Lipase-catalyzed enantioselective reaction of amines with carboxylic acids under reduced pressure in non-solvent system and in ionic liquids

Article abstract of DOI:10.1016/j.tetlet.2003.10.210

Lipase-catalyzed enantioselective acylation of 1-phenylethylamine and 2-phenyl-1-propylamine was performed by reacting the amines with carboxylic acids in a non-solvent system or in ionic liquids as reaction media. The reaction equilibrium was shifted toward amide synthesis by the removal of formed water under reduced pressure.

Full text of DOI:10.1016/j.tetlet.2003.10.210

Tetrahedron
Letters
Tetrahedron Letters 45 (2004) 523–525
Lipase-catalyzed enantioselective reaction of amines with
carboxylic acids under reduced pressure in non-solvent system and
in ionic liquids
Roxana Irimescu and Katsuya Kato*
Bio-functional Ceramics Group, Ceramics Research Institute, National Institute of Advanced Industrial Science and Technology
(AIST), 2266-98 Anagahora, Shimoshidami, Moriyama-ku, Nagoya 463-8560, Japan
Received 14 October 2003; revised 31 October 2003; accepted 31 October 2003
Abstract—Lipase-catalyzed enantioselective acylation of 1-phenylethylamine and 2-phenyl-1-propylamine was performed by
reacting the amines with carboxylic acids in a non-solvent system or in ionic liquids as reaction media. The reaction equilibrium was
shifted toward amide synthesis by the removal of formed water under reduced pressure.
Ó 2003 Elsevier Ltd. All rights reserved.
Lipase-catalyzed acylation is one of the currently
employed methods for the resolution of primary amines.
Unlike the alcohols, for which vinyl esters are the most
common acylating agents, less reactive compounds are
required for the much more nucleophilic amines to
avoid spontaneous non-enzymatic reactions. Simple
esters or carbonates are usually employed.1 Carboxylic
acids have been scarcely used as acyl donors as it was
supposed that the spontaneous formation of an unre-
active salt with the amine would prevent the formation
of the acyl-enzyme complex. However, the salt forma-
tion is a reversible reaction with the salt being in equi-
librium with some amounts of free acid and amine. This
small amount of acid is available to the enzyme, and
Montet et al.2 demonstrated that the amide bond was
synthesized by lipase in hexane, though with a very low
reaction rate: 60% conversion of amine in 12 days. The
acylation reaction is also a reversible reaction with the
amide and water on the synthetic side of the equilibrium.
In this study, we tried to develop an effective method for
enantioselective acylation of primary amines with car-
boxylic acids catalyzed by lipase in a non-solvent system
or in ionic liquids as reaction media. The reaction
equilibrium was displaced in favor of amide formation
by the continuous removal of the by-product water
under low pressure (Scheme 1). Two primary amines:
1-phenylethylamine (1) with the chiral center at the
a-carbon atom, and 2-phenyl-1-propylamine (2) with
the chiral center at the b-carbon atom were used as
substrates in the reactions catalyzed by immobilized
Candida antarctica lipase B (CALB).
The immobilized CALB catalyst (Chirazyme L-2, c.-f,
C2, Lyo.) (EC 3.1.1.1.) was purchased from Roche
Diagnostics (Mannheim, Germany). The experiments in
non-solvent system were carried out as follows: the
lipase (23 mg) was added to a mixture of amine (1 mmol)
and acid (1 mmol) under stirring at a specified temper-
ature and 5 mm Hg vacuum was applied to the system.
The reaction was stopped by neutralizing the unreacted
acid with NaOH solution (2 N) followed by extraction
of amine and amide with diethyl ether. The experiments
in ionic solvents were carried out with the same amounts
as above by adding the lipase to a mixture of amine and
acid in ionic solvent (1.5 mL) and connecting the system
to a vacuum pump. The reaction was stopped by the
extraction of the reaction mixture with diethyl ether.
The amine and amide were extracted separately
according to Gonzalez-Sabin et al.3 The enantiomeric
compositions of the residual amine and the formed
amide were determined by HPLC on a Chiralpak AD-H
(Dicel Chemical Industries, Tokyo, Japan) column
eluted with a mixture of hexane/2-propanol/diethyl-
amine with a volumetric ratio of 95:5:0.05. The
Keywords: Enzymatic amide synthesis; Ionic liquids; Kinetic resolu-
tion; Lipase; Non-solvent reaction system; Primary amine.
* Corresponding author. Tel.: +81-52-736-7275; fax: +81-52-736-7405;
e-mail: katsuya-kato@aist.go.jp
0040-4039/$ - see front matter Ó 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2003.10.210

