Synthesis of clustered xenotransplantation antagonists using palladium-catalyzed cross-coupling of prop-2-ynyl α-D-galactopyranoside

Article abstract of DOI:10.1039/b101234g

Hyperacute rejection of pig liver transplantation can be antagonized using high affinity anti α-galactoside epitopes (Galα1-3Galβ, Galili antigen). To this end, clusters containing the Galili antigen were synthesized using palladium cross-coupling reactions. Thus, benzyl 3-(O-α-D-galactopyranosyl)-β-D-galactopyranoside derivative 6 was prepared using perbenzylated thiophenyl β-D-galactopyranosyl donor 5 and benzyl 2,6-di-O-benzoyl-α-D-galactopyranosyl acceptor 3. Aglycone interchange from benzyl to prop-2-ynyl disaccharide 8 was successively achieved by hydrogenolysis, peracetylation, regioselective anomeric de-O-acetylation, trichloroacetimidate activation, and finally coupling to propargyl alcohol with triflic acid. The resulting prop-2-ynyl disaccharide 8 was transformed in high yields (>85%) into dimer 10, trimer 12, and tetramer 17 using oxidative homocoupling (10) and palladium-catalyzed aryl iodide cross-coupling (12,17), respectively. Ester protecting group removal under transesterification conditions afforded fully deprotected α-Gal clusters.

Full text of DOI:10.1039/b101234g

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Synthesis of clustered xenotransplantation antagonists
using palladium-catalyzed cross-coupling of prop-2-ynyl
ꢀ-D-galactopyranoside
1
Bingcan Liu and René Roy*
Department of Chemistry, Centre for Research in Biopharmaceuticals, University of Ottawa,
Ottawa, ON, Canada K1N 6N5. Fax: ϩ1 613 562 5170
Received (in Cambridge, UK) 7th February 2001, Accepted 1st March 2001
First published as an Advance Article on the web 23rd March 2001
Hyperacute rejection of pig liver transplantation can be antagonized using high affinity anti α-galactoside epitopes
(Galα1-3Galβ, Galili antigen). To this end, clusters containing the Galili antigen were synthesized using palladium
cross-coupling reactions. Thus, benzyl 3-(O-α--galactopyranosyl)-β--galactopyranoside derivative 6 was prepared
using perbenzylated thiophenyl β--galactopyranosyl donor 5 and benzyl 2,6-di-O-benzoyl-α--galactopyranosyl
acceptor 3. Aglycone interchange from benzyl to prop-2-ynyl disaccharide 8 was successively achieved by
hydrogenolysis, peracetylation, regioselective anomeric de-O-acetylation, trichloroacetimidate activation, and finally
coupling to propargyl alcohol with triflic acid. The resulting prop-2-ynyl disaccharide 8 was transformed in high
yields (>85%) into dimer 10, trimer 12, and tetramer 17 using oxidative homocoupling (10) and palladium-catalyzed
aryl iodide cross-coupling (12, 17), respectively. Ester protecting group removal under transesterification conditions
afforded fully deprotected α-Gal clusters.
be obtained by oxidative homocoupling, while trimer 12 and
tetramer 17 could be obtained by a Sonogashira cross-coupling
between 8 and aryl iodides 11 and 16, respectively.
Initial attempts to introduce the suitably protected prop-2-
ynyl β--galactopyranoside as a glycosyl acceptor were met
with some difficulties due to either incompatibility of the
alkyne moieties toward glycosylating reagents such as N-iodo-
succinimide–triflic acid† (NIS–TfOH) or to the lack of selec-
tive benzyl ether deprotection. Therefore a feasible strategy
was designed in which the required prop-2-ynyl aglycone was
introduced at the very end of the disaccharide synthesis. Thus,
benzyl 2,6-di-O-benzoyl-α--galactopyranoside acceptor 3 was
Introduction
Galili antigens are cell surface carbohydrate motifs containing
Galα1-3Galβ non-reducing ends responsible for hyperacute
rejection following pig to human xenotransplantation.1 As a
result of organ donor shortage, pig’s livers have been highly
considered as a viable alternative since their size, availability,
and low risk of viral transmission have been well established.2
Unfortunately, most humans possess abundant anti α-Gal anti-
bodies (1–2% IgG, 3–8% IgM) as a result of prior exposure to
bacterial or viral infections.3 Thus, within minutes to hours of
surgery and in spite of immunosuppressive treatments, the
transplanted organs are rapidly rejected by most recipients. To
synthesized using an established protocol (Scheme 1). Starting
overcome this problem, a possible strategy has been to develop
antagonizing anti α-Gal inhibitors by infusing large concen-
trations of synthetic α-Gal oligosaccharides or by using
affinity columns to deplete the patient’s serum of α-Gal anti-
bodies.4 Polymeric α-Gal epitopes have also been designed to
increase the antagonizing activity of simple, low affinity
oligosaccharides.5,6
In the present study, the synthesis of various α-Gal clusters is
described for the purpose of creating smaller size, high affinity
anti-α-Gal antibody antagonists. Previous investigations by us7
and others,8 including the findings by Magnusson et al.9 on
nanomolar inhibition of bacterial receptors by galactobiose
clusters, strongly support the concept of multivalent inhibitors.
Scheme 1 Reagents and conditions: i) 2,2-dimethoxypropane, PTSA,
rt, 5 h, 85%; ii) BzCl, pyridine, DMAP, rt, 2 h, 90%; iii) 60% AcOH,
Results and discussion
80 ЊC, 5 h, 95%.
Synthesis of Galili epitope (Galꢀ1-3Galꢁ-OR)
from the known12 benzyl α--galactopyranoside 1, 3,4-diol free
galactosyl acceptor 3 was obtained by regioselective acetalation
of 1 with 2,2-dimethoxypropane in the presence of toluene-p-
sulfonic acid (PTSA) to provide the corresponding acetal 2a in
Recent investigations have shown that propynyl to aryl halide
palladium-catalyzed cross-coupling of saccharides bearing
these functionalities provided an efficient entry into glyco-
clusters having high affinity.10 The rationale for rigid-linkers is
85% yield. Treatment of the resulting diol 2a with benzoyl
based on the findings that conformationally restricted ligands
chloride (pyridine, 4-(N,N-dimethylamino)pyridine (DMAP))
are exposed to a lesser entropic loss during binding to their
homologous receptors.11 To reach this goal for the Galili antigen,
we chose prop-2-ynyl disaccharide 8 from which dimer 10 could
† The IUPAC name for triflic acid is trifluoromethanesulfonic acid.
