Synthesis of a core trisaccharide as a versatile building block for N-glycans and glycoconjugates

Article abstract of DOI:10.1002/chem.200390156

N-Linked oligosaccharides from glycoproteins (N-glycans) can be conveniently assembled with a building block approach. A protected form of the core trisaccharide (β-mannosyl chitobiose) was identified as a key building block. The chitobiose part of the co

Full text of DOI:10.1002/chem.200390156

FULL PAPER
Synthesis of a Core Trisaccharide as a Versatile Building Block
for N-Glycans and Glycoconjugates
Carlo Unverzagt*[a,b]
Dedicated to Professor Lutz Tietze on the occasion of his 60th birthday
Abstract: N-Linked oligosaccharides
from glycoproteins (N-glycans) can be
conveniently assembled with a building
block approach. A protected form of the
core trisaccharide (b-mannosyl chito-
biose) was identified as a key building
block. The chitobiose part of the core
trisaccharide was built from a glycosyl
fluoride, which served as a precursor for
the reducing GlcNAc azide and the
inner GlcNAc moiety. b-Mannosylation
was accomplished at the trisaccharide
stage by intramolecular inversion of a b-
glucosyl chitobiose. The benzylidene
protection of the b-mannoside and the
azido group at the reducing end of the
core trisaccharide allow modular syn-
thesis of N-glycans and their glycocon-
jugates.
Keywords: b-mannosides ¥ carbo-
hydrates
¥
glycosylation
¥
glyco-
proteins ¥ oligosaccharide
9]
N-glycans,[7 the last decade has seen considerable increase
Introduction
in activity.[10
Up to now several strategies have been
15]
Asparagine-linked oligosaccharides (N-glycans) are a com-
mon feature of eucaryotic proteins that are surface bound or
secreted.[1] The biological properties of glycoproteins are
influenced by the structure of their oligosaccharides. Despite
a wide knowledge of structural details of N-glycans little is
known about detailed structure activity relationships of
entire glycoproteins.[2] One of the few glycoproteins studied
thoroughly is recombinant human erythropoietin (EPO), used
in the treatment of cancer and kidney patients. The serum
halflife of EPO and its modification (NESP) can be tuned by
the extent of N-glycosylation.[3]
developed leading to complete N-glycans or partial struc-
tures.[16] The key challenges of the syntheses have been the
construction of the b-mannoside[17] followed by the incorpo-
ration of terminal sialic acid and the deprotection of the final
products. Besides the chemical approaches the core trisac-
charide has also been synthesized by enzymatic methods.[18]
Results and Discussion
A combinatorial biochemical machinery is installing N-gly-
cans as a posttranslational modification leading to a large
variety of possible oligosaccharide structures. The resulting
glycoproteins are therefore heterogeneous mixtures of indi-
vidual glycoforms. The diversity of natural N-glycans is thus
impairing the isolation of these compounds from glycopro-
The total synthesis of N-glycans and their derivatives can be
facilitated by combining chemical and enzymatic ap-
proaches.[19] Our initial goal in N-glycan synthesis was the
heptasaccharide asparagine conjugate 1,[20] which can serve
both as a building block for glycopeptides[21] and as an
acceptor for enzymatic elongation of the sugar chains.[19]
Retrosynthetic analysis of 1 (Scheme 1) suggested a division
into building blocks of general applicability.[22] A protected
core trisaccharide was desired which would permit good
coupling to various building blocks for the antennae and allow
the convenient construction of glycoconjugates through an
amino function at the reducing end. Furthermore, the
protected core trisaccharide should be available in amounts
over 10 g on a typical laboratory scale to yield sufficient
quantities of N-glycans after deprotection. The building block
meeting these requirements was found in trisaccharide 2 (see
Scheme 1). An optimized approach to this versatile com-
pound is described in the following.
6]
teins.[4 Therefore, the synthesis of N-glycans has been used
to overcome the shortage of material for biological studies.
While only few groups pioneered the chemical synthesis of
[a] Prof. Dr. C. Unverzagt
Institut f¸r Organische Chemie und Biochemie
Technische Universit‰t M¸nchen
Lichtenbergstrasse 4, 85748 Garching (Germany)
[b] Current address:
Bioorganische Chemie
Universit‰t Bayreuth
Geb‰ude NW1, 95440 Bayreuth (Germany)
Fax : (‡49)921-55-5365
E-mail: Carlo.Unverzagt@uni-bayreuth.de
Chem. Eur. J. 2003, 9, No. 6
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1369

C. Unverzagt
FULL PAPER
Rf value of compounds 8 and 10
required monitoring of the re-
1
action by H NMR spectrosco-
py. The cleavage appears to be
sterically hindered[33] and led to
phthalimido ring opening dur-
ing the prolonged reaction
time. Similar difficulties occur-
red for the deacetylations at the
di- and trisaccharide stage,
which suggested the synthesis
of an alternative set of building
blocks carrying chloroacetyl
groups. After introduction of
the chloroacetyl moiety[34] (5)
the following transformations
to fluoride 7 and azide 9 pro-
Scheme 1. Retrosynthetic analysis of the heptasaccharide-asparagine 1 suggests the disconnection to the core
trisaccharide 2 as the key building block for N-glycans.
Synthesis of chitobiosylazide: The early introduction of an
azido functionality[23, 24] at the reducing end of the core
trisaccharide 2 is advantageous because this moiety remains
neutral under most reaction conditions applied in carbohy-
drate chemistry. Glycosyl azides can also be converted into
glycosyl donors after derivatization.[25, 26] The stable azido
group at the anomeric center protects the labile amino
function required for the attachment of aspartic acid[27] or
spacers. Glycosyl azides are obtained from the corresponding
fluorides[28] in high yield. The latter can be generated
conveniently from thioglycosides.[29, 30]
The starting material for the intermediate chitobiosyl azide
13 (Scheme 2) was the thioglycoside 3,[31, 32] a useful inter-
mediate, which is accessible on a 100 g scale in a five-step
sequence starting from glucosamine hydrochloride. First, the
hydroxyl group was acetylated and the resulting thioglycoside
4 was converted to fluoride 6 with N-bromosuccinimide
(NBS) and HF/pyridine according to Nicolaou.[29] When
activated with BF3 ¥ Et2O the fluoride[28] 6 readily reacted
with trimethylsilylazide to give the b-azide 8. The deacetyla-
tion using potassium methoxide in dioxane/methanol 1:1
proceeded slowly and gave only 67% yield. A nearly identical
ceeded analogously in yields over 90%. With a base lability
increased by three orders of magnitude,[35] a dechloroacety-
lation occurs much faster than the cleavage of the corre-
sponding acetate. The improved cleavage conditions permit-
ted the deprotection of compound 9 to furnish the acceptor 10
in 92% yield.
