Design and synthesis of dimeric heparinoid mimetics

Article abstract of DOI:10.1021/jo049092r

Synthetic oligosaccharide constructs exhibiting tailored and well-defined heparan sulfate (HS) like sequences offer the potential to modulate dynamic HS-dependent biomolecular recognition processes. We report an efficient strategy for the generation of HS

Full text of DOI:10.1021/jo049092r

Design and Synthesis of Dimeric Heparinoid Mimetics
Shyam M. Rele,* Suri S. Iyer, Subramanian Baskaran, and Elliot L. Chaikof*
Departments of Surgery and Biomedical Engineering, Emory University, Atlanta, Georgia 30322,
and School of Chemical and Biomolecular Engineering, Georgia Institute of Technology,
Atlanta, Georgia 30322
echaiko@emory.edu; smrele@emory.edu
Received May 30, 2004
Synthetic oligosaccharide constructs exhibiting tailored and well-defined heparan sulfate (HS) like
sequences offer the potential to modulate dynamic HS-dependent biomolecular recognition processes.
We report an efficient strategy for the generation of HS-like fragments [GlcA-â-(1,4)-GlcNAc] and
related dimerized (gemini) disaccharides (4a and 4b) via n-pentenyl glycoside formation. When a
convergent synthetic approach was utilized, construction of target molecules was achieved through
a combination of chemoselective protection/deprotection protocols, imidate and n-pentenyl glyco-
sylations, and functional group manipulations followed by ozonolysis and reductive amination. For
example, glycosylation of a 2-azido glycoside (25) with a trichloroacetimidate glucuronic acid donor
(13), using a catalytic amount of TMSOTf, furnished heparin-like disaccharides (28a and 28b) that
were equipped with an n-pentenyl tether at the anomeric end. In turn, heparinoid-like gemini
disaccharides (4a and 4b) were produced by selective transformation of the olefinic unit in the
n-pentenyl glycoside to the four-carbon aldehyde followed by reductive amination with ethylene-
diamine. The described synthetic approach provides access to structural variants of small heparinoid
oligomers as versatile building blocks for generating novel HS mimetic pharmacotherapeutics,
diagnostic reagents, and biomaterials.
Introduction
preparation of simpler HS oligosaccharides and their
analogues remains an area of active investigation. In this
regard, a variety of synthetic methods directed at the
preparation of H/HS fragments have been reported in the
literature, including, for example, the total synthesis of
ATIII-binding oligosaccharides.3-5 In addition, approaches
involving the modular synthesis of larger heparin oli-
gosaccharides,6 heparin-related fragments varying in
length and composition,7 and the synthesis of heparin
disaccharide subunits have also been disclosed.8
Heparin/heparan sulfate like (H/HS, 1)1,2 glycosami-
noglycans (GAGs), found in the extracellular matrix, are
complex linear biopolymers; more specifically, they are
structurally heterogeneous, polyanionic polysaccharides
that sequester and modulate the activity of a variety of
soluble, membrane-bound, and matrix proteins. As such,
H/HS oligosaccharides play a key role in regulating the
biological activity of a variety of critical protein modula-
tors of coagulation and inflammatory cascades, cell
growth, migration, and proliferation, and bacterial and
viral recognition. Characteristically, H/HS consists of
repeating disaccharide units comprised of an alternating
uronic acid [D-glucuronic acid (D-GlcA) or the epimeric
L-iduronic acid (L-IdoA)] linked to a D-glucosamine
(GlcNAc) through a (1,4) glycosidic bond.1,2 Unique
structural modifications of these disaccharide building
blocks add to the structural heterogeneity and molecular
complexity associated with H/HS (Figure 1).
Although most structural and biochemical studies
have been focused on sulfated H/HS oligosaccharide
variants,4d,9-11 nonsulfated HS-derived di- and trisaccha-
(3) (a) Jacquient, J.-C.; Petitou, M.; Duchaussoy, P.; Lederman, I.;
Choay, J.; Torri, G.; Sinay, P. Carbohydr. Res. 1984, 130, 221-241.
(b) Sinay, P.; Jacquient, J.-C.; Petitou, M.; Duchaussoy, P.; Lederman,
I.; Choay, J.; Torri, G. Carbohydr. Res. 1984, 132, C5-C9.
(4) (a) Dreef-Tromp, C. M.; Basten, J. E. M.; Broekhoven, M. A.;
van Dinther, T. G.; Petitou, M. C.; van Boeckel, A. A. Bioorg. Med.
Chem. Lett. 1998, 8, 2081-2086. (b) Kovensky, J.; Duchaussoy, P.;
Bono, F.; Salmivirta, M.; Sizun, P.; Herbert, J.-M.; Petitou, M.; Sinay,
P. Bioorg. Med. Chem. 1999, 7, 1567-1580. (c) Kuberan, B.; Lech, M.
Z.; Beeler, D. L.; Wu, Z. L. L.; Rosenberg, R. D. Nat. Biotechnol. 2003,
21, 1343-1346. (d) Tabeur, C.; Mallet, J.-M.; Bono, F.; Herbert, J.-M.;
Petitou, M.; Sinay, P. Bioorg. Med. Chem. Lett. 1999, 8, 2003-2012.
(e) de Paz, J.-L.; Ojeda, R.; Erichardt, N.; Ma´rtin-Lomas, M. Eur. J.
Org. Chem. 2003, 68, 3308-3324.
Given the potential significance of therapies based on
the control of GAG-protein interactions, the chemical
* Address correspondence to Elliot L. Chaikof, M.D., Ph.D., or
Shyam M. Rele, Ph.D., 1639 Pierce Drive, Woodruff Memorial Research
Building, Room 5105, Emory University, Atlanta, GA 30322. Phone:
(404) 727-8413. Fax: (404) 727-3660.
(1) Conrad, H. E. Heparin Binding Proteins; Academic Press: New
York, 1998.
(5) (a) Petitou, M.; Duchaussoy, P.; Driguez, P.-A.; Jaurand, J.;
He´rault, J.-P.; Lormeau, J.-C.; Van Boeckel, C. A. A.; Herbert, J. M.
Angew. Chem., Int. Ed. 1998, 37, 3009-3014. (b) Petitou, M.; He´rault,
J.-P.; Bernat, A.; Driguez, P.-A.; Duchaussoy, P.; Lormeau, J.-C.;
Herbert, J.-M. Nature 1999, 398, 417-422.
(2) For recent reviews, see: (a) Capila, I.; Linhardt, R. Angew.
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Lindahl, U. Adv. Carbohydr. Chem. Biochem. 2001, 57, 159-206.
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Palmacci, E. R.; Seeberger, P. H. Chem.sEur. J. 2003, 9, 140-169.
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10.1021/jo049092r CCC: $27.50 © 2004 American Chemical Society
Published on Web 11/17/2004
J. Org. Chem. 2004, 69, 9159-9170
9159

Rele et al.
FIGURE 1. Structure of natural heparin/heparan sulfate (H/HS), 1.
ride motifs, which are abundant in H/HS, may also
exhibit significant biological activity. For example, Ornitz
and co-workers have shown that nonsulfated disaccha-
rides (2) and trisaccharides can stimulate or inhibit
(fibroblast growth factor) FGF-1 and FGF-2.12 Although
the potency of these oligosaccharides appeared to be less
pronounced than that of H/HS, both binding of FGF to
FGF receptors and FGF-mediated mitogenic activity were
significantly enhanced. Similarly, a diverse library of
nonsulfated glucuronic acid (monosaccharide) derivatives
(3) has been recently generated, several members of
which competitively inhibited heparin binding to FGF-
2.13 Consequently, these results support the notion that
tailored and precisely defined nonsulfated HS-related
disaccharide modules or their homologues could serve as
a significant tool for studying receptor protein binding
in GAG-mediated events (Figure 2).
FIGURE 2. Glucuronic acid derivatives capable of inhibiting
the bioactivity of FGF-2.
To this end, we have recently postulated that head-
to-head tethering of GlcA-â-(1,4)-GlcNAc by a flexible
molecular spacer might provide a rational strategy for
the synthesis of HS-like glycomimetic probes for studying
GAG-dependent protein-binding events. In response to
this goal, a synthetic route was designed for the insertion
of an n-pentenyl group at the anomeric terminus of a
glycosidic acceptor, in order to facilitate direct access to
higher ordered glycoconjugates.14,15 As an initial assess-
ment of this approach, gemini disaccharides were pro-
duced via a reductive amination strategy with ethylene-
diamine, after conversion of the terminal olefinic bond
to an aldehyde (Scheme 1 and Figure 3).
(7) (a) Poletti, L.; Lay, L. Eur. J. Org. Chem. 2003, 2999-3024 and
references therein. (b) Petitou, M.; Driguez, P.-A.; Duchaussoy, P.;
Herault, J.-P.; Lormeau, J.-C.; Herbert, J.-M. Bioorg. Med. Chem. Lett.
1999, 9, 1161-1166. (c) Koshida, S.; Suda, Y.; Sobel, M.; Kusumoto,
S. Tetrahedron Lett. 2001, 42, 1289-1292. (d) Suda, Y.; Koshida, S.;
Arano, A.; Morichika, T.; Fukui, Y.; Kusumoto, S.; Sobel, M. Polym.
Prepr. (Am. Chem. Soc., Div. Plym. Chem.) 2000, 41, 1624-1625. (e)
Tromp, C. M.; Basten, J. E. M.; Broekhoven, M. A.; van Dinther, T.
G.; Petitou, M.; van Boeckel, C. A. A. Bioorg. Med. Chem. Lett. 1998,
8, 2081-2086. (f) Buijsman, R. C.; Basten, J. E.; Tromp, C. M.; Marel,
G. A.; van Boeckel, C. A. A.; Boom, J. H. Bioorg. Med. Chem. Lett.
1999, 7, 1881-1890. (g) Codee, J. D. C.; Marel, G. A.; van Boeckel, C.
A. A.; Boom, J. H. Eur. J. Org. Chem. 2002, 3954-3965.
(8) (a) Gavard, O.; Hersant, Y.; Alais, J.; Duverger, V.; Dilhas, A.;
Bascou, A.; Bonnaffe, D. Eur. J. Org. Chem. 2003, 18, 3603-3620. (b)
Yu, H. N.; Furukawa, J.; Ikeda, T.; Wong, C.-H. Org. Lett. 2004, 6,
723-726. (c) Haller, M.; Boons, G. J. Eur. J. Org. Chem. 2002, 2033-
2038. (d) Ferla, B. L.; Lay, L.; Guerrini, M.; Poletti, L.; Panza, L.; Russo,
G. Tetrahedron 1999, 55, 9867-9880.
(9) (a) Walker, A.; Turnbull, J. E.; Gallagher, J. T. J. Biol. Chem.
1994, 269, 931-935. (b) Ornitz, D. M.; Yayon, A.; Flanagan, J. G.;
Svahn, C. M.; Levi, E.; Leder, P. Mol. Cell. Biol. 1992, 12, 240-247.
(c) Wang, H.; Toda, T.; Kim, Y. S.; Capila, I.; Hileman, R. Y.; Bernfield,
M.; Linhardt, R. J. Biochem. Biophys. Res. Commun. 1997, 235, 369-
373. (d) Ishihara, M.; Tyrell, D. J.; Stauber, G. B.; Brown, S.; Cousens,
L. S.; Stack, R. J. J. Biol. Chem. 1993, 268, 4675-4683. (e) Avizer, D.;
Levy, E.; Safran, M.; Svahn, C.; Buddecke, E.; Schmidt, A.; David, G.;
Vlodavsky, I.; Yayon, A. J. Biol. Chem. 1994, 269, 114-121.
(10) (a) Ornitz, D. M.; Yayon, A.; Flanagan, J. G.; Svahn, C. M.;
Levi, E.; Leder, P. Mol. Cell. Biol. 1992, 12, 240-247. (b) Yayon, A.;
Klagsbrun, M.; Esko, J. D.; Leder, P.; Ornitz, D. M. Cell 1991, 64, 841-
848. (c) Rapraeger, A. C.; Krufka, A.; Olwin, B. B. Science 1991, 252,
1705-1708. (d) Ornitz, D. M.; Xu, J.; Colvin, J. S.; McEwen, D. G.;
MacArthur, C. A.; Coulier, F.; Gao, G.; Goldfarb, M. J. Biol. Chem.
1996, 271, 15292-15297.
Results and Discussion
The success of any strategy that targets the synthesis
of a HS mimetic depends on the preparation of a suitable
glycosyl acceptor and donor and the use of appropriate
protecting groups, as well as careful consideration of the
order of glycosidic bond formation, facile removal of
protecting groups, and incorporation of amino or sulfate
functionalites where appropriate. On the basis of a
retrosynthetic analysis of 4 (Figure 4), a strategy was
devised to obtain the desired imidate donor (B) and the
n-pentenyl glycosyl acceptor (C) on a multigram scale.
The synthesis of heparinoids is often complicated by
the inherent difficulty associated with the formation of
a â-(1,4) glycosidic linkage between uronic acid and
hexosamine residues because of the low nucleophilicity
and steric deactivation of C-4 hydroxyl groups.16 Conse-
quently, the synthesis of a HS mimetic requires the
proper installation and protection of a 2-deoxy-2-amino
functionality. In developing an efficient route to GluA-
â-(1,4)-GlcNAc, we postulated that the ready availability
(11) de Paz, J.-L.; Angulo, J.; Lassaletta, J.-M.; Nieto, P. M.;
Redondo-Horcajo, M.; Lozano, R. M.; Gime´nez-Gallego, G.; Ma´rtin-
Lomas, M. ChemBioChem 2001, 2, 673-685 and references therein.
(12) (a) Ornitz, D. M.; Herr, B. A.; Nilsson, M.; Westman, J.; Svahn,
C. M.; Waksman, G. Science 1995, 268, 432-436. (b) Westman, J.;
Nilsson, M.; Ornitz, D. M.; Svahn, C. M. J. Carbohydr. Chem. 1995,
14, 95-113.
(13) Murphy, P. V.; Pitt, N.; O’Brien, A.; Enright, P. M.; Dunne, A.;
Wilson, S. J.; Duane, R. M.; O’Boyle, K. M. Bioorg. Med. Chem. Lett.
2002, 12, 3287-3290.
(14) Buskas, T.; Soderbegh, E.; Konradsson, P.; Fraser-Reid, B. J.
Org. Chem. 2000, 65, 958-963.
(15) (a) Allen, R. J.; Danishefsky, S. J. J. Am. Chem. Soc. 1999, 121,
10875-10882 and references therein. (b) Nishimura, I.; Matsuoka, K.
Macromolecules 1994, 27, 157-163. (c) Nishimura, I.; Furuike, T.;
Matsuoka, K.; Maruyama, K.; Nagata, K.; Kurita, K.; Nishi, N.; Tokura,
S. Macromolecules 1994, 27, 4876-4880.
(16) Crich, D.; Dudkin, V. J. Am. Chem. Soc. 2001, 123, 6819-6825
and references therein.
9160 J. Org. Chem., Vol. 69, No. 26, 2004

