Candida antarctica lipase A - A powerful catalyst for the resolution of heteroaromatic β-amino esters

Article abstract of DOI:10.1016/S0957-4166(02)00637-7

Enantioselective acylations of 3-amino-3-heteroarylpropanoates (ArCH(NH₂)CH₂CO₂Et; Ar=2- or 3-thienyl or -furyl) were performed in the presence of Candida antarctica lipase A. As a result of the excellent chemo- and enantioselectivities (E >100), gram-scale resolutions were carried out in ethyl butanoate. The hydrochloride salts of the unreacted R substrates and the butanamides of the reactive S enantiomers were thus prepared.

Full text of DOI:10.1016/S0957-4166(02)00637-7

TETRAHEDRON:
ASYMMETRY
Pergamon
Tetrahedron: Asymmetry 13 (2002) 2383–2388
Candida antarctica lipase A—a powerful catalyst for the
resolution of heteroaromatic b-amino esters
Magdolna Solyma´r,a,b Ferenc Fu¨lo¨pb and Liisa T. Kanervaa,*
aLaboratory of Synthetic Drug Chemistry and Department of Chemistry, University of Turku, Lemminka¨isenkatu 2,
FIN-20520 Turku, Finland
bInstitute of Pharmaceutical Chemistry, University of Szeged, PO Box 121, H-6701 Szeged, Hungary
Received 13 September 2002; revised 4 October 2002; accepted 8 October 2002
Abstract—Enantioselective acylations of 3-amino-3-heteroarylpropanoates (ArCH(NH2)CH2CO2Et; Ar=2- or 3-thienyl or -furyl)
were performed in the presence of Candida antarctica lipase A. As a result of the excellent chemo- and enantioselectivities (E
>100), gram-scale resolutions were carried out in ethyl butanoate. The hydrochloride salts of the unreacted R substrates and the
butanamides of the reactive S enantiomers were thus prepared. © 2002 Elsevier Science Ltd. All rights reserved.
1. Introduction
because E values are not given and enantiomeric puri-
ties either refer to those obtained after recrystallization
or are known for only one of the resolution products.
Accordingly, we considered it of interest to study the
CAL-A-catalysed resolution of heteroaryl-substituted
b-amino esters 1–4 (Scheme 1). Furthermore, incorpo-
ration of an appropriate (S)-3-amino-3-heteroaryl-
propanoic acid (or ester) into nipecotic acid is known
to enhance antithrombotic activity in platelet fibrinogen
receptor antagonists.19,20
Some b-amino acids and esters exhibit interesting phar-
macological effects and many b-amino acids are of
significance as constituents of natural products and
pharmaceutical drug compounds.1–6 b-Amino acids are
used for the preparation of b-peptides, which adopt
well-defined three-dimensional structures1,6–10 and they
also serve as precursors for b-lactams and heterocyclic
compounds.6 Consequently, enantiopure b-amino acids
have become important and challenging targets in syn-
thetic organic chemistry.
CAL-A is a calcium dependent, thermostable lipase
that is reported to be highly active in a non-specific
manner.21 The enzyme is unique because of its reported
SN2 preference towards triacylglycerols.22 Although
only rarely used for enantioselective reactions, the suc-
cess of CAL-A for the resolution of amino esters is
conspicuous.11,13,23,24 In this work, it was essential that
CAL-A adsorbed on Celite in the presence of sucrose
was used. For instance, the results for the acylation of
ethyl cis-2-aminocyclohexyl carboxylate (the first appli-
cation of CAL-A in our laboratory)11 with 2,2,2-tri-
fluoroethyl acetate in the presence of native (vo=3 mmol
mg−1 min−1; E=7) and immobilized (vo=12 mmol mg−1
min−1; E=31) enzyme in diethyl ether show consider-
able reactivity and enantioselectivity enhancements for
the immobilized enzyme.25 The addition of sucrose was
earlier shown to exert its own positive effect on enzy-
matic activity.26 A possible explanation for the effect of
sucrose lies in its highly hydrophilic nature which can
help the enzyme to maintain a high water level in
Enzymes have gained wide acceptance as asymmetric
catalysts in organic chemistry, especially with the aim
of the preparing enantiopure compounds. As concerns
b-amino esters, our research has long been focused on
the lipase-catalysed kinetic resolutions of alicyclic 2-
aminocarboxylic and 3-amino-3-alkyl- or 3-amino-3-
phenylpropanoic esters.11–14 In these reactions, CAL-A
(lipase A from Candida antarctica) has proven excep-
tionally effective for acylation reactions in organic sol-
vents.11,13 Other studies relate to catalysis with
penicillin and penicillin G acylase for N-phenylacetyla-
tion of the amino group or hydrolysis of the N-phenyl-
acetyl group in 3-aminopropanoic acid analogues
containing aliphatic, aromatic and heteroaromatic sub-
stituents at position 2 or 3.15–20 The level of enantiose-
lectivity in the acylase methods is somewhat obscure
* Corresponding author.
