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Introduction of Mass Spectrometry in a First-Semester General Chemistry Laboratory Course: Quantification of MTBE or DMSO in Water Mike Solow Department of Chemistry, City College of San Francisco, San Francisco, CA 94112;
[email protected] An early introduction to the theory and practice of mass spectrometry will serve first-year general chemistry students well later in their careers. Currently many general chemistry students do not perform their own mass spectrometry experiments although it is typically described early in first-year general chemistry texts as a technique that can be used to determine the atomic mass of an element (1). As a testament to the technique’s versatility, mass spectrometry can be used to determine abundance of various isotopes of an element, to determine the quantities of drugs or drug metabolites in urine or blood, and to verify the sequence of recombinant proteins (2–5). Mass spectrometry is increasingly employed to analyze samples of significance to environmental and biological studies, two areas that many general chemistry students pursue in depth later in their academic studies (6). New techniques such as MALDI–TOF and electrospray ionization are expanding the use of mass spectrometry into emerging fields such as biotechnology and nanotechnology (7, 8). Many first-year general chemistry students now have access to mass spectrometers as a result of the rapid decline in cost of gas chromatograph– mass spectrometers (GC–MS). While many excellent experiments have been developed for more advanced undergraduate chemistry students (9), the confluence of these developments described above suggests that more laboratory experiments involving hands-on use of mass spectrometry be developed for first-year general chemistry students (10). At City College of San Francisco two related experiments have been developed to introduce first-semester general chemistry students to mass spectrometry. The goals of these experiments are • to supplement the classroom lecture presentation of this technique with a hands-on activity • to prepare students to understand the role of GC–MS in organic structure determination in second-year organic chemistry courses • to help students appreciate the synergetic relationship between the various disciplines of science.
Learning Outcomes In performing this experiment students should learn: (i) how the mass of an atom or molecule is determined, (ii) the effect of the presence of different isotopes on molecular mass, (iii) the role of an internal standard, and (iv) how mass spectrometry is used in answering various scientific questions. Quantification of DMSO or MTBE in Water Using GC–MS Two compounds, DMSO (C2H6SO, dimethyl sulfoxide) and MTBE (C5H12O, methyl tert-butyl ether) have been quantified by our general chemistry students in separate but very similar experiments. In each case, students use an isoto1172
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Table 1. Quantities of Water, DMSO, DMSO-d6, and Unknown Used by Groups of Four Students Vial
Water/ µL
DMSO/ µL
DMSO-d6/ µL
Unknown/ µL
1
990
10
10
0000
2
975
25
10
0000
3
960
40
10
0000
4
000
00
10
1000
pically labeled version of the compound as an internal standard. The experiment using DMSO, rather than MTBE, has been used more extensively owing to the low cost and general ease of sample preparation in this experiment. The experiment involving the quantification of DMSO in water will be described in this section. Students working in small groups (typically four students each) prepare three standards containing known quantities of DMSO and the internal standard, deuterated DMSO, or DMSO-d6 (C2D6SO). As shown in Table 1, the quantity of DMSO in 1.000 mL of standard solution varies from 10 to 40 µL. Ten microliters of DMSO-d6 is added to each of the standard solutions. An unknown solution of DMSO in water is given to each group of students. The students add 10 µL of DMSO-d6 to their unknown. All samples are prepared using micropipettes and GC–MS snap cap vials. Once students have labeled and prepared their samples they place them into a HP G1800C GCD Series II GC–MS sample carousel. In groups of eight, students operate the GC– MS instrument and program the instrument to acquire data using a prescribed method. The method involves split injection of 1 µL of sample to a helium flow rate of 1mL兾min through a HP-5MS Crosslinked 5% Ph Me Siloxane column (30.0-m × 0.25-mm × 0.25-µm film thickness) at 100 ⬚C. The inlet and detector temperatures are 250 ⬚C and 280 ⬚C, respectively. The mass range analyzed is 45–200 m兾z with a solvent delay of one minute. The total time for the analysis is two minutes. Sample acquisition for two groups of students (a total of eight samples) is complete in roughly 30 minutes. Students save their data to a floppy disk and proceed to a computer studio to analyze their data. A single peak is observed in the total ion chromatogram corresponding to the elution of both DMSO and DMSOd6 roughly 1.65 minutes after injection. (Water is not observed as the mass range for the detector under these conditions extends down only to 45 m兾z.) Students then examine the mass spectrum produced at this time and observe molecular ion peaks at both 78 m兾z (DMSO) and 84 m兾z (DMSO-d6). Students next extract the ion chromatograms for each of these species and record the peak area for these
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In the Laboratory
