Enzymatic resolution of five membered heterocyclic bromohydrins

Article abstract of DOI:10.1016/j.tetasy.2013.04.012

The lipase catalyzed resolution of trans-3,4-tetrahydrofuran and pyrrolidine bromohydrins by acylation or hydrolysis of their acylated derivatives has been studied. For both heterocycles, the best enantioselectivity was obtained using Candida antarctica l

Full text of DOI:10.1016/j.tetasy.2013.04.012

Tetrahedron: Asymmetry 24 (2013) 694–698
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Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
Enzymatic resolution of five membered heterocyclic bromohydrins
⇑
⇑
Ángela Villar-Barro, Vicente Gotor , Rosario Brieva
Departamento de Química Orgánica e Inorgánica and Instituto Universitario de Biotecnología de Asturias, Universidad de Oviedo, 33006 Oviedo (Asturias), Spain
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 6 April 2013
Accepted 18 April 2013
The lipase catalyzed resolution of trans-3,4-tetrahydrofuran and pyrrolidine bromohydrins by acylation
or hydrolysis of their acylated derivatives has been studied. For both heterocycles, the best enantioselec-
tivity was obtained using Candida antarctica lipase B as the catalyst in the hydrolytic processes. The enan-
tiomerically pure bromohydrins are useful intermediates for the preparation of 3,4-fuctionalized cis-
heterocycles.
Ó 2013 Elsevier Ltd. All rights reserved.
1. Introduction
2. Results and discussion
Optically active 3,4-disubstituted tetrahydrofurans and pyrrol-
idines are common structural subunits found in many natural com-
pounds and synthetic derivatives that exhibit interesting biological
activities. For example, enantiomerically pure tetrahydrofurans
can serve as monosaccharide mimetic structures while some cis-
3,4-pyrrolidine derivatives have been described as inhibitors of
the natural enzyme renin.1 Their presence as substituents in quin-
oline antibacterial derivatives,2 or thiopenems,3 is crucial for their
biological activity. Moreover, non-racemic vicinal heterocyclic
diols, diamines, or aminoalcohols, occupy a central role in catalytic
asymmetry synthesis as chiral auxiliaries or ligands.
On the other hand, easily accessible vicinal trans-bromohydrins,
are common intermediates in numerous synthetic routes for the
preparation of pharmaceuticals, agrochemicals, and other fine
chemicals. The SN2 displacement reaction of the bromo functional-
ity in 3,4-disubstituted tetrahydrofurans and pyrrolidines yields
heterocycles with a cis-relative configuration.
As part of our research on the enzymatic preparation of opti-
cally active functionalized heterocycles,4 we are interested in
extending biocatalytic methods to the resolution of bromohydrin
heterocyclic structures. We have studied the enzymatic resolution
of trans-3-bromo-4-hydroxytetrahydrofuran and its analogue pyr-
rolidine derivative.
We have also carried out the SN2 displacement reaction of the
bromo functionality with sodium azide in order to obtain the enan-
tiomerically pure cis-hydroxyazide derivatives. The comparison of
the specific rotation of these compounds to those described in
the literature5 led us to determine the absolute configuration of
the enantiomerically pure bromohydrins obtained.
Initially, we studied the enzymatic hydrolysis of ( )-trans-3-
bromo-4-(2-phenylacetoxy)tetrahydrofuran, ( )-trans-3a, which
was prepared in high yield by treatment of 2,5-dihydrofuran 1
with N-bromosuccinimide and subsequent acylation of the bro-
mohydrin obtained with 2-phenylacetylchloride (Scheme 1).
O
O
Br
HO
Br
Ph
Ph
NBS
H2O
Cl
K2CO3
CH3CN
O
O
O
O
(
)-trans-2
1
( )-trans-3a
Scheme 1. Synthesis of bromohydrins ( )-trans-2 and ( )-trans-3a.
We started testing the enzymatic hydrolysis of ( )-trans-3a
(Scheme 2) using two different reaction conditions: with 10 equiv
of water in organic solvent (tBuOMe) or in a 1:2 ratio of water/or-
ganic solvent. The obtained results are summarized in Table 1.
O
Br
HO
Br
O
Br
Lipase
tBuOMe/H2O
30ºC
Ph
Ph
+
O
O
O
O
O
( )-trans-3a
(3S,4S)-(+)-3a
(3R,4R)-(−)-2
Scheme 2. Enzymatic hydrolysis of ( )-trans-3a (the configuration shown was
obtained using CAL-B).
