New mannotriosides and trimannosides as potential ligands for mannose-specific binding proteins

Article abstract of DOI:10.1139/v02-120

The α-D-mannopyranosyl and 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranosyl neoglycolipids 3-6 and the branched and cluster trimannosidic acids 38 and 41 have been made in connection with studies of liposomes as transporters of antigens to dendritic cells

Full text of DOI:10.1139/v02-120

964
New mannotriosides and trimannosides as
potential ligands for mannose-specific binding
proteins
Richard H. Furneaux, Zbigniew Pakulski, and Peter C. Tyler
Abstract: The α-D-mannopyranosyl and 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranosyl neoglycolipids 3–6 and
the branched and cluster trimannosidic acids 38 and 41 have been made in connection with studies of liposomes as
transporters of antigens to dendritic cells.
Key words: mannotrioside, trimannoside, neoglycoconjugate, glycolipid, synthesis.
Résumé : Dans le cadre d’études sur les liposomes comme transporteurs d’antigènes dans des cellules dendritiques, on
a préparé les néoglycolipides α-D-mannopyranosyl- et 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopyranosyle (3–6) ainsi
que les acides trimannosidiques ramifié et en agrégat (38 et 41).
Mots clés : mannotrioside, trimannoside, néoglycoconjugué, glycolipide, synthèse.
[Traduit par la Rédaction] Furneaux et al. 972
Introduction
mannopyranosyl trichloroacetimidate as glycosylating agent
has been described (4).
Terminal α-D-mannopyranosyl units commonly occur in
the glycoproteins of pathogenic bacteria, yeasts, viruses, and
various parasites and are recognised by receptors of macro-
phages (1) and dendritic cells (2, 3). Because the branched
trisaccharide 3,6-di-O-(α-D-mannopyranosyl)-α-D-mannopy-
ranosyl ligand binds with higher affinity to the mannose re-
ceptors of human macrophages and dendritic cells than do
mono- and linear oligo-mannosyl ligands (4), we began
studies of synthetic neoglycoconjugates and liposomes con-
taining this trimer with a view to using the liposomes to
transport antigens to dendritic cells.
We now report (Scheme 1) the preparation of the neogly-
colipids 3–6 which differ from analogues 1 and 2 in having
four-carbon (compounds 3 and 5) and six-carbon (com-
pounds 4 and 6) as opposed to nine-carbon atom spacer
groups. Additionally, and with further neoglycolipids in
view, the polyoxyethylene glycosides 30 and 34 and the
mannotriose-containing acid 38 were made, as well as the
alternative cluster type of trimannosyl glycosidic (8) acid 41
(Scheme 2). Considerable effort has gone into the study of
this last kind of trimannoside (9).
In the first report of this work (5), we exploited the earlier
finding (6, 7) that various α-D-mannopyranosides could be
di-O-mannosylated concurrently and with some selectivity at
O-3 and O-6 by use of tetra-O-acetyl-α-D-mannopyranosyl
bromide to give the required branched-chain mannotriose
derivatives, and we introduced the new tetra-O-benzoyl-α-D-
mannopyranosyl trichloroacetimidate as glycosylating agent.
By its use 8-(methoxycarbonyl)octyl α-D-mannopyranoside
was converted to the 3,6-bis-O-(2,3,4,6-tetra-O-benzoyl-α-D-
mannopyranosyl)-mannopyranoside in 55% yield, and the
neoglycolipids 1 and 2 (Scheme 1) were made (5). More re-
cently the 3,6-di-O-α-D-mannosylation of p-nitrophenyl α-D-
mannopyranoside in 42% yield by use of tetra-O-acetyl-α-D-
Results and discussion
The neoglycolipids 3 and 4 (Scheme 1) were made fol-
lowing glycosylations of methyl 4-hydroxybutanoate and
methyl 6-hydroxyhexanoate, respectively, by use of tetra-O-
benzoyl-α-D-mannopyranosyl trichloroacetimidate. The first
products 7 and 11 were converted successively to the re-
quired neoglycolipids by the previously used processes (5),
that is, via the debenzoylated glycosides 8 and 12, the
carboxylic acids 9 and 13 and the N-succinimido derivatives
10 and 14, respectively. The final step involved coupling of
the activated acids 10 and 14 with dipalmitoyl-1-α-phospha-
tidyl ethanolamine (DPPE) to give products 3 and 4.
Selective bis-(tetra-O-benzoyl-α-D-mannosylation) of gly-
cosides 8 and 12, by use of the glycosylating agent men-
tioned above, afforded the trisaccharide derivatives 15 and
21 which were isolated following chromatographic separa-
tion of their derived diacetates 16 (23% from 8) and 22
(33% from 12), respectively. Compounds 16 and 22 were
used to prepare the neoglycolipids 5 and 6, respectively,
Received 30 April 2002. Published on the NRC Research
Press Web site at http://canjchem.nrc.ca on 5 August 2002.
Dedicated to the memory of Professor Raymond U. Lemieux.
R.H. Furneaux,1 Z. Pakulski,2 and P.C. Tyler. Industrial
Research Ltd, PO Box 31–310, Lower Hutt, New Zealand
1Corresponding author (e-mail: r.furneaux@irl.cri.nz).
2On leave from the Institute of Organic Chemistry, Polish
Academy of Sciences, Warsaw, Poland.
by the routes used for compounds
3 and 4, i.e.,
16→18→19→20→5 and 22→24→25→26→6 as also out-
lined in Scheme 1. The 1→3-, 1→6-linked trisaccharide
Can. J. Chem. 80: 964–972 (2002)
DOI: 10.1139/V02-120
© 2002 NRC Canada

Furneaux et al.
965
Scheme 1.
R2O
RO
R3O
R2O
O
O
HO
OR1
OH
RO
CO2R4
14
( )
O
O
(
O
O
( )n
O
)
n
O
14
OPO
ONa
( )
N
H
O
O
R3
R1
R2
n
R4
Compound
R
Compound
n
8
8
3
5
3
5
H
Man
H
H
Man
Man
1
2
3
4
5
6
Bz
H
H
H
Bz
H
H
H
H
Ac
Ac
H
H
H
Bz
H
H
H
Bz
H
H
H
Man Bz
Man Bz
Man Bz
Man
Man
Man
Man Bz
Man Bz
Man Bz
Man
Man
Man
Me
Me
H
Suc
Me
Me
H
Suc
Me
Me
Me
Me
H
Suc
Me
Me
Me
Me
H
3
7
8
9
Bz
H
H
H
Bz
H
H
H
H
Ac
Man Bz
H
H
H
H
Ac
i
3
3
3
5
5
5
5
3
3
3
3
3
3
5
5
5
5
5
5
ii
iv
iv
iii
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
i
v
ii
iii
iv
iv
vi
vii
ii
v
iii
H
vi
vii
ii
Ac
Ac
H
H
H
Man Bz
H
H
H
Man = α−D-mannopyranosyl
Man Bz = tetra-O-benzoyl-α-
Suc = N-succinimido
D-mannopyranosyl
Suc
iii
Reagents: i, K2CO3, MeOH; ii, NaOH (aq.); iii, N-hydroxysuccinimide; DCC; iv,
1,2-hexadecanoyl-sn-glycero-3-phosphoethanolamine, NaHCO3; v,
trichloroacetimidate; vi, Ac2O, Py; vii, NaOMe, MeOH
tetra-O-benzoyl-D-mannopyranosyl
structures of the products 15 and 21 followed from the
1H NMR characteristics of their diacetates 16 and 22, the
H-2 and H-4 resonances being ca. 1 ppm deshielded with re-
spect to those of H-3, and the J2,3 and J3,4 coupling constant
values being 3.4 and 9.8 Hz, respectively. These data com-
pare closely with those obtained for the corresponding tri-
saccharide octa-O-benzoyl diacetate prepared in the course
of making compound 2 (5).
In all the above cases the glycosylation reactions gave no
observable orthoester by-products.
En route to alternative types of neoglycolipids the mono-
and trisaccharide glycosides 27 and 35 (Scheme 2) were
made by standard condensations of tetra-O-benzoyl-α-D-
mannopyranosyl trichloroacetimidate with the corresponding
alcohols whereas, in our hands, the corresponding acetylgly-
cosyl bromide with mercury(II) cyanide was more effective
for the preparation of triglycoside 39 from [tris(hydroxy-
methyl)]methyl-(5-methoxycarbonyl)pentanoamide (8).
From 27 and 35 the acids 30 and 38 were made by standard
methods, and from 27 the extended-chain compound 34 was
produced via the azide 31. The dimannosylation of com-
pound 29 gave the 1→3-,1→6-linked trisaccharide derivative
35 in 40% yield, the positions of substitution being deter-
mined from the 1H,1H coupling constants of the H-2 and H-4
resonances of the derived diacetate 36. De-esterification of
the substituted triglycoside 39 gave acid 41 via methyl ester 40.