Synthesis of Dimeric Heparinoid Mimetics
FIGURE 3. Gemini oligosaccharides of H/HS-derived disaccharides.
SCHEME 1. Strategy for Functionalization of n-Pentenyl Glycoside To Obtain Tethered Head-to-Head
H/HS Mimetic Homodimers
and low cost of N-acetyl â-D-glucosamine (5) would
provide an ideal starting point for the preparation of the
nucleophilic glycosyl acceptor. Furthermore, we specu-
lated that the presence of a 2-N-acyl group on the hex-
osamine unit would allow us to incorporate the 2-aceta-
mido functionality into GluA-â-(1,4)-GlcNAc during gly-
cosylation, thereby eliminating the need for a potentially
cumbersome N deprotection and acetylation sequence at
the conclusion of dimer synthesis. Thus, our initial efforts
involved the substitution of the anomeric center in
N-acetyl D-glucosamine (5) using n-pentenyl alcohol, with
camphorsulfonic acid (CSA) as a catalyst, to give 6
(Scheme 2). The R/â (3:1) anomers were separated using
column chromatography (9:1 CHCl3/MeOH).17
The C-4 and C-6 hydroxyl groups of the â isomer were
then capped with benzylidene acetal to yield the 4,6-
benzylidene alkenyl glycoside (7). The 3-OH position was
then protected using a standard benzylation procedure
to yield the 3-O-benzyl derivative (8). Deprotection of the
benzylidene group using a 2:1 TFA/H2O mixture yielded
the 4,6-diol (9), which was regioselectively acetylated at
the C-6 hydroxyl position using acetyl chloride/pyridine,
furnishing the N-acetyl D-glucosamine glycosyl acceptor
(10).
The highly reactive glucuronic acid imidate donor was
synthesized in 50% yield in two steps from commercially
available acetobromo-R-D-glucuronic acid methyl ester
(11; Scheme 3). The methyl ester 11 was converted to
its corresponding free hemiacetal 12 using CdCO3/H2O,
which was subsequently converted to the R-imidate
glycosyl donor 13 using trichloroacetonitrile and 1,8-
diazabicyclo[5.4.0]undec-7-ene. Unfortunately, glycosy-
lation of the n-pentenyl-terminated N-acetyl D-glu-
cosamine acceptor 10 with either glycosyl donor 11 or
13 was unsuccessful (Scheme 4 and Table 1).
As an alternative approach, a 2-azido-2-deoxy glycoside
acceptor was synthesized (Scheme 5). Specifically, 2-azido-
2-deoxy-3,4,6-tri-O-acetyl-D-glucopyranosyl acetate (16)
was initially synthesized from TfN3 and D-glucosamine
hydrochloride (15).18 Although the synthesis of an n-
pentenyl glycoside from the mannose analogue, 2-azido-
2-deoxy-3,4,6-tri-O-acetyl-D-mannosepyranosyl acetate,19
using 4-penten-1-ol and BF3‚Et2O was successful, similar
attempts using compound 16 in the presence of BF3‚Et2O,
SnCl4, or TMSOTf as the promoter did not succeed. Thus,
selective hydrolysis of the anomeric acetate to the hy-
droxyl group 17 was performed using hydrazine acetate.
The hydroxyl group was subsequently converted to the
trichloroacetimidate derivative 18 in 74% yield. The
(17) Grande, D.; Baskaran, S.; Baskaran, C.; Gnanou, Y.; Chaikof,
E. L. Macromolecules 2000, 33, 1123-1125.
(18) Vasella, A.; Witzig, C.; Chiara, J.-L.; Ma´rtin-Lomas, M. Helv.
Chim. Acta 1991, 74, 2073-2077.
(19) Li, Q.; Li, H.; Cai, M. S.; Li, Z. J.; Zhou, R. L. Tetrahedron:
Asymmetry 1999, 10, 2675-2683.
FIGURE 4. Retrosynthetic strategy for the synthesis of bis-
carbohydrates possessing â-(1,4)-linked H/HS disaccharides
(P1-P5 ) protecting groups).
J. Org. Chem, Vol. 69, No. 26, 2004 9161

Rele et al.
SCHEME 2. Synthesis of an N-Acetyl D-Glucosamine Glycosyl Acceptor 10a
a Reaction conditions: (a) CSA, 1-penten-1-ol, DMF, 75 °C, yield ) 50% [R and â isomers were separated by silica gel column
chromatography, R/â (3:1)]; (b) CSA, dry THF, C6H5CH(OMe)2, reflux, 8 h, yield ) 75%; (c) NaH, C6H5CH2Br, dry THF, reflux, 8 h, 75%;
(d) 2:1 TFA/H2O, CH2Cl2, 0 °C, 5 h, yield: 85%; (e) AcCl, pyridine, 25 °C, 15 h, yield ) 80%.
SCHEME 3. Synthesis of Imidate Donor 13a
a Reaction conditions: (a) CdCO3, H2O (2 equiv), CH3CN, 70 °C, 4 h, yield ) 75%; (b) CCl3CN, 1,2-dichloroethane, 1,8-DBU, -10 to 0
°C, 1 h, yield ) 60%.
SCHEME 4. Attempted Strategy for the Synthesis of Heparin Disaccharide 14
TABLE 1. Glycosylation Reaction between n-Pentenyl
Protection of C-4 and C-6 hydroxyl groups as a ben-
zylidene acetal provided 20 in 75% yield. The R (20a) and
â (20b) anomers were separated using column chroma-
tography with 40:60 EtOAc/hexane as the eluant. To
explore the influence of 3-position neighboring group
participation on the rate of the glycosylation reaction, the
3-OH group in 20 was functionalized using different
protecting groups. Specifically, benzylation of R/â isomers
20a/20b and acetylation of the â derivative 20b at the
3-OH position using NaH/benzyl bromide and acetyl
chloride/pyridine afforded the 3-O-benzyl derivative 21a/
21b and the 3-O-acetyl derivative 22b, respectively.
Deprotection of the benzylidene acetal afforded the 4,6-
dihydroxylated products 23a/23b and 24b. Regioselective
acetylation of the 6-OH position using acetyl chloride/
pyridine gave the orthogonally protected acceptor 25a/
25b and 3,6-diacetylated glycoside 26b. Further reactions
were carried out independently for each anomer (25a,
25b, and 26b).
Glycosylation of the n-pentenyl glycoside acceptors 25a/
25b and 26b was carried out separately for both R and â
isomers using the trichloroacetimidate donor 13 in the
presence of acid promoters. The preparation of the
disaccharide is outlined in Scheme 6, and the results of
coupling reactions are summarized in Table 2. Attempts
to glycosylate the sugar alcohol 25b with imidate 13 in
the presence of TBDSOTf or TMSOTf and molecular
Glycoside Acceptor 10 and Glycosyl Donors
entry reactantsa activatorb reaction conditionsc yield (%)d
1
2
3
10 + 11
10 + 13
10 + 13
AgOTf
-15 to +25 °C,
4.0 h
-15 to +25 °C,
5.0 h
-15 to +25 °C,
3.5 h
No reaction
e
BF3‚OEt2
g
g
TMSOTf e,f
a Ratio of acceptor/donor ) 1:1.8-2.0 equiv. b Equivalents of
activator: AgOTf (0.14 equiv), TMSOTf (0.1 equiv, 0.22 M solution
in CH2Cl2), BF3‚OEt2 (1.5 equiv). c All reactions were carried out
in anhydrous CH2Cl2/4 Å molecular sieves. d Reactions were
monitored at regular intervals by TLC by removing small aliquots
of the reaction mixture with a syringe. e No disaccharide formation
was observed even after carrying out the glycosidation reaction
with an excess amount of activators BF3‚OEt2 (2.0 and 3.0 equiv)
and TMSOTf (0.2 and 0.3 equiv). f Reaction when carried out in
the absence of molecular sieves did not have any appreciable effect
on the disaccharide product formation. g A trace amount (less than
10%) of ortho ester was isolated. NMR of the intermediate ortho
ester showed characteristic resonance signals at δ 1.74 (s, 3H, CH3)
and 5.92 (d, 1H, H-1′, J ) 4.4 Hz) corresponding to the methyl
and H-1′ anomeric protons, respectively.
n-pentenyl group was then introduced at the anomeric
position using a catalytic amount of TMSOTf at 0 °C.
Despite variations in temperature, promoter, and sol-
vents, the selectivity ratio for R/â isomers (2:3) did not
substantially change. Removal of the ester protecting
group by Zemplen conditions20 produced the n-pentenyl
glycoside triol 19 in 80% yield.
(20) Zemplen, G.; Pacsu, E. Ber. Dtsch. Chem. Ges. 1929, 62, 1613-
1614.
9162 J. Org. Chem., Vol. 69, No. 26, 2004