Acceptor 10 was treated with the glycosylfluorides 6 and 7
activated by BF3 ¥ Et2O to give the chitobiosyl azides 11 and 12
in good yields. In the following deacylation step the chlor-
oacetyl moiety could be readily removed. Compound 13 is
equipped with a combination of protective groups suitable for
the construction of the b-mannoside. The neighboring benzyl
groups increase the reactivity[36] of the OH-4'' group; the
phthalimido groups are known to be inert during nucleophilic
substitutions and glycosylation reactions.
Synthesis of b-mannosides: The b-mannosidic linkage[17] was
established following a procedure introduced by Kunz.[32, 37, 38]
This method is based on the intramolecular inversion of 4,6-
benzylidenated b-glucosides, which excludes unwanted
anomeric or epimeric side products. For the synthesis of
N-glycans with title compound 2 it was highly advantageous
that the b-mannosyl moiety was
not blocked by permanent pro-
tective groups, for example
benzyl ethers, but could be
easily debenzylidenated at the
desired time under mild condi-
tions.[32]
To introduce the b-glucosyl
moiety the Kunz group used a
glucosyl bromide,[32] which gave
also orthoesters under glycosy-
lation conditions. Therefore, we
converted peracetylated 3-phe-
nylcarbamoyl glucose 14[32] to
the anomeric fluoride 15 by
reaction with HF/pyridine
(Scheme 3).[39] Elongation of
the chitobiosyl acceptor 13 with
the donor 15 under BF3 ¥ Et2O
activation yielded 85% of the
OBzl
OBzl
OBzl
c
d
O
O
O
RO
BzlO
RO
BzlO
RO
BzlO
SEt
F
N3
NPht
NPht
NPht
8: R = Ac
9: R = CA
6: R = Ac
3: R = H
a
b
7: R = CA
4: R = Ac
e
f
10: R = OH
5: R = CA
g
h
OBzl
O
6 + 10
7 + 10
11: R = Ac
12: R = CA
13: R = H
NPht
O
BzlO
RO
O
BzlO
N3
i
k
NPht
OBzl
11-13
Scheme 2. a) Ac2O/pyridine (quant.); b) chloroacetic anhydride/pyridine, CH2Cl2 (93%); c) HF/pyridine, NBS,
CH2Cl2 (93% 6), (96% 7); d) TMS-N3, BF3 ¥ OEt2, CH2Cl2 (91% 8), (92% 9); e) MeOH, dioxane, NaOMe
(67%); f) K2CO3, MeOH, CH2Cl2 (92%); g) BF3 ¥ OEt2, CH2Cl2 (83%); h) BF3 ¥ OEt2, CH2Cl2 (86%); i) K2CO3,
MeOH, dioxane (79%); k) K2CO3, MeOH, CH2Cl2 (94%).
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Chem. Eur. J. 2003, 9, No. 6

Core Trisaccharide Building Blocks
1369 1376
nylcarbamoyl glucose 18 could not be acetalized selectively
with benzaldehyde dimethylacetal. However, the traditional
procedure with benzaldehyde in the presence of zinc chloride
avoided the overreaction and gave the crystalline benzylidene
acetal 19. After chloroacetylation the anomeric center was
liberated chemoselectively with hydrazine acetate and con-
verted to the trichloroacetimidate 20 according to Schmidt.[42]
The imidate 20 could be converted to the b-fluoride 21, which
was formed stereoselectively through neighboring-group
participation.
OAc
OAc
a
O
O
AcO
AcO
H
H
OAc
O
O
N
N
F
OAc
OAc
O
O
14
15
b
OAc
OBzl
NPht
O
O
BzlO
O
BzlO
H
AcO
O
N3
O
N
O
O
NPht
OAc
Initial experiments to glycosylate the chitobiosyl azide 13
with the imidate 20 showed only little conversion when BF3 ¥
Et2O was employed. Use of trimethylsilyl triflate[42]
(TMSOTf) gave the b-glucoside 22 in 62% yield. The quality
of the reagent was crucial for the success of the reaction. Only
freshly opened trimethylsilyl triflate could suppress orthoest-
er formation, which suggests that the deactivating 2-chloroa-
cetyl moiety allows for the reaction to proceed via an
intermediate orthoester. Furthermore, the yields were low-
ered by trimethylsilylation of the acceptor 13, which was
desilylated and recycled after large-scale reactions. These
problems were avoided using the fluoride 21 as a donor. The
glucosyl fluoride could be activated with BF3 ¥ Et2O and did
not show formation of the orthoester. From compounds 21
and 13 the chloroacetylated trisaccharide 22 was obtained in
45% yield, which was conveniently deacylated (17) by mild
base treatment. For the removal of the chloroacetyl moiety
alternative methods[43] are known, however, the rapid and
easily adjustable procedure favored the use of potassium
carbonate in methanol/dichloromethane.
The conversion of the b-gluco configured trisaccharide 17
to the b-mannoside 2 was achieved by a four-step procedure.
