Synthesis of a ring-oxygenated variant of the 2-carboxy-6- hydroxyoctahydroindole core of aeruginosin 298-A from glucose

Article abstract of DOI:10.1021/jo0507901

The design and synthesis of a new core structure, a ring-oxygenated variant of 2-carboxy-6-hydroxyoctahydroindole (Choi) from D-glucose, is reported. Choi, a rigid bicyclic unnatural amino acid, is the core structure of about 15 aeruginosins natural compounds. These compounds are thrombin, trypsin, and factor VIIa inhibitors and Choi is important for their biological activity. The ring-oxygenated variant of 2-carboxy-6-hydroxyoctahydroindole can potentially be used as a surrogate of Choi in the design and synthesis of aeruginosin-based thrombin inhibitors. & 2005 American Chemical Society.

Full text of DOI:10.1021/jo0507901

Synthesis of a Ring-Oxygenated Variant of the
2-Carboxy-6-hydroxyoctahydroindole Core of Aeruginosin 298-A
from Glucose
Xiaoping Nie and Guijun Wang*
Department of Chemistry, University of New Orleans, New Orleans, Louisiana 70148
gwang2@uno.edu
Received April 19, 2005
The design and synthesis of a new core structure, a ring-oxygenated variant of 2-carboxy-6-
hydroxyoctahydroindole (Choi) from D-glucose, is reported. Choi, a rigid bicyclic unnatural amino
acid, is the core structure of about 15 aeruginosins natural compounds. These compounds are
thrombin, trypsin, and factor VIIa inhibitors and Choi is important for their biological activity.
The ring-oxygenated variant of 2-carboxy-6-hydroxyoctahydroindole can potentially be used as a
surrogate of Choi in the design and synthesis of aeruginosin-based thrombin inhibitors.
Introduction
ing site are potentially more selective and could have
better therapeutic profiles. Flexible inhibitors can adopt
multiple conformations but they need to rearrange and
fit into a protease binding site. Conformational strained
thrombin inhibitor with an indolizidinone motif 3 and
fused piperazinone system 4 showed good inhibitory
activity.
Thrombin (factor IIa) is a key trypsin-like serine
protease of the blood coagulation cascade, it is also the
most potent activator of platelet aggregation. Thrombin
inhibitors are important agents in treating or preventing
thrombosis disorders. Many thrombin inhibitors have
been designed and synthesized in recent years.1-4 Sub-
strate analogue strategy has been used in the design of
reversible thrombin inhibitor D-Phe-Pro-Argininal (1)5
and the irreversible potent inhibitor chromethyl ketone
D-Phe-Pro-ArgCH2Cl (2) (PPACK).6 These compounds
and the crystal structures of thrombin complexes with
inhibitors including PPACK were often used as the
starting point for structure optimizations of novel throm-
bin inhibitors.7,8 Conformational restricted inhibitors that
can preorganize into optimal conformations for the bind-
Despite much research effort, it is still very challenging
to find an orally active, safe, and effective agent for
treating or preventing thrombosis disorders. The major
anticoagulants heparin and warfrin were introduced
about 50 years ago. Currently recombinant hirudin and
argatroban are approved for the treatment of thrombosis
associated with heparin induced thrombocytopenia (HIT).
Hirudin (66 amino acids) and its analogue hirulog (20
amino acids) are polypetides isolated from medicinal
leeches.9 Argatroban is a synthetic small molecule throm-
bin inhibitor, it was recently approved for use as an
anticoagulant for treating prophylaxis, HIT, and associ-
ated thrombotic complications.10 Like heparin, both hiru-
din and argatroban are administered parenterally. Ximel-
agatran is an orally active thrombin inhibitor and
* Address correspondence to this author. Phone: 504 280-1258.
Fax: 504 280-6860.
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2005, 25, 66-92.
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10.1021/jo0507901 CCC: $30.25 © 2005 American Chemical Society
Published on Web 09/27/2005
J. Org. Chem. 2005, 70, 8687-8692
8687

Nie and Wang
FIGURE 1. Structures of aeruginosian 298-A, occillarin,
FIGURE 2. X-ray crystal structures of thrombin inhibited by
oscillarin. This is obtained from protein data bank code
1R1W.22 The Choi ring is number labeled in the molecular
structure (a) and the crystal structure (b). The substrate
binding sites are also labeled in the oscillarin structure.
dysinosin A, and aeruginosin EI 461.
