Synthesis of isoxazolin-5-one glucosides by a cascade reaction

Article abstract of DOI:10.1021/jo4023155

A novel synthetic route was developed for the construction of isoxazolin-5-one glucosides using a cascade reaction. An X-ray crystal structure analysis of a isoxazolin-5-one glucoside confirmed the structure and stereochemistry of the heterocycle. The properties of the α- and β-anomers of the isoxazolin-5-one glucosides were compared. The first synthesis of 2-[6′-(3″-nitropropanoyl)-β-d-glucopyranosyl]-3- isoxazolin-5-one was realized by direct enzymatic esterification without need of further protective groups.

Full text of DOI:10.1021/jo4023155

Article
pubs.acs.org/joc
Synthesis of Isoxazolin-5-one Glucosides by a Cascade Reaction
Tobias Becker,† Helmar Gorls,‡ Gerhard Pauls,† Roland Wedekind,† Marco Kai,† Stephan H. von Reuß,†
̈
and Wilhelm Boland*,†
†Max Planck Institute for Chemical Ecology, Beutenberg Campus, Hans-Knoell-Straße 8, D-07745 Jena, Germany
‡Institute for Inorganic and Analytical Chemistry, Humboldtstraße 8, D-07743 Jena, Germany
S
* Supporting Information
ABSTRACT: A novel synthetic route was developed for the
construction of isoxazolin-5-one glucosides using a cascade
reaction. An X-ray crystal structure analysis of a isoxazolin-5-
one glucoside confirmed the structure and stereochemistry of the
heterocycle. The properties of the α- and β-anomers of the
isoxazolin-5-one glucosides were compared. The first synthesis of
2-[6′-(3″-nitropropanoyl)-β-D-glucopyranosyl]-3-isoxazolin-5-one
was realized by direct enzymatic esterification without need of
further protective groups.
strategies for the construction of isoxazolin-5-one moieties
have been reported.19−24 Van Rompuy et al. described a
nucleophilic substitution of organohalides with the sodium salt
of isoxazolin-5-one.20−22 This approach requires complicated
purification procedures and provides low yields because of the
formation of many side products when applied to the synthesis
of glucose derivatives.22 Hence, Baldwin and co-workers
developed an alternative method based on a 5-endo-dig
cyclization reaction to synthesize amino acid derivatives of
isoxazolin-5-one.23−26
INTRODUCTION
■
Isoxazolin-5-one glucosides are natural compounds that were
identified as metabolites in legumes (Fabaceae) as well as in
certain species of leaf beetles (Chrysomelidae).1−6 2-(β-D-
Glucopyranosyl)-3-isoxazolin-5-one (1) and its nitropropanoyl
derivative 2 have been reported to be major components of the
defensive secretions of adult leaf beetles.1−3 Seedlings of
Lathyrus odoratus show concentrations of compound 1 of up to
0.8% of their dry mass (figure 1).4
Herein we report a novel direct synthetic route for
compound 1 based on a cascade reaction consisting of a 6-
exo-trig ring closure followed by a 5-endo-dig reaction. The
synthesis of 2-[6′-(3″-nitropropanoyl)-β-D-glucopyranosyl]-3-
isoxazolin-5-one (2) was achieved by the regioselective
transesterification of an activated ester of 3-NPA to compound
1. Furthermore our novel synthetic strategy allows for the first
time the synthesis of α-configured 3,4-unsaturated isoxazolin-5-
one glucosides.
Figure 1. Isoxazolin-5-one glucosides.
A variety of different glucose derivatives of 3-nitropropanoic
acid (3-NPA) have been found in several plant, insect, and
fungi families.7 It has been shown that the acid component 3-
NPA, is an irreversible inhibitor of succinate dehydrogenase.8
As a consequence the mitochondrial citric acid cycle is
inhibited, causing neurodegenerative symptoms.9 The esters
of 3-NPA serve as pretoxins and storage molecules in different
organisms and represent an important class of defensive
compounds.10−13 In order to provide standards for studies
concerning the biosynthesis, quantification, and hydrolysis
kinetics of 2, the synthesis of compounds 1 and 2 is of interest.
In the case of compound 1, no efficient synthesis is known, and
no synthesis of the 3-NPA derivative 2 has been reported.