J. Chem. Soc., Perkin Trans. 1, 2001, 773–779
DOI: 10.1039/b101234g
773
This journal is © The Royal Society of Chemistry 2001

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gave 2b (90%). Acetal hydrolysis in hot 60% aqueous acetic acid
(80 ЊC) afforded galactosyl acceptor 3 in 95% yield.
syl donor 5b using NIS–triflic acid as promoter gave disacchar-
ide 6 as an α–β mixture (6 : 1, 76%) from which the pure
α-anomer (δ H-1Ј 4.84 ppm, J1Ј,2Ј 3.8 Hz) could be isolated in
65% yield after careful column chromatography separation.
The desired prop-2-ynyl disaccharide 8 was then readily syn-
thesized through a sequence of straightforward manipulations.
Benzyl glycoside 6 was hydrogenolyzed (H2, Pd-C, MeOH,
95%) and peracetylated using acetyl chloride to give 7a
(pyridine, DMAP, 90%). The anomeric mixture was then
regioselectively de-O-acetylated using hydrazinium acetate to
provide the reducing disaccharide 7b that was transformed into
trichloroacetimidate 7c (Cl3CCN, DBU, CH2Cl2, 90%). Finally,
treatment of 7c with propargyl‡ alcohol with triflic acid
afforded β-galactoside 8 in 86% yield (δ H-1 4.90 ppm, J1,2 7.9
Hz).
Perbenzylated phenyl β--thiogalactosyl donor 5b13 was pre-
pared from known peracetylated β--galactopyranoside 4 (i)
NaOMe, MeOH; ii) NaH, BnBr, DMF, 85%) obtained under
phase transfer catalyzed conditions (PTC) previously developed
in our group (Scheme 2).14 Glycosidation of diol 3 with galacto-
Synthesis of ꢀ-Gal containing clusters
In order to obtain Galili antigens as small rigid clusters that
may have the potential to form stable cross-linked complexes
with naturally occurring anti α-Gal antibodies, a series of
palladium catalyzed transformations were then effected. The
goal was to obtain di- to tetra-mers analogous to those previ-
ously prepared for galabiosides that have shown nanomolar
inhibitory properties against various pathogens with carbo-
hydrate binding lectins on their surfaces.9
Dimer 9 was successfully prepared from prop-2-ynyl glyco-
side 8 by oxidative homocoupling using palladium catalyzed
conditions ((Ph3P)2PdCl2, CuI, DMF–TEA (1 : 1), rt, 95%)
(Scheme 2).15 Interestingly, these conditions were found to be
milder than the classical Glaser reaction16 previously used by us
in analogous circumstances.17 Acetylenic dimer 9 was then sub-
jected to Zemplèn transesterification (NaOMe, MeOH) to
provide fully deprotected dimer 10 in 95% yield.
Trimer 13 was then obtained using a Sonogashira reaction18
similar to the one recently described for the synthesis of
mannoside clusters.10,15 Thus, palladium catalysts such as
(PPh3)2PdCl2, (PPh3)4Pd and Pd2(dba)3 were all successfully
applied for the cross-coupling of prop-2-ynyl glycoside 8 and
1,3,5-triiodobenzene (11)19 which provided trimer 12 with more
or less analogous yields (85% with (PPh3)2PdCl2) (Scheme 3).
As previously observed,15 it is noteworthy to mention that CuI
is not essential for the successful cross-coupling in this reaction.
In fact, without CuI, the coupling reaction proceeds under heat-
ing conditions (60 ЊC) while the reaction can function at room
Scheme 2 Reagents and conditions: i) NaOMe, MeOH, quant. (95%
for 10); ii) BnBr, NaH, DMF, rt, 2 h, 85%; iii) 3, NIS, TfOH, CH2Cl2,
4 Å MS, Ϫ40 ЊC, 1 h, α–β (6 : 1), 76%; iv) H2, Pd-C, MeOH, rt, 2 days,
95%; v) AcCl, pyridine, DMAP, rt, 4 h, 90%; vi) H2NNH2–HOAc,
CH2Cl2, rt, 2 h, 75%; vii) Cl3CCN, DBU, CH2Cl2, rt, 5 h, 90%; viii)
propargyl alcohol, CH2Cl2, TfOH, 4 Å MS, rt, 4 h, 86%; ix) (Ph3P)2-
PdCl2, CuI, DMF–TEA 1 : 1, rt, 3 h, 95%.
‡ The IUPAC name for propargyl is prop-2-ynyl.
Scheme 3 Reagents and conditions: i) (Ph3P)2PdCl2, DMF–TEA 1 : 1, 60 ЊC, 5 h, 81%; ii) NaOMe, MeOH, 24 h, rt, 95%.
J. Chem. Soc., Perkin Trans. 1, 2001, 773–779
774

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temperature in the presence of CuI, albeit in lower yields. How-
ever in the absence of CuI, the oxidative homodimerization of
the alkynyl moiety is abolished. These observations are particu-
larly important when the desired cross-coupling products and
the homodimers are difficult to separate. Finally, trimer 12 was
fully deprotected to its corresponding analog 13 using NaOMe
in methanol (95%).
To provide sufficient inter-sugar distances required for
efficient protein cross-linking, a tetrameric core was con-
structed using pentaerythritol (14) as a seed molecule. Tetra-p-
iodobenzylated precursor 16 was prepared by a classical
etherification procedure using p-iodobenzyl bromide 15,
tetrabutylammonium iodide, NaH, and DMF in moderate
yield (47%) (Scheme 4). In spite of several efforts, the yield
DMF-TEA (1 : 1), 60 ЊC, 5 h) afforded fully protected tetramer
17 (Scheme 5). Usual deprotection (NaOMe, MeOH) of all
ester protecting groups in 17 gave tetramer 18 (95%).