Synthesis of Dimeric Heparinoid Mimetics
SCHEME 5. Synthesis of a D-Glucosamine Glycosyl Acceptora
a Reagents and conditions: (a) (1) TfN3, MeOH, DMAP, 25 °C, 18 h, (2) Ac2O, pyridine, 0 °C, 10 h, yield ) 75%; (b) H2NNH2‚AcOH,
DMF, 0 to 25 °C, 45 min, yield ) 70%; (c) anhydrous K2CO3, CCl3CN, CH2Cl2, 25 °C, 48 h, yield ) 74%; (d) (1) TMSOTf, 4-penten-1-ol,
dry CH2Cl2, 0 °C, 1 h, (2) MeONa, MeOH, 0 °C to 25 °C, 6 h, yield: 80% over two steps; (e) CSA, THF, C6H5CH(OMe)2, reflux, 6 h, yield
) 75%; (f) NaH, C6H5CH2Br, dry THF, reflux, 10 h, yield ) 82%; (g) AcCl, pyridine, 25 °C, 15 h, yield: 86%; (h) 2:1 TFA/H2O, CH2Cl2,
0 °C, 5 h; (i) AcCl, pyridine, 25 °C, 12 h.
SCHEME 6. Generation of a Protected GlcA-â-(1,4)-GlcN Disaccharide
sieves produced an undesired ortho ester intermediate
(27b) as the major product. The exclusive formation of
ortho ester was confirmed by 1H NMR resonance signals
at δ 5.96 (doublet, J ) 4.2 Hz) and 1.78 (singlet),
corresponding to H-1 and methyl protons along with
three acetyl peaks. We believe that the molecular sieves,
which are alumino silicates and are inherently basic in
nature, hindered acid-catalyzed 1,2-trans glycosidic bond
formation. Indeed, in the absence of molecular sieves,
boron trifluoride etherate mediated glycosylation of 25b
with imidate 13 afforded the thermodynamically favored
â-(1,4) disaccharide without a trace of ortho ester, albeit
in 30% yield after 14 h. Significantly, TMSOTf-catalyzed
conditions provided the most efficient route for coupling
of the acceptor 25b with glycosyl donor 13, producing the
fully protected â-(1,4)-linked disaccharide 28b in moder-
ate (60%) yield. Of note, the ortho ester 27b could be
converted to disaccharide 28b using an excess amount
of acid catalyst in the absence of molecular sieves.
Similarly, we also observed that replacing the 3-O benzyl
protecting group by a 3-O acetyl functionality had a
retarding influence on â-(1,4) glycosidic bond formation
(Table 2, compare entries 5 and 7). Likely, the electron-
withdrawing nature of the 3-O benzyl group enhanced
the nucleophilicity of the glycosyl acceptor at the 4
position.
Comparing the glycosylation reaction pattern between
the imidate donor 13 and either N-acetyl 10 or azido 25
acceptors revealed that the 4-hydroxy of a 2-azido-2-
deoxyglucose derivative was a more reactive nucleophile
J. Org. Chem, Vol. 69, No. 26, 2004 9163

Rele et al.
TABLE 2. Influence of Activators on the Glycosylation Reaction Involving 2-Azido Acceptors and
Tricholoroacetimidate Donor 13
product (% yield)b,c
entry
reactantsa
activator
temperature (°C), time (h)
ortho ester disaccharide
1
2
3
4
5
6
7
8
25b + 13
25b + 13
25b + 13
25b + 13
25b + 13
27b
TBDSOTf + 4 Å mol sieves
TMSOTf + 4 Å mol sieves
TMSOTf + 4 Å mol sieves
BF3‚OEt2
0, 5.0
No reaction
27b (40)
27b (65)
0, 1.5
0-25, 6.5
0-25, 14
0-25, 4.5
0-25, 4.0
0-25, 24
0-25, 4.5
28b (30)
28b (60)
28b (58)
29b (25)e
28a (60)
TMSOTf
TMSOTfd
26b + 13
25a + 13
TMSOTf
TMSOTf
a Ratio of acceptor/donor ) 1:1.5-2.0 equiv. b Isolated yields. c Reactions were monitored at regular intervals to confirm the conversion
of the intermediate 1,2-ortho ester derivative to the 1,2-trans disaccharide using 1H NMR. d Reaction was carried out in the absence of
molecular sieves. e Reaction is not clean; a number of byproducts are observed.
FIGURE 5. Structures of glucosamine acceptors in decreasing order of reactivity.
than that of the corresponding N-acetyl derivative (Fig-
ure 5). We suspect that the absence of a hydrogen-
bonding moiety in the azido precursor, such as the
acetamido group, enhances the nucleophilicity of the C-4
hydroxy group. Indeed, recent experimental evidence by
Crich and Dudkin16 involving â-mannosylation reactions
with various glucosamine acceptors suggests that the
lack of reactivity of the N-acetyl derivative in comparison
to azido (N3) and pthalimido (NPth) groups is primarily
a result of amide NH-induced intramolecular hydrogen
bonding with the free 4-hydroxy group.
Further synthetic manipulation of the n-pentenyl
spacer group was necessary to obtain the desired final
target molecule 4 (Scheme 7). The presence of unsatura-
tion in 28b precluded debenzylation by hydrogenation.
As a result, deacetylation and saponification of 28b using
3 M NaOH in a MeOH/water mixture yielded the tetraol
glucuronic acid derivative in which the terminal olefinic
unit was transformed to an aldehyde using reductive
ozonolysis, yielding 30b. We had anticipated that hydro-
genolysis of 30b by 10% Pd-C/H2 would reduce the azide
to a free amine and debenzylate the C-3 position while
preserving the chemical integrity of the aldehyde group.
Unfortunately, several attempts to carry out the hydro-
genation reaction of 30b afforded a complex mixture of
products.21-23
In response to this limitation, we converted the azido
group to an acetamido functionality. To this end, initial
attempts to reduce the azido group to the acetamido
functionality using standard Staudinger reaction24 condi-
tions (PPh3/CH2Cl2/Ac2O) led to very low yields. Alter-
natively, conversion of the azido functionality to the
acetamido (-NHAc) group was performed using thioace-
tic acid,25 which furnished 32b in appreciable yield (55%),
with 1H NMR spectroscopy demonstrating the NHCOCH3
peak at δ 1.83. Subsequent saponification of 32b, using
3 M NaOH in 9:1 MeOH/water, afforded the disaccharide
33b in 86% yield. The crude product was purified using
Sephadex LH-20 with MeOH as the eluant, and the
structure was confirmed by 13C NMR, which showed the
presence of two carbonyl groups (CdO) at δ 176 and 175,
and by FAB-MS (m/z ) 562.24 for M+ + Li).
The presence of an n-pentenyl group in compound 33b
provides a versatile handle for diverse conjugation reac-
tions by transformation of the terminal olefin to either
an aldehyde, carboxylic acid, ester, thioether, thioester,
or hydroxyl group.14 In the current reaction scheme,
ozonolysis of 33b afforded the terminal aldehyde glyco-
side intermediate 34b. The formation of 34b was con-
firmed by FAB-MS (m/z ) 570.23 for M+ + 2Li - H)
and the appearance of an aldehyde signal at δ 9.72 and
203 in 1H and 13C NMR spectra, respectively, along with
the disappearance of the olefinic double bond peaks at δ
5.85 (δc 132.1) and 5.1-4.9 (δc 118.1). Subsequent de-
benzylation of the four-carbon aldehyde 34b using 10%
Pd-C/H2 afforded 35b, which was confirmed by NMR
and FAB-MS (m/z ) 474.25 for M+ + Li).
(21) Rele, S. M.; Iyer, S. S.; Chaikof, E. L. Unpublished results.
Reactions were carried out using different stoichiometries (2, 5, and
7.5 equiv) of Pd-C reagents including Pearlmans catalyst (20%); 10%
Pd-C still afforded complex mixtures. In some cases, according to the
NMR spectrum, the benzyl group appeared to stay intact despite using
an excess of reagent. Use of anhydrous FeCl3 to selectively debenzylate
the 3-O benzyl group in the presence of the n-pentenyl group, as
reported by Fraser-Reid (see Tetrahedron Lett. 1996, 37, 5477-5478),
again resulted in a complex mixture. Also, reaction of the saponified
product obtained from 28b with NiCl2‚6H2O/NaBH4/MeOH converted
the N3 to NH2 and reduced the terminal olefinic double bond without
knocking off the benzyl group.
(22) Removal of benzyl groups was found to be problematic during
fluorescence labeling of carbohydrates. For reference, see: France, R.
R.; Cumpstey, I.; Butters, T. D.; Fairbanks, A. J.; Wormald, M. R.
Tetrahedron: Asymmetry 2000, 11, 4985-4994.
(23) Reduction of the disaccharide 28b under similar hydrogenation
conditions followed by N acetylation using AcCl/pyridine showed the
disappearance of benzyl peaks (δ 7.4-7.2) and terminal olefinic double
bonds (δ 5.84 and 5.02) in the 1H NMR spectrum. In addition, a new
Reductive amination of 35b (Scheme 8) with ethylene-
diamine produced the homodimerized â-(1,4)-linked H/HS
set of resonance signals appeared in the reduced product 31 at δ 0.85
and 2.01, the integration of which corresponded to terminal methyl
protons and acetamido protons, indicating the conversion of the azido
group to an amino functionality. Thus, the outcome of the hydrogena-
tion reaction of 30b might be subject to the collective influence of
electronic, steric, and coordination effects of the azido, benzyl, and
terminal aldehyde groups.
(24) (a) Staudinger, H.; Meyer, J. Helv. Chim. Acta 1919, 2, 635-
646. (b) Gololobov, Y. G.; Kasukhin, L. F. Tetrahedron 1992, 48, 1353-
1406.
(25) Jacquinet, J. C. Carbohydr. Res. 1990, 199, 153-181.
9164 J. Org. Chem., Vol. 69, No. 26, 2004

