Application and limitations of the methyl imidate protection strategy of N-acetylglucosamine for glycosylations at O-4: synthesis of Lewis A and Lewis X trisaccharide analogues

Article abstract of DOI:10.1016/j.carres.2008.08.025

We describe here the synthesis of the allyl Le^a trisaccharide antigen as well as that of an analogue of the Le^x trisaccharide antigen, in which the galactose residue has been replaced by a glucose unit. Although successful fucosylations at O-4 of N-acetylglucosamine acceptors have been reported using perbenzylated thioethyl fucosyl donors under MeOTf activation, such conditions led in our case to the conversion of our acceptor to the corresponding alkyl imidates. Indeed, in this synthesis of the Le^a analogue, we demonstrate that the temporary protection of the N-acetyl group as a methyl imidate is advantageous to fucosylate at O-4. In contrast, we report here that glucosylation at O-4 of an N-acetylglucosamine monosaccharide acceptor using the α-trichloroacetimidate of peracetylated glucopyranose as a donor proceeded in better yields under activation with excess BF₃·OEt₂ than that of the corresponding methyl imidate. Therefore, we conclude that activation of thioglycoside donors by MeOTf to glycosylate at O-4 of a glucosamine acceptor is best accomplished following the temporary protection of the N-acetyl group as a methyl imidate, especially when the donors are highly reactive and prone to degradation. In contrast, if donor and acceptor can withstand multiple equivalents of BF₃·OEt₂, glycosylations at O-4 of a glucosamine acceptor with a trichloroacetimidate donor does not benefit from the temporary protection of the N-acetyl group as a methyl imidate.

Full text of DOI:10.1016/j.carres.2008.08.025

Carbohydrate Research 343 (2008) 2914–2923
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Carbohydrate Research
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Application and limitations of the methyl imidate protection strategy
of N-acetylglucosamine for glycosylations at O-4: synthesis of Lewis
A and Lewis X trisaccharide analogues
*
Jenifer L. Hendel, Anderson Cheng, France-Isabelle Auzanneau
Department of Chemistry, University of Guelph, Guelph, Ontario, Canada N1G 2W1
a r t i c l e i n f o
a b s t r a c t
We describe here the synthesis of the allyl Lea trisaccharide antigen as well as that of an analogue of the
Lex trisaccharide antigen, in which the galactose residue has been replaced by a glucose unit. Although
successful fucosylations at O-4 of N-acetylglucosamine acceptors have been reported using perbenzylat-
ed thioethyl fucosyl donors under MeOTf activation, such conditions led in our case to the conversion of
our acceptor to the corresponding alkyl imidates. Indeed, in this synthesis of the Lea analogue, we dem-
onstrate that the temporary protection of the N-acetyl group as a methyl imidate is advantageous to
fucosylate at O-4. In contrast, we report here that glucosylation at O-4 of an N-acetylglucosamine mono-
Article history:
Received 5 August 2008
Received in revised form 21 August 2008
Accepted 25 August 2008
Available online 31 August 2008
Keywords:
Methyl imidate
Lewis A
saccharide acceptor using the
a-trichloroacetimidate of peracetylated glucopyranose as a donor pro-
ceeded in better yields under activation with excess BF3ꢀOEt2 than that of the corresponding methyl
imidate. Therefore, we conclude that activation of thioglycoside donors by MeOTf to glycosylate at O-4
of a glucosamine acceptor is best accomplished following the temporary protection of the N-acetyl group
as a methyl imidate, especially when the donors are highly reactive and prone to degradation. In contrast,
if donor and acceptor can withstand multiple equivalents of BF3ꢀOEt2, glycosylations at O-4 of a glucos-
amine acceptor with a trichloroacetimidate donor does not benefit from the temporary protection of the
N-acetyl group as a methyl imidate.
Lewis X
N-Acetylglucosamine acceptor
Thioglycoside donor
Tricholoroacetimidate donor
Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction
the synthesis of the allyl Lea trisaccharide 1 that will be subse-
quently conjugated to carrier proteins and used in binding stud-
Our group is involved in the design of new anti-cancer vaccines
based on the Tumor Associated Carbohydrate Antigens (TACAs)
LeaLex and dimeric Lex (dimLex).1,2 These tumor specific antigens
consist of hexasaccharides that display either the Lea or Lex trisac-
charide antigen linked to O-300 of the galactose residue of another
Lex trisaccharide (Chart 1). Thus, employing these hexasaccharides
to raise immune responses against cancer cells is also likely to trig-
ger immune responses against these non-reducing end trisaccha-
ride antigens and will eventually lead to the auto-immune
destruction of the non-cancerous cells that display the natural
Lea or Lex antigens.3
We have therefore embarked on the quest to discover analogues
of these TACAs that would no longer display the Lea or Lex antigens
at their reducing end but that would still be able to trigger the de-
sired anti-LeaLex or anti-dimLex immune responses.1a,2a,2f In that
context, we have reported the synthesis of Lea analogues in which
either one or both the galactosyl and fucosyl residues have been re-
placed by glucose and rhamnose, respectively.4 We describe here
ies.5 In addition, we have also reported the chemical synthesis of
Lex analogues6 in which either one or both the N-acetylglucosami-
nyl and fucosyl residues have been replaced by glucose and rham-
nose, respectively. We report here the synthesis of an additional
Lex analogue (2), in which the galactose residue is replaced by a
glucose unit.
Although our syntheses of the Lex analogues6 have relied on the
glycosylation of lactosyl6a or 2-azido lactosaminyl acceptors,6b and
thus did not entail glycosylation at O-4 of N-acetylglucosamine,
our work on the Lea analogues involved either a rhamosylation
or fucosylation at this position.4 However, glycosylations at O-4
of N-acetylglucosamine are notoriously difficult to carry out suc-
cessfully.4,7,8 The difficulties encountered in such reactions have
been attributed to steric hindrance,7 hydrogen bond network for-
mation involving the amide group and reducing the nucleophilicity
at O-4,8 or to the formation of unwanted stable glycosyl imi-
dates.4a,b,9 More recently we have reported4b that when thioethyl
rhamnoside donors were activated with methyl triflate to promote
glycosylation at O-4 of an N-acetylglucosamine acceptor, the for-
mation of the acceptor methyl imidate could be observed prior
or instead of its glycosylation. Turning this result to our advantage
* Corresponding author. Tel.: +1 519 824 4120x53809; fax: +1 519 766 1499.
E-mail address: fauzanne@uoguelph.ca (F.-I. Auzanneau).
0008-6215/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2008.08.025

J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
2915
HO
OH
OH
OH
O
OH
HO
HO
O
LeaLex R =
O
O
O
OH
HO
O
OH
NHAc
O
OH
O
O
O
O
OH
R
HO
HO
OH
OH
O
NHAc
O
O
O
DimLex
OH
R =
O
OH
OH
OH
HO
NHAc
O
OH
HO
O
HO
OH
OH
OH
O
OH
HO
O
O
O
OH
HO
OMe
OH
HO
HO
OH
OH
O
O
NHAc
O
OAll
O
O
OH
NHAc
OH
HO
2
1
Chart 1.
we have developed10 a new method for temporarily protecting the
amide group of an N-acetylglucosamine acceptor immediately
prior to its glycosylation at O-4. We showed that this temporary
protecting group strategy was compatible with thioglycoside and
trichloroacetimidate glycosyl donors as well as with NIS–TMSOTf
(or TfOH), MeOTf, or TMSOTf activation conditions.10 Although suc-
cessful fucosylations at O-4 of N-acetylglucosamine acceptors have
been reported,4b,11 we demonstrate here that in some cases
employing the temporary protection of the N-acetyl group as a
methyl imidate is advantageous even when attempting to intro-
duce a simple fucosyl unit at O-4. In contrast and while preparing
analogue 2, we describe here that glucosylation at O-4 of an N-ace-
tylglucosamine acceptor can proceed in good yields if using an ex-
cess of BF3ꢀOEt2 as Lewis acid to activate a trichloroacetimidate
glucosyl donor.
Thus, the known12 p-methoxybenzylidene acceptor 3 was galac-
tosylated with the peracetylated bromide13 4 under Helferich acti-
vation and gave disaccharide 5 in 82% yield (Scheme 1).
The p-methoxybenzylidene protecting group was hydrolyzed
off using mild acidic conditions and the resulting diol (6) was sub-
mitted to selective silylation at O-6 using t-butyldiphenyl silyl
chloride (TBDPSCl) and imidazole in acetonitrile to give the mono-
silylated disaccharide 7 in 67% yield. The regioselectivity of the
silylation at O-6 of diol 6 was confirmed via the acetylation of
the remaining free hydroxyl group in compound 7 to give disaccha-
ride 8. 1H NMR spectroscopy of disaccharide 8 showed a signal for
H-4 at 4.86 ppm, while the same signal in the alcohol 7 was found
upfield at approximately 3.8 ppm, thus supporting that O-4 was
free in disaccharide 7 because it became acetylated in disaccharide
8. Having in hand the acceptor 7, we prepared fucosyl donors 10
and 11 to attempt glycosylation at O-4 (Scheme 2).
Silylation of the known14,15 thioglycoside 9 with TBDPSCl and
imidazole gave the thiofucoside donor 10 while its p-methoxyben-
zylation with PMBCl and NaH following literature procedures gave
the known15 donor 11.
2. Results and discussion
Applying the synthetic strategy originally established by Lemi-
eux et al.11a,b and that we have followed to prepare other Lea ana-
logues,4 we decided to prepare the allyl Lea trisaccharide 1 via the
galactosylation at O-3 followed by the fucosylation at O-4 of a suit-
ably protected N-acetylglucosamine allyl glycoside derivative.
It has been established11a-c in previous syntheses of Lea ana-
logues that fucosyl donors could successfully glycosylate at O-4
of the N-acetylglucosamine residue of a disaccharide such as
AcO
AcO
OAc
R2O
AcO
AcO
OAc
PMP
O
O
HO
R1O
O
O
O
O
a
OAll
O
OAll
AcO
NHAc
AcO
NHAc
Br
1
2
5
6
R ,R = >CHPhOMe
3
4
b
1
R
= R2 = H
c
7 R1 = H, R2 = TBDPS
8 R1 = Ac, R2 = TBDPS
d
Scheme 1. Reagents and conditions: (a) 2 equiv 4, 2 equiv Hg(CN)2, toluene, CH3NO2, MS 4 Å, 50 °C, 3 h, 82%; (b) 50% aq AcOH, 60 °C, 1 h, 67%; (c) TBDPSCl 1 equiv, imidazole
6 equiv, CH3CN, rt, 15 min, 67%; (d) Ac2O, pyridine, DMAP, 50 °C, 18 h, 78%.

2916
J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
dates giving m/z peaks at [M+H]+ 844.3576 and 858.3402 for 12a
(calcd 844.3591) and 12b (calcd 858.3732), respectively.