0957-4166/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0957-4166(02)00637-7

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M. Solyma´r et al. / Tetrahedron: Asymmetry 13 (2002) 2383–2388
anhydrous organic solvents. On the basis of these ear-
lier results, CAL-A on Celite when adsorbed in the
presence of sucrose was used throughout the work
reported herein.
CAL-A is also obvious for the reactions of racemic 1–4
with the studied achiral acetate and butanoate esters
and consequently, 1b–4b are the only new products
detected by GC and GC–MS (Table 1 and Scheme 1).
The acylations of 1–4 under various reaction conditions
in the presence of CAL-A proceed smoothly to 50%
conversion and at this point results in highly enantio-
pure resolution products (Table 1). The data (and more
clearly the progression curves; not given here) further
indicate that the reactivities (the time needed to reach a
certain conversion) are practically independent of the
substrate structure for given reaction conditions (Table
1, entries 2, 10, 17 and 20; entries 5, 13, 18 and 21;
entries 14, 19 and 22). Substrate 1 with a 2-thienyl ring
2. Results and discussion
One of the most impressive properties of CAL-A is its
absolute chemoselectivity in the N-acylation of b-amino
esters under conditions where other lipases tend to lead
to competition between N-acylation at the amino group
and transesterification at the ester group, the extent of
competition being controlled by the structure of the
amino ester.11–14,23,24 The absolute chemoselectivity of
Scheme 1. R=Me or Pr; R1=Et, CH2CF3 or Bu.
Table 1. Acylation of 1–4 (0.05 M) by CAL-A (15 mg/ml) with achiral esters (0.1 M or neat) at room temperature (25°C)
Entry
Substrate
RCO2R1
Solvent
Time (h)
Conv. (%)
eea (%)
eeb (%)
E
1
2
3
4
5
6
7
8
1
1
1
1
PrCO2CH2CF3
PrCO2CH2CF3
PrCO2CH2CF3
PrCO2CH2CF3
PrCO2Et
PrCO2Et
PrCO2Bu
MeCO2Et
PrCO2CH2CF3
PrCO2CH2CF3
PrCO2CH2CF3
PrCO2CH2CF3
PrCO2Et
PrCO2Et
PrCO2Bu
MeCO2Et
PrCO2CH2CF3
PrCO2Et
tBuOMe
iPr2O
MeCN
0.33
1.5
1
8
3
48
50
50
49
50
50
45
49
40
50
52
47
48
50
49
51
47
48
50
49
47
51
89
99
95
92
96
97
82
88
62
96
99
86
90
94
94
97
88
90
93
95
87
\99
98
97
95
95
97
98
99
90
94
96
89
97
96
92
97
92
98
97
94
99
99
96
380
380
130
120
305
410
580
60
60
210
90
210
150
90
220
100
245
220
110
510
460
470
THF
1
PrCO2Et
PrCO2Et
PrCO2Bu
MeCO2Et
tBuOMe
iPr2O
1a
1
6
0.5
60
0.67
1.3
1.3
11
3
10
1
66
1.5
4
1
2
2
2
9
10
11
12
13
14
15
16
17
18
19
20
21
22
MeCN
THF
2
2
PrCO2Et
PrCO2Et
PrCO2Bu
MeCO2Et
iPr2O
PrCO2Et
PrCO2Et
iPr2O
2a
2
2
3
3
3a
4
PrCO2Et
PrCO2CH2CF3
PrCO2Et
11
1
2
4
PrCO2Et
PrCO2Et
4a
PrCO2Et
11
a Substrate 0.2 M.