The regioselective synthesis of pyranose esters via trans-
esterification has been intensively studied.14−18 Different
RESULTS AND DISCUSSION
■
We commenced our work with the synthesis of (E)- and (Z)-
2,3,4,6-tetra-O-benzyl-D-glucose oxime (3) starting from
commercially available 2,3,4,6-tetra-O-benzyl-D-glucopyranose
(TOBG).27 An excess of hydroxylamine formed in situ via
deprotonation of its hydrochloride salt by sodium ethanolate
solution in ethanol28 afforded isomers (E/Z)-3 in quantitative
yield (Scheme 1).
Received: October 22, 2013
Published: November 22, 2013
© 2013 American Chemical Society
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6 are formed in a ratio of ca. 1:3. Compounds 5 and 6 were
isolated by column chromatography, and their structures were
confirmed by NMR, IR, and UV spectroscopy as well as
HRMS. In addition, an X-ray crystal structure analysis of
compound 6 was performed after crystallization from ethanol/
ethyl acetate (Figure 2); this represents the first crystal
Scheme 1. Solvent-Dependent Formation of 3 and 4
A solvent-dependent equilibrium between the aldoxime
forms (E/Z)-3 and the N-hydroxylamine pyranose isomers
1
α,β-4 was observed. H NMR spectra in CDCl3 showed a
relative 3/4 ratio of 3:1, while in CD3CN the cyclic isomers
α,β-4 were absent. Because of the complexity of the mixture,
the α/β-ratio of isomers 4 was not determined.
We found (E/Z)-3 to be transformed into isoxazolin-5-one
derivatives 5 and 6 in a one-pot synthesis when propynoic acid
was applied under Steglich conditions29 in MeCN (Scheme 2).
Scheme 2. Proposed Mechanism and Reaction Pathway for
the Cascade Reaction
Figure 2. Ellipsoid plot of glucoside 6 (50% probability). Gray,
carbon; red, oxygen; blue, nitrogen; green, hydrogen. Hydrogen atoms
have been drawn with arbitrary units.
structure of an isoxazolin-5-one glycoside. The resulting bond
lengths and angles confirmed the structure of the heterocycle as
an isoxazolin-5-one instead of its isoxazolin-3-one isomer, as
shown. This observation rules out a possible intramolecular
propynoyl transfer from O-1 to N-2 in ii. Furthermore, the
absolute configuration of compound 6 was confirmed.
1
The H NMR spectra of compounds 5 and 6 show doublet
signals with chemical shifts and coupling constants typical for
isoxazolin-5-one moieties (8.3−8.1 and 5.3−5.1 ppm; J3,4 = 3.7
Hz)21,23,24 and anomeric protons of the glucose ring (α: 5.53
ppm, J1′,2′ = 5.7 Hz and β: 4.90 ppm, J1′,2′ = 9.1 Hz). The 13C
NMR spectra show signals at 154.8 (Cα-3), 155.2 (Cβ-3), 89.3
(Cα-4), 92.8 (Cβ-4), 85.9 (Cα-1′), and 89.2 ppm (Cβ-1′). The
glucosides 5 and 6 show UV absorptions around 205 nm (ε =
39 000 L mol−1 cm−1) and 260 nm (ε = 12 100 L mol−1 cm−1)
corresponding to the benzyl and isoxazolin-5-one moieties. The
IR spectra show two characteristic absorptions at 1750 cm−1
(CO stretch) and 1550 cm−1 (CC stretch of the
isoxazolin-5-one double bond). The spectroscopic data are
consistent with literature values.21−24
Deprotection of compounds 5 and 6 was achieved using
boron trichloride at −84 °C in dichloromethane32 to give 2-(α-
D-glucopyranosyl)-3-isoxazolin-5-one (8) and 2-(β-D-glucopyr-
anosyl)-3-isoxazolin-5-one (1) in high yields and purity
(Scheme 3). The spectral data for compound 1 are identical
with literature values.3,22 The 1H NMR spectrum of compound
8 shows doublets at 8.52 and 5.39 ppm (J3,4 = 3.7 Hz)
corresponding to the isoxazolin-5-one moiety, which can be
discriminated from those of the β-anomer 1 (8.49 and 5.52
ppm, J3,4 = 3.7 Hz). The anomeric proton in compound 8 (Hα-
1′) shows a doublet at 5.73 ppm (J1′,2′ = 6.1 Hz), while the Hβ-
1′ doublet shows a signal at 5.14 ppm (J1′,2′ = 9.2 Hz). The
The reaction is initiated by a chemoselective acylation of
aldoxime 3 with propynoic acid to produce intermediates (E/
Z)-i followed by two consecutive ring-closure steps to provide a
mixture of 5 and 6. The course of the reaction was followed by
1H and 13C NMR measurements at rt, which provided evidence
for the formation of intermediates (E/Z)-i by a low-field shift of
the oxime doublets from 7.41 ppm (HE-1) and 6.87 ppm (HZ-