Synthesis of Dimeric Heparinoid Mimetics
SCHEME 7a
a Reagents and conditions: (a) 3 M NaOH, 9:1 MeOH/H2O, 25 °C, 2 h, yield: 82%; (b) O3 (-78 °C), Me2S, -78 to +25 °C, 24 h, yield
) 85%; (c) Pd-C/H2, MeOH, 25 °C, 24 h; (d) thioacetic acid, 25 °C, 24 h, yield: 55%; (e) 3 M NaOH, 9:1 MeOH/H2O, 25 °C, 2 h, yield: 86%;
(f) O3 (-78 °C), Me2S, -78 to +25 °C, 24 h, yield: 90%.
SCHEME 8a
a Reagents and conditions: (a) NH2(CH2)2NH2 (0.5 equiv), 3 h, NaCNBH3 (4 equiv), 25 °C, 24 h, yield ) 67%.
neoglycoconjugate 4b (yield 67%). The crude product was
purified using a Sephadex G-10 gel filtration column
using water as the eluant. The structure was confirmed
by heteronuclear multiple-quantum coherence (HMQC)
13C NMR, which showed the presence of four anomeric
carbons, and by MALDI-TOF analysis (m/z ) 963.45 for
M+ + H; Figure 6).
The biological response of receptor proteins depends,
in part, on their interaction with particular structural
motifs of GAGs. Computational modeling procedures26
have suggested that the intrinsic binding affinity of
oligosaccharide mimetics to target proteins depends on
the distance and appropriate orientation of the sugar
epitopes, as well as on the carbohydrate conformational
flexibility.27 Moreover, small and subtle differences in
stereochemistry, conformation, and functionality can
have a profound influence on the biological activity of
glycoconjugates. To this end, while the â-linked (1,2-
trans) neoglycoconjugate 4b resembles the naturally
occurring structure of GlcA-â-(1,4)-GlcNAc within GAG
(26) (a) Kolb, H.; Ernst, B. Chem.sEur. J. 1997, 3, 1571-1578. (b)
Hanessian, S.; Reddy, G. V.; Huynh, H.; Pan, J.; Pedatella, S.; Ernst,
B.; Kolb, H. C. Bioorg. Med. Chem. Lett. 1997, 7, 2729-2734.
J. Org. Chem, Vol. 69, No. 26, 2004 9165

Rele et al.
FIGURE 6. HMQC spectrum highlighting the anomeric region indicating the presence of four anomeric carbons and the MALDI-
TOF spectrum of 4b.
residue. Subsequent flash column chromatography of the crude
extract (40:60 EtOAc/hexane, 1% Et3N) furnished an off-white
compound 13 (60%). 1H NMR (CDCl3, 400 MHz) δ: 8.71 (s,
1H, NH), 6.58 (d, 1H, J ) 3.6 Hz), 5.62 (t, 1H, J ) 10 Hz),
5.26 (t, 1H, J ) 10 Hz), 5.11 (dd, 1H, J ) 3.2 Hz), 4.43 (d, 1H,
J ) 10.2 Hz), 3.74 (s, 3H, COOMe), 2.05 (s, 6H, OAc), 2.02 (s,
3H). HRMS-FAB: [M+] calcd for C15H18O10NCl3, 478.9956;
found, 478.9968.
chains, R-linked (1,2-cis) disaccharides may well exhibit
different binding affinities. Thus, the glycosidation pro-
cedure was repeated with the R acceptor (25a) to obtain
the R-linked disaccharide module 28a, which was further
elaborated sequentially to obtain 32a, 33a, 34a, and 35a.
Reductive amination of 35a afforded the homodimeric R
product 4a. In summary, beginning with acceptors 25a/
25b, gemini homodimers 4a/4b possessing head-to-head
orientation were obtained in six steps with an overall
yield of 7%.
Synthesis of 2-Azido-2-deoxy-3,4,6-tri-O-acetyl-r,â-D-
glucopyranosyl Acetate (16). To a round-bottomed flask
containing NaN3 (0.43 mol, 28 g) and equipped with an argon
balloon were added water (80 mL) and CH2Cl2 (75 mL). The
emulsion was stirred and cooled to 0 °C followed by dropwise
addition of dry Tf2O (0.088 mol, 15 mL) via syringe. The
reaction mixture was stirred at 0 °C for 2 h and at 25 °C for
15 min. The organic layer was separated, and the aqueous
layer was extracted with CH2Cl2 (2 × 25 mL). The combined
organic layers were neutralized with a cold saturated solution
of NaHCO3, washed with water, dried over MgSO4, and
filtered. This stock solution contains ca. 0.1 mol of TfN3.
In a separate three-necked flask equipped with an argon
inlet containing D-glucosamine hydrochloride (15; 0.042 mol,
9 g) dissolved in methanol (250 mL), NaOMe (80 mL of a 0.5
M solution in MeOH) was added and stirred for 30 min. DMAP
(5 g) was added to it, and the reaction mixture was diluted
with an additional 100 mL of MeOH. The freshly prepared
solution of TfN3 was added to this reaction mixture via a
cannula under positive pressure of argon and left to stir at
room temperature for 18 h. The solvent was removed in vacuo
to give a sticky, yellow paste. The residue obtained was then
acetylated at 0-4 °C for 10 h using anhydrous pyridine (250
mL) and dry acetic anhydride (120 mL). Upon completion of
the reaction, the reaction mixture was coevaporated with
toluene (3 × 50 mL) to give a light-brown, sticky residue. Flash
chromatography (60:40 EtOAc/hexane) of the crude reaction
mixture using silica gel gave the desired product 16. Yield:
Conclusions
In summary, the synthesis of n-pentenyl bearing GlcA-
â-(1,4)-GlcNAc disaccharides and related bis-heparinoid
dimers is described. Investigations of carbohydrate-
protein binding interactions are ongoing and will be
reported in due course. We believe the synthetic approach
described herein provides a useful foundation for the
rational design and synthesis of small-molecule glycomics
capable of modulating HS-dependent protein-binding
events.
Experimental Section
Synthesis of Trichloroacetimidate Donor Methyl (2,3,4-
Tri-O-acetyl-r-D-glucopyranosyl trichloacetimidate) Glu-
coronate (13). Acetobromo-R-D-glucuronic acid methyl ester
(11; 0.025 mol, 10 g) was added to a flame-dried, three-necked
flask containing CdCO3 (0.026 mol, 4.5 g), CH3CN (150 mL),
and 0.8-1.0 mL of degassed H2O. The reaction mixture was
stirred and heated to 70 °C for a period of 4 h under argon,
filtered through Celite, washed with 50 mL of CH3CN, and
concentrated in vacuo. Purification of the crude residue by
flash chromatography (50:50 EtOAc/hexane) afforded the
hydrolyzed product (white solid), which was subsequently
dissolved in 200 mL of dichloroethane; Cl3CCN (10 equiv, 0.142
mol, 13.6 mL) was then added to it under argon. After cooling
to 0 °C, 1,8-diazabicyclo[5.4.0]undec-7-ene (1,8-DBU; 0.28
equiv, 3.96 mmol, 595 µL) was added in a dropwise manner.
The reaction mixture was allowed to stir for 1 h, and the
mixture was concentrated to afford a sticky, dark-brown
1
11.5 g (75%). H NMR (CDCl3) δ: 6.30 (d, 1H, H-1R, J ) 3.6
Hz), 5.55 (d, 1H, H-1â, J ) 8.7 Hz), 5.43 (t, 1H, J ) 8.0 Hz),
5.18-5.04 (m, 3H), 4.34-4.27 (m, 2H), 4.11-4.05 (m, 4H),
3.68-3.64 (m, 2H), 2.19 (s, 6H), 2.11 (s, 3H), 2.10 (s, 3H), 2.08
(s, 6H), 2.05 (s, 3H), 2.02 (s, 3H).
Synthesis of 2-Azido-2-deoxy-3,4,6-tri-O-acetyl-r,â-D-
glucopyranoside (17). To an ice-cooled solution of 16 (0.0241
mol, 9.0 g) dissolved in 100 mL of dry DMF was added solid
hydrazine acetate (2.5 equiv, 0.06 mol, 5.5 g) under an argon
atmosphere. The ice bath was then removed, and the reaction
mixture was allowed to stir for 45 min. Upon completion (TLC),
the reaction mixture was quenched using ethyl acetate (50 mL)
and water (50 mL). The organic layer was then separated, and
(27) (a) Alibes, R.; Bundle, D. R. J. Org. Chem. 1998, 63, 6288-
6301 and references therein. (b) Wacowich-Sgarbi, S.; Bundle, D. R.
J. Org. Chem. 1998, 64, 9080-9089 and references therein. (c)
Kretzschmar, G. Tetrahedron 1998, 54, 3765-3780.
9166 J. Org. Chem., Vol. 69, No. 26, 2004