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R. Irimescu, K. Kato / Tetrahedron Letters 45 (2004) 523–525
CH3
CH3
CH3
NH2
+
Ph
Ph
Ph
Ph
NHCO-R
NH2
rac-1
lipase,
3
4
vacuum
or
+
or
R COOH
H2O
+
non-solvent
or
ionic liquid
CH3
CH3
CH3
+
NH2
NH2
NHCO-R
Ph
Ph
5
rac-2
6
-
R = C7H15, C11H23, CH2=CHCH2CH2
Scheme 1. Lipase-catalyzed acylation of primary amines.
(R)-enantiomer was preferentially acylated for amine 1
while for amine 2, it was the (S)-enantiomer. The
absolute configuration of amine 5 was determined by
comparison with the retention times of the pure enan-
tiomer standards on chiral HPLC. The enantiomeric
excess of amine 1 was determined after its transforma-
tion into the corresponding acetamide (acetic anhydride,
pyridine). The (S)-configuration of amine 3 isolated
from the reaction mixture and transformed in acetamide
was determined after comparison of the retention times
with the enantiomerically pure (R)- and (S)-acetamides
of 1 prepared chemically from pure enantiomers of 1.
The absolute configuration of the formed amides 4 and 6
was determined similarly using chemically prepared
amide enantiomer standards (carboxylic acid, DCC,
DMAP in dichloromethane).
the enzyme for the acid than for the corresponding ethyl
ester.5 Dodecanoic acid was a better acyl donor: the
reaction was faster and highly enantioselective. A simi-
lar effect was observed for these acids in our previous
study on enantioselective esterification of secondary
alcohols with free fatty acids in non-solvent system.5 The
pent-4-enoyl group is an amine protecting group that is
readily cleaved under mild conditions6 and therefore, it
is worth investigating the use of 4-pentenoic acid as acyl
donor for enzymatic acylation of amines. The reaction
with 4-pentenoic acid was very sluggish for amine 1
(entry 5), but proceeded with a moderate rate for amine
2 (entry 7). Amine 2 can access more easily the active site
of the enzyme due to the remote stereogenic center.
Therefore, the reaction is faster, but the enantioselec-
tivity is much lower than for 1.
First, the feasibility of a non-solvent system was inves-
tigated. The reactions with equimolar amounts of amine
1 and octanoic acid (Table 1, entry 1), and dodecanoic
acid (entry 2) were performed. For both, the reaction
mixtures were very viscous and the reaction temperature
had to be raised to 55 °C in order to achieve an efficient
stirring in the system. Although a much slower reaction
was expected, the rate and enantioselectivity for the
reaction with octanoic acid were close to that obtained
in a previous study, which used ethyl octanoate as acyl
donor in a non-solvent system under reduced pressure.4
An explanation might be that the negative effect on the
reaction rate produced by the formation of the salt with
the amine was compensated by the higher specificity of
Ionic liquids are increasingly used as reaction media in
organic syntheses as they offer a wide range of advan-
tages over classical organic solvents.7 Recently, enzy-
matic transformations were also performed successfully
and in many cases, with remarkably improved results in
ionic liquids.8 Lau et al.8a reported the first examples of
lipase-catalyzed reactions performed in ionic liquids in
the absence of water. First asymmetric enzymatic reac-
tions in ionic liquids were reported almost at the same
8c
time by Itoh et al.8b and Schofer et al. Both studies
€
dealt with kinetic resolution of secondary alcohols by
acylation with vinyl acetate and demonstrated that the
enzyme suspended in the ionic liquids could be reused
efficiently several times. In a further development of the
Table 1. Lipase-catalyzed enantioselective acylation of primary amines with carboxylic acids
Entry
Amine
Acid
Solvent
Temperature Initial rate Reaction
(°C)
(lmol/h mg)a time (h)
Conversion
amine (%)
EeS
(%)b
EeP
(%)c
E
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
2
2
2
Octanoic
None
None
55
55
55
55
30
30
30
30
30
0.57
0.88
1.67
0.38
0.005
0.07
0.31
1.92
0.71
24
19
19
24
24
24
24
5
31.6
44.5
48.9
20.7
0.6
44.8
80.3
95.9
26.2
0.6
97.1
106
Dodecanoic
Dodecanoic
Dodecanoic
4-Pentenoic
4-Pentenoic
4-Pentenoic
4-Pentenoic
4-Pentenoic
>99.0
>99.0
>99.0
>99.0
>99.0
59.8
>500
>500
>500
>500
>500
6
[bmim]PF6
[emim]BF4
None
[bmim]PF6
None
8.0
8.7
38.2
48.1
17.8
36.9
24.5
5.3
[bmim]PF6
[emim]BF4
48.1
25.7
2
5
2
a Amount of amine (lmol) esterified per hour and milligram of immobilized enzyme.
b Enantiomeric excess of unreacted amine.
c Enantiomeric excess of resulting amide.

