LABORATORY EXPERIMENT pubs.acs.org/jchemeduc
The Use of Gas Chromatography and Mass Spectrometry To Introduce General Chemistry Students to Percent Mass and Atomic Mass Calculations Brian W. Pfennig* and Amy K. Schaefer Department of Chemistry, Ursinus College, Collegeville, Pennsylvannia 19426, United States
bS Supporting Information ABSTRACT: A general chemistry laboratory experiment is described that introduces students to instrumental analysis using gas chromatographymass spectrometry (GCMS), while simultaneously reinforcing the concepts of mass percent and the calculation of atomic mass. Working in small groups, students use the GC to separate and quantify the percent composition in a mixture of dichloromethane and chloroform dissolved in toluene by the injection of known quantities of each pure substance dissolved in toluene and the determination of the individual instrumental response factor. The relative abundances of the chlorine-35 and chlorine-37 isotopes can also be determined from several different groupings on the mass spectra of the compounds, allowing for the calculation of the atomic mass of chlorine. This collaborativelearning experiment enhances student understanding through the hands-on use of chemical instrumentation and helps to develop their problem solving skills in the analysis of several different types of experimental data. KEYWORDS: First-Year Undergraduate/General, Laboratory Instruction, Physical Chemistry, Collaborative/Cooperative Learning, Hands-On Learning/Manipulatives, Atomic Properties/Structure, Gas Chromatography, Mass Spectrometry
T
he use of modern chemical instrumentation in the laboratory curriculum during the first year of an undergraduate student’s educational training can not only enhance student understanding of basic chemical concepts, but it can also help to motivate student interest by providing them with a hands-on kinesthetic learning experience. One of the more commonly available and versatile instrumental techniques employed in chemistry, forensics, and environmental science is gas chromatographymass spectrometry (GCMS). As a result, a wide variety of GCMS laboratory experiments have been reported in this Journal over the last several decades; however, most of these projects are geared toward more advanced chemistry courses, such as organic chemistry or instrumental analysis.1 Only a small number of GCMS experiments have been developed that are suitable for use in the general chemistry laboratory curriculum.2 Given the widespread importance of the technique and the availability of reasonably priced GCMS instruments on the market, there is a growing need for well-designed general chemistry projects that utilize the GCMS, so that undergraduate students can be exposed to this instrumentation early in their chemical training. Chemistry is often defined as the study of matter and its interconversions. Many introductory courses therefore cover the material using a bottom-up approach, beginning with a discussion of the atoms and atomic masses and then working their way up to the more macroscopic properties of matter and the stoichiometric relationships involved in chemical transformations.3 This Copyright r 2011 American Chemical Society and Division of Chemical Education, Inc.
particular order of topics provides an excellent opportunity for instructors to introduce the theoretical concepts underlying gas chromatography (the separation of substances in a mixture on the basis of their physical properties) and mass spectrometry (the determination and identification of substances on the basis of their molecular masses and fragmentation patterns, the isotopic distribution of the elements, and the calculation of atomic masses) in the laboratory environment. Accordingly, the goals of this experiment are to: (a) determine the percent mass of each component in a mixture of dichloromethane and chloroform from the retention times and instrumental response factors of each of the pure substances; (b) to determine the natural abundances of the two most common isotopes of chlorine (35Cl and 37Cl) from the mass spectrum of the halogenated compounds; and (c) to calculate the atomic mass of chlorine as it appears on the periodic table based on a weighted average of the masses of these two isotopes. Some other objectives of the project are to give students practical experience with the injection and use of the GCMS, to work collaboratively on their calculations using their experimental data, and to encourage students to think about different ways in which the calculations can be performed. Although a variety of analogous chemical demonstrations and laboratory projects accomplishing similar objectives have been
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peaks. Because the peak having an atomic mass of ∼86 amu corresponds to either CH235Cl37Clþ or CH237Cl35Clþ, the percentage of the total area of this cluster of peaks should be equal to 2xy. This provides a good check on the two former calculations for x and y. Using the natural abundances of each of the two isotopes and their calculated atomic masses from the mass spectrum of dichloromethane, the atomic mass of chlorine can be calculated as a weighted average of the two isotopes. Students are encouraged to identify a different group of peaks in the mass spectrum for CH2Cl2, corresponding to the fragmented ions CH235Clþ and CH237Clþ, to determine the natural abundances of each isotope from this cluster of peaks, and to compare their calculated values with those from the CH2Cl2þ cluster and with the literature values.5 The mass spectrum of chloroform can also be used to determine the natural abundances and weighted average atomic mass of chlorine, and this should be left as an exercise for the students to perform outside of the laboratory.