M. Solyma´r et al. / Tetrahedron: Asymmetry 13 (2002) 2383–2388
2385
tends to react somewhat faster than the other com-
pounds, although this can be clearly seen only for the
reaction in ethyl butanoate when more concentrated
(0.1 M) substrate is used (entry 6 as compared with
entries 14, 19 and 22). The reactions of 1 and 2 are
faster in butyl butanoate (entries 7 and 15) and much
slower in ethyl acetate (entries 8 and 16) than the
corresponding reactions in ethyl butanoate (entries 5
and 13), indicating that butanoates are favoured by the
enzyme. An unexpected feature of the present results is
the considerable reactivity drop for the acylation of 1–4
in ethyl butanoate at substrate concentration of 0.1 M
(entries 6, 14, 19 and 22) as compared with the gener-
ally used concentration of 0.05 M (entries 5, 13, 18 and
21). A probable reason is substrate inhibition, although
this was not studied in detail in the present work.
with the generally accepted finding that lipases do not
constrain amide bonds to react. The CAL-A-catalysed
acylations of 1 and 2 with alkyl-activated 2,2,2-trifl-
uoroethyl butanoate in organic solvents were also stud-
ied (Table 1, entries 1–4 and 9–12). Solvent effects on
the reactivity and enantioselectivity are clear, the best
t
i
solvents being BuOMe and Pr2O for 1 (entries 1 and
i
2), and Pr2O (entries 10 and 12) for 2. No benefit of
using 2,2,2-trifluoroethyl butanoate over the more eco-
nomical ethyl butanoate can be seen.
It may be concluded, that CAL-A-catalysed acylation is
an excellent and highly selective method of resolution
for various types of b-amino esters,11,13 the present
results adding 3-heteroaryl-substituted propanoate
esters to the list. On the basis of the results, the
enantiomers of 1–4 were resolved on a gram scale in
ethyl butanoate, as shown in the experimental section
(Scheme 1). In spite of the longer reaction times needed
to reach 50% conversion, a substrate concentration of
0.1 M was used in the cases of 1 and 4.
We earlier found CAL-A to be a highly enantioselective
catalyst for the acylation of various b-amino esters.11,13
Excellent enantioselectivities in terms of the enantiomer
ratio (E >100) are also observed for the reactions of 1–4
with various butanoate esters (Table 1). On the other
hand, for the CAL-A-catalysed acylation of 3-amino-3-
phenylpropanoate with 2,2,2-trifluoroethyl butanoate in
diisopropyl ether (E=75) and with butyl butanoate in
the neat ester (E=30), much lower enantioselectivities
and reactivities have been detected.13 It is possible that
the heteroatoms in the thienyl and furyl rings of com-
pounds 1–4 interact favourably with the protein struc-
ture of the enzyme, thus enhancing both the reactivity
and enantioselectivity. The difference in the aromatic
3-substituents is that the polarity increases with
decreasing aromatic character (i.e. in the sequence
phenyl>furyl>thienyl). The accuracy of the E values in
Table 1 is open to question because it is well known
that for E >100 even minor errors in ee may cause a
significant variation in E. However, the given values
more or less correctly describe the differences in E from
one substrate to another under various conditions.
There is still a question about the absolute configura-
tions of 1a–4a and 1b–4b to be solved. We herein rely
on the observed enantioselectivity of CAL-A for the
acylation of aliphatic b-substituted b-amino esters in
the previous work13 and expect the S absolute configu-
ration for 1b–4b. To support this, the Candida antarc-
tica lipase B (CAL-B)-catalysed acylations of the
present substrates were conducted in neat ethyl
butanoate. The opposite selectivity observed for CAL-B
relative to that for CAL-A justifies the expectation, as
opposite enantiodiscrimination was observed earlier for
the enzymes when aliphatic b-amino esters were used as
substrates.13,14,24 Otherwise, very slow reactions, with E
values of only 4, 10, 3 and 2 for the reactions of 1–4,
respectively, make CAL-B useless as a catalyst for
kinetic resolution of the present substrates. The abso-
lute configurations of 1a was also proved via reductive
desulphurization by Raney nickel, resulting in 3-amino-
heptanoic acid.27 The measured specific rotation of the
product in comparison with the literature value28 indi-
cates S absolute configuration; therefore the R absolute
configuration for 1a and S for 1b correlate with the
observed enantioselectivity of CAL-A for the acylation
of aliphatic b-substituted b-amino esters.13
Previous results relating to alicyclic b-amino esters indi-
cated the enhanced enantioselectivity and reactivity of
CAL-A with increasing carbon chain length of the acyl
part of an achiral acyl donor.11 Substrates 1 and 2 were
subjected to enzymatic acylation conditions both in
neat ethyl acetate and in ethyl butanoate. The reactions
in the latter medium (Table 1, entries 5 and 13) were
not only faster, but also more enantioselective than in
the acetate ester (entries 8 and 16), structural effects
being observed in addition to solvent effects. Accord-
ingly, butanoate esters were generally used in this work.