SEt
OR
O
SEt
OH
O
a or b
O
Although we have reported the formation of methyl and ethyl
imidates when treating N-acetylglucosamine O-4 acceptors with
MeOTf in Et2O, we were surprised that no fucosylation took place
during the attempted coupling of acceptor 7 and fucosyl donor
10. We hypothesized that this glycosylation did not give the de-
sired trisaccharide because of steric hindrance resulting from the
presence of the two TBDPS groups on O-2 of fucose and O-6 of
N-acetylglucosamine. However, glycosylation of acceptor 7 with
the less sterically demanding 2-PMB fucosyl donor 11 had a similar
outcome than that using the TBDPS donor 10, that is, no reaction
was observed when 5 equiv of MeOTf was added, and upon further
addition of up to 15 equiv of MeOTf, TLC showed the formation of
imidates 12a and 12b, which, in this case, were not isolated. Thus,
we concluded that fucosylation of acceptor 7 with either thioglyco-
side donors 10 or 11 under MeOTf activation could not occur at the
low MeOTf concentration usually needed to activate such reactive
donors (0.1 M), and that extended reaction times with increased
concentrations of MeOTf (up to 0.4 M) was leading to the forma-
tion of the alkyl imidates and concurrent degradation of the fucosyl
donors. Indeed, treatment of acceptor 7 with MeOTf (0.5 M) in Et2O
according to the conditions that we have previously established10
gave a 7:3 mixture of methyl and ethyl imidates 12a,b in excellent
98% yield (Scheme 2). Given this excellent yield and the fact that
O
O
O
10 R = TBDPS
9
11
R = PMB
OTBDPS
O
AcO
AcO
OAc
HO
O
c
O
+
10 or 11
7
OAll
N
AcO
12a,b
OR
d
R = Me, Et
Scheme 2. Reagents and conditions: (a) TBDPSCl 2 equiv, imidazole 5 equiv,
CH3CN, rt, 18 h, 10 64%; (b) Literature procedure15 PMBCl, NaH, DMF, 11 76%; (c)
10 or 11 (5 equiv), MeOTf up to 15 equiv, Et2O, MS 4 Å, with 10 and 7: 12a 26%, 12b
16%, with 11 and 7: formation of 12a, 12b observed by TLC; (d) MeOTf 25 equiv
0.5 M, Et2O, MS 4 Å, 12a/12b 7:3 98%.
acceptor 7. Thus, using the conditions that were successful in our
previous synthesis of the methyl Lea trisaccharide,4b we attempted
the coupling of alcohol 7 with thioglycoside 10 under activation
with 5 equiv of MeOTf (0.12 M) at room temperature in Et2O
(Scheme 2). In these conditions, the acceptor remained unreacted
and it is only after adding up to 15 equiv (0.36 M) of MeOTf por-
tionwise that we observed the disappearance of alcohol 7 and
the formation of two less polar products by TLC. Although no tri-
saccharide was isolated, the two new products formed in this reac-
tion were separated by chromatography and identified by NMR to
be the methyl and ethyl imidates, 12a and 12b that were isolated
in 26% and 16% yield, respectively.
We have already described the formation of a mixture of methyl
and ethyl imidates when treating N-acetylglucosamine O-4 accep-
tors with 0.5 M MeOTf in diethyl ether10 and their characteristic
NMR signals have been well documented in our previous pa-
pers.4b,10 Thus, 12a gave, as expected, no NH signal in the 1H
NMR spectrum but an additional OCH3 singlet around 3.65 ppm
and a signal shifted up field from the acetates methyl group and
corresponding to the acetimidate methyl group. In addition, 13C
NMR spectroscopy for methyl imidate 12a gave signals at 164.8,
16.4, and 52.4 ppm, corresponding, respectively, to the C@N as well
as CH3 signals for the acetimido group and to the additional O-
methyl group. NMR spectroscopy of compound 12b also showed
no NH signal but a quaternary C@N at 164.4 ppm as well as a
methyl signal for an acetimido group at 16.1 and 1.90 ppm in the
1H and 13C NMR spectra, respectively. In addition, 13C NMR spec-
troscopy for ethyl imidate 12b showed signals corresponding to
an O-ethyl group at 14.2 and 60.6 ppm and that correlated, respec-
tively, in the HSQC, with a CH3 triplet at 1.25 ppm and a multiplet
at 4.03 ppm. Finally, ESI-HRMS confirmed the structure of the imi-
further attempts at coupling donor 11 and O-4 acceptor
7
using other promoters (NIS/TfOH) also failed, we decided to inves-
tigate the reactivity of methyl imidate 12a in fucosylation
reactions.
Indeed, we have established10 that such alkyl imidates are com-
patible with various donors and glycosylation methods for glyco-
sylation at O-4 of N-acetylglucosamine acceptors. Although we
have demonstrated previously that both methyl and ethyl imidates
could be successfully glycosylated,10 using mixtures such as 12a,b
as acceptors in glycosylation reactions leads to complex TLC, and
we thus decided to prepare methyl imidate 12a as a pure acceptor.
Furthermore, because imidates are unstable and difficult to purify
without degradation the intermediate trisaccharide imidate was
not purified but directly treated with AcOH/Ac2O to convert the
acetimidate to the acetamide as we have described previously.4b
Thus, treatment of acceptor 7 with 0.5 M MeOTf in CH2Cl2 gave
the methyl imidate 12a, which was isolated pure in 92% yield fol-
lowing silica gel chromatography using CHCl3 and MeOH as the
solvent system. Glycosylation of methyl imidate 12a with fucosyl
donor 10 under activation with 0.2 M MeOTf in Et2O at room tem-
perature for 24 h was followed by regeneration of acetamide (Ac2O,
AcOH, 55 °C) and gave the desired trisaccharide 13 in 67% yield
over the two steps (Scheme 3).
It is important to point out that quenching the glycosylation
reaction with NEt3 had to be performed slowly and at 0 °C to avoid
the unwanted degradation of the trisaccharide. Although we
O
O
OTBDPS
O
b
c
a
7
12a
OTBDPS
(R = Me)
d
AcO
AcO
OAc
e
O
O
1
O
OAll
O
NHAc
AcO
13
Scheme 3. Reagents and conditions: (a) MeOTf 25 equiv 0.5 M, CH2Cl2, MS 4 Å, 12a 92%; (b) 10 (5 equiv), MeOTf 10 equiv, 0.2 M, Et2O, MS 4 Å; (c) Ac2O, AcOH, 55 °C, 18 h, 13
67% over two steps; (d) (1) MeOTf 25 equiv, 0.5 M, CH2Cl2, MS 4 Å, 18 h rt then in situ 10 (5 equiv) in Et2O rt up to 24 h, (2) Ac2O, AcOH, 55 °C, 18 h, 13 8% over three steps (e)
(1) TBAF 7 equiv, THF, (2) Ac2O, pyridine, (3) 90% aq AcOH, 80 °C, (4) MeONa, MeOH, 1 77% over four steps.

J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
2917
postulated earlier that the high concentration of MeOTf (0.4 M)
used when attempting to couple the acetamido 7 and fucosyl
donors 10 or 11 led to degradation of the donors prior to their reac-
tion with the acceptors 7 or 12a,b, coupling of the purified methyl
imidate 12a with donor 10 was successful using a fairly high
(0.2 M) concentration of MeOTf in Et2O. Thus, we wondered if addi-
tion of the donor after formation of the alkyl imidates 12a,b in situ
would allow the two steps: imidate formation and glycosylation, to
be carried out successively in one pot. Thus, the acetamide accep-
tor 7 was converted to the imidates 12a,b with 0.5 M MeOTf in
Et2O and when TLC showed that the starting material had been
consumed, a solution of thioglycoside 10 (5 equiv) in Et2O was
added to the reaction mixture (Scheme 3). Unfortunately, TLC
showed that the glycosylation of the alkyl imidates 12a,b to the
corresponding trisaccharide imidates did not proceed to comple-
tion and after an extended reaction time and following the conver-
sion of the acetimide to the acetamide, trisaccharide 13 was only
isolated in 8% yield. Thus, we concluded that in 0.5 M MeOTf, the
donor 10 was too unstable and degraded faster than it reacted with
the imidate acceptors thus not allowing this one pot strategy to be
successful.
Trisaccharide 13 was, in turn, deprotected in four steps to give
the desired allyl Lea trisaccharide 1 (Scheme 3). Because the fucos-
idic bond is known to be acid labile when it is not stabilized at O-2
by an electron-withdrawing substituent,6a,6b,17 the silyl groups
were first removed with TBAF, and the resulting diol was acety-
lated. Subsequent acid hydrolysis of the isopropylidene group fol-
lowed by Zemplèn deacetylation gave the desired deprotected allyl
Lea (1) in 77% yield over four steps and after Biogel P2 gel exclusion
chromatography. The synthesis of the Lea analogue 1 that we have
described above constitutes a good application of the imidate syn-
thetic strategy that we have reported previsouly.10 Indeed, it illus-
trates that the temporary conversion of N-acetyl groups to alkyl
imidates when attempting to glycosylate at O-4 of N-acetylgluco-
samine is particularly useful when using a reactive thioglycoside
donor such as fucoside 10 under mild activation with MeOTf.