reported separately in this and other journals,4 this experiment is specifically designed for use in the general chemistry curriculum, it is one of the first experiments of the semester so that students immediately acquire hands-on familiarity with modern chemical instrumentation, and it combines a number of introductory chemical concepts (volume percent, mass percent, isotopes and their natural abundances, atomic masses) in a single, wellintegrated project. Students need to carefully think about the different data obtained in the experiment (retention times and peak areas, mass spectral data and abundances) to arrive at the results. The use of raw data by the students helps to reinforce concepts learned in the lecture setting and to develop strong problem-solving skills.
’ THE APPROACH In this experiment, students use the gas chromatograph to calculate the mass percent of dichloromethane and chloroform in an unknown mixture (where both substances are dissolved in toluene) and to examine various groupings of peaks in the mass spectrum of each of the halogenated compounds to determine the isotopic composition and the atomic mass of chlorine. After a brief introduction to the two experimental techniques, students work collaboratively in small groups to determine the retention times and instrumental response factors for pure samples of dichloromethane and chloroform by injecting a known volume of a 0.1% (v/v) solution of the halogenated compound (dissolved in toluene) into the GC and recording the retention time and area of the relevant peak. The instrumental response factor can then be calculated by the quotient of the integrated area over the volume injected. The students then inject an unknown mixture containing the two compounds (also dissolved in toluene). The component peaks can be identified based on a one-to-one correlation with the retention times of the pure substances. Using the appropriate instrumental response factor for each peak in the gas chromatogram, the corresponding volume of each component in the mixture can then be ascertained. After researching (or experimentally measuring) the densities of dichloromethane and chloroform at room temperature, students can convert each volume into mass and then calculate the mass percentage of the two components in the mixture. The m/z spectrum of each of the pure halogenated substances is recorded using the mass spectrometer. Given that the isotopic composition of carbon is essentially 100% 12C (which by definition has an atomic mass of exactly 12 amu) and that hydrogen is largely 1H (with an atomic mass of 1.007 amu), students are asked to determine the relative abundances of the two major isotopes of chlorine (35Cl and 37Cl) from different groupings of peaks on the mass spectra of the two compounds. For instance, in the mass spectrum of CH2Cl2, there is a grouping of peaks having m/z = 84, 86, and 88 amu, which correspond to the singly ionized species containing CH235Cl35Cl, CH235Cl37Cl, and CH237Cl37Cl, respectively. With a little coaching, students should be able to assign these peaks to the corresponding molecular ions. Using the relative areas of the three peaks, students can then calculate the natural abundances of 35Cl and 37 Cl. For example, the decimal percent of 35Cl isotope (x) is equal to the square root of the area of the peak having m/z = 84 amu divided by the total area under all three of these peaks. Likewise, the decimal percent of 37Cl isotope (y) can be calculated from the square root of the area of the peak having m/z = 88 amu divided by the total area of all three CH2Cl2þ
’ EXPERIMENTAL SECTION Chemicals and Sample Preparation
HPLC-grade dichloromethane and chloroform, as well as reagent-grade toluene, were obtained from Sigma-Aldrich. Each of the pure halogenated substances was diluted in toluene to make a 0.1% (v/v) solution. In each case, a micropipet was used to deliver 100 μL of the pure reagent to a 100 mL volumetric flask, which was then filled to the mark with toluene and mixed several times by inversion. An unknown containing 6.00 mL of dichloromethane and 4.00 mL of chloroform was made using a buret. A small volume, 100 μL, of the unknown mixture was then dissolved in toluene using a 100 mL volumetric flask. Each of the solutions was capped and wrapped with Parafilm. To prevent changes in the concentrations due to evaporation of the volatile components, the solutions were made by the instructor on the same day as the experiment and were stored in the capped volumetric flasks until immediately preceding their injection into the GCMS. Instrumentation