This reaction sequence was previously established for mono-
or disaccharides[32] and was successfully applied to the
trisaccharide 17. First, the 2''-OH function was converted to
a good leaving group with trifluoromethanesulfonic acid
anhydride and pyridine (Scheme 5). The intermediate triflate
17a was substituted by an intramolecular nucleophilic attack
of the neighboring carbonyl group of the phenylcarbamoyl
moiety. This was achieved by warming a solution of the triflate
in DMF/pyridine to 658C. The initially formed b-manno
configured iminocarbonate 17b was unstable and was con-
OBzl
16
c
OBzl
NPht
O
O
O
O
BzlO
O
O
O
BzlO
H
N3
O
N
NPht
OH
O
OBzl
17
Scheme 3. a) HF/pyridine (56%); b) 13, BF3 ¥ OEt2, CH2Cl2 (85%);
c) 1) K2CO3, MeOH, dioxane, 2) benzaldehyde dimethylacetal, p-TosOH,
CH3CN (44% over two steps).
b-gluco configured trisaccharide 16. Prior to the inversion of
the b-glucoside to the b-mannoside the 2-OH'' group had to
be selectively activated. Removal of the acetates and regio-
selective benzylidenation of the intermediate triol gave the
trisaccharide 17. This compound was suitably functionalized
for the ensuing inversion sequence to the b-mannoside.
Unfortunately, compound 17 was obtained in only 44%
yield after the two-step reaction. The main reason was again a
difficult removal of acetyl groups, in particular of the sterically
hindered 2''-acetate.[40] It is known that the deprotection of
esters in the vicinity of sterically demanding residues is
impaired. During the prolonged reaction times (1.5 d) the
base labile phthalimido groups were also cleaved.
To circumvent these difficulties the novel b-glucosyl donor
20 (Scheme 4) was constructed, which was already equipped
with a benzylidene acetal and carried an easily removable
chloroacetyl group[41] in position 2. Thus, the critical change of
protective groups at the trisaccharide stage was avoided. In
contrast to 3-phenylcarbamoyl b-glucosides, the free 3-phe-
OH
O
O
O
O
O
HO
O
O
H
H
O
H
c
a
F
O
O
N
N
O
OH
OR1
N
OH
OR2
CAO
O
O
19: R1=R2=H
18
21
b
20: R1=C(NH)CCl3, R2=CA
d
20 + 13
21 + 13
OBzl
NPht
O
O
O
CAO
O
BzlO
O
O
O
f
O
BzlO
H
N3
17
O
N
NPht
OBzl
e
22
Scheme 4. a) Benzaldehyde, ZnCl2 (34%); b) 1) chloroacetic anhydride/pyridine, CH2Cl2, 2) hydrazine acetate, DMF; 3) trichloroacetonitrile, DBU,
CH2Cl2 (over three steps 55%); c) HF/pyridine, CH2Cl2 (91%); d) TMSOTf, CH2Cl2 (62%); e) BF3 ¥ OEt2, CH2Cl2 (45%); f) K2CO3, MeOH/CH2Cl2 (89%).
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C. Unverzagt
FULL PAPER
performed on silica gel 60 (230 400 mesh, Merck Darmstadt). The
reactions were monitored by thin-layer chromatography on coated
aluminum plates (silica gel 60 GF254, Merck Darmstadt). Spots were
detected by UV light or by charring with a 1:1 mixture of 2n H2SO4 and
0.2% resorcine monomethyl ether in ethanol.
OBzl
NPht
O
O
O
O
BzlO
O
O
H
O
BzlO
N3
O
N
NPht
OH
O
OBzl
Ethyl 4-O-acetyl-3,6-di-O-benzyl-2-deoxy-2-phthalimido-1-thio-b-d-gluco-
pyranoside (4): Compound 3[31, 32] (13.31 g, 24.9 mmol) was dissolved in
pyridine (20 mL) and acetic anhydride (10 mL). After 20 h at ambient
temperature the reaction mixture was concentrated and codistilled three
times with toluene (50 mL). The residue was dried in high vacuo to yield 4
(14.35 g, quant.). [a]2D3 ˆ ‡77.58 (c ˆ 1.0 in CH2Cl2); Rf ˆ 0.34 (hexane/
acetone 2:1); 1H NMR (250 MHz, [D6]DMSO): d ˆ 7.87 7.79 (m, 4H, Pht),
7.25 (m, 5H, Ph), 6.92 (m, 5H, Ph), 5.30 (d, J1,2 ˆ 10.4 Hz, 1H, H-1b), 5.00
17
1.
OBzl
NPht
O
O
O
O
BzlO
O
O
O
H
N
N3
O
BzlO
O
NPht
17a
OTf
OBzl
(dd, J4,5 ˆ 9.8 Hz, 1H, H-4), 4.56 4.44 (m, 3H, CH2O), 4.35 (dd, J3,4
8.1 Hz, 1H, H-3), 4.26 (d, Jgem ˆ 12.2 Hz, 1H, CH2O), 4.05 (dd, J2,3
ˆ
ˆ
2.
N
10.4 Hz, 1H, H-2), 3.90 3.83 (m, 1H, H-5), 3.59 3.47 (m, 2H, H-6a,b),
2.57 (m, 2H, SCH2), 2.02 (s, 3H, OAc), 1.09 (t, J ˆ 7.4 Hz, CH3); 13C NMR
O
O
OBzl
O
NPht
O
ˆ
ˆ
O
(62.5 MHz, [D6]DMSO): d ˆ 168.8 (C O Ac), 166.9, 166.5 (C O, Pht),
137.6, 136.9 (C-i Ph), 134.3 (C-4/5 Pht), 130.1 (C-1/2 Pht), 127.6, 127.4, 127.0,
126.8 (C-Ar), 122.9 (C-3/6 Pht), 80.0 (C-1), 77.1 (C-3), 76.1 (C-5), 73.0, 71.08
(CH2O), 70.9 (C-4), 68.2 (C-6), 53.9 (C-2), 23.4 (CH2), 20.7 (OAc), 15.0
(CH3); elemental analysis calcd (%) for C32H33NO7S (575.68): C 66.77, H
5.78, N 2.43; found: C 66.69, H 5.56, N 2.57.
O
BzlO
O
O
BzlO
N3
O
NPht
OBzl
17b
3.