In conclusion, efficient oxidative homocoupling or palladium
catalyzed cross-coupling reactions of alkynyl glycosides with
aryl iodides provided an efficient entry into novel “rigid-like”
glycoclusters that may have the potential to inhibit various
carbohydrate binding proteins (lectins and antibodies). Work is
now in progress to evaluate the above series of clusters bearing
the Galili epitope against human anti α-Gal antibodies.1
Experimental
General
Dichloromethane was stored over 4 Å MS after drying over
P2O5 and distillation. Propargyl alcohol was from Fluka and
p-iodobenzyl bromide was from KARL INDUSTRIES INC.
All the other reagents were purchased from Aldrich. Thin layer
chromatography was performed on Silica Gel F254 (Merck) pre-
coated aluminium sheets and visualized with molybdenum
solution and an UV lamp. Column chromatography was run on
Ultra Pure Silica Gel (SILICYCLE). Elemental analyses were
measured on a CE-2500 instrument. Melting points were
determined on a Gallenkamp melting point apparatus without
temperature correction. Optical rotations were measured on a
PERKIN-ELMER 241 polarimeter and are given as 10Ϫ1 deg
cm2 gϪ1. All the NMR spectra (500 MHz for 1H and 125.7 MHz
for 13C) were recorded on a Bruker AMX-500 spectrometer.
The resonances were assigned based on 1H, 13C, 1H–1H COSY,
DEPT, and HMQC experiments. Chemical shifts are refer-
enced to CDCH3 (δH 7.29 and δC 77.0 ppm). J values are given
in Hz. ESI-MS analyses were carried out on a MICROMASS
Quattro LC. FAB-MS spectra were recorded on KRATOS
Concepts IIH with Csϩ beam. MALDI-TOF MS was acquired
on a PerSeptive Biosystems Elite-STR (Framingham, MA,
USA).
Scheme 4 Reagents and conditions: i) NaH, TBAI, DMF, rt, 5 h, 47%.
could not be improved further. However, for reasons not yet
fully understood, 14 is known to be reluctant to full derivatiz-
ation under Williamson etherification. Moreover, p-iodobenzyl
bromide is light sensitive and unstable in various solvents which
may contribute to the low yield.
Glycosyl acceptor 3
Using the cross-coupling conditions described above for the
synthesis of trimer 12 without CuI, treatment of propynyl
glycoside 8 with tetrakis p-iodobenzyl ether 16 ((PPh3)2PdCl2),
Benzyl 3,4-O-isopropylidene-ꢀ-D-galactopyranoside 2a. Benzyl
α--galactopyranoside 112 (246 mg, 1 mmol) was dissolved into
acetone (5 ml) to which was added 2,2-dimethoxypropane (250
Scheme 5 Reagents and conditions: i) (Ph3P)2PdCl2, DMF–TEA (1 : 1), 60 ЊC, 5 h, 85%; ii) NaOMe, MeOH, 24 h, rt, 95%.
J. Chem. Soc., Perkin Trans. 1, 2001, 773–779
775

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µl, 2 mmol) and a catalytic amount of toluene-p-sulfonic acid.
The mixture was stirred at room temperature for 5 h. After
removal of the solvent under reduced pressure, the residue
was purified by silica gel column chromatography using ethyl
acetate–hexane (1.5 : 1) to give 2a as a thick liquid in 85% yield,
[α]D ϩ110 (c 3.2, CHCl3); δH (500 MHz; CDCl3) 7.28–7.36 (5 H,
m, aromatic), 4.99 (1 H, d, J = 3.8, H-1), 4.77 (1 H, d, J = 11.8,
PhCH2), 4.57 (1 H, d, J = 11.8, PhCH2), 4.26 (1 H, t, J = 6.2,
H-4), 4.21 (1 H, dd, J = 3.3, 7.2, H-3), 4.07–4.09 (1 H, m, H-5),
3.89 (1 H, dd, J = 6.3, 11.8, H-6a), 3.82 (1 H, dd, J = 3.8, 7.2,
H-2), 3.77 (1 H, dd, J = 6.3, 11.8, H-6b), 1.48 (3 H, s, CH3), 1.32
(3 H, s, CH3); δC (125 MHz; CDCl3) 137.0, 128.5, 128.1, 128.0
and 109.8 (aromatic), 96.9 (C-1), 76.1 (C-4), 73.9 (C-3), 69.4
(PhCH2), 69.3 (C-2), 68.4 (C-5), 62.7 (C-6), 27.5 and 25.8
(CH3); m/z (FAB-MS) 349.2 (Found: [M ϩ Kϩ]. C16H22O6
requires [M ϩ Kϩ] 349.1).
The solution was stirred overnight at room temperature. After
neutralization of the reaction mixture with Amberlite (IR-120,
Hϩ), the solution was filtered through a cotton plug. Methanol
was evaporated under reduced pressure to provide 5a in a quan-
titative yield and the product was used directly in the next step
without further purification.
Compound 5a was dissolved into 30 ml of DMF to which was
added sodium hydride (440 mg, 11 mmol). After stirring at
room temperature for 20 minutes, benzyl bromide (1.3 ml, 10.9
mmol) was added to the suspension during a period of 10 min-
utes. The mixture was further stirred at room temperature for
2 hours. Excess sodium hydride was destroyed with several
drops of methanol. The solution was diluted with 100 ml of
ether and washed with 3 × 100 ml of water. The organic phase
was dried over anhydrous sodium sulfate and the solvent was
removed under reduced pressure. The residue was purified by
silica gel column chromatography using ethyl acetate–hexane
(1 : 10) to provide 5b as a white solid with a mp 88.9 to 90.1 ЊC
and a [α]D 1 (c 1.0, CHCl3), literature13 mp 88–89 ЊC and a [α]D 1
(c 1.0, CHCl3).
Benzyl 2,6-di-O-benzoyl-3,4-O-isopropylidene-ꢀ-D-galacto-
pyranoside 2b. Compound 2a (310 mg, 1 mmol) was dissolved
into pyridine (5 ml) and a catalytic amount of DMAP (10 mg)
was added followed by benzoyl chloride (280 µl, 2.2 mmol). The
mixture was stirred at room temperature for 2 h. After removal
of pyridine under reduced pressure, the residue was dissolved
into 50 ml of ether which was washed with water (3 × 30 ml).
