In the Laboratory
Decomposition Kinetics of Hydrogen Peroxide: Novel Lab Experiments Employing Computer Technology
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Dorota A. Abramovitch,* Latrice K. Cunningham, and Mitchell R. Litwer Division of Mathematics and Sciences, Anderson College, Anderson, SC 29621; *
[email protected] The quantitative decomposition of hydrogen peroxide in the presence of catalyst 2H2O2(aq) → 2H2O(l) + O2(g)
(1)
has been used in many general chemistry experiments (1–4) to: (i) generate oxygen gas to study its properties, (ii) study the gas laws, (iii) determine the proportionality constant, R, in the ideal gas equation, (iv) examine the stoichiometry of the decomposition, (v) investigate the role of an enzyme and temperature in this process, and (vi) find the rate and order of the decomposition. The typical procedure involves measuring the volume of oxygen produced in the reaction (eq 1) under constant pressure, usually by water displacement, following with the calculations of a wanted unknown. Using a LabWorks II pressure sensor interfaced with a computer, a novel way to study all of the above (i–vi) has been developed. This reaction is carried out in an airtight vessel and the change in pressure at constant volume is measured. The experiment should help the students understand that there is usually more than one way to design scientific experiments to obtain the desired data and the scientist has to decide which method is more suitable (safer, cheaper, and more accurate). To emphasize this point, our students determined the value of R using both methods: monitoring the change in volume at constant pressure by water displacement (5–7) or using a friction-free barrel syringe1 and, in a separate experiment, measuring the increase in pressure at constant volume using the pressure probe interfaced with a computer (8). The students then calculated the percent error for each method and commented on the safety issues and time involved. The average value of R from this set of experiments was 0.085 atm L mol᎑1 K᎑1, and the percent error ranged from 0.5% to 8.5%. The same process can be adapted to examine the stoichiometry of the reaction, using both the volume and then the pressure as variables. This gives students a better appreciation of the relationship between volume and pressure in the ideal gas equation. LabWorks II software (SCI Technologies; ref 9) and an appropriate sensor allow the collection and monitoring of real-time data. An example of the graph of time versus the pressure change in a reaction vessel as it appears on the monitor screen is shown in Figure 1. The final pressure, as determined from the plateau region of the pressure, accounts for the total number of moles of gas in the vessel: oxygen produced in the reaction and gas (air, water vapor) present before the decomposition started, at atmospheric pressure. The LabWorks II program allows students to actually observe the increase in pressure during the reaction progress, save the data, carry out data analysis, and import the data to Microsoft Excel to do additional calculations. These features aid in the kinetics investigation. The objectives of the kinetics experiments are: (a) to find the decomposition rate of hydrogen peroxide aqueous solution using this same pressure 790
probe, (b) to investigate the influence of the temperature on the reaction rate (at room temperature and at 32 ⬚C), and (c) to reexamine the kinetic order of decomposition. The reaction is carried out at several different concentrations of the substrate, keeping the amount of enzyme (Baker’s yeast) constant, to confirm that the reaction is first order in hydrogen peroxide. Using different concentrations of enzyme, bovine liver catalase (Sigma, Inc.), and keeping the substrate concentration constant, the effect of the catalyst concentration on the reaction rate is determined and the overall order of reaction is found to be second order. This experiment was introduced into the freshman chemistry lab in the middle of the second semester