In the work described below, we further studied the reactivity
of an imidate versus that of the corresponding acetamido acceptor
while preparing the Lex analogue 2. As we have mentioned above,
although fucosylation at O-4 of an N-acetylglucosamine acceptor is
usually straightforward,11 it is generally known that glycosylations
using other donors are often difficult and result in poor yields.7–9
However, there are a few successful reports of such glycosylation
reactions involving mono- or disaccharide acceptors.18 Thus, our
synthetic approach to the Lex analogue 2 involved first the glucosy-
lation at O-4 followed by the fucosylation at O-3 of a suitably pro-
tected glucosamine acceptor. Chloroacetylation (ClAcCl, pyridine,
0 °C) at O-3 of the known19 benzylidene 14 gave the chloroacetate
15 (81%), which was reductively opened (NaCNBH3, HCl, Et2O) to
give the acetamido acceptor 16 in 79% yield. The acetamido 16
was subsequently converted to the methyl imidate 17 in 77% yield
using 0.5 M MeOTf in CH2Cl2 and both acceptors 16 and 17 were
As we had observed for other protected Lea analogs,16 the N-
acetylglucosamine ring in trisaccharide 13 assumed a conforma-
tional equilibrium between the usual 4C1 chair and a 1S5 or 3S5
skewed conformations in CDCl3. This conformational behavior
was characterized by a JH-1,H-2 of 4.8 Hz in the 1H NMR spectrum
that was smaller than that usually observed for a 4C1 conformation
(ꢁ8.5 Hz). As expected based on our previous study,16 1H NMR for
trisaccharide 13 in CD3CN gave a larger JH-1,H-2 coupling constant
(7.9 Hz) suggesting that this hydrogen bond-accepting solvent
the N-acetylglucosamine ring assumed mostly the usual 4C1
conformation.
submitted to glucosylation at O-4 with either the
a
- or b-trichloro-
Table 1
OR3
OBn
O
O
R2O
R1O
HO
OMe
OMe
OMe
ClAcO
NHAc
N
14 R1 = H; R2, R3 = >CHPh
15 R1 = ClAc; R2, R3 = >CHPh
16 R1 = ClAc, R2 = H, R3 = Bn
17
OAc
O
OBn
OAc
AcO
AcO
O
O
O
AcO
AcO
OMe
ClAcO
OC(NH)CCl3
OAc
NHAc
OAc
19
18α
18
β
Entry
Acceptor
Donor (equiv)
Promoter (equiv)
Temp (°C)
Time
19 (%)
b
1
2
3
4
5
6
7
8
9
17
17
17
17
17
17
17
16
16
16
16
18b (3)
18b (4)
18b (4)
18b (5)
TMSOTf (0.1–1.0)a
AgOTf (1.5)a
ꢂ78 to rt
18 h
18 h
1 h
5 min
24 h
24 h
24 h
10 min
10 min
1 h
—
ꢂ 40
11c
15c
27c
50c
75c
55c
88
90
95
71
BF3ꢀOEt2 (4–10)d
BF3ꢀOEt2 (5)e
BF3ꢀOEt2 (4)d
BF3ꢀOEt2 (4)d
BF3ꢀOEt2 (2)d
BF3ꢀOEt2 (2)f
BF3ꢀOEt2 (2)g
BF3ꢀOEt2 (2)g
BF3ꢀOEt2 (2)g
rt
40
40
40
40
40
40
rt
18
18
18
18
18
18
18
a
a
a
a
a
a
a
(5)
(10)
(5)
(5)
(5)
10
11
(5)
(1.5)
40
10 min
a
Reagents and conditions: (i) CH2Cl2, MS 4 Å; (ii) AcOH, Ac2O, 55 °C.
Traces observed by TLC.
Isolated yield over two steps.
b
c
d
e
f
Reagents and conditions: (i) CH2Cl2; (ii) AcOH, Ac2O, 55 °C.
Reagents and conditions: (i) CH2Cl2, MW 150 W, 200 psi; (ii) AcOH, Ac2O, 55 °C.
Reagents and conditions: CH2Cl2, MW 150 W, 200 psi.
Reagents and conditions: CH2Cl2.
g

2918
J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
OAc
O
OBn
AcO
AcO
OAc
O
OBn
O
O
O
AcO
AcO
OMe
HO
OAc
O
NHAc
OMe
O
a
b
OAc
20
+
2
NHAc
O
OBn
22
O
SEt
OBn
OBn
BnO
OBn
BnO
21
Scheme 4. Reagents and conditions: (a) 21 2 equiv, CuBr2 2 equiv, Bu4NBr 2 equiv, CH2Cl2, DMF, MS 4 Å, 14 h rt, 70%; (b) 1. MeONa, MeOH, 2. Pd/C, 100 psi H2, MeOH 2 83%
over two steps.
acetimidate donors 18
a
and 18b (Table 1). To avoid unnecessary
ing to 1.5 the number of equiv of donor 18a still led to an accept-
degradation of the intermediate disaccharide imidates, glucosyla-
tion of the methyl imidate 17 was always followed by the subse-
quent conversion of the methyl imidate to the acetamido group
by treatment with AcOH and Ac2O at 55 °C.
able yield of the desired disaccharide 19 (71%).
The reactions reported in Table 1 clearly indicate that when
using an excess of a mild Lewis acid such as BF3ꢀOEt2, the acetam-
ido 16 is more reactive toward glycosylation at O-4 than the
methyl imidate 17. In fact, glucosylation of acceptor 17 using
2 equiv of BF3ꢀOEt2 only gave a 55% yield of disaccharide 19 after
24 h of reaction at 40 °C, although under the same conditions
acceptor 16 gave disaccharide 19 in 90% yield after 10 min of reac-
tion (Table 1, entries 7 and 9). These results are not very surprising
because the methyl imidate substituent in acceptor 17 is more
electron-withdrawing than the acetamido group in acceptor 16
and may therefore further decrease the nucleophilicity of O-4.
However, it is interesting to notice that even though the coupling
Entries 1 to 4 in Table 1 summarize the reaction conditions that
were attempted for the glycosylation of monosaccharide imidate
acceptor 17 with the b-trichloroacetimidate donor20 18b. Applying
conditions that we have found10 to be successful for the glycosyl-
ation of a similar methyl imidate acceptor with a rhamnosyl tri-
chloroacetimidate, we first attempted to couple imidate acceptor
17 and donor 18b at low temperature under activation with
0.1 equiv of TMSOTf (Table 1, entry 1). Under these mild condi-
tions, no reaction was apparent by TLC and thus the amount of pro-
moter was increased to 1.0 equiv, and the temperature allowed to
reach ambient temperature overnight. However, after conversion
of the imidate to the acetamido, only a trace amount of disaccha-
ride 19 was observed by TLC and the majority of the acceptor
was recovered as the acetamido 16. Using AgOTf (entry 2) as a pro-
moter at ꢂ40 °C was moderately more successful as the desired
disaccharide 19 was isolated in 11% yield.
of acceptor 16 with trichloroacetimidate 18a required relatively
high temperatures to proceed, the N-acetyl group in acceptor 16
did not seem to impact negatively the glycosylation at O-4 as, for
example, we did not observe the formation of glycosyl imidate.
Therefore, we propose that the additional equivalent of Lewis acid
(BF3ꢀOEt2) present in the reaction mixture effectively allowed gly-
cosylation at O-4 by interacting with the N-acetyl group and reduc-
ing its nucleophilicity4b as well as its ability to form a hydrogen
bond network that would impede glycosylation at O-4.8
The synthesis of analogue 2 was then easily completed (Scheme
4). The chloroacetate in disaccharide 19 was removed with DABCO
in EtOH to give acceptor 20 in 71% yield and fucosylation of accep-
tor 20 with the known21 donor 21 under activation with CuBr2 and
tetra-n-butylammonium bromide gave the protected trisaccharide
22 in 70% yield.
Hypothesizing that low temperature reduced the reactivity of
the acceptor toward glycosylation, we investigated the use of the
milder Lewis acid (BF3ꢀOEt2) at higher temperatures (entries 3–7)
to glucosylate imidate 17. Although activation at rt required up
to 10 equiv of BF3ꢀOEt2 to proceed, and only led to 15% yield of
disaccharide 19, activation under microwave irradiation at 40 °C
for 5 min in the absence of molecular sieves gave an improved
27% yield of disaccharide 19. Because these rather low yields were
accompanied by fast degradation of the donor 18b, we investigated
The deprotection of trisaccharide 22 was then accomplished in
two steps: Zemplèn deacetylation followed by hydrogenolysis (H2-
Pd/C) gave the Lex analogue 2 in 83% yield over the two steps.
the coupling of the more stable a a with
-tricholoracetimidate20 18
imidate acceptor 17 (entries 5–7). Indeed, unlike the reactions with
donor 18b, TLC analysis of these reactions did not show fast degra-
dation of donor 18
a and a marked yield increase was obtained
3. Conclusions
when the glycosylation was allowed to proceed for 24 h at 40 °C
in the presence of 4 equiv of BF3ꢀOEt2. In these conditions, the de-
sired disaccharide 19 was isolated in 50% yield (entry 5); a result,
which was further improved to 75% by using 10 equiv of the donor
The syntheses described above shed some additional insight on
the value and limitation of the methyl imidate strategy that we
have developed to assist in difficult glycosylations at O-4 of N-ace-
tylglucosamine. Although fucosylations at this position are usually
easy to achieve, our synthesis of the Lea analogue 1 illustrate that it
is not always the case. Indeed the glycosylation of disaccharide 7
with either thiofucosyl donors 10 or 11 was unsuccessful. Although
the donors were activated with 0.2 M MeOTf they failed to react
with the acetamido acceptor, which upon further addition of MeO-
Tf was converted to a mixture of methyl and ethyl imidates. Thus,
it appeared that the nucleophilicity of the 4-OH toward the acti-
vated fucosyl donors was insufficient to allow glycosylation while
that of the N-acetyl group toward the MeOTf led to the formation
of the alkyl imidates. In contrast, once the acetamido was con-
verted to a methyl imidate the nucleophilicity of the 4-OH toward
18a
(entry 6).
We then investigated the coupling of donor 18
amido 16 using conditions similar to those that we had just estab-
lished for the coupling of 18 with the imidate 17. In fact, such
coupling appeared to be much easier (entries 8–11) than that of
17 with 18
a
with the acet-
a
a
and only required 2 equiv of BF3ꢀOEt2 to proceed. In-
deed, whether the reaction was left to proceed at 40 °C for
10 min, under microwave irradiation or in a conventional oil bath
(Table 1, entries 8 and 9), or whether it was left to proceed for 1 h
at room temperature (entry 10) the disaccharide 19 was isolated in
more than 88% yield when using 2 equiv of activator and 5 equiv of
donor 18a. Finally, as can be seen in Table 1, entry 11, even reduc-

J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
2919
the activated donors 10 led to acceptable yields of the desired tri-
saccharide. Therefore, we conclude that activation of thioglycoside
donors by MeOTf to glycosylate at O-4 of a glucosamine acceptor is
best accomplished following the temporary protection of the N-
acetyl group as a methyl imidate, especially when the donors are
highly reactive and prone to degradation.
The synthesis of trisaccharide 2 shed light on the relative reac-
tivity of methyl imidate and acetamido glucosamine monosaccha-
ride acceptors when coupled to a relatively stable trichloro-
2 equiv) and the known13 peracetylated galactosyl bromide 4
(1.17 g, 2.85 mmol, 2 equiv) were added and the reaction mixture
was stirred under N2 for 3 h at 50 °C. The reaction mixture was
then filtered on Celite and the solids were washed with CH2Cl2
(200 mL). The combined filtrate and washing was washed sequen-
tially with satd aq NaHCO3 (200 mL) and brine (200 mL), and the
aq phases were re-extracted with CH2Cl2 (100 mL). The organic
solutions were combined, dried, and concentrated. Flash chroma-
tography of the residue (8:1 EtOAc–hexanes) gave the pure
acetimidate glycosyl donor such as 18
a
under activation with an
disaccharide 5 as a
colorless glass (0.831 g, 82%). 1H NMR
excess of BF3ꢀOEt2. Indeed, in these conditions, the acetamido
acceptor proved to react faster and in better yields than the corre-
sponding methyl imidate, even though such reactions required at
least 2 equiv of promoter at room temperature to proceed. We pro-
pose that in these conditions 1 equiv of BF3ꢀOEt2 interacts non-
covalently with the nucleophilic N-acetyl group and that the sec-
ond one promotes glycosylation. Therefore, as long as donor and
acceptor can withstand activation with multiple equivalents of
BF3ꢀOEt2 at elevated temperatures, glycosylations at O-4 of a
glucosamine acceptor with a trichloroacetimidate donor do not
seem to benefit from the temporary protection of the N-acetyl
group as a methyl imidate. In fact, the presence of a more electron
withdrawing methyl imidate at C-2 of the acceptor further
decreases its reactivity.