Five of the lipases tested catalyzed the hydrolysis of substrate
( )-trans-3a: lipases A and B from Candida antarctica (CAL-A), Pseu-
domonas fluorescens (AK), Burkholderia cepacia (PS-SD), and
⇑
Corresponding authors. Tel.: +34 985 102 994; fax: +34 985 103 448.
E-mail addresses: vgs@uniovi.es (V. Gotor), rbrieva@uniovi.es (R. Brieva).
0957-4166/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetasy.2013.04.012

Ángela Villar-Barro et al. / Tetrahedron: Asymmetry 24 (2013) 694–698
695
Table 1
Lipase catalyzed hydrolysis of ( )-trans-3a, in tBuOMe at 30 °C
a
a
Entry
Lipase
H2O
t (h)
ees (%) trans-3a
Configuration
eep (%) trans-2
Configuration
cb (%)
Eb
1
2
3
4
5
6
7
CAL-A
CAL-B
AK
CAL-A
CAL-B
TL-IM
PS-SD
10 equiv
10 equiv
10 equiv
22
16
22
17
17
22
59
>99
33
86
97
16
4
(3R,4R)
(3S,4S)
(3S,4S)
(3R,4R)
(3S,4S)
(3S,4S)
(3R,4R)
27
>99
61
31
83
(3S,4S)
(3R,4R)
(3R,4R)
(3S,4S)
(3R,4R)
(3R,4R)
(3S,4S)
69
50
35
73
54
30
16
3
>200
6
5
45
3
tBuOMe/H2O 2:1
tBuOMe/H2O 2:1
tBuOMe/H2O 2:1
tBuOMe/H2O 2:1
37
21
8 (day)
2
a
Determined by chiral HPLC.
b
Conversion, c = ees/(ees + eep), enantiomeric ratio, E = ln[(1 ꢀ c)(1 ꢀ ees)]/ln [(1 ꢀ c)((1 + ees)].6
Thermomyces lanuginosus (TL IM). Using the latter two enzymes,
the product was only detected using a proportion of solvent/H2O
(2:1), but low enantioselectivity and conversion were achieved
(Table 1, entries 6 and 7).
O
Br
HO
Br
R
O
CbzCl
CH2Cl2
Py
NBS
H2O
R
Cl
O
K2CO3
CH3CN
N
N
H
N
N
In general, better results were obtained using a small amount of
water (10 equiv), particularly in the case of the hydrolysis cata-
lyzed by CAL-B (Table 1, entry 2). Using this enzyme, 50% conver-
sion was achieved after 16 h of reaction and both the product and
the remaining substrate were obtained in enantiopure form.
A different stereochemical preference of some lipases was ob-
served. Lipases CAL-B, AK, and TL-IM catalyzed the hydrolysis of
the (3R,4R)-configured ester while CAL-A and PS-D catalyzed the
hydrolysis of the (3S,4S)-configured ester. The assignation of the
absolute configurations of the products and the remaining sub-
strate is described later.
Cbz
Cbz
Cbz
5
4
(
)-trans-6
( )-trans-7a-c
7a
R = benzyl
7b R = methyl
7c R = metoxymethyl
Scheme 4. Synthesis of bromohydrins ( )-trans-6 and ( )-trans-7a–c.
HO
Br
R
O
Br
R
O
Br
Lipase
+
O
We also examined the acylation process of substrate ( )-trans-2,
using CAL-B as catalysts, but a lower reaction rate and enantiose-
lectivity were achieved in these processes (Scheme 3). The results
obtained are summarized in Table 2.
O
tBuOMe/H2O
30ºC
N
N
N
Cbz
Cbz
Cbz
6
(3S,4S)-7a-c
(3R,4R)-
( )-trans-7a-c
7a
R = benzyl
7b R = methyl
7c R = metoxymethyl
HO
Br
R
O
Br
HO
Br
O
CAL-B
+
+
R'
tBuOMe
30ºC
Scheme 5. Lipase catalyzed hydrolysis of ( )-trans-7a–c (the configuration shown
O
R
O
was obtained using CAL-B).
O
O
O
2
3a,b
(3R,4R)-
( )-trans-2
(3S,4S)-
Scheme 3. CAL-B catalyzed acylation of ( )-trans-2.
Under the same reaction conditions, lipase CAL-A catalyzed the
process in a moderate yield but with low enantioselectivity
(Table 3, entry 2). The lipases AK and TL IM also catalyzed the
hydrolysis but the product was only detected when using sol-
vent/H2O in a ratio of 2:1 and low enantioselectivity and conver-
sion were achieved (Table 3, entries 4 and 5).