Synthesis of Dimeric Heparinoid Mimetics
the aqueous layer was extracted with EtOAc (3 × 25 mL).
Finally, the combined organic layers were washed with
saturated NaHCO3 and brine and dried over MgSO4. Concen-
tration of the solvent in vacuo gave a light-yellow gel, which
on flash column chromatography (80:20 EtOAc/ hexane)
furnished 17 as a white solid. Yield: 5.9 g (70%). 1H NMR
(CDCl3) δ: 5.35 (t, 2H, J ) 10.4 Hz, J ) 9.6 Hz), 5.23 (d, 1H,
H-1R, J ) 3.2 Hz), 4.88 (m, 2H), 4.58 (d, 1H, H-1â, J ) 8.0
Hz), 4.12 (m, 2H), 3.94 (m, 2H), 3.80 (br s, 2H, OH), 3.35-
3.28 (m, 2H), 3.20 (s, 2H), 2.09 (s, 6H), 2.08 (s, 6H), 2.08 (s,
6H). 13C NMR (CDCl3) δ: 171.3, 171.2, 170.5, 170.4, 170.2,
170.0, 96.1, 91.9, 72.5, 71.7, 70.5, 68.7, 68.5, 67.2, 64.8, 62.2,
61.4, 20.7, 20.6.
Synthesis of 2-Azido-2-deoxy-3,4,6-tri-O-acetyl-r,â-D-
glucopyranosyl Trichloroacetimidate (18). To the reaction
flask containing compound 17 (0.024 mol, 8 g) dissolved in dry
CH2Cl2 (35 mL) was added anhydrous K2CO3 (0.06 mol, 8.33
g). This was followed by the dropwise addition of Cl3CCN
(0.144 mol, 14.5 mL), after which the reaction mixture was
stirred for 48 h under an argon atmosphere. Upon completion
(TLC), the reaction mixture was cooled to 0 °C, diluted with
25 mL of CH2Cl2, and quenched with 20 mL of ice-cold water.
The organic layer was separated, and the aqueous layer was
extracted with CH2Cl2 (2 × 20 mL). The combined organic
layers were washed with brine, dried over Na2SO4, and
concentrated to yield a light-yellow solid. Purification of the
crude mixture by column chromatography (60:40 EtOAc/
hexane) gave compound 18 as an off-white solid. Yield: 8.5 g
(74%). 1H NMR (CDCl3) δ: 8.81 (s, 1H, NH), 8.79 (s, 1H, NH),
6.43 (d, 1H, H-1R, J ) 3.6 Hz), 5.68 (d, 1H, H-1â, J ) 7.8 Hz),
5.55 (dd, 2H, J ) 9.6 Hz, J ) 9.6 Hz), 5.11 (t, 1H, J ) 10.0
Hz), 5.09 (m, 1H), 4.23 (m, 4H), 4.05 (m, 2H), 3.76 (dd, 2H, J
) 3.6 Hz), 2.05 (s, 3H), 2.04 (s, 3H), 2.01 (s, 3H), 2.0 (s, 3H),
1.99 (s, 3H), 1.96 (s, 3H). 13C NMR (CDCl3) δ: 170.7, 170.6,
170.0, 169.8, 169.7, 160.6, 96.5, 94.1, 90.6, 72.8, 72.7, 70.8, 70.2,
68.1, 63.4, 61.5, 60.7, 20.8, 20.7.
pound 19 (0.011 mol, 3.2 g) dissolved in anhydrous THF (25
mL) was added 200 mg of (+)-CSA as a catalyst, followed by
the dropwise addition of benzaldehyde dimethyl acetal (0.0197
mol, 2.95 mL). The reaction mixture was refluxed under an
argon atmosphere for 6 h (monitored by the complete disap-
pearance of the starting material), cooled to room temperature,
diluted with 50 mL of EtOAc, and quenched with 10 mL of a
saturated solution of NaHCO3. The organic layer was sepa-
rated, and the aqueous layer was extracted with 3 × 25 mL of
EtOAc. The organic layers were combined and washed with
brine, dried over Na2SO4, and concentrated to yield a light-
yellow solid. Purification of the crude product by flash chro-
matography (5:95 EtOAc/hexane to 30:70 EtOAc/hexane) gave
20 as a mixture of R and â isomers (yield: 3.17 g, 75%). The
anomeric isomers 20a (R) and 20b (â) were separated at this
stage by silica gel column chromatography using a 60:40
hexane/ethyl acetate mixture as the eluant. Pent-4-enyl-2-
azido-2-deoxy-4,6-O-benzylidene-r,D-glucopyranoside
1
(20a). [R]25D: -37.31 (c ) 0.98, CHCl3). H NMR (CDCl3) δ:
7.50-7.45 (m, 2H, C6H5), 7.39-7.27 (m, 3H, C6H5), 5.81 (m,
1H, CHdCH2), 5.59 (s, 1H, CHPh), 5.08-4.99 (m, 2H, CHd
CH2), 4.89 (d, 1H, H-1R, J ) 3.6 Hz), 4.28-4.21 (m, 2H), 3.87-
3.83 (m, 1H), 3.75-3.70 (m, 2H), 3.53-3.47 (m, 2H), 3.24 (dd,
1H, J ) 4 Hz, J ) 3.6 Hz), 2.80 (br s, 1H, OH), 2.19-2.16 (m,
2H), 1.77-1.73 (m, 2H). 13C NMR (CDCl3) δ: 138.1, 137.1,
129.6, 128.6, 126.5, 115.3, 102.3, 98.6, 82.1, 69.0, 68.8, 68.1,
63.2, 62.6, 30.4, 28.5. HRMS-FAB: [M+] calcd for C18H23O5N3,
361.3918; found, 361.3924. Pent-4-enyl-2-azido-2-deoxy-4,6-
O-benzylidene-â-D-glucopyranoside (20b). [R]25D: +33.6 (c
) 1.5, CHCl3). 1H NMR (CDCl3) δ: 7.48-7.45 (m, 2H, Ph),
7.38-7.26 (m, 3H, Ph), 5.83-5.79 (m, 1H, CHdCH2), 5.56 (s,
1H, CHPh), 5.06-4.97 (m, 2H, CHdCH2), 4.33 (d, 1H, H-1â,
J ) 7.8 Hz), 4.29 (m, 1H), 3.89 (m, 1H), 3.73 (t, 1H, J ) 10.0
Hz), 3.56-3.48 (m, 3H), 3.45-3.38 (m, 2H), 2.65 (br s, 1H, OH),
2.17-2.15 (m, 2H), 1.76-1.73 (m, 2H). 13C NMR (CDCl3) δ:
138.1, 137.1, 129.6, 128.6, 126.5, 115.4, 102.7, 102.1, 80.8, 72.0,
70.1, 68.7, 66.7, 66.3, 30.22, 28.94. HRMS-FAB: [M+] calcd
for C18H23O5N3, 361.3918; found, 361.3926.
Synthesis of 3-O-Benzyl-Protected Derivative Pent-
4-enyl-2-azido-2-deoxy-3-O-benzyl-4,6-O-benzylidene-r/â-
D-glucopyranoside (21a/21b). To a suspension of NaH (12
equiv, 49.8 mmol, 1.2 g) in anhydrous THF under argon was
added compound 20a/20b (4.15 mmol, 1.5 g) via a cannula,
and the resulting solution was stirred for 10 min. Benzyl
bromide (4 equiv, 16.62 mmol, 2.0 mL) was added to the
mixture, and the reaction was refluxed under an argon
atmosphere for 10 h. Upon completion (TLC), the reaction
mixture was diluted with EtOAc and quenched slowly with
an ice-cold solution of saturated NaHCO3, and the organic
layer was separated from the aqueous layer. The aqueous
fraction was further extracted with small portions of EtOAc
(three times); the combined organic layers were collected,
subsequently washed with water and brine, and dried over
Na2SO4. Removal of the solvent in vacuo followed by column
chromatography (silica gel, 5:95 EtOAc/hexane to 25:75 EtOAc/
hexane) gave the solid product 21a/21b (R/â) (yield: 1.63 g,
82%). 21a. 1H NMR (CDCl3) δ: 7.5-7.25 (m, 10H, Ph), 5.85-
5.78 (m, 1H, CHdCH2), 5.57 (s, 1H), 5.12-4.95 (m, 2H, CHd
CH2), 4.88 (d, 1H, H-1R, J ) 3.6 Hz), 4.82 (d, 2H, J ) 10.8
Hz), 4.35-4.25 (m, 1H), 4.15-4.07 (m, 1H), 3.97-3.86 (m, 1H),
3.82-3.88 (m, 3H), 3.52-3.48 (m, 1H), 3.37-3.34 (m, 1H),
2.13-2.08 (m, 2H), 1.80 (m, 2H). HRMS-FAB: [M+] calcd for
C25H29O5N3, 451.5128; found, 451.5142. 21b. 1H NMR (CDCl3)
δ: 7.5-7.25 (m, 10H, Ph), 5.85-5.78 (m, 1H, CHdCH2), 5.57
(s, 1H), 5.12-4.75 (m, 4H), 4.37 (d, 1H, H-1â, J ) 8.0 Hz),
4.36 (m, 1H), 4.15-4.07 (m, 1H), 3.97-3.86 (m, 1H), 3.83-
3.64 (m, 2H), 3.62-3.40 (m, 2H), 3.38-3.28 (m, 1H), 2.14-
2.08 (m, 2H), 1.80 (m, 2H). HRMS-FAB: [M+] calcd for
C25H29O5N3, 451.5130; found, 451.5142.
Synthesis of Pent-4-enyl-2-azido-2-deoxy-r,â-D-glu-
copyranoside (19). To an oven-dried flask containing 18
(0.016 mol, 7.5 g) dissolved in anhydrous CH2Cl2 (25 mL) was
added 4-penten-1-ol (0.0195 mol, 2.0 mL) via syringe. The
reaction mixture was initially stirred for 10 min at room
temperature and then cooled to 0 °C under an argon atmo-
sphere. A freshly prepared solution of TMSOTf (1.6 mmol, 7.4
mL of a 0.22 M solution in anhydrous CH2Cl2) was added
dropwise through a syringe to the reaction mixture and stirred
at 0 °C. Upon completion (TLC), the reaction mixture was
quenched with Et3N, filtered through a pad of Celite, and
concentrated under reduced pressure to give a semicrystalline
material. Flash chromatography (50:50 EtOAc/hexane) af-
forded a light-yellow, semicrystalline material, which was
directly used for the next step. (The compound obtained was
contaminated with trace amounts of 4-penten-1-ol.) The above
mixture was dissolved in anhydrous MeOH followed by the
addition of NaOMe (10 mL of a 0.5 M solution in MeOH, 0.05
mol) at 0 °C under an argon atmosphere. The reaction mixture
was stirred at room temperature for 6 h (monitored by TLC)
and quenched with a Dowex 50W X 8-200 (H+) ion-exchange
resin at 0 °C. The resin was filtered and, upon evaporation of
the solvent, gave a light-brown syrup, which on flash chro-
matography (50:50 EtOAc/hexane to 10:90 MeOH/CHCl3)
furnished 19 as a light-yellow compound. Yield over two
1
steps: 3.56 g (80%). H NMR (CDCl3) δ: 5.81 (m, 1H, CHd
CH2), 5.02-4.92 (m, 2H, CHdCH2), 4.87 (d, 0.3H, H-1R, J )
3.2 Hz), 4.6 (br s, 3H, OH), 4.34 (d, 0.7H, H-1â, J ) 7.6 Hz),
3.98-3.74 (m, 3H), 3.68-3.31 (m, 4H), 3.28-3.20 (m, 1H),
2.11-2.05 (m, 2H, CH2CHdCH2), 1.80-1.65 (m, 2H, CH2). 13
C
NMR (CDCl3) δ: 138.7, 115.1, 102.2, 98.1, 75.2, 74.6, 71.4, 70.1,
70.05, 69.5, 67.7, 66.0, 62.6, 61.4, 61.1, 30.3, 30.1, 28.8, 28.6.
HRMS-FAB: [M+] calcd for C11H19O5N3 , 273.1228; found,
273.1238.
Synthesis of the 3-O-Acetyl-Protected Derivative Pent-
4-enyl-2-azido-2-deoxy-3-O-acetyl-4,6-O-benzylidene-â-D-
glucopyranoside (22b). The compound 20b (4.15 mmol, 1.5
Synthesis of Pent-4-enyl-2-azido-2-deoxy-4,6-O-ben-
zylidene-r,â-D-glucopyranoside (20). To a solution of com-
J. Org. Chem, Vol. 69, No. 26, 2004 9167