1) to 7.95 and 7.85 ppm, respectively. We observed that the H-
1 oxime signals in 3 disappeared nearly quantitatively after the
application of DCC/propynoic acid. Over a period of several
days the integrals of the oxime doublets decreased while the
intensities of the isoxazolin-5-one doublets increased. This
observation can be explained by an intramolecular nucleophilic
attack of the 5′-OH group on the 1′-carbon of the oxime
moiety in (E/Z)-i (6-exo-trig reaction) to form the α-and β-
anomers (α,β-ii). The isomers α,β-ii are transformed into the
corresponding isoxazolin-5-one glucosides by a second
nucleophilic attack of the nitrogen atom on the β-position of
the propynoyl group (5-endo-dig reaction). For these
sequential nucleophilic processes, the terms nucleophilic
cascade reaction30 or homodomino reaction31 can be used.
The integration of the heterocyclic protons (H-3 and H-4) in
the 1H NMR spectra indicated that the α- and β-anomers 5 and
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Methanol was added as a reference for 13C NMR spectra in D2O.
Assignment of peaks was carried out using 2D NMR experiments
(COSY, HSQC, and HMBC). The multiplicities are given as follows:
br, broad; s, singlet; d, doublet; t, triplet; dd, doublet of doublets; ddd,
doublet of doublets of doublets; m, multiplet. High-resolution mass
spectra were recorded on a UHR-qTOF and an APCI-OrbitrapXL
mass spectrometer. The intensity data for the X-ray analysis were
collected on a diffractometer using graphite-monochromatized Mo Kα
radiation. Data were corrected for Lorentz and polarization effects but
not for absorption effects.33,34 The structures were solved by direct
methods (SHELXS35) and refined by full-matrix least-squares
Scheme 3. Reaction Conditions for the Synthesis of
Compounds 1, 2, 7, and 8
techniques against Fo (SHELXL-9735). HPLC-MS analyses were
2
carried out using the APCI mode (vaporizer temperature 500 °C,
capillary temperature 300 °C) connected to an HPLC system
equipped with an RP18 column. Thin-layer chromatography was
performed on TLC silica gel 60 F254 aluminum sheets. Compounds
containing nitro groups were visualized using the modified Griess
assay. Preparative column chromatography was carried out using silica
gel (30−60 μm). All chemicals were purchased in the highest purity
that was commercially available and used without further purification.
All solvents except for diethyl ether were purchased in HPLC grade
and used without further purification. Dichloromethane, tert-butyl
alcohol, and acetonitrile were dried over activated molecular sieves (4
Å) under an argon atmosphere. Diethyl ether was distilled before use.
Synthesis of 2,3,4,6-Tetra-O-benzyl-2-(α-D-glucopyranosyl)-
3-isoxazolin-5-one (5) and 2,3,4,6-Tetra-O-benzyl-2-(β-D-glu-
copyranosyl)-3-isoxazolin-5-one (6). To a solution of propynoic
acid (1.12 g, 16 mmol, 1.1 equiv) in 1 mL of dry MeCN, a solution of
oxime 3 (8.0 g, 14.4 mmol) in 7 mL and a solution of DCC (3.09 g, 15
mmol, 1.04 equiv) in 5.3 mL of dry MeCN were added dropwise over
20 min simultaneously at rt. After 7 days of stirring at rt, the mixture
was concentrated under reduced pressure at 30 °C, and the residue
was taken up in diethyl ether (10 mL). The colorless precipitate was
filtered off, and the filtrate was concentrated under reduced pressure at
30 °C. The crude product was purified by column chromatography
(DCM/MeCN 100:1 and CHCl3/ethyl acetate 95:5). Methanol was
added, and the solvents were removed under reduced pressure. This
procedure was repeated five times to yield the title compound 6 as a
colorless solid (2.5 g, 4.11 mmol, 28.5%). Crystals suitable for an X-ray
crystal structure analysis were obtained via recrystallization from
corresponding 13C NMR signals appear at 154.8 (Cα-3), 155.2
(Cβ −3), 88.2 (Cα-4), 92.8 (Cβ-4), 87.1 (Cα-1′), and 89.2 ppm
(Cβ-1′). NMR spectra showed that compounds 1 and 8 are
stable in D2O and CD3OD at rt and neutral pH. The glucosides
1 and 8 exhibit strong UV absorption at 261 nm (ε = 10 800 L
mol−1 cm−1) as well as IR absorptions typical for isoxazolin-5-
one moieties at 1720−1730 cm−1 (CO stretch) and 1550
cm−1 (CC stretch).
