In the Laboratory edited by
The Microscale Laboratory
R. David Crouch Dickinson College Carlisle, PA 17013-2896
Mosher Amides: Determining the Absolute Stereochemistry of Optically-Active Amines Damian A. Allen, Anthony E. Tomaso, Jr., and Owen P. Priest* Department of Chemistry, Northwestern University, Evanston, IL 60208; *
[email protected] David F. Hindson and Jamie L. Hurlburt Department of Chemistry, Hobart and William Smith Colleges, Geneva, NY 14456
Experimental Overview
Although there are labs available to help students explore the concepts of enantiomers, diastereomers, optical rotation, and chiral resolution, a search of this Journal’s archives has yielded no experiments where undergraduates can determine the absolute stereochemistry of an unknown starting material (1). In fields such as medicinal chemistry, drug discovery, and natural products, to name a few, it is critical to be able to determine the absolute stereochemistry of a molecule. X-ray diffractometry is often not able to assist us because (a) the molecule of interest may not form a uniform crystal and (b) the cost can be prohibitive. The use of chiral reagents for the derivatization of opticallyactive amines and alcohols for the purpose of determining their enantiomeric purity or absolute configuration is a tool used by natural-products and synthetic chemists. Among the techniques used, Mosher’s method is one of the most reliable and one of the most often used. In 1969 Harry Mosher proposed that the analysis of 1H or 19F NMR spectra of methoxy(trifluoromethyl) phenylacetyl (MTPA) derivatives of chiral amines and alcohols would result in the determination of the absolute configuration of the optically-active compound (2). Mosher’s method begins with the derivatization of an optically-active amine with both (R)- and (S)-MTPA-Cl1 to form a pair of diastereomeric amides, which can be distinguished by 1H or 19F NMR spectroscopy.
Herein we describe a simple procedure for derivatizing and analyzing optically-active amines. An example reaction pair is shown in Scheme I. The reactions described are high yielding and easy to carry out. All chemicals used are commercially available and the reaction products are easily and conveniently purified by micro-column chromatography performed in a Pasteur pipet. The experiments can be performed in a regular four-hour laboratory period, although collecting and analyzing the 1H NMR data requires additional time. Students work in teams of two and each team is assigned an optically-active amine, 1, of known structure but unknown stereochemistry.2 One member of the team derivatizes their amine with the (R)-MTPA-Cl, (R)-2, and the other member of the team derivatizes their amine with the (S)-MTPA-Cl, (S)-2. By comparing spectral data for the two diastereomers, (R, R)-3 and (R, S)-3, the students can deduce the structure of their starting amine (Scheme I). Students are told that if their starting amine is in the free-base form, 1.2 equivalents of Hunig’s base (N,Ndiisopropylethylamine) should be used in the reaction.3 On the other hand, if their starting amine is in the HCl-salt form, an extra equivalent of Hunig’s base should be used in the reaction so
O
O NH2
+
CF3
Cl
Hunig's base
Ph OMe
Scheme I. Procedure for derivatizing and analyzing optically-active amines.
Ph (R, R)-3
+
CF3
Cl MeO
(R)-1 (assigned as unknown)
OMe
O
O NH2
698
H
(S)-2
(R)-1 (assigned as unknown)
CF3
N
Ph
(R)-2
Hunig's base
N H MeO
CF3 Ph
(R, S)-3
Journal of Chemical Education • Vol. 85 No. 5 May 2008 • www.JCE.DivCHED.org • © Division of Chemical Education
In the Laboratory
that the free-base is released. After stirring the reagents in methylene chloride at room temperature for 15 minutes, reactions are worked up and amides are purified by micro-flash column chromatography on silica gel in a Pasteur pipet. All of the amides in our library are soluble in CDCl3. Amides are dissolved in CDCl3 and NMR spectra are collected and analyzed. The students assign all peaks in their NMR spectra. After first determining the chemical shifts of any protons that are near the stereocenter in the amide diastereomers, they determine the Δδ values for the shifts and compare their results to models of the compounds. They then deduce the absolute stereochemistry of the amine they were assigned. Hazards Methylene chloride is harmful if swallowed, inhaled, or absorbed through skin; it is a suspected carcinogen. Silica gel is harmful and irritating if inhaled. Hydrochloric acid is corrosive and highly toxic. Hexanes and ethyl acetate are flammable irritants. Standard safety precautions, including the use of safety goggles and gloves, should be followed during the experiment. The reactions and micro-chromatography should be performed in a hood. Discussion In recent years Mosher’s method has been modified and analyzed with molecular mechanics and semiempirical calculations (3–5). Today, Mosher’s method is considered to be a reliable and effective method for analyzing the various proton chemical shift differences in derivatized molecules, thus allowing for the determination of their absolute configuration. The Mosher argument assumes that the most stable conformation of the (S)-amide, 5, is as shown in Figure 1 (6). When analyzing the 1H NMR spectra of pairs of diastereomers, the students will observe differences in chemical shifts for protons located near the molecule’s stereocenter. For example, in Figure