is in a phase III clinical trial as an antithrombotic
drug.11
In recent years, a new class of marine nature products,
the aeruginosins, were isolated from cyanobacterial
blooms.12-20 They are small linear peptides containing
unnatural amino acids and they are found to be serine
protease inhibitors. The approximately 16 compounds in
this family share a common new bicyclic amino acid core
structure 2-carboxy-6-hydroxyoctahydroindole (Choi) (5).
stereochemistry observed in aeruginosin 298-A, the op-
posite configuration of the Choi bicycle has also been
reported. Aeruginosin EI46123 contains one less amino
acid unit and the Choi has opposite stereochemistry to
that of aeruginosin 298-A. Dysinosin A contains a new
Choi structure with 5,6-dihydroxyl substituents, and it
is a factor VIIa inhibitor (Ki ) 0.11 µM) and thrombin
inhibitor (Ki ) 0.45 µM).25,26
Structure Design
The X-ray crystal structures of thrombin inhibiting by
several aeruginosins have been obtained through the
inhibitor and thrombin-hirugen complexes.21,22,25 The
binding mode of aeruginosin 298-A resembles closely that
of D-Phe-Pro-Arg chloromethyl ketone6 and other serine
protease inhibitors.1-4 Aeruginosin 298-A binds to the
active site of thrombin in a noncovalent way forming an
antiparallel strand with thrombin.21 The five-membered
ring of the Choi residue occupies the hydrophobic binding
site, while its six-membered ring projects out and loosely
interacts with Try 60 and Tyr 60 from thrombin. The
active site where the Choi group interacts has many
degrees of freedom. The crystal structures of oscillarin
and dysinosin-A thrombin complexes revealed similar
binding patterns as aeruginosin 298-A. Dysinosin-A
forms a hydrogen bonding network with thrombin com-
plex; however, the 5,6-dihydroxyoctahydroindole group
appears not to have hydrogen bonding interactions with
the P2 binding pocket even though there are polar amino
acid residues in the pocket.25 The crystal structure of
oscillarin with thrombin complex is shown in Figure 2.
The amide NH from the octahydroindole carboxamide
forms a hydrogen bond with Ser 256; however, no
interactions were observed with the 6-hydroxyl group of
Aeruginosins are thrombin, trypsin, and factor VIIa
inhibitors and this new amino acid is the critical deter-
minant of their biological activity. The 14 initially
reported members mostly contain the configuration of
2S,3aS,6R,7aS in their azabicyclic core. The structures
of several aeruginosins are shown in Figure 1. Aerugi-
nosin 298-A exhibits thrombin and trypsin inhibition at
IC50 of 0.5 and 1.7 µM but does not significantly affect
papain, chymotrypsin, and plasmin.15,16 Oscillarin is a
relatively newer member of the family and it is a potent
thrombin inhibitor with IC50 ) 28 nM.22 It has the same
Choi structure as in aeruginosin 298-A, but with different
amino acids. The crystal structure of thrombin inhibited
by oscillarin was also published. Other than the typical
(11) Gustafsson, D.; Bylund, R.; Antonsson, T.; Nilsson, I.; Nystrom,
J. E.; Eriksson, U.; Bredberg, U.; Teger-Nilsson, A. C. Nat. Rev. Drug
Discovery 2004, 3, 649-659.
(12) Shin, H. J.; Matsuda, H.; Murakami, M.; Yamaguchi, K. J. Org.
Chem. 1997, 62, 1810-1813.
(13) Matsuda, H.; Okino, T.; Murakami, M.; Yamaguchi, K. Tetra-
hedron 1996, 52, 14501-14506.
(14) Murakami, M.; Ishida, K.; Okino, T.; Okita, Y.; Matsuda, H.;
Yamaguchi, K. Tetrahedron Lett. 1995, 36, 2785-2788.
(15) Murakami, M.; Okita, Y.; Matsuda, H.; Okino, T.; Yamaguchi,
K. Tetrahedron Lett. 1994, 35, 3129-3132.
(16) Ishida, K.; Okita, Y.; Matsuda, H.; Okino, T.; Murakami, M.
Tetrahedron 1999, 55, 10971-10988.
(21) Steiner, J. L. R.; Murakami, M.; Tulinsky, A. J. Am. Chem. Soc.
1998, 120, 597-598.
(17) Kodani, S.; Ishida, K.; Murakami, M. J. Nat. Prod. 1998, 61,
1046-1048.
(22) Hanessian, S.; Tremblay, M.; Petersen J. F. W. J. Am. Chem.
Soc. 2004, 126, 6064-6071.
(18) Ploutno, A.; Shoshan, M.; Carmeli, S. J. Nat. Prod. 2002, 65,
973-978.
(23) Valls, N.; Vallribera, M.; Carmeli, S.; Bonjoch, J. Org. Lett.