3. Materials and methods
2,2,2-Trifluoroethyl butanoate was prepared from
butanoyl chloride and 2,2,2-trifluoroethanol. CAL-A
(Candida antarctica lipase A, Chirazyme L5) and CAL-
B (Candida antarctica lipase B, Chirazyme L2) were
purchased from Boehringer–Mannheim. Before use
CAL-A and sucrose (0.24 g) were dissolved in Tris–HCl
buffer (pH 7.9), the solution was added on Celite (4.0 g)
and thereafter left to dry at room temperature.26 The
final lipase content in the enzyme preparation was 10%
(w/w). The solvents were of the highest analytical grade
from Lab Scan Ltd and were dried over molecular
sieves before use. The progress of the reactions and the
ee values were followed by taking samples (0.1 ml) at
For kinetic enzymatic resolution in organic solvents,
special attention is usually paid to the nature of the
achiral acyl donor in order to avoid reverse enzymatic
catalysis, which can become important with the accu-
mulation of products such as 1b–4b. The reverse reac-
tion, if it occurs, can easily be seen via the lowering of
the enantiopurities of the less reactive enantiomers at
higher conversions. In the present N-acylations, eea
>99% remains unchanged after the more reactive enan-
tiomer has reacted at somewhat over 50% conversion.
The absence of the reverse reaction is in accordance

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M. Solyma´r et al. / Tetrahedron: Asymmetry 13 (2002) 2383–2388
intervals and analysing them by gas chromatography
on a Chrompack CP-Chirasil- -Val column (25 m). For
3.1.4. 3-Amino-3-(3-furyl)propanoic acid. Dark-brown
crystals, mp >250°C, 1H NMR (400 MHz, D2O): l
2.78–2.84 (2H, m, CH2CO), 4.66 (H, t, J=7.00 Hz
CHCH2CO), 6.58 (1H, d, J=1.20 Hz, aromatic), 7.57
(1H, t, J=1.80 Hz, aromatic), 7.66 (1H, s, aromatic).
L
good baseline separation, the unreacted amino group in
the sample was derivatized with acetic or propanoic
anhydride in the presence of pyridine containing 1%
4-N,N-dimethylaminopyridine (DMAP) before injec-
tions. In a typical small-scale experiment, one or other
of the substrates 1–4 (0.05 M if not otherwise stated)
was dissolved in the reaction medium containing an
acyl donor (2 equiv. in an organic solvent or the solvent
itself) and the CAL-A preparation (15 mg/ml) was
added. The reactions in the case of CAL-B (40 mg/ml)
were performed in the same way. All reactions pro-
ceeded at room temperature (25°C). The determination
of E was based on the equation E=ln[1−eea)/(1−eea/
eeb)]/ln[(1−eea)/(1+eea/eeb)] with c=eeS/(eeS+eeP) as
derived from the original equations of Chen et al.29
For the preparation of amino esters 1–4, the above
acids were esterified with ethanol in the presence of
thionyl chloride, followed by the bubbling of ammonia
through the solution in CH2Cl2 for 2 h. Ammonium
chloride was filtered off and the filtrate was evaporated.
The free base was eluted from a silica column, using
CH2Cl2:MeOH (95:5) as eluent.
3.1.5. Ethyl 3-amino-3-(2-thienyl)propanoate, 1. 1H
NMR (400 MHz):
l 1.23 (3H, t, J=7.15 Hz,
OCH2CH3), 2.70–2.90 (2H, m, CH2CO), 4.22 (2H, q,
J=7.19, OCH2CH3), 4.75–4.80 (1H, m, CHCH2CO),
6.92 (1H, d, J=3.42 Hz, aromatic), 7.18 (2H, t, J=6.41
Hz, aromatic).
1H NMR spectra were recorded on solutions in CDCl3
or D2O at ambient temperature on a JEOL L400 or a
Bruker AM200 spectrometer. Chemical shifts are given
in l (ppm) relative to TMS as internal standard; multi-
plicities were recorded as s (singlet), d (doublet), dd
(double doublet), t (triplet) or m (multiplet). MS spec-
tra were recorded on a VG Analytical 7070E instru-
ment. Elemental analyses were performed with a
Perkin–Elmer CHNS-2400 Ser II Elemental Analyser.
Optical rotations were measured with a Perkin–Elmer
241 polarimeter, and [h]D values are given in units of
10−1 deg cm2 g−1.