(400 MHz, CDCl3): d 7.36, 6.87 (2d, 2 ꢃ 2H, J = 8.7 Hz, Ar); 5.84
(m, 1H, –CH@CH2); 5.74 (br d, 1H, J = 6.9 Hz, NH); 5.45 (s, 1H,
CHPh); 5.32–5.20 (m, 4H, H-1, H-40, –CH@CH2); 5.12 (dd, 1H,
J = 8.0, 10.3 Hz, H-20); 4.88 (dd, 1H, J = 3.4, 10.4 Hz, H-30); 4.76 (d,
1H, J = 8.0 Hz, H-10); 4.72 (t, 1H, J = 9.3 Hz, H-3); 4.33–4.25 (m,
2H, H-6a, OCHH allyl); 4.12–3.97 (m, 2H, H-6a0, OCHH allyl); 3.90
(dd, 1H, J = 5.9, 11.0 Hz, H-6b0); 3.80 (s, 3H, OCH3); 3.75 (t, 1H,
J = 10.2 Hz, H-6b); 3.62 (t, 1H, J = 8.8 Hz, H-4); 3.52 (m, 2H, H-5,
H-50); 3.01 (m, 1H, H-2); 2.09, 1.97, 1.96, 1.94, 1.93 (5s, 5 ꢃ 3H,
5 ꢃ CH3CO). 13C NMR (100 MHz, CDCl3): d 170.8, 170.7, 170.2,
170.1, 169.5 (C@O); 133.5 (–CH@); 127.4, 113.7 (Ar); 118.1
(@CH2); 101.5 (CHPMP); 100.1 (C-10); 98.4 (C-1); 80.8 (C-4); 76.5
(C-3); 71.0 (C-30); 70.6 (OCH2CH@); 69.3, 65.8 (C-5, C-50); 68.7
(C-6); 66.8 (C-40); 60.9 (C-60); 58.5 (C-2); 55.2 (OCH3); 23.7, 20.7,
20.6, 20.5 (CH3CO). HRMS calcd for C33H43NO16 [M+H]+ 710.2660.
Found: 710.2659.
4. Experimental
4.3. Allyl 2-acetamido-3-O-(2,3,4,6-tetra-O-acetyl-b-
D-
4.1. General methods
galactopyranosyl)-2-deoxy-b- -glucopyranoside (6)
D
1H (600.13, 400.13 or 300.13 MHz) and 13C NMR (150, 100 or
75 MHz) spectra were recorded at 300 K for solution in CDCl3
The benzylidene 5 (590 mg, 0.832 mmol) was dissolved in 90%
aq AcOH (50 mL) and the solution was stirred for 1 h at 60 °C.
The mixture was then co-concentrated with toluene
(3 ꢃ 250 mL), and chromatography (20:1 CHCl3–MeOH) of the
resulting residue gave pure diol 6 (331 mg, 67%) as a colorless
glass. 1H NMR (400 MHz, CDCl3): d 6.29 (d, 1H, J = 8.2 Hz, NH);
5.87 (m, 1H, –CH@); 5.35 (br d, 1H, J = 3.2 Hz, H-40); 5.32–5.17
(m, 3H, H-20, –CH@CH2); 5.03 (dd, 1H, J = 3.4, 6.5 Hz, H-30); 4.94
(d, 1H, J = 8.3 Hz, H-1); 4.59 (d, 1H, J = 8.1 Hz, H-10); 4.41 (dd, 1H,
J = 8.1, 10.0 Hz, H-3); 4.34 (dd, 1H, J = 10.1, 15.6 Hz, OCHH allyl);
4.18-3.96 (m, 3H, OCHH allyl, H-6a, H-6b); 3.91 (dd, 1H, J = 3.3,
12.1 Hz, H-6a0); 3.80 (dd, 1H, J = 4.0, 11.8 Hz, H-6b0); 3.51 (t, 1H,
J = 9.3 Hz, H-5); 3.45–3.39 (m, 1H, H-50); 3.11 (dd, 1H, J = 8.2,
14.7 Hz, H-2); 2.17, 2.10, 2.07, 2.00, 1.99 (5s, 5 ꢃ 3H, 5 ꢃ CH3CO).
13C NMR (100 MHz, CDCl3): d 170.9, 170.5, 170.0, 169.2 (C@O);
133.7 (–CH@CH2); 117.9 (–CH@CH2); 101.3 (C-10); 98.5 (C-1);
83.2 (C-3); 75.2, 70.0 (C-5, C-50); 71.0 (C-4); 70.7 (C-3); 70.3
(OCH2CH@); 68.8 (C-20); 66.9 (C-40); 62.6 (C-60); 61.5 (C-6); 57.3
(C-2); 23.6, 20.7, 20.6, 20.5 (CH3CO). HRMS calcd for C25H37NO15
[M+H]+ 592.2241. Found: 592.2253.
(internal standard, for 1H residual CHCl3 d 7.24; for 13C CDCl3
d
77.0), DMF-d7 (internal standard, for 1H residue DMF d 2.75; for
13C DMF-d7 d 29.8), CD3CN (internal standard, for 1H residual
CH3CN d 1.93) or D2O [external standard 3-(trimethylsilyl)-propi-
onic acid-d4, sodium salt (TSP) for 1H d 0.00, for 13C d 0.00]. 1H
NMR and 13C NMR chemical shifts are reported in parts per million
(ppm). Coupling constants (J) are reported in Hertz (Hz). Chemical
shifts and coupling constants were obtained from a first-order
analysis of one-dimensional spectra. Assignments of proton and
carbon resonances were based on two dimensional 1H–1H and
13C –1H correlation experiments. Multiplicities are abbreviated as
follows: singlet (s), doublet (d), triplet (t), quartet (q), multiplet
(m), and broadened (b). Analytical thin-layer chromatography
(TLC) was performed using Silica Gel 60 F254 precoated plates
(250 lm) with a fluorescent indicator, visualized under UV and
charred with 10% sulfuric acid in ethanol. Compounds were puri-
fied by flash chromatography with Silica Gel 60 (230–400 mesh)
unless otherwise stated. Solvents were distilled and dried accord-
ing to standard procedures,22 and organic solutions were dried
over Na2SO4 and concentrated below 40 °C, under reduced pres-
sure. Molecular sieves were activated by heating at high tempera-
ture over P2O5 under vacuum. IR experiments were conducted
using BOMEN MB-100 FT-IR. High-resolution electrospray ioniza-
tion mass spectra (HR-ESI-MS) were recorded by the analytical ser-
vices of the McMaster Regional Center for Mass Spectrometry,
Hamilton, Ontario.
4.4. Allyl 2-acetamido-3-O-(2,3,4,6-tetra-O-acetyl-b-D-
galactopyranosyl)-6-O-tert-butyldiphenylsilyl-2-deoxy-b-D-
glucopyranoside (7)
Diol 6 (1.02 g, 1.69 mmol) and imidazole (702 mg, 10.3 mmol,
6.1 equiv) were dissolved in CH3CN (20 mL). t-Butyldiphenylsilyl
chloride (TBDPSCl, 441 lL, 1.75 mmol, 1.0 equiv) was added into
the solution and the reaction mixture was stirred for 15 min at
rt. The reaction mixture was then concentrated and the residue
was dissolved in CH2Cl2 (250 mL) and washed with brine
(50 mL). The aq phase was re-extracted with CH2Cl2 (250 mL)
and the combined organic solutions were dried and concentrated.
Flash chromatography (3:2 EtOAc–hexanes) of the residue afforded
the pure silyl ether 7 (950 mg, 67%) as a colorless glass. The regi-
oselectivity of the silylation was confirmed through the acetylation
4.2. Allyl 2-acetamido-3-O-(2,3,4,6-tetra-O-acetyl-b-D-
galactopyranosyl)-2-deoxy-4,6-O-p-methoxybenzylidene-b-D-
glucopyranoside (5)
The known12 acceptor 3 (549 mg, 1.43 mmol) was dissolved in a
1:1 mixture of toluene and nitromethane (10 mL). Activated pow-
dered molecular sieves 4 Å (1.6 g), Hg(CN)2 (722 mg, 2.86 mmol,

2920
J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
of an analytical sample of alcohol 7 as described in Section 4.4.1. 1H
NMR (400 MHz, CDCl3): d 7.72–7.64 (m, 4H, Ar); 7.43–7.32 (m, 6H,
Ar); 5.89 (m, 1H, –CH@); 5.62 (d, 1H, J = 6.0 Hz, NH); 5.36 (br d, 1H,
J = 2.7 Hz, H-40); 5.29–5.14 (m, 4H, H-1, H-20, @CH2); 4.97 (t, 1H,
J = 8.8 Hz, H-30); 4.55 (d, 1H, J = 8.1 Hz, H-1); 4.41 (t, 1H,
J = 6.7 Hz, H-3); 4.32 (dd, 1H, J = 3.5, 11.3 Hz, OCHH allyl); 4.13–
3.95 (m, 3H, H-6a, H-6b, OCHH allyl); 3.90–3.72 (m, 3H, H-6a0, H-
6b0, H-4); 3.51–3.38 (m, 2H, H-5, H-50); 3.04 (dd, 1H, J = 4.8,
7.9 Hz, H-2); 2.13, 2.06, 1.99, 1.96, 1.96 (5s, 3H, 5 CH3CO); 1.03
(s, 9H, (CH3)3C). 13C NMR (100 MHz, CDCl3): d 170.7, 170.4, 170.1,
170.0, 169.1 (C@O); 135.7, 135.6, 129.5, 127.6 (Ar); 133.9
(–CH@); 117.8 (@CH2); 101.3 (C-10); 97.8 (C-1); 83.6 (C-3); 76.2,
69.5 (C-5, C-50); 71.0 (C-4); 70.8 (C-30); 69.7 (OCH2CH@); 68.9 (C-
20); 66.9 (C-40); 63.6 (C-60); 61.5 (C-6); 57.8 (C-2); 26.7 ((CH3)3C);
23.8, 20.8, 20.6, 20.5 (CH3CO); 19.3 ((CH3)3C). IR (NaCl):
1753 cmꢂ1 (4 ꢃ C@O acetates), 1655 cmꢂ1 (C@O acetamide). HRMS
calcd for C41H55NO15Si [M+H]+: 830.3419. Found: 830.3402.
