In the Laboratory
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Deuterium Exchange in Ethyl Acetoacetate: An Undergraduate GC–MS Experiment C. D. Heinson, J. M. Williams, W. N. Tinnerman, and T. B. Malloy* Department of Chemistry, University of St. Thomas, Houston, TX 77006; *
[email protected] Gas chromatography–mass spectrometry (GC–MS) has become available in the undergraduate laboratory in the past 25 years with the advent of less-expensive benchtop instruments. This has been accompanied by an increase in articles on GC–MS in this Journal. Of over 50 articles in this Journal in the past quarter century, over 30 have appeared in the last 5 years. Examples range from illustrating isotope ratios (1–4), elucidating molecular structure (5–8), and various quantitative analyses (9–14) including environmental and health and safety applications. Some involved the use of stable isotopes for structure elucidation and the investigation of fragmentation patterns in mass spectrometry (15, 16). In recent years, student researchers and faculty at St. Thomas have developed or adapted several GC–MS experiments that have been integrated into the laboratory courses. These have been both qualitative and quantitative. This has included an experiment using deuterium exchange to demonstrate the use of base catalysis and keto–enol tautomerism in carbonyl-containing compounds similar to that suggested in ref 15. One particular molecule that did not immediately lend itself for inclusion was ethyl acetoacetate. Acidity of the methylene hydrogens of the acetoacetate moiety is exploited in the acetoacetic ester synthesis of alkyl-substituted acetones (e.g., refs 17, 18). These readily exchange under mild conditions. More challenging is to exchange all five enolizable hydrogens. Use of increasingly severe reaction conditions leads to side reactions, such as deuterolysis of the ester portion if D2O is used, as shown in Scheme I. Decarboxylation of the β-keto acid formed may also occur. One solution is to use ethanol O-d as the source of deuterium and KOD for the base with moderate heat for the exchange reaction. The role of ethanol O-d in nullifying the deuterolysis may be demonstrated by determining that transesterification of methyl acetoacetate to the ethyl ester occurs as well as deuterium exchange of the five acetoacetate hydrogens (Scheme II).
O H3C
O
ODⴚ
O
CH2
O
D2O
H3C
O O
CD2
heat
O H3C
O OD
CD2
+
OD
Scheme I. Side reaction leading to loss of product.
A O H3C
O
O
C2H5OD
O
CH2
KOD heat
D3C
O O
CD2
B O H3C
O O
CH2
O
C2H5OD KOD heat
D3C
O CD2
O
Scheme II. (A) Exchange of five protons and (B) transesterification of methyl acetoactate to yield the deuterated product.
Experimental Procedure A number of experimental conditions may be used to illustrate the basic concepts involved. Comparison of the mass spectra of ethyl acetoacetate deuterated to varying degrees to the mass spectrum of the undeuterated species yields insight into the degree of exchange and the identity of the fragment ions (Figure 1A–D). Deuterium exchange of ethyl acetoacetate directly with D2O or C2H5OD to produce the d2 derivative proceeds readily at room temperature without the presence of base (Figure 1B). Including base and heating in D2O for extended periods, causes the product to be lost. The mass spectra in Figures 1C and D result when approximately 5 µL of ethyl acetoacetate and 5 µL of 40% KOD (> 98% D) in D2O were added to 1 mL of ethanol O-d in each of two capped vials. In the former case, Figure 1C, the reaction www.JCE.DivCHED.org
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Figure 1. Mass spectra of ethyl acetoacetate: (A) d0, (B) d2, (C) mixture of isotopomers between d2 and d5, and (D) d5.
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In the Laboratory
O A X 3C
OX
O Y2C
O
CY 2
was allowed to proceed at room temperature, whereas in the latter, Figure 1D, the vial was placed in a heated sand bath at approximately 70 ⬚C. After 20 minutes, the latter sample was cooled and 1 mL of D2O was added to each vial, followed quickly by 2 mL of methylene chloride that had been pretreated (saturated) with D2O. After vigorous mixing followed by phase separation, a portion of the methylene chloride layers were transferred to autosampler vials and analyzed with a Hewlett Packard Model 1800A GCD equipped with a 7673A autosampler. Explicit procedures are given in the Supplemental Material.W
O
X X 2C
O
O
O X 3C
B CY 2
CY2
O
O
O X 3C
O
Hazards
O CY2
OH
H O X 3C
O CY2
CH2
Results and Discussion
O
Scheme III. Mechanism for the rearrangements resulting in additional mass spectra peaks: (A) McLafferty rearrangement and loss of ketene (B)
Loss of ethylene
Table 1. Some of the Major Fragment Ions in the Mass Spectra of Ethyl Acetoacetate-d0, -d2, and -d5 X
Y
Mass/ Da
X3C⫺C(O)
3H
---
43
3D
---
46
X3C⫺C(O)⫺CY2⫺C(O)
3H
2H
85
3H
2D
87
Moiety
Y2C⫽C(OX)⫺O⫺C2H5
X3C⫺C(O)⫺CY2⫺C(O)⫺OH
X3C⫺C(O)⫺CY2⫺C(O)⫺OC2H5
788
3D
2D
90
1H
2H
88
1H
2D
90
1D
2D
91
3H
2H
102
3H
2D
104
3D
2D
107
3H
2H
130
3H
2D
132
3D
2D
135
Journal of Chemical Education
The reagents and solvents should be handled in a wellventilated hood with eye and hand protection. Ethyl and methyl acetoacetate are irritants and combustible. Ethanol O-d is flammable. KOD in D2O is corrosive and methylene chloride is carcinogenic and an embryofetotoxin.