O
4-O-Acetyl-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosyl-
fluoride (6): Thioglycoside 4 (12.6 g, 21.9 mmol) was dissolved in dichloro-
methane (130 mL) and cooled to 08C. N-Bromosuccinimide (5.83 g,
32.7 mmol) was added to the stirred solution followed by HF/pyridine
complex (12.6 mL). The reaction was complete after 10 min (TLC: hexane/
acetone 1.5:1). Subsequently, dichloromethane (130 mL) was added and
the reaction mixture was extracted in a Teflon separatory funnel with
water, aq HCl (3 Â ) and aq Na2CO3. The organic phase was dried (MgSO4),
concentrated and purified by flash chromatography (hexane/acetone 2:1)
to yield 6 (10.83 g, 92.7%). The product was recrystallized from acetone/
hexane. M.p. 1008C; Rf ˆ 0.45 (hexane/acetone 1.5:1); [a]2D3 ˆ ‡76.48 (c ˆ
0.5 in CH2Cl2); 1H NMR (250 MHz, [D6]DMSO): d ˆ 7.88 (m, 4H, Pht),
7.35 (m, 5H, Ph), 6.93 (m, 5H, Ph), 5.93 (dd, J1,2 ˆ 8.0, J1,F ˆ 53.6 Hz, 1H,
H-1b), 5.10 (dd, J4,5 ˆ 9.5 Hz, 1H, H-4), 4.57 4.44 (m, 3H, CH2O), 4.39
(dd, J2,3 ˆ J3,4 ˆ 9.9 Hz, 1H, H-3), 4.27 (d, Jgem ˆ 12.0 Hz, 1H, CH2O), 4.14
O
O
OBzl
O
NPht
O
O
O
BzlO
O
O
N3
O
BzlO
NPht
OBzl
17c
4.
OBzl
OH
O
NPht
O
O
O
BzlO
O
O
HO
O
BzlO
N3
NPht
OBzl
2
Scheme 5. 1) Tf2O, pyridine, CH2Cl2; 2) DMF, pyridine, 658C; 3) AcOH,
dioxane, water; 4) NaOMe, MeOH, CH2Cl2 (overall yield 70%).
(m, 1H, H-2), 4.04 (m, 1H, H-5), 3.58 (m, 2H, H-6a,b), 2.05 (s, 3H, OAc);
13
ˆ
C NMR (62.5 MHz, [D6]DMSO): d ˆ 169.2, 167.2 (C O), 137.9, 137.3 (C-i
Ph), 134.9 (C-4/5 Pht), 130.5 (C-1/2 Pht), 128.2 127.5 (C-Ar), 123.5 (C-3/6
Pht), 104.0 (d, JC-1,F ˆ 212.3 Hz, C-1), 75.9 (d, JC-3,F ˆ 9.7 Hz, C-3), 73.4
verted to the carbonate 17c by mild acid hydrolysis. Depro-
tection of the carbonate 17c by methanolysis gave the desired
core trisaccharide 2 in 70% yield over the reaction se-
quence.[22] The inversion at C-2'' was conducted as a one-pot
reaction and every step was monitored by TLC. NMR
spectroscopy confirmed the newly formed b-mannosidic
linkage via the C-1''/H-1'' coupling constant of 163.3 Hz[44, 45]
determined from an HMQC spectrum without decoupling.[46]
Following this approach the protected core trisaccharide 2
was obtained in overall amounts of more than 10 g. This
building block has proven to be a key compound in the
synthesis of several families of natural N-glycans[15, 47] with
different substitution patterns.
(CH2O), 72.5 (C-5), 72.4 (CH2O), 70.7 (C-4), 68.1 (C-6), 55.1 (d, JC-2,F
ˆ
21.8 Hz, C-2), 20.6 (OAc); elemental analysis calcd (%) for C30H28FNO7
(533.55): C 67.53, H 5.29, N 2.63; found: C 67.62, H 5.48, N 2.52.
4-O-Acetyl-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosyl-
azide (8): Trimethylsilylazide (2.35 mL, 17.7 mmol) was added to a stirred
solution of fluoride 6 (4 g, 7.5 mmol) in dry dichloromethane (70 mL). The
reaction was initiated with BF3 ¥ OEt2 (300 mL, 2.4 mmol). After complete
reaction (TLC: hexane/acetone 1.5:1) dichloromethane was added fol-
lowed by extraction with dilute K2CO3. The organic phase was dried
(MgSO4), evaporated and purified by flash chromatography (hexane/
acetone 2:1) to afford
8
(3.81 g, 91.3%). [a]2D3 ˆ ‡49.58 (c ˆ 0.5 in
dichloromethane); Rf ˆ 0.48 (hexane/acetone 1.5:1); 1H NMR (250 MHz,
[D6]DMSO): d ˆ 7.9 7.7 (m, 4H, Pht), 7.4 7.25 (m, 5H, Ph), 6.94 (m, 5H,
Ph), 5.54 (d, J1,2 ˆ 9.5 Hz, 1H, H-1b), 5.04 (dd, J3,4 ˆ J4,5 ˆ 9.5 Hz, 1H, H-4),
4.57 4.47 (m, 3H, CH2O), 4.37 (dd, J2,3 ˆ 10.1 Hz, 1H, H-3), 4.26 (d, Jgem
ˆ
12.0 Hz, 1H, CH2O), 3.98 (m, 1H, H-5), 3.94 (dd, 1H, H-2), 3.6 3.5 (m,
2H, H-6a,b), 2.03 (s, 3H, OAc); 13C NMR (125 MHz, [D6]DMSO): d ˆ
ˆ
169.3, 167.0 broad (C O), 138.1, 137.4 (C-i Ph), 134.9 (C-4/5 Pht), 130.6 (C-
1/2 Pht), 128.2, 127.9, 127.6, 127.50, 127.47, 127.42 (C-Ar), 123.5 (C-3/6 Pht),
85.0 (C-1), 76.4 (C-3), 74.6 (C-5), 73.5 (CH2O), 71.1 (C-4), 72.4 (CH2O),
71.1 (C-4), 68.3 (C-6), 54.6 (C-2), 20.7 (OAc); elemental analysis calcd (%)
for C30H28N4O7 (556.57): C 64.74, H 5.07, N 10.07; found: C 64.71, H 5.00, N
10.00.
