Studies on the chemoenzymatic synthesis of 3-phenyl-GABA and 4-phenyl-pyrrolid-2-one: The influence of donor of the alkoxy group on enantioselective esterification

Article abstract of DOI:10.1016/j.tetasy.2013.02.016

A new chemoenzymatic method for the synthesis of enantiomerically pure 3-phenyl-γ-aminobutyric acid 1 and 4-phenyl-pyrrolid-2-one 9 based on the enzymatic kinetic resolution of 3-phenyl-4-pentenoic acid 2 is described herein. Enzymatic resolution of the r

Full text of DOI:10.1016/j.tetasy.2013.02.016

Tetrahedron: Asymmetry 24 (2013) 427–433
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Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
Studies on the chemoenzymatic synthesis of 3-phenyl-GABA
and 4-phenyl-pyrrolid-2-one: the influence of donor of the
alkoxy group on enantioselective esterification
⇑
Anna Brodzka, Dominik Koszelewski, Malgorzata Cwiklak, Ryszard Ostaszewski
Institute of Organic Chemistry, PAS, Kasprzaka 44/52, 01-224 Warsaw, Poland
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 12 November 2012
Accepted 20 February 2013
A new chemoenzymatic method for the synthesis of enantiomerically pure 3-phenyl-c-aminobutyric acid
1 and 4-phenyl-pyrrolid-2-one 9 based on the enzymatic kinetic resolution of 3-phenyl-4-pentenoic acid
2 is described herein. Enzymatic resolution of the racemic substrate provided products with good enanti-
oselectivity upon esterification. In these reactions, a new class of alkoxy group donor—orthoesters, acetals
and ketals were used. The best results of the enzymatic kinetic resolution were obtained for triethyl
orthoacetate in toluene solvent.
Crown Copyright Ó 2013 Published by Elsevier Ltd. All rights reserved.
1. Introduction
3-Phenyl-4-pentenoic acid is a substrate for the synthesis of 4-
phenyl-pyrrolid-2-one 9. This class of compounds are selective
3-PhenylGABA (phenibut) 1 is a lipophilic analogue of GABA
-aminobutyric acid), the main neurotransmitter of the central
inhibitors of cAMP phosphodiesterase. They are used in the ther-
(c
apy of ischaemic heart disease.11 Compound 9 is a precursor for
nervous system layout (CNS).1 The disorders in the synthesis of
GABA in an organism often lead to epilepsy, as well as other dis-
eases such as Huntington’s disease or Parkinson’s disease2 and
other psychiatric disorders such as anxiety and pain. The direct
application of GABA in treatment therapy is limited due to its
hydrophilic nature. GABA has a low permeability through the
blood–brain barrier.3 Phenibut was discovered and introduced into
clinical practice in Russia in the 1960s. The biological activity of
this compound depends on its absolute configuration. The (R)-iso-
mer is much more active than the (S)-isomer, and it is twice as po-
tent as the racemate.4
the synthesis of c-aminobutyric acid (GABA) analogues, which
are of great interest due to their importance in various nervous
system functions
2. Results and discussion
We focused our attention on a novel irreversible method for the
enzymatic esterification of carboxylic acids. We used orthoesters,
acetals and ketals as new donors of alkoxy groups. The mechanism
of this reaction is shown on Scheme 1. The racemic carboxylic acid
E reacts with the hydroxyl group of enzyme C to form the
acylenzyme intermediate D and a water molecule. In the next step
the intermediate D reacts with alcohol B to give ester A. The alco-
hol B derives from the reaction between water and compound F to
form compound G, which in the next step is transformed into com-
pound H and alcohol B. The last step is irreversible.
Herein our aim was to develop enzymatic methods for the
enantioselective synthesis of ester derivatives of 3-phenyl-4-
pentenoic acid. In our previous study, we looked at the enzymatic
esterification of 3-phenyl-4-pentenoic acid using triethyl orthoac-
etate as a donor of the alkoxy group.12 The results obtained were
very promising so we decided to expand upon this methodology
using the other donors of alkoxy groups and enzymes.
Previously, enantiomerically pure phenibut 1 has been synthe-
sized via chemical methods from
a,b-unsaturated oxazolines 3
(Fig. 1).5 There are several chemical routes in which asymmetric
catalysts are used (with substrates 4,6 57 and 68).