Three research-grade GCMS instruments were available for the laboratory: (i) a Hewlett-Packard G1800A GCD series gas chromatograph with an electron ionization detector, (ii) a Hewlett-Packard 5890 series II gas chromatograph with an HP 5971A mass selective detector, and (iii) a Hewlett-Packard G1800 GCD series gas chromatograph with an electron ionization detector. All three instruments were equipped with a Supelco 28089-U Equity-5 bonded, poly(5% diphenyl, 95% dimethylsiloxane) 30 m, 0.25 mm diameter column as the stationary phase. The helium carrier gas was set at a flow rate of 1.0 mL/min, the inlet and detector temperatures were both set at 150 °C, and the split injection option was employed. The mass range of the mass spectrometer was set to 35:150 (m/z). In each case, the following GC method was employed: the oven temperature was initially held at 35 °C for a period of 2 min, the temperature was then ramped at 10 °C/min to a maximum oven temperature of 55 °C. The total run time was only 4 min in duration, providing ample time for three groups of approximately three students to rotate through each series of injections on the three instruments in a 3-h lab period. Procedure
Because this experiment was performed during the first weeks of the semester, coinciding with the lecture coverage of atomic
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Table 1. Retention Times for Dichloromethane and Chloroform and the Volume Percentages for Each Component in the 60:40 (v/v) Unknown Retention Time/min a
Student Group
CHCl3
CH2Cl2
CHCl3
A
2.35
3.10
62.7
37.3
B
2.15
2.79
65.8
34.2
C
2.40
3.09
58.7
41.3
D
12.16
2.80
65.3
34.7
2.95 (0.17)
63.1 (3.3)
36.9 (3.3)
Ave (σ) a
CH2Cl2
Volume (%)
2.27 (0.13)
General chemistry students using instrument (i). Figure 1. A representative student-generated gas chromatogram for the 60:40 (v/v) unknown mixture of dichloromethane and chloroform using instrument (i).
masses, the students who were not immediately involved with the instrument were either checking into their lockers or collaboratively working on a worksheet (included in the Supporting Information). Each student in the group of three was able to make a single injection into the GCMS. Separate injections of the “pure” samples containing 0.1% (v/v) dichloromethane or chloroform dissolved in toluene were made. In each case, a 1.0 μL injection was employed. While the students were working up the data, the oven temperature was manually raised to 150 °C to bake off the toluene solvent in preparation for the next injection. The retention times and integrated areas (manual integration was employed) and the mass spectrum data (relative abundance and m/z ratios) for the two compounds were recorded in the students’ notebooks. On instrument (ii), the mass spectrum data were also compared with those in the instrument’s software database. A third injection containing the unknown mixture was then performed, recording only the retention times and integrated areas of the two component peaks. The cycle of three injections, which lasted approximately 40 min per group, was then repeated with the next group of students.
Figure 2. Mass spectrum of CH2Cl2 obtained using instrument (i) under the experimental conditions listed in the text.
The average dichloromethane/chloroform ratio calculated by these four groups of students using instrument (i) was 63.1:36.9 (v/v), in good agreement with the actual volume percent of each component in the unknown mixture. Using densities of 1.327 g/mL for dichloromethane and 1.483 g/mL for chloroform,6 the mass percent of dichloromethane was calculated as 60.5%. The element chlorine is particularly well suited for atomic mass calculations because there are principally only two different isotopes and the percent composition of both isotopes is large enough to be readily observed in the mass spectrum of halogenated compounds. A typical mass spectrum for dichloromethane using instrument (i) is shown in Figure 2. Representative data for the relative percentages of 35Cl and 37Cl determined from the percent abundances of the m/z peaks at approximately 84, 86, and 88 amu for four groups of students using instrument (i) are reported in Table 2. Using the experimentally determined relative abundances and masses of each chlorine isotope for this grouping of peaks in the mass spectrum of dichloromethane, the weighted average atomic mass of chlorine was calculated, and these values are also included in the table. The average experimentally determined atomic mass for chlorine is 35.44 amu, in good agreement with the accepted value of 35.45 amu.5 The students are also asked to use MS data from the grouping of mass spectral peaks corresponding to the CH235Clþ and CH237Clþ fragments, having m/z = 49 and 51 amu, respectively, to calculate the relative abundances of the two chlorine isotopes and the atomic mass of chlorine. Using the average data from four
’ HAZARDS Dichloromethane, chloroform, and toluene are eye and skin irritants, and gloves should be worn in the preparation of the samples. In addition, dichloromethane and chloroform are potentially carcinogenic. Excess samples should be disposed of in waste containers according to federal and local regulations. Care should also be exercised in the handling of syringes and with respect to the hot injection port on the gas chromatograph. ’ RESULTS The retention times of the pure samples of dichloromethane and chloroform using instrument (i) for four groups of students are reported in Table 1, along with the data for the volume percent of the unknown mixture. The relatively large standard deviations associated with the data sample reflect the inexperience of first-year students timing the injection with pushing the start button on the GC. Nonetheless, both compounds are fully eluted within the 4-min window of the programmed temperature ramp and with sufficient separation to allow students to separately integrate the areas under peak. A representative gas chromatogram for the unknown mixture containing a 60:40 (v/v) ratio of dichloromethane to chloroform is shown in Figure 1. Using the instrumental response factors determined from the pure samples of compound, the volume percent of each component in the mixture could be calculated. 972