The biological evaluation of compounds described above
will be reported separately.
During the isolation of the acetylated trisaccharide deriva-
tives 16 and 22 the (3,4,6-tri-O-mannosyl)mannoside by-
products 17 and 23, containing small proportions of other
1
oligosaccharide derivatives, were also obtained. The H NMR
spectra of monoacetates 17 and 23 showed δ values for the
H-2 resonances of the central mannose residues of 5.34 and
5.43, respectively. The less-deshielding acetyl groups are
consequently concluded to be substituted at O-2 since, for
both of the compounds, the nine protons at the secondary
benzoylated ring positions resonated within the range δ 5.79–
6.27. The tetrasaccharide by-products are therefore 3,4,6-
tris-O-(tetra-O-benzoyl-mannopyranosyl)-mannopyranosides.
© 2002 NRC Canada

966
Can. J. Chem. Vol. 80, 2002
Scheme 2.
R2O
O
R1
R1O
R2O
O
O
OR1
CO2R2
R1
N
H
O
R1O
O
X
O
O
R1
Bz
R2
Compound
R1
R2
X
Compound
39
40
41
Me
Me
H
Man Ac
Man
Cl
Bz
Bz
H
27
28
29
30
ix
iii
i
CN
Bz
H
ii
Man
iv
CN
iii
H
H
CO2H
N3
Bz
Bz
Bz
H
31
32
33
34
35
36
37
38
Bz
Bz
Bz
H
v
vi
ii
NH2
vii
NHCO(CH2)4CO2Me
NHCO(CH2)4CO2Me
H
CN
Man Bz
Man Bz
Man
viii
ii
Ac
H
CN
CN
Man Ac = tetra-O-acetyl-α-
For other abbreviations see Scheme 1.
D-mannopyranosyl.
iii
H
CO2H
Man
Reagents: i, KCN, DMSO; ii, K2CO3, MeOH; iii, NaOH (aq.); iv, NaN3, DMF; v, NaBH4, NiCl2, EtOH;
vi, methyl adipate, 2-ethoxy-1-ethoxycarbonyl-1,2-didydroquinoline ; vii, tetra-O-benzoyl-D-mannopyranosyl
trichloroacetimidate; viii, Ac2O, Py; ix, NaOMe, MeOH.
methyl 4-hydroxybutanoate (10) (760 mg, 6.4 mmol) in
CH2Cl2 (30 mL) was stirred for 30 min at room temperature
over molecular sieves (4 Å, 700 mg, finely ground). The so-
lution was cooled in an ice bath and TmsOTf (60 µL) was
added. After 20 min the reaction was quenched by addition
of Et3N (0.5 mL), and the solvent was evaporated in vacuo.
Column chromatography (hexane–EtOAc, 2:1) gave the title
compound 7 (3.60 g, 92%, contaminated with 10% of insep-
arable methyl 4-hydroxybutanoate): [α]2D0 −47 (c 1.0, CHCl3,
Experimental
General methods
All glycosylation reactions were performed under argon in
dry CH2Cl2. TLC was performed on silica gel HF-254 and
column chromatography on silica gel 230–400 mesh
(Merck). 1H NMR spectra were recorded with a Bruker
Avance 300 MHz NMR spectrometer equipped with a 5 mm
o.d. Quattro nucleus probe or (when indicated) a Varian
Unity 500 MHz spectrometer equipped with 5 mm Universe
probe. Resonance assignments were by use of DEPT, H,H-
COSY, inverse C,H-COSY, and H,H-TOCSY experiments.
All spectra were consistent with those expected for the struc-
tures assigned to the respective compounds; unresolved reso-
nances and those of substituent groups and carbonyl
groups (13C spectra) are frequently not listed. High resolu-
tion mass spectra (HR-MS) were measured with a VG70-
250S double focussing magnetic sector mass spectrometer or
a Mariner 8105 electrospray TOF mass spectrometer. Opti-
cal rotations were measured with a PerkinElmer 241 auto-
matic polarimeter. 2-[2-(2-Chloroethoxy)ethoxy]ethanol was
purchased from Aldrich.
corrected for the impurity). 1H NMR (CDCl3) δ: 6.10 (t, J3,4
,
J4,5 = 10 Hz, 1H, H-4), 5.88 (dd, J2,3 = 3.3 Hz, 1H, H-3),
5.69 (dd, J1,2 = 1.8 Hz, 1H, H-2), 5.08 (d, 1H, H-1), 4.69
(dd, J5,6 = 2.5 Hz, J6,6′ = 12.0 Hz, 1H, H-6), 4.50 (dd, J5,6′
=
4.4 Hz, 1H, H-6′), 4.42 (m, 1H, H-5), 3.90 (dt, J = 9.8,
6.2 Hz, 1H, OCHH), 3.73 (s, 3H, Me), 3.61 (dt, J = 9.8,
6.3 Hz, 1H, OCHH), 2.53 (m, 2H, CH2), 2.06 (m, 2H, CH2).
13C NMR (CDCl3) δ: 97.65 (C-1), 70.43, 70.09, 68.93, 67.48
(CH2), 66.91, 62.87 (CH2), 51.69 (CH3), 30.78 (CH2), 24.73
(CH2). FAB-HR-MS calcd. for C39H37O12 [M + H]+ (m/z):
697.2285; found: 697.2330.
3-(Methoxycarbonyl)propyl α-D-mannopyranoside (8)
To a solution of compound 7 (3.6 g) in methanol (60 mL)
K2CO3 (300 mg) was added and the mixture was stirred
overnight then neutralized with Amberlyst 15 resin (H+),
filtered and the filtrate was evaporated to dryness. Column
chromatography (CHCl3–MeOH, 3:1) of the residue gave
3-(Methoxycarbonyl)propyl 2,3,4,6-tetra-O-benzoyl-α-D-
mannopyranoside (7)
A
solution of 2,3,4,6-tetra-O-benzoyl-α-D-mannopy-
ranosyl trichloroacetimidate (5) (4.45 g, 6.0 mmol) and
© 2002 NRC Canada

Furneaux et al.
967
8 (1.03 g, 71% after two debenzoylation treatments):
12.0 Hz, 1H, H-6), 4.50 (dd, J5,6′ = 4.5 Hz, 1H, H-6′ ), 4.42
(m, 1H, H-5), 3.84 (dt, J = 6.6, 6,6, 9.6 Hz, 1H, OCHH),
3.68 (s, 3H, CH3), 3.58 (dt, J = 6.6, 6,6, 9.6 Hz, 1H,
OCHH), 2.36 (t, 2H, CH2), 1.71 (m, 4H, 2 × CH2), 1.46 (m,
2H, CH2). 13C NMR (CDCl3) δ: 98.08 (C-1), 71.02, 70.55,
69.28, 68.83 (CH2), 67.46, 63.37 (CH2), 51.87 (CH3), 34.32
(CH2), 29.46 (CH2), 26.08 (CH2) 25.05 (CH2). FAB- HR-
MS calcd. for C41H41O12 [M + H]+: 725.2598; found:
725.2582.
1
[α]2D0 +54.2 (c 0.59, MeOH). H NMR (CD3OD) δ: 4.74 (d,
J
1,2 = 1.5 Hz, 1H, H-1), 3.68 (s, Me), 2.44 (t, 2H, CH2), 1.91
(m, 2H, CH2). 13C NMR (CD3OD) δ: 176.03 (C=O), 101.97
(C-1), 75.07, 73.04, 72.56, 69.02, 67.92 (CH2), 63.33 (CH2),
52.47 (CH3), 32.19 (CH2), 26.40 (CH2). FAB-HR-MS calcd.
for C11H21O8 [M + H]+: 281.1236; found: 281.1243.
3-Carboxypropyl α-D-mannopyranoside (9)
NaOH (0.75 mL, 3M aq) was added to a solution of the
methyl ester 8 (517 mg, 1.84 mmol) in THF (15 mL) and
water (7 mL), The reaction mixture was stirred for 24 h,
neutralised with Amberlyst 15 resin (H+), filtered, concen-
trated in vacuo and freeze-dried to afford the acid 9
(440 mg, 90%): [α]2D0 +46.1 (c 0.41, water). 1H NMR (D2O)
δ: 4.78 (d, J1,2 = 1.2 Hz, 1H, H-1), 3.46–3.87 (m, 8H), 2.42
(t, 2H, CH2), 1.86 (m, 2H, CH2). 13C NMR (D2O) δ: 178.73
(C=O), 100.11 (C-1), 73.12, 71.00, 70.44, 67.22 (CH2),
67.15, 61.31 (CH2), 31.36 (CH2), 24.55 (CH2). FAB-HR-MS
calcd. for C10H19O8 [M + H]+: 267.1080; found: 267.1084.