There are also some examples of the enzymatic method of syn-
thesis of enantiomerically pure 3-phenyl-GABA. Wang et al. de-
scribed chemoenzymatic preparation of phenibut (85% ee) via
hydrolysis of 3-phenylglutaronitrile 7 catalysed by Rhodococcus
sp. AJ270 followed by Curtius rearrangement and consequent
acidic hydrolysis.9 Felluga et al. prepared compound 1 via enzy-
matic hydrolysis of racemic
c
-nitroester 8 with
a
-chymotrypsin
and subsequent reduction with Raney Nickel.10
At first, the racemic esters 11 and 13 were obtained in only 11%
and 3% yields, respectively (Scheme 2).13 In this reaction, a trialkyl
orthoester was used. The addition of an orthoester shifts the
equilibrium in favour of the ester side, due to the consumption
of water formed through hydrolysis of the orthoester.
⇑
Corresponding author. Tel.: +48 22 343 2120.
E-mail
addresses:
ryszard.ostaszewski@icho.edu.pl,
rysza@icho.edu.pl
(R. Ostaszewski).
0957-4166/$ - see front matter Crown Copyright Ó 2013 Published by Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetasy.2013.02.016

428
A. Brodzka et al. / Tetrahedron: Asymmetry 24 (2013) 427–433
O
Ph
NO2
PhO
CO2tBu
N
CHO
4
5
O
6
CN
O
N
NH2
CN
3
Ph
OH
1
7
O
COOH
COOMe
NH
2
8
O
NO2
9
Figure 1. Substrates for the synthesis of 3-phenylGABA.
OR1
R1O
R3
R1 = Me, Et
RCOOH
H2O
R2 = H, Me
E
R3 = H, Me, OMe, OEt
R2
F
O
Enz-OH
C
Enz-O
R
D
O
OR1
R3
HO
R1OH
B
R
OR1
A
R2
G
O
R1OH
+
R2
R3
B
H
Scheme 1. The proposed mechanism for the enzymatic esterification.
R1
O
O
O
O
toluene, 110°C
R1
OR1
+
OH
R
O
R1
2
11: R1 = Et
13:
R1 = Me
1
10:
R = Et
a: R = CH3
b:
R = H
12: R1 = Me
a:
R = CH3
b: R = H
Scheme 2. Synthesis of racemic esters of 3-phenyl-4-pentenoic acid.

A. Brodzka et al. / Tetrahedron: Asymmetry 24 (2013) 427–433
429
2.1. Comparison of various solvents
(Table 2, entries 3–11 and 13–15), while triethyl orthoformate
10b gives a reasonable enantioselectivity for only one enzyme
(Table 2, entry 18). However, the results obtained using trimethyl
orthoacetate 12a and trimethyl orthoformate 12b as donors, reveal
very low enantioselectivity despite a twofold time extension
(Table 2, entries 22–29).
It is noteworthy that using trimethyl orthoformate as a donor of
the methoxy group and GLAP as a biocatalyst (Table 2, entry 29),
the absolute configuration of the product was opposite (R). It is
interesting to note that simple change of the alkoxy group donor
can reverse the stereochemical course of the enzymatic kinetic
resolution.
Previously, we have found that the best biocatalyst for the enzy-
matic esterification was lipase from Wheat germ.12 It is well docu-
mented that the solvent has a strong influence on biocatalytic
reactions. In order to determine the best solvent for the kinetic res-
olution of 3-phenyl-4-pentenoic acid 2, different solvents listed in
Table 1, were used. These reactions, catalysed by lipase from
Wheat germ, were conducted in an organic solvent at 40 °C for
24 h. Triethyl orthoacetate was used as a donor of the alkoxy
group.
The experimental data show that the best enantioselectivity
was obtained for toluene (Table 1, entry 2). Tetrahydrofuran,
dimethylformamide and acetonitrile (Table 1, entries 5, 10 and
11) also exhibited good enantioselectivity but the substrate con-
version was substantially lower.
2.3. Acetals and ketals as donors of alkoxy group
Based on the mechanism of enzyme catalysed reaction, we
noted that another class of alkoxy group donors for enzymatic
esterification reaction can be used. Therefore, in the next stage of
our studies, we used acetals 14 and ketals 15 (Fig. 2). The reactions
were conducted in toluene at 40 °C for 24–48 h. The results ob-
tained are shown in Table 3.
The results show that the acetal donors of ethoxy group 14a are
better than methoxy 14b and 15, as in the case of orthoesters. In
some cases, 1,1-diethoxyethane 14a gave good enantioselectivity
(Table 3, entries 4, 7 and 10). 1,1-Dimethoxyethane 14b and 2,2-
dimethoxypropane 15 exhibited lower enantioselectivity, however
these values are acceptable for several enzymes (Table 3, entries
13, 14 and 16).
