In the Laboratory
Teaching Experimental Design Using a GC–MS Analysis of Cocaine on Money: A Cross-Disciplinary Laboratory
W
Christopher A. Heimbuck and Nathan W. Bower* Chemistry Department, Colorado College, Colorado Springs, CO 80903; *
[email protected] The goal of problem-based learning is to help students make their own connections with the subject matter. This objective is most easily accomplished when the students have a genuine interest in the material. Forensic analysis of drug money appeals to a wide audience and the lessons encountered are appropriate for undergraduate courses in analytical and environmental chemistry or forensic science (1). Such analyses also offer an opportunity to segue to the social sciences by looking at factors that affect drug use and by initiating discussions of what constitutes proof in a legal versus a scientific sense (2). Moral and ethical issues arise in how we should treat individuals who use drugs. Different countries and cultures approach these issues in various ways. Through discussion and research on the Web, students can gain a more international perspective. Although concentrations of cocaine on U.S., Canadian, and U.K. currencies have been reported (3–5), cocaine levels on currency from other parts of the world are hard to find. There is no standard procedure for the analysis of cocaine on currency, and the role of the currency’s age—degree of wear—is not fully characterized. These uncertainties provide an opportunity for students to develop and use experimental designs to refine and optimize a procedure from the literature. It also allows the students to develop their own questions after the basic principles of chromatography and solid-phase extraction (SPE) have been introduced. Experimental Procedure In order to introduce experimental designs and to develop the students’ analytical skills, an optimization of the procedure is initially conducted for three parameters: extraction method—methanol versus HCl/NH 3 /SPE; time of extraction—one half versus one minute; and GC–MS detection method—total ion chromatography versus extracted ion chromatography.1 Additional sample parameters added to the design include denomination and age of the currency. Percent recoveries of cocaine from solution are measured by doing successive extractions from a single bill, and detection limits of the method are determined from both the uncertainty of the intercept of the calibration curve and from the background levels. The instrumental procedure found in the paper by Negrusz et al. (6) is used with a HP6890/5973 GC–MS for quantitative and qualitative analysis. This method uses a sample inlet of 270 ⬚C and a HP-5MS column (30 m × 0.25 mm × 0.25 µm) with the temperature program: 130 ⬚C for 1 min, ramp at 12 ⬚C/min to 280 ⬚C with He carrier at 2 mL/min. The ion source is kept at 230 ⬚C and the quadrupole at 106 ⬚C. Sample sizes of approximately 2 µL are manually injected, or sample sizes of 2 µL are injected by an auto sampler via a split-splitless capillary inlet system in splitless mode. 1254
Pairs or small groups of students develop short research projects of their own after developing a sample preparation procedure. The students prepare technical papers by working in small groups using appropriate portions of class data for a final write-up (further details are presented in the JCE Online lab documentation).W Students easily conceive a number of independent projects, such as further modifications of the experimental procedure or measuring cocaine levels in currency found in their local environment (e.g., on- or off-campus). The results reported here are from two of these independent projects that developed into socially-interesting discussions. The first project involved the determination of normal cocaine levels on currency so that questions of innocence might be addressed. The second project examined cocaine levels on currencies from around the world, with the students initially hypothesizing higher levels for countries where cocaine originates. Hazards Methanol, heptane, HCl, NH3, and CH2Cl2 are irritants to the skin, eyes, and nose. Methanol and heptane are volatile and flammable. In addition, CH3OH consumption may cause blindness, and CH2Cl2 has been found to cause cancer in animals. The use of gloves and a hood are recommended. Waste should be disposed of according to local environmental regulations. Results and Discussion A fractional factorial experimental design (FEED) measures the effect of more than one parameter (factor) at a time so that the effect of each parameter as well as selected interactions between factors can be determined while minimizing the number of trials required (7). A worked example is presented in JCE Online lab documentation.W Normally there are few if any replicates used with FEEDs, so it is important that the analysis errors are small compared to the differences being measured. Typical GC–MS relative standard deviations (RSDs) for this analysis are 5–10% if care is taken to keep liners clean and septa replaced. Variation within a bill cut in half vertically is much higher (RSD 32%), so using spiked, uncirculated bills for the method development stage may be desirable. Although this within-bill RSD appears high, it is still significantly lower than the differences found between the new and used bills. The data presented here are based on bills cut in half so that each level of a parameter could be tested in the same bill. The optimization experiment indicates that a oneminute extraction was better than a half-minute extraction. A two-minute extraction did not offer much more for the waffle folding we used, but was somewhat better for rolled
Journal of Chemical Education • Vol. 79 No. 10 October 2002 • JChemEd.chem.wisc.edu
In the Laboratory Table 1. Optimization Results (n = 3)
Table 2. Method Detection Limits
Parameters Compared
Area Ratios
Method
CH3OHa vs SPE
0.83 ± 0.24
TIC (background)a
6.2
0.76 ± 0.16
a
0.3
0.5 min vs 1 min Counts EIC vs TIC
EIC (background) b
TIC (regression)
0.18 ± 0.05
XF vs VG bill
0.01 ± 0.01
$1 vs $10 bill
0.67 ± 0.50
EIC (regression)
b
15 4.2
a
Pure standard cocaine spike on an uncirculated bill, 3σ limits. b From pure standard calibration curve using 3σ of the intercept.
a
CH3OH corrected for for its lower concentration.