Table 2
CAL-B catalyzed acylation of ( )-trans-2 in tBuOMe, using 5 equiv of the acylating
agent
a
a
Entry Product
R
R’
t (h) ees (%) eep (%) cb (%) Eb
Dioxane, toluene, acetonitrile, and THF were also examined as
solvents; the rate and enantioselectivity were not improved in
1
2
3a
3b
Benzyl
Methyl vinyl
ethyl
48 38
48 14
56
42
40
25
5
3
t
comparison with the results obtained in BuOMe.
a
Determined by chiral HPLC.
As in the case of the tetrahydrofuran derivative, a different
stereochemical preference of some lipases was observed. Lipases
CAL-B, AK, and TL-IM catalyzed the hydrolysis of the (3R,4R)-con-
figured ester while CAL-A catalyzed the hydrolysis of the (3S,4S)-
configured ester.
b
Conversion, c = ees/(ees + eep), enantiomeric ratio, E = ln[(1 ꢀ c)(1 ꢀ ees)]/ln
[(1 ꢀ c)((1 + ees)].6
Taking into account the results obtained in the resolution of the
tetrahydrofuran bromohydrin ( )-trans-3a, we studied the enzy-
matic hydrolysis of ( )-trans-1-benzyloxycarbonyl-3-bromo-4-(2-
phenylacetoxy)pyrrolidine, ( )-trans-7a, which was prepared using
the same methodology as for the tetrahydrofuran derivative
(Scheme 4).
The CAL-B catalyzed hydrolysis of ( )-trans-7a was slower than
for substrate ( )-trans-3a: under the same reaction conditions,
only 22% conversion was achieved after 12 days and the enantiose-
lectivity was also lower, E = 23, (Scheme 5, Table 3, entry 1). We
then attempted to improve these results by testing different li-
pases and analyzing the influence of different reaction parameters.
Since the results obtained for substrate ( )-trans-7a were not
satisfactory, we prepared two new derivatives ( )-trans-7b and
( )-trans-7c and carried out the hydrolysis processes catalyzed by
CAL-B under the best conditions found for derivative ( )-trans-7a
(Table 3, entries 6 and 7). For both substrates, the rate and enanti-
oselectivity of the process were higher in comparison with those of
substrate ( )-trans-7a, particularly for the methoxyacetylated
derivative ( )-trans-7c. In this process, 50% conversion was
achieved after 9 h of reaction and both the product and the remain-
ing substrate were obtained in enantiopure form.
Finally, we examined the enzymatic acylation of the bromohy-
drin ( )-trans-6. When vinyl acetate was used as the acylating

696
Ángela Villar-Barro et al. / Tetrahedron: Asymmetry 24 (2013) 694–698
Table 3
Lipase catalyzed hydrolysis of ( )-trans-7a–c, in tBuOMe at 30 °C
a
a
Entry
Substrate
Lipase
H2O
t (h)
ees (%) trans-7a–c
Configuration
eep (%) trans-6
Configuration
cb (%)
Eb
1
2
3
4
5
6
7
( )-trans-7a
( )-trans-7a
( )-trans-7a
( )-trans-7a
( )-trans-7a
( )-trans-7b
( )-trans-7c
CAL-B
CAL-A
CAL-B
AK
TL-IM
CAL-B
CAL-B
10 equiv
10 equiv
12 (days)
24
7 (days)
7 (days)
7 (days)
48
25
5
17
1
6
76
>99
(3S,4S)
(3R,4R)
(3S,4S)
(3S,4S)
(3S,4S)
(3S,4S)
(3S,4S)
89
12
88
15
66
(3R,4R)
(3S,4S)
(3R,4R)
(3R,4R)
(3R,4R)
(3R,4R)
(3R,4R)
22
31
16
8
8
46
50
23
1
18
1
5
50
>200
tBuOMe /H2O 2:1
tBuOMe /H2O 2:1
tBuOMe /H2O 2:1
10 equiv
91
>99
10 equiv
9
a
Determined by chiral HPLC.
b
Conversion, c = ees/(ees + eep), enantiomeric ratio, E = ln[(1 ꢀ c)(1 ꢀ ees)]/ln [(1 ꢀ c)((1 + ees)].6
agent, very low enantioselectivity was observed for all of the
catalysts tested. Scheme 6 shows the best results, obtained in the
process catalyzed by CAL-B.
process. In order to avoid possible racemization, the hydroxyl
group was protected as a butyldimethylsilylated derivative (ꢀ)-
t
10, and then treated with sodium azide in DMSO to yield the cor-
responding cis-azido derivative (+)-11, with an enantiomeric ex-
cess higher than 99%. The deprotection of (+)-11 yielded the
azido alcohol (ꢀ)-12. The comparison of the specific rotation of this
compound to that described in the literature5a led us to determine
the (3S,4R)-absolute configuration for the azido alcohol (ꢀ)-12; the
product of the enzymatic hydrolysis was therefore established as
the (3R,4R)-enantiomer (Scheme 8).