1 the (S)–amide, 5, protons in the L3 portion of the molecule are shifted upfield relative to the protons in the L3 portion of the molecule in the (R)-amide, 6. Organic chemistry students will recognize this to be the result of anisotropic shielding caused by the phenyl ring introduced during the derivatization of the amine (or alcohol). By knowing the absolute stereochemistry of the Mosher’s acid chloride used in the experiment and adhering to Mosher’s model, it is possible to “back deduce” the absolute stereochemistry of the starting amine. As an example, in diastereomers 7 and 8, the proton chemical shift of the benzylic methyl groups are 1.55 and 1.51 ppm, respectively (Scheme II). With respect to the MTPA plane (see Figure 1), in molecule 8 the methyl group in question is syn to the phenyl ring on the opposite end of the molecule. In molecule 7 the methyl group in question is anti to the phenyl ring on the opposite end of the molecule. Therefore, it must be the case that molecule 9 is of the (S)-configuration (as shown) or else the chemical shift differences would be the reverse of those observed, as they are when starting with ent-9, which has the (R)-configuration. Conclusions This lab is appropriate for an undergraduate organic chemistry course. It reinforces the concepts of optical activity, enantiomer versus diastereomer, and shielding versus deshielding in 1H NMR spectroscopy. The lab provides the opportunity for students to learn the importance of working in teams in addition to using 1H NMR spectroscopy for solving structure-elucidation problems. By derivatizing amines of unknown stereochemistry with Mosher’s acid chlorides of known stereochemistry and studying the 1H NMR spectra of the pairs of diastereomeric derivatives, students can determine the absolute stereochemistry of their starting amines. We have taught this lab for a number of years and, in our experience, students have found it to be both challenging and highly rewarding.
E = 1.55
E = 1.51 O
O N H
CF3
+
N
Ph OMe
H MeO
7
L3
L3 H O F3C
NH (S) L2 OMe 5
F3C
NH
Ph
8 (R)- and (S)MTPA
MTPA plane
H O
CF3
OMe (R)
NH2
L2
6
Figure 1. Stereo structures of the diastereomeric amides.
9
Scheme II. Assignment of the benzylic methyl proton shift in the diastereomeric amides, which leads to the configuration of the amine starting reagent.
© Division of Chemical Education • www.JCE.DivCHED.org • Vol. 85 No. 5 May 2008 • Journal of Chemical Education
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In the Laboratory
Acknowledgments We thank the Hewlett Foundation, Northwestern University’s Office of Undergraduate Research, and the Provost’s Office of Hobart and William Smith Colleges for generous support of this project. Notes 1. MTPA-Cl is methoxy(trifluoromethyl)phenylacetyl chloride, otherwise known as Mosher’s acid chloride. 2. For a chart detailing the full library of derivatives we have synthesized and use in our course, 18 in all, see the supplemental information. We feel that the library is sufficiently large enough that the experiment can be run in both small and large lab sections. 3. The Hunig’s base is added to prevent HCl, generated during the reaction, from reacting with the optically-active amine to form the ammonium chloride salt, thus deactivating it.
3. Ohtani, I.; Kusumi, T.; Kashman, Y.; Kakisawa, H. J. Am. Chem. Soc. 1991, 113, 4092–4096. 4. Latypov, S. H.; Seco, J. M.; Quiñoá, E.; Riguera, R. J. Org. Chem. 1995, 60, 1538. 5. Seco, J. M.; Latypov, S. K.; Quinoa, E.; Riguera, R. J. Org. Chem. 1997, 62, 7569–7574. 6. Rieser, M. J.; Hui, Y.; Rupprecht, J. K.; Kozlowski, J. F.; McLaughlin, J. L.; Hanson, P. R.; Zhuang, Z.; Hoye, T. R. J. Am. Chem. Soc. 1992, 114, 10203.
Supporting JCE Online Material
http://www.jce.divched.org/Journal/Issues/2008/May/abs698.html Abstract and keywords Full text (PDF) Links to cited JCE articles Supplement
Literature Cited
Microscale procedure
1. (a) Ball, D. B. J. Chem. Educ. 2006, 83, 101–105. (b) Amburgey-
A list of optically-active amines assigned (9 total), a library of Mosher amides synthesized (18 total)
NMR spectral assignments and copies of 1H NMR spectra for all amines in our library
A list of chemicals with CAS registry numbers
Notes to the instructor
Peters, J. C.; Haynes, L. W. J. Chem. Educ. 2005, 82, 1051–1052. (c) Baar, M. R.; Cerrone-Szakal, A. L. J. Chem. Educ. 2005, 82, 1040–1042. 2. (a) Dale, J. A.; Dull, D. L.; Mosher, H. S. J. Org. Chem. 1969, 34, 2543–2549. (b) Dale, J. A.; Mosher, H. S. J. Amer. Chem. Soc. 1973, 95, 512–519.
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Journal of Chemical Education • Vol. 85 No. 5 May 2008 • www.JCE.DivCHED.org • © Division of Chemical Education