1
3.1.6. Ethyl 3-amino-3-(2-furyl)propanoate, 2. H NMR
(400 MHz): l 1.21 (3H, t, J=7.12 Hz, OCH2CH3),
2.60–2.75 (1H, m, CH2CO), 2.75–2.90 (1H, m,
CH2CO), 4.15 (2H, q, J=7.12, OCH2CH3), 4.35–4.40
(1H, m, CHCH2CO), 6.10–6.15 (1H, m, aromatic),
6.25–6.30 (1H, m, aromatic), 7.30–7.35 (1H, m,
aromatic).
3.1.7. Ethyl 3-amino-3-(3-thienyl)propanoate, 3. 1H
NMR (200 MHz):
l 1.25 (3H, t, J=7.12 Hz,
OCH2CH3), 2.50–2.80 (2H, m, CH2CO), 4.15 (2H, q,
J=7.15 Hz, OCH2CH3), 4.45–4.60 (1H, m,
CHCH2CO), 7.07 (1H, d, J=5.06 Hz, aromatic), 7.17
(1H, d, J=2.52 Hz, aromatic), 7.25–7.35 (1H, m,
aromatic).
3.1. Preparation of racemic ethyl 3-amino-3-heteroaryl
propanoates, 1–4
3-Amino-3-heteroarylpropanoic acids were synthesized
in a modified Rodionov synthesis from the correspond-
ing aldehyde through condensation with malonic acid
(1 equiv.) in the presence of ammonium acetate (2
equiv.), the mixture being heated under reflux in
iPrOH:H2O (9:1).30–32 The amino acids were precipi-
tated from the mixture, filtered off and washed with
ethanol. The properties of the products are as follows.
1
3.1.8. Ethyl 3-amino-3-(3-furyl)propanoate, 4. H NMR
(400 MHz): l 1.24 (3H, t, J=7.15 Hz, OCH2CH3),
2.50–2.75 (2H, m, CH2CO), 4.10–4.25 (2H, m,
OCH2CH3), 4.30–4.40 (1H, m, CHCH2CO), 6.30–6.40
(1H, m, aromatic), 7.20–7.30 (1H, m, aromatic), 7.30–
7.40 (1H, m, aromatic).
3.1.1. 3-Amino-3-(2-thienyl)propanoic acid. White crys-
tals, mp 225–228°C, lit. mp:33 207–208°C, 1H NMR
(400 MHz, D2O): l 2.84–3.00 (2H, m, CH2CO), 4.97
(H, t, J=7.00 Hz CHCH2CO), 7.12 (1H, dd, J=3.60,
5.20 Hz, aromatic), 7.26 (1H, d, J=3.60 Hz, aromatic),
7.52 (1H, d, J=4.80 Hz aromatic).
3.2. Gram-scale resolution of ethyl 3-amino-3-(2-
thienyl)propanoate, 1
Racemic 1 (1.0 g, 5.03 mmol) was dissolved in ethyl
butanoate (50 ml), and CAL-A (15 mg/ml) was added.
The mixture was stirred at room temperature. The
reaction was stopped at 51% conversion (ee1a=98%
and ee1b=96%) after 8 h by filtering off the enzyme.
After evaporation, the residue was purified by column
chromatography using CH2Cl2:MeOH (95:5) as an elu-
ent to separate compounds 1a and 1b.
3.1.2. 3-Amino-3-(2-furyl)propanoic acid. Brown crys-
1
tals, mp 219–221°C, lit. mp:34 206°C, H NMR (400
MHz, D2O): l 2.90 (2H, d, J=7.20 Hz, CH2CO), 4.76
(1H, t, J=7.20 Hz, CHCH2CO), 6.47–6.52 (1H, m,
aromatic), 6.52–6.56 (1H, m, aromatic), 7.56–7.60 (1H,
m, aromatic).
Dry HCl gas was bubbled through the solution of 1a
for 2–3 h and the solvent was evaporated. 1a·HCl was
crystallized from diethyl ether in the cold, the filtrate
was recrystallized from a mixture of absolute ethanol
and diethyl ether, and the oily product was washed with
diethyl ether, resulting in (R)-1a·HCl (0.33 g, 1.40
mmol) as a slowly crystallizing light-yellow oil: [h]2D5=
3.1.3. 3-Amino-3-(3-thienyl)propanoic acid. White crys-
1
tals, mp 239–242°C, lit. mp:35 203°C, H NMR (400
MHz, D2O): l 2.81–2.94 (2H, m, CH2CO), 4.77 (1H, t,
J=7.20 Hz, CHCH2CO), 7.22 (1H, dd, J=2.00, 4.80
Hz, aromatic), 7.51–7.56 (2H, m, aromatic).