HO
Br
O
Br
HO
Br
O
CAL-B, tBuOMe
+
+
O
30ºC, 24h
c = 52, E = 2
O
N
N
N
Cbz
Cbz
Cbz
(
)-trans-6
(3S,4S)-6
(3R,4R)-7b
It should be noted that for both substrates, CAL-B preferentially
catalyzed the hydrolysis of the (3R,4R) enantiomer, as expected by
Kazlauskas’ rule.7
Scheme 6. CAL-B catalyzed acylation of ( )-trans-6.
Using less activated acylating agents such as ethyl 2-phenylac-
etate, only lipase CAL-A catalyzed the process, but low conversion
and enantioselectivity were observed.
3. Conclusions
In order to determine the absolute configuration of the bromo-
hydrins obtained we carried out an SN2 displacement reaction of
the bromo functionality with sodium azide and then subsequent
hydrolysis of the ester to yield the corresponding enantiomerically
pure cis-hydroxyazides (Schemes 7 and 8). In the case of the tetr-
ahydofuran bromohydrins, we used the remaining enantiomeri-
cally pure substrate (+)-3a obtained in the lipase-catalyzed
kinetic resolution. The treatment of this compound with sodium
azide in DMSO yielded the corresponding cis-azido ester (+)-8, with
an enantiomeric excess higher than 99%. The hydrolysis of (+)-8
yielded the azido alcohol (ꢀ)-9, whose specific rotation sign was
compared with that reported for the (3R,4R)-enantiomer;5b the
Herein we have described an easy method for the resolution of
trans-3-bromo-4-hydroxytetrahydrofuran and the analogous pyr-
rolidine derivatives via a CAL-B catalyzed hydrolytic process. These
compounds could be useful intermediates in the preparation of
enantiomerically pure 3,4-disubstituted heterocycles.
The best substrates for the resolution are the phenylacetylated
derivative of the bromohydrin, in the case of the tretrahydrofuran
ring, and the methoxyacetylated derivative in the case of the pyr-
rolidine. Good yields and high enantioselectivities can be achieved
by the appropriate selection of the reaction parameters. The two
enantiomers of both substrates can be obtained in good yield and
in enantiomerically pure forms. On the other hand, enzymatic
hydrolysis of an ester derivative is preferable to acylation of the
hydroxyl group of the bromohydrins.
O
Br
O
N3
HO
N3
Ph
Ph
ii
i
4. Experimental
4.1. General
O
O
O
O
O
(3S,4S)-(+)-3a
(3R,4R)-(+)-8
(3R,4R)-(−)-9
Scheme 7. Reagents and conditions: (i) NaN3, DMSO, 110 °C, 7 h; (ii) K2CO3, MeOH/
H2O, rt, 2 h.
Enzymatic reactions were carried out in a Gallenkamp incuba-
tory orbital shaker. Immobilized Candida antarctica lipase B, CAL-B
(Novozym 435, 7300 PLU/g), was a gift from Novo Nordisk co.,
immobilized CAL-A (lipase NZL-101, 6.2 U/g) is commercialized by
Codexis, and immobilized Burkholderia cepacia lipase (PS-SD,
23000 U/g) is commercialized by Amano Pharmaceuticals. Pseudo-
monas fluorescens (AK, 22100 U/g)), and Thermomyces lanuginosus
(TL IM, 560 TBU/g) are commercialized by Novo Nordisk. Chemical
reagents were commercialized by Aldrich, Fluka, Lancaster or Pro-
labo. Solvents were distilled over an appropriate desiccant under
nitrogen. Flash chromatography was performed using Merck silica
gel 60 (230–400 mesh). Optical rotations were measured using a
Perkin–Elmer 343 polarimeter and are quoted in units of
HO
Br TBDMSO
Br
HO
N3
TBDMSO
N3
i
ii
iii
N
N
N
N
Cbz
Cbz
Cbz
Cbz
−
6
(3R,4R)-( )-
(3R,4R)-(−)-10
+
11
−
12
(3S,4R)-( )-
(3S,4R)-( )-
Scheme 8. Reagents and conditions: (i) TBDMSCl, imidazole, CH2Cl2, rt, 24 h; (ii)
NaN3, DMSO, 110 °C, 7 h; (iii) CSA, anhydrous MeOH, rt, overnight.