Experimental Section
General methods: Solvents were dried according to standard methods.
Melting points were determined on a B¸chi apparatus and are uncorrected.
Optical rotations were measured on a Perkin Elmer 241 polarimeter at
589 nm. NMR spectra were recorded on Bruker AC 250 and AMX 500
instruments. Coupling constants are reported in Hz. For mass spectra a
Varian CH5 instrument was used in the fast atom bombardment mode
(FAB) with a thioglycerine/HOAc (MB) or a m-nitrobenzylalcohol matrix
(NBA). ESI-TOF mass spectra were recorded on a Micromass LCT
instrument coupled to an Agilent 1100 HPLC. Flash chromatography was
3,6-Di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosylazide
(10):
a) Startingfrom 8: NaOMe (100 mg) was added to a stirred solution of
acetate 8 (3.7 g, 6.65 mmol) in dioxane/methanol 1:1 (40 mL). The course
of the reaction was monitored by 1H NMR spectroscopy following the
disappearance of the signal of the acetyl group. Two further portions of
sodium methylate (100 mg each) were added after 30 min intervals. After
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Chem. Eur. J. 2003, 9, No. 6

Core Trisaccharide Building Blocks
1369 1376
complete reaction the solution was neutralized with the acidic ion exchange
resin Amberlyst 15H , filtered and concentrated. The residue was purified
by flash chromatography (hexane/acetone 2.2:1) and gave 10 (2.3 g,
67.2%).
washed with aq K2CO3, dried (MgSO4) and concentrated. The residue was
purified by flash chromatography (hexane/acetone 2:1) to yield 9 (38.2 g,
91.8%). Crystals were obtained by trituration of the product with
methanol. M.p. 86 À 918C; [a]2D3 ˆ ‡45.08 (c ˆ 0.5 in dichloromethane);
Rf ˆ 0.37 (hexane/acetone 2:1); 1H NMR (250 MHz, [D6]DMSO): d ˆ 7.85
(m, 4H, Pht), 7.35 (m, 5H, Ph), 6.92 (m, 5H, Ph), 5.57 (d, J1,2 ˆ 9.4 Hz, 1H,
H-1b), 5.15 (dd, J4,5 ˆ 9.5 Hz, 1H, H-4), 4.6 4.4 (m, 6H, CH2O, CH2Cl,
‡
b) Startingfrom 9: K2CO3 (200 mg) was added to a stirred solution of
chloroacetate
9 (12.0 g, 20.3 mmol) in methanol/dichloromethane 1:2
(120 mL). After complete reaction (TLC: hexane/ethyl acetate 1.5:1) the
solution was filtered, neutralized with acidic acid and concentrated. The
residue was purified by flash chromatography (hexane/ethyl acetate 5:3)
and gave 10 (9.6 g, 91.9%). [a]2D3 ˆ ‡17.28 (c ˆ 0.5 in CH2Cl2); Rf ˆ 0.45
(hexane/ethyl acetate 1.5:1); 1H NMR (500 MHz, [D6]DMSO): d ˆ 7.9 7.7
H-3), 4.25 (d, Jgem ˆ 12.0 Hz, 1H, CH2O), 4.03 (m, 1H, H-5), 3.97 (dd, J2,3
ˆ
10.6 Hz, 1H, H-2), 3.63 (m, 2H, H-6a,b); 13C NMR (62.5 MHz,
ˆ
[D6]DMSO): d ˆ 167.2, 166.4 (C O, ClAc, Pht), 138.0, 137.3 (C-i Ph),
134.9 (C-4/5 Pht), 130.5 (C-1/2 Pht), 128.2, 127.9, 127.5 (C-Ar), 123.5 (C-3/6
Pht), 84.9 (C-1), 76.2 (C-3), 74.3 (C-5), 73.6 (CH2O), 72.6 (C-4), 72.4
(CH2O), 68.0 (C-6), 54.5 (C-2), 40.9 (CH2Cl); ESI-MS: m/z: calcd for
(m, 4H, Pht), 7.4 7.33 (m, 5H, Ph), 6.93 6.86 (m, 5H, Ph), 5.74 (d, J4,OH
ˆ
7.2 Hz, 1H, OH-4), 5.42 (d, J1,2 ˆ 9.5 Hz, 1H, H-1b), 4.78 (d, Jgem ˆ 12.2 Hz,
1H, CH2O), 5.58 (s, 2H, CH2O), 4.42 (d, Jgem ˆ 12.2 Hz, 1H, CH2O), 4.14
(dd, J3,4 ˆ J2,3 ˆ 9.6 Hz, 1H, H-3), 3.86 3.82 (m, 2H, H-6a, H-2), 3.75 3.68
(m, 2H, H-6b, H-5), 3.54 (ddd, J4,5 ˆ 8.5 Hz, 1H, H-4); 13C NMR (125 MHz,
‡
C30H27ClN4O7: 590.16; found: 613.26 [M‡Na] .