M. Solyma´r et al. / Tetrahedron: Asymmetry 13 (2002) 2383–2388
2387
+4.6 (c 1.0, MeOH); M=199 according to MS. Anal.
calcd for C9H14ClNO2S: C, 45.86; H, 5.99; Cl, 15.04; N,
5.94; S, 13.60. Found: C, 45.45; H, 6.30; Cl, 16.18; N,
3.4.1. (R)-3a·HCl. Yield: 0.52 g, 2.21 mmol; white
crystals, mp 106–108°C, [h]2D5=+1.00 (c 1.0, MeOH);
M=199 according to MS. Anal. calcd for
C9H14ClNO2S: C, 45.86; H, 5.99; Cl, 15.04; N, 5.94; S,
13.60. Found: C, 48.00; H, 7.07; Cl, 14.03; N, 5.39; S,
1
6.30; S, 15.02%. H NMR (200 MHz): l 1.21 (3H, t,
J=7.13 Hz, OCH2CH3), 3.15–3.35 (2H, m,
CHCH2CO), 4.14 (2H, q, J=7.14 Hz, OCH2CH3),
4.95–5.10 (1H, m, CHCH2CO), 6.95–7.05 (1H, m, aro-
matic), 7.25–7.40 (2H, m, aromatic), 8.90 (3H, br s,
1
12.42%. H NMR (200 MHz): l 1.20 (3H, t, J=7.13
Hz, OCH2CH3), 2.95–3.35 (2H, m, CH2CO), 4.12 (2H,
q, J=7.13 Hz, OCH2CH3), 4.75–4.90 (1H, m,
CHCH2CO), 7.25–7.35 (2H, m, aromatic), 7.52–7.58
NH3 Cl−) ppm.
+
(1H, m, aromatic), 8.82 (3H, br s, NH3 Cl−) ppm.
+
After evaporation of the eluent, (S)-1b (0.62 g, 2.30
mmol) was obtained as a yellow oil: [h]2D5=−84.2 (c 1.0,
MeOH); M=269 according to MS. Anal. calcd for
C13H19NO3S: C, 57.97; H, 7.11; N, 5.20; S, 11.90.
3.4.2. (S)-3b. Yield: 0.69 g, 2.56 mmol; an orange oil,
[h]2D5=−83.1 (c 1.0, MeOH); M=269 according to MS.
Anal. calcd for C13H19NO3S: C, 57.97; H, 7.11; N, 5.20;
S, 11.90. Found: C, 56.12; H, 7.59; N, 5.75; S, 11.55%.
1H NMR (200 MHz): l 0.96 (3H, t, J=7.33 Hz,
CH3CH2CH2), 1.20 (3H, t, J=6.17 Hz, OCH2CH3),
1.55–1.80 (2H, m, CH3CH2CH2), 2.20 (2H, t, J=7.46
Hz, CH3CH2CH2), 2.75–3.00 (2H, m, CH2CO), 4.10
(2H, q, J=7.12 Hz, OCH2CH3), 5.40–5.60 (1H, m,
CHCH2CO), 6.50–6.65 (1H, d, J=8.16 Hz, NH), 6.98–
7.05 (1H, m, aromatic), 7.05–7.15 (1H, m, aromatic),
7.30–7.35 (1H, m, aromatic) ppm.
1
Found: C, 60.46; H, 8.01; N, 5.72; S, 13.56%. H NMR
(400 MHz): l 0.94 (3H, t, J=7.37 Hz, CH3CH2CH2),
1.19 (3H, t, J=7.15 Hz, OCH2CH3), 1.55–1.75 (2H, m,
CH3CH2CH2), 2.18 (2H, t, J=7.48 Hz, CH3CH2CH2),
2.80–3.00 (2H, m, CH2CO), 5.65–5.75 (1H, m,
CHCH2CO), 6.62 (1H, d, J=8.55 Hz, NH), 6.85–6.95
(1H, m, aromatic), 7.15–7.20 (2H, m, aromatic) ppm.
3.3. Gram-scale resolution of ethyl 3-amino-3-(2-
furyl)propanoate, 2
3.5. Gram-scale resolution of ethyl 3-amino-3-(3-
furyl)propanoate, 4
Racemic 2 (1.0 g, 5.46 mmol) was dissolved in ethyl
butanoate (109 ml), and CAL-A (15 mg/ml) was added.