10ꢀ1 deg cm2 gꢀ1 1H NMR, 13C NMR, and DEPT spectra were re-
.
configuration of the remaining substrate in the enzymatic process
was therefore established as (3S,4S)-(+)-3a.
In the case of the pyrrolidine bromohydrins, we started with the
enantiomerically pure product (ꢀ)-6 obtained via the biocatalytic
corded in a Bruker AC-300, Bruker AC-300 DPX, or Bruker NAV-
400 spectrometer using CDCl3 as solvent. The chemical shift values
(d) are given in ppm. APCI+ and ESI+ using a Hewlett–Packard 1100

Ángela Villar-Barro et al. / Tetrahedron: Asymmetry 24 (2013) 694–698
697
chromatograph mass detector or EI+ with a Hewlett–Packard 5973
4.3.4. ( )-trans-1-Benzyloxycarbonyl-3-acetoxy-4-bromo
pyrrolidine ( )-trans-7b
mass spectrometer were used to record mass spectra (MS). IR spec-
tra were recorded in a UNICAM Mattson 3000 FT. The enantiomeric
excesses were determined by chiral HPLC analysis on a Hewlett–
Packard 1100, LC liquid chromatograph, using a CHIRALPAK OJ-H
column (4.5 ꢁ 250 mm) and CHIRALPAK IC column (4.0 ꢁ 250 mm).
Yellow oil, yield 50%. 1H NMR (CDCl3, 300.13 MHz): d 7.39 (br s,
5H), 5.33 (m, 1H), 5.18 (br s, 2H), 4.41–4.24 (m, 1H), 4.12–3.86 (m,
3H), 3.71–3.5 (m, 1H), 2.09 (s, 3H); 13C NMR (CDCl3, 75.5 MHz): d
169.6 (CO), 154.7 (CO), 136.4 (C), 128.5 (CH), 128.2 (CH), 128.0
(CH), 128.6 (CH), 78.3 (CH), 67.3 (CH2), 53.4 (CH2), 59.56 (CH2),
46.9 (CH), 20.9 (CH3); IR (neat, NaCl):
m
1744, 1704 cmꢀ1; HRMS-
4.2. General procedure for the preparation of bromohydrins
ESI+ calcd for [C14H16BrNO4Na]+ (M+Na)+ 364.0155 m/z, found:
364.0169.
2,5-Dihydrofuran (175 mg, 2.50 mmol) or 1-benzyloxycar-
bonyl-3-pyrroline (500 mg, 2.46 mmol) was added to a stirred
solution of N-bromosuccinimide (NBS) (650 mg, 3.7 mmol) in
water and cooled on an ice bath. The resulting mixture was stirred
4 h at rt. After this time, 15 mL of ethyl ether was added and the
resulting mixture was washed with brine separated and dried over
Na2SO4. The solvent was removed under reduced pressure to give a
yellow oil as a crude residue in a 70–80% yield. The crude product
obtained was used for the chemical acylation without further puri-
fication. When the pure bromohydrin was required, the crude was
purified by flash chromatography on silica gel (hexane/EtOAc 6:4).
4.3.5. ( )-trans-1-Benzyloxycarbonyl-3-bromo-4-(2-methoxy
acetyl)pyrrolidine ( )-trans-7c
Yellow oil, yield 65%. 1H NMR (CDCl3, 300.13 MHz): d 7.52–7.27
(m, 5H), 5.42 (m, 1H), 5.17 (br s, 2H), 4.38–4.24 (m, 1H), 4.21–3.84
(m, 5H), 3.62 (m, 1H), 3.44 (s, 3H); 13C NMR (CDCl3, 75.5 MHz): d
168.1 (CO), 154.6 (CO), 136.3 (C), 128.5 (C), 128.2 (CH), 128.0
(CH), 78.5 (CH), 69.5 (CH2), 67.3 (CH2), 59.5 (CH3), 53.3 (CH2),
49.4 (CH2), 46.3 (CH); IR (neat, NaCl):
m
1758, 1704 cmꢀ1; HRMS-
ESI+ calcd for [C15H18BrNO5Na]+ (M+Na)+ 394.0261 m/z, found:
394.0258.