O-(4-O-Acetyl-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyrano-
syl)-(1 ! 4)-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosyl-
azide (11): A suspension of azide 10 (3.0 g, 5.8 mmol), fluoride 6 (3.9 g,
7.3 mmol) and ground molecular sieves 4 ä (7 g) in dry dichloromethane
(70 mL) was stirred for 30 min at room temperature. BF3 ¥ OEt2 (150 mL,
1.2 mmol) was added and the reaction was allowed to proceed for 1 h
(TLC: hexane/acetone 1.5:1). The suspension was filtered over Celite,
washed with dichloromethane followed by extraction with aq Na2CO3. The
organic phase was dried (MgSO4), concentrated and the residue was
purified by flash chromatography (hexane/acetone 2.3:1) to give 11 (5.0 g,
83.4%). [a]2D3 ˆ ‡26.58 (c ˆ 0.5 in CH2Cl2); Rf ˆ 0.42 (hexane/acetone
1.5:1); 1H NMR (500 MHz, [D6]DMSO): d ˆ 8.0 7.7 (m, 8H, Pht), 7.4 7.2
(m, 10H, Ph), 6.95 6.80 (m, 10H, Ph), 5.31 (d, J1,2 ˆ 7.2 Hz, 1H, H-1'b),
5.29 (d, J1,2 ˆ 9.0 Hz, 1H, H-1b), 5.01 (dd, J3,4 ˆ J4,5 ˆ 9.5 Hz, 1H, H-4'), 4.81
(d, Jgem ˆ 12.3 Hz, 1H, CH2O), 4.54 4.36 (m, 7H, CH2O, H-3'), 4.26 (d,
Jgem ˆ 12.0 Hz, 1H, CH2O), 4.12 4.07 (m, 3H, H-2', H-3, H-4), 3.80 (dd,
J2,3 ˆ 10.6 Hz, 1H, H-2), 3.62 3.54 (m, 3H, H-5, H-5', H-6a'), 3.49 (d,
Jgem ˆ 10.9 Hz, 1H, H-6a), 3.41 (dd, Jvic ˆ 5.5, Jgem ˆ 11.2 Hz, 1H, H-6b'),
3.37 (dd, Jvic ˆ 3.8 Hz, 1H, H-6b), 2.03 (OAc); 13C NMR (125 MHz,
ˆ
[D6]DMSO): d ˆ 167.3 (C O), 138.4, 138.1 (C-i Ph), 134.7 (C-4/5 Pht), 130.6
(C-1/2 Pht), 128.2 127.1 (C-Ar), 123.5 (C-3/6 Pht), 84.9 (C-1), 78.3 (C-3),
77.8 (C-5), 73.6, 72.3 (CH2O), 71.1 (C-4), 68.9 (C-6), 54.6 (C-2); elemental
analysis calcd (%) for C28H26N4O6 (514.53): C 65.36, H 5.09, N 10.89; found:
C 65.45, H 5.33, N 10.78.
Ethyl 3,6-di-O-benzyl-4-O-chloroacetyl-2-deoxy-2-phthalimido-1-thio-b-
d-glucopyranoside (5): Compound 3[31, 32] (97 g, 181.8 mmol) and pyridine
(100 mL) were dissolved in dichloromethane (500 mL) and cooled to 08C.
Chloroacetic acid anhydride (45 g, 263.2 mmol) was added and the reaction
was maintained at 08C until the starting material was consumed (TLC:
hexane/ethyl acetate 1:1). Subsequently, the solution was diluted with
dichloromethane (500 mL) and extracted with dilute K2CO3, dilute HCl
(3 Â ) and dilute K2CO3. The organic phase was dried (MgSO4), concen-
trated and purified by flash chromatography (hexane/acetone 2:1) to give 5
(103.4 g, 93.2%). [a]2D3 ˆ ‡50.98 (c ˆ 1 in dichloromethane); Rf ˆ 0.66
(hexane/EtOAc 1:1); 1H NMR (250 MHz, [D6]DMSO): d ˆ 7.9 7.7 (m,
4H, Pht), 7.34 (m, 5H, Ph), 6.92 (m, 5H, Ph), 5.30 (d, J1,2 ˆ 10.4 Hz, 1H,
H-1b), 5.10 (dd, J4,5 ˆ 9.5 Hz, 1H, H-4), 4.6 4.35 (m, 6H, CH2O, CH2Cl,
H-3), 4.26 (d, Jgem ˆ 12.2 Hz, 1H, CH2O), 4.07 (dd, J2,3 ˆ 10.3 Hz, 1H, H-2),
3.92 (m, 1H, H-5), 3.58 (m, 2H, H-6a,b), 2.68 2.47 (m, 2H, SCH2), 1.09 (t,
J ˆ 7.4 Hz, CH3); 13C NMR (62.5 MHz, [D6]DMSO): d ˆ 167.4, 167.0, 166.4
ˆ
[D6]DMSO): d ˆ 169.4, 167.1 (C O), 138.2, 138.0, 137.9, 137.4 (C-i Ph),
134.9, 134.8 (C-4/5 Pht), 130.6 (C-1/2 Pht), 128.2 127.0 (C-Ar), 123.5 (C-3/6
Pht), 96.4 (C-1'), 84.8 (C-1), 76.3 (C-3'), 76.1 (C-3), 75.6 (C-5), 75.1 (C-4),
73.7, 73.5 (CH2O), 73.0 (C-5'), 72.5, 71.7 (CH2O), 71.5 (C-4'), 68.3 (C-6'),
67.6 (C-6), 55.7 (C-2'), 54.6 (C-2), 20.6 (OAc); elemental analysis calcd (%)
for C58H53N5O13 Â 0.5H2O (1028.08): C 67.17, H 5.25, N 6.75; found: C 67.07,
H 5.07, N 6.42.
ˆ
(C O), 138.4, 137.4 (C-i Ph), 134.9 (C-4/5 Pht), 130.6 (C-1/2 Pht), 128.2
127.4 (C-Ar), 123.5 (C-3/6 Pht), 80.5 (C-1), 77.5 (C-3), 76.3 (C-5), 73.7
(CH2O), 74.1 (C-4), 72.5 (CH2O), 68.5 (C-6), 54.4 (C-2), 41.0 (CH2Cl), 23.5
(CH2), 15.1 (CH3); elemental analysis calcd (%) for C32H32ClNO7S
(610.12): C 63.0, H 5.29, N 2.30; found: C 62.85, H 5.48, N 2.07.