The regioselective acylation of glucosides can be achieved
without the use of protective groups by enzymatic trans-
esterification reactions in organic solvents.14 2,2,2-Trichlor-
oethyl 3-nitropropanoate (7) was synthesized applying Steglich
conditions29 to commercially available 3-NPA and 2,2,2-
trichloroethanol (TCE) and purified by chromatography.
Esterification of 1 was realized using immobilized Candida
antarctica lipase B (CALB) and 7 as an activated acyl transfer
agent to obtain 2-[6′-(3″-nitropropanoyl)-β-D-glucopyranosyl]-
3-isoxazolin-5-one (2) (Scheme 3). A regioselectivity of about
95% was determined. The nonconverted 2-(β-D-glucopyrano-
syl)-3-isoxazolin-5-one 1 could be recovered by chromatog-
raphy. The spectral data of compound 2 were identical with the
literature values.3
ethanol/ethyl acetate. [α]D22 −16.6 (c 0.55, CHCl3); Rf = 0.30 (ethyl
1
acetate/CHCl3 5:95); H NMR (500 MHz, CD3CN) δ 8.16 (d, J3,4
=
3.7 Hz, 1H, H-3), 7.36−7.20 (m, 20H, Ar−H), 5.33 (d, J3,4 = 3.7 Hz,
1H, H-4), 4.90 (d, J1′,2′ = 9.1 Hz, 1H, H-1′), 4.87 (d, J = 11.2 Hz, 1H,
OCH2Ph), 4.84 (d, J = 11.2 Hz, 1H, OCH2Ph), 4.78 (d, J = 10.9 Hz,
1H, OCH2Ph), 4.76 (d, J = 11.0 Hz, 1H, OCH2Ph), 4.63 (d, J = 11.0
Hz, 1H, OCH2Ph), 4.57 (d, J = 10.9 Hz, 1H, OCH2Ph), 4.52 (d, J =
12.1 Hz, 1H, OCH2Ph), 4.48 (d, J = 12.1 Hz, 1H, OCH2Ph), 3.90 (t,
CONCLUSION
■
In conclusion, the first total synthesis of naturally occurring 2-
[6′-(3″-nitropropanoyl)-β-D-glucopyranosyl]-3-isoxazolin-5-
one (2) using an efficient four-step synthetic route is reported.
A novel cascade reaction pathway was found to incorporate 3,4-
unsubstituted isoxazolin-5-one moieties at the 1-position of
glucose to afford 2-(β-D-glucopyranosyl)-3-isoxazolin-5-one
(1). In addition, the first synthesis of α-configured isoxazolin-
5-one glucosides (5 and 8) has been described. Finally, we have
presented the first crystal structure of an isoxazoline-5-one
glycoside.