4.3. General procedure for the acylation of bromohydrins
4.4. General procedure for the enzymatic hydrolysis
To a solution of the crude bromohydrin (3 mmol) in acetonitrile
(30 mL), the corresponding acyl chloride (4.6 mmol) and K2CO3
(623 mg, 3 mmol) were added. The mixture was stirred at room
temperature for 12 h. Next, the solvent was removed under re-
duced pressure and ethyl acetate (25 mL) was added to the crude
residue; the mixture was washed with water (3 ꢁ 15 mL) and the
organic phase was dried over Na2SO4. The solvent was removed
under reduced pressure and the crude residue was purified by flash
chromatography on silica gel (hexane/EtOAc 8:2) to afford the cor-
responding ester.
The reaction mixture, which contained the corresponding acyl-
ated bromohydrin (20 mg), the lipase (20 mg), and H2O (10 equiv)
in tBuOMe (2.5 mL), was shaken at 30 °C and 250 rpm in an orbital
shaker. The progress of the reaction was monitored by TLC (hex-
ane/EtOAc 8:2) until required conversion was achieved. Next, the
enzyme was removed by filtration and washed with BuOMe. The
crude residue was purified by flash chromatography on silica gel
(hexane/EtOAc 8:2).
t
4.4.1. (3S,4S)-3-Bromo-4-hydroxytetrahydrofuran (3R,4R)-2
Yellow oil, yield 48%; ½a D25
¼ ꢀ27:6 (c 0.08, CH2Cl2), ee >99%.
ꢂ
4.3.1. ( )-trans-3-Bromo-4-(2-phenylacetoxy)-tetrahydrofuran
( )-trans-3a
4.4.2. (3S,4S)-3-Bromo-4-(2-phenylacetoxy)tetrahydrofuran
(3S,4S)-3a
Yellow oil, yield 80%. 1H NMR (CDCl3, 300.13 MHz): d 7.49–7.18
(m, 5H), 5.40 (d, 1H, JHH 6.6), 4.30 (m, 3H), 4.09 (m, 1H), 3.87 (m,
1H), 3.66 (s, 2H); 13C NMR (CDCl3, 75.5 MHz): d 170.9 (CO), 133.6
(C), 129.6 (CH), 129.1 (CH), 127.8 (CH), 81,0 (CH), 75.0 (CH2), 71.7
Yellow oil, yield 49%; ½a D25
¼ þ73:1 (c 0.1, CH2Cl2), ee >99%.
ꢂ
4.4.3. (3R,4R)-1-Benzyloxycarbonyl-3-bromo-4-hydroxypyrro-
lidine (3R,4R)-6
(CH2), 48.4 (CH), 41.5 (CH2); IR (neat, NaCl):
m
1739 cmꢀ1; HRMS-
ESI+ calcd for [C12H13BrO3Na]+ (M+Na)+ 306.9940 m/z, found:
Yellow oil, yield 47%; ½a D20
¼ ꢀ5:0 (c 0.1, CHCl3), ee >99%.
ꢂ
306.9913.
4.4.4. (3S,4S)-1-Benzyloxycarbonyl-3-bromo-4-(2-methoxyace-
tyl)pyrrolidine (3S,4S)-trans-7c
4.3.2. ( )-trans-3-Acetoxy-4-bromotetrahydrofuran ( )-trans-
3b
Yellow oil, yield 49%; ½a D20
¼ ꢀ10:0 (c 0.05, CHCl3), ee >99%.
ꢂ
Yellow oil yield 82%. 1H NMR (CDCl3, 300.13 MHz): d 5.29–5.19
(m, 1H), 4.28–4.15 (m, 3H), 3.97 (m, 1H), 3.74 (m, 1H), 1.98 (s, 3H).
13C NMR (CDCl3, 75.5 MHz): d 169.7 (CO), 81,2 (CH), 74.4 (CH2),
4.5. Determination of the enantiomeric excess by HPLC analysis
Chiralpak OJ-H, 30 °C, hexane/2-propanol (95:5), UV 210 nm,
0.8 mL minꢀ1, tR 26.96 min (3S,4S)-3a, tR 23.76 min (3R,4R)-3a.
71.3 (CH2), 48.2 (CH), 20.7 (CH3); IR (neat, NaCl):
m ;
1742 cmꢀ1
HRMS-ESI+ calcd for [C6H9BrO3Na]+ (M+Na)+ 230.9627 m/z, found:
Chiralpak IC; hexane/2-propanol (90:10) UV 210 nm, 0.8 mL min-
232.9618.
1
,
tR 15.77 min (3R,4R)-6, tR 13.5 min (3S,4S)-6; tR 56.5 min
(3R,4R)-7c; tR 47.38 min (3S,4S)-7c.