2.2. Enantioselective synthesis of esters of 3-phenyl-4-
pentenoic acid using various trialkyl orthoesters
Herein we have focused our attention on the synthesis of chiral
non-racemic esters of 3-phenyl-4-pentenoic acid. We decided to
test other orthoesters 10b, 12a, and 12b as donors of an alkoxy
group. In our studies commercially available lipases were used as
biocatalysts as well as acetone powders [beef liver acetone powder
(BLAP), turkey liver acetone powder (TLAP), chicken liver acetone
powder (CLAP), deer liver acetone powder (JLAP), beef kidney ace-
tone powder (BKAP) and goose liver acetone powder (GLAP)].
These reactions were conducted in toluene at 40 °C for 24–48 h
(Scheme 3). The results obtained are shown in Table 2.
Comparing the results obtained for orthoesters and acetals we
noticed that orthoesters are better donors of alkoxy group for the
esterification of carboxylic acids than acetals.
The results show that triethyl orthoesters 10 are better donors
of the alkoxy group than trimethyl orthoesters 12. In most cases
triethyl orthoacetate 10a exhibits very good enantioselectivity
2.4. Synthesis of 3-phenylGABA
The retrosynthetic analysis of 3-phenylGABA is shown in
Figure 3.
Table 1
Results obtained in enzymatic esterification 3-phenyl-4-pentenoic acid in different
3-PhenylGABA A can be obtained by functional group intercon-
version (FGI) from the corresponding azide B synthon obtained di-
rectly from the corresponding alcohol C. The hydroxyl group in
alcohol C can be obtained via functional group interconversion of
the double bond present in the 3-phenyl-4-pentenoic acid 2
structure.15
Stereodifferentiation was planned to take place during the
enzymatic functionalization of acid 2. For the synthesis of 3-phe-
nylGABA, the obtained ethyl ester (R)-11 was used (Scheme 4).
Ozonolysis of the ester (R)-11 double bond gives an aldehyde 16
(40% yield). Using sodium cyanoborohydride to reduce the car-
bonyl group did not lead to a desirable product. After many trials,
we found that sodium cyanoborohydride reduces the aldehyde 16
efficiently and provides alcohol 17 (76% yield). In the next step, the
reaction of alcohol 17 with methanesulfonic acid chloride led to
the formation of compound 18 in 90% yield. Compound 18 was
converted into azide 19 in 66% yield whose hydrogen/palladium
reduction gave the respective amine 20 in 58% yield. The last stage
solventsa
Entry
Solvent
Product
c (%)
eeb (%)
Ec
1
2
3
4
5
6
7
8
—
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
30.5
45
19.5
27
16
65
2.5
9.7
67
3.7
16
69
93
52
3.7
90
50
21
45
41
91
96
14
7
63
4
Toluene
Hexane
Chloroform
THF
Dichloromethane
Ethyl ether
Dioxane
tert-Butyl-methyl ether
DMF
Acetonitrile
DMSO
1
23
10
2
3
6
23
53
1
9
10
11
12
38.5
a Conditions: 2 (1 mmol), 10a (3 mmol), solvent (1.21 mL), lipase from Wheat germ
(10 mg), 40 °C, 24 h.
b
Determined by HPLC on a Daicel Chiracel OD-H column.
Calculated
c
according
to
Chen
et
al.,14
using
the
equation:
E = (ln[1 ꢀ c(1 + eep)])/(ln[1 ꢀ c(1 ꢀ eep)]).
O
OR
O
O
OR
OR
toluene, 40°C, 24-48h
enzyme
R1
OR +
+
OH
OH
2
10a: R = Et, R1 = CH3
1
10b:
(R)-2
R = Et, R = H
(S)-11: R = Et
12a: R = Me, R1 = CH3
13:
(S)-
R = Me
1
12b:
R = Me, R = H
Scheme 3. Enzymatic kinetic resolution of compound 2 using different trialkyl orthoesters.

430
A. Brodzka et al. / Tetrahedron: Asymmetry 24 (2013) 427–433
Table 2
Kinetic resolution of compound 2 using orthoestersa
Entry
Enzyme
Donor of the alkoxy group
Time (h)
Product
c (%)
% eeb
Ec
1
2
3
4
5
6
7
8
—
10a
10a
10a
10a
10a
10a
10a
10a
10a
10a
10a
10a
10a
10a
10a
10b
10b
10b
10b
10b
10b
12a
12a
12a
12a
12b
12b
12b
12b
24
24
24
24
24
24
24
24
24
24
24
24
24
24
24
24
24
24
24
24
24
48
48
48
48
48
48
48
48
11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
11
(S)-11
(S)-11
(S)-11
11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
2
11
50
52
23
0
0
—
—
Hog pancreas
Penicilium roqueforti
Aspergillus
Wheat germ
Pseudomonas cepacia
Pseudomonas sp
Mucor miehei
Candida rugosa
BLAP
TLAP
CLAP
JLAP
BKAP
90
92
93
92
93
91
90
90
82
—
83
93
97
0
68
89
71
27
44
—
776
34
63
30
50
40
29
26
41
—
45
15.4
37.5
40.9
30.1
24
54
9.3
52
25.7
25
3
12.2
5.9
43
3.2
6
0
17
9
5.8
8.9
0
9.2
18.5
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
32
37
82
—
GLAP
—
Aspergillus
Pseudomonas cepacia
Wheat germ
Mucor miehei
Candida rugosa
—
Wheat germ
Penicillium roqueforti
GLAP
6
19
10
2
3
—
1
11
1
—
—
(S)-13
(S)-13
(S)-13
13
0.5
82
1.5
0
—
82
16
—
Wheat germ
Penicillium roqueforti
GLAP
2
(S)-13
(R)-13
11
1.4
a
Conditions: 2 (1 mmol), 10/12 (3 mmol), toluene (1.21 mL), the enzyme (10 mg), 40 °C, 24–48 h.