The organic layer was dried over anhydrous sodium sulfate and
the solvent was removed under reduced pressure. The residue
was purified by silica gel column chromatography using ethyl
acetate–hexane (1 : 4) to provide 2b as a liquid in 90% yield, [α]D
ϩ90.2 (c 1, CHCl3); δH (500 MHz; CDCl3) 8.03–8.09 (4 H, m,
aromatic), 7.55–7.59 (6 H, m, aromatic), 7.15–7.20 (5 H, m,
aromatic), 5.20 (1 H, dd, J = 3.7, 7.9, H-2), 5.17 (1 H, d, J = 3.7,
H-1), 4.77 (1 H, d, J = 12.3, PhCH2), 4.67 (1 H, dd, J = 4.8,
11.8, H-6a), 4.63 (1 H, dd, J = 7.5, 11.8, H-6b), 4.50 (1 H, dd,
J = 5.5, 7.9, H-3), 4.50 (1 H, d, J = 11.8, PhCH2), 4.47–4.51
(1 H, m, H-5), 4.38 (1 H, dd, J = 2.6, 5.5, H-4), 1.56 (3 H, s,
CH3), 1.36 (3 H, s, CH3); δC (125 MHz; CDCl3) 166.3, 165.9
(PhCO), 136.8, 133.1, 133.0, 130.0, 129.8, 129.7, 129.6, 128.4,
128.3, 128.2, 127.8, 127.5 and 110.1 (aromatic), 95.3 (C-1), 73.6
(C-2), 72.1 (C-4), 69.6 (PhCH2), 66.1 (C-3 and C-5), 64.0 (C-6),
27.9 and 26.3 (CH3); m/z (FAB-MS) 557.0 (Found: [M ϩ Kϩ].
C30H30O8 requires [M ϩ Kϩ] 557.0).
Benzyl (2,3,4,6-tetra-O-benzyl-ꢀ-D-galactopyranosyl)-(1→3)-
2,6-di-O-benzoyl-ꢀ-D-galactopyranoside 6. Glycosyl donor 5b
(200 mg, 0.32 mmol) and glycosyl acceptor 3 (230 mg, 0.48
mmol) and 4 Å MS were put into a dry flask. After flushing the
flask with dry nitrogen, 20 ml of dry CH2Cl2 was injected into
it. The suspension was stirred for 30 min at Ϫ40 ЊC, then NIS
(106.6 mg, 0.48 mmol) and a catalytic amount of triflic acid was
added to the solution. One hour later, the mixture was neutral-
ized with DIPEA and filtered over Celite. The filtrate was
washed with diluted sodium thiosulfate solution and the
organic phase was dried over anhydrous sodium sulfate. The
solvent was evaporated on a rotavapor. The residue was sub-
jected to silica gel column chromatography using ethyl acetate–
hexane (2 : 3) to afford 6 as a white foam in 65% yield, [α]D
ϩ84.4 (c 1.8, CHCl3); δH (500 MHz; CDCl3) 8.08–8.11 (4 H, m,
aromatic), 7.08–7.60 (31 H, m, aromatic), 5.45 (1 H, J = 3.8,
10.0, dd, H-2), 5.24 (1 H, d, J = 3.8, H-1), 4.88 (1 H, d, J = 11.5,
PhCH2), 4.84 (1 H, d, J = 3.8, H-1Ј), 4.82 (1 H, d, J = 11.5,
PhCH2), 4.70 (1 H, d, J = 12.5, PhCH2), 4.69 (2 H, s, PhCH2),
4.65 (1 H, d, J = 11.5, PhCH2), 4.63 (1 H, dd, J = 6.3, 11.6,
H-6a), 4.56 (1 H, dd, J = 7.6, 11.6, H-6b), 4.48 (1 H, d, J = 12.5,
PhCH2), 4.45 (1 H, d, J = 11.5, PhCH2), 4.27 (1 H, t, J = 6.6,
H-5), 4.25 (1 H, dd, J = 3.4, 10.0, H-3), 4.16 (1 H, d, J = 11.5,
PhCH2), 4.08 (1 H, d, J = 11.5, PhCH2), 4.02 (1 H, dd, J = 3.8,
10.0, H-2Ј), 4.01 (1 H, m, H-4), 3.91–3.94 (2 H, m , H-3Ј, H-5Ј),
3.86 (1 H, br d, J = 1.5, H-4Ј), 3.47 (1 H, t, J = 8.6, H-6aЈ), 3.22
(1 H, dd, J = 5.5, 8.8, H-6bЈ); δC (125 MHz; CDCl3), 166.3 and
165.8 (PhCO2), 138.6, 138.3, 137.9, 137.5, 137.0, 136.5, 135.5,
133.0, 131.3, 130.1, 129.8, 129.6, 129.4, 128.7, 128.5, 128,4,
128.3, 128.2, 128.1, 128.0, 127.7, 127.6, 127.5, 127.4 and 127.3
(aromatic), 96.1 (C-1), 95.4 (C-1Ј), 79.5 (C-3Ј or C-5Ј), 75.6
(C-2Ј), 74.9 and 74.8 (PhCH2), 74.5 (C-3), 74.4 (C-4Ј), 73.2 and
72.6 (PhCH2), 69.8 (C-5Ј or C-3Ј), 69.6 (C-2), 69.3 (PhCH2),
Benzyl 2,6-di-O-benzoyl-ꢀ-D-galactopyranoside 3. Benzyl 2,6-
di-O-benzoyl-3,4-O-isopropylidene-α--galactopyranoside 2b
(518 mg, 1 mmol) was placed into a round-bottomed flask with
30 ml of 60% aqueous acetic acid. The mixture was heated and
stirred at 80 ЊC for 5 h. After removing the solvent under
reduced pressure, the residue was purified by silica gel column
chromatography using ethyl acetate–hexane (1.5 : 1) to yield 3
as a white solid in 95% yield, [α]D ϩ111.6 (c 1, CHCl3); δH (500
MHz; CDCl3) 8.02–8.11 (4 H, m, aromatic), 7.53–7.58 (2 H, m,
aromatic), 7.39–7.46 (4 H, m, aromatic), 7.16–7.29 (5 H, m,
aromatic), 5.33 (1 H, dd, J = 3.8, 10.2, H-2), 5.21 (1 H, d,
J = 3.8, H-1), 4.72 (1 H, d, J = 12.2, PhCH2), 4.66 (1 H, dd,
J = 6.0, 11.4, H-6a), 4.52 (1 H, d, J = 12.2, PhCH2), 4.50 (1 H,
dd, J = 7.0, 11.4, H-6b), 4.27 (1 H, d, J = 1.4, H-5), 4.26 (1 H,
dd, J = 3.6, 10.2, H-3), 4.10 (1 H, dd, J = 1.0, 3.6, H-4), 2.97
(2 H, br s, -OH); δC (125 MHz; CDCl3) 166.9 and 166.6
(PhCO2), 137.0, 133.3, 133.2, 129.9, 129.6, 128.4, 128.3, 127.8
and 127.6 (aromatic), 95.7 (C-1), 72.1 (C-2), 69.6 (PhCH2), 69.5
(C-4), 68.4 and 68.2 (C-3 and C-5), 63.5 (C-6); m/z (ESI-MS)
68.3 (C-6Ј), 67.8 (C-5), 66.2 (C-4), 64.5 (C-6); m/z (ESI-MS)
ϩ
1018.1 (Found: [M ϩ NH4ϩ]. C61H64NO13 requires [M ϩ NH4
]
1018.4).