4.3.3. ( )-trans-1-Benzyloxycarbonyl-3-bromo-4-(2-phenyl
acetoxy)pyrrolidine ( )-trans-7a
4.6. General procedure for the enzymatic acylation
Yellow oil yield 53%. 1H NMR (CDCl3, 300.13 MHz): d 7.52–7.19
(m, 10H), 5.35 (d, 1H, 3JHH 4.3), 5.19 (s, 2H), 4.2 (br s, 1H), 4.11–4.00
(m, 1H), 3.94 (m, 1H), 3.75–3.53 (m, 3H); 13C NMR (CDCl3,
75.5 MHz): d 170.7 (CO), 155.1 (CO), 137.8 (C), 133.5 (C), 129.5
(CH), 129.2 (CH), 129.0 (CH), 128.6 (CH), 128.4 (CH), 127.88 (CH),
78.9 (CH), 67.7 (CH2), 52.85 (CH2), 51.53 (CH2), 49.9 (CH), 41.5
The reaction mixture containing the corresponding bromohy-
drin (50 mg), the lipase (100 mg), and the acylating agent (5 equiv)
t
in BuOMe (1 mL) was shaken at 30 °C and 250 rpm in an orbital
shaker. The progress of the reaction was monitored by TLC (hex-
ane/EtOAc 8:2) until the required conversion. The enzyme was
then removed by filtration and washed with BuOMe. The solvent
was evaporated under reduced pressure and the crude residue
m
1740, 1704 cmꢀ1; HRMS-ESI+ calcd for
t
(CH2); IR (neat, NaCl):
[C20H20BrNO4Na]+ (M+Na)+ 440.0468 m/z, found 440.0457.

698
Ángela Villar-Barro et al. / Tetrahedron: Asymmetry 24 (2013) 694–698
was purified by flash chromatography on silica gel (hexane/EtOAc
8:2).
(CH3); IR (neat, NaCl): m ;
1708.8 cmꢀ1 HRMS-ESI+ calcd for
[C18H28BrNO3SiNa]+ (M+Na)+ 436.0914 m/z, found: 436.0899.
4.7. Assignment of the absolute configuration of
tetrahydrofuran derivatives
4.8.2. Synthesis of (3S,4R)-(+)-(-1-benzyloxycarbonyl-3-azido-4-
(tert-butyldimethylsilyloxy)pyrrolidine (3S,4R)-(+)-11
To a solution of (ꢀ)-trans-10 (30 mg, 0.1 mmol) in DMSO
(3 mL), sodium azide (23.4 mg, 0.36 mmol) was added under a
nitrogen atmosphere at room temperature. The mixture was then
stirred at 110 °C for 12 h. Next, the solvent was removed under re-
duced pressure and the crude residue was purified by flash chro-
matography on silica gel (hexane/EtOAc 8:2) to afford the
4.7.1. Synthesis of (3R,4R)-(+)-3-azido-4-phenylacetoxytetrahy-
drofuran, (3R,4R)-(+)-8
To a solution of the enantiomerically pure substrate (+)-trans-
3a (30 mg, 0.1 mmol), obtained from the CAL-B catalyzed hydroly-
sis, in DMSO (2 mL), sodium azide (33 mg, 0.5 mmol) was added
under a nitrogen atmosphere at room temperature. The mixture
was stirred at 110 °C for 7 h. Next, the solvent was removed under
reduced pressure. The crude residue was purified by flash chroma-
tography on silica gel (hexane/EtOAc 7:3) to afford the product as a
product as a colorless oil in 57% yield. ½a D25
¼ þ35:0 (c 0.1, CHCl3),
ꢂ
ee >99%. 1H NMR (CDCl3, 300.13 MHz): d 7.38 (br s, 5H), 5.16 (s,
2H), 4.43 (m, 1H), 4.05–3.70 (m, 1H), 3.50 (m, 4H), 0.94 (s, 9H),
ꢀ0.29 (s, 6H); 13C NMR (CDCl3, 75.5 MHz): d 154.8 (CO), 136.6
(C), 128.5 (CH), 128.1 (CH), 128.0 (CH), 72.5 (CH), 67.1 (CH2),
61.4 (CH), 50.4 (CH2), 47.63 (CH2), 25.29 (CH3), 18.1 (C), ꢀ5.0
yellow oil in 52% yield. ½a D25
ꢂ
¼ þ1:4 (c 0.08, CH2Cl2), ee >99%. 1H
NMR (CDCl3, 300.13 MHz): d 7.40–7.28 (m, 5H), 5,20 (m, 1H)
4.11 (m, 1H), 4.08–3.96 (m, 2H), 3.90–3.72 (m, 2H), 3.67 (s, 2H);
13C NMR (CDCl3, 75.5 MHz): d 171.1 (CO), 133.2 (C), 129.3 (C),
128.7 (CH), 127.3 (CH), 73.7 (CH), 70.8 (CH2), 70.1 (CH2), 60.7
(CH3); IR (neat, NaCl):
m
2104.3, 1704.5 cmꢀ1. HRMS-ESI+ calcd
for [C18H29N4O3Si]+ (M+1)+ 377.2003 m/z, found: 377.1996.