5-(Methoxycarbonyl)pentyl α-D-mannopyranoside (12)
Debenzoylation was carried out on compound 11 (5.16 g)
as with the ester 7 to give glycoside 12 (1.735 g, 93%, two
treatments): [α]D20 +44.1 (c 0.76, MeOH). 1H NMR
(CD3OD) δ: 4.74 (d, J1,2 = 1.5 Hz, 1H, H-1), 3.84 (dd, J5,6
=
2.3 Hz, J6,6′ = 11.7 Hz, 1H, H-6), 3.80 (dd, J2,3 = 3.2 Hz,
1H, H-2), 3.68–3.77 (m, 3H, H-3, H-6′, OCHH), 3.67 (s,
3H, CH3), 3.62 (t, J3,4, J4,5 = 9.2 Hz, 1H, H-4), 3.54 (m, 1H,
H-5), 3.43 (dt, J = 6.6, 6,6, 9.6 Hz, 1H, OCHH), 2.35 (t, J =
7.4 Hz, 2H, CH2), 1.64 (m, 4H, 2 × CH2), 1.44 (m, 2H,
CH2). 13C NMR (CD3OD) δ: 176.32 (C=O), 101.98 (C-1),
75.02, 73.09, 72.67, 69.08, 68.73(CH2), 63.36 (CH2), 52.38
(CH3), 35.12 (CH2), 30.62 (CH2), 27.27 (CH2), 26.19 (CH2).
FAB-HR-MS calcd. for C13H25O8 [M + H]+: 309.1549;
found: 309.1540.
3-[Carboxy-2-(1,2-dihexadecanoyl-sn-glycero-3-
phospho)ethanolamido]propyl α-D-mannopyranoside
sodium salt (3)
A mixture of acid 9 (119 mg, 0.45 mmol), DCC (248 mg,
1.5 mmol), N-hydroxysuccinimide (139 mg, 1.5 mmol) in
THF (6.5 mL) – CHCl3 (3 mL) – methanol (6.5 mL) was
stirred at room temperature for 24 h. The solvent was re-
moved and column chromatography (CHCl3–MeOH, 4:1) of
the residue gave the crude amide 10 (115 mg, 71%). FAB-
HR-MS calcd. for C14H12NO10 [M + H]+: 364.1244; found:
364.1225.
To a suspension of 1,2-dihexadecanoyl-sn-glycero-3-
phosphoethanolamine (DPPE, 200 mg, 0.284 mmol) in
CHCl3–MeOH (2:1, 6 mL), water (0.4 mL), and NaHCO3
(120 mg, 1.42 mmol) were added, and the mixture was
stirred at 30°C for 30 min. A solution of 10 (94 mg,
0.27 mmol) in CHCl3–MeOH (2:1, 6 mL) was added and
stirring was continued at 30°C overnight. The solvents were
evaporated and column chromatography (CHCl3–MeOH, 3:1)
of the residue gave glycolipid 3, 125 mg (47%): [α]2D0 +17.1
(c 0.38, CHCl3–MeOH, 3:1). 13C NMR (CDCl3–CD3OD,
3:1) δ: 174.33, 173.93, 173.61 (C=O), 100.09 (C-1),
72.59, 71.39, 71.15, 70.75, 70.50, 66.55, 66.55 (CH2), 64.39
(CH2), 63.80 (CH2), 62.66 (CH2), 61.89 (CH2), 61.18 (CH2),
60.89 (CH2), 57.62 (CH2), 51.60, 40.36 (CH2), 34.26 (CH2),
34.11 (CH2), 31.94 (CH2), 29.70 (CH2), 29.36 (CH2), 29.22
(CH2), 25.34 (CH2), 24.94(CH2), 22.67 (CH2), 13.96 (CH3).
FAB-HR-MS calcd. for C47H89NO15P [M]+: 938.5970;
found: 938.6012.
5-Carboxypentyl α-D-mannopyranoside (13)
Ester hydrolysis of 12 (785 mg, 2.54 mmol) was carried
out as for compound 8 and gave 13 (750 mg, 100%): [α]D20
1
+53.2 (c 0.34, MeOH). H NMR (CD3OD) δ: 4.76 (d, J1,2
=
1.5 Hz, 1H, H-1), 3.33–3.89 (m, 8H), 2.28 (t, 2H, CH2), 1.65
(m, 4H, 2 × CH2), 1.42 (m, 2H, CH2). 13C NMR (CD3OD) δ:
179.48 (C=O), 101.95 (C-1), 74.91, 73.05, 72.65, 69.01,
68.85 (CH2), 63.17 (CH2), 36.42 (CH2), 30.71 (CH2),
27.43(CH2), 26.68 (CH2). FAB-HR-MS calcd. for C12H23O8
[M + H]+: 295.1393; found: 295.1398.
5-(N-Succinimidyloxycarbonyl)pentyl α-D-mannopyrano-
side (14)
Amide formation was effected with acid 13 (148 mg,
0.5 mmol) as for analogue 9 and gave amide 14 (170 mg,
89%). 13C NMR (CD3OD) δ: 101.97 (C-1), 75.00, 73.08,
72.66, 69.09, 68.70 and 63.35 (CH2), 52.73, 32.55 (CH2),
30.53 (CH2), 27.12 (CH2), 26.92 (CH2), 25.94 (CH2).
FAB-HR-MS calcd. for C16H26NO10 [M + H]+: 392.1557;
found: 392.1542.
5-[Carboxy-2-(1,2-dihexadecanoyl-sn-glycero-3-
phospho)ethanolamido]pentyl α-D-mannopyranoside
sodium salt (4)
5-(Methoxycarbonyl)pentyl 2,3,4,6-tetra-O-benzoyl-α-D-
mannopyranoside (11)
The title glycolipid (170 mg, 87%) was made from amide
14 (77 mg, 0.19 mmol) in like manner to its analogue 3:
[α]D20 +13.1 (c 0.58, CHCl3–MeOH, 2:1). 13C NMR (CDCl3–
CD3OD) δ: 101.94, 74.76, 73.12, 72.63, 69.05, 68.82 (CH2),
63.34 (CH2), 52.62, 50.11, 35.44 (CH2), 35.25 (CH2), 33.51
(CH2), 31.20 (CH2), 31.01 (CH2), 30.92 (CH2), 30.81 (CH2),
30.66 (CH2), 27.22 (CH2), 26.50 (CH2), 24.19 (CH2), 15.09
(CH3). FAB-HR-MS calcd. for C49H93NNaO15P [M]+:
989.6180; found: 989.6157.
Glycosylation of methyl 6-hydroxyhexanoate (11) (0.94 g,
6.4 mmol) was carried out with the glycosylating agent
(4.45 g, 6.0 mmol) used in the preparation of compound 7
and gave 5.16 g of crude title compound contaminated with
6-hydroxyhexanoate. A chromatographically purified sample
had: 1H NMR (CDCl3) δ: 6.10 (t, J3,4, J4,5 = 10 Hz, 1H, H-4),
5.92 (dd, J2,3 = 3.3 Hz, 1H, H-3), 5.69 (dd, J1,2 = 1.8 Hz,
1H, H-2), 5.08 (d, 1H, H-1), 4.69 (dd, J5,6 = 2.4 Hz, J6,6′
=
© 2002 NRC Canada

968
Can. J. Chem. Vol. 80, 2002
CH2). 13C NMR (125 MHz, CDCl3) δ: 100.68 (C-1′), 100.43
(C-1′′), 98.53 (C-1′′′), 97.14 (C-1), 79.61 (C-3), 76.52 (C-4),
72.29 (C-2), 71.86 (C-2′′); 71.66 (C-5), 71.33 (C-2′), 70.78
(C-2′′′), 70.48 (C-3′′′), 70.27, 70.16, 69.56 (C-5′, C-5′′,
C-5′′′), 69.77, 69.73 (C-3′, C-3′′); 68.80 (CH2), 68.80,
67.25, 67.24 (C-4′, C-4′′, C-4′′′), 66.97 (C-6), 63.49, 63.08,
62.95 (C-6′, C-6′′, C-6′′′), 51.95 (CH3), 31.11 (CH2), 25.12
(CH2), 21.38 (CH3CO). FAB-HR-MS calcd. for
3-(Methoxycarbonyl)propyl 2,4-di-O-acetyl-(2,3,4,6-tetra-
O-benzoyl-α-D-mannopyranosyl)-(1→3)-[(2,3,4,6-tetra-O-
benzoyl-α-D-mannopyranosyl)-(1→6)]-α-D-mannopy-
ranoside (16) and 3-(methoxycarbonyl)propyl 2-O-acetyl-
(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-(1→3)-
[(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-(1→4)]-
{[(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-(1→6)]}-
α-D-mannopyranoside (17)
A solution of glycoside 8 (832 mg, 3.0 mmol) in CH2Cl2
(50 mL) was stirred at room temperature over molecular
sieves (4 Å, 500 mg, finely ground) for 30 min. The solution
was cooled to −40°C and TmsOTf (200 µL) was added, fol-
lowed by 2,3,4,6-tri-O-benzoyl-α-D-mannopyranosyl tri-
chloroacetimidate (4.41 g, 5.95 mmol) in CH2Cl2 (30 mL)
dropwise over 30 min. The solution was stirred for further
30 min, treated with Et3N (0.5 mL), and evaporated to dry-
ness. Column chromatography (hexane–EtOAc, 1.5:1, then
1:1) afforded a mixture of trisaccharide (15) (FAB-HR-MS
calcd. for C79H72O26Cs [M + Cs]+: 1569.3366; found
1569.3367) and tetrasaccharide derivatives (2.863 g).