2003, 5, 447-450.
(19) Carroll, A. R.; Pierens, G. K.; Fechner, G.; de Almeida Leone,
P.; Ngo, A.; Simpson, M.; Hyde, E.; Hooper, J. N. A.; Bostrom, S. L.;
Musil, D.; Quinn, R. J. J. Am. Chem. Soc. 2002, 124, 13340-13341.
(20) Sandler, B.; Murakami, M.; Clardy, J. J. Am. Chem. Soc. 1998,
120, 595-596.
(24) Valls, N.; Vallribera, M.; Lo´pez-Canet, M.; Bonjoch, J. J. Org.
Chem. 2002, 67, 4945-4950.
(25) Carroll, A. R.; Pierens, G. K.; Fechner, G.; de Almeida Leone,
P.; Ngo, A.; Simpson, M.; Hyde, E.; Hooper, J. N. A.; Bostrom, S.-L.;
Musil, D.; Quinn, R. J. J. Am. Chem. Soc. 2002, 124, 13340-13341.
8688 J. Org. Chem., Vol. 70, No. 22, 2005

Variant of 2-Carboxy-6-hydroxyoctahydroindole
SCHEME 1. Synthetic Plan To Obtain the Choi Analogues from Methyl r-D-Glucopyranose
SCHEME 2. Synthesis of the Mesylate Bromide Intermediate 8 from D-Glucose
the Choi moiety. The terminal phenyl group has no
interaction with the enzyme.22
been carried out.23-24,26-30 The Choi core structure is
crucial for their antithrombotic activity. Several methods
have been developed for the synthesis of Choi.26-31 The
majority of strategies involve the use of amino acids as
starting materials. These include tyrosine oxidation-
rearrangement27 and Birch reduction of O-methyltyrosine
followed by benzylation and cyclization.28-30 A third
method involves glutamic acid as the starting material
through intramolecular N-acyloxyiminium ion carbocyl-
ization reactions.22 Other methods include using a pyr-
rolidine as starting material32 and a synthesis that
utilized catalytic phase transfer alkylation strategy.30,33
To optimize thrombin inhibitor structure and discover
novel antithrombosis drugs, we designed and synthesized
novel analogues of the unnatural amino acid Choi for the
preparation of aeruginosin-based thrombin inhibitors.
The Choi analogues 6 and 6′ can be prepared from readily
available D-glucose 7, through an intermediate mesylate
bromide 8 (Scheme 1). The mesylate 8 can be used to
synthesize the products by extending the molecule with
a two carbon unit that can be converted to the cyclic
amino acid later.
The synthesis of intermediate mesyalte bromide 8 is
shown in Scheme 2. The D-glucose 7 is converted to the
bromosugar 9. Reduction of 9 with tributyltin hydride
gave the deoxygenated tetraacetate 10. Deacetylating
with ethanol and sodium ethoxide afforded tetraol 11 in
quantitative yield. The selective protection of 11 to the
acetal 12 was carried out smoothly with dimethoxyl ben-
zylidene acetal in DMF. Dibenzylation and deprotection
to free the 4,6-dihydroxyl groups followed by dimesylation
gave dimesylate 15 with excellent yields. Monodisplace-
On the basis of the crystal structures and the binding
sites interactions, we can design novel inhibitors with
similar core structure motifs but with different substit-
uents and functionalization to enhance binding, activity,
and pharmacokinetic properties. In all the crystal struc-
tures of aeruginosin-thrombin complexes so far, Choi
moieties of the aeruginosins which occupy the S2 binding
site generally have a good degree of freedom. Thrombin
generally can tolerate imprecise binding from different
molecules. The rigid bicyclic amino acid structure is very
important in defining the conformation of the molecule
and it is essential in their antithrombin activity. The
6-hydroxyl group projects out and the Choi core is not
interacting strongly with residues at the active site. Thus
modifications on the core structure in this area should
not impact negatively on binding to the protein while
some changes will increase contact and enhance binding.
Such changes include introducing an oxygen at the C-4
position and/or adding an additional hydroxyl group to
the C-5 or C-7 position, these lead to structures 6 and 6′
as surrogates for 5. Because pharmacokinetics are very
unpredictable, these changes will provide other possibili-
ties that might provide scope for improving the pharma-
cokinetic profile of this compound class if required. The
availability of different stereoisomers is also important
in elucidating structure-activity relationships. Studies
to rationalize electronic and steric influence of different
inhibitors have not given good correlations so far. Modi-
fication from the natural analogues can allow us to
understand the stereoelectronic factors that determine
the activity of these inhibitors. These proposed Choi
analogues are novel structures and will have great
potential in discovering better thrombosis inhibitors.