Prop-2-ynyl
(2,3,4,6-tetra-O-acetyl-ꢀ-D-galactopyranosyl)-
(1→3)-4-O-acetyl-2,6-di-O-benzoyl-ꢁ-D-galactopyranoside 8. To
compound 6 (1 g, 1 mmol) dissolved in 30 ml of methanol was
added a catalytic amount of 10% Pd/C and 2 drops of acetic
acid. The suspension was subjected to hydrogenolysis at rt for
2 days. After the reaction was completed, the solution was fil-
tered through a pad of Celite. After removal of methanol under
reduced pressure, the dry residue was dissolved into 10 ml of
pyridine to which was added acetyl chloride (0.52 ml, 6 mmol)
and a catalytic amount of DMAP (10 mg) at 0 ЊC. The mixture
was stirred at room temperature for 4 h. Pyridine was then
removed under reduced pressure. The residue was dissolved
ϩ
496.0 (Found: [M ϩ NH4ϩ]. C27H30NO8 requires [M ϩ NH4
]
496.2).
ꢀ-Gal epitope 8
Phenyl 2,3,4,6-tetra-O-benzyl-1-thio-ꢁ-D-galactopyranoside
5b. Phenyl 2,3,4,6-tetra-O-acetyl-1-thio-β--galactopyranoside
(4)14 (1 g, 2.27 mmol) was dissolved into 25 ml of methanol to
which was added a catalytic amount of sodium methoxide.
776
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into 60 ml of ether and washed with water (3 × 40 ml). The
organic layer was dried over anhydrous sodium sulfate. After
removal of ether under reduced pressure, the residue was puri-
fied by silica gel column chromatography using ethyl acetate–
hexane (1 : 1) to provide 7a (α–β) as a white foam in 90% yield
which was used in the next step without further purification.
Compound 7a (800 mg, 1 mmol) was dissolved into 20 ml of
dichloromethane to which was added hydrazinium acetate (135
mg, 1.5 mmol). The solution was stirred at room temperature
for 2 h. After removal of the solvent under reduced pressure,
the residue was subjected to silica gel column chromatography
using ethyl acetate–hexane (1.5 : 1) to give 7b as a white foam
in 75% yield. Compound 7b was obtained as an anomeric
α–β mixture, which was used directly in the next step without
further purification.
Compound 7b (760 mg, 1 mmol), trichloroacetonitrile (0.15
ml, 1.5 mmol), and DBU (30 µl, 0.2 mmol) were added to 20 ml
of dry dichloromethane. The solution was stirred at room tem-
perature for 5 h. After removal of the solvent under reduced
pressure, the residue was subjected to silica gel column chrom-
atography using ethyl acetate–hexane (1 : 1) to give 7c as a white
foam in 90% yield. Compound 7c existed as an α–β mixture
(~10 : 1 ratio), which was used directly in the next step without
further purification.
Crude trichloroacetimidate 7c (200 mg, 0.22 mmol), propar-
gyl alcohol (25.8 µl, 0.44 mmol) and 4 Å MS were added into 20
ml of dry dichloromethane. The mixture was stirred at room
temperature for 30 minutes to which was added a catalytic
amount of triflic acid. Four hours later the suspension was neu-
tralized with DIPEA and filtered over Celite. After evaporation
of the solvent, the residue was purified by silica gel column
chromatography using ethyl acetate–hexane (1 : 1) to provide 8
as a white foam in 86% yield, [α]D ϩ117.0 (c 1, CHCl3); δH (500
MHz; CDCl3) 7.98–8.06 (4 H, m, aromatic), 7.53–7.57 (2 H, m,
aromatic), 7.40–7.45 (4 H, m, aromatic), 5.52 (1 H, dd, J = 7.9,
10.1, H-2), 5.50 (1 H, br d, J = 2.4, H-4), 5.21 (1 H, br d, J ~3.5
Hz, H-1Ј), 5.19 (1 H, dd, J = 3.5, 10.5, H-2Ј), 5.03 (1 H, dd,
J = 3.3, 10.5, H-3Ј), 4.91 (1 H, m, H-4Ј), 4.90 (1 H, d, J = 7.9,
H-1), 4.53 (1 H, dd, J = 6.6, 11.3, H-6a), 4.41 (1 H, dd, J = 2.4,
16.1, CH2-CCH), 4.34 (1 H, dd, J = 2.4, 16.1, CH2-CCH), 4.32
(1 H, dd, J = 6.7, 11.3, H-6b), 4.10 (1 H, dd, J = 3.2, 10.1, H-3),
4.03 (1 H, m, H-5), 4.00 (1 H, m, H-5Ј), 3.80 (1 H, dd, J = 6.8,
12.1, H-6aЈ), 3.72 (1 H, dd, J = 6.6, 12.1, H-6bЈ), 3.45 (1 H, t,
J = 3.4, acetylenic), 2.20, 2.04, 2.00, 1.89 and 1.86 (each 3 H, 5s,
5 CH3CO2); δC (125 MHz; CDCl3) 170.2, 169.9, 169.8 and 169.3
(CH3CO2), 165.7 and 165.0 (PhCO2), 133.4 (2), 129.7, 129.6,
129.3, 129.2, 128.5 and 128.4 (aromatic), 98.7 (C-1), 95.7 (C-1Ј),
78.2 and 75.3 (acetylenic), 73.7 (C-3), 71.1 (C-5), 69.8 (C-2),
67.6 (C-4Ј), 66.9 (C-3Ј and C-5Ј), 66.5 (C-2Ј), 65.2 (C-4), 61.8
(C-6), 61.1 (C-6Ј), 55.9 (C-1Љ), 20.7, 20.5, 20.3 (3) (CH3CO2);
m/z (FAB-MS) 837.2 (Found: [M ϩ Kϩ]. C39H42O8 requires
[M ϩ Kϩ] 837.2).