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Table 2. Student-Generated Mass Spectral Data for the Calculation of the Atomic Mass of Chlorine Using Data from the m/z = 84, 86, and 88 amu Cluster of Peaks in the Mass Spectrum of Dichloromethane Student Group
a
35
Cl Mass/amu
35
Cl (%)
37
Cl Mass/amu
37
Cl (%)
Atomic Mass of Cl/amu
A
35.02
75.6
37.02
24.3
35.47
B C
34.94 34.98
75.9 75.9
36.89 37.03
24.1 24.1
35.41 35.47
D
34.90
75.6
36.90
24.4
35.39
Ave (σ) Literaturea
34.96 (0.05) 34.97
75.8 (0.2) 75.78
36.96 (0.08) 36.97
24.2 (0.2) 24.22
35.44 (0.04) 35.45
The literature values are from ref 5.
Table 3. A Comparison of Scores on a Quiz Before and After the Experiment Pretesta,,b
Metric Performance on the quiz (%) Confidence rank (14)
47 2.34
Posttesta,,b 69 2.82
Increase (%) 46 21
a
The background theory for the quiz was presented in the classroom setting before either quiz was administered. Self-reported scores on student confidence in the material are also presented (see text and Supporting Information for further information). b Data were only available for the most recent semester.
are initially daunting, our experience has been that properly coached students can readily grasp the material and extract relevant information from the data set. By having students work in small groups, the project also develops reasoning and problemsolving skills of the students and gets them engaged collaboratively in the calculations. A principal pedagogical advantage of this experiment are that it explores several different aspects of stoichiometric relationships from the calculation of mass percent for the two components in the unknown mixture using the GC to the calculation of the atomic mass of chlorine from different groupings of peaks on the mass spectra of dichloromethane and chloroform. When students obtain essentially the same result for the atomic mass of chlorine using two or more different data sets, it gives them confidence in the validity of their analytical-thinking skills. Furthermore, the experiment can be easily performed in a 3-h laboratory period by rotating groups of students on the GCMS, while other students are independently completing a related worksheet on the calculation of atomic masses using MS data. In addition, very little sample preparation is necessary, and the students only need to make the injections, collect the data, and perform the relevant calculations. We have performed this experiment in our advanced general chemistry lab for three years. A pretest (included in the Supporting Information) involving the calculation of the percent by volume in the mixture from sample GC data, the natural abundances of the two chlorine isotopes from the MS data for dichloromethane, and a calculation of the weighted average of chlorine was given to the students during the prelaboratory lecture. The students had been exposed to the concept of gas chromatography as a separation technique in the “composition of matter” lecture and to the use of mass spectrometry as a means of calculating atomic mass in the “atomic masses” lecture. Students were also asked to rate their understanding of the material on a scale of 1—I do not know what is going on, 2—I understand it when I have guidance, 3—I feel comfortable with the material on
Figure 3. Mass spectrum of CHCl3 obtained using instrument (i) under the experimental conditions listed in the text.
groups of students using instrument (i), the percent 35Cl is 76.7%, the percent 37Cl is 23.3%, and the atomic mass is 35.43 amu, all of which are also in excellent agreement with the literature values.5 By using the data from two different groupings of peaks on the mass spectrum of CH2Cl2, students can gain confidence in the validity of their calculations. A representative mass spectrum of CHCl3 is shown in Figure 3. As an out-of-class exercise, students are expected to assign each of the major peaks to their appropriate ion fragments. For example, the peaks at m/z = 47 and 49 amu correspond with the C35Clþ and C37Clþ fragments; the m/z = 48 and 50 amu are for CH35Clþ and CH37Clþ; the weak grouping of peaks having m/z = 82, 84, and 86 amu are for C35Cl35Clþ, C35Cl37Clþ, and C37Cl37Clþ; and the more intense peaks having m/z = 83, 85, and 87 amu can be assigned to the CH35Cl35Clþ, CH35Cl37Clþ, and CH37Cl37Clþ fragments, respectively. The more advanced students in this course should be able to calculate the relative abundances of each chlorine isotope and the atomic mass of chlorine using data from the different clusters of related peaks in the mass spectrum of chloroform. This is left as an exercise for interested students to pursue outside of the laboratory setting.
