ** ||
 * 0 || 100 || 100 ||
 * 2 || 103 || 105 ||
 * 4 || 105 || 104 ||
 * 6 || 108 || 128 ||
 * 8 || 116 || 162 ||


 * Analysis Questions****

1. What do your results tell you about how your variable affected catalase activity? Include specific data from your experiment to help you answer this question. ** The results show that the amount of hydrogen peroxide will induce changes in the gas released from the experiment. As every 2 mL of hydrogen peroxide were added, the graph of the pressure would steepen. The gas pressure which 6 mL of hydrogen peroxide yielded was measured to be about 125 kPa after about one minute of testing. After one minute of testing the pressure from 8 mL of hydrogen peroxide, the gas pressure was measured to be about 160 kPa. This shows how adding a mere 2 mL of the fluid results in a 35 kPa increase. **

2. What were some sources of error and how might they have affected your data? What changes would you make to your procedure –or—what changes did your peer editor suggest that you make? ** Though originally believed to be one of the more simple experiments, this test extracted the consequences of carelessness. Some sources of error may lie in the amounts of hydrogen peroxide added, and at what time the gas pressure started to be monitored. It was difficult to measure out the specified amount of hydrogen peroxide in pipettes, and reflecting back on the decision, it was probably a better option to use a graduated cylinder. Due to the fact that it is hard to hold more than one 2 mL pipette per hand, for certain tests the hydrogen peroxide was not all added at exactly the same time. This results in the gas pressure not being measured from the exact start of the first reaction. It is actually measured from “0 min/sec” after the last bit of fluid is added. In regards to the experiment, I don’t believe much else could have gone wrong. To correct this minor decisional error, the procedure should clarify that the experiment would yield more accurate results if the hydrogen peroxide would be measured with a graduated cylinder. **

3. Catalase is located in a cell organelle called the peroxisome. What do you think is the function of the peroxisome? Explain your reasoning. ** The peroxisome “digests” hydrogen into oxygen, and then releases hydrogen peroxide as a byproduct. The peroxisomes are also capable of turning hydrogen peroxide into water through the uses of enzymes. The peroxisome is also able to break down break down toxic materials in the cell, like lysosomes. Their name indicates that they are like lysoSOMEs, and are able to create and break down hydrogen PEROXIde.

If a person takes ecstasy at a party, they will have an increase in energy in their body caused by the decreased release of gas. More importantly, the ecstasy prevents the liver from absorbing and processing the toxins at a sufficient rate. The drug does that in any situation. Adding a warm environment, such as a party, the effect of ecstasy increases; meaning that the liver is starting to considerably worsen at the job unique to itself: oxidize the alcohol in a person’s system at a reasonable rate. The alcohol will not be metabolized, which can cause serious damage such as liver disease, cancer, and malnutrition. Malnutrition has less to do with the liver than the actual drug. The euphoria caused by ecstasy suppresses any appetite a person may have. This could ultimately cause either eating disorders, or malnutrition. **
 * 4. Researchers studying the effects of ecstasy (methylenedioxymethamphetamine or MDMA) on mice, found that ecstasy decreased catalase activity in the liver. They also noticed that ecstasy’s effects on catalase were increased as the temperature of the mouse’s environment increased. Based on this information, can you draw any conclusions about what might be going on inside a person’s body when he/she takes ecstasy at a party? What risks are involved? **

5. Catalase is used commercially whenever hydrogen peroxide is used as a germicide. For example, it breaks down the H2O2 that was used to pasteurize milk prior to cheese-making (Chu //et al//. 1975). **

In the factory, it would be best to keep the temperature stable. If would also be important to add hydrogen peroxide if necessary, because this will create more of a reaction within the peroxisomes and their catalases. More reaction results in more gas, which is found when the amount of H2O2 is increased. It would be important that the catalase temperature never rose above a certain temperature, to make sure that it would not denature. Essentially cooking the catalase, high temperatures take away any special or required assets the catalase needs to perform.
 * a.** **As a cheese-maker, what conditions would you want to have in your cheese factory to ensure that catalase was working best? (use the class findings to help you answer this question)**
 * b.** **Why would you need to make sure that the temperature of the catalase never went above a certain temperature?**

During this experiment, we tested the effects of different amounts of hydrogen peroxide on liver. Liver contains the enzyme catalase. Catalase catalyzes the reaction which breaks down hydrogen peroxide into water and oxygen. Our hypothesis for the experiment was correct; as more hydrogen peroxide is added to the flask with the liver, the resulting oxygen pressure increases. In the control trial, with liver but no hydrogen peroxide, the pressure began at 100 kPa, and after five minutes, the pressure was still 100 kPa. In the trial with 2 ml of hydrogen peroxide, the pressure started at approximately 103 kPa, and after 1 minute, it had risen to 105 kPa. The pressure remained at 105 kPa for the remaining four minutes. In the trial with 4 ml of hydrogen peroxide, the pressure began at 105 kPa. There was some fluctuation in the pressure throughout the five minutes, and after 5 minutes, the pressure was 104 kPa. In the trial with 6 ml of hydrogen peroxide, the pressure began at approximately 108 kPa, there was significant fluctuation in the pressure, and the pressure after 5 minutes was 128 kPa. In the trial with 8 ml of hydrogen peroxide, the pressure started at approximately 116 kPa, increased to 162 kPa in the first minute, and remained constant for the remaining four minutes. Although there are considerable fluctuations in the data, there is a definite pattern of an increase in air pressure as the amount of hydrogen peroxide increases.
 * Conclusion **

The fluctuations in the data could be due to a variety of sources of error. The largest source of error was most likely in the air pressure measurement. As the air pressure increased, it became more difficult to hold the stopper in place. We physically held the stopper in place at the top of the flask, but pushing down on the stopper could have increased the air pressure. In addition, if we did not hold it down with consistent pressure, this could be the cause of the slight fluctuations in the graphs. Also, if the stoppers did not completely block the top of the flask, or there was a gap between the plastic tubing and the valve on the gas pressure sensor, the air pressure could have decreased. Another possible source of error could be the measurement of the hydrogen peroxide. Although we intended to use micropipettes to ensure accurate measurements, they were not available, and instead we used regular pipettes, which are less accurate. Additionally, because we wanted to start measuring the pressure immediately, we measured the hydrogen peroxide quickly, and the measurements could be slightly inaccurate.

Catalase is found in nearly all organisms that utilize oxygen. Its main purpose is to prevent the accumulation of toxic levels of hydrogen peroxide. Hydrogen peroxide is a by-product of many metabolic processes. Catalase can stimulate the breakdown of hydrogen peroxide into oxygen and water, therefore eliminating the toxic substance. Our conclusion, that a high amount of hydrogen peroxide results in a high oxygen pressure, is very logical. When the body is performing many metabolic reactions, a large amount of hydrogen peroxide is released. Because it is dangerous for the hydrogen peroxide to accumulate, catalase must catalyze the reaction to break down hydrogen peroxide. The by-products of this reaction, oxygen and water, will be produced in higher quantities when there is a large amount of hydrogen peroxide in the body. Therefore, it makes sense that when we added a lot of hydrogen peroxide to the liver, the oxygen release, and consequently, the oxygen pressure, increased. (The pressure increases because the amount of oxygen is increasing, but the volume of the flask remains constant). This reaction occurs in humans, as well as in many other organisms.