some recent bacterial resolutions of closely related molecules
(cyclopropanes6 and oxiranes7) bearing nitrile or amide
functionalities.
Table 1. Bacterial Preparation of Enantiopurea N-Substituted
Aziridine-2-carboxamides 1a-d
We now report our first findings in this field, namely, the
enantioselective hydrolysis of several unactivated 1-benzyl-
[(()-1a] or 1-arylaziridine-2-carboxamides [(()-1b-d] by
the amidase-containing, commercially available bacterium
Rhodococcus rhodochrous IFO 15564,8 leading to the easy
preparation of enantiopure amides (1R,2S)-1a-d. Moreover,
some regio- and stereoselective NROs of (1R,2S)-1a and its
LAH reduction product are also described.
As the slow or blocked pyramidal inversion in aziridines
introduces a stereogenic nitrogen,1a the possible existence
of cis-trans diastereomers (invertomers) has to be taken into
account for racemic and enantiopure compounds 1. However,
according to previous H NMR studies,9 our own H and
13C NMR spectra, including NOESY correlations, reveal the
sole presence of the trans invertomers. DFT calculations
(B3LYP method, 6-31G* basis set) also corroborate the
existence of these structures.10
product
R
time (h)
yieldb (%)
(1R,2S)-1a
(1R,2S)-1b
(1R,2S)-1c
(1R,2S)-1d
PhCH2
67
45
47
46
45
1
1
Ph
4.5
4.1
6.7
4-MeOC6H4
4-F3CC6H4
a ee >99.5% (HPLC). bIsolated yield after column chromatography (from
rac-1a-d as starting materials).
Biotransformations of racemic substrates trans-1 were
carried out with a standard cell concentration of R. rhodo-
chrous IFO 1556411 in the metabolic resting phase [ap-
proximately 2.7 mg/mL of aqueous 0.10 M, pH 7.0 potas-
sium phosphate (equivalent to A65012 ) 3.0)]. The reactions
were stopped after HPLC analysis (Chiralcel OD) showed
the presence of only one out of the two enantiomers of the
corresponding aziridinecarboxamide 1. All the enantiopure
amides were obtained in this way with very good yields,
considering that the upper limit for any kinetic resolution is
50% yield (Table 1).
Biotransformations can also start from the N-substituted
aziridine-2-carbonitriles (()-2a-c, due to the concomitant
presence in the bacterium of a nitrile hydratase. In such cases,
yields (42-43%) were slightly lower than those obtained
from (()-1. The nitrile hydratase step was found to be too
fast and poorly enantioselective, as has been generally
reported for Rhodococci strains.13,14
and also 2a). A possible reason for such a great difference
in reaction rates may lie in the fact that aryl substituents are
stiffer than the benzyl group, thus fitting more efficiently
into the active site of the amidase.
None of the expected optically active aziridinecarboxylic
acids 3 could be isolated after biotransformations. The
instability of some aziridine-2-carboxylic acids [including
that of 1-(R-methylbenzyl)aziridine-2-carboxylic acid, closely
related to 3a] has been explicitly or implicitly established
several times,15 though without any indication as to their
decomposition pathways. By means of complementary
experiments, however, we were able to discard a concerted
decarboxylation mechanism similar to that well-known for
oxiranecarboxylic acids.16
To rule out possible incompatibility between amino acids
3 and aqueous media, we carried out a Candida antarctica
lipase (CAL-B) catalyzed hydrolysis of the ethyl ester of 3a
(0.5 mmol) in anhydrous THF (4 mL), adding a minimal
amount of water (54 µL). Although 1H and 13C NMR spectra
of the crude material were not conclusive as to the presence
of the amino acid 3a, it can be discarded from the mass
spectrum (ESI). After silica gel column chromatography, the
remaining ester showed a modest enantiomeric excess (ee
) 20%), an indication that enzymatic hydrolysis proceeded,
but the tail fractions consisted of a very complex mixture of
It should be noted that N-aryl racemic substrates 1b-d
(and also 2b,c) produce the corresponding enantiopure
amides at least ten times faster than N-benzyl substrates (1a
(6) (a) Wang, M.-X.; Feng, G.-Q. Tetrahedron Lett. 2000, 41, 6501-
6505. (b) Wang, M.-X.; Feng, G.-Q. J. Org. Chem. 2003, 68, 621-624.
(c) Wang, M.-X.; Feng, G.-Q.; Zheng, Q.-Y. Tetrahedron: Asymmetry 2004,
15, 347-354.
(7) (a) Wang, M.-X.; Lin, S.-J.; Liu, C.-S.; Zheng, Q.-Y.; Li, J.-S. J.
Org. Chem. 2003, 68, 4570-4573. (b) Wang, M.-X.; Deng, G.; Wang, D.-
X.; Zheng, Q.-Y. J. Org. Chem. 2005, 70, 2439-2444.
(8) Formerly known as Rhodococcus butanica ATCC 21197. (a) Kakeya,
H.; Sakai, N.; Sugai, T.; Ohta, H. Tetrahedron Lett. 1991, 32, 1343-1346.
(b) Effenberger, F.; Bo¨hme, J. Bioorg. Med. Chem. 1994, 2, 715-721.
(9) (a) Saitoˆ, H.; Nubada, K.; Kobayashi, T.; Morita, K.-i. J. Am. Chem.
Soc. 1967, 89, 6605-6611. (b) Manatt, S. L.; Elleman, D. D.; Brois, S. J.
J. Am. Chem. Soc. 1965, 87, 2220-2225. (c) Bouteville, G.; Gelas-Mialhe,
Y.; Vessie`re, R. Bull. Soc. Chim. Fr. 1971, 3264-3270.
(10) trans-1 compounds are 2.7-4.8 kcal/mol more stable than cis-1. In
trans-1, an intramolecular hydrogen bond (amide N-H as a proton donor,
cyclic N as a proton acceptor) accounts for their extra stabilization in relation
to cis-1 compounds.
(14) In a complementary experiment, R. rhodochrous was grown in the
presence of diethyl phosphoramidate (an amidase inhibitor11), suspended
in the usual phosphate buffer with an A650 of only 0.30, and then exposed
to (()-1-benzylaziridine-2-carbonitrile (2a) during 34 min. From the crude
product (a 56:44 mixture of 2a/1a; crude yield, 90%), we were able to
isolate the amide (1S,2R)-1a with 8% ee. Therefore, the nitrile hydratase
has a poor 2R-selectivity, the same as the amidase’s high 2R-selectivity.
This observation accounts for the fact that the yields in (1R,2S)-1 obtained
from (()-2 were slightly lower than those obtained from (()-1.
(15) See ref 5c and also: Lambert, C.; Viehe, H. G. Tetrahedron Lett.
1985, 26, 4439-4442. In ref 5e, such acids have to be formed, but no
comment is made about them.
(11) Gotor, V.; Liz, R.; Testera, A. M. Tetrahedron 2004, 60, 607-
618.
(12) Absorbance or optical density at 650 nm.
(13) Beard, T.; Cohen, M. A.; Parratt, J. S.; Turner, N. J. Tetrahedron:
Asymmetry 1993, 4, 1085-1104.
(16) Singh, S. P.; Kagan, J. J. Org. Chem. 1970, 35, 2203-2207 and
references therein.
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Org. Lett., Vol. 9, No. 3, 2007