(26) Hanessian, S.; Margarita, R.; Hall, A.; Johnstone, S.; Tremblay,
M.; Parlanti, L. J. Am. Chem. Soc. 2002, 124, 13342-13343.
(27) Wipf, P.; Methot, J. L. Org. Lett. 2000, 2, 4213-4216.
(28) Valls, N.; Lopez-Canet, M.; Vallribera, M.; Bonjoch, J. J. Am.
Chem. Soc. 2000, 122, 11248-11249.
(29) Valls, N.; Lopez-Canet, M.; Vallribera, M.; Bonjoch, J. Chem.
Eur. J. 2001, 7, 3446-3460.
(30) Ohshima, T.; Gnanadesikan, V.; Shibuguchi, T.; Fukuta, Y.;
Nemoto, T.; Shibasaki, M. J. Am. Chem. Soc. 2003, 125, 11206-11207.
(31) Bonjoch, J.; Catena, J.; Isabal, E.; Lopez-Canet, M.; Valls, N.
Tetrahedron: Asymmetry 1996, 7, 1899-1902.
(32) Toyoka, N.; Okumura, M.; Himiyama, T.; Nakazawa, A.;
Nemoto, H. Synlett 2003, 55-58.
(33) Fukuta, Y.; Ohshima, T.; Gnanadesikan, V.; Shibuguchi, T.;
Nemoto, T.; Kisugi, T.; Okino, T.; Shibasaki, M. Proc. Natl. Acad. Sci.
U.S.A. 2004, 101, 5433-5438.
Results and Discussion
Because of the important biological activity of aerugi-
nosins, the total syntheses of these natural products have
J. Org. Chem, Vol. 70, No. 22, 2005 8689

Nie and Wang
SCHEME 3. Alkylation of Intermediate 8 by Diethyl N-Acetylmalonate
SCHEME 4. Alkylation of Intermediate 8 by
Diethyl N-Boc Malonate to Choi Analogues
Intermediates 24 and 25 are the protected versions of 6
and 6′. The fact that they can be separated easily allows
us to study the stereochemical influences of the Choi core
structures to the aeruginosin analogues’ properties.
The stereochemistry of compounds 24 and 25 is con-
firmed by their 1H NMR and 2D-NOESY spectra. Figure
3 shows the assigned structures and the expected NOE
ment of the primary mesylate was achieved with sodium
bromide in DMSO at 60 °C to furnish intermediate 8.
To extend intermediate 8 with the two-carbon glycine
unit, several experiments were carried out. These include
the alkylation using the anion from diethyl malonate.
Diethyl acetamidomalonate 16 was first used as the two-
carbon unit and the alkylation of the anion from 16 with
bromide 8 led to the formation of intermediate 17
(Scheme 3). Bromide 8 reacted with diethyl acetamido-
malonate sluggishly with low yield under typical reaction
conditions (e.g., NaOEt/EtOH, reflux; NaH, THF or tolu-
ene reflux). Successful alkylation was achieved in 54%
yield by adding TBAI as a catalyst presumably through
a more reactive iodide intermediate and the nucleophi-
licity of the anion is enhanced by interacting with TBAI.
However, the acetamide was not acidic enough to be
deprotonated with a relatively mild base and afford the
SN2 displacement of the mesylate to give intermediate
18. Various conditions (NaH or KO-t-Bu in THF, toluene,
DMF, or DMSO) were testedsin the best case only about
10% yield of product was formed, possibly due to partial
ester hydrolysis and/or mesylate decomposition.
With the promising result of diethyl malonate, we
anticipated that the cyclization problem could be avoided
by using free amine to displace the mesylate. In this route
(Scheme 4), N-Boc protected diethyl aminomalonate 21
was used as the coupling reagent, and the anion gener-
ated from 21 was alkylated with 8 under slightly modified
reaction conditions (NaH, with TBAI as a catalyst, reflux
overnight in toluene). The alkylation product 22 was
obtained in 56% yield. The monodecarboxylation of 22
was carried out with a sodium hydroxide in water and
ethanol mixture to hydrolyze one ester group followed
by refluxing to remove the carboxylate to give an insepa-
rable diastereomer mixture of intermediate 23 in 86%
yield. The deprotection of 23 with TFA and cyclization
of the resulting amine under refluxing condition with
TEA as catalyst furnished the cyclized Choi analogues
24 and 25 in very good yield. The two diasteromers 24
and 25 can be isolated with flash chromatography.
FIGURE 3. Structures of compounds 24 and 25 and the NOE
among different protons.
among hydrogens from the five-membered ring. The
2D-NOESY spectra of the two diastereomers’ aliphatic
regions are shown in the Supporting Information.