Acetylation of this crude mixture under standard conditions
followed by column chromatography (hexane–EtOAc, 1:1)
gave the following title compounds.
C
115H100O36Cs [M + Cs]+: 2189.5049; found: 2189.5031.
3-(Methoxycarbonyl)propyl (α-D-mannopyranosyl)-(1→3)-
[(α-D-mannopyranosyl)-(1→6)]-α-D-mannopyranoside (18)
To a solution of the perester 16 (1.050 g, 0.65 mmol) in
MeOH (30 mL) was added NaOMe (8 mmol, 1 M in
MeOH), and the mixture was stirred at room temperature for
24 h, taken to pH ~ 5 with Amberlyst 15 (H+), filtered, and
evaporated to dryness to afford the title trisaccharide
glycoside 18 (420 mg, 100%): [α]2D0 +74.7 (c 0.38, H2O).
1H NMR (D2O) δ: 5.07 (d, J1,2 = 1.4 Hz, 1H, H-1), 4.86 (d,
J1,2 = 1.5 Hz, 1H, H-1), 4.77 (d, J1,2 = 1.4 Hz, 1H, H-1),
3.43–4.03 (m, 26H), 2.46 (t, 2H, CH2), 1.93 (m, 2H, CH2).
13C NMR (D2O) δ: 177.24 (C=O), 102.77, 100.32, 99.83
(C-1), 79.00, 73.74, 73.11, 71.51, 71.03, 70.81, 70.49, 70.39,
70.08, 67.35 (CH2), 67.16, 66.12, 65.74 (CH2), 61.38 (2 ×
CH2), 52.65 (CH3), 31.19 (CH2) and 24.54 (CH2). FAB-HR-
MS calcd. for C23H41O18 [M + H]+: 605.2293; found: 605.2321.
16:
1
(1.055 g, 23%): ([α]D20 –21.3 (c 0.65, CHCl3). H NMR
(500 MHz, CDCl3) δ: 6.18 (t, J3′′,4′′, J4′′,5′′ = 10 Hz, 1H, H-4′′),
6.13 (t, J3′,4′, J4′,5′ = 10 Hz, 1H, H-4′), 5.95 (dd, J2′,3′ = 3.3 Hz,
1H, H-3′), 5.82 (dd, J2′′,3′′ = 3.2 Hz, 1H, H-3′′), 5.75 (dd,
1H, H-2′), 5.53 (dd, 1H, H-2′′), 5.44 (m, 2H, H-2,4), 5.37
(d, J1′′,2′′ = 1.7 Hz, 1H, H-1′′), 5.16 (d, J1′,2′ = 1.7 Hz, 1H,
H-1′), 4.88 (d, J1,2 = 1.7 Hz, 1H, H-1), 4.71 (m, 2H, H-6′a,
H-6′′a), 4.59 (m, 2H, H-5′, H-5′′), 4.52 (m, 2H, H-6′b,
H-6′′b), 4.37 (dd, J2,3 = 3.4 Hz, J3,4 = 9.8 Hz, 1H, H-3), 4.02
(m, 2H, H-5,6a), 3.89 (dt, J = 6.6, 6,6, 9.6 Hz, 1H, OCHH),
3.69 (m, 1H, H-6b), 3.63 (s, 3H, CH3), 3.57 (dt, J = 6.6, 6,6,
9.6 Hz, 1H, OCHH), 2.45 (m, 2H, CH2), 2.32, 2.27 (2s, 2 ×
3 H, OAc), 2.00 (m, 2H, CH2). 13C NMR (125 MHz,
CDCl3) δ: 99.46 (C-1′′), 97.87 (C-1′), 97.80 (C-1), 75.39
(C-3), 71.39 (C-2), 71.19 (C-2′′), 70.86 (C-2′), 70.39 (C-3′),
70.10, 70.07 (C-5, C-5′′), 69.78 (C-3′′), 69.41 (C-5′), 68.89
(C-4), 67.70 (OCH2), 67.29 (C-6, C-4′), 66.93 (C-4′′), 63.35,
63.02 (C-6′, C-6′′), 51.95 (CH3), 31.19 (CH2), 25.08 (CH2),
21.41 (CH3CO), 21.22 (CH3CO). FAB-HR-MS calcd. for
C83H76O28Cs [M + Cs]+: 1653.3577; found: 1653.3583.
3-Carboxypropyl (α-D-mannopyranosyl)-(1→3)-[(α-D-
mannopyranosyl)-(1→6)]-α-D-mannopyranoside (19)
To a solution of glycoside 18 (121 mg, 0.20 mmol) in
H2O (3 mL), NaOH (0.5 mL, 3 M, aq) was added. The reac-
tion mixture was stirred for 2 days, neutralized with
Amberlyst 15 resin (H+), filtered, and freeze-dried to afford
the acid 19 (120 mg, 101%): [α]2D0 +85.5 (c 0.31, MeOH).
13C NMR (CD3OD) δ: 170.34 (C=O), 104.27, 102.17,
101.81 (C-1), 81.13, 75.32, 74.76, 73.90, 73.06, 72.89, 72.48
(2×CH), 71.81, 69.22, 69.01, 68.17 (CH2), 67.89, 67.68
(CH2), 63.29 (2 × CH2), 32.27 (CH2), 26.44 (CH2). FAB-
HR-MS calcd. for C22H39O18 [M + H]+: 591.2136; found:
591.2159.
3-(N-Succinimidyloxycarbonyl)propyl (α-D-mannopyra-
nosyl)-(1→3)-[(α-D-mannopyranosyl)-(1→6)]-α-D-manno-
pyranoside (20)
By use of the procedure described for making amide 10,
and using CHCl3 – MeOH – H2O (5:4:1) as eluent for col-
umn chromatography, acid 19 (115 mg, 0.19 mmol) was
converted to amide 20 (85 mg, 64%): [α]2D0 +87.8 (c 0.23,
MeOH). 13C NMR (CDCl3) δ: 104.36, 102.28 and 101.81
(C-1, C-1′, C-1′′), 81.13, 75.32, 74.76, 73.90, 73.06, 72.89,
72.48, 69.22, 69.01, 67.89, 67.68 (2×CH2), 63.29 (2×CH2),
32.27 (CH2), 30.16, (CH2), 28.69 (CH2), 26.17 (CH2). FAB-
HR-MS calcd. for C26H42NO20 [M + H]+: 688.2300; found:
688.2317.