J
1′,2′ = 9.1 Hz, 1H, H-2′), 3.76−3.72 (m, 1H, H-3′), 3.68−3.62 (m, 2H,
H-6′), 3.61−3.56 (m, 2H, H-4′ and H-5′); 13C NMR (125 MHz,
CD3CN) δ 171.6 (C-5), 155.2 (C-3), 139.6 (Ar-Cq), 139.3 (Ar-Cq),
139.3 (Ar-Cq), 139.0 (Ar-Cq), 129.4 (Ar-C), 129.4 (Ar-C), 129.3 (Ar-
C), 129.3 (Ar-C), 129.2 (Ar-C), 129.2 (Ar-C), 129.1 (Ar-C), 129.0
(Ar-C), 128.9 (Ar-C), 128.8 (Ar-C), 128.8 (Ar-C), 128.8 (Ar-C),
128.7 (Ar-C), 128.6 (Ar-C), 128.6 (Ar-C), 128.5 (Ar-C), 92.8 (C-4),
89.2 (C-1′), 86.0 (C-3′), 78.9 (C-2′), 78.3 (C-4′ or C-5′), 77.7 (C-4′
or C-5′), 76.2 (OCH2Ph), 75.6 (OCH2Ph), 75.4 (OCH2Ph), 73.7
(OCH2Ph), 69.4 (C-6′); HRMS (APCI-Orbitrap) m/z calcd for
C37H37NO7Na 630.2462 [M + Na]+, found 630.2449; IR (thin film,
cm−1) 3063 (m), 3031 (m), 2961 (m), 2915 (m), 2869 (m), 1750 (s),
1556 (m), 1090 (s); UV (MeOH) λmax/nm (ε/L mol−1 cm−1) 205
(39340 650), 261 (12140 200); mp 97−99 °C. Compound 6
crystallized with two symmetrically independent molecules per
asymmetric unit. The two molecules were identical except for the
orientation of one benzyl group. The hydrogen atoms of the
isoxazolin-5-one ring were located by difference Fourier synthesis
and refined isotropically. All other hydrogen atoms were included at
calculated positions with fixed thermal parameters. All non-hydrogen
atoms were refined anisotropically.35 XP was used for structure
representations. Crystal data for 6: C37H37NO7, Mr = 607.703 g mol−1,
EXPERIMENTAL SECTION
■
General Experimental Methods. Melting points were deter-
mined using a capillary melting point apparatus. Infrared spectra were
measured with a IR spectrometer over the 700−4000 cm−1 range in
transmission mode with a spectral resolution of 2 cm−1. Optical
rotations were measured at 589 nm and 22 °C. Ultraviolet spectra
were recorded on a UV spectrophotometer over the range from 190 to
300 nm. NMR spectra were measured using a spectrometer operating
at 500 MHz (1H) and 125 MHz (13C). Chemical shifts (δ) are quoted
in parts per million (ppm) and are referenced to the signals of residual
protonated solvents (CD2HCN at 1.94 ppm and CHCl3 at 7.26 ppm).
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colorless prism, size 0.10 mm × 0.10 mm × 0.09 mm, triclinic, space
group P1, a = 10.6443(5) Å, b = 12.4845(6) Å, c = 12.8966(6) Å, α =
104.609(2)°, β = 96.556(2)°, γ = 104.520(2)°, V = 1576.38(13) Å3, T
= 23 °C, Z = 2, ρcalcd = 1.280 g cm−3, μ(Mo Kα) = 0.88 cm−1, F(000)
= 644, 70 532 reflections in h(−12/12), k(−14/14), l(−15/15)
(C-2′), 69.5 (C-4′), 61.0 (C-6′); HRMS (ESI-TOF) m/z calcd for
C9H13NO7Na 270.0584 [M + Na]+, found 270.0575; IR (thin film,
cm−1) 3370 (br, s), 2932 (m), 1725 (s), 1544 (s), 1104 (s), 1040 (s);
UV (MeOH) λmax/nm (ε/L mol−1 cm−1) 262 (10800 200). The
spectral data were in agreement with the literature values.3,22
measured in the range 2.72° ≤ Θ ≤ 25.03°, completeness Θmax
=
Synthesis of 2-(α-D-Glucopyranosyl)-3-isoxazolin-5-one (8).
2,3,4,6-Tetra-O-benzyl-2-(α-D-glucopyranosyl)-3-isoxazolin-5-one (5)
(100 mg, 0.165 mmol) was dissolved in dry DCM (8 mL) and cooled
to −84 °C under an argon atmosphere. Boron trichloride (2 mL, 1 M,
heptane, 10 equiv) was added dropwise. After 1.5 h at −84 to −79 °C,
the mixture was warmed to rt and stirred for 1.5 h. The reaction was
quenched with methanol (4 mL) at −75 °C and stirred for 30 min.