The signals corresponding to methylene protons 3s and
3t can be assigned unambiguously from 2D-NOESY
spectra. There should be more NOE interactions for
proton 3t than 3s (Figure 3). In both isomers, there is an
additional NOE cross-peak for H3t and H8. H3s will not
have NOE interactions with H8. With this assignment
confirmed, we can further assign the stereochemistry at
C-2 base on the coupling constant differences of the two
1H NMR spectra. For compound 24, proton 3t is a ddd
centered at 2.03 ppm, 3s is a dd centered at 2.30 ppm,
the coupling constants of H3s are 13.7 Hz for the geminal
coupling and 8.8 Hz for the H-C-C-H J3 coupling with
H2. Molecular modeling of 24 indicated that the di-
hedral angle of H2-C2-C3-H3s is about 12° and that of
H9-C9-C3-H3s is about 86°; these explain the splitting
pattern and that the J3 of H9 and H3s is very small,
therefore the NMR signals degenerate to a dd. For
diastereomer 25, proton 3t is a multiplet with pattern
ddd centered at 2.21 ppm, and proton 3s is a dd centered
at 2.31 ppm. The coupling constants of H3s are 13.7 Hz
for the geminal coupling and 1.9 Hz for the H-C-C-H
coupling with H2 and/or H9. The molecular model of
compound 25 indicated that the H9-C9 bond is almost
perpendicular to the C3-H3s bond, and the J3 for
H3s-H9 is therefore very small. The dihedral angle for
H2-C2-C3-H3s is about 131°, which explains the ob-
served splitting pattern of dd and a smaller coupling
constant J3 for H3s-H2.
Compounds 24 and 25 can be used in the synthesis of
aeruginosin analogues directly by coupling at the amino
8690 J. Org. Chem., Vol. 70, No. 22, 2005

Variant of 2-Carboxy-6-hydroxyoctahydroindole
crystal. Mp 64-67 °C, [R]D +27.7 (c 1.1, CHCl3). 1H NMR (500
MHz, CDCl3) δ 7.34-7.27 (m, 10H), 5.07 (d, 1H, J ) 11.1 Hz),
4.68 (d, 1H, J ) 11.1 Hz), 4.65 (d, 1H, J ) 11.5 Hz), 4.58 (d,
1H, J ) 11.5 Hz), 4.36 (t, 1H, J ) 9.1 Hz), 4.10-4.06 (m, 1H),
3.75-3.64 (m, 2H), 3.51 (dt, 1H, J ) 7.2, 2.2 Hz), 3.41 (dd,
1H, J ) 11.3, 7.3 Hz), 3.26-3.21 (m, 1H), 2.81 (s, 3H); 13C NMR
(125 MHz, CDCl3) δ 137.6, 137.4, 128.6, 128.5, 128.2, 127.9,
127.8, 127.7, 82.4, 79.8, 78.7, 77.8, 75.3, 73.2, 67.8, 38.6, 31.8.
HRMS calcd for C21H25BrO6S + Na 507.0453, found 507.0453.
group. They can also be converted to the unprotected Choi
analogues 6 and 6′. We are currently synthesizing
aeruginosin 298-A and oscillarin analogues containing
the novel Choi core structures.
Conclusions
We have designed and synthesized ring-oxygenated
variants of Choi analogues from D-glucose in short
sequences efficiently. The two protected ring oxygenated
variants of Choi synthesized can be used in the synthesis
of aeruginosin 298-A analogues. The synthetic strategies
are versatile in constructing inhibitor core structures
with different stereochemistry. Because of the abundance
of chiral hydroxyl groups in carbohydrate raw materials,
different stereoisomers can be synthesized readily from
different sugar molecules by similar methods. During the
ring cyclization step to form the bicyclic structure, both
configurations of the amino acids can be obtained in one
step. These isomers are diastereomers of each other, thus
they can be readily separated by chromatography. This
allows quick access to different stereoisomers at a very
late stage of the synthesis. The availability of novel Choi
core structures with different stereochemistries will be
useful in helping us to understand the stereochemical
influences of substituents to the inhibitor properties. The
biochemical information obtained from these isomers can
be used to establish structure-activity relationships. Our
future efforts will be the design and synthesis of deriva-
tives from these novel core structures and the study of
their antithrombotic activities. This will potentially lead
to the discovery of novel thrombin inhibitor drugs that
are useful in the treatment of blood coagulation.
