Analysis of the Effects of the Head vs the Stem of Asparagus on Enzyme Catalysis of Hydrogen Peroxide

Analysis of the Effects of the Head vs the Stem of Asparagus on Enzyme Catalysis of Hydrogen Peroxide

L. Berndt, L. Buckel, H. Cassriel, and E. McAvenia

AP Biology, Period 1

Miramonte High School, Orinda, CA

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Abstract

This experiment aims to determine what part of asparagus acts as the strongest catalyst in a hydrogen peroxide reaction. We predicted that the head of the asparagus would serve as a better catalyst by creating greater amounts of oxygen when placed in a solution of H2O2. The results of our experiment proved our hypothesis to be false, as the stem of the asparagus acted as a better catalyst to the reaction.

Introduction

Our group, tested the catalyzing ability of different parts of an asparagus stalk. We hypothesized that because asparagus contains very high levels of glutathione peroxidase, an enzyme our bodies use to create antioxidants and the asparagus uses to create “leaves” where photosynthesis takes place, a lot of enzymes are needed in this location to complete the chemical reaction to make ATP.  The stem of the asparagus, which is mainly used for water storage and structure, would contain fewer enzymes in order to complete their duties. Therefore, we believe that the leaf end (now referred to as head) of the asparagus will be a better enzyme catalysis than the sliced end (now referred to as end).

In order to test our hypothesis, we measured the increase in pressure, over a given time period, of a piece of asparagus (head or end) placed in hydrogen peroxide.  With the equation PV = nrt, we were able to calculate the change in the number of moles of gas (O2) by subtracting the final number of moles of the container’s gas from the initial number of moles.

Materials

Material	Quantity
Asparagus (Safeway)	One bundle, about 25 full pieces of asparagus
Hydrogen peroxide (Safeway)	186 mL, 3% concentrate
PASPort Chemistry Sensor (Pasco)	1 volume and temperature gauge
PASPort USB Link (Pasco)	1
Glass chamber (shown in Figure 2)	1, holding 38 mL
Plastic tube	1, holding 5 mL, length = 60.07cm, diameter = .30cm
Glass chamber top (shown in Figure 6)	1 (plastic screw on piece with rubber nozzle; nozzle is cut so glass tube can be placed in hole)
Scalpel	1
Scale	1
Computer	1, with Data Studio software

Figure 1: Hydrogen Peroxide Bottle Figure 2: Glass Chamber

                                                              

Figure 3: Plastic Tube Figure 4: PASPort Chemistry Sensor

                                             

Figure 5: PASPort USB Link

Figure 6: Glass Chamber Top

            

Procedure

1. We went to Safeway and bought one bundle of asparagus.  Once in the laboratory, we cut the asparagus into sections that weighed .98 grams.  These sections were classified as either head pieces (pieces with the plant’s cladodes) or end pieces (stem with one open end).  
1. We cut the pieces not the same size, but the same weight due to the fact that we could not measure the surface area of the head pieces because of the multitude of cladode protrusions.  We now realize that we could have crushed or cut up the pieces to get equal surface areas on each where the reaction could take place. That however, could have killed many of the cells.
2. Along with the weight, we should have measured the volumes of the pieces of asparagus through water displacement.  Because we did not do this, our calculations were affected because we could not subtract the volume of the asparagus from the volume that the gas could occupy.
2. To set up our experiment, we brought a computer, glass chamber, gas chamber top, plastic tube, PASPort Chemistry Sensor, and PASPort USB Link to our lab table (set up shown in Figure 7).  We then connected the sensor and our computer through the USB Link.  Through Data Studio on the computer, we set up a graph and table to record the pressure (in atm) throughout our experiments.  We also recorded the starting temperature of our control test.
1. In this step, we should have set up the computer to record the changes in temperature throughout all of our experiments, instead of just taking a starting temperature.

Figure 7: Lab Set-up (not including computer)

3. To get the initial volume of the glass chamber, glass chamber top, and plastic tube, we first attached the glass chamber top to the plastic tube (shown in Figure 8).  Then, we filled the glass chamber and tube-top apparatus with water from a sink. We then poured the water from each into separate graduated cylinders and measured the amount of water each held. We then added the volumes together to calculate the total volume of the system.

Figure 8: Plastic Tube attached to Glass Chamber Top for Volume Measurements

4. After pouring the water out of the glass chamber and plastic tube (attached to the glass chamber top), we connected our plastic tube to the PASPort Chemistry Sensor.
5. We poured 8 mL of hydrogen peroxide into the glass chamber.
6. We placed a head piece of asparagus in the glass chamber and secured the lid, which was still connected to the plastic tube, and started collecting data through Data Studio on the computer.
1. During this step, we realized that our piece of asparagus was not fully submerged in the hydrogen peroxide.  To fix this, we decided to use 18 mL of hydrogen peroxide for trials 3-7.

FIgure 9: Placing Head Piece into Peroxide Figure 10: Head in Chamber with Peroxide                                        

7. We collected data for 4 minutes.
8. We repeated steps 5 through 7 for a middle piece of asparagus.
1. After noticing the large increase in pressure with the middle piece, we thought this might be the result of an increase in surface area (middle piece had two open ends, the head piece only had one).  To fix this, we decided to use the end pieces of our asparagus, which only had one open end.