17:
(1.670 g, 27%, containing small proportions of other
1
oligosaccharide derivatives). H NMR (500 MHz, CDCl3) δ:
6.21 (t, J3′′,4′′, J4′′,5′′ = 10.1 Hz, 1H, H-4′′), 6.12, 6.10 (dt,
2H, H-4′, H-4′′′), 6.04 (m, 2H, H-2′, H-3′), 5.95 (dd, J2′′,3′′
2.7 Hz, 1H, H-3′′), 5.91 (dd, J2 ′′′,3 ′′′ = 3.2 Hz, J3 ′′′,4 ′′′
=
=
10.2 Hz, 1H, H-3 ′′′), 5.88 (m, 1H, H-2′′), 5.79 (m, 1H,
H-2′′′), 5.68 (d, J1′′,2′′ = 1.7 Hz, 1H, H-1′′), 5.51 (d, J1′,2′
=
1.5 Hz, 1H, H-1′), 5.34 (dd, J2,3 = 3.0 Hz, 1H, H-2), 5.26 (d,
J1 ′′′,2 ′′′ = 1.7 Hz, 1H, H-1′′′), 4.83 (d, J1,2 = 1.7 Hz, 1H, H-1),
4.43–4.73 (m, 9H, H-5′, 6′a, 6′b, H-5 ′′, 6′′a, 6′′b, H-5 ′′′,
6 ′′′a, 6′′′b), 4.38 (m, 2H, H-3, H-4), 4.23 (dd, J5,6a = 5 Hz,
3-[Carboxy-2-(1,2-dihexadecanoyl-sn-glycero-3-
phospho)ethanolamido]propyl (α-D-mannopyranosyl)-
(1→3)-[(α-D-mannopyranosyl)-(1→6)]-α-D-
mannopyranoside sodium salt (5)
J6a,6b = 12 Hz, 1H, H-6a), 4.18 (dd, J5,6b = 1.5 Hz, 1H, H-6b),
4.00 (m, 1H, H-5), 3.82, 3.48 (2m, 2H, OCH2CH2), 3.58 (s,
3H, CH3), 2.38 (m, 2H, CH2), 2.18 (s, 3H, Ac), 1.92 (m, 2H,
From amide 20 (83 mg, 0.12 mmol), according to the pro-
cedure described for making glycolipid 3, the analogue 5
© 2002 NRC Canada

Furneaux et al.
969
(105 mg, 70%, containing small proportions of 1,2-hexa-
decanoyl-sn-glycero-3-phosphoethanolamine) was obtained
following column chromatography with CHCl3–MeOH–H2O
(5:4:1) as eluent: [α]D20 +26.4 (c 0.25, CHCl3–MeOH, 3:1).
13C NMR (CDCl3) δ: 106.47, 104.85 and 104.04 (C-1, C-1′,
C-1′′), 83.33, 77.55, 76.92, 75.90, 75.30, 75.00, 74.65,
71.32, 71.10, 70.17, 69.78, 68.63, 68.04, 66.85, 66.01,
65.27, 44.72, 38.48, 38.33, 36.92, 36.15, 33.93, 33.58,
33.40, 29.53, 29.14, 26.88 and 18.17. FAB-HR-MS calcd.
for C59H110NNaO25P [M]+: 1286.7002; found: 1286.6958.
2.9 Hz, H-3′′), 5.99 (dd, J2 ′′′,3 ′′′ = 3.4 Hz, J3 ′′′,4 ′′′ = 10.0 Hz,
1H, H-3′′′), 5.96 (m, 1H, H-2′′), 5.86 (dd, 1H, J1 ′′′,2 ′′′
=
1.7 Hz, H-2′′′), 5.76 (d, J1′′,2′′ = 1.7 Hz, 1H, H-1′′), 5.59 (d,
J1′,2′ = 1.5 Hz, 1H, H-1′), 5.43 (dd, J2,3 = 2.9 Hz, 1H, H-2),
5.35 (d, 1H, H-1′′′), 4.90 (d, J1,2 = 1.7 Hz, 1H, H-1), 4.52–
4.82 (m, 9H, H-5′, 6′a, 6′b, H-5′′, 6′′a, 6′′b, H-5′′′, 6′′′a,
6′′′b), 4.50 (dd, 1H, H-3), 4.42 (t, J = 9.8 Hz, 1H, H-4),
4.31 (dd, J5,6a = 5.0 Hz, J6a,6b = 11.8 Hz, 1H, H-6a), 4.17
(dd, J5,6b = 1.5 Hz, 1H, H-6b), 4.07 (m, 1H, H-5), 3.81, 3.47
(2m, 2H, CH2), 3.61 (s, 3H, CH3), 2.34 (m, 2H, CH2), 2.27
(s, 3H, Ac), 1.69 (m, 4H, 2 × CH2), 1.45 (m, 2H, CH2).
13C NMR (125 MHz, CDCl3) δ: 100.68 (C-1′), 100.46 (C-1′′),
98.53 (C-1′′′), 97.10 (C-1), 79.77 (C-3), 76.57 (C-4), 72.45
(C-2), 71.89 (C-2′′), 71.57 (C-5), 71.37 (C-2′), 70.76 (C-2′′′),
70.51 (C-3′′′), 70.30, 70.26, 69.79, 69.76 and 69.53 (C-3′,
C-3′′, C-5′, C-5′′, C-5′′′), 68.65 (CH2), 67.84, 67.44, 67.28
(C-4′, C-4′′, C-4′′′), 67.02 (C-6), 63.52, 63.10, 62.97 (C-6′,
C-6′′, C-6′′′), 51.77 (CH3), 34.33 (CH2), 29.49 (CH2), 26.12
(CH2), 25.14 (CH2), 21.43 (CH3CO). FAB-HR-MS calcd.
for C117H104O36Cs [M + Cs]+: 2217.5362; found: 2217.5339.
5-(Methoxycarbonyl)pentyl 2,4-di-O-acetyl-(2,3,4,6-tetra-
O-benzoyl-α-D-mannopyranosyl)-(1→3)-[(2,3,4,6-tetra-O-
benzoyl-α-D-mannopyranosyl)-(1→6)]-α-D-mannopyrano-
side (22) and 5-(methoxycarbonyl)pentyl 2-O-acetyl-
(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-(1→3)-
[(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-(1→4)]-
{[(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-(1→6)]}-
α-D-mannopyranoside (23)
A solution of glycoside 12 (917 mg, 2.97 mmol) in
CH2Cl2 (50 mL) was mannosylated as for the analogue 8,
and column chromatography (hexane–EtOAc, 1.5:1, then
1:1) afforded two main fractions (692 mg) and (2.73 g) (in
order of elution), both as mixtures of products mainly con-
taining trisaccharide derivative 21 and a tetrasaccharide ana-
logue. Acetylation of the second fraction under standard
conditions followed by column chromatography (hexane–
EtOAc, 1:1) gave the trisaccharide glycoside perester 22 and
23. The latter contained small proportions of impurities —
probably tri- and tetra-saccharide isomers of the main products.
5-(Methoxycarbonyl)pentyl (α-D-mannopyranosyl)-(1→3)-
[(α-D-mannopyranosyl)-(1→6)]-α-D-mannopyranoside (24)
To a solution of diacetate 22 (1.502 g, 0.97 mmol) in
MeOH (30 mL) was added NaOMe (X M in MeOH, 12 mL)
and the mixture was stirred at room temperature for 24 h,
neutralized to pH ~ 5 with Amberlyst 15 (H+), filtered, and
evaporated to dryness to afford the de-esterified glycoside 24
1
(575 mg, 94%): [α]D20 +79.7 (c 0.33, H2O). H NMR (D2O)
δ: 5.09 (d, J1,2 = 1.4 Hz, 1H, H-1′), 4.88 (d, J1,2 = 1.4 Hz,
1H, H-1′′), 4.80 (d, J1,2 = 1.4 Hz, 1H, H-1), 3.67 (s, 3H,
CH3), 3.50–4.07 (m, 7H, H-2,3,4,5,6, OCH2CH2), 2.39 (t,
J = 7.4 Hz, 2H, CH2), 1.61 (m, 4H, 2 × CH2), 1.37 (m, 2H,
CH2). 13C NMR (D2O) δ: 177.98, 102.76, 100.29, 99.82
(C-1, C-1′, C-1′′), 79.00, 73.74, 73.12, 71.51, 71.06, 70.79,
70.42, 70.41, 70.13, 68.24 (CH2), 67.18, 67.17, 66.21, 65.79
(CH2), 61.39 (2 CH2), 52.53 (CH3), 34.02 (CH2), 28.54
(CH2), 25.40 (CH2), 24.40 (CH2). FAB-HR-MS calcd. for
C25H45O18 [M + H]+: 633.2606; found: 633.2630.
22:
(1.543 g, 33% from 12): [α]2D0 –17.7 (c 0.52, CHCl3).