Preparation of Compound 17. Diethyl acetamidoma-
lonate 16 (214 mg, 0.99 mmol) was dissolved in 5 mL of
anhydrous THF, then NaH (40 mg, 0.99 mmol, 60% oil
immersion) was added to the solution. The reaction mixture
was stirred at room temperature for half an hour. Then
compound 8 (160 mg, 0.33 mmol) in 5 mL of anhydrous THF
was injected and the reaction mixture was stirred at room
temperature for 1 h. TBAI (37 mg, 0.1 mmol) was added and
the reaction mixture was stirred at room temperature for
another hour before the reaction mixture was heated to reflux
for 2 days. The reaction mixture was cooled to 0 °C and 5 mL
of 0.5 N HCl was added. The organic layer was separated and
the aqueous layer was extracted with EtOAc (3 × 15 mL). The
combined organic layer was washed with brine and dried over
Na2SO4. Solvent was removed under reduced pressure and the
crude was purified by flash chromatography to give the product
as a white solid (110 mg, 54%). Mp 107-109 °C, [R]D +13.3 (c
1
0.7, CHCl3). H NMR (400 MHz, CDCl3) δ 7.37-7.24 (m, 10
H), 6.66 (s, 1H), 4.84 (ABq, 2H, J ) 9.8 Hz), 4.64 (d, 1H, J )
11.7 Hz), 4.55 (d, 1H, J ) 11.7 Hz), 4.36-4.31 (m, 1H), 4.25-
4.11 (m, 4H), 3.81-3.77 (m, 1H), 3.61-3.54 (m, 2H), 3.30 (dd,
1H, J ) 10.7, 9.8 Hz), 3.06-2.95 (m, 2H), 2.98 (s, 3H), 2.37
(dd, 1H, J ) 14.7, 9.8 Hz), 2.02 (s, 3H), 1.23-1.18 (m, 6H); 13
C
NMR (100 MHz, CDCl3) δ 169.3, 168.4, 167.3, 137.8, 137.7,
128.4, 128.3, 128.2, 127.9, 127.8, 127.7, 82.7, 80.1, 78.5, 75.3,
74.6, 73.3, 67.7 64.1, 62.7, 62.3, 39.2, 34.6, 23.0, 13.9, 13.7.
Preparation of Compound 22. Diethyl 2-[(tert-butyloxy-
carbonyl)amino]malonate (21) was prepared according to
literature procedure.34 Compound 21 (1.4 g, 5.1 mmol) was
dissolved in 15 mL of dry toluene under N2. NaH (204 mg, 5.1
mmol) was then added to the solution. The reaction mixture
was left stirring at room temperature for half an hour. Then
compound 8 (820 mg, 1.7 mmol) was added and the reaction
mixture was stirred at room temperature for an hour. TBAI
(188 mg, 0.51 mmol) was added and the reaction mixture was
heated to reflux for 17 h. The reaction mixture was cooled to
0 °C and 10 mL of 0.5 N HCl was added. The organic layer
was separated and the aqueous layer was extracted with
EtOAc (3 × 75 mL). The combined organic layer was washed
with brine and dried over Na2SO4. Solvent was removed under
reduced pressure and the crude was purified by flash chro-
matography to give the product as a viscous colorless oil (630
mg, 56%). [R]D +8.3 (c 0.93, CHCl3). 1H NMR (400 MHz, CDCl3)
mixture of conformers δ 7.42-7.20 (m, 10 H), 6.03 (br s, 0.3H),
5.87 (br s, 0.7H), 5.13-4.96 (m, 0.3H), 4.86 (br s, 1.4H), 4.70-
4.54 (m, 2.3H), 4.35-4.09 (complex, 5H), 3.78 (d, 1H, J ) 9.8
Hz), 3.68-3.52 (m, 2H), 3.39 (t, 1H, J ) 9.8 Hz), 3.08-2.78
(complex, 5H), 2.37 (dd, 1H, J ) 14.7, 10.7 Hz), 1.42 (s, 9H),
1.24-1.18 (complex, 6H); 13C NMR (100 MHz, CDCl3) δ 168.2,
167.7, 153.9, 137.8, 128.4, 128.3, 128.2, 127.9, 127.7, 127.4,
82.7, 81.8, 80.5, 80.3, 80.1, 79.0, 78.6, 75.4, 75.1, 74.4, 73.2,
73.0, 67.0, 64.0, 62.6, 62.1, 39.2, 38.6, 35.3, 35.0, 28.1, 27.8,
13.9, 13.7. HRMS calcd for C33H45NO12S + Na 702.2560, found
702.2568.