R. Irimescu, K. Kato / Tetrahedron Letters 45 (2004) 523–525
525
ionic liquid system, Itoh et al.8d used reduced pressure to
remove resulting methanol from the enantioselective
transesterification of 5-phenyl-1-penten-3-ol with methyl
esters and improved the reaction rates and yields. The
reasons for investigating the use of ionic liquids as
reaction media for our system are the following: (1) they
have no detectable vapor pressure and therefore can be
used under reduced pressure, and (2) they are very good
solvents for a wide range of organic compounds. Ionic
liquids have the ability to dissolve compounds with
relatively different polarities (in this work: the substrates
and the resulting amide) so that they can improve the
properties of the reaction mixture and alleviate the dif-
fusional limitations of a very viscous reaction mixture,
as in our case. In an attempt to improve the reaction
performance, two ionic liquids with different properties,
the hydrophobic 1-butyl-3-methylimidazolium hexaflu-
orophosphate ([bmim]PF6) and the hydrophilic 1-ethyl-
3-methylimidazolium tetrafluoroborate ([emim]BF4),
were tested as reaction media. The initial rate for the
reaction of amine 1 with dodecanoic acid doubled in
[bmim]PF6 (entry 3), but reduced to half in [emim]BF4
(entry 4). The rate was raised more than an order by
performing the reaction of 1 with 4-pentenoic acid in
[bmim]PF6 (entry 6). The enantioselectivity remained
very high for the reactions of 1 in both ionic liquids.
Different results were obtained for the reactions of
amine 2 with 4-pentenoic acid. In both ionic liquids, the
initial reaction rate increased (entries 8 and 9) (with a
more remarkable effect for [bmim]PF6), but the enantio-
selectivity decreased to less than half of the value in non-
solvent system.
ates.10 Better reaction rates were achieved by the use of
ionic liquids as reaction media. We intend to further
improve the performances of the system by expanding
this study to a larger variety of ionic liquids. Finding the
best reaction medium is quite a complicated and tedious
task as its performances depend on the nature of each
substrate and expected product and in many cases, the
most performing reaction medium is very different even
for similar compounds.10 This work is made easier by
the fact that the ionic liquid properties (polarity, vis-
cosity, density, solubility, etc.) can be finely or drasti-
cally modified according to the oneÕs purpose by
changing the anion or the components of the cation.7;8;10
Acknowledgements
A postdoctoral research fellowship from the Japanese
Society for Promotion of Science for Roxana Irimescu is
gratefully acknowledged.
References and Notes
1. Bornscheuer, U. T.; Kazlauskas, R. J. Hydrolases in
Organic Synthesis; Willey-VCH: Weinheim, 1999. pp 103–
106.
2. Montet, D.; Pina, M.; Graille, J.; Renard, G.; Grimaud, J.
Fat. Sci. Technol. 1989, 91, 14–18.
3. Gonzalez-Sabin, J.; Gotor, V.; Rebolledo, F. Tetrahedron:
Asymmetry 2002, 13, 1315–1320.
€
4. Orner, N.; Orrenius, C.; Mattson, A.; Norrin, T.; Hult, K.
To our knowledge, the results presented in this study
demonstrated for the first time the amide synthesis in
non-solvent system by the direct lipase-catalyzed reac-
tion of amines with carboxylic acids and its use for
kinetic resolution of primary amines. This system is the
enzymatic equivalent to the general procedure for amide
synthesis involving the heating of a mixture of amine
and acid at about 200 °C.9 The present enzymatic pro-
cedure offers some important advantages over the
chemical one for the application in industrial processes.
The reaction temperatures are significantly reduced and
as a result its applications can be extended to thermally
labile compounds. In addition, the enantioselectivity of
lipase allows the kinetic resolution of chiral substrates.
In comparison to other lipase-catalyzed systems used to
date, the reaction rates, conversions and enantioselec-
tivities are comparable to the reactions employing other
more expensive acyl donors such as esters or carbon-
Enzyme Microb. Technol. 1996, 19, 328–331.
5. Irimescu, R.; Saito, T.; Kato, K. J. Am. Oil Chem. Soc.
2003, 80, 659–663.
6. Madsen, R.; Roberts, C.; Fraser-Reid, B. J. Org. Chem.
1995, 60, 7920–7926.
7. Welton, T. Chem. Rev. 1999, 99, 2071–2083.
8. For some examples of lipase-catalyzed transformations in
ionic solvents, see: (a) Madeira Lau, R.; van Rantwijk, F.;
Seddon, K. R.; Sheldon, R. A. Org. Lett. 2000, 2, 4189–
4191; (b) Itoh, T.; Akasaki, E.; Kudo, K.; Shirakami, S.
€
Chem. Lett. 2001, 262–263; (c) Schofer, S. H.; Kaftzik, N.;
Wasserscheid, P.; Kragl, U. Chem. Commun. 2001, 425–
426; (d) Itoh, T.; Akasaki, E.; Nishimura, Y. Chem. Lett.
2002, 154–155.
9. Beckwith, A. L. J. The Chemistry of Amides. In Synthesis
of Amides; Zabicky, J., Ed.; Interscience Publishers:
London, 1970; pp 105–109.
10. van Rantwijk, F.; Hacking, M. A. P. J.; Sheldon, R. A.
Monatsh. Chem. 2000, 131, 549–569.