4.6. Allyl 3-O-(2,3,4,6-tetra-O-acetyl-b-
tert-butyldiphenylsilyl-2-deoxy-2-methylacetimido-b-
glucopyranoside (12a) and allyl 3-O-(2,3,4,6-tetra-O-acetyl-b-
galactopyranosyl)-6-O-tert-butyldiphenylsilyl-2-deoxy-2-
ethylacetimido-b-D-glucopyranoside (12b)
D-galactopyranosyl)-6-O-
D-
D-
4.6.1. Glycosylation of acceptor 7 with donor 10
A mixture of acceptor 7 (45 mg, 0.054 mmol), thioglycoside 10
(122 mg, 0.250 mmol, 4.6 equiv), and activated powdered 4 Å
molecular sieves (150 mg) in Et2O (2 mL) was stirred for 1 h at rt
under N2. MeOTf (29 lL, 0.256 mmol, 4.6 equiv) was added and
the reaction mixture was stirred at rt for 5 h. Up to 15 equiv of
MeOTf was added to the reaction mixture over the following
24 h while the reaction was kept at rt under N2. When TLC (7:3
EtOAc–hexanes) showed no remaining starting acceptor, the reac-
tion mixture was quenched with Et3N (147 lL). The reaction mix-
ture was filtered, the solids were washed with CH2Cl2, and the
combined organic solutions were washed with aq satd NaHCO3
and brine. The aq phases were re-extracted with CH2Cl2 and the
combined organic layers were dried and concentrated. Flash chro-
matography (3:7 EtOAc–hexanes) of the residue gave first the pure
ethyl imidate 12b as a colorless glass (7.3 mg, 16%) then the pure
methyl imidate 12a also as a colorless glass (12.1 mg, 26%).
4.4.1. Acetylation of an analytical sample of alcohol 7
An analytical sample of alcohol 7 (20 mg, 24
solved in Ac2O (1 mL) and pyridine (1 mL), and DMAP (5 mg,
41 mol, 1.7 equiv) was added. The reaction solution was stirred
lmol) was dis-
l
at 50 °C for 18 h, diluted in CH2Cl2 (30 mL), and washed with 2 M
HCl (2 ꢃ 30 mL) and satd aq NaHCO3 (2 ꢃ 30 mL). The organic layer
was dried and concentrated, and flash chromatography (3:2
EtOAc–hexanes) of the residue gave the pure acetylated disaccha-
ride 8 (16 mg, 78% yield). 1H NMR (400 MHz, CDCl3): d 7.67–7.62
(m, 4H, Ar); 7.43–7.33 (m, 6H, Ar); 5.89 (m, 1H, –CH@); 5.65 (d,
1H, J = 7.4 Hz, NH); 5.32 (d, 1H, J = 2.7 Hz, H-40); 5.25 (dd, 1H,
J = 1.5, 17.2 Hz, CH@CHH); 5.18 (dd, 1H, J = 1.2, 10.3 Hz, CH@CHH);
5.05 (dd, 1H, J = 7.9, 10.4 Hz, H-20); 4.97 (d, 1H, J = 8.0 Hz, H-1);
4.93 (dd, 1H, J = 3.4, 7.7 Hz, H-30); 4.86 (t, 1H, J = 9.2 Hz, H-4);
4.53 (d, 1H, J = 7.9 Hz, H-10); 4.49 (t, 1H, J = 9.1 Hz, H-3); 4.29 (dd,
1H, J = 5.3, 12.7 Hz, OCHH allyl); 4.12–4.01 (m, 3H, H-6a0, H-6b0,
OCHH allyl); 3.84 (t, 1H, J = 6.3 Hz, H-50); 3.76–3.64 (m, 2H, H-6a,
H-6b); 3.55 (m, 1H, H-5); 3.17 (m, 1H, H-2); 2.11, 2.04, 2.03,
2.00, 1.94, 1.88 (6s, 6 ꢃ 3H, 6 CH3CO); 1.02 (s, 9H, (CH3)3C). 13C
NMR (100 MHz, CDCl3): d 170.7, 170.2, 170.2, 169.2, 168.9
(C@O); 135.6, 135.6, 129.6, 127.6 (Ar); 133.7 (CH@); 133.3 (Ar);
117.9 (@CH2); 100.6 (C-10); 97.8 (C-1); 77.1 (C-3); 74.8 (C-5);
71.1 (C-30); 70.5 (C-5); 69.8 (OCH2C@); 69.3 (C-4, C-20); 66.9 (C-
4); 63.2 (C-6); 61.1 (C-60); 58.2 (C-2); 26.7 ((CH3)3C); 23.7, 20.8,
20.8 20.6, 20.6, 20.5 (CH3CO); 19.2 ((CH3)3C).
4.6.2. Reaction of acceptor 7 with MeOTf in Et2O gave a mixture
12a,b
Acceptor 7 (25 mg, 0.030 mmol) was dissolved in Et2O (1.5 mL);
activated molecular sieves 4 Å (150 mg) were added and the mix-
ture was stirred under N2 at rt for 1 h. MeOTf (85 lL, 0.75 mmol,
25 equiv, 0.5 M) was then added and the reaction mixture was stir-
red at rt under N2 for 18 h. The reaction mixture was quenched
slowly with Et3N (106 lL, 0.76 mmol, 25 equiv), the solids were fil-
tered off and washed with CH2Cl2 (40 mL). The organic solution
was washed with satd aq NaHCO3 (40 mL), dried, and concen-
trated. Chromatography (50:1 CHCl3–MeOH) gave 7:3 mixture of
the methyl and ethyl imidates 12a and 12b, respectively (25 mg,
98%).
4.6.3. Reaction of acceptor 7 with MeOTf in CH2Cl2 gave methyl
imidate 12a
The acceptor 7 (100 mg, 0.12 mmol) was dissolved in CH2Cl2
(6 mL) containing activated molecular sieves 4 Å (340 mg) and
after stirring under N2 at rt for 1 h, MeOTf (341 lL, 3.0 mmol,
4.5. Ethyl 2-O-tert-butyldiphenylsilyl-3,4-O-isopropylidene-b-
thiofucopyranoside (10)
L
-
25 equiv, 0.5 M) was added. The reaction was allowed to proceed
at rt under N2 for 18 h and worked up as described above in Section
4.6.2. Chromatography (50:1 CHCl3–MeOH) gave the pure methyl
imidate 12a (95 mg, 92%) as a colorless glass.
The known14 alcohol
9 (1.05 g, 4.2 mmol) and imidazole
(1.48 g, 21.8 mmol, 5.2 equiv) were dissolved in CH3CN (35 mL).
TBDPSCl (2.16 mL, 8.4 mmol, 2.0 equiv) was added to the solution
and the reaction mixture was stirred for 18 h at rt. The solution
was diluted with CHCl3 (150 mL), and washed with brine
(150 mL) and 1 M aqueous NaOH (150 mL). The organic layer
was dried and concentrated, and flash chromatography of the res-
idue (3:100 EtOAc–hexanes) gave the pure silyl ether 10 (1.31 g,
64%). 1H NMR (300 MHz, CDCl3): d 7.78–7.65 (m, 4H, Ar); 7.44–
7.30 (m, 6H, Ar); 4.44 (d, 1H, J = 8.0 Hz, H-1); 4.17 (t, 1H,
J = 5.9 Hz, H-3); 4.02 (dd, 1H, J = 1.9, 6.0 Hz, H-4); 3.91–3.82 (m,
1H, H-5); 3.72 (dd, 1H, J = 5.7, 8.0 Hz, H-2); 2.59–2.33 (m, 2H,
SCH2CH3); 1.31 (d, 3H, J = 6.3 Hz, H-6); 1.22, 1.13 (2s, 2 ꢃ 3H,
(CH3)2C); 1.07 (s, 9H, (CH3)3C). 13C NMR (75 MHz, CDCl3): d
136.4, 136.4, 129.6, 129.4, 127.4, 127.2 (Ar); 109.5 (Ar quater-
nary); 85.3 (C-1); 79.2 (C-3); 76.0 (C-4); 73.7 (C-2); 71.6 (C-5);
27.2, 26.5 ((CH3)3C and (CH3)2C); 24.6 (SCH2CH3); 19.7
((CH3)3C); 16.8 (SCH2CH3). 13.5 (C-6); HRMS calcd for C27H38O4SSi
[M+Na]+ 509.2158. Found: 509.2140.
4.6.4. Analytical data for methyl imidate 12a
1H NMR (400 MHz, CDCl3): d 7.73–7.68 (m, 4H, Ar); 7.41–7.29
(m, 6H, Ar); 5.86 (m, 1H, –CH@); 5.35 (br d, 1H, J = 3.0 Hz, H-40);
5.29–5.12 (m, 3H, H-2, @CH2); 4.95 (dd, 1H, J = 3.7, 10.4 Hz, H-
30); 4.57 (d, 1H, J = 8.0 Hz, H-10); 4.39–4.27 (m, 2H, H-1, OCHH al-
lyl); 4.14–3.85 (m, 6H, H-6a, H-6b, H-50, H-6a0, H-6b0, OCHH allyl);
3.65–3.50 (m, 5H, H-3, H-4, OCH3); 3.42 (m, 1H, H-5); 3.30 (t, 1H,
J = 8.2 Hz; H-2); 2.14, 2.00, 1.98, 1.97 (4s, 4 ꢃ 3H, 4 ꢃ CH3CO); 1.91
(s, 3H, CH3CN imidate); 1.06 (s, 9H, (CH3)3C). 13C NMR (100 MHz,
CDCl3): d 170.4, 170.1, 170.1, 169.4 (C@O); 164.8 (C@N); 135.7,
135.6, 129.5, 127.6 (Ar); 134.2 (–CH@CH2); 133.8 (Ar quaternary);
116.7 (–CH@CH2); 102.0 (C-10); 101.7 (C-1); 89.4 (C-3); 77.3 (C-5);
71.1 (C-30); 70.8 (C-50); 69.5 (OCH2 allyl); 68.7 (C-4); 68.4 (C-20);
66.8 (C-40); 63.8 (C-60); 63.7 (C-2); 61.5 (C-6); 52.4 (OCH3); 26.8
((CH3)3C); 20.6, 20.5, 20.3, 20.2 (CH3CO); 19.3 ((CH3)3C); 16.4
(CH3CN). IR (NaCl): 1754 cmꢂ1 (C@O), 1686 cmꢂ1 (C@N). HRMS
calcd for C42H57NO15Si [M+H]+ 844.3576. Found: 844.3591.