J = 7.1, H-5Ј), 3.77 (2 H, dd, J = 7.1, 11.1, H-6aЈ), 3.68 (2 H, dd,
J = 6.6, 11.1, H-6bЈ), 2.19, 2.03, 2.00, 1.86 and 1.83 (each 6 H,
5s, 10 CH3CO2); δC (125 MHz; CDCl3) 170.2, 169.9, 169.8,
169.7, 169.3 (CH3CO2), 166.0, 165.1 (PhCO2), 133.5, 133.4,
129.7, 129.3, 129.1, 128.5, 128.4 and 128.1 (aromatic), 99.2
(C-1), 93.4 (C-1Ј), 74.3 (acetylenic), 73.7 (C-3), 71.2 (C-5), 70.3
(acetylenic), 69.8 (C-2), 67.6 (C-4Ј), 66.9 (C-3Ј and C-5Ј), 66.5
(C-2Ј), 65.2 (C-4), 61.7 (C-6), 61.0 (C-6Ј), 56.5 (C-1Љ), 20.7, 20.5
and 20.4 (CH3CO2); m/z (FAB-HRMS) 1633.4570 (Found:
[M ϩ Kϩ]. C78H82O36 requires [M ϩ Kϩ] 1633.4223).
Fully deprotected ꢀ-Gal homodimer 10
α-Gal homodimer 9 (80 mg, 0.050 mmol) was dissolved into 30
ml of methanol. A catalytic amount of sodium methoxide was
then added to the solution. The mixture was stirred at room
temperature for 24 h. Sodium methoxide was neutralized with
AMBERLITE IR-120 (Hϩ). After careful filtration of the
resin through a loose cotton plug, methanol was removed under
reduced pressure. After evaporation of the methanol under
reduced pressure, the dried residue was extracted with ether
3 × 10 ml to remove methyl benzoate to afford the deprotected
glycocluster 10 as a white powder in 95% yield, [α]D 0.0 (c 2.1,
H2O); δH (500 MHz; H2O) 5.22 (2 H, d, J = 3.9, H-1Ј), 4.69
(2 H, d, J = 7.9, H-1), 4.65 (4 H, br s, H-1Љ), 4.25 (4 H, m, H-5
and H-5Ј), 4.08 (2 H, m, H-4Ј), 4.02 (2 H, dd, J = 3.4, 10.4,
H-3Ј), 3.92 (2 H, dd, J = 3.9, 10.4, H-2Ј), 3.84–3.94 (6 H, m, H-3
and H-6 or H-3 and H-6Ј), 3.76–3.81 (6 H, m, H-4 and H-6 or
H-4 and H-6Ј), 3.72 (2 H, dd, J = 7.9, 10.0, H-2); δC (125 MHz;
D2O) 100.6 (C-1), 94.8 (C-1Ј), 76.8 (C-3), 74.6 (C-2), 74.5 (acet-
ylenic), 70.42 (C-5 or C-5Ј), 70.0 (acetylenic), 68.8 (C-3Ј), 68.7
(C-4Ј), 68.6 (C-4), 67.7 (C-2Ј), 64.3 (C-5 or C-5Ј), 60.5 and 60.4
(C-6 and C-6Ј), 56.5 (C-1Љ); m/z (FAB-MS) 797.1 (Found:
[M ϩ Kϩ]. C30H46O22 requires [M ϩ Kϩ] 797.2).
ꢀ-Gal trimer 12
To a solution of compound 8 (100 mg, 0.12 mmol) in 5 ml of
DMF–TEA (1 : 1) was added Pd(PPh3)2Cl2 (4.3 mg, 0.006
mmol (5 mol%)) and 1,3,5-triiodobenzene (11)19 (15.8 mg,
0.035 mmol). The solution was stirred under nitrogen at 60 ЊC
for 5 h. The solvent and TEA were evaporated under reduced
pressure and the resulting mixture was purified by silica gel
column chromatography using ethyl acetate–hexane (2 : 1) to
provide compound 12 as a white foam in a yield of 85%, [α]D
ϩ93.4 (c 1.8, CHCl3); δH (500 MHz; CDCl3) 7.98–8.00 (12 H,
m, aromatic), 7.32–7.47 (18 H, m, aromatic), 5.53 (3 H, dd,
J = 8.0, 10.0, H-2), 5.46 (3 H, d, J = 3.0, H-4), 5.19 (3 H, d,
J < 1, H-1Ј), 5.18 (3 H, dd, J = 3.5, 10.5, H-2Ј), 5.04 (3 H, dd,
J = 3.3, 10.5, H-3Ј), 4.91 (3 H, d, J = 8.0, H-1), 4.90 (3 H, m,
H-4Ј), 4.60 (6 H, s, H-1Љ), 4.55 (3 H, dd, J = 6.4, 11.3, H-6a),
4.32 (3 H, dd, J = 6.6, 11.3, H-6b), 4.12 (3 H, dd, J = 3.1, 10.0,
H-3), 4.04 (3 H, t, J = 6.7, H-5), 3.97 (3 H, t, J = 6.9, H-5Ј), 3.74
(3 H, dd, J = 7.1, 11.1, H-6aЈ), 3.68 (3 H, dd, J = 6.6, 11.1,
H-6bЈ), 2.19, 2.03, 2.00, 1.86 and 1.72 (each 9 H, 5s, 15
CH3CO2); δC (125 MHz; CDCl3) 170.2, 169.9, 169.8, 169.7 and
169.3 (CH3CO2), 166.0 and165.1 (PhCO2), 134.7, 133.4, 133.3,
129.7, 129.6, 129.3, 129.1, 128.5, 128.4 and 122.1 (aromatic),
99.2 (C-1), 94.0 (C-1Ј), 85.4 and 84.5 (acetylenic), 73.9 (C-3),
71.1 (C-5), 70.1 (C-2), 67.6 (C-4Ј), 66.9 (C-3Ј), 66.5 (C-2Ј and
C-5Ј), 65.3 (C-4), 61.8 (C-6), 61.0 (C-6Ј), 56.5 (C-1Љ), 20.7, 20.6,
20.4 and 20.3 (CH3CO2); m/z (ESI-MS) 2485.1 (Found:
[M ϩ NH4ϩ]. C123H130NO54 requires [M ϩ NH4ϩ] 2484.7).