1H NMR (500 MHz, CDCl3) δ: 6.18 (t, J3 ′′,4 ′′, J4 ′′,5 ′′
=
10.0 Hz, 1H, H-4′′), 6.12 (t, J3′,4′, J4′,5′ = 10.0 Hz, 1H, H-4′),
5.94 (dd, J2′,3′ = 3.4 Hz, 1H, H-3′), 5.81 (dd, J2 ′′,3 ′′
=
3.2 Hz, 1H, H-3′′), 5.74 (dd, J1′,2′ = 1.7 Hz, 1H, H-2′), 5.52
(dd, J1 ′′,2 ′′ = 1.7 Hz, 1H, H-2′′), 5.43 (m, 2H, H-2,4), 5.37
(d, 1H, H-1′′), 5.16 (d, 1H, H-1′), 4.87 (d, J1,2 = 1.5 Hz, 1H,
H-1), 4.71 (dd, J5′,6′a = 2.3 Hz, J6′a,6′b = 12.1 Hz, 1H, H-6′a),
4.68 (dd, J5′′,6′′a = 2.3 Hz, J6 ′′a,6 ′′b = 12.3 Hz, 1H, H-6′′a),
4.58 (m, 2H, H-5′, H-5′′), 4.53 (dd, J5 ′′,6 ′′b = 3.7 Hz, 1H,
H-6′′b), 4.51 (dd, J5 ′,6′b = 3.8 Hz, 1H, H-6′b), 4.38 (dd,
J2,3 = 3.4, J3,4 = 9.8 Hz, 1H, H-3), 4.01 (m, 2H, H-5,6a),
3.84 (dt, J = 6.6, 6,6, 9.6 Hz, 1H, OCHH), 3.69 (m, 1H, H-
6b), 3.59 (s, 3H, CH3), 3.50 (dt, J = 6.6, 6,6, 9.6 Hz, 1H,
OCHH), 2.29 (m, 8H, CH2, 2 × Ac), 1.66 (m, 4H, 2 × CH2),
1.42 (m, 2H, CH2). 13C NMR (125 MHz, CDCl3) δ: 99.36
(C-1′′), 97.71, 97.67 (C-1, C-1′), 75.44 (C-3), 71.41 (C-2),
71.10 (C-2′′), 70.74 (C-2′), 70.30 (C-3′), 69.95 (C-5, C-
5′′), 69.71 (C-3′′), 69.26 (C-5′), 68.78 (C-4), 68.33 (CH2),
67.24, 67.21 (C-4′, C-6), 66.82 (C-4′′), 63.25 (C-6′), 62.92
(C-6′′), 51.65 CH3), 34.18 (CH2), 29.29 (CH2), 26.04 (CH2),
24.99 (CH2), 21.35 (CH3CO), 21.13 (CH3CO). FAB-HR-MS
calcd. for C85H80O28Cs [M + Cs]+: 1681.3890; found:
1681.3899.
5-Carboxypentyl (α-D-mannopyranosyl)-(1→3)-[(α-D-
mannopyranosyl)-(1→6)]-α-D-mannopyranoside (25)
Acid 25 (774 mg, 96%) was obtained from methyl ester
24 (821 mg, 1.30 mmol) by use of the procedure applied to
make analogue 9: [α]2D0 +87.3 (c 0.75, H2O). 13C NMR
(D2O) δ: 179.56 (C=O), 102.71, 100.26, 99.78 (C-1, C′-1,
C′′-1), 79.00, 73.68, 73.06, 71.47, 71.05, 70.79, 70.47,
70.40, 70.13, 68.20 (CH2), 67.14, 66.20, 65.77 (CH2), 61.37
(2 CH2), 34.37 (CH2), 28.57 (CH2), 25.41 (CH2), 24.51
(CH2). FAB-HR-MS calcd. for C24H43O18 [M + H]+:
619.2449; found: 619.2468.
5-[Carboxy-2-(1,2-dihexadecanoyl-sn-glycero-3-
phospho)ethanolamido]pentyl (α-D-mannopyranosyl)-
(1→3)-[(α-D-mannopyranosyl)-(1→6)]-α-D-
mannopyranoside sodium salt (6)
Fom acid 25 (125 mg, 0.202 mmol), following the proce-
dure described to make amide 14, crude succinimidyl deriva-
tive (26) was obtained, and was used without further
purification, following the method described for making the
glycolipid 5. The sodium salt 6 (132 mg, 50% from acid 25)
23:
1
(1.033 g, 17% from 12). H NMR (500 MHz, CDCl3) δ:
6.27 (t, J3 ′′,4 ′′, J4 ′′,5 ′′ = 10.1 Hz, 1H, H-4′′), 6.17 (2t, 2H,
H-4′, H-4′′′ ), 6.12 (m, 2H, H-2′, H-3′), 6.07 (dd, 1H, J2′′,3′′
=
© 2002 NRC Canada

970
Can. J. Chem. Vol. 80, 2002
was isolated after column chromatography on silica gel with
CHCl3–MeOH–H2O (5:4:1) as eluent: [α]2D0 +37.0 (c 0.6,
CHCl3–MeOH, 2:1). 13C NMR (CDCl3–CD3OD) δ: 102.71,
100.26 and 99.78 (C-1, C-1′, C-1′′), 79.00, 73.68, 73.06,
71.47, 71.05, 70.79, 70.47, 68.20, 67.14, 66.20, 63.87 (CH2),
63.35 (CH2), 62.38 (CH2), 61.35 (CH2), 40.03 (CH2), 35.84
(CH2), 33.93 (CH2), 33.78 (CH2), 33.42 (CH2), 31.67 (CH2),
29.40 (CH2), 29.08 (CH2), 28.86 (CH2), 25.69 (CH2), 25.25
(CH2), 24.65 (CH2), 22.36 (CH2), 13.41 (CH3). FAB-HR-
MS calcd. for C61H113NNa2O25P [M + Na]+: 1336.7135;
found: 1336.7076.
tralized with Amberlyst 15 (H+) resin, filtered, and freeze-
dried to afford acid 30 (81 mg, 100%): [α]2D0 +38.0 (c 0.41,
H2O). 13C NMR (D2O) δ: 176.89 (C=O), 100.37 (C-1),
73.16, 72.14 (CH2), 70.92, 70.37, 69.98 (CH2), 67.18, 66.87
(CH2), 61.35 (CH2), 60.81 (CH2) 57.80 (CH2), 37.34 (CH2).
FAB-HR-MS calcd. for C13H25O10 [M + H]+: 341.1448;
found: 341.1449.
2-[2-(2-Azidoethoxy)ethoxy]ethyl 2,3,4,6-tetra-O-benzoyl-
α-D-mannopyranoside (31)
A mixture of chloro-compound 27 (2.785 g, 3.72 mmol),
sodium azide (2.6 g 40 mmol), and DMF (40 mL) was
stirred under argon at 65°C for 19 h. Water (60 mL) was
added, and the mixture was extracted with EtOAc (3 ×
30 mL). The combined organic extracts were washed with
water (10 mL), dried (Na2SO4), and evaporated to dryness.
Column chromatography (hexane–EtOAc, 1:1) of the residue
2-[2-(2-Chloroethoxy)ethoxy]ethyl 2,3,4,6-tetra-O-benzoyl-
α-D-mannopyranoside (27)
2-[2-(2-Chloroethoxy)ethoxy]ethanol (1.00 g, 5.95 mmol)
was glycosylated according to the procedure described for
making glycoside 7, to give product 27 (4.07 g, 91%): [α]D20
−48.2 (c 0.57, CHCl3). H NMR (CDCl3) δ: 6.11 (t, J3,4
1
,
gave of azide 31 (2.655 g, 95%): [α]2D0 −43.7 (c 0.6, CHCl3).
J4,5 = 10.1 Hz, 1H, H-4), 5.94 (dd, J2,3 = 3.3 Hz, 1H, H-3),
5.73 (dd, J1,2 = 1.8 Hz, 1H, H-2), 5.16 (d, 1H, H-1), 4.70 (m,
1H, H-6), 4.50 (m, 2H, H-5,6′), 4.00–3.61 (m, 12H, spacer
protons). 13C NMR (CDCl3) δ: 98.22 (C-1), 71.83 (CH2),
71.20 (CH2), 71.14 (CH2), 70.90, 70.61 (CH2), 70.47, 69.25,
68.05 (CH2), 67.39, 63.27 (CH2), 43.18 (CH2). FAB-HR-MS
calcd. for C40H40ClO12 [M + H]+: 747.2208; found: 747.2184.
1
IR (film) (cm–1): 2105. H NMR (CDCl3) δ: 6.11 (t, J3,4
,
J4,5 = 10.0 Hz, 1H, H-4), 5.95 (dd, J2,3 = 3.3 Hz, 1H, H-3),
5.73 (dd, J1,2 = 1.8 Hz, 1H, H-2), 5.16 (d, 1H, H-1), 4.70 (m,
1H, H-6), 4.50 (m, 2H, H-5,6′), 4.0–3.6 (m, 10H, spacer H),
3.38 (t, J = 5.0 Hz, 2H, CH2N3). 13C NMR (CDCl3) δ: 98.23
(C-1), 71.28 (CH2), 71.15 (CH2), 70.90, 70.63 (CH2), 70.50,
69.25, 68.06 (CH2), 67.40, 63.26 (CH2), 51.10 (CH2), 14.57
(CH2). FAB-HR-MS calcd. for C40H39N3O12Cs [M + Cs]+:
886.1588; found: 886.1538.