J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
2921
4.6.5. Analytical data for ethyl imidate 12b
20.6 (CH3CO); 19.3, 19.2 ((CH3)3C); 16.0 (C-600). HRMS calcd for
1H NMR (400 MHz, CDCl3): d 7.73–7.68 (m, 4H, Ar); 7.41–7.29
(m, 6H, Ar); 5.86 (m, 1H, –CH@); 5.35 (br d, 1H, J = 3.0 Hz, H-40);
5.29–5.12 (m, 3H, H-2, @CH2); 4.95 (dd, 1H, J = 3.7, 10.4 Hz, H-
30); 4.53 (d, 1H, J = 8.0 Hz, H-10); 4.39–4.27 (m, 2H, H-1, OCHH al-
lyl); 4.14–3.85 (m, 8H, H-6a, H-6b, H-50, H-6a0, H-6b0, OCHH allyl,
OCH2CH3 imidate); 3.60–3.50 (m, 2H, H-3, H-4); 3.42 (m, 1H, H-
5); 3.30 (t, 1H, J = 8.2 Hz; H-2); 2.14, 2.00, 1.98, 1.97 (4s, 4 ꢃ 3H,
4 ꢃ CH3CO); 1.90 (s, 3H, CH3CN imidate); 1.25 (t, 3H, J = 7.1 Hz,
OCH2CH3 imidate); 1.06 (s, 9H, (CH3)3C). 13C NMR (100 MHz,
CDCl3): d 170.4, 170.1, 170.1, 169.4 (C@O); 164.4 (C@N); 135.7,
135.6, 129.5, 127.6 (Ar); 134.2 (–CH@CH2); 133.8 (Ar quaternary);
116.7 (–CH@CH2); 102.1 (C-10); 101.7 (C-1); 89.4 (C-3); 77.3 (C-5);
71.1 (C-30); 70.8 (C-50); 69.5 (OCH2 allyl); 68.7 (C-4); 68.4 (C-20);
66.8 (C-40); 63.8 (C-60); 63.7 (C-2); 61.5 (C-6); 60.6 (OCH2 imidate);
26.8 ((CH3)3C); 20.6, 20.5, 20.3, 20.2 (CH3CO); 19.3 ((CH3)3C); 16.1
(CH3CO); 14.2 (CH3CH2 imidate). HRMS calcd for C43H59NO15Si
[M+H]+ 858.3732. Found: 858.3714.
C66H87NO19Si2 [M+H]+ 1254.5489. Found: 1254.5430.
4.8. Allyl 2-acetamido-2-deoxy-3-O-(
a-L-fucopyranosyl)-4-O-(b-
D
-galactopyranosyl)-b- -glucopyranoside (1)
D
Trisaccharide 13 (29 mg, 0.023 mmol) was dissolved in THF
(3 mL) and a 1 M solution of TBAF in THF (169 L, 0.169 mmol,
l
7.3 equiv) was added. The reaction mixture was stirred at rt for
1.5 h and concentrated. Ac2O (1 mL) and pyridine (1 mL) were
added to the dry residue and acetylation was allowed to proceed
for 2.5 h. The mixture was co-concentrated with toluene
(3 ꢃ 10 mL), and the residue was submitted to flash chromatogra-
phy (8:2 EtOAc–hexanes). The fractions containing the diol were
pooled, concentrated, and the residue was dissolved in 90% aq
AcOH (2 mL). The solution was stirred for 0.5 h at rt, the temper-
ature was raised to 80 °C in 1 h and stirring at 80 °C was contin-
ued for 6 h. The solution was co-concentrated with toluene
(2 ꢃ 10 mL), and the residue was dissolved in anhyd MeOH
4.7. Allyl 2-acetamido-3-O-(2,3,4,6-tetra-O-acetyl-b-
galactopyranosyl)-6-O-tert-butyldiphenylsilyl-4-O-(2-O-tert-
butyldiphenylsilyl-3,4-O-isopropylidene- -fucopyranosyl)-2-
deoxy-b- -glucopyranoside (13)
D-
(2 mL). A 1 M solution of NaOMe in MeOH (40 lL) was added
to the solution and the reaction mixture was stirred for 18 h at
rt. The solution was deionized with DOWEXÒ H+ resin, the resin
was filtered and rinsed with MeOH (10 mL). The pooled filtrate
and washings were concentrated, and the dry residue was puri-
a
-L
D
A solution of the acceptor 12a (23 mg, 0.027 mmol) and thiogly-
coside 10 (68 mg, 0.140 mmol, 5.2 equiv) in anhyd Et2O (1.4 mL)
containing activated molecular sieves 4 Å (100 mg) was stirred un-
fied by gel permeation chromatography on a Biogel P2
(100 ꢃ 1 cm) column eluted with water. The pure allyl Lea
1
(10.3 mg, 77%) was obtained as a white amorphous powder after
der N2 for 1 h at rt. MeOTf (32
l
L, 0.28 mmol, 10 equiv, 0.2 M) was
freeze-drying. [
a
]
D
ꢂ55.6 (c 0.5, H2O). 1H NMR (300 MHz, D2O): d
added to the reaction mixture that was then stirred for 24 h at rt
under N2. The reaction mixture was cooled to 0 °C, diluted with an-
hyd Et2O (10 mL), and the reaction was quenched by adding Et3N
5.80 (m, 1H, CH@); 5.18 (m, 2H, @CH2); 4.92 (d, 1H, J = 3.7 Hz, H-
100); 4.75 (m, 1H, H-500); 4.46 (d, 1H, J = 8.4 Hz, H-1); 4.38 (d, 1H,
J = 7.6 Hz, H-10); 4.23 (m, 1H, OCHH allyl); 4.05 (m, 1H, OCHH al-
lyl); 4.00–3.3 (m, 15H, H-2, H-3, H-4, H-5, H-6a, H-6b, H-20, H-30,
H-40, H-50, H-6a0, H-6b0, H-200, H-300, H-400); 1.92 (s, 3H, CH3CO);
1.07 (d, 3H, J = 6.5 Hz, H-600). 13C NMR (75 MHz, D2O): d 174.5
(C@O); 133.3 (–CH@); 118.2 (@CH2); 102.9 (C-10); 99.5 (C-1);
98.1 (C-100); 76.1, 75.4, 74.8, 72.4, 72.3, 71.9, 70.5, 69.1, 68.4,
67.8 (C-3, C-4, C-5, C-20, C-30, C-40, C-50, C-200, C-300, C-400); 70.5
(CH2 allyl); 66.8 (C-500); 61.6, 59.7 (C-6, C-60); 55.7 (C-20); 22.3
(CH3CO); 15.4 (C-600). HRMS calcd for C23H39NO15 [M+H]+
570.2398. Found: 570.2401.
(40 lL) slowly to the mixture. It was then filtered, solids were
washed with CH2Cl2, and the combined filtrate and washings were
concentrated to dryness. The residue was dissolved in Ac2O
(2.0 mL) and AcOH (0.7 mL) and the solution was stirred at 55 °C
for 18 h. The mixture was co-concentrated with toluene
(3 ꢃ 20 mL), and the residue was dissolved in CH2Cl2 (40 mL),
and washed with satd aq NaHCO3. The aq phases were extracted
with CH2Cl2 (40 mL), and the combined organic solutions were
dried and concentrated. Flash chromatography (3:2 EtOAc–hex-
anes) of the residue gave pure trisaccharide 13 as a colorless glass
(23 mg, 67%). 1H NMR (400 MHz, CDCl3): d 7.73–7.57 (m, 8 H, Ar);
7.41–7.26 (m, 12H, Ar); 6.46 (d, 1H, J = 9.4 Hz, NH); 5.67 (m, 1H, –
CH@); 5.33 (br d, 1H, J = 3.2 Hz, H-40); 5.24–4.99 (m, 4H, H-20, H-30,
@CH2); 4.72 (d, 1H, J = 8.0 Hz, H-10); 4.68 (d, 1H, J = 2.9 Hz, H-1);
4.65 (d, 1H, J = 3.3 Hz, H-100); 4.31–4.00 (m, 9H, H-2, H-3, H-4, H-
6a, H-6b, H-6a0, H-300, H-400, OCHH allyl); 3.95–3.65 (m, 6H, H-5,
H-50, H-6b0, H-200, H-500, OCHH allyl); 2.01 (s, 3H, CH3CO); 2.00 (s,
9H, 3 ꢃ CH3CO); 1.98 (s, 3H, CH3CO); 1.26–1.21 (m, 9H, H-6,
(CH3)2C); 1.06, 1.01 (2s, 2 ꢃ 9H, 2 (CH3)3C). 1H NMR (400 MHz,
CD3CN): d 7.78–7.23 (m, 20H, Ar); 6.47 (d, 1H, J = 9.5 Hz, NH);
5.87 (m, 1H, –CH@); 5.39 (br d, 1H, J = 3.1 Hz, H-40); 5.28–5.02
(m, 4H, H-20, H-30, = CH2); 5.01 (br d, 1H, J = 3.6 Hz, H-100); 4.93
(m, 2H, H-10, H-500); 4.45 (d, 1H, J = 7.9 Hz, H-1); 4.38–3.69 (m,
13H, H-2, H-3, H-4, H-6a, H-6b, H-50, H-6a0, H-6b0, H-200, H-300, H-
400, OCH2 allyl); 3.44 (m, 1H, H-5); 2.09, 2.07, 2.01, 1.98, 1.93 (5s,
5 ꢃ 3H, 5 ꢃ CH3CO); 1.29 (d, 3H, J = 6.6 Hz, H-600); 1.20 (s, 3H, one
of (CH3)2C); 1.02, 1.01 (2s, 2 ꢃ 9H, 2 of (CH3)3C); 0.93 (s, 3H, one
of (CH3)2C). 13C NMR (100 MHz, CDCl3): d 170.2, 170.1, 170.0,
169.4, 169.2 (C@O); 136.4, 135.9, 135.6, 129.8, 120.5, 127.6,
127.5 (Ar); 133.9 (–CH@); 133.8 (Ar quaternary); 117.1 (@CH2);
100.7 (C-10); 97.2 (C-1); 93.8 (C-100); 76.3, 67.9, 67.3 (C-3, C-4,
C-300); 75.4 (OCH2 allyl); 72.0, 71.2 (C-5, C-50); 70.5 (C-30); 69.9
(C-200); 67.9 (C-4, C-500); 67.6 (C-20); 66.3 (C-40); 62.7 (C-60), 60.0
(C-6); 48.9 (C-2); 27.1, 26.9 ((CH3)3C); 26.3 ((CH3)2C); 23.7, 20.8,
4.9. Methyl 2-acetamido-4,6-O-benzylidene-3-O-chloroacetyl-
2-deoxy-b-D-glucopyranoside (15)
Pyridine (2.8 mL, 34 mmol, 15 equiv) was added to a solution of
the known19 alcohol 14 (750 mg, 2.31 mmol) in anhyd CH2Cl2
(83 mL), and the mixture was cooled down to 0 °C under N2. Chlo-
roacetyl chloride (430 lL, 5.4 mmol, 2.3 equiv) was then slowly
added to the mixture over 15 min. The reaction mixture was al-
lowed to warm up to rt over 2 h and was stirred for 18 h at rt.