The reaction was stopped at 51% conversion (ee2a=
96% and ee2b=93%) after 7 h by filtering off the
enzyme, followed by the work-up as above.
Racemic 4 (0.5 g, 2.73 mmol) was dissolved in ethyl
butanoate (27 ml) and CAL-A (15 mg/ml) was added.
The reaction was stopped at 50% conversion (ee4a >99%
and ee4b=99%) after 4 h by filtering off the enzyme,
followed by the work-up as above.
3.3.1. (R)-2a·HCl. Yield: 0.45 g, 2.05 mmol; a slowly
crystallizing brown oil, [h]2D5=+9.5 (c 1.0, MeOH); M=
183 according to MS. Anal. calcd for C9H14ClNO3: C,
49.21; H, 6.42; Cl, 16.14; N, 6.38. Found: C, 48.16; H,
3.5.1. (R)-4a·HCl. Yield: 0.30 g, 1.36 mmol; a slowly
crystallizing brown oil, [h]2D5=+7.25 (c 1.0, MeOH);
M=183 according to MS. Anal. calcd for C9H14ClNO3:
C, 49.21; H, 6.42; Cl, 16.14; N, 6.38. Found: C, 49.70;
1
6.86; Cl, 15.42; N, 6.68%. H NMR (200 MHz): l 1.34
(3H, t, J=7.13 Hz, OCH2CH3), 3.15–3.50 (2H, m,
CH2CO), 4.26 (2H, q, J=7.13 Hz, OCH2CH3), 4.85–
5.15 (1H, m, CHCH2CO), 6.40–6.50 (1H, m, aromatic),
6.60–6.80 (1H, m, aromatic), 7.45–7.65 (1H, m, aro-
1
H, 7.72; Cl, 13.83; N, 5.64%. H NMR (400 MHz): l
1.20 (3H, t, J=7.16 Hz, OCH2CH3), 2.58–3.10 (1H, m,
CH2CO), 3.10–3.25 (1H, m, CH2CO), 4.00–4.25 (2H,
m, OCH2CH3), 4.65–4.80 (1H, m, CHCH2CO), 6.00–
6.75 (1H, m, aromatic), 7.30–7.40 (1H, m, aromatic),
matic), 9.03 (3H, br s, NH3 Cl−) ppm.
+
7.55–7.65 (1H, m, aromatic), 8.74 (3H, br s, NH3 Cl−)
+
3.3.2. (S)-2b. Yield: 0.53 g, 2.09 mmol; an orange oil,
[h]2D5=−84.5 (c 1.0, MeOH); M=253 according to MS.
Anal. calcd for C13H19NO4: C, 61.64; H, 7.56; N, 5.53.
ppm.
1
Found: C, 62.43; H, 8.95; N, 6.34%. H NMR (200
3.5.2. (S)-4b. Yield: 0.19 g, 0.75 mmol; a yellow oil,
[h]2D5=−57.1 (c 1.0, MeOH); M=253 according to MS.
Anal. calcd for C13H19NO4: C, 61.64; H, 7.56; N, 5.53.
MHz): l 0.95 (3H, t, J=7.37 Hz, CH3CH2CH2), 1.23
(3H, t, J=7.13 Hz, OCH2CH3), 1.60–1.80 (2H, m,
CH3CH2CH2), 2.20 (2H, t, J=7.20 Hz, CH3CH2CH2),
2.70–3.00 (2H, m, CH2CO), 4.12 (2H, q, J=7.07 Hz,
OCH2CH3), 5.45–5.65 (1H, m, CHCH2CO), 6.15–6.20
(1H, m, aromatic), 6.25–6.35 (1H, m, aromatic), 6.44
(1H, d, J=8.34 Hz, NH), 7.30–7.35 (1H, m, aromatic)
ppm.
1
Found: C, 59.74; H, 8.65; N, 6.45%. H NMR (200
MHz): l 0.96 (3H, t, J=7.35 Hz, CH3CH2CH2), 1.23
(3H, t, J=7.12 Hz, OCH2CH3), 1.60–1.80 (2H, m,
CH3CH2CH2), 2.19 (2H, t, J=7.44 Hz, CH3CH2CH2),
2.70–2.95 (2H, m, CH2CO), 4.13 (2H, q, J=7.09 Hz,
OCH2CH3), 5.30–5.48 (1H, m, CHCH2CO), 6.31–6.38
(1H, m, aromatic), 6.49 (1H, d, J=8.49 Hz, NH),
7.30–7.45 (2H, m, aromatic) ppm.