Figure 11: Middle in Chamber with Peroxide

9. We repeated our experiment (steps 5-7) four more times with a few changes: 18 mL of hydrogen peroxide instead of 8 mL, and end pieces of the asparagus instead of middle pieces.  These four trials consisted of two with head pieces and two with end pieces.
1. Because our first two trials had problems with both the amount of hydrogen peroxide we used and the pieces of asparagus we used, we decided to disregard the results from these two trials.

Results

Volume of glass chamber = 38 mL

Volume of plastic tube = 5 mL

Total initial volume = 43 mL

Temperature = 19.2 degrees Celsius

Table 1: Change in Pressure

Trial	Amount of Hydrogen Peroxide (mL)	Initial Volume (mL)	Initial Pressure (atm)	Final Pressure (atm)	Change in Pressure (atm)
Control (no asparagus)	18	25	.980	.991	.011
Head 1	18	25	.977	.996	.0194
End 1	18	25	.979	1.056	.077
Head 2	18	25	.978	.994	.016
End 2	18	25	.976	1.041	.065

Table 2: Rate of Moles of Gas Created

Trial	n(of gas created) = (V/rt)*(Pf-Pi) (moles)	Time (min)	Moles created per minute (mol/min)
Control (no asparagus)	1.745 x 10^-4	4	4.364 x 10^-5
Head 1	3.015 x 10^-4	4	7.537 x 10^-5
End 1	12.22 x 10^-4	4	30.54 x 10^-5
Head 2	2.539 x 10^-4	4	6.347 x 10^-5
End 2	10.31 x 10^-4	4	25.78 x 10^-5

Fig. 13

Fig. 14

Fig. 15

Discussion

The data we collected did not support our hypothesis and in fact, proved the exact opposite. We hypothesized that the heads of the asparagus would act as a better catalyst for the H2O2 reaction than the stems would. Our hypothesis was based on the notion that the heads of the asparagus contained enzymes needed for photosynthesis and would serve as strong catalysts for the H2O2 reaction. Conversely, we thought that because the stems function primarily as structure and water storage for the asparagus, there would not be as many enzymes to catalyze the reaction. The data that refutes our hypothesis is shown by the greater number of moles of oxygen produced in the stem trials (1.221 x 10^-3 moles and 1.031 x 10^-3 moles) vs. the head trials (3.015 x 10^-4 moles and 2.539 x 10^-4 moles).

This conclusion is important to society because we consume vast amounts of produce similar to asparagus. Fresh, raw fruits and vegetables contain differing levels of glutathione peroxidase, and the amount may vary depending on the plant's stage of growth. In general, asparagus has been shown to have the highest levels of this non-phenolic antioxidant precursor. Other foods high in glutathione peroxidase include avocados, spinach, tomatoes, apples, carrots, grapefruit and purslane. Cooking the vegetables will remove most of the glutathione peroxidase, and processing of fruits and vegetables also causes significant loss of the compound necessary to make the antioxidant (Live Strong). Essentially, to consume the maximum amount of antioxidants, asparagus and similar foods should be consumed raw and at the bottom of the stock.

In the original trial with only 8 mL of H2O2, the asparagus was not fully submerged in the solution. This could have reduced the effect of the catalyst because a smaller surface area was utilized with part of the asparagus not in direct contact with the H202. In order to correct this, we conducted the subsequent trials using 18 mL of H2O2 to maximize the surface area/volume ratio. As mentioned in the procedure, we did not account for surface area in measuring the amounts of asparagus. This variable was difficult to control because if the asparagus was crush up, many of the cells would have been killed and unable to act as catalysts to the reaction. Another flaw in our use of experiment is that we measured mass as opposed to volume. We should have controlled volume by placing the pieces of asparagus in water and measuring the displacement in mL in order to have the correct units in our calculations. Error could have also been avoided had we dried the inside of the tubes before using them in the reaction. There were still remnants of water on the inside from when we measured their volume and the excess droplets could of had an effect on the data. Another way we could have recorded more accurate data is if we had conducted shorter trials to get a larger sample size. It would have been more efficient to run many 1 minute trials than a few 4 minute trials and we could have collected more data. Another possible source of error is that the control test we conducted on Tuesday was done with a different bottle of H2O2. It may have not had the exact same 3% concentration as the previously used bottle, thus leading to slightly different results.

References

Caligiore, Paul, B. Sc., et al. "Peroxidase levels in food: relevance to colorectal cancer screening." Original Research Communications. Ed. Paul Caligiore, B. Sc. N.p., n.d. Web. 21 Oct. 2013 <http://ajcn.nutrition.org/content/35/6/1487.full.pdf>.

Magoro 1 Magdeline, Beryl Zondagh 2. 2012. A Perspective of South African Foods and Food Habits3 Nutrition 216. E-Campus. Oregon State University, Corvallis, OR 97331,USA.

Wickham, Erica, M.S. "Food Sources of Glutathione." Live Strong. Demand Media, 15 Oct. 2013. Web. 15 Oct. 2013. <http://www.livestrong.com/article/335859-food-sources-of-glutathione/>.