Rele et al.
g) was dissolved in dry pyridine and stirred at 0 °C. Using a
syringe, acetyl chloride (1.5 equiv, 6.23 mmol, 0.43 mL) was
added dropwise to the reaction mixture and stirred overnight
under an argon atmosphere. Upon completion (TLC), the crude
reaction mixture was concentrated, diluted with CHCl3, and
extracted with water. After repeated extraction of the aqueous
phase with CHCl3 (two times), the organic fractions were
pooled together and concentrated to afford a dark-brown
residue. Flash chromatography of the crude product (30:70
EtOAc/hexane) gave the desired 3-O-acetylated product 22b
(â). Yield: 1.43 g (86%). 1H NMR (CDCl3) δ: 7.6-7.25 (m, 5H,
Ph), 5.82-5.75 (m, 1H, CHdCH2), 5.65-5.59 (m, 1H), 5.50 (s,
1H), 5.12-4.94 (m, 2H, CHdCH2), 4.42 (d, 1H, H-1â, J ) 7.6
Hz), 4.32-4.24 (m, 1H), 4.01-3.92 (m, 1H), 3.82-3.72 (m, 2H),
3.65-3.47 (m, 2H), 3.18 (dd, 1H, J ) 3.6 Hz), 2.14 (m, 2H),
2.03 (s, 3H), 1.77 (m, 2H). 13C NMR (CDCl3) δ: 170.1, 137.9,
137.2, 129.3, 128.4, 126.4, 115.5, 101.9, 99.2, 79.7, 69.1, 69.0,
68.2, 62.9, 61.8, 30.4, 28.7, 21.1. HRMS-FAB: [M+] calcd for
C20H25O6N3, 403.1764; found; 403.1752.
enyl-2-azido-2-deoxy-3-O-benzyl-6-O-acetyl-â-D-glucopy-
ranoside (25b). 1H NMR (CDCl3, 400 MHz) δ: 7.42-7.32 (m,
5H, ArH), 5.86-5.74 (m, 1H, dCH), 5.06-4.98 (m, 2H, dCH2),
4.90 (d, 1H, J ) 11.6 Hz), 4.75 (d, 1H, J ) 11.6 Hz), 4.44 (dd,
1H, J ) 4.4 Hz), 4.27 (d, 1H, J ) 8.0 Hz), 4.22 (d, 1H, J ) 2.0
Hz), 3.93-3.90 (m, 1H), 3.57-3.52 (m, 1H), 3.48-3.42 (m, 1H),
3.38-3.34 (m, 2H), 3.22 (m, 1H), 2.6 (s, 1H, -OH), 2.17-2.14
(m, 2H), 2.08 (s, 3H), 1.78-1.71 (m, 2H). 13C NMR (CDCl3,
400 MHz) δ: 171.8, 138.2, 138.1, 128.8, 128.3, 128.2, 115.3,
102.4, 82.4, 75.4, 73.8, 70.2, 69.8, 66.1, 63.4, 30.2, 28.9, 21.1.
HRMS-FAB: [M+ + H] calcd for C20H27O6N3, 406.1968; found,
406.1960. Pent-4-enyl-2-azido-2-deoxy-3,6-di-O-acetyl-â-
D-glucopyranoside (26b). 1H NMR (CDCl3, 400 MHz) δ:
5.82-5.79 (m, 1H, CHdCH2), 5.28-5.23 (m, 1H), 5.01-4.96
(m, 2H, CHdCH2), 4.48 (dd, 1H, J ) 4.5 Hz), 4.40 (d, 1H, H-1â,
J ) 8.0 Hz), 4.28 (dd, 1H, J ) 2.1 Hz), 3.88-3.82 (m, 1H),
3.75-3.69 (m, 1H), 3.55-3.48 (m, 2H), 3.22 (dd, 1H, J ) 3.6
Hz), 2.99 (dd, 1H, J ) 4.5 Hz), 2.12 (m, 2H), 2.11 (s, 3H), 2.03
(s, 3H), 1.8-1.76 (m, 2H). 13C NMR (CDCl3, 400 MHz) δ: 172.3,
170.9, 137.9, 115.5, 102.9, 73.4, 70.4, 69.7, 68.9, 63.1, 60.9, 30.3,
28.6, 21.1, 21.0. HRMS-FAB: [M+ + H] calcd for C15H23O7N3,
358.1554; found, 358.1572.
Synthesis of â-(1,4)-Linked Disaccharide Units. (A)
Formation of Ortho Ester 27b. The n-pentenylazido glyco-
side acceptor 25b (2.47 mmol, 1.0 g) dissolved in dry CH2Cl2
was added to a flask already containing 4 Å molecular sieves
and was stirred at 0 °C under an argon atmosphere. To this
solution was added dropwise a freshly prepared solution of
TMSOTf (0.13 mmol of a 0.22 M solution in dry CH2Cl2, 0.561
mL) over a period of 5 min via syringe. The imidate donor 13
(1.8 equiv, 4.5 mmol, 2.2 g), dissolved in dry CH2Cl2, was then
added to the reaction mixture via a cannula, and the reaction
mixture was stirred at 0 °C for 30 min, then allowed to warm
to room temperature, and stirred for 4.0 h. Upon completion
(TLC), the reaction mixture was quenched with N,N-diisopro-
pylamine, filtered over Celite, concentrated, and subjected to
column chromatography (30:70 EtOAc/hexane) to give the
intermediate ortho ester derivative 27b as an off-white solid.
Yield: 1.16 g (65%). 1H NMR (CDCl3, 300 MHz) δ: 7.42-7.28
(m, 5H, ArH), 5.96 (d, 1H, H-1′, J ) 4.2 Hz), 5.88-5.74 (m,
1H, dCH), 5.09-4.91 (m, 2H, dCH2), 4.86 (d, 1H, J ) 10.2
Hz), 4.76 (d, 1H, J ) 10.2 Hz), 4.39-4.33 (m, 2H), 4.23 (d, 1H,
H-1, J ) 7.2 Hz), 4.19-4.13 (m, 1H), 3.92-3.87 (m, 1H), 3.75
(s, 3H, COOMe), 3.65-3.52 (m, 2H), 3.39-3.29 (m, 3H), 2.10
(m, 2H, CH2), 2.09 (s, 3H, COCH3), 2.08 (s, 3H, COCH3), 1.97
(s, 3H, COCH3), 1.78 (s, 3H, ortho ester CH3), 1.77-1.72 (m,
2H, CH2). HRMS-FAB: [M+] calcd for C33H43O15N3,721.2748;
found, 721.2728.
General Procedure for Selective Acetylation of the
6-OH Position (25a/25b and 26b). To an ice-cooled solution
of the 3-O benzyl derivative 21a/21b (3.61 mmol, 1.63 g) or
the 3-O acetyl derivative 22b (3.54 mmol, 1.43 g) in anhydrous
CH2Cl2 was added dropwise a 2.5:1 TFA/water mixture. The
mixture was stirred at room temperature for 5 h, monitored
by TLC, and upon disappearance of the starting compound
(TLC) was cooled to 0 °C followed by quenching with a
saturated NaHCO3 solution. The organic layer was separated
from the aqueous layer that was further extracted with CH2-
Cl2 (two times). The combined organic fractions were washed
with water and brine and dried over Na2SO4. Evaporation of
the solvent gave a pale-yellow gel, which was then subjected
to column chromatography to yield the debenzylidenated
product 23a/23b (yield: 1.02 g, 80%) or 24b (yield: 0.93 g,
1
83%). 23a. H NMR (CDCl3, 400 MHz) δ: 7.38-7.30 (m, 5H,
Ph), 5.82-5.76 (m, 1H, CHdCH2), 5.08-4.97 (m, 3H), 4.95 (d,
1H, H-1R, J ) 3.6 Hz), 4.84-4.71 (m, 1H), 3.94-3.73 (m, 3H),
3.61-3.54 (m, 2H), 3.36-3.19 (m, 3H), 2.12-2.06 (m, 2H),
1.82-1.74 (m, 2H). 23b. 1H NMR (CDCl3, 400 MHz) δ: 7.38-
7.32 (m, 5H, Ph), 5.82-5.74 (m, 1H, CHdCH2), 5.08-4.95 (m,
3H), 4.71 (d, 1H, J ) 11.7 Hz), 4.34 (d, 1H, H-1â, J ) 7.8 Hz),
3.92-3.73 (m, 3H), 3.61-3.54 (m, 2H), 3.35-3.17 (m, 3H),
2.10-2.06 (m, 2H), 1.86-1.74 (m, 2H). 24b. 1H NMR (CDCl3,
400 MHz) δ: 5.82-5.79 (m, 1H, CHdCH2), 5.33-5.27 (m, 1H),
5.07-4.97 (m, 2H, CHdCH2), 4.38 (d, 1H, H-1â, J ) 8.0 Hz),
3.86-3.84 (m, 2H), 3.75-3.70 (m, 3H), 3.51-3.46 (m, 1H), 3.23
(dd, 1H, J ) 3.6 Hz), 2.16-2.12 (m, 2H), 2.03 (s, 3H), 1.82-
1.78 (m, 2H). 13C NMR (CDCl3) δ: 171.2, 137.9, 115.5, 98.1,
74.1, 71.8, 70.3, 68.1, 62.2, 61.0, 30.4, 28.7, 21.1.
(B) Synthesis of 3-O-Benzyl-Protected â-(1,4)-Linked
n-Pentenyl Disaccharide (28a/28b). The acceptor 25a/25b
(2.47 mmol, 1.0 g) was dissolved in dry CH2Cl2 and stirred at
0 °C under an argon atmosphere. To this solution was added
dropwise a freshly prepared solution of TMSOTf (0.05 equiv,
of 0.22 M TMSOTf solution in dry CH2Cl2, 0.13 mmol, 0.51
mL) over a period of 5 min via syringe. The imidate donor 13
(1.5 equiv, 3.71 mmol, 1.78 g), dissolved in dry CH2Cl2, was
then added to the reaction mixture via a cannula, and the
reaction mixture was warmed to room temperature over a
period of 30 min and stirred for an additional 4 h. Upon
completion (TLC), the reaction mixture was quenched with
N,N-diisopropylamine, concentrated, and subjected to column
chromatography (30:70 EtOAc/hexane) to give the desired
disaccharide 28a/28b as an off-white solid (yield: 1.07 g, 60%).
The deprotected product 23a/23b (2.91 mmol, 1.02 g), or 24b
(2.95 mmol, 0.93 g), obtained above was dissolved in 25 mL of
dry pyridine and cooled to 0 °C. Dropwise addition of acetyl
chloride (1.1 equiv) to the above solution resulted in the
immediate formation of a white precipitate. The reaction
mixture was allowed to stir for 12 h at room temperature,
quenched with MeOH, and concentrated to give light-brown
syrup. The crude product was extracted with water and CH2-
Cl2 (two times), followed by subsequent washings with water
and brine, and finally dried over Na2SO4. Purification of the
crude mixture using column chromatography (30:60 EtOAc/
hexane) gave the selective 6-O acetylated product 25a/25b
(yield: 1.0 g, 85%), or 26b (yield: 0.63 g, 60%). Pent-4-enyl-
2-azido-2-deoxy-3-O-benzyl-6-O-acetyl-r-D-glucopyrano-
1
1
side (25a). H NMR (CDCl3, 400 MHz) δ: 7.42-7.28 (m, 5H,
28a. H NMR (CDCl3, 400 MHz) δ: 7.35-7.24 (m, 5H, ArH),
ArH), 5.85-5.74 (m, 1H, dCH), 5.06-4.97 (m, 2H, dCH2),
4.89-4.81 (m, 3H), 4.37 (dd, 1H, J ) 4.8 Hz), 4.22 (d, 1H, J )
12 Hz), 3.85-3.73 (m, 2H), 3.71-3.64 (m, 1H), 3.47-3.44 (m,
2H), 3.23-3.20 (m, 2H), 2.14 (m, 2H), 2.14 (s, 3H, COCH3),
1.78-1.68 (m, 2H, CH2). 13C NMR (CDCl3, 400 MHz) δ: 171.8,
138.2, 138.1, 128.8, 128.3, 128.2, 115.3, 98.2, 79.6, 75.36, 70.9,
5.84-5.78 (m, 1H, dCH), 5.17-5.15 (m, 2H, dCH2), 5.03-4.90
(m, 3H), 4.82 (d, 1H, H-1, J ) 3.6 Hz), 4.75 (d, 1H, J ) 11.6
Hz), 4.73 (d, 1H, H-1′, J ) 8.0 Hz), 4.42 (dd, 1H, J ) 2.4 Hz),
4.11 (dd, 1H, J ) 4.4 Hz, J ) 4.8 Hz), 3.92 (dd, 1H, J ) 8.4
Hz, J ) 8.8 Hz), 3.85-3.81 (m, 2H), 3.77-3.66 (m, 3H), 3.48
(s, 3H, COOMe), 3.47-3.45 (m, 1H), 3.25 (dd, 1H, J ) 3.6 Hz,
J ) 3.6 Hz), 2.15-2.08 (m, 2H, CH2), 2.09 (s, 3H, COCH3),
2.05 (s, 3H, COCH3), 1.99 (s, 3H, COCH3), 1.98 (s, 3H, COCH3),
70.1, 67.9, 63.4, 62.9, 30.4, 28.7, 21.1. HRMS-FAB: [M+
+
H] calcd for C20H27O6N3, 406.1968; found, 406.1958. Pent-4-
9168 J. Org. Chem., Vol. 69, No. 26, 2004