ꢀ-Gal homodimer 9
To a solution of compound 8 (79.8 mg, 0.1 mmol) in 5 ml of
DMF–TEA (1 : 1) was added Pd(PPh3)2Cl2 (3.6 mg, 5 mol%)
and CuI (3.8 mg, 20 mol%). The solution was stirred under
nitrogen at room temperature for 3 h. The solvent and TEA
were evaporated under reduced pressure. The resulting mixture
was purified by silica gel column chromatography using ethyl
acetate–hexane (1.5 : 1) to provide compound 9 as a white foam
in 95% yield, [α]D ϩ115.6 (c 1, CHCl3); δH (500 MHz; CDCl3)
7.98–8.00 (8 H, m, aromatic), 7.32–7.47 (12 H, m, aromatic),
5.49–5.53 (4 H, m, H-2, H-4), 5.19 (2 H, d, J < 1, H-1Ј), 5.18
(2 H, dd, J = 3.5, 10.5, H-2Ј), 5.02 (2 H, dd, J = 3.3, 10.5, H-3Ј),
4.90 (2 H, m, H-4Ј), 4.83 (2 H, d, J = 7.9, H-1), 4.54 (2 H, dd,
J = 6.6, 11.3, H-6a), 4.46 (2 H, d, J = 16.1, H-1aЉ), 4.41 (2 H, d,
J = 16.1, H-1bЉ), 4.30 (2 H, dd, J = 6.6, 11.3, H-6b), 4.11 (2 H,
dd, J = 3.1, 10.1, H-3), 4.04 (2 H, t, J = 6.7, H-5), 3.96 (2 H, t,
Fully deprotected ꢀ-Gal trimer 13
α-Gal trimer 12 (90 mg, 0.036 mmol) was dissolved into 30 ml
of methanol. A catalytic amount of sodium methoxide was
added to the solution. The mixture was stirred at room tem-
perature for 24 h. Sodium methoxide was neutralized with
AMBERLITE IR-120 (Hϩ). After careful filtration of the
J. Chem. Soc., Perkin Trans. 1, 2001, 773–779
777

View Article Online
resin through a loose cotton plug, methanol was removed under
reduced pressure. The dried residue was extracted with ether
3 × 10 ml to remove methyl benzoate to provide the fully depro-
tected glycocluster 13 as an off-white foam in 95% yield, [α]D 0.0
(c 1.5, H2O); δH (500 MHz; D2O) 7.67 (3 H, s, aromatic), 5.23
(3 H, d, J = 3.8, H-1Ј), 4.79 (6 H, br s, H-1Љ), 4.75 (3 H, d,
J = 7.9, H-1), 4.27 (6 H, m, H-5 and H-5Ј), 4.09 (3 H, m, H-4Ј),
4.04 (3 H, dd, J = 3.3, 10.4, H-3Ј), 3.94 (3 H, dd, J = 3.9, 10.4,
H-2Ј), 3.76–3.89 (21 H, m, H-2, H-3, H-4, H-6 and H-6Ј);
δC (125 MHz; D2O) 134.4 and 122.2 (aromatic), 100.8 (C-1),
94.9 (C-1Ј), 85.2 and 84.6 (acetylenic), 76.9 (C-3), 74.6 (C-2 or
C-4), 72.3 (C-5 or C-5Ј), 70.4 (C-3Ј), 68.9 and 68.7 (C-3Ј, C-4Ј
and C-4 or C-3Ј, C-4Ј and C-2), 67.8 (C-2Ј), 64.4 (C-5 or C-5Ј),
60.5 (C-6 and C-6Ј), 56.7 (C-1Љ); m/z (ESI-MS) 1230.0 (Found:
[M ϩ NH4ϩ]. C51H76NO33 requires [M ϩ NH4ϩ] 1230.4).
ml of methanol to which a catalytic amount of sodium methox-
ide was added. The mixture was stirred at room temperature for
24 h. Sodium methoxide was neutralized with AMBERLITE
IR-120 (Hϩ). After careful filtration of the resin through a
loose cotton plug, methanol was removed under reduced pres-
sure, the dried residue was extracted with ether 3 × 10 ml to
remove methyl benzoate to afford the deprotected glycocluster
18 as an off-white foam in 95% yield, [α]D 0.0 (c 1.8, H2O);
δH (500 MHz; D2O) 7.30 (8 H, br s, aryl), 6.98 (8 H, br s, aryl),
5.21 (4 H, br s, H-1Ј), 4.82 (8 H, s, PhCH2 or H-1Љ), 4.63 (8 H, s,
PhCH2 or H-1Љ), 4.60 (4 H, br s, H-1), 4.23 (8 H, br s, H-5
and H-5Ј), 4.06 (4 H, br s, H-4Ј), 4.01 (4 H, br d, J = 10.8,
H-3Ј), 3.93 (4 H, dd, J = 2.9, 10.1, H-2Ј), 3.77–3.83 (28 H, m,
H-2, H-3, H-4, H-6 and H-6Ј), 3.42 (8 H, s, C(CH2OR)4);
δC (125 MHz; D2O) 138.5, 131.3, 126.8 and 120.6 (aromatic),
101.1 (C-1), 95.0 (C-1Ј), 85.2 and 84.5 (acetylenic), 77.2,
70.3, 68.9, 68.7, 67.8, 64.2, 60.5, 60.2, 56.6 (C-4, C-5, C-6, C-2Ј,
C-3Ј, C-4Ј, C-5Ј, C-6Ј, PhCH2, C-1Љ); m/z (ESI-MS) 1022.2
(Found: [M ϩ 2NH4ϩ]. C93H132N2O48 requires [M ϩ 2NH4ϩ],
1022.4).