The solution was co-concentrated with toluene (3 ꢃ 15 mL) and
the solid obtained was recrystallized from hot ethanol (50 mL).
The mixture was filtered and the solid was washed with cold EtOH
(10 mL). The pure benzylidene acetal 15 was obtained as a white
powder (745 mg, 81%). Mp = 257–259 °C, [
a
]
ꢂ3.80 (c 1.0, DMF).
D
1H NMR (600 MHz, DMF-d7): d 8.07 (d, 1H, J = 9.1 Hz, NH); 7.47–
7.39 (m, 5H, Ar); 5.73 (s, 1H, CHPh); 5.36 (dd, 1H, J = 9.8, 9.9 Hz,
H-3); 4.75 (d, 1H, J = 8.4 Hz, H-1); 4.45–4.31 (m, 3H, ClCH2–, H-
6a); 3.98 (dd, 1H, J = 9.1, 9.6 Hz, H-2); 3.92–3.85 (m, 2H, H-4, H-
6b); 3.64 (m, 1H, H-5); 3.45 (s, 3H, OCH3); 1.86 (s, 3H, COCH3).
13C NMR (150 MHz, DMF-d7): d 171.2, 168.3 (C@O); 139.3 (Ar,
quat); 130.1, 129.3, 127.5 (Ar); 103.6 (C-1); 102.1 (CHPh); 79.7
(C-4); 75.2 (C-3); 69.0 (C-6); 67.2 (C-5); 57.2 (OCH3); 55.4 (C-2);
42.1 (ClCH2–); 23.3 (COCH3). HRMS calcd for C18H22ClNO7 [M+H]+
400.1163. Found: 400.1165.

2922
J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
4.10. Methyl 2-acetamido-6-O-benzyl-3-O-chloroacetyl-2-
deoxy-b- -glucopyranoside (16)
washed with satd aq NaHCO3 (20 mL), and the aqueous layer was
re-extracted with CH2Cl2 (20 mL). The combined organic layers
were dried and concentrated, and the residue was submitted to
flash chromatography (8:2 EtOAc–hexanes) yielding the pure
disaccharide 19 (80 mg, 75%) that was obtained as a colorless glass.
D
A solution of benzylidene acetal 15 (80 mg, 0.163 mmol), NaC-
NBH3 (128 mg, 2.03 mmol, 12.5 equiv), and methyl orange indica-
tor (2 mg) in anhyd in THF (3.6 mL) containing activated MS 3 Å
(150 mg) was cooled to 0 °C under N2 and stirred for 0.5 h. M HCl
in Et2O (ꢁ1.6 mL) was then added to the reaction mixture at 0 °C
until the methyl red indicator turned pink, remained as such for
10 min, and H2 (g) was no longer generated. The reaction mixture
was then stirred for 1 h at room temperature and filtered over Cel-
iteÒ. The solids were washed with THF (3 ꢃ 5 mL), and the pooled
filtrate and washings were concentrated. Flash chromatography
(25:1 CHCl3–MeOH) of the residue gave pure alcohol 16 (69 mg,
4.12.2. Glycosylation of the acetamido 16
BF3ꢀOEt2 (13
lL, 0.080 mmol, 2.0 equiv) was added to a solution
of the acceptor 16 (20 mg, 0.049 mmol) and the glucosyl donor 18
a
(122 mg, 0.248 mmol, 5.0 equiv) in CH2Cl2 (2.5 mL) that was stir-
red under N2 at rt. The reaction mixture was stirred for 1 h at rt un-
der N2 and quenched with Et3N (15 lL, 0.11 mmol, 2.3 equiv). The
mixture was diluted with CH2Cl2 (10 mL) and washed with satd aq
NaHCO3 (10 mL). The aqueous layer was re-extracted with CH2Cl2
(2 ꢃ 10 mL), and the combined organic layers were dried and con-
centrated. Purification, as described above Section 4.12.1, gave the
pure disaccharide 19 (35 mg, 95%) as a colorless glass.
79%) as a colorless glass. [
a]
ꢂ42.1 (c 1.0, CHCl3). 1H NMR
D
(600 MHz, CDCl3): d 7.35–7.29 (m, 5H, Ar); 5.69 (d, 1H, J = 9.2 Hz,
NH); 5.14 (dd, 1H, J = 10.6, 9.2 Hz, H-3); 4.60 (d, 1H, J = 11.9 Hz,
CH-Ph); 4.55 (d, 1H, J = 11.9 Hz, CH–Ph); 4.45 (d, 1H, J = 8.3, H-1);
4.10 (m, 2H, ClCH2CO–); 3.94–3.90 (m, 1H, H-2); 3.81–3.73 (m,
3H, H-4, H-6a, H-6b); 3.46 (s, 3H, OCH3); 1.91 (s, 3H, COCH3). 13C
NMR (150 MHz, CDCl3): d 170.5, 168.3 (C@O); 137.4 (Ar quat);
128.6, 128.0, 127.8 (Ar); 101.63 (C-1); 77.3 (C-3); 73.8 (CH2Ph);
73.6 (C-3); 70.9 (C-4); 70.3 (C-6); 56.7 (OCH3); 53.9 (C-2); 40.9
(ClCH2CO–); 23.4 (COCH3). HRMS calcd for C18H24ClNO7 [M+H]+
402.1320. Found: 402.1323.
4.12.3. Analytical data for disaccharide 19
[a]
D ꢂ7.0 (c 1.0, CHCl3). 1H NMR (400 MHz, CDCl3): d 7.45–7.24
(m, 5H, Ar); 5.64 (d, 1H, J = 9.3 Hz, NH); 5.09 (dd, 1H, J = 10.4,
9.1 Hz, H-30); 5.97 (m, 2H, H-40, H-30); 4.81–4.74 (m, 2H, H-20,
PhCH); 4.49–4.36 (m, 4H, PhCH, H-6a, H-1, H-10); 4.19–3.89 (m,
5H, ClCH2CO-, H-6b, H-2, H-4); 3.72 (d, 2H, J = 2.31 Hz, H-6a0, H-
6b0); 3.50–3.38 (m, 4H, OCH3, H-5); 3.34 (m, 1H, H-50); 2.08, 1.98,
1.95, 1.93, 1.91 (4s, 4 ꢃ 3H, 4 ꢃ CH3). 13C NMR (100 MHz, CDCl3):
d 170.6, 170.1, 169.3, 169.1, 168.3 (C@O); 138.0 (Ar quat); 128.7,
128.2, 128.1 (Ar); 101.8 (C-10); 100.1 (C-1); 74.7 (C-4); 74.5 (C-3,
C-5); 73.7 (CH2Ph); 72.9 (C-30); 71.8 (C-50); 71.4 (C-20); 67.4 (C-
40); 67.1 (C-6); 61.0 (C-60); 56.7 (OCH3); 53.6 (C-2); 40.7
(ClCH2CO-); 23.3 (NCOCH3); 20.6, 20.5 (OCOCH3). HRMS calcd for
C33H44ClNO16 [M+H]+ 732.2319. Found: 732.2270.
4.11. Methyl 6-O-benzyl-3-O-chloroacetyl-2-deoxy-2-
methylacetimido-b-D-glucopyranoside (17)
A mixture of the acetamido 16 (450 mg, 1.12 mmol), CH2Cl2
(53 mL), and activated powdered MS 4 Å (2.65 g) was stirred at rt
for 1 h. MeOTf (3.2 mL, 28 mmol, 25 equiv) was added and the
mixture was stirred at rt for 3 h. The reaction mixture was cooled
to 0 °C and was quenched slowly with Et3N (4.6 mL, 34 mmol,
30 equiv). The reaction mixture was then filtered over CeliteÒ
and the solids were washed with CH2Cl2 (2 ꢃ 25 mL). The filtrate
and washings were combined and washed with satd aq NaHCO3
(80 mL). The aq layer was re-extracted with CH2Cl2 (20 mL), and
the combined organic phases were dried and concentrated. Col-
umn chromatography (30:1 CHCl3–MeOH) of the residue gave pure
4.13. Methyl 2-acetamido-4-O-(2,3,4,6-tetra-O-acetyl-b-
D-
glucopyranosyl)-6-O-benzyl-2-deoxy-b- -glucopyranoside (20)
D
A mixture of disaccharide 19 (30 mg, 0.04 mmol) and DABCO
(114 mg, 1.02 mmol, 26 equiv) in EtOH (2.6 mL) was stirred for
18 h at 55 °C. The reaction mixture was diluted with MeOH
(10 mL) and deionized with Dowex H+ resin. The resin was filtered
off, washed with MeOH (5 mL), and the combined filtrate and
washing was concentrated. Chromatography (50:1 CHCl3–MeOH)
of the residue gave pure acceptor 20 (18 mg, 71%) as a colorless
methyl imidate 17 (350 mg, 77%) as a colorless glass. [
a
]
D ꢂ14.1 (c
1.0, CHCl3). 1H NMR (400 MHz, CDCl3): d 7.32–7.24 (m, 5H, Ar);
5.14 (dd, 1H, J = 9.4, 9.5 Hz, H-3); 4.59 (q, 2H, J = 10.9 Hz, CH2Ph);
4.34 (d, 1H, J = 7.48 Hz, H-1); 4.00 (d, 2H, J = 2.5 Hz, ClCH2CO–);
3.83–3.73 (m, 3H, H-6a, H-6b, H-4); 3.61–3.53 (m, 4H, @COCH3,
H-5); 3.44 (s, 3H, OCH3); 3.31 (dd, 1H, J = 9.7, 7.7 Hz, H-2); 2.91
(br s, 1H, OH); 1.88 (s, 3H, COCH3). 13C NMR (100 MHz, CDCl3): d
167.3 (C@O); 147.7 (C@N); 137.5 (Ar quat); 128.5, 127.9, 127.8
(Ar); 103.9 (C-1); 79.5 (C-3); 73.8 (CH2Ph); 73.7 (C-5); 70.9 (C-4);
70.5 (C-6); 62.9 (C-2); 57.3, 54.1 (OCH3); 40.7 (ClCH2–); 15.7
(OCH3 imidate). HRMS calcd for C19H26ClNO7 [M+H]+ 416.1476.