The solvents were removed at 25 °C under reduced pressure to yield a
colorless oil. The residue was dissolved in methanol (5 mL), and silica
(500 mg) was added. The solvent was removed, and the residual
colorless powder was applied to a silica gel column. After elution of the
column (DCM/MeOH 5:1 to 2:1), the product fractions were
concentrated under high vacuum at rt to yield 8 as a colorless oil (33.3
mg, 0.135 mmol, 81.8%). [α]2D2 +85.3 (c 1.5, MeOH); Rf = 0.49
99.9%, 11066 independent reflections, Rint = 0.0209, 10606 reflections
with Fo > 4σ(Fo), 1107 parameters, 3 restraints, R1(obs) = 0.0253,
wR2(obs) = 0.0614, R1(all) = 0.0273, wR2(all) = 0.0626, GOF = 1.033,
Flack parameter = 0.0(3), largest difference peak/hole 0.161/−0.155 e
Å−3. Crystallographic data (excluding structure factors) have been
deposited with the Cambridge Crystallographic Data Centre as
supplementary publication CCDC-961971. Copies of the data can
be obtained free of charge from the CCDC, 12 Union Road,
Cambridge CB2 1EZ, U.K. (E-mail: deposit@ccdc.cam.ac.uk).
The α-anomer 5 was isolated as a colorless oil (605 mg, 0.99 mmol,
6.8%). [α]2D2 +78.5 (c 0.51, CHCl3); Rf = 0.41 (ethyl acetate/CHCl3
5:95); 1H NMR (500 MHz, CD3CN) δ 8.26 (d, J3,4 = 3.7 Hz, 1H, H-
3), 7.36−7.21 (m, 20H, Ar−H), 5.53 (d, J1′,2′ = 5.7 Hz, 1H, H-1′), 5.17
(d, J3,4 = 3.7 Hz, 1H, H-4), 4.88 (d, J = 11.1 Hz, 1H, OCH2Ph), 4.80
(d, J = 11.1 Hz, 2H, OCH2Ph) 4.65 (s, 2H, OCH2Ph), 4.57 (d, J =
11.0 Hz, 1H, OCH2Ph), 4.51 (d, J = 11.9 Hz, 1H, OCH2Ph), 4.46 (d, J
= 11.9 Hz, 1H, OCH2Ph), 4.10 (t, J3′,4′ = 9.1 Hz, 1H, H-3′), 3.95 (dd,
J2′,3′ = 9.6 Hz, J1′,2′ = 5.8 Hz, 1H, H-2′), 3.89−3.85 (m, 1H, H-5′),
3.68−3.62 (m, 2H, H-6′), 3.60 (dd, J4′,5′ = 10.2 Hz, J3′,4′ = 8.7 Hz, 1H,
H-4′); 13C NMR (125 MHz, CD3CN) δ 171.7 (C-5), 154.8 (C-3),
139.7 (Ar-Cq), 139.4 (Ar-Cq), 139.3 (Ar-Cq), 138.7 (Ar-Cq), 129.4
(Ar-C), 129.3 (Ar-C), 129.2 (Ar-C), 129.1 (Ar-C), 128.9 (Ar-C),
128.9 (Ar-C), 128.8 (Ar-C), 128.6 (Ar-C), 128.5 (Ar-C), 89.3 (C-4),
85.9 (C-1′), 82.8 (C-3′), 79.7 (C-2′), 78.2 (C-4′), 75.9 (OCH2Ph),
75.5 (OCH2Ph), 75.4 (C-5′), 74.2 (OCH2Ph), 73.8 (OCH2Ph), 69.7
(C-6′); HRMS (APCI-Orbitrap) m/z calcd for C37H41N2O7 625.2908
[M + NH4]+, found 625.2892; IR (thin film, cm−1) 3087 (m), 3062
(m), 3030 (m), 2923 (m), 2867 (m), 1749 (s), 1552 (m), 1093 (s);
UV (MeOH) λmax/nm (ε/L mol−1 cm−1) 204 (36740 600), 264
(12110 200).