(CH), 40.9 (CH2; IR (neat, NaCl):
m
2355.5, 1738.05 cmꢀ1; HRMS-
4.8.3. Synthesis of (3S,4R)-(ꢀ)-(-1-benzyloxycarbonyl-3-azido-4-
hydroxypyrrolidine (3S,4R)-(ꢀ)-12
ESI+ calcd for [C12H13N3O3Na]+ (M+Na)+ 270.0855 m/z, found
270.0849.
To a solution of (+)-cis-11 (10 mg, 0.03 mmol), in dry methanol
(300
lL), (ꢀ)-Camphor sulfonic acid, CSA, (35 mg, 0.15 mmol) was
added. The mixture was stirred overnight at room temperature.
Then, the solvent was removed under reduced pressure and the
crude residue was purified by flash chromatography on silica gel
(hexane/EtOAc 6:4) to afford the product as a colorless oil in 71%
yield. The specific rotation sign of the pure product was in accor-
dance with that reported for the (3S,4R)-(ꢀ)-(-1-benzyloxycar-
bonyl-3-azido-4-hydroxypyrrolidine5a
4.7.2. Synthesis of (3R,4R)-3-azido-4-hydroxytetrahydrofuran,
(3R,4R)-(ꢀ)-9
A solution of (+)-cis-8 (8 mg, 0.03 mmol) and K2CO3 in metha-
nol/water 2:1 (1.5 mL) was stirred at room temperature for 6 h.
Next, ethyl acetate (10 mL) was added, the resulting mixture was
washed with water (3 ꢁ 15 mL) and the organic phase was dried
over Na2SO4. The solvent was removed under reduced pressure
to afford the product as a colorless oil in 75% yield. The crude res-
idue was purified by flash chromatography on silica gel (hexane/
EtOAc 7:3). The specific rotation sign of the pure product was in
accordance with that reported for the (3R,4R)-(ꢀ)-3-azido-4-
hydroxytetrahydrofuran.5b
Acknowledgments
This work was supported by the Ministerio de Ciencia e Innova-
ción of Spain (Project CTQ 2011-24237). A. V.-B. thanks Principado
de Asturias for a predoctoral fellowship.
4.8. Assignment of the absolute configuration of pyrrolidine
derivatives
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4.8.1. Synthesis of (3R,4R)-(ꢀ)-1-benzyloxycarbonyl-3-bromo-4-
(tert-butyldimethylsilyloxy)pyrrolidine (3R,4R)-(ꢀ)-10
To a solution of enantiomerically pure product (ꢀ)-trans-6
(25 mg, 0.083 mmol), obtained from the CAL-B catalyzed hydroly-
sis, in CH2Cl2 (8.8 mL), imidazole (7.48 mg, 0.11 mmol), and t-
butyldimethylsilyl chloride, TBDMSCl (15 mg, 0.1 mmol), was
added under a nitrogen atmosphere at room temperature. The
mixture was then stirred for 24 h. The crude residue was purified
by flash chromatography on silica gel (hexane/EtOAc 75:35) to af-
ford the product as a colorless oil in 88% yield. ½a D25
¼ ꢀ3:0 (c 0.08,
ꢂ
CHCl3), ee >99%. 1H NMR (CDCl3, 300.13 MHz): d 7.38 (m, 5H), 5.18
(s, 2H), 4.41 (m, 1H), 4.18–4.01 (m, 2H), 3.97–3.72 (m, 2H), 3.40 (m,
1H), 0.89 (s, 9H), 0.15 (s, 6H); 13C NMR (CDCl3, 75.5 MHz): d 155.4
(CO), 137.1 (C), 128.9 (CH), 128.4 (CH), 128.3 (CH), 77.6 (CH), 67.4
(CH2), 53.4 (CH), 52.7 (CH2), 50.7 (CH2), 26.02 (CH3), 18.3 (C), ꢀ4.3
6. Chen, C. S.; Fujimoto, Y.; Girdaukas, G.; Sih, C. J. J. Am. Chem. Soc. 1982, 104, 7294.
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