bills (data not shown). Data in Table 1 suggests that 5 mL of 0.1 M HCl extractant followed by 2 mL of 0.2 M NH3, SPE, and 2 mL of CH3OH eluent is better than 5 mL of methanol alone. The single methanol extract was simpler, but the final solution was only 40% as concentrated relative to the SPE procedure and the background counts were worse.2 Some of the HCl or CH3OH was lost when the bills were removed from the extractant. Combining successive rinses of a bill or weighing the solution can improve the analysis if necessary. Cocaine levels shown here are based on single rinses in order to shorten lab time and reduce tedium. Specialty cartridges for drug separations are available at little extra cost over generic C-18 SPE. They contain mixed bed resins that may give better performance or an additional variable to optimize. However, C-18 is satisfactory for this application and introducing both SPE and organic liquid extraction provides important exposure to sample preparation at little additional cost. The comparison of extracted-ion versus total-ion chromatography (EIC versus TIC) demonstrated that EIC offers a 10-fold improvement in detection limits, particularly in the presence of background compounds that might co-elute (Table 2). Such co-elution was a larger problem with the C18 SPE as a small amount of the solid phase was also eluted. However, cocaine was easily resolved from major peaks by the GC–MS temperature program, and both TIC and EIC were able to detect cocaine even on many new bills. The detection limit methods used in Table 2 underestimate the realistic limits, so more rigorous methods should be introduced in advanced courses (8). The age of a bill was estimated by grading the wear pattern of bills on a nine-point scale from uncirculated to poor, with most bills falling in the very good (VG) to extra fine (XF) range (9). Sleeman et al. attribute most of the cocaine found on bills to indirect transfer such as from contaminated counting machines in banks rather than direct contact with drugs (10). A comparison of data from new and old bills suggests that a given $1 bill passes through a counting machine about once a week, gaining about 10 ± 4 ng of cocaine each time. In this limited study of twelve bills from Colorado Springs, about 25% of the bills showed levels that were significantly above the age-dependent baseline level. Various denominations of U.S. currencies were found to have the following average cocaine levels (n = 5): $1, 7.4 ± 5.1; $5, 11.1 ± 5.5; $10, 12.2 ± 12.3; and $20, 5.0 ± 4.2 µg/bill. These values are not significantly different, but the relative
ng/Bill
newness of the $20 bills (all XF versus VG–VF for the other denominations in age evaluations) confounded this experiment. Using a log (concentration +1) to normalize the data increases the significance found between bills, as the widely varying concentrations are log-normally distributed. The study of world currencies also produced interesting results (see data in lab documentation in JCE Online).W The number of countries represented was small (n = 20), and the number of samples from each country was also small (average n = 2). However, there were enough data to easily disprove the students’ initial hypothesis; the data indicated that the major-user countries clearly had higher levels of cocaine on bills than countries where coca is grown (4.1 versus 0.9 µg/ bill). If a single bill from the Netherlands Antilles is left out of the regression and values from user countries are corrected for age of the bills, cocaine levels correlate significantly with per capita gross domestic product as reported by the U.S. Central Intelligence Agency (CIA) (11). Other information from the CIA about individual countries, such as the CIA’s evaluation of the international impact of drug trafficking through each country, prompted class discussion. The ensuing evaluation of the data and discussion helped students tie together chemical analyses with real-life issues. Additional enthusiasm outside the class was generated when a local television station asked to interview a student who subsequently presented the class findings on the evening news. Acknowledgments We are indebted to the Barnes Trust and the Fairchild Foundation for providing the instrumentation. We also thank Chad Kraus of the Hallenbeck Coin Gallery for help with the numismatic grading and Peter Neifert for loaning us the European currencies. W
Supplemental Material
Instructions for the student, procedural details, and instrumental settings are provided in lab documentation available in this issue of JCE Online. Notes 1. Extracted ion chromatography is a post-run data manipulation that allows the experimenter to view the chromatogram as if only one (or a limited set) of ions had been monitored. True single-
JChemEd.chem.wisc.edu • Vol. 79 No. 10 October 2002 • Journal of Chemical Education
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In the Laboratory ion monitoring is conducted by setting up the mass spectrometer before the analysis so that it collects only that ion, offering significant improvements in the counts obtained. 2. A third alternative, extraction with 2 mL of heptane or CH2Cl2 from 5 mL of 0.1 M HCl, gave the best recovery: see supplementary material.W
Literature Cited 1. Tarr, M. J. Chem. Educ. 2001, 78, 61. 2. Rothchild, R. J. Chem. Educ. 1979, 56, 757. 3. Oyler, J.; Darwin, W. D.; Cone, E. J. J. Anal. Toxicol. 1996, 20, 213. 4. Hudson, J. C. J. Can. Soc. Forensic Sci. 1989, 22, 203.
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5. Sleeman, R.; Burton, F.; Carter, J.; Roberts, D. Analyst 1999, 124, 103. 6. Negrusz, A.; Perry, J. L.; Moore, C. M. J. Forensic Sci. 1998, 43 (3), 626. 7. Strange, R. J. Chem. Educ. 1990, 67, 113. 8. Burdge, J.; MacTaggart, D.;, Farwell, S. J. Chem. Educ. 1999, 76, 434. 9. Krause, C. L.; Lemke, R. F. In Standard Catalog of United States Paper Money, 6th ed.; Wilhite, R. E., ed.; Krause Publications: Iola, WI, 1987; p 2. 10. Sleeman, R.; Burton, F.; Carter, J.; Roberts, D.; Hulmston, P. Anal. Chem. 2000, 72 (11), 397A. 12. Central Intelligence Agency. http://www.cia.gov/cia/publications/factbook/index.html (accessed July 2002).
Journal of Chemical Education • Vol. 79 No. 10 October 2002 • JChemEd.chem.wisc.edu