b
Determined by HPLC on a Daicel Chiracel OD-H column.
c
Calculated according to Chen et al.,14 using the equation: E = (ln[1 ꢀ c(1 + eep)])/(ln[1 ꢀ c(1 ꢀ eep)]).
was the acidolysis of the ester group. In this reaction we obtained
the desired product as hydrochloride 21 in 89% yield.
R2
OR
OR
The assignment of the absolute configuration of the 3-phenylG-
ABA stereogenic centre was based on the sign of specific rotation of
hydrochloride 21. The absolute configuration at carbon 3 was as-
signed as (S) by comparison of the specific rotation with the fol-
R1
1
2
14a:
R = Et, R = CH3, R = H
14b: R = Me, R1 = CH3, R2 = H
lowing literature value: [
a
]
D = ꢀ2.6 (c 0.86, MeOH), [(S)-isomer].7
1
2
15
: R = Me, R = R = CH3
It is noteworthy that amine 20 was converted into (S)-(+)-4-
phenyl-pyrrolid-2-one 9 with 60% yield. Compound 9 is an impor-
tant substance for the study of the nervous system function.11
Figure 2. Selected donors of the alkoxy group.
Table 3
Enzymatic kinetic resolution of 2 using acetals and ketalsa
Entry
Enzyme
Donor of alkoxy group
Time (h)
Product
c (%)
% eeb
Ec
1
2
3
4
5
6
7
8
—
14a
14a
14a
14a
14a
14a
14a
14a
14a
14a
14a
14a
14b
14b
14b
15
24
24
24
24
24
24
24
24
24
24
24
24
48
48
48
48
48
48
11
(S)-11
(R)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
(S)-11
11
1
0
46
33
>99
16
30
98
12
18
98
0
—
3
2
202
1
2
106
1
2
104
—
—
16
19
4
19
4
Wheat germ
P. roqueforti
Aspergillus
6.2
6.7
1.5
6.6
23
Papaine
Hog pancreas
Rhizopus arrhizus
Candida rugosa
Mucor javanicus
Rhizomucor miehei
Novozym
6.7
25
9
5.8
4.4
2.5
2.8
2.6
1.3
1.6
11.3
1.1
1.1
10
11
12
13
14
15
16
17
18
GLAP
11
0
Wheat germ
Penicillium roqueforti
GLAP
Wheat germ
Penicillium roqueforti
GLAP
(S)-13
(S)-13
(S)-13
(S)-13
(R)-13
(R)-13
88
90
58
89
57
72
15
15
6
a
Conditions: 2 (1 mmol), 14/15 (3 mmol), toluene (1.21 mL), the enzyme (10 mg), 40 °C, 24–48 h.
b
Determined by HPLC on a Daicel Chiracel OD-H column.
c
Calculated according to Chen et al.,14 using the equation: E = (ln[1 ꢀ c(1 + eep)])/(ln[1 ꢀ c(1 ꢀ eep)]).

A. Brodzka et al. / Tetrahedron: Asymmetry 24 (2013) 427–433
431
NH2
N3
OH
OH
OPG
OPG
OH
O
O
O
O
2
B
C
A
Figure 3. Retrosynthetic analysis of 3-phenylGABA.