Pentaerythritol tetrakis(p-iodobenzyl) ether 16
Pentaerythritol (14) (27.2 mg, 0.2 mmol) was dissolved into 10
ml of dry DMF, then a catalytic amount of tetrabutylam-
monium iodide and 46 mg of NaH (0.96 mmol, 1.2 eq./OH)
were added to the solution. After one hour, p-iodobenzyl brom-
ide (15) (286 mg, 0.96 mmol) was added. The mixture was
stirred at room temperature for 5 h. Excess NaH was slowly
quenched with cold water. The solution was washed with 3 × 20
ml of ether and the organic phases were combined together and
washed with 3 × 20 ml of water. After drying over anhydrous
sodium sulfate the solvent was evaporated and the residue was
carefully separated by silica gel column chromatography using
hexane–ether (7 : 1) to give 16 as a white solid in 47% yield,
mp 136–138 ЊC; δH (500 MHz; CDCl3) 7.58 (8 H, d, J = 8.1,
aromatic), 6.95 (8 H, d, J = 8.1, aromatic), 4.36 (8 H, s, PhCH2),
3.45 (8 H, s, C(CH2-)4); δC (125 MHz; CDCl3) 138.2, 137.3,
129.2 and 92.8 (aromatic), 72.5 (PhCH2), 69.1 (C(CH2-)4), 45.5
(C(CH2-)4); m/z (FAB-MS) 1001.2 (Found: [M ϩ Hϩ]. C33H33O4
requires [M ϩ Hϩ] 1001.0) (Anal. Calcd for C33H32O4I4: C,
39.63; H, 3.22. Found: C, 39.83; H, 3.19%).
Acknowledgements
Dedicated to the late Professor Göran Magnusson who was an
inspiration throughout my career in carbohydrate chemistry.
We thank the National Science and Engineering Research
Council of Canada (NSERC) for financial support.
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ꢀ-Gal tetramer 17
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To a solution of compound 8 (150 mg, 0.188 mmol) in 10 ml of
DMF–TEA (1 : 1) were added Pd(PPh3)2Cl2 (6.7 mg, 0.009
mmol (5 mol%)) and 16 (39.1 mg, 0.039 mmol). The solution
was stirred under nitrogen at 60 ЊC for 5 h. The solvent and
TEA were evaporated under reduced pressure. The resulting
mixture was purified by silica gel column chromatography using
ethyl acetate–hexane (2 : 1) to provide compound 17 as a white
foam in 81% yield, [α]D ϩ97.5 (c 1, CHCl3); δH (500 MHz;
CDCl3) 7.98–8.02 (16 H, m, aromatic), 7.40–7.55 (16 H, m,
aromatic), 7.14–7.29 (20 H, m, aromatic), 5.54 (4 H, dd, J = 8.0,
10.1, H-2), 5.51 (4 H, d, J = 2.8, H-4), 5.20 (4 H, d, J < 1, H-1Ј),
5.18 (4 H, dd, J = 3.5, 10.4, H-2Ј), 5.03 (4 H, dd, J = 3.3, 10.4,
H-3Ј), 4.90 (4 H, d, J = 8.0, H-1), 4.88 (4 H, m, H-4Ј), 4.58 (8 H,
s, H-1Љ), 4.55 (4 H, dd, J = 6.4, 11.3, H-6a), 4.45 (8 H, s,
PhCH2), 4.32 (4 H, dd, J = 6.6, 11.3, H-6b), 4.10 (4 H, dd,
J = 3.1, 10.1, H-3), 4.04 (4 H, t, J = 6.7, H-5), 3.96 (4 H, t,
J = 6.5, H-5Ј), 3.73 (4 H, dd, J = 6.6, 11.2, H-6aЈ), 3.66 (4 H, dd,
J = 6.6, 11.2, H-6bЈ), 3.56 (8 H, s, C(CH2OR)4), 2.20, 2.03, 1.99,
1.86 and 1.72 (each 12 H, 5s, 20 CH3CO2); δC (125 MHz;
CDCl3) 170.2, 169.9, 169.8, 169.7 and 169.3 (CH3CO2), 166.0,
165.0 (PhCO2), 139.5, 133.4, 133.3, 132.1, 132.0, 131.7, 129.7,
129.4, 129.3, 128.5, 128.4, 127.1, 127.0 and 121.1 (aromatic),
98.9 (C-1), 93.7 (C-1Ј), 86.7 and 83.5 (acetylenic), 73.7 (C-3),
72.8 (PhCH2), 71.0 (C-5), 70.0 (C-2), 69.5 (C(CH2OR)4), 67.6
(C-4Ј), 66.9 (C-3Ј), 66.5 (C-2Ј), 66.4 (C-5Ј), 65.1 (C-4), 61.7
(C-6), 61.2 (C-6Ј), 56.8 (C-1Љ), 45.7 (C(CH2OR)4), 20.7, 20.4
and 20.3 (CH3CO2); m/z (MALDI-TOF MS) 3706.55 (Found:
[M ϩ Naϩ]. C189H196O78 requires [M ϩ Naϩ] 3706.68).
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Fully deprotected ꢀ-Gal tetramer 18
α-Gal tetramer 17 (95 mg, 0.026 mmol) was dissolved into 40
778
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J. Chem. Soc., Perkin Trans. 1, 2001, 773–779
779