2-[2-(2-Cyanoethoxy)ethoxy]ethyl 2,3,4,6-tetra-O-benzoyl-
α-D-mannopyranoside (28)
A mixture of chloro-compound 27 (3.98 g, 5.33 mmol),
potassium cyanide (3.26 g 50 mmol), and DMSO (40 mL)
was stirred under argon at 65°C for 22 h. Water (100 mL)
was added, and the mixture was extracted with EtOAc (4 ×
50 mL). The combined organic extracts were washed with
H2O (20 mL), dried (Na2SO4), and evaporated to dryness.
Column chromatography (hexane–EtOAc, 1:1) of the residue
gave the nitrile 28 (2.77 g, 70%): [α]2D0 –37.2 (c 0.78,
CHCl3). IR (film) (cm–1): 2253. 1H NMR (CDCl3) δ: 6.11 (t,
2-{2-[2-(5-Methoxycarbonylpentanoylamino)ethoxy]etho-
xy}ethyl α-D-mannopyranoside (34)
To a solution of azide 31 (1.30 g, 1.72 mmol) in EtOH
(25 mL), NaBH4 (325 mg, 8.5 mmol) was added followed
by a solution of NiCl2 in EtOH (1.5 mL, 0.2 M), and the
mixture was stirred at room temperature for 45 min, filtered
through a pad of silica gel (thickness ~ 5 cm) and washed
with CHCl3–MeOH (2:1). The filtrate and washings were
taken to dryness and purified by column chromatography
(CHCl3–MeOH, 3:1) to give the amine 32 (660 mg, 53%).
13C NMR (CD3OD) δ: 99.53 (C-1), 72.15, 71.99 (CH2),
71.90, 71.64 (CH2), 71.49 (CH2), 70.50, 68.91 (CH2), 68.41,
68.33 (CH2), 64.00 (CH2), 41.05 (CH2). A solution of amine
32 (200 mg, 0.27 mmol), monomethyl adipate (48 mg,
0.3 mmol), and 2-ethoxy-1-ethoxycarbonyl-1,2-dihydro-
quinoline (12) (74 mg, 0.3 mmol) in EtOH (10 mL) was
heated under reflux for 3 h and taken to dryness in vacuo.
Column chromatography (CHCl3–MeOH, 50:1) of the resi-
due gave amide 33 (244 mg, 100%). 13C NMR (CDCl3) δ:
98.23 (C-1), 71.04, 70.90 (CH2), 70.72 (CH2), 70.55, 70.49
(CH2), 70.33 (CH2), 69.28, 68.06 (CH2), 67.35, 63.26 (CH2),
51.84 (CH3), 39.75 (CH2), 36.51 (CH2), 34.11 (CH2), 25.46
(CH2), 24.81 (CH2). A mixture of amide 33 (1.615 g,
18.6 mmol) and K2CO3 (200 mg) in MeOH (20 mL) was
stirred overnight, neutralized with Amberlyst 15 (H+), fil-
tered, and taken to dryness in vacuo. Column chromatogra-
phy (CHCl3–MeOH, 3:1) of the residue gave the de-
esterified glycoside 34 (530 mg, 63%): [α]2D0 +32.1 (c 0.57,
MeOH). 13C NMR (CD3OD) δ: 102.17 (C-1), 75.03, 73.01,
72.54, 72.02 (CH2), 71.82 (CH2), 71.71 (CH2), 71.06 (CH2),
69.07, 68.13 (CH2), 63.40 (CH2), 52.43 (CH3), 40.79 (CH2),
37.00 (CH2), 34.86 (CH2), 26.77 (CH2), 25.91 (CH2). FAB-
J
3,4, J4,5 = 10.1 Hz, 1H, H-4), 5.93 (dd, J2,3 = 3.3 Hz, 1H,
H-3), 5.72 (dd, J1,2 = 1.8 Hz, 1H, H-2), 5.16 (d, 1H, H-1),
4.70 (m, 1H, H-6), 4.50 (m, 2H, H-5,6′), 4.00–3.70 (m, 12H,
spacer H), 2.63 (t, J = 6.4 Hz, 2H, CH2CN). 13C NMR
(CDCl3) δ: 118.34 (CN), 98.23 (C-1), 71.19 (2 × CH2),
70.88, 70.64 (CH2), 70.47, 69.28, 68.02 (CH2), 67.39, 66.44
(CH2), 63.26 (CH2), 19.28 (CH2). FAB-HR-MS calcd. for
C41H40NO12 [M + H]+: 738.2550; found: 738.2573.
2-[2-(2-Cyanoethoxy)ethoxy]ethyl α-D-mannopyranoside
(29)
De-esterification of tetrabenzoate 28 (320 mg,
0.43 mmol), following the procedure described for making
glycoside 8, gave glycoside 29 (128 mg, 92%): [α]2D0 +47.0
(c 0.61, MeOH). IR (film) (cm–1): 2252.; 13C NMR
(CD3OD) δ: 120.16 (CN), 102.13 (C-1), 74.99, 72.99, 72.51,
72.02 (CH2), 71.95 (CH2), 71.85 (CH2), 69.03, 68.18 (CH2),
67.51 (CH2), 63.35 (CH2), 19.66 (CH2). FAB-HR-MS calcd.
for C13H24NO8 [M + H]+: 322.1502; found: 322.1500.
2-[2-(2-α-D-Carboxyethoxy)ethoxy]ethyl α-D-mannopyra-
noside (30)
A mixture of nitrile 29 (79 mg, 0.24 mmol) in NaOH
(1 mL, 3 M) was stirred at room temperature for 24 h, neu-
© 2002 NRC Canada

Furneaux et al.
971
HR-MS calcd. for C19H36NO11 [M + H]+: 454.2288; found:
454.2326.
FAB-HR-MS calcd. for C25H44NO18 [M + H]+: 646.2558;
found: 646.2560.
2-[2-(2-Carboxyethoxy)ethoxy]ethyl (α-D-mannopyran-
osyl)-(1→3)-[(α-D-mannopyranosyl)-(1→6)]-α-D-manno-
pyranoside (38)
2-[2-(2-Cyanoethoxy)ethoxy]ethyl (2,3,4,6-tetra-O-benzoyl-
α-D-mannopyranosyl)-(1→3)-[(2,3,4,6-tetra-O-benzoyl-α-D-
mannopyranosyl)-(1→6)]-α-D-mannopyranoside (35)
Hydrolysis of the nitrile 37 (90 mg) was carried out in so-
dium hydroxide solution (2 mL, 3 M, aq) over 2 days at
20°C. Water (5 mL) was added and the base was neutralized
to pH ~ 5 with Amberlyst resin (H+). Evaporation of the sol-
vent gave the title acid (95 mg, 102%): [[α]2D0 +70.0 (c 0.43,
H2O). 13C NMR (D2O) δ: 105.31, 103.12, 102.38 (C-1, C-1′,
C-1′′), 81.50, 76,29, 75.66, 74.70 (CH2), 74.03, 73.57,
73.34, 72.93, 72.53 (2 CH2), 69.70 (CH2), 68.65, 68.24
(CH2), 63.95 (2CH2), 63.37 (CH2), 37.92 (CH2). FAB-HR-
MS calcd. for C25H45O20 [M+]: 665.2504; found: 665.2508.
Glycoside 29 (580 mg, 1.8 mmol) and 2,3,4,6-tetra-O-
benzoyl α-D-mannopyranosyl trichloroacetimidate (2.667 g,
3.6 mmol) were treated according to the procedure described
for making compound 21, and gave mixed products
(1.585 g). Column chromatography (hexane–EtOAc, 1:1) af-
forded the title trisaccharide derivative 35 (1.070 g, 40%):
[α]2D0 –24.4 (c 0.36, CHCl3). IR (film) (cm–1): 2252.;
1H NMR (CDCl3) δ: 6.12 (t, J = 10 Hz, 2H, H-4′, H-4′′),
6.00 (m, 2H, H-3′, H-3′′), 5.86 (dd, J1′,2′ = 1.7 Hz, J2′,3′
=
3.0 Hz, 1H, H-2′), 5.79 (dd, J1 ′′,2 ′′ = 1.7 Hz, J2 ′′,3 ′′ = 3.2 Hz,
1H, H-2′′), 5.44 (d, 1H, H-1′′), 5.33 (d, 1H, H-1′), 4.83 (d,
1H, H-1), 3.90–4.95 (m, 12H), 3.60–3.72 (m, 10H, spacer
H), 2.58 (t, J = 6.4 Hz, 2H, CH2CN). 13C NMR (CDCl3) δ:
118.05 (CN), 100.07, 99.75, 97.62 (C-1,1′,1′′), 82.54, 71.41,
70.73, 70.56, 70.22, 69.49, 68.91, 67.50, 66.98, 66.87,
65.90, 63.26, 62.96 (CH2), 18.85 (CH2CN); (other CH2 car-
bon resonances not identified). FAB-HR-MS calcd. for
C81H76NO26 [M + H]+: 1478.4656; found: 1478.4629.