ions. The peak area corresponds to the number of ions of either 78 amu or 84 amu that strike the detector during an analysis. The ratio of the area of the peak generated by the species weighing 78 amu (DMSO) over the area of the peak generated by the species weighing 84 amu (DMSO-d6) is plotted against the known concentration of DMSO for the standards on Microsoft Excel. The standard curves students typically produce are quite good (R 2 = 0.99 or better). Students determine the concentration of the unknown solution of DMSO in water algebraically using the equation for the best-fit line. When students are not preparing samples, acquiring, or analyzing their data they are asked to go to the computer studio and identify Internet resources related to mass spectrometry as well as answer questions about the theory and practical applications of mass spectrometry. The experiment requires two three-hour laboratory periods. As many as 200 students per semester perform this experiment on one GC– MS instrument. Quantification of MTBE in Water, Part of a Guided-Inquiry Activity The experiment involving MTBE is essentially the same as the one involving DMSO described above with several exceptions. The internal standard is MTBE-d3 rather than DMSO-d6. Owing to the more volatile nature of MTBE, both the MTBE and MTBE-d3 are injected directly into rubber septum-capped GC–MS vials using a glass syringe. The molecular ion is not observed for either MTBE or MTBEd3. The base peak (73 m兾z or 76 m兾z) of each species is used to assess the quantity of MTBE in the unknown sample. This experiment has been used as part of a guided-inquiry activity in which students are presented with a hypothetical scenario involving the sudden decline of a waterfowl population. Students propose various types of water contamination as the root source of the numerous bird deaths and then eventually discover MTBE present in the fictitious “lake” water. This discovery is followed by the quantification of the MTBE described above. A discussion of the environmental effects of using MTBE as a gasoline additive (11) flows naturally from this experiment. The Role of Gas Chromatography While students are using a gas chromatograph–mass spectrometer for these experiments, the role of gas chromatography in these experiments is not important and there is generally no discussion of theory of gas chromatography in conjunction with these experiments. This topic is typically discussed during second-year organic chemistry courses. Hazards DMSO and MTBE can cause health problems if inhaled or ingested. Contact with skin should be avoided as it can cause irritation. MTBE is a high fire hazard and should be kept away from flames and sparks. The small quantities of these compounds used here minimize these risks. Conclusion Quantification of a contaminant in water provides students with a tangible application of mass spectrometry. The early introduction of mass spectrometry in a general chemiswww.JCE.DivCHED.org
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try lab lays the groundwork for experiments of greater complexity later in general and organic chemistry courses, such as organic structure determination. The relevance of chemistry to assessing and solving environmental problems is highlighted for students when they perform this experiment. Acknowledgments This project was supported in part by the Division of Undergraduate Education at the National Science Foundation, grant number DUE-9553786, to the San Francisco Bay Area Collaborative for Excellence in Teacher Preparation, known locally as MASTEP (Math and Science Teacher Education Program). The National Science Foundation DUEILI program provided City College of San Francisco with funding for the purchase of the GC–MS and computers used in the data analysis and Web exploration portions of this experiment (DUE-9851317, DUE-9751604). Supplemental Material Instructor notes, four student handouts, stockroom instructions, and GC–MS methods for analysis of DMSO and MTBE are available in this issue of JCE Online. W
Literature Cited 1. For example: Zumdahl, S. S. Chemical Principles, 3rd ed.; Houghton Mifflin Company: Boston, MA, 1998. 2. Morrison, J.; Brockwell, T.; Merren, T.; Fourel, F.; Phillips, A. M. Anal. Chem. 2001, 73, 3570. 3. Ashley, D. L.; Bonin, M. A.; Cardinali, F. L.; McCraw, J. M.; Holler, J. S.; Needham, L. L.; Patterson, D. G., Jr. Anal. Chem. 1992, 64, 102. 4. Zubritsky, E. Anal. Chem. 1998, 70, 733. 5. Tsarbopoulos, A.; Her, G.-R.; Pramanik, B. N.; Trotta, P. P.; Nagabhushan, T. L. Anal. Chem. 1992, 64, 2303. 6. Perkel. J. M, The Scientist 2001, 15, 31. 7. Wang, Y.; Gross, M. L.; Taylor, J.-S. Biochemistry 2001, 40, 11785. 8. (a) Stewart, T.; Zetlmeisl, M.; Leppin, L.; Rodi, C.; McGinniss, M. J. Am. J. Hum. Genet. 2001, 69, 430. (b) Zhang, B.; Liu, H.; Karger, B. L.; Foret, F. Anal. Chem. 1999, 71, 3258. 9. (a) Slawson, C.; Stewart, J.; Potter, R. J. Chem. Educ. 2001, 78, 1533. (b) Sadoski, R. C.; Shipp, D.; Durham, B. J. Chem. Educ. 2001, 78, 665. (c) Bergen H. R., III; Benson, L. M.; Naylor, S. J. Chem. Educ. 2000, 77, 1325. (d) Pelter, M. W.; Macudzinski, R. M. J. Chem. Educ. 1999, 76, 826. (e) Quach, D. T.; Ciszkowski, N. A.; Finalyson-Pitts. B. J. J. Chem. Educ. 1998, 75, 1595. (f ) Kjonaas, R. A.; Soller, J. L.; McCoy, L. A. J. Chem. Educ. 1997, 74, 1104. (g) Rubinson, J. F.; NeyerHilvert, J. J. Chem. Educ. 1997, 74, 1106. (h) Yang, M. J.; Orton, M. L.; Pawliszyn, J. J. Chem. Educ. 1997, 74, 1130. (i) Guisto-Norkus, R.; Gounili, G.; Wisniecki, P.; Hubball, J. A.; Smith, S. R.; Stuart, J. D. J. Chem. Educ. 1996, 73, 1176. (j) Sproch, N.; Begin, K. J.; Morris, R. J. J. Chem. Educ. 1996, 73, A33. (k) Kostecka, K. S.; Palmer, C. F., Jr.; Rabah, A. J. Chem. Educ. 1995, 72, 853. (l) Grandler, J. R.; Kittredge, K. W.; Saunders, O. L. J. Chem. Educ. 1995, 72, 855. 10. (a) Reeves, P. C.; Pamplin, K. L. J. Chem. Educ. 2001, 78, 368. (b) Van Ryswyk, H. J. Chem. Educ. 1997, 74, 842. (c) Amenta, D. S.; DeVore, T. C.; Gallaher, T. N.; Zook, C. M.; Mosbo, J. A. J. Chem. Educ. 1996, 73, 572. 11. Occurernce of MTBE Fact Sheet. http://sd.water.usgs.gov/ nawqa/pubs/factsheet/fs114.95/fact.html (accessed Apr 2004).
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