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Laboratories and Demonstrations

Reductive Amination
of Pyruvate Esters: A
Microscale Synthesis
of N-Benzylalanine
Esters
R. DAVID CROUCH*, MICHAEL S. HOLDEN*, AND TOM M.
WEAVER
Dickinson College
Carlisle, PA 17013-2896
[email protected]; [email protected]

T
he reductive amination of pyruvate esters using
When students benzylamine and sodium triacetoxyborohydride is
work in pairs, described. Students isolate the N-benzylalanine ester
and determine its structure using NMR and IR
structure spectroscopy. Computational methods using CAChe provide
determination, information on reactive sites in the pyruvate substrate and
reaction intermediates and allow for insight into the reaction
though mechanism. This experiment is safer than the more traditional
reductive amination techniques, and it is appropriate for the
challenging, is
introductory organic laboratory.
within the reach
of introductory Reductive amination of aldehydes and ketones is an excellent
method for the synthesis of amines and usually receives some
organic students. coverage in organic chemistry textbooks [1–9]. However, a
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FIGURE 1. REDUCTIVE ANIMATION WITH NaBH(OAc)3.

search of the chemical education literature and some popular organic laboratory texts
[10–14] does not reveal any instructional laboratory experiments using this important
method. A review of the chemical literature shows reductive amination to be a widely
used technique [15]; unfortunately, the toxicity of sodium cyanoborohydride, the
preferred reducing agent, precludes the use of these reactions in an undergraduate
setting. Recently, however, Abdel-Magid and co-workers [16, 17] described the use of
sodium triacetoxyborohydride, NaBH(OAc)3 [18, 19] as a safe and efficient alternative
to NaBH3CN [20]. We describe herein our adaptation of their procedure to the
reductive amination of pyruvate esters with benzylamine (Figure 1).

The original work of Abdel-Magid, et al. was quite extensive [16] and provided us
with a number of potential carbonyl–amine pairs from which to choose. Our choice of
pyruvate esters and benzylamine was based on a number of factors. The reaction times
were short enough (30 minutes) to allow easy incorporation of the experiment into a
typical laboratory period. Reaction yields were high; students typically obtained yields
of 60–80%. The product of the reaction is an N-protected alanine ester, a compound of
interest to the many biology and biochemistry majors in our course.
1
Structure determination was made via analysis of the H NMR spectrum of the
product. Due to overlap of signals, we found that it was best to have students work in
pairs, one using methyl pyruvate and the other using ethyl pyruvate. In both products,
the benzylic protons are diastereotopic and appear as two doublets due to the newly
created chiral center. The product formed from methyl pyruvate showed (300 MHz) an
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FIGURE 2. COMPARISON OF PRODUCT NMR DATA.

overlap of the signals from the methyl ester singlet and the benzylic signals. The
product formed from ethyl pyruvate gave an overlap of the signals from the methyl
triplet of the ethyl ester and the doublet from the methyl β to the ester. However, when
they compared spectra, the students could “subtract out” the alkoxy signals and
structure elucidation was simplified (Figure 2).

Experimental
Methyl pyruvate and ethyl pyruvate were obtained from Aldrich and Lancaster,
respectively, and used as received. Sodium triacetoxyborohydride was obtained from
Acros and used as received. Benzylamine was obtained from Baker and distilled by a
1 13
teaching assistant prior to the laboratory session. H and C NMR spectra were
recorded on a Bruker DPX-300 spectrometer at 300 MHz and 75.5 MHz, respectively.
Infrared spectra were recorded on a Perkin-Elmer Spectrum 1000 FT-IR spectrometer.

Molecular modeling was performed using CAChe (version 3.7) [21] which calculated
nucleophilic susceptibility after optimizing geometry with Augmented MM2 followed
by MOPAC [22]using PM3 parameters. Nucleophilic susceptibility “reveals reactive
sites based on the electron distribution of a set of active orbitals near the
LUMO” [21, 23].

General Procedure
To a 10-mL round-bottom flask containing a magnetic stir bar was added 1.00 mmol of
alkyl pyruvate, 1.00 mmol of benzylamine, and 3.5 mL of CH2Cl2. The mixture was
stirred for approximately 5 minutes, and 1.43 mmol of NaBH(OAc)3 was added in one
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portion. After stirring at room temperature for 30 minutes, the reaction was quenched
by addition of 3.5 mL of saturated NaHCO3. After stirring for 5 minutes, the reaction
mixture was transferred to a centrifuge tube, the layers were separated, and the
aqueous phase was extracted twice with 1 mL of CH2Cl2. The combined organic layers
were passed through a plug of anhydrous MgSO4. Solvent was removed, using a rotary
evaporator, to yield the alkyl N-benzylalaninate as a colorless oil. After recording the
mass of the isolated product, an IR spectrum was recorded and a sample was sumitted
for NMR analysis.