Synthesis of Dimeric Heparinoid Mimetics
1.78-1.71 (m, 2H, CH2). 13C NMR (CDCl3, 400 MHz) δ: 170.6,
170.3, 169.5, 168.8, 166.8, 138.6, 137.9, 128.5, 127.6, 127.4,
115.4, 101.1, 97.8, 79.1, 78.1, 74.9, 72.8, 72.3, 71.9, 69.4, 68.6,
68.1, 63.1, 62.3, 52.9, 30.3, 28.6, 21.0, 20.7, 20.6. HRMS-
FAB: [M+ + Li] calcd for C33H43O15N3, 728.2854; found,
3.58 (t, 2H, J ) 9.2 Hz, J ) 9.6 Hz), 3.43-3.35 (m, 3H), 3.25-
3.23 (m, 1H), 1.68-1.58 (m, 4H). HRMS-FAB: [M+ + Li] calcd
for C23H31O12N3, 548.2068; found, 548.2053.
Synthesis of 32a/32b. In a flame-dried flask, compound
28a/28b (1.48 mmol, 1.07 g) was dissolved in 5 mL of thioacetic
acid (CH3COSH). The reaction mixture was then allowed to
stir for 24 h under an argon atmosphere. Upon completion of
the reaction, the excess thioacetic acid was removed in vacuo,
and the reaction mixture was subjected to column chromatog-
raphy (100:0 CHCl3/CH3OH to 2.5:95 CHCl3/CH3OH) to give
the N-acetylated product 32a/32b as a pale-yellow crystalline
material (yield: 605 mg, 55%). 32a. 1H NMR (CDCl3, 400 MHz)
δ: 7.28-7.21 (m, 5H, ArH), 5.62-5.42 (m, 1H, dCH), 5.18-
5.11 (m, 3H), 5.02-4.94 (m, 4H), 4.67-4.65 (m, 2H), 4.54 (d,
1H, J ) 12.4 Hz), 4.42 (d, 1H, J ) 11.6 Hz), 4.12-4.05 (m,
2H), 3.88 (d, 1H, J ) 9.2 Hz), 3.75-3.72 (m, 2H), 3.59-3.56
(m, 2H), 3.50 (s, 3H, COOMe), 3.32-3.17 (m, 1H), 2.07 (s, 3H,
COCH3), 2.04 (m, 2H, CH2), 2.03 (s, 3H, COCH3), 1.97 (s, 3H,
COCH3), 1.96 (s, 3H, COCH3), 1.72 (s, 3H, NHCOCH3), 1.78-
1.62 (m, 2H, CH2). 13C NMR (CDCl3, 400 MHz) δ: 170.7, 170.2,
170.0, 169.6, 169.3, 167.0, 139.1, 138.0, 128.6, 127.9, 127.7,
115.3, 101.2, 97.4, 79.6, 78.1, 74.8, 72.6, 72.4, 71.9, 69.4, 68.8,
67.7, 62.2, 52.9, 52.5, 30.4, 28.6, 23.5, 21.1, 20.7, 20.6. HRMS-
FAB: [M+ + Li] calcd for C35H47O16N, 744.3055; found,
1
728.2858. 28b. H NMR (CDCl3, 400 MHz) δ: 7.38-7.28 (m,
5H, ArH), 5.84-5.75 (m, 1H, dCH), 5.16-5.12 (m, 2H, dCH2),
5.03-4.91 (m, 4H), 4.83 (d, 1H, J ) 11.2 Hz), 4.73 (d, 1H, H-1′,
J ) 8.0 Hz), 4.42 (dd, 1H, J ) 2.4 Hz), 4.23 (d, 1H, H-1, J )
7.8 Hz), 4.07 (dd, 1H, J ) 5.2 Hz), 3.88-3.84 (s, 1H), 3.78 (d,
1H, J ) 9.2 Hz), 3.73-3.65 (m, 1H), 3.52 (m, 1H), 3.50 (s, 3H,
COOMe), 3.48-3.35 (m, 3H), 2.12-2.06 (m, 2H, CH2), 2.03 (s,
3H, COCH3), 2.02 (s, 3H, COCH3), 1.97 (s, 3H, COCH3), 1.96
(s, 3H, COCH3), 1.72-1.60 (m, 2H, CH2). 13C NMR (CDCl3,
400 MHz) δ: 170.7, 170.3, 170.1, 168.8, 166.4, 138.5, 137.2,
128.5, 127.8, 127.3, 115.1, 102.2, 101.6, 81.6, 79.8, 75.1, 72.8,
72.6, 72.1, 70.0, 69.7, 66.2, 62.1, 52.4, 30.2, 29.6, 20.8, 20.4.
HRMS-FAB: [M+ + Li] calcd for C33H43O15N3, 728.2854;
found, 728.2824.
(C) Synthesis of 3-O-Acetyl-Protected â-(1,4)-Linked
n-Pentenyl Disaccharide (29b). This procedure is the same
as that above. Stoichiometric ratio: 26b (1.76 mmol, 0.63 g),
13 (2 equiv, 3.52 mmol, 1.68 g), TMSOTf (0.05 equiv of a 0.22
M solution, 0.088 mmol, 400 µL); yield: 260 mg (25%). 29b.
1H NMR (CDCl3, 400 MHz) δ: 5.76-5.72 (m, 2H, dCH + ring
proton), 5.63 (m, 1H), 5.48-5.42 (m, 2H), 5.02-4.91 (m, 3H),
4.58 (d, 1H, J ) 8.0 Hz), 4.42 (d, 1H, J ) 7.6 Hz), 4.3-4.14
(m, 1H), 3.96-3.89 (m, 2H), 3.73-3.66 (m, 4H, COOMe + ring
proton), 3.50-3.48 (m, 2H), 2.95 (dd, 1H, J ) 3.2 Hz), 2.15-
1.95 (m, 17H, 5 × COCH3 + CH2), 1.75-1.71 (m, 2H, CH2).
13C NMR (CDCl3, 400 MHz) δ: 170.9, 170.1, 169.5, 168.6,
168.5, 144.8, 137.8, 116.0, 115.4, 102.1, 96.2, 73.2, 72.6, 69.1,
68.3, 68.0, 65.7, 62.6, 61.2, 53.0, 30.3, 28.6, 21.2, 21.1, 21.0,
20.9. HRMS-FAB: [M+ + H] calcd for C28H39O16N3, 674.2364;
found, 674.2330.
Synthesis of Deprotected â-(1,4)-Linked 2-Azido
Disaccharide 30b. Compound 28b (1.48 mmol, 1.07 g) was
dissolved in a 9:1 MeOH/H2O mixture (5 mL) and stirred at 0
°C. A 3 M NaOH solution was added, and the reaction mixture
was allowed to stir at room temperature for 2 h (monitored
by TLC). The reaction was then quenched using a strongly
acidic Dowex H+ ion-exchange resin and filtered, and the
filtrate was concentrated in vacuo. The crude product was then
purified by gel filtration using a Sephadex LH-20 column with
methanol as the eluant. Removal of the solvent followed by
subsequent lyophilization gave the saponified product as a
white solid. Yield: 650 mg (82%). 1H NMR (CD3OD, 400 MHz)
δ: 7.43 (d, 2H, ArH, J ) 6.8 Hz), 7.41 (m, 3H), 5.84-5.79 (m,
1H, dCH), 5.06 (d, 1H, J ) 10.4 Hz), 5.05-4.98 (m, 2H, d
CH2), 4.66 (d, 1H, J ) 10.8 Hz), 4.60 (d, 1H, H-1′, J ) 8.0 Hz),
4.34 (d, 1H, H-1, J ) 7.8 Hz), 3.98-3.85 (m, 4H), 3.74 (d, 1H,
J ) 9.6 Hz), 3.59-3.54 (m, 2H), 3.33-3.24 (m, 3H), 2.16-2.12
(m, 2H, CH2), 1.69-1.66 (m, 2H, CH2). 13C NMR (CD3OD, 400
MHz) δ: 171.1, 138.1, 128.4, 128.0, 127.3, 114.2, 103.4, 102.0,
81.6, 76.9, 76.1, 75.7, 75.5, 75.1, 74.2, 71.9, 69.0, 66.6, 60.0,
30.1, 28.8. HRMS-FAB: [M+ + Li] calcd for C24H32O11N3,
546.2275; found, 546.2269.
The above deprotected compound (1.21 mmol, 650 mg) was
dissolved in 3 mL of dry methanol and cooled to -78 °C. Ozone
was bubbled through the light-yellow solution for 1.5 h. The
pale-yellow solution appeared to turn pale blue. To this
solution was added 3 mL of dimethyl sulfide at -78 °C, and
the reaction was stirred and allowed to warm to room tem-
perature overnight. The reaction mixture was concentrated,
and the crude product was purified using Sephadex LH-20
with methanol as the eluant. The pure compound 30b obtained
was then lyophilized to give an off-white solid. Yield: 554 mg
(85%). 30b. 1H NMR (CD3OD, 400 MHz) δ: 9.72 (s, 1H, CHO),
7.43 (d, 2H, ArH, J ) 7.6 Hz), 7.32-7.26 (m, 3H, ArH), 5.06
(d, 1H, J ) 10.8 Hz), 4.65 (d, 1H, J ) 10.8 Hz), 4.60 (d, 1H, J
) 8.0 Hz), 4.56-4.54 (m, 0.5H), 4.4-4.38 (m, 0.5H), 4.35 (d,
1H, J ) 8.0 Hz), 3.94-3.85 (m, 4H), 3.75 (d, 1H, J ) 10 Hz),
1
744.3048. 32b. H NMR (CDCl3, 400 MHz) δ: 7.30-7.21 (m,
5H, ArH), 5.9 (br m, 1H, NH), 5.81-5.68 (m, 1H, dCH), 5.23-
5.15 (m, 2H, dCH2), 5.02-4.89 (m, 3H), 4.78 (d, 1H, J ) 12.4
Hz), 4.70 (d, 1H, J ) 5.4 Hz), 4.65 (d, 1H, J ) 12.4 Hz), 4.58
(d, 1H, H-1, J ) 8.4 Hz), 4.45 (dd, 1H, J ) 4 Hz, J ) 3.2 Hz),
4.23 (dd, 1H, J ) 6 Hz), 3.95-3.90 (m, 2H), 3.81-3.64 (m, 4H),
3.60 (s, 3H, COOMe), 3.42-3.38 (m, 1H), 2.06 (s, 3H, COCH3),
2.04 (s, 3H, COCH3), 2.03-1.99 (m, 2H, CH2), 1.98 (s, 3H,
COCH3), 1.97 (s, 3H, COCH3), 1.83 (s, 3H, NHCOCH3), 1.66-
1.58 (m, 2H, CH2). 13C NMR (CDCl3, 400 MHz) δ: 170.7, 170.5,
170.1, 169.9, 169.6, 166.9, 138.7, 138.2, 128.5, 127.7, 115.1,
100.4, 100.0, 76.9, 73.6, 72.8, 72.5, 71.8, 71.5, 69.5, 69.1, 63.6,
53.2, 53.1, 30.2, 28.8, 23.4, 21.1, 20.7, 20.6. HRMS-FAB: [M+
+ H] calcd for C35H47O16N, 738.2973; found, 738.2964.
Synthesis of 34a/34b. To a solution of acetamido derivative
32a/32b (0.82 mmol, 605 mg) dissolved in a 9:1 MeOH/H2O
mixture (3 mL) was added a 3 M NaOH solution at 0 °C, and
the reaction mixture was allowed to stir at room temperature
for 2 h (monitored by TLC). The addition of a strongly acidic
Dowex H+ ion-exchange resin to the reaction mixture not only
neutralized the excess base but also changed the pH of the
solution from basic to acidic. Subsequently, the reaction
mixture was filtered, concentrated, and subjected to gel
filtration using a Sephadex LH-20 column with methanol as
the eluant. Removal of the solvent followed by subsequent
lyophilization gave the deacteylated and deesterified product
1
as a white solid 33a/33b (yield: 391 mg, 86%). 33a. H NMR
(CD3OD, 400 MHz) δ: 7.9 (d, 1H, NH, J ) 9.2 Hz), 7.35-7.24
(m, 5H, ArH), 5.85-5.76 (m, 1H, dCH), 5.03-4.99 (m, 4H),
4.71 (d, 1H, H-1, J ) 3.6 Hz), 4.61 (t, 3H, J ) 7.2 Hz′, J )
10.8 Hz), 4.12-4.02 (m, 1H), 4.02-3.88 (m, 2H), 3.87-3.76 (m,
2H), 3.70-3.67 (m, 2H), 3.52 (t, 1H, J ) 8.8 Hz, J ) 8.8 Hz),
3.42-3.35 (m, 2H), 3.29-3.26 (m, 1H), 2.18-2.10 (m, 2H, CH2),
1.87 (s, 3H, NHCOCH3), 1.71-1.65 (m, 2H, CH2). 13C NMR
(CD3OD, 400 MHz) δ: 176.1, 176.0, 143.1, 142.1, 131.9, 131.2,
118.7, 107.4, 101.4, 83.1, 81.2, 80.1, 79.4, 78.2, 75.9, 75.5, 71.1,
64.2, 57.0, 34.1, 32.5, 25.5. HRMS-FAB: [M+ + Li] calcd for
1
C26H37O12N, 562.2476; found, 562.2465. 33b. H NMR (CD3-
OD, 400 MHz) δ: 7.32-7.22 (m, 5H, ArH), 5.85-5.76 (m, 1H,
dCH), 5.01-4.97 (m, 4H), 4.61 (d, 1H, H-1, J ) 7.6 Hz), 4.55
(d, 1H, J ) 11.2 Hz), 4.45 (d, 1H, J ) 7.6 Hz), 3.98-3.83 (m,
4H), 3.76-3.67 (m, 3H), 3.56-3.54 (m, 1H), 3.48-3.36 (m, 2H),
3.29-3.26 (m, 1H), 2.08-2.06 (m, 2H, CH2), 1.80 (s, 3H,
NHCOCH3), 1.62-1.58 (m, 2H, CH2). 13C NMR (CD3OD, 400
MHz) δ: 176.0, 175.1, 143.1, 142.1, 132.2, 132.1, 131.8, 118.1,
107.5, 105.3, 85.2, 81.5, 80.1, 79.5, 78.3, 78.2, 75.9, 72.6, 64.3,
59.3, 33.9, 32.7, 25.8. HRMS-FAB: [M+ + Li] calcd for
C26H37O12N, 562.2476; found, 562.2465.
J. Org. Chem, Vol. 69, No. 26, 2004 9169