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In Figures 1A and 1B, in addition to the shift of the molecular ion from 130 Da to 132 Da, the fragments at 87 and 90 Da have increased by two from their values of 85 (loss of ethylene) and 88 Da (McLafferty rearrangement and loss of ketene) (19) in the undeuterated molecule, while the acetyl fragment ion remains unchanged at 43 Da. Evidence for the partial exchange of the aceto methyl group may be seen in Figure 1B from the ion at mass 133, too intense to be entirely from 13C and by the CH2DCO+ fragment ion at 44 Da, which is almost nonexistent in the spectrum of the undeuterated molecule. The mass spectrum in Figure 1C shows partial exchange of the aceto methyl group, but is clearly a complex mixture of isotopically substituted species. The mass spectrum in Figure 1D shows successful exchange of all five enolizable hydrogens. The major ions in the mass spectra in Figures 1A, B, and D, are identified in Table 1. The mechanism for the fragments that result from rearrangements are shown in Scheme III. In conclusion, this series of experiments illustrates the significant acidity of the methylene protons in the acetoacetate group, the efficacy of base catalysis, the role of elevated temperature in increasing the reaction rate, and the nullification of the effect of interfering side reactions. Acknowledgment The Welch Foundation is acknowledged for support of this research. W
Supplemental Material
A student handout including explicit experimental instructions and a list of questions and notes for the instructor are available in this issue of JCE Online.
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In the Laboratory
Literature Cited 1. O’Malley, Rebecca M. J. Chem. Educ. 1982, 59, 1073. 2. O’Malley, Rebecca M.; Lin, Hsiao C. J. Chem. Educ. 1999, 76, 1547. 3. Schildcrout, Steven M. J. Chem. Educ. 2000, 77, 501. 4. Blauch, David N.; Schuh, Merlyn D.; Carroll, Felix A. J. Chem. Educ. 2002, 79, 584. 5. Amenta, D. S.; Devore, T. C.; Gallaher, T. N.; Zook, C. M.; Mosbo, J. A. J. Chem. Educ. 1996, 73, 572. 6. Gandler, Joseph R.; Kittredge, Kevin W.; Saunders, Oliver L. J. Chem. Educ. 1995, 72, 855. 7. Sadoski, Robert C.; Shipp, David; Durham, Bill. J. Chem. Educ. 2001, 78, 665. 8. Anam, Kishorekumar T.; Curtis, Michael P.; Irfan, Muhammad J.; Johnson, Michael P.; Royer, Andrew P.; Shahmohammadi, Kianor; Vinod, Thottumkara K. J. Chem. Educ. 2002, 79, 629. 9. Hill, Devon W.; Mcsharry, Brian T.; Trzupek, Larry S. J. Chem. Educ. 1988, 65, 907.
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10. Quach, Dinh T.; Ciszkowski, Nancy A.; Finlayson-Pitts, Barbara J. J. Chem. Educ. 1998, 75, 1595. 11. Fleurat-Lessard, Paul; Pointet, Karine; Renou-Gonnord, Marie-France. J. Chem. Educ. 1999, 76, 962. 12. Nahir, Tal M. J. Chem. Educ. 1999, 76, 1695. 13. O’Hara, Patricia B.; Sanborn, Jon A.; Howard, Meredith. J. Chem. Educ. 1999, 76, 1673. 14. Witter, A. E.; Klinger, D. M.; Fan, X.; Lam, M.; Mathers, D. T.; Mabury, S. A. J. Chem. Educ. 2002, 79, 1257. 15. Rowland, Alex T. J. Chem. Educ. 1995, 72, A160. 16. Zahedkargaran, Hengameh; Smith, Leverett R. J. Chem. Educ. 2001, 78, 1379. 17. McMurry, John. Fundamentals of Organic Chemistry, 5th ed.; Brooks/Cole: Monterey, CA, 2002. 18. Solomons, T. W. Graham; Fryhle, Craig B.Organic Chemistry, 8th ed.; John Wiley & Sons: New York, 2003. 19. McLafferty, Fred W.; Turecek, Frantisek. Interpretation of Mass Spectra, 4th ed.; University Science Books: Sausalito, CA, 1993.
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