O-(3,6-Di-O-benzyl-4-O-chloroacetyl-2-deoxy-2-phthalimido-b-d-gluco-
pyranosyl)-(1 ! 4)-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyra-
nosylazide (12): A suspension of azide 10 (13.0 g, 25.3 mmol), fluoride 7
(17.0 g, 29.9 mmol) and ground molecular sieves 4 ä (22 g) in dry dichloro-
methane (350 mL) was stirred for 30 min at room temperature. BF3 ¥ OEt2
(1.46 mL, 11.9 mmol) was added and the reaction was allowed to proceed
for 1 h (TLC: hexane/acetone 1.5:1). The suspension was filtered over
Celite, washed with dichloromethane followed by extraction with aq
Na2CO3. The organic phase was dried (MgSO4), concentrated and the
residue was purified by flash chromatography (hexane/acetone 2:1)
furnishing 12 (23.12 g, 86.1%). [a]2D3 ˆ ‡17.68 (c ˆ 0.5 in dichloromethane);
Rf ˆ 0.36 (hexane/acetone 1.5:1); 1H NMR (500 MHz, [D6]DMSO): d ˆ
7.9 7.7 (m, 8H, Pht), 7.4 7.2 (m, 10H, Ph), 6.92 6.79 (m, 10H, Ph), 5.31
3,6-Di-O-benzyl-4-O-chloroacetyl-2-deoxy-2-phthalimido-b-d-glucopyra-
nosylfluoride (7): Thioglycoside 5 (112.0 g, 183.6 mmol) was dissolved in
dichloromethane (800 mL) and cooled to 08C. N-Bromosuccinimide
(42.5 g, 239.8 mmol) was added to the stirred solution followed by HF/
pyridine complex (20 mL). The reaction was complete after 20 min (TLC:
hexane/acetone 1.5:1). Subsequently, the reaction mixture was transferred
to a Teflon separatory funnel and extracted with a mixture of water and ice
(2 Â ), dilute HCl (3 Â ) and aq Na2CO3. The organic phase was dried
(MgSO4), concentrated and purified by flash chromatography (hexane/
ethyl acetate 2:1) to yield 7 (99.6 g, 95.5%). The product was recrystallized
from acetone/hexane. M.p. 1158C; [a]2D3 ˆ ‡75.48 (c ˆ 1 in CH2Cl2); Rf ˆ
1
(d, J1,2 ˆ 7.9 Hz, 1H, H-1'b), 5.29 (d, J1,2 ˆ 9.2 Hz, 1H, H-1b), 5.12 (dd, J3,4
ˆ
0.33 (hexane/acetone 2:1); H NMR (250 MHz, [D6]DMSO): d ˆ 7.85 (m,
J4,5 ˆ 9.4 Hz, 1H, H-4'), 4.81 (d, Jgem ˆ 12.3 Hz, 1H, CH2O), 4.56 (d, Jgem
ˆ
4H, Pht), 7.35 (m, 5H, Ph), 6.93 (m, 5H, Ph), 5.96 (dd, J1,2 ˆ 7.8, J1,F
ˆ
12.0 Hz, 1H, CH2O), 4.50 (d, Jgem ˆ 12.0 Hz, 1H, CH2O), 4.44 4.38 (m,
7H, CH2O, CH2Cl, H-3'), 4.26 (d, Jgem ˆ 12.1 Hz, 1H, CH2O), 4.16 4.05 (m,
3H, H-2', H-3, H-4), 3.81 (dd, J2,3 ˆ 10.5 Hz, 1H, H-2), 3.63 3.55 (m, 3H,
H-5, H-5', H-6a'), 3.49 (d, Jgem ˆ 10.8 Hz, 1H, H-6a), 3.45 (dd, Jvic ˆ 5.0,
Jgem ˆ 11.1 Hz, 1H, H-6b'), 3.37 (dd, Jvic ˆ 3.7 Hz, 1H, H-6b); 13C NMR
53.4 Hz, 1H, H-1b), 5.22 (dd, J3,4 ˆ J4,5 ˆ 9.4 Hz, 1H, H-4), 4.6 4.4 (m, 6H,
CH2O, CH2Cl, H-3), 4.27 (d, Jgem ˆ 12.0 Hz, 1H, CH2O), 4.2 (m, 1H, H-2),
4.13 (m, 1H, H-5), 3.64 (m, 2H, H-6a,b); 13C NMR (62.5 MHz,
ˆ
[D6]DMSO): d ˆ 167.2, 166.4 (C O, Ac, Pht), 137.9, 137.2 (C-i Ph), 134.9
(C-4/5 Pht), 130.5 (C-1/2 Pht), 128.2, 127.9, 127.6, 127.5 (C-Ar), 123.6 (C-3/6
ˆ
(125 MHz, [D6]DMSO): d ˆ 168.1, 167.1, 166.5 (C O), 138.3, 138.1, 138.0,
Pht), 104.5 (d, JC-1,F ˆ 212.4 Hz, C-1), 75.3 (d, JC-3,F ˆ 9.7 Hz, C-3), 73.6
137.4 (C-i Ph), 134.9, 134.8 (C-4/5 Pht), 130.6 (C-1/2 Pht), 128.3 127.0 (C-
Ar), 123.6, 123.4 (C-3/6 Pht), 96.5 (C-1'), 84.8 (C-1), 76.2 (C-3', C-3'), 75.6
(C-5), 75.2 (C-4), 73.8, 73.7 (CH2O), 73.1 (C-4'), 72.7 (C-5'), 72.6, 71.7
(CH2O), 68.1 (C-6'), 67.6 (C-6), 55.8 (C-2'), 54.6 (C-2), 41.0 (CH2Cl);
elemental analysis calcd (%) for C58H52ClN5O13 (1062.53): C 65.56, H 4.93,
N 6.59; found: C 65.39, H 4.89, N 6.50.
(CH2O), 72.5 (CH2O), 72.3 (C-4), 72.2 (C-5), 68.0 (C-6), 55.1 (d, JC-2,F
ˆ
22.0 Hz, C-2), 40.9 (CH2Cl); elemental analysis calcd (%) for
C30H27ClFNO7 Â H2O (586.01): C 61.49, H 4.99, N 2.39; found: C 61.82,
H 4.94, N 2.66.