Experimental Section
1,5-Anhydro-2,3-di-O-benzyl-4,6-di-O-methylsulfonyl-
D-glucitol (15). Compound 14 (2 g, 5.8 mmol) was dissolved
in 10 mL of DCM and 8 mL of pyridine. Methanesulfonyl
chloride (1.8 mL, 23.3 mmol) was then added to the reaction
mixture, which was left stirring under anhydrous condition
until completion (usually 12 h). The reaction mixture was then
poured into a beaker containing 15 g of crushed ice and 15
mL of saturated aq NaHCO3 and was left stirring in an ice
bath for 1 h. The organic layer was extracted with ethyl acetate
three times and the combined organic layer was washed with
water and brine,and then dried over Na2SO4. Concentration
and purification by flash chromatography gave 15 as colorless
crystals (2.8 g, 97% yield). Mp 104-105 °C, [R]D +26.0 (c 1.0,
CHCl3). 1H NMR (500 MHz, CDCl3) δ 7.34-7.26 (m, 10H), 5.06
(d, 1H, J ) 11.1 Hz), 4.70 (d, 1H, J ) 11.1 Hz), 4.64 (d, 1H, J
) 11.5 Hz), 4.64 (d, 1H, J ) 11.5 Hz), 4.49-4.43 (m, 2H), 4.29
(dd, 1H, J ) 11.3, 5.26 Hz), 4.08-4.04 (m, 1H), 3.72-3.65 (m,
2H), 3.59 (ddd, 1H, J ) 9.8, 5.2, 2.3 Hz), 3.24-3.20 (m, 1H),
3.02 (s, 3H), 2.82 (s, 3H); 13C NMR (125 MHz, CDCl3) δ 137.5,
137.3, 128.6, 128.5, 128.1, 127.9, 127.8, 127.7, 82.4, 78.4, 77.1,
76.1, 75.3, 73.2, 67.8, 67.7, 38.5, 37.5; HRMS calcd for
C22H28O9S2 + Na 523.1072, found 523.1083.
1,5-Anhydro-2,3-di-O-benzyl-6-bromo-4-O-methylsulfo-
nyl-D-glucitol (8). The mixture of the dimesylate 15 (1.34 g,
2.7 mmol), NaBr (1.65 g, 16.0 mmol), TBAB (0.26 g, 0.80
mmol), and DMSO (10 mL) was stirred at 60 °C for 12 h;
usually by then the synthesis was completed. The reaction
mixture was cooled to room temperature then poured into 20
mL of water, the water phase was extracted with ethyl acetate
4 times, and the combined organic phase was dried over
sodium sulfate. Concentration and purification on a silica gel
column afford the pure product (1.26 g, 93% yield) as a white
Preparation of Compound 23. NaOH (1 N, 2 mL) in 10
mL of ethanol was added dropwise at room temperature to a
stirred solution of compound 22 (360 mg, 0.53 mmol). After
the reaction was complete (TLC analysis, usually 5 h), the
reaction mixture was acidified with 1 N HCl to pH 2-3 and
extracted with chloroform five times. The combined organic
(34) Schneider, H.; Sigmund, G.; Schricker, B.; Thirring, K.; Berner,
H. J. Org. Chem. 1993, 58, 683-689.
J. Org. Chem, Vol. 70, No. 22, 2005 8691

Nie and Wang
layer was dried over Na2SO4. Solvent was removed under
reduced pressure to afford crude product, which was subse-
quently decarboxylated by heating in 15 mL of toluene at 75
°C for 2 h. Solvent was removed under reduced pressure and
the crude was purified by flash chromatography. Compound
23 (322 mg, 86%) was obtained as viscous oil (a mixture of
inseparable diasteromers): 1H NMR (400 MHz, CDCl3) (the
NMR spectrum is complicated due to the presence of two
diasteromers) δ 7.36-7.19 (m, 10 H), 5.24-5.19 (m, 1H), 5.07
(d, 0.6H, J ) 11.7 Hz), 5.02 (d, 0.4H, J ) 11.7 Hz), 4.73-4.37
(complex, 4H), 4.25-3.91 (complex, 4H), 3.70-3.58 (m, 2H),
3.43 (dt, 0.5H, J ) 9.8, 2.0 Hz), 3.32 (t, 0.5H, J ) 9.8 Hz),
3.18-3.10 (m, 1H), 2.81 (s, 1.2H), 2.77 (s, 1.8H), 2.30 (dd, 0.5H
J ) 14.7, 6.8 Hz), 2.09 (m, 0.5H), 1.97-1.84 (m, 1H), 1.43
(singlet with shoulder, 9H), 1.24 (t, 1.2H, J ) 6.84 Hz), 1.23
(t, 1.8H, J ) 6.8 Hz); 13C NMR (100 MHz, CDCl3) δ 172.2,
171.9, 155.5, 154.9, 137.7, 137.6, 137.5, 137.4, 128.55, 128.47,
128.4, 128.1, 128.0, 127.9, 127.8, 127.7, 127.6, 82.6, 81.1, 80.9,
79.8, 78.9, 77.2, 75.6, 75.4, 75.2, 73.1, 67.8, 67.7, 61.4,
61.3, 38.67, 38.58, 34.7, 34.2, 28.2, 14.1, 14.0. HRMS calcd for
C30H41NO10S + Na 630.2349, found 630.2359.