1. O3/DCM
-78°C
1. NaBH3CN
AcOEt/AcOH
MeSO2Cl
O
OH
2. NH4Cl
76%
2. Me2S, -10°C
40%
Et3N/DCM
90%
OEt
OEt
OEt
O
11
O
O
(+)-17
(R)-(+)-
16
1. H2/Pd.C
EtOH
SO2
NH3Cl
OH
20: 58%
N3
DABCO/NaN3
O
OEt
OEt
DCM, reflux
66%
2. 6M HCl/AcOH
89%
O
O
O
18
(+)-
(S)-(-)-21
(-)-19
1. H2/Pd.C
EtOH
20: 58%
NH
O
2. Et3N/PhH
reflux
9
(S)-(+)-
60%
Scheme 4. Formal synthesis of 3-phenylGABA.
a pre-column (4 mm ꢁ 10 mm, 5 m) using an LC-6A Varian ProStar
apparatus with UV Varian ProStar 330 detector and Chromatopac
C-R6A analyzer. The elemental analyses were performed on CHN
Perkin–Elmer 240 apparatus. All reactions were monitored by
TLC on Merck silica gel Plates 60 F254. The lipases were purchased
from Fluka. Novo sp 435A was purchased from Novo Industri A/S.
The acetone powders were prepared in our laboratory.17 All of
the chemicals were obtained from commercial chemical sources.
The solvents were of analytical grade.
3. Conclusion
We have studied the enzyme-catalysed synthesis of enantiome-
rically pure ester derivates of 3-phenyl-4-pentenoic acid. Our re-
sults demonstrate that several enzymes catalysed these reactions
with excellent enantioselectivity, when we used triethyl orthoace-
tate as the donor of the alkoxy group. Trialkyl orthoesters were
used previously in the kinetic resolution of racemic carboxylic
acids, catalysed by enzymes, although this reactions required a
few days and the enantiomeric excesses were lower.16 We proved
that other donors such acetals and ketals can be used in kinetic res-
olution of racemic 3-phenyl-4-pentenoic acid. In most cases, the
proper choice of the enzyme and the alkoxy group donor is re-
quired to achieve excellent enantioselectivity.
The solvent effect in the reaction studied was investigated in an
attempt to find suitable reaction conditions; the best result was
obtained in toluene.
We have also provided a novel and convenient chemoenzymatic
method for the preparation of biologically active compounds: 3-
phenylGABA 1 and 4-phenyl-pyrrolid-2-one 9. These two six-step
syntheses have been developed starting from the readily available
precursor (R) ethyl-3-phenyl-4-pentenoate 11, through an efficient
enzymatic kinetic resolution.
4.2. Synthesis of 3-phenyl-4-pentenoic acid 2
3-Phenyl-4-pentenoic acid was synthesized according to the
method described by Bermejo et al.18 A mixture of cinnamyl alco-
hol (0.25 mol; 33.7 g), triethyl orthoacetate (0.25 mol; 46.1 mL)
and hexanoic acid (1.5 mmol; 0.19 mL) was heated in an oil bath.
The solution was placed in a flask with a Claisen head (for distilling
off ethanol), condenser and thermometer. After 3 h, 0.1 mL of hex-
anoic acid was added. An additional portion (0.1 mL) of hexanoic
acid was added at 3.5 and 4.5 h. After 6 h the temperature rose
to 166 °C. After this time, 27 mL of ethanol were distilled off and
TLC analysis indicated that no cinnamyl alcohol remained.
The mixture was cooled, and a solution of potassium hydrox-
ide (0.35 mol; 19.7 g) in water (25 mL), and methanol (75 mL)
were added. The mixture was heated at reflux for 1 h under
nitrogen. After cooling, the solution was washed with ethyl ether
and acidified with concentrated HCl to pH 1. The acidic solution
was extracted with ethyl ether (3ꢁ) and the organic layer was
dried over MgSO4, filtered and concentrated under reduced
pressure.
4. Experimental
4.1. General
The HPLC analyses were performed on a Chiracel OD-H column
(4.6 mm ꢁ 250 mm, from Diacel Chemical Ind., Ltd) equipped with

432
A. Brodzka et al. / Tetrahedron: Asymmetry 24 (2013) 427–433
The crude acid was purified by crystallization from hexane (75%
crude product was purified by silica gel flash chromatography
using hexane/ethyl acetate (99.5/0.5; v/v) as an eluent to afford
the corresponding product 13 as a colourless oil; Rf = 0.5 (hex-
ane/EtOAc, 9:1; v/v); HPLC analysis [hexane/iPrOH; 9:1;
k = 232 nm; 1.0 mL/min] tR(S) = 4.70 min; tR(R) = 6.70 min.
yield) to give a product that melted at 46 °C.10 1H NMR (200 MHz,
CDCl3): 2.74 (dd, J = 15.0, 7.5 Hz, 1H), 2.78 (dd, J = 15.0, 7.8 Hz, 1H),
3.90 (q, J = 7.2 Hz, 1H), 5.00–5.18 (m, 2H), 5.90–6.10 (m, 1H), 7.19–
7.40 (m, 5H); 13C NMR (200 MHz, CDCl3): 40.2, 45.5, 115.3, 127.1,
127.8, 128.9, 140.2, 142.2, 178.3.