N-[Tris-(2,3,4,6-tetra-O-acetyl-α-D-mannopyranosyl-
oxymethyl)]methyl-(5-methoxycarbonyl)pentanoamide (39)
A mixture of tetra-O-acetyl-α-D-mannopyranosyl bromide
(3.69 g, 8.97 mmol), N-[tris(hydroxymethyl)]methyl-(5-
methoxycarbonyl)pentanoamide (8) (590 mg, 2.24 mmol)
and HgCN2 (2.3 g, 9.1 mmol) in CH3NO2–toluene (1:1 v/v,
60 mL) was heated at 60°C for 3.5 h. The mixture was evap-
orated in vacuo to a syrup, dissolved in CHCl3 (150 mL),
and the organic layer was washed with NaCl solution (3 ×
60 mL, sat., aq), H2O (50 mL), dried (Na2SO4), and evapo-
rated to dryness. Column chromatography (CHCl3–MeOH,
30:1) of the residue gave the title compound (1.555 g, 55%):
2-[2-(2-Cyanoethoxy)ethoxy]ethyl (α-D-mannopyranosyl)-
(1→3)-[(α-D-mannopyranosyl)-(1→6)]-α-D-mannopyra-
noside (37)
Standard acetylation of an analytical sample of the tri-
saccharide derivative 35 yielded 2-[2-(2-cyanoethoxy)ethoxy]ethyl
2,4-di-O-acetyl-(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-
(1→3)-[(2,3,4,6-tetra-O-benzoyl-α-D-mannopyranosyl)-(1→6)]-
α-D-mannopyranoside (36): [α]D20 –20.2 (c 0.54, CHCl3). IR
1
[α]2D0 +30.7 (c 0.88, CHCl3). H NMR (CDCl3) δ: 5.33–5.17
(m, 9H, H-2,3,4), 4.85 (d, J1,2 = 1.3 Hz, 3H, H-1), 4.34 (dd,
J5,5 = 4.9 Hz, J6,6′ = 12.3 Hz, 3H, H-6), 4.13 (m, J5,6′
=
2.4 Hz, H-6′, 6H, OCHHCN), 3.96 (m, 3H, H-5), 3.73 (d,
2JH,H = 10.1 Hz, 3H, OCHHCN), 3.66 (s, 3H, CH3), 2.35 (m,
2H, CH2), 2.11 (m, 2H, CH2), 1.71 (m, 4H, CH2), 2.14, 2.12,
2.04, 1.98 (4s, 36H, Ac). 13C NMR (CDCl3) δ: 103.06 (C-1),
73.84, 73.7, 71.11 (CH2), 70.52, 66.78 (CH2), 63.74 (C),
56.19 (CH3), 41.25 (CH2), 38.15 (CH2), 29.72 (CH2), 28.81
(CH2), 25.31 (CH3CO). FAB-HR-MS calcd. for C53H76NO33
[M + H]+: 1254.4300; found: 1254.4381.
1
(film) (cm–1): 2252. H NMR (500 MHz, CDCl3) δ: 6.19 (t,
J3 ′′,4 ′′, J4 ′′,5 ′′ = 10.1 Hz, 1H, H-4′′), 6.12 (t, J3 ′,4 ′, J4 ′,5 ′
=
10.1 Hz, 1H, H-4′), 5.92 (dd, J2 ′,3 ′ = 3.3 Hz, 1H, H-3′), 5.81
(dd, J2 ′′,3 ′′ = 3.3 Hz, 1H, H-3′′), 5.73 (dd, J1 ′,2 ′ = 1.7 Hz,
1H, H-2′), 5.51 (dd, J1 ′′,2 ′′ = 1.7 Hz, 1H, H-2′′), 5.47 (dd,
J2,1 = 1.7 Hz, J2,3 = 3.4 Hz, 1H, H-2), 5.43 (t, J3,4, J4,5
=
10.0 Hz, 1H, H-4), 5.36 (d, 1H, H-1′′), 5.17 (d, 1H, H-1′),
4.94 (d, 1H, H-1), 4.69–4.75 (m, 2H, H-6′a, H-6′′a), 4.55–
4.63 (m, 2H, H-5′, H-5′′), 4.49 (m, 2H, H-6′b, H-6′′b), 4.40
(dd, 1H, H-3), 4.10 (m, 1H, H-5), 4.01 (dd, J5,6 = 6.8 Hz,
J6a,6b = 10.7 Hz, 1H, H-6a), 3.60–3.73 (m, 11H, H-6b and
spacer H), 2.55 (t, J = 6.5 Hz, 2H, CH2CN). 13C NMR
(125 MHz, CDCl3) δ: 99.40 (C-1′′), 97.87 (C-1), 97.73 (C-1′),
75.33 (C-3), 71.17 (C-2), 71.08 (C-2′′, spacer CH2), 70.90
(spacer CH2), 70.73 (C-2′), 70.49 (C-3′, CH2), 70.02
(C-5′′), 69.85 (C-5), 69.77 (C-3′′), 69.38 (C-5′), 68.90 (C-4),
67.43, 67.30 (C-6, spacer CH2), 67.24 (C-4′), 66.87 (C-4′′),
66.31 (spacer CH2), 63.29 (C-6′), 62.91 (C-6′′), 21.42 (Ac),
21.21 (Ac), 19.16 (CH2CN).
A mixture of perester 35 (895 mg, 0.605 mmol) and
K2CO3 (250 mg) in MeOH (30 mL) was stirred at room tem-
perature for 22 h, neutralized to pH ~ 5 with Amberlyst 15
(H+), filtered, and evaporated to dryness to afford title
compound 37 (230 mg, 59%): [α]2D0 +73.9 (c 0.41, H2O).
13C NMR (D2O) δ: 120.35 (CN), 102.78, 100.58 and 99.85
(C-1,1′,1′′) 79.01, 73.74, 73.11, 71.49, 71.04, 70.81, 70.50,
70.38, 70.09 (CH2), 69.97 (2 × CH2), 67.17 (CH, CH2),
66.10, 65.79 (2 × CH2), 61.40 (2 × CH2), 18.52 (CH2CN).
N-[Tris-(α-D-mannopyranosyloxymethyl)]methyl-(5-
methoxycarbonyl)pentanoamide (40)
To a solution of perester 39 (2.45 g, 1.95 mmol) in metha-
nol (30 mL) was added NaOMe (2 drops, X M in MeOH)
and the mixture was stirred at room temperature overnight.
Removal of the solvent afforded the triglycoside 40 (1.47 g,
100%): [α]2D0 +45.4 (c 2.52, MeOH). 13C NMR (CD3OD) δ:
176.48 and 176.33 (C=O), 102.77 (C-1), 75.24, 73.16,
72.39, 69.22, 67.30 (CH2), 63.22 (CH2), 61.03 (C), 50.29
(CH3), 37.60 (CH2), 34.85 (CH2), 26.92 (CH2), 25.91 (CH2).
FAB-HR-MS calcd. for C29H52NO21 [M + H]+: 750.3032;
found: 750.3067.
N-[Tris-(α-D-mannopyranosyloxymethyl)]methyl-5-
carboxypentanoamide (41)
To a solution of methyl ester 40 (1.47 g, 1.95 mmol) in
THF (30 mL) and H2O (15 mL), sodium hydroxide (1.3 mL,
3 M aq) was added. The mixture was stirred for 48 h, neu-
tralized with Amberlyst 15 resin (H+), filtered, evaporated in
vacuo, and freeze-dried to afford acid 41 (1.45 g, 100%):
© 2002 NRC Canada

972
Can. J. Chem. Vol. 80, 2002
[α]D20 +42.7 (c 0.66, H2O). 13C NMR (D2O) δ: 180.43 and
177.00 (C=O), 100.77 (C-1), 73.42, 71.10, 70.37, 67.16,
65.87 (CH2), 61.25 (CH2), 59.81 (C), 36.52 (CH2), 35.00
(CH2), 25.55 (CH2), 24.61 (CH2). FAB-HR-MS calcd. for
C28H50NO21 [M + H]+: 736.2875; found: 736.2829.
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Acknowledgements
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We thank the New Zealand Foundation for Research, Sci-
ence & Technology for funding, Dr H. Wong for NMR spec-
troscopy, HortResearch, Palmerston North, New Zealand for
mass spectral analyses, and Prof. R.J. Ferrier for assistance
in the preparation of this manuscript.
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© 2002 NRC Canada