Rele et al.
The saponified product obtained, 33a/33b (0.704 mmol, 391
mg), was dissolved in 3 mL of dry methanol and cooled to -78
°C. Ozone was bubbled through the light-yellow solution for
1.5 h. The pale-yellow solution appeared to turn pale blue. To
this solution was added 3 mL of dimethyl sulfide at -78 °C,
and the reaction was stirred and allowed to warm to room
temperature overnight. The reaction mixture was concen-
trated, and the crude product was purified using Sephadex
LH-20 with methanol as the eluant. The pure compound
obtained was then lyophilized to give an off-white solid
(yield: 353 mg, 90%). 34a. 1H NMR (CD3OD, 400 MHz) δ: 9.70
(s, 1H, CHO), 7.96 (d, 1H, NH, J ) 9.2 Hz), 7.40-7.19 (m, 5H,
ArH), 5.05 (d, 2H, J ) 11.2 Hz), 4.72 (d, 1H, H-1, J ) 3.2 Hz),
4.62 (t, 2H, H-1′ merged), 4.52 (m, 1H), 4.12 (m, 1H), 3.94-
3.91 (m, 2H), 3.83-3.66 (m, 4H), 3.56-3.49 (m, 1H), 3.47-
3.37 (m, 2H), 3.29-3.26 (m, 1H), 1.88 (s, 3H, NHCOCH3),
1.68-1.66 (m, 4H, CH2). 13C NMR (CD3OD, 400 MHz) δ: 203.6,
172.7, 172.1, 139.1, 128.1, 128.0, 127.3, 103.4, 98.4, 97.5, 79.2,
77.2, 76.3, 75.4, 74.4, 72.1, 71.6, 67.5, 60.4, 53.1, 33.4, 33.3,
24.6, 21.5. HRMS-FAB: [M+ + 2Li - H] calcd for C25H35O13N,
54.1, 52.7, 29.4, 24.7, 23.1. HRMS-FAB: calcd for C18H29O3N,
474.2556; found, 474.2552. 35b. 1H NMR (DMSO-d6, 400 MHz)
δ: 9.72 (br s, 1H, -CHO), 7.82 (br s, 1H, -NH, J ) 9.2 Hz),
5.4-5.2 (br peak, OH), 4.40 (d, 1H, H-1, J ) 7.6 Hz), 4.34 (d,
1H, H-1′, J ) 8.0 Hz), 4.30-4.25 (m, 1H), 3.81-3.54 (m, 4H),
3.42-3.38 (m, 4H), 3.33-3.30 (m, 1H), 3.22-3.11 (m, 2H),
3.02-2.96 (m, 1H), 1.75 (s, 3H, NHCOCH3), 1.68-1.48 (m, 4H,
CH2). 13C NMR (CD3OD, 400 MHz) δ: 197.8, 171.2, 169.5,
104.3, 103.5, 101.5, 80.7, 76.5, 75.6, 75.4, 73.6, 72.7, 72.1, 68.8,
60.7, 55.4, 52.9, 29.1, 24.8, 23.5. HRMS-FAB: [M+ + Li] calcd.
for C18H29O3N, 474.2556; found, 474.2550.
Synthesis of Gemini Heparinoid-Mimetic Disaccha-
rides 4a/4b. Compound 35a/35b (0.14 mmol, 65 mg) was
dissolved in 0.5 mL of dry methanol, and ethylenediamine (0.5
equiv, 0.07 mmol, 4.67 µL) was added in a dropwise manner.
There was an immediate precipitation of a white solid. The
reaction mixture was stirred for 3 h at room temperature,
NaBH3CN (4 equiv, 56 mmol, 36 mg) was then added , and
the reaction mixture was allowed to stir overnight. Removal
of the solvent gave a residue, which was purified using
Sephadex G-25 with water as the eluant and subsequently
lyophilized to give a pale-yellow powder (4a/4b) (yield: 45 mg,
67%). 4a. 1H NMR (D2O, 400 MHz) δ: 7.86 (d, J ) 7.2 Hz), [in
DMSO-d6, the R-anomeric peak appears at 4.6 (d, 2H, H-1, J
) 3.2 Hz)], 4.39 (d, 1H, H-1′, J ) 8.0 Hz), 4.37 (d, 1H, H-1′, J
) 7.6 Hz), 3.82-3.50 (m, 20H), 3.36-3.10 (m, 12H), 2.98 (t,
2H), 2.67 (t, 2H), 1.86 (s, 6H, NHCOCH3), 1.62-1.48 (m, 8H,
CH2). 13C NMR (D2O, 400 MHz) δ: 173.1, 170.2, 104.4, 103.4,
96.9, 96.8, 80.8, 76.7, 74.2, 73.7, 72.4, 71.4, 69.4, 60.5, 54.1,
53.2, 30.1, 23.9, 22.1. HRMS-FAB: [M+ + H] calcd for
C38H67O24N4, 963.4239; found, 963.4214. 4b. 1H NMR (D2O,
400 MHz) δ: 4.46 (d, 1H, J ) 8 Hz), 4.40-4.36 (merged d,
3H, J ) 8.0 Hz), 3.81-3.74 (m, 3H), 3.66 (dd, 2H, J ) 4.8 Hz,
J ) 4.8 Hz), 3.58-3.22 (m, 12H), 3.19-3.15 (m, 12H), 2.96 (t,
2H), 2.85 (m, 2H), 2.72 (t, 2H), 1.85 (s, 6H, NHCOCH3), 1.52-
1.41 (m, 8H, CH2). 13C NMR (D2O, 400 MHz) δ: 175.6, 174.4,
104.7, 104.6, 102.3, 101.1, 79.1, 75.9, 75.4, 74.8, 73.1, 72.4, 71.8,
69.9, 60.2, 55.2, 53.3, 53.0, 28.5, 23.7, 22.2. HRMS-FAB: [M+
+ H] calcd for C38H67O24N4, 963.4430; found, 963.4482.
1
570.2358; found, 570.2352. 34b. H NMR (CD3OD, 400 MHz)
δ: 9.72 (s, 1H, CHO), 7.98 (d, 1H, NH, J ) 7.6 Hz), 7.33-7.23
(m, 5H, ArH), 4.64 (s, 1H), 4.62 (d, 1H, H-1, J ) 8.0 Hz), 4.56
(d, 1H, J ) 11.6 Hz), 4.48 (br s, 1H), 4.46 (d, 1H, H-1′, J ) 7.8
Hz), 3.94-3.84 (m, 4H), 3.77-3.63 (m, 2H), 3.58-3.55 (m, 2H),
3.45-3.39 (m, 2H), 3.29-3.26 (m, 1H, H-2), 1.83 (s, 3H,
NHCOCH3), 1.64-1.58 (br m, 4H, CH2). 13C NMR (CD3OD,
400 MHz) δ: 204.2, 172.2, 171.2, 139.2, 128.2, 128.1, 127.3,
103.5, 101.3, 98.4, 89.5, 81.4, 77.6, 76.2, 75.6, 74.4, 74.3, 72.0,
69.2, 60.4, 55.3, 33.2, 33.1, 28.9, 22.0. HRMS-FAB: [M+
2Li - H] calcd for C25H35O13N, 570.2360; found, 570.2352.
+
Synthesis of 35a/35b. A solution of 34a/34b (0.64 mmol,
353 mg) in 8:2 MeOH/H2O was hydrogenated in the presence
of 10% Pd-C (4 equiv). After 24 h and upon completion of the
reaction (TLC), the suspension was filtered through a pad of
Celite and concentrated to give the debenzylated product. Final
purification of the compound was carried out on Sephadex LH-
20 using MeOH as the eluant. The absence of aromatic peaks
in the 1H NMR spectrum indicated complete debenzylation
(yield: 207 mg, 70%). 35a. 1H NMR (DMSO, 400 MHz) δ: 9.64
(s, 1H, CHO), 7.81 (d, 1H, NH, J ) 9.2 Hz), 5.2-4.98 (br peak,
OH), 4.60 (d, 1H, H-1, J ) 2.8 Hz), 4.34 (d, 1H, H-1′, J ) 8.0
Hz), 4.32-3.94 (m, 1H), 3.68-3.52 (m, 6H), 3.48-3.35 (m, 2H),
3.28-3.24 (t, 1H, J ) 8.8 Hz, J ) 9.2 Hz), 3.2-3.12 (m, 2 H),
3.01-2.98 (m, 1H), 1.78 (s, 3H, NHCOCH3), 1.52-1.48 (m, 4H,
CH2). 13C NMR (DMSO, 400 MHz) δ: 204.1, 170.8, 170.1,
103.6, 96.9, 80.9, 76.5, 75.6, 73.7, 72.0, 71.3, 69.3, 67.4, 60.3,
Supporting Information Available: General experimen-
tal details and 1H and 13C NMR spectra of compounds 4a, 4b,
and 13-35b are available free of charge via the Internet at
http://pubs.acs.org.
JO049092R
9170 J. Org. Chem., Vol. 69, No. 26, 2004