4.7. Enantioselective synthesis of ethyl 3-phenyl-4-pentenoate
11 in different solvents
4.3. Synthesis of racemic ethyl 3-phenyl-4-pentenoate 11
Compound 11 was synthesized as described by Gopalan.13 To a
solution of 3-phenyl-4-pentenoic acid (1 mmol) in toluene, the
corresponding triethyl orthoester 10 (3 mmol) was added. The
reaction mixture was heated at reflux (110 °C) for 24 h.
After cooling, 2 M HCl was added (1.1 mL). The organic layer
was washed with saturated NaHCO3 (1ꢁ) and brine (1ꢁ), and dried
over anhydrous MgSO4. The excess reagents and solvent were
evaporated under vacuum.
The crude product was purified by silica gel flash chromatogra-
phy using hexane/ethyl acetate (99.5/0.5; v/v) as an eluent to af-
ford the corresponding product 11 as a colourless oil; Rf = 0.8
(hexane/EtOAc, 8:2; v/v); HPLC analysis [hexane/iPrOH; 9:1;
k = 232 nm; 1.0 mL/min] tR(S) = 9.77 min; tR(R) = 11.03 min.
To a solution of 3-phenyl-4-pentenoic acid (1 mmol) in an or-
ganic solvent, triethyl orthoacetate 10a (3 mmol) and Lipase from
Wheat germ (10 mg) were added. The reaction mixture was then
stirred for 24 h at 40 °C. After cooling, 2 M HCl was added
(1.1 mL). The organic layer was washed with saturated NaHCO3
(1ꢁ) and brine (1ꢁ), and dried over anhydrous MgSO4. The excess
reagents and solvent were evaporated under vacuum. The crude
product was purified by silica gel flash chromatography using hex-
ane/ethyl acetate (99.5/0.5; v/v) as an eluent to afford product 11
as a colourless oil; Rf = 0.8 (hexane/EtOAc, 8:2; v/v); HPLC analysis
[hexane/iPrOH; 9:1; k = 232 nm; 1.0 mL/min] tR(S) = 9.77 min;
tR(R) = 11.03 min.
4.8. Synthesis of the ethyl ester of 3-phenyl-4-oxobutyric acid
16
4.4. Enantioselectivity synthesis of ethyl 3-phenyl-4-pentenoate
11
A solution of the ethyl ester of (R)-(+)-3-phenyl-4-pentenoic
acid (12.2 mmol) in dry dichloromethane was cooled to ꢀ78 °C.
Next, this solution was saturated by ozone for 30 min in ꢀ78 °C.
Next, air was blown for 5 min and dimethyl sulfide (1 mL) was
added. After evaporating the excess solvent, the crude product
was purified by silica gel flash chromatography using hexane/ethyl
acetate as an eluent to afford product 16 as a colourless oil (40%
yield). 1H NMR (200 MHz, CDCl3): d 1.21 (t, J = 7.12 Hz, 3H), 2.60
(dd, J = 7.0, 15.0 Hz, 1H), 3.18 (dd, J = 7.0, 15.0 Hz, 1H), 3.94–4.40
(m, 2H), 5.10 (dd, J = 7.0, 15.0 Hz, 1H), 7.10–7.58 (m, 5H), 9.60 (s,
1H); 13C NMR (50 MHz, CDCl3): d 14.4, 34.9, 54.9, 61.1, 127.8,
128.3, 128.8, 129.0, 129.1, 129.5, 171.8, 198.8; ESI-MS HR,
[M+Na]+, calcd for C12H14O3Na: 229.083, found (m/z) 229.082
(100%).
To a solution of 3-phenyl-4-pentenoic acid (1 mmol) in toluene,
the corresponding triethyl orthoester 10 or acetal 14 (3 mmol) and
enzyme (10 mg) were added. The reaction mixture was stirred for
24 h at 40 °C.
After cooling, 2 M HCl was added (1.1 mL). The organic layer
was washed saturated NaHCO3 (1ꢁ) and brine (1ꢁ), and dried over
anhydrous MgSO4. The excess reagents and solvent were evapo-
rated under vacuum.
The crude product was purified by silica gel flash chromatogra-
phy using hexane/ethyl acetate (99.5/0.5; v/v) as an eluent to af-
ford the corresponding product 11 as a colourless oil; Rf = 0.8
(hexane/EtOAc, 8:2; v/v); HPLC analysis [hexane/iPrOH; 9:1;
k = 232 nm; 1.0 mL/min] tR(S) = 9.77 min; tR(R) = 11.03 min.