1
(DCM/MeOH 2:1); H NMR (500 MHz, D2O) δ 8.52 (d, J3,4 = 3.6
Hz, 1H, H-3), 5.73 (d, J1′,2′ = 6.1 Hz, 1H, H-1′), 5.39 (d, J3,4 = 3.6 Hz,
1H, H-4), 4.10 (t, J2′,3′ = 9.6 Hz, 1H, H-3′), 4.02 (dd, J2′,3′ = 10.0 Hz,
J1′,2′ = 6.2 Hz, 1H, H-2′), 3.87−3.80 (m, 2H, H-5′ and HA-6′), 3.78−
3.73 (m, 1H, HB-6′) 3.53 (t, J3′,4′ = 9.5 Hz, 1H, H-4′); 13C NMR (125
MHz, D2O) δ 174.9 (C-5), 154.8 (C-3), 88.2 (C-4), 87.1 (C-1′), 77.1
(C-5′), 74.1 (C-3′), 70.6 (C-2′), 69.7 (C-4′), 61.1 (C-6′); HRMS
(APCI-Orbitrap) m/z calcd for C9H14NO7 248.0765 [M + H]+, found
248.0762; IR (thin film, cm−1) 3382 (br, s), 2962 (m), 2923 (m), 1727
(s), 1552 (s), 1192 (s), 1063 (s); UV (MeOH) λmax/nm (ε/L mol−1
cm−1) 261 (10760 200).
Synthesis of 2-[6′-(3″-Nitropropanoyl)-β-D-glucopyranosyl]-
3-isoxazolin-5-one (2). A mixture of 2-(β-D-glucopyranosyl)-3-
isoxazolin-5-one (1) (100 mg, 0.404 mmol), 2,2,2-trichloroethyl 3-
nitropropanoate (7) (160.1 mg, 0.639 mmol), immobilized C.
antarctica lipase B (150 mg), and 4 Å molecular sieves was suspended
in dry tert-butyl alcohol (7 mL). The suspension was stirred at 50 °C
under an argon atmosphere for 18 h. The enzyme was filtered off, and
the filter cake was washed with tert-butyl alcohol (2 × 5 mL) at 40 °C
and cold methanol (5 mL). The filtrate was concentrated under
reduced pressure at 25 °C, and the residue was taken up in methanol.
Silica was added, and the solvent was evaporated under reduced
pressure at 25 °C to obtain a colorless powder. The dry powder was
added to a silica column, and the product was purified by column
chromatography (ethyl acetate/MeOH/DCM 10:1:1 to 2:1:0). The
solvent was removed to yield 2 as a colorless solid (50 mg, 0.144
mmol, 35.6%). Nonconverted glucoside 1 could be recovered (31 mg,
0.125 mmol, 31%). [α]2D2 +30.1 (c 0.36, MeOH); Rf = 0.20 (ethyl
acetate/MeOH/DCM 10:1:1); 1H NMR (500 MHz, D2O) δ 8.47 (d,
Synthesis of 2,2,2-Trichloroethyl 3-Nitropropanoate (7). 3-
Nitropropanoic acid (687 mg, 5.77 mmol), 2,2,2-trichloroethanol
(3.45 g, 23.08 mmol, 4 equiv), and DMAP (63.4 mg, 0.52 mmol, 9
mol %) were dissolved in dry DCM (5.77 mL). The mixture was
cooled to 0 °C, and DCC (1.308 g, 6.35 mmol, 1.1 equiv) was added
all at once. After 10 min at 0 °C, the mixture was heated to rt and
stirred for 3 h. After purification by flash column chromatography
(CHCl3) and removal of the solvent at 40 °C under reduced pressure,
a colorless powder of 7 (834 mg, 3.33 mmol, 57.7%) was obtained. Rf
= 0.78 (CHCl3); 1H NMR (500 MHz, CDCl3) δ 4.81 (s, 2H,
CH2CCl3), 4.72 (t, J2,3 = 6.1 Hz, 2H, CH2NO2), 3.16 (t, J2,3 = 6.1 Hz,
2H, CH2CO2R); 13C NMR (125 MHz, CDCl3) δ 168.13, 94.46, 74.59,
69.35, 30.97; HRMS (APCI-Orbitrap) m/z calcd for C5H7Cl3NO4
249.9435 [M + H]+, found 249.9429; IR (thin film, cm−1) 3012 (w),
2961 (m), 2926 (m), 1747 (s), 1549 (s), 1088 (s); mp 35−36 °C.
Synthesis of 2-(β-D-Glucopyranosyl)-3-isoxazolin-5-one (1).