’ LEARNING OUTCOMES This general chemistry experiment gives students firsthand access to the GCMS in the undergraduate laboratory curriculum. Because the experiment is performed early in the semester, it can be used to reinforce such introductory general chemistry concepts as the separation of matter (GC) and the calculation of atomic masses (MS), as well as to introduce students early in their academic careers to the use of modern chemical instrumentation. Although some of the data collection and calculations 973
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my own, or 4—I understand the full concept of the material and could teach others. The same test was given during the class immediately following the experiment (with only the numbers in the sample data having been changed). The results of this sample of 16 students are shown in Table 3. Although data were only available for the most recent semester, there was a 46% increase in their ability to perform these types of calculations on the posttest over the pretest, as well as a 21% increase in their confidence level with the material. Taken together with anecdotal comments on end-of-the-semester student evaluations, students seem to benefit from the experiment and routinely comment on their appreciation of the hands-on use of chemical instrumentation in the lab course.
’ ASSOCIATED CONTENT
bS
Supporting Information The laboratory procedure given to students; a worksheet for the students to complete; notes to the instructor for preparation of the analytes and instrumental conditions; a copy of the testing used to evaluate the effectiveness of the experiment on student learning, and safety considerations. This material is available via the Internet at http://pubs.acs.org.
’ AUTHOR INFORMATION Corresponding Author
*E-mail:
[email protected].
’ REFERENCES (1) Some representative examples (and their respective courses) include (a) Hope, W. W.; Johnson, C.; Johnson, L. P. J. Chem. Educ. 2004, 81, 1182 (analytical chemistry). (b) Knupp, G.; Kusch, P.; Neugebauer, M. J. Chem. Educ. 2002, 79, 98 (instrumental analysis). (c) Schultz, E.; Pugh, M. E. J. Chem. Educ. 2001, 78, 944 (biochemistry). (d) Atterholt, C.; Butcher, D. J.; Bacon, J. R.; Kwochka, W. R.; Woosley, R. J. Chem. Educ. 2000, 77, 1550 (environmental chemistry). (e) Bender, J. D.; Catino, A. J., III; Hess, K. R.; Lassman, M. E.; Leber, P. A.; Reinard, M. D.; Strotman, N. A.; Pike, C. S. J. Chem. Educ. 2000, 77, 1466 (organic chemistry). (2) A few recent examples include (a) McKay, S. E.; Lashlee, R. W., III; Petrie, G. A.; Sariah, M. Chem. Educ. 2006, 11, 319. (b) Fong, L. K. J. Chem. Educ. 2004, 81, 103. (c) Solow, M. J. Chem. Educ. 2004, 81, 1172. (3) For example (a) Brown, T. L.; LeMay, H. E., Jr.; Bursten, B. E. Chemistry: The Central Science, 8th ed.; Prentice Hall: Upper Saddle River, NJ, 2000. (b) McMurry, J.; Fay, R. C. Chemistry, 4th ed.; Pearson: Prentice Hall: Upper Saddle River, NJ, 2004. (c) Atkins, P.; Jones, L. Chemical Principles: The Quest for Insight, 4th ed.; W. H. Freeman & Co.: New York, 2008. (4) (a) Reeves, P. C.; Pamplin, K. L. J. Chem. Educ. 2001, 78, 368. (b) Blauch, D. N.; Schuh, M. D.; Carroll, F. A. J. Chem. Educ. 2002, 79, 584. (5) Commission on Atomic Weights and Isotopic Abundances Report for the International Union of Pure and Applied Chemistry. Isotopic Compositions of the Elements. Pure Appl. Chem., 1998, 70, 217. (6) Weast, R. C., ed. CRC Handbook of Chemistry and Physics, 64th ed.; CRC Press: Boca Raton, FL, 1983.
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