4.5. Synthesis of racemic methyl 3-phenyl-4-pentenoate 13
4.9. Synthesis of the ethyl ester of (+)-3-phenyl-4-
hydroxybutyric acid 17
Compound 13 was synthesized as described by Gopalan.13 To a
solution of 3-phenyl-4-pentenoic acid (1 mmol) in toluene, the
corresponding trimethyl orthoester 12 (3 mmol) was added. The
reaction mixture was then heated at reflux (110 °C) for 48 h.
After cooling, 2 M HCl was added (1.1 mL). The organic layer
was washed with saturated NaHCO3 (1ꢁ) and brine (1ꢁ), and dried
over anhydrous MgSO4. The excess reagents and solvent were
evaporated under vacuum. The crude product was purified by silica
gel flash chromatography using hexane/ethyl acetate (99.5/0.5; v/
v) as an eluent to afford the corresponding product 13 as a colour-
less oil; Rf = 0.5 (hexane/EtOAc, 9:1; v/v); HPLC analysis [hexane/
To a solution of acid 16 (3.0 mmol) in ethyl acetate/acetic acid
(9/1, v/v, 5 mL) was added sodium cyanoborohydride (3 mmol)
and stirred in room temperature for 20 min after which a saturated
solution of ammonium hydrochloride was added. The reaction
mixture was then extracted with ethyl acetate (3 ꢁ 5 mL). The or-
ganic layer was dried over anhydrous MgSO4 and the excess sol-
vent was evaporated under vacuum. The crude product was
purified by silica gel flash chromatography using hexane/ethyl ace-
tate as an eluent to afford product 17 as a colourless oil (76% yield).
1H NMR (200 MHz, CDCl3): d 1.22 (t, J = 7.2 Hz, J = 14.2 Hz, 3H),
2.71 (dd, J = 7.0, 15.0 Hz, 1H), 2.91 (dd, J = 7.0, 15.0 Hz, 1H), 3.33–
3.47 (m, 1H), 3.73–3.89 (m, 2H), 4.12 (q, J = 7.0 Hz, 2H), 5.07
(1H), 7.22–7.44 (m, 5H); 13C NMR (50 MHz, CDCl3): d 14.3, 21.0,
37.7, 44.7, 60.9, 67.0, 127.3, 128.0, 128.7, 128.9, 141.2, 173.0,
176.2; ESI-MS HR, [M+Na]+, calcd for C12H16O3Na: 231.099, found
iPrOH;
9:1;
k = 232 nm;
1.0 mL/min]
tR(S) = 4.70 min;
tR(R) = 6.70 min.
4.6. Enantioselective synthesis of methyl 3-phenyl-4-
pentenoate 13
(m/z): 231.099; ½a D25
ꢂ
= +4.8 (c 1.0, CHCl3).
To a solution of 3-phenyl-4-pentenoic acid (1 mmol) in toluene,
the corresponding trimethyl orthoester 12 or acetal 14 or ketal 15
(3 mmol) and enzyme (10 mg) were added. The reaction mixture
was then stirred for 48 h at 35–40 °C. After cooling, 2 M HCl was
added (1.1 mL). The organic layer was washed with saturated NaH-
CO3 (1ꢁ) and brine (1ꢁ), and dried over anhydrous MgSO4. The ex-
cess reagents and solvent were evaporated under vacuum. The
4.10. Synthesis of the ethyl ester of (+)-3-phenyl-4-
methansulfonyloxybutyric acid 18
A solution of acid 17 (0.96 mmol, [a]D = +4.8) and triethylamine
(2.89 mmol) in dry dichloromethane was cooled to 0 °C and
methanesulfonic acid chloride was added (2.74 mmol). The reac-

A. Brodzka et al. / Tetrahedron: Asymmetry 24 (2013) 427–433
433
tion mixture was stirred at room temperature for 6 h, filtered and
the excess solvent evaporated under vacuum. The crude product
was purified by silica gel flash chromatography using hexane/ethyl
acetate as an eluent to afford the corresponding product 18 (90%
yield). 1H NMR (200 MHz, CDCl3): d 1.24 (t, J = 7.2 Hz, J = 14.2 Hz,
3H), 2.70–3.00 (m, 1H), 2.90 (s, 3H), 3.57–3.73 (m, 1H), 4.13 (q,
J = 7.4 Hz, J = 7.0 Hz, 2H), 4.48–4.52 (m, 2H), 7.28–7.45 (m, 5H);
13C NMR (50 MHz, CDCl3): d 14.4, 37.0, 37.5, 41.7, 61.0, 72.8,
128.0, 128.0, 129.1, 139.3, 171.5; ESI-MS HR, [M+Na]+, calcd for
138.6, 175.8—according to the literature;10 ESI-MS HR, [M]+, calcd
for C10H14NO2: 180.102, found (m/z): 180.102, ½a D25
= ꢀ0.7 (c 1.0,
ꢂ
H2O).