2,3,4,6-Tetra-O-benzyl-2-(β-D-glucopyranosyl)-3-isoxazolin-5-one (6)
(1.73 g, 2.85 mmol) was dissolved in dry DCM (150 mL) and cooled
to −84 °C under an argon atmosphere. Boron trichloride (20 mL, 1
M, heptane, 7 equiv) was added dropwise. After 17 h at −84 to −79
°C, the mixture was quenched with methanol (20 mL) and warmed to
rt. The solvents were removed at 25 °C under reduced pressure to
yield a colorless oil. The residue was dissolved in methanol (5 mL),
and silica (2 g) was added. The solvent was removed, and the residual
colorless powder was applied to a silica gel column. After elution of the
column (DCM/MeOH 5:1 to 2:1), the pure fractions were
concentrated under high vacuum at rt to yield 1 as a colorless solid
(522 mg, 2.11 mmol, 74.2%). [α]2D2 +13.5 (c 0.53, MeOH); Rf = 0.49
J3,4 = 3.7 Hz, 1H, H-3), 5.52 (d, J3,4 = 3.7 Hz, 1H, H-4), 5.15 (d, J1′,2′
=
9.2 Hz, 1H, H-1′), 4.82 (t, J2″,3″ = 5.8 Hz, 2H, H-3″), 4.51 (dd, JA6′,B6′
= 12.3 Hz, J5′,A6′ = 2.2 Hz, 1H, HA-6′), 4.33 (dd, JA6′,B6′ = 12.4 Hz,
J5′,B6′ = 5.2 Hz, 1H, HB-6′), 3.91 (t, J1′,2′ = 9.2 Hz, 1H, H-2′), 3.78
(ddd, J4′,5′ = 10.0 Hz, J5′,B6′ = 5.2 Hz, J5′,A6′ = 2.2 Hz, 1H, H-5′), 3.62 (t,
J2′,3′ = 9.3 Hz, 1H, H-3′), 3.50 (t, J3′,4′ = 9.5 Hz, 1H, H-4′), 3.13 (t,
J
2″,3″ = 5.8 Hz, 2H, H-2″); 13C NMR (125 MHz, D2O) δ 174.7 (C-5),
172.4 (C-1″), 155.0 (C-3), 91.7 (C-4), 88.7 (C-1′), 76.5 (C-5′), 76.1
(C-3′), 70.7 (C-3″), 69.9 (C-2′), 69.5 (C-4′), 63.9 (C-6′), 31.6 (C-
2″); HRMS (ESI-TOF) m/z calcd for C12H16N2O10Na 371.06972 [M
+ Na]+, found 371.06958; IR (thin film, cm−1) 3374 (br, s), 2924 (m),
2887 (m), 1725 (s), 1549 (s), 1067 (br, s); UV (MeOH) λmax/nm (ε/
L mol−1 cm−1) 201 (6250 110), 261 (11040 200). The spectral
data were in agreement with the literature values.3
1
ASSOCIATED CONTENT
(DCM/MeOH 2:1); H NMR (500 MHz, D2O) δ 8.49 (d, J3,4 = 3.7
■
Hz, 1H, H-3), 5.50 (d, J3,4 = 3.7 Hz, 1H, H-4), 5.14 (d, J1′,2′ = 9.2 Hz,
1H, H-1′), 3.92−3.88 (m, 2H, HA-6′ and H-2′), 3.73 (dd, J1 = 6.9 Hz,
J2 = 5.6 Hz, 1H, HB-6′), 3.64−3.57 (m, 2H, H-3′ and H-5′), 3.49 (t,
S
* Supporting Information
Crystallographic data for compound 6 (CIF); NMR spectra for
compounds 1, 2, and 5−8; HRMS spectra of compounds 5−8;
J
3′,4′ = 9.5 Hz, 1H, H-4′); 13C NMR (125 MHz, D2O) δ 174.8 (C-5),
1
154.8 (C-3), 91.1 (C-4), 88.8 (C-1′), 78.8 (C-5′), 76.7 (C-3′), 70.0
HPLC-MS analyses of 1 and 8; and H NMR spectra of the
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dx.doi.org/10.1021/jo4023155 | J. Org. Chem. 2013, 78, 12779−12783

The Journal of Organic Chemistry
Article
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formation of compounds 5 and 6. This material is available free
of charge via the Internet at http://pubs.acs.org.
AUTHOR INFORMATION
■
Corresponding Author
*E-mail: boland@ice.mpg.de.
Notes
The authors declare no competing financial interest.
(34) Otwinowski, Z.; Minor, W. Methods Enzymol. 1997, 276, 307−
ACKNOWLEDGMENTS
■
326.
We thank Dr. Maritta Kunert and Kerstin Ploss for the MS
measurements and Mina Shaheed for the IR measurements.
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