N-benzylalanine methyl ester (82% yield): H NMR (CDCl3, δ): 7.32 (m, 5H), 3.80 (d,
1

J = 12.8 Hz, 1H), 3.73 (s, 3H), 3.67 (d, J = 12.8 Hz, 1H), 3.40 (q, J = 7.0 Hz, 1H), 2.00
(br s, 1H), 1.32 (d, J = 7.0 Hz, 3H); C NMR (CDCl3, δ): 176.1, 139.6, 128.3, 128.1,
13

-1
127.0, 55.8, 51.8, 51.7, 19.0; IR (neat, cm ): 3330, 2952, 1736, 1454, 1199, 1152.

N-benzylalanine ethyl ester (85% yield): H NMR (CDCl3, δ): 7.29 (m, 5H), 4.19 (q, J
1

= 7.1 Hz, 2H), 3.81 (d, J = 12.8 Hz, 1H), 3.67 (d, J = 12.8 Hz, 1H), 3.37 (q, J = 7.0 Hz,
13
1H), 2.06 (br s, 1H), 1.32 (d, J = 7.0 Hz, 3H), 1.29 (t, J = 7.1 Hz, 3H); C NMR
(CDCl3, δ): 175.6, 139.7, 128.3, 128.1, 126.9, 60.5, 55.8, 51.8, 19.0, 14.2; IR (neat,
-1
cm ): 3329, 2980, 1732, 1453, 1187, 1153.

Discussion
Although most of the experiments in our organic sequence are of a synthetic nature,
we have incorporated elements of computational chemistry into a number of
experiments as a means of emphasizing differences in substrate reactivity [24, 25]. So,
in addition to the synthesis of a protected amino acid and the accompanying structure
elucidation, the selectivity of the reagents is a key discussion point in this experiment.
Benzylamine reacts exclusively with the ketone carbonyl in pyruvate while
NaBH(OAc)3 selectively reduces the imine of the intermediate. The selectivity of both
steps of this reaction can be investigated using molecular modeling with CAChe.

In the formation of the imine intermediate, the results (Figure 3) of an assessment of

the “nucleophilic susceptibility” clearly show that the ketone carbonyl is much more
reactive toward a nucleophile than the ester carbonyl, and formation of the imine thus
results. Similarly, once the imine has formed, NaBH(OAc)3 will attack the more
susceptible carbon and, again, nucleophilic susceptibility (as determined by CAChe)
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FIGURE 3. CALCULATED NUCLEOPHILIC SUSCEPTIBILITIES.

suggests that the imine will be reduced selectively. However, the CAChe results
indicate that, if the imine does not form in the brief period prior to introduction of the
reducing agent, reduction of the ketone by NaBH(OAc)3 should compete with
reduction of the imine, resulting in a product mixture containing alkyl 2-
hydroxypropionate. But, this outcome is not observed. In fact, when NaBH(OAc)3 is
mixed with pyruvate prior to the addition of benzylamine, the product is the same as
that obtained using the method described in the Experimental Section.

The solution to this paradox lies in the inherent basicity of the imine intermediate.
Imines will protonate even when the pH of a reaction is 6–8 and the resultant iminium
ion is more susceptible to reduction than the carbonyl of a ketone or an ester [16, 26]
Indeed, the iminium ion intermediate is in equilibrium with the imine and it is the
protonated species that undergoes reduction (Figure 4).

Sodium triacetoxyborohydride is also an unusually selective reducing agent. While

NaBH4 reduces aldehydes and ketones in the presence of esters, NaBH(OAc)3 has been
used to reduce aldehydes in the presence of ketones [27]. This additional selectivity
has been attributed to the sterically bulky and inductively electron withdrawing
acetoxy groups that stabilize the B–H bond [16, 27]. The success of reductive
aminations using this reagent is undoubtedly due, in part, to this selectivity. A more
powerful reducing agent could be expected to reduce the ketone before the iminium
ion intermediate could form.
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FIGURE 4. REDUCTION MECHANISM.

Conclusion
The reductive amination of pyruvate esters using benzylamine and NaBH(OAc)3 has
1
proven to be an operationally simple experiment that requires students to analyze H
NMR and IR spectra and use computational methods to compare the reactivities of
ketone and ester carbonyls as well as imines and iminium ions. Chemical yields
typically range from 60% to 80%. When students work in pairs, structure
determination, though challenging, is within the reach of introductory organic
students.

ACKNOWLEDGEMENT

We wish to thank the National Science Foundation for partial funding of Dickinson
College’s Bruker DPX-300 NMR spectrometer (DUE #9351330) and Molecular
Modeling Lab, including CAChe software and accompanying hardware
(DUE #9450995).

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