Found: 416.1502.
glass. [
a]
ꢂ3.9 (c 1.0, CHCl3). 1H NMR (400 MHz, CDCl3): d 7.43–
D
7.29 (m, 5H, Ar); 5.67 (d, 1H, J = 8.0 Hz, NH); 5.10 (dd, 1H, J = 9.5,
9.6 Hz, H-30); 4.99 (dd, 1H, J = 9.9, 9.5 Hz, H-40); 4.93 (dd, 1H,
J = 9.6, 8.1 Hz, H-20); 4.70 (d, 1H, J = 12.1 Hz, CHPh); 4.63 (d, 1H,
J = 8.2 Hz, H-1); 4.49 (d, 1H, J = 9.1 Hz, H-10); 4.48 (d, 1H,
J = 12.0 Hz, CHPh); 4.14 (d, 2H, J = 3.3 Hz, H-6a0, H-6b0); 3.92 (dd,
1H, J = 9.7, 8.6 Hz, H-3); 3.72–3.59 (m, 4H, H-4, H-50, H-6a, H-
6b); 3.51–3.39 (m, 5H, OCH3, H-2, H-5); 2.08, 2.05, 2.01, 1.97,
1.94 (5s, 5 ꢃ 3H, 5 ꢃ COCH3). 13C NMR (100 MHz, CDCl3): d 170.6,
170.1, 169.3, 169.1 (C@O); 137.9 (Ar quat); 128.5, 127.9, 127.8
(Ar); 101.1 (C-1); 100.7 (C-10); 80.9 (C-4); 73.9 (C-5); 73.6 (CH2Ph);
72.6 (C-30); 71.9 (C-50); 71.5 (C-3); 71.1 (C-20); 68.1 (C-40); 67.9 (C-
6); 61.6 (C-60); 56.7 (OCH3); 56.7 (C-2); 23.7 (NCOCH3); 20.5
(OCOCH3). HRMS calcd for C30H41NO15 [M+H]+ 678.2402. Found:
678.2374.
4.12. Methyl 2-acetamido-4-O-(2,3,4,6-tetra-O-acetyl-b-D-
glucopyranosyl)-6-O-benzyl-3-O-chloroacetyl-2-deoxy-b-D-
glucopyranoside (19)
4.12.1. Glycosylation of the methyl imidate 17
BF3ꢀOEt2 (73
of methyl imidate 17 (60 mg, 0.144 mmol) and known20 glucosyl
donor 18 (711 mg, 1.44 mmol, 10 equiv) in CH2Cl2 (4 mL) stirred
at 40 °C. The reaction mixture was stirred for 18 h at 40 °C and
quenched by addition of Et3N (93 L, 0.64 mmol, 4.5 equiv). A mix-
ture of Ac2O and AcOH (1:1, 3 mL) was added and heated for 18 h
at 55 °C. The reaction mixture was diluted with CH2Cl2 (20 mL),
l
L, 0.58 mmol, 4.0 equiv) was added to a solution
4.14. Methyl 2-acetamido-4-O-(2,3,4,6-tetra-O-acetyl-b-
glucopyranosyl)-6-O-benzyl-3-O-(2,3,4-tri-O-benzyl-
fucopyranosyl)-2-deoxy-b- -glucopyranoside (22)
D-
a-L-
a
D
l
Disaccharide acceptor 20 (68 mg, 0.104 mmol) and known21
fucosyl donor 21 (99 mg, 0.208 mmol, 2.0 equiv) were dissolved
in a mixture of anhyd CH2Cl2 (1.5 mL) and anhyd DMF (1.5 mL)

J. L. Hendel et al. / Carbohydrate Research 343 (2008) 2914–2923
2923
containing activated powdered MS 4 Å (150 mg), and the mixture
Acknowledgments
was stirred for 1 h at rt. Cu(II)Br2 (46 mg, 0.208 mmol, 2.0 equiv)
and n-Bu4NBr (64 mg, 0.218 mmol, 2.1 equiv) were then added to
the reaction mixture and stirring was continued for 14 h at rt. The
reaction mixture was filtered over CeliteÒ and the solids were
washed with CH2Cl2 (40 mL). The combined filtrate and washings
were washed with brine (35 mL) and satd aq NaHCO3 (6 ꢃ 30
mL). The aq phases were re-extracted with CH2Cl2 (40 mL), and
the combined organic solutions were dried and concentrated.
Flash chromatography (3:2 EtOAc–hexanes) of the residue gave
The authors are grateful for financial support by NSERC, the
Canada Foundation for Innovation, and the Ontario Innovation
Trust.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.carres.2008.08.025.
pure trisaccharide 22 (77 mg, 70%) as a colorless glass. [a]
D
ꢂ33.3 (c 1.0, CHCl3). 1H NMR (400 MHz, CDCl3): d 7.36–7.26 (m,
20H, Ar); 6.05 (d, 1H, J = 7.7 Hz, NH); 5.15 (d, 1H, J = 3.7 Hz, H-
10); 5.09 (dd, 1H, J = 9.6, 9.5 Hz, H-300); 4.99–4.88 (m, 3H, H-200,
H-400, CHHPh); 4.84 (d, 1 H, J = 11.7 Hz, CHHPh); 4.80–4.70 (m,
2H, 2 CHHPh); 4.69–4.61 (m, 5H, H-1, H-100, 4 ꢃ CHHPh); 4.44
(d, 1H, J = 12.0 Hz, CHHPh); 4.33 (dd, 1H, J = 12.4, 4.3 Hz, H-6a00);
4.18 (q, 1H, J = 6.5 Hz, H-50); 4.12 (dd, 1H, J = 10.1, 3.7 Hz, H-20);
4.05 (dd, 1H, J = 6.4, 6.3 Hz, H-3); 3.97–3.84 (m, 4H, H-4, H-6a,
H-30, H-6b00); 3.79 (dd, 1H, J = 10.3, 4.0 Hz, H-6b); 3.73–3.63 (m,
2H, H-2, H-40); 3.59 (m, 1H, H-5); 3.40–3.33 (m, 4H, H-500,
OCH3); 2.02, 2.01, 2.00, 1.93 (4s, 4 ꢃ 3H, 4 ꢃ OCOCH3); 1.80
(NCOCH3); 1.16 (d, 3 H, J = 6.5, H-60). 13C NMR (100 MHz, CDCl3):
d 170.9. 170.6, 169.9 (C@O); 139.2, 139.0, 138.5 (Ar quat); 129.0,
128.9, 128.8, 128.6, 128.3, 128.1, 127.9, 127.6 (Ar); 101.3 (C-100);
99.6 (C-1); 97.1 (C-10); 79.9 (C-300); 78.1 (C-40); 77.5 (C-20); 75.1
(CH2Ph, C-20); 74.7 (C-4);74.6 (C-5); 73.8 (CH2Ph, C-6); 73.7 (C-
3); 73.6 (CH2Ph, C-30); 73.1 (CH2Ph, C-40); 72.9 (C-300); 72.3 (C-
500); 71.6 (C-200); 69.3 (C-6); 68.5 (C-400); 67.2 (C-50); 62.1 (C-600);
57.0 (OCH3); 53.2 (C-2); 23.6 (NCOCH3); 21.1, 21.0 (OCOCH3);
17.1 (C-60). HRMS calcd for C57H69NO19 [M+H]+ 1072.4581.
Found: 1072.4542.
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4.15. Methyl 2-acetamido-2-deoxy-3-O-(
a-L-fucopyranosyl)-4-
O-(b- -glucopyranosyl)-b- -glucopyranoside (2)
D
D
8. Crich, D.; Dudkin, V. J. Am. Chem. Soc. 2001, 123, 6818–6825.
Na (11 mg, 0.5 mmol) was reacted with MeOH (4 mL) at 0 °C.
The protected trisaccharide 22 (85 mg, 0.079 mmol) was dissolved
in 2 mL of anhyd MeOH and added to the solution of NaOMe in
MeOH. The reaction mixture was stirred at rt for 1 h, diluted with
MeOH (5 mL) and deionized with Dowex H+ resin. The resin was
filtered off and washed with MeOH (5 mL). The combined filtrate
and washing was concentrated to dryness, and MeOH (7 mL) and
10% Pd/C (250 mg) were added to the residue. The mixture was
stirred under H2 (100 psi) for 24 h and diluted with MeOH
(10 mL). Solids were filtered off, washed with MeOH (5 mL), and
the combined filtrate and washings were concentrated. The residue
was purified by gel permeation on a Biogel P2 column eluted with
water and gave pure trisaccharide 2 (35 mg, 83%), which was ob-
9. (a) Hindsgaul, O.; Norberg, T.; Le Pendu, J.; Lemieux, R. U. Carbohydr. Res. 1982,
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2008, in press.
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18. For monosaccharide acceptors see for example: (a) Herzner, H.; Kunz, H.
Carbohydr. Res. 2007, 342, 541–557; (b) Nicolaou, K. C.; Hummel, C. W.;
Bockovic, N. J.; Wong, C.-H. J. Chem. Soc., Chem. Commun. 1991, 870–872; For a
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tained as a white amorphous powder after freeze-drying. [a]
D
+35.5 (c 0.8, H2O). 1H NMR (600 MHz, D2O): d 5.16 (d, 1H,
J = 4.0 Hz, H-10); 4.87 (q, 1H, J = 6.6 Hz, H-50); 4.56 (d, 1H,
J = 7.9 Hz, H-100); 4.52 (d, 1H, J = 7.9 Hz, H-1); 4.05 (dd, 1H,
J = 12.3, 2.1 Hz, H-6a); 4.00–3.88 (m, 5H, H-2, H-4, H-3, H-30, H-
6b, H-6a00); 3.81 (d, 1H, J = 2.3 Hz, H-40); 3.75 (dd, 1H, J = 10.4,
4.0 Hz, H-20); 3.76–3.58 (m, 2H, H-5, H-6b00); 3.58–3.52 (m, 4H,
H-300, OCH3); 3.49–3.55 (m, 1H, H-500); 3.28–3.22 (m, 2H, H-200, H-
400); 2.07 (s, 3H, COCH3); 1.22 (d, 3H, J = 6.6, H-60). 13C NMR
(150 MHz, D2O): d 174.6 (C@O); 101.9 (C-1); 101.5 (C-100); 98.9
(C-10); 76.2 (C-500); 75.7 (C-300); 75.5 (C-5); 75.4 (C-3); 74.0 (C-4);
73.8 (C-200); 72.1 (C-40); 70.6 (C-400); 69.4 (C-30); 67.9 (C-20); 66.6
(C-50); 61.8 (C-600); 59.9 (C-6); 57.2 (OCH3); 55.8 (C-2); 22.4
(COCH3); 15.5 (C-60). HRESIMS calcd for C21H37NO15 [M+H]+
544.2264. Found: 544.2241.
19. (a) Roth, W.; Pigman, W. J. Am. Chem. Soc. 1960, 82, 4608–4611; (b) Hesek, D.;
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ˇ
. Synthesis
21. Lönn, H. Carbohydr. Res. 1985, 139, 105–113; Ruttens, D.; Kovác, P
2004, 2505–2508.
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