3.4. Gram-scale resolution of ethyl 3-amino-3-(3-
thienyl)propanoate, 3
3.6. Reductive desulphurization of 1
Racemic 3 (1.0 g, 5.03 mmol) was dissolved in ethyl
butanoate (100 ml), and CAL-A (15 mg/ml) was added.
The reaction was stopped at 51% conversion (ee3a >99%
and ee3b=96%) after 5 h by filtering off the enzyme,
followed by the work-up as above.
Racemic 1 (0.3 g, 1.5 mmol) was dissolved in water
with the addition of a few drops of ethanol and Raney
nickel (4 g) was suspended. The mixture was stirred at
70°C under 40 atm H2 pressure in an autoclave. After

2388
M. Solyma´r et al. / Tetrahedron: Asymmetry 13 (2002) 2383–2388
4 days the catalyst was filtered off and the solvent was
Shishkina, I. P.; Galushko, S. V.; Sorochinsky, A. E.;
Kukhar, V. P.; Savchenko, M. V.; Svedas, V. K. Tetra-
hedron: Asymmetry 1995, 6, 1601–1610.
evaporated off. A white solid of 3-aminoheptanoic acid
1
formed which was crystallized from acetone. H NMR
(400 MHz, D2O): l 0.90 (3H, t, J=7.00 Hz, CH3),
1.25–1.45 (4H, m, CH2CH2CH3), 1.55–1.75 (2H, m,
CH2CH), 2.35–2.50 (H, m, CH2CO), 2.50–2.65 (H, m,
CH2CO), 3.43–3.52 (H, m, CHCH2CO).
16. Cardillo, G.; Gentilucci, L.; Tolomelli, A.; Tomasini, C.
J. Org. Chem. 1998, 63, 2351–2353.
17. Cardillo, G.; Tolomelli, A.; Tomasini, C. Eur. J. Org.
Chem. 1999, 155–161.
18. Roche, D.; Prasad, K.; Repic, O. Tetrahedron Lett. 1999,
40, 3665–3668.
Reductive desulphurization of 1a (R-configuration
expected) was carried out under the conditions applied
for the racemic compound. [h]2D0=+12.4 (c 0.8, H2O)
was determined for the resulting (S)-3-aminoheptanoic
acid (the reduction product contained some 20% impu-
rities, but the characteristic lines for 3-aminoheptanoic
acid were identical with the 1H NMR spectrum of
racemate) in good accordance with the literature value25
for (S)-3-aminoheptanoic acid: [h]2D2=+33.3 (c 0.91,
H2O), ee >90%.
19. Hoekstra, W. J.; Maryanoff, B. E.; Andrade-Gordon, P.;
Cohen, J. H.; Costanzo, M. J.; Damiano, B. P.;
Haertlein, B. J.; Harris, B. D.; Kauffman, J. A.; Keane,
P. M.; McComsey, D. F.; Villani, F. J., Jr.; Yabut, S. C.
Bioorg. Med. Chem. Lett. 1996, 6, 2371–2376.
20. Hoekstra, W. J.; Maryanoff, B. E.; Damiano, B. P.;
Andrade-Gordon, P.; Cohen, J. H.; Costanzo, M. J.;
Haertlein, B. J.; Hecker, L. R.; Hulshizer, B. L.; Kauff-
man, J. A.; Keane, P.; McComsey, D. F.; Mitchell, J. A.;
Scott, L.; Shah, R. D.; Yabut, S. C. J. Med. Chem. 1999,
42, 5254–5265.
21. (a) Nielsen, T. B.; Ishii, M.; Kirk, O. Biotechnol. Appl.
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Acknowledgements
M.S. is grateful for a grant from the Centre for Interna-
tional Mobility (CIMO) in Finland. We also thank the
Hungarian Research Foundation (OTKA) for financial
support.
22. Rogalska, E.; Cudrey, C.; Ferrato, F.; Verger, R. Chiral-
ity 1993, 5, 24–30.
23. Liljeblad, A.; Lindborg, J.; Kanerva, A.; Katajisto, J.;
Kanerva, L. T. Tetrahedron Lett. 2002, 43, 2471–2474.
24. Solymar, M.; Liljeblad, A.; La´za´r, L.; Fu¨lo¨p, F.;
Kanerva, L. T. Tetrahedron: Asymmetry 2002, 13, 1923–
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