of general chemistry. By this time stoichiometry of solutions, gas laws, and thermochemistry (eq 1 had been used to illustrate these topics) had been covered. The laboratory coincided with the lecture on kinetics. The kinetics studies that involve finding the rate law are too long for one laboratory period and, thus, were spread over two or three lab periods. It is possible to limit this lab to one period if different students collect the data for different concentrations and then share these data to perform the necessary calculations. The experiment is also suitable for a 200-level biochemistry course, with the following modifications: (i) students use their own enzyme preparations from different sources, such as liver or potato, and use the enzyme to determine their activity, and (ii) students examine the effect of temperature on the reaction rate, as well as on the enzyme activity. The follow-up search for other methods of determining all of the above—for example, spectroscopic methods —could further engage the students in examining data collection. Experimental Procedure
Determining the Initial Rates Using the LabWorks II Pressure Probe The catalytic decompositions of hydrogen peroxide were carried out in aqueous solution in a 25 × 150 mm test tube at room temperature and at 32 ⬚C. Two procedures were used. In procedure A, Baker’s yeast was used as the catalyst source (40 mg each time) and the concentration of the aqueous hydrogen peroxide solution (4 mL), obtained through serial dilutions, varied from 0.28 to 1.75% weight/volume. In procedure B, the concentration of aqueous hydrogen peroxide solution (0.60% weight) was kept constant and bovine liver catalase concentration, prepared in 50 mM phosphate buffer, pH 7.0, varied from 32 to 128 units/mL in a total volume of 3 mL. The test tube was connected to the pressure probe interfaced with a PC computer via the LabWorks II interface box by a narrow (3 mm i.d.), 10-cm long rubber tubing. The system was airtight as the reaction went to completion (about 500–600 seconds). The catalytic decomposition was started
Journal of Chemical Education • Vol. 80 No. 7 July 2003 • JChemEd.chem.wisc.edu
In the Laboratory Table 1. Substrate Concentration versus Initial Rate
Pressure / atm
1.3
Initial Concentration of H2O2 /M
Initial Rate /atm s᎑1
0.071
0.0012
0.11
0.0020
0.14
0.0026
0.18
0.0034
0.21
0.0045
0.28
0.0055
0.35
0.0072
1.2
1.1
1.0 0
50
100
150
200
250
300
350
Time / s Figure 1. Pressure plotted as a function of reaction time for the decomposition of 0.42% aqueous solution of hydrogen peroxide in the presence of Baker’s yeast.
NOTE: A constant amount of Baker‘s yeast (40 mg) was used as the catalyst (procedure A, room temperature). Table 2. Enzyme Concentration versus Initial Rate Concentration of Catalase / units mL᎑1
Initial Rate / atm s᎑1
32
0.00198
64
0.00654
0.007
96
0.00823
0.006
128
Initial Rate / (atm s-1)
0.008
0.004
r 2 = .9969
Y
0.003
Predicted Y
0.002
Linear (Y)
0.001 0.000 0.00
0.05
0.10
0.15
0.20
0.25
0.30
0.35
H2O2 Concentration / M
Initial Rate / (atm s-1)
Figure 2. Hydrogen peroxide concentration versus initial rate.
0.012
0.008
r 2 = .9587 Y Predicted Y Linear (Y)
0.004
0.000 0
0.0108
NOTE: Concentration of H2O2 kept constant (procedure B, room temperature).
0.005
20
40
60
80
100
Catalase Concentration / (units
120
140
mL-1)
Figure 3. Catalase concentration versus initial rate.
either by inserting a glass rod coated with yeast into the hydrogen peroxide solution (5), or by mixing the two solutions and immediately stoppering the test tube (procedure B). Simultaneously the LabWorks II program was started, which allowed the progress of the reaction to be monitored on the computer. When the graph plotting the pressure versus time reached a plateau, data collection was terminated (Figure 1). As a cautionary note, some students had trouble maintaining airtight reaction vessels. Students should be warned
that all tubing connections must fit snugly and a student must apply pressure to the stopper (via a thumb pushing on the stopper).