Preparation of Compounds 24 and 25. Compound 23
(270 mg, 0.44 mmol), 12 mL of DCM, and 3 mL of TFA were
mixed at 0 °C. The reaction mixture was warmed to room
temperature and stirred for 1 h before the solvent was
evaporated under reduced pressure. Saturated NaHCO3 (15
mL) and 25 mL of EtOAc were added to the residue. The
aqueous layer was extracted with EtOAc five times. The
combined organic layer was dried over Na2SO4. Solvent was
removed under reduced pressure to afford crude product, which
was dissolved in 10 mL of toluene. TEA (70 µL, 0.50 mmol)
was added to the reaction mixture which was then heated to
reflux for 2 h. After this period of time, solvent was removed
under reduced pressure and the crude was purified by flash
chromatography to give pure diasteoromers 24 (80 mg, 44%)
and 25 (84 mg, 46%). Compound 24 was obtained as a colorless
syrup. [R]D +11.0 (c 0.7, CHCl3). 1H NMR (400 MHz, CDCl3) δ
7.39-7.24 (m, 10 H), 4.78 (q, 2H, J ) 2.0 Hz), 4.69 (q, 2H, J
) 11.7 Hz), 4.17 (q, 2H, J ) 6.84 Hz,), 4.01-3.91 (m, 4H), 3.65
(dd, 1H, J ) 8.8, 4.9 Hz), 3.60 (dd, 1H, J ) 4.9, 2.0 Hz), 3.13-
3.07 (m, 1H), 2.39 (br, 1H), 2.30 (dd, 1H, J ) 13.7, 8.8 Hz),
2.03 (ddd, 1H, J ) 13.7, 6.8, 4.9 Hz), 1.26(t, 3H, J ) 6.8 Hz);
13C NMR (100 MHz, CDCl3) δ 175.6, 138.6, 138.5, 128.3, 127.6,
127.55, 127.50, 80.3, 78.2, 74.6, 73.3, 71.7, 67.5, 61.0, 60.3, 56.9,
36.9, 14.2. HRMS calcd for C24H29NO5 + H 412.2124, found
412.2130.
Compound 25 was obtained as a colorless syrup. [R]D +94.8
(c 0.7, CHCl3). 1H NMR (400 MHz, CDCl3) δ 7.40-7.24 (m, 10
H), 4.84 (q, 2H, J ) 2.0 Hz), 4.65 (q, 2H, J ) 5.9 Hz), 4.16(q,
2H, J ) 6.8 Hz), 3.91-3.74 (m, 5H), 3.58 (dd, 1H, J ) 4.9, 2.0
Hz), 3.02 (t, 1H, J ) 10.7 Hz), 2.55 (br, 1H), 2.31 (dd, 1H, J )
13.7, 2.0 Hz,), 2.21 (ddd, 1H, J ) 13.7, 9.8, 3.9 Hz), 1.23 (t,
3H, J ) 6.8 Hz,); 13C NMR (100 MHz, CDCl3) δ 174.4, 138.6,
138.4, 128.4, 127.7, 127.6, 79.6, 77.4, 74.1, 73.7, 70.8, 67.2, 61.1,
58.21, 37.5, 14.2. HRMS calcd for C24H29NO5 + H 412.2124,
found 412.2133.
Acknowledgment. We are grateful for the financial
support from American Heart Association Award No.
0430285N and the start-up fund from the University of
New Orleans. We thank Kristopher Williams and Lousi
Dakessian for their assistance in purification of some
intermediates.
Note Added after ASAP Publication. References
24, 25, 26, and 31 were incorrectly cited in the text in
the version published ASAP September 27, 2005; the
corrected version was published ASAP September 29,
2005.
Supporting Information Available: The procedures
1
for preparing compounds 9-14, the H and 13C NMR spectra
of compounds 8-15, 17, and 22-25, and the 2D NOESY,
2D-COSY spectra for compounds 24 and 25. This material is
available free of charge via the Internet at http://pubs.acs.org.
JO0507901
8692 J. Org. Chem., Vol. 70, No. 22, 2005