4.14. Synthesis of the ethyl ester of (S)-(+)-4-phenyl-2-
pyrollidone 9
Compound 20 (0.04 mmol) was suspended in toluene (3 mL)
and triethylamine was dropped (0.04 mmol). The reaction mixture
was then heated at reflux for 24 h, after which the excess solvent
was evaporated under vacuum. The residue was washed with
dichloromethane. The organic layer was washed with hydrochloric
acid (10%, 3 ꢁ 5 mL) and water (3 ꢁ 5 mL). The combined organic
layers were dried over anhydrous MgSO4 and the excess solvent
evaporated. The crude product was purified by crystallization from
hexane/ethyl acetate (60% yield). ESI-MS HR, [M]+, calcd for
C10H11NO: 161.084, found (m/z): 161.085; mp 78 °C [lit. 76–77].11
C
13H18O5NaS: 309.076, found (m/z): 309.076; ½a D25
ꢂ
= +1.05 (c 1.0,
CHCl3).
4.11. Synthesis of the ethyl ester of (-)-3-phenyl-4-azidobutyric
acid 19
A solution of ester 18 (1.64 mmol), sodium azide (3.28 mmol),
diazabicycloundecane (2.62 mmol), DMAP (10 mg) and crown
ether B18C6 (10 mg) in dry dichloromethane (10 mL) was heated
at reflux for 72 h. After filtration, the excess solvent was evapo-
rated under vacuum. The crude product was purified by silica gel
flash chromatography using hexane/ethyl acetate as an eluent to
afford product 19 (66% yield). 1H NMR (200 MHz, CDCl3): d 1.18–
1.30 (m, 3H), 2.64–3.12 (m, 2H), 3.40–3.90 (m, 4H), 4.06–4.24
(m, 1H), 7.24–7.47 (m, 5H); 13C NMR (50 MHz, CDCl3): d 14.4,
38.1, 38.2, 49.0, 56.5, 127.7, 127.8, 127.9, 128.9, 129.0, 140.8,
140.9, 171.8; ESI-MS HR, [M+Na]+, calcd for C12H15N3O2Na:
Acknowledgments
This work was supported by the State Committee for Scientific
Research, KBN Grant PBZ-MIN-007/P04/2003 and by the project
‘Biotransformations for pharmaceutical and cosmetics industry’
No. POIG.01.03.01-00-158/09-01 part-financed by the European
Union within the European Regional Development Fund.
256.105, found (m/z): 256.104; ½a D25
ꢂ
= ꢀ0.6 (c 1.0, CHCl3).
References
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4.12. Synthesis of the ethyl ester of 3-phenyl-4-aminobutyric
acid 20
To a solution of ester 19 (0.38 mmol) in ethanol (5 mL) was
added palladium on carbon. The reaction mixture was then stirred
under hydrogen for 10 h, filtered and the excess solvent evapo-
rated under vacuum. Product 20 was obtained in 58% yield. 1H
NMR (200 MHz, CDCl3): d 1.16 (t, J = 7.2 Hz, 3H), 2.75 (dd, J = 5.5,
9.8 Hz, 1H), 2.84 (dd, J = 5.5, 16.1, 1H), 3.25 (q, 1H), 3.40 (m, 2H),
4.06 (q, J = 7.2 Hz, 2H), 7.37 (m, 3H), 7.43 (m, 2H); 13C NMR
(50 MHz, CDCl3): d 14.4, 22.1, 38.0, 38.9, 48.9, 126.6, 127.0,
127.7, 127.8, 128.1, 128.7, 128.9, 129.1, 140.5, 172.2; ESI-MS HR,
[M+Na]+, calcd for C12H18NO2: 208.133, found (m/z): 208.132.
4.13. Synthesis of (S)-3-phenyl-4-aminobutyric acid
hydrochloride (phenylGABA) 21
To a solution of ester 20 (0.58 mmol) in acetic acid (1 mL) was
added hydrochloric acid (6 M, 0.1 mL). The reaction mixture was
then stirred for 24 h at room temperature, after which the excess
solvent was evaporated under vacuum. The crude product was
purified by crystallization from benzene (89% yield). 1H NMR
(400 MHz, D2O): d 2.75 (dd, J = 5.5, 9.8 Hz, 1H), 2.84 (dd, J = 5.5,
16.1, 1H), 3.25 (m, 1H), 3.40 (m, 2H), 7.37 (m, 3H), 7.43 (m, 2H);
13C NMR (50 MHz, D2O): d 38.5, 40.2, 44.1, 128.1, 128.6, 129.6,
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