Graphing and Analyzing Data After the data had been collected, the graphs representing the increase in pressure of generated oxygen versus time at different concentrations of hydrogen peroxide were analyzed using the LabWorks II software. The initial rate (slope of the rising part of the graph, see Figure 1) was determined for each initial concentration of hydrogen peroxide. These data were organized in a table (Tables 1 and 2) and imported to Microsoft Excel. Using Excel’s linear regression analysis, the initial rates were regraphed as a function of initial concentration of hydrogen peroxide or catalase concentration. The best-fit lines were generated (Figures 2 and 3) and the correlation coefficients, r2, were determined. Hazards Hydrogen peroxide [CAS #7722-84-1] may cause skin irritation or respiratory and digestive tract irritation, but it does not present a significant health hazard in this experiment because of the low concentrations used. Students are instructed to follow general laboratory safety rules, including the wearing of safety goggles. Data and Results Using the LabWorks II pressure sensor and data analysis software, the initial rates within the range of 0.001–0.008 atms with respect to oxygen were obtained, corresponding to molar concentrations of hydrogen peroxide from 0.07–0.35 M (Table 1). We chose these concentrations of hydrogen peroxide for three reasons: (i) they produced pressures below 2
JChemEd.chem.wisc.edu • Vol. 80 No. 7 July 2003 • Journal of Chemical Education
791
In the Laboratory
atm, within the pressure probe’s linear range, (ii) they limited reaction times to a few minutes, and (iii) higher pressures cause more deviations from the ideal gas properties. Eighteen students (9 pairs) used 0.60% weight/volume aqueous solution of hydrogen peroxide and obtained initial rates ranging from 0.00198 to 0.0108 atms with respect to oxygen for the enzyme concentration from 32 to 128 unitsmL. This H2O2 concentration worked well, minimizing problems owing to pressure loss during a run when a student took his or her finger off the stopper. Linear regression analysis (Figure 2) demonstrated direct proportionality between the concentration of the H2O2 and reaction rate with excellent correlation, r2 = .9969, confirming that the reaction is first order in the substrate. The same analysis (Figure 3) proved the same first order in enzyme concentration. Conclusions Employing computer technology (LabWorks II), general chemistry laboratory experiments were developed that present a simple and dependable method for students to determine the rate of decomposition of hydrogen peroxide and the order of this reaction, using pressure as a variable. Extensions of this procedure, including the study of the activities of catalase enzyme from different sources and the temperature effect on enzyme activity, can be adopted by introductory biochemistry laboratory curricula. The LabWorks II experiments complement the labs described in the literature on the decomposition of hydrogen peroxide, allowing students to monitor pressure of generated oxygen at constant volume and giving them the option of using different variables in data collection. This successfully demonstrates that there is usually more than one way to collect data needed for research. The benefits of the experiment are its versatility and its ability to introduce the students to statistical analysis. Acknowledgments We thank the NSF (DUE-9950817, Instrumentation and Laboratory Improvement) for funding the project and the students who performed these experiments.
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Supplemental Material
A handout containing detailed instruction for students, including samples of calculations and postlab questions, is available in this issue of JCE Online. Note 1. We used the same test tube, connected to a gas tight syringe, equipped with a special friction-free barrel (B-D Brand Multifit Reusable Syringe, 20-mL size, from Fisher) to demonstrate another way of determining the volume of generated oxygen at constant pressure.
Literature Cited 1. Hein, M.; Best, L.; Miner, R. L.; Ritchey, J. M.; Pattison, S.; Arena, S. Introduction to General, Organic, and Biochemistry in the Laboratory, 7th ed., Brooks/Cole Publishing: Pacific Grove, CA, 2001. 2. Nelson, J. H.; Kemp, K. C., Laboratory Experiments, Chemistry: The Central Science, 8th ed.; Prentice-Hall, Inc.: New York, 2000. 3. Tyner, K. Exploring Chemistry in Today’s World; Wm. C. Brown Publishers: Dubuque, Iowa, 1993. 4. Summerlin, L.; Ealy, J. Chemical Demonstrations: A Sourcebook for Teachers, Volume 1; American Chemical Society: Washington DC, 1985. 5. Bedenbaugh, J. H.; Bedenbaugh, A. O.; Heard, T. S. Module Prop 379; Chemical Education Resources, Inc.: Palmyra, PA, 1990. 6. Bedenbaugh, J. H.; Bedenbaugh, A. O.; Heard, T. S. J. Chem. Educ. 1988, 65, 455. 7. Bedenbaugh, J. H.; Bedenbaugh, A. O.; Heard, T. S. J. Chem. Educ. 1989, 66, 679. 8. Abramovitch, D.; Cunningham, L.; Litwer, M. NSF Catalyzed Innovations; 222nd National Meeting of the American Chemical Society, Chicago, IL, Aug 26–30, 2001. 9. LabWorks II Sensors and Software, SCI Technologies, 105 Terry Drive, Suite 120, Newtown, PA 18940.
Journal of Chemical Education • Vol. 80 No. 7 July 2003 • JChemEd.chem.wisc.edu