3,6-Di-O-benzyl-4-O-chloroacetyl-2-deoxy-2-phthalimido-b-d-glucopyra-
nosylazide (9): TMS-N3 (20 mL, 150.6 mmol) followed by BF3 ¥ OEt2 (2 mL,
16.3 mmol) was added to a stirred suspension of fluoride 7 (40.0 g,
70.4 mmol) and ground molecular sieves 4 ä (20 g) in dry dichloromethane
(400 mL). After complete reaction (TLC: hexane/acetone 2:1) the mixture
was diluted with dichloromethane and filtered. The organic phase was
O-(3,6-Di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosyl)-(1 ! 4)-
3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosylazide
(13):
a) Startingfrom 11: Disaccharide 11 (4.1 g, 3.99 mmol) was dissolved
in dioxane/methanol 1:1 (80 mL) and stirred for 36 h at ‡48C with
Chem. Eur. J. 2003, 9, No. 6
¹ 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
0947-6539/03/0906-1373 $ 20.00+.50/0
1373

C. Unverzagt
FULL PAPER
potassium carbonate (350 mg). The suspension was filtered, neutralized
with acetic acid and concentrated. The residue was purified by flash
chromatography (hexane/acetone 2:1) and gave title compound 13 (3.1 g,
78.7%).
68.4 (C-4''), 67.6 (C-6), 67.5 (C-6'), 61.7 (C-6''), 55.8 (C-2'), 54.7 (C-2), 20.4,
20.3 (OAc); elemental analysis calcd (%) for C75H72N6O21 (1393.42): C
64.65, H 5.21, N 6.03; found: C 64.81, H 5.24, N 5.66.
4,6-O-Benzylidene-3-O-(N-phenylcarbamoyl)-d-glucopyranoside
(19):
b) Startingfrom 12: Disaccharide 12 (14.0 g, 13.2 mmol) was dissolved in
methanol/dichloromethane 1:2 (120 mL) and stirred with potassium
carbonate (30 mg). After complete reaction (TLC: hexane/acetone 1.5:1)
the suspension was filtered, neutralized with acetic acid and concentrated.
The residue was purified by flash chromatography (hexane/acetone 1.5:1)
and yielded 13 (12.2 g, 93.7%). [a]2D3 ˆ À3.38 (c ˆ 0.5 in CH2Cl2); Rf ˆ 0.22
(hexane/acetone 1.5:1); 1H NMR (500 MHz, [D6]DMSO): d ˆ 7.95 7.7 (m,
Compound 18[32] (101 g, 337 mmol) and anhydrous zinc chloride (25 g)
were suspended in benzaldehyde (270 mL) and stirred for 3 d at ambient
temperature (TLC: hexane/acetone 1:1). The reaction was added to ethyl
acetate/water 1:1 (1.6 L). After thorough extraction a first fraction of
product crystallized. The crystals were filtered off and washed with water
and diethyl ether. The organic phase was dried (MgSO4) and diluted with
hexane. A second crop of crystalline product was filtered off and washed
with water and diethyl ether. The fractions of crystalline 19 were combined
(45 g, 34.4%). M.p. 1798C; [a]2D3 ˆ ‡2.28 (1, MeOH/DMF 5:1); Rf ˆ 0.42
(hexane/acetone 1:1); in DMSO compound 19 was present as a mixture of
anomers (a:b ˆ 2:1); a-anomer: 1H NMR (500 MHz, [D6]DMSO): d ˆ 9.61
(s, 1H, NH), 7.5 7.2 (m, 9H, Ph), 6.95 (m, 1H, Ph), 6.88 (d, J1,OH ˆ 4.8 Hz,
8H, Pht), 7.4 7.2 (m, 10H, Ph), 6.95 6.80 (m, 10H, Ph), 5.63 (d, J4,OH
ˆ
7.2 Hz, 1H, OH-4), 5.29 (d, J1,2 ˆ 9.1 Hz, 1H, H-1b), 5.27 (d, J1,2 ˆ 7.1 Hz,
1H, H-1'b), 4.82 (d, Jgem ˆ 12.2 Hz, 1H, CH2O), 4.77 (d, Jgem ˆ 12.2 Hz, 1H,
CH2O), 4.60 (d, Jgem ˆ 12.4 Hz, 1H, CH2O), 4.48 (d, Jgem ˆ 12.4 Hz, 1H,
CH2O), 4.45 4.38 (m, 4H, CH2O), 4.15 (dd, J3,4 ˆ 8.4 Hz, 1H, H-3'), 3.98
(dd, J2,3 ˆ 10.5 Hz, 1H, H-2'), 3.85 3.80 (m, 2H, H-2, H-6a'), 3.60 (m, 1H,
H-5), 3.56 3.48 (m, 3H, H-6b', H-4', H-6a), 3.43 (m, 1H, H-5'), 3.37 (dd,
Jvic ˆ 3.6, Jgem ˆ 11.3 Hz, 1H, H-6b); 13C NMR (125 MHz, [D6]DMSO): d ˆ
ˆ
1H, OH-1), 5.59 (s, 1H, CH-Ph), 5.15 (dd, J2,3 ˆ J3,4 ˆ 9.7 Hz, 1H, H-3),
5.08 (dd, J1,2 ˆ 3.6 Hz, 1H, H-1a), 5.02 (d, J2,OH ˆ 7.7 Hz, 1H, OH-2), 4.16
(dd, Jgem ˆ 10.2, Jvic ˆ 4.5 Hz, 1H, H-6a), 3.93 (m, 1H, H-5), 3.74 (dd, Jvic
ˆ
ˆ
168.2, 167.1 (C O), 138.8, 138.3, 138.1, 138.0 (C-i Ph), 134.8 (C-4/5 Pht),
10.2 Hz, 1H, H-6b), 3.62 (m, 1H, H-4), 3.55 (m, 1H, H-2); b-anomer:
1H NMR (500 MHz, [D6]DMSO): d ˆ 9.63 (s, 1H, NH), 7.5 7.2 (m, 9H,
130.8, 130.6 (C-1/2 Pht), 128.2 127.0 (C-Ar), 123.4 (C-3/6 Pht), 96.5 (C-1'),
84.9 (C-1), 78.4 (C-3'), 76.1 (C-3, C-5'), 75.8 (C-5), 74.8 (C-4), 73.6, 72.4,
71.71 (CH2O), 71.65 (C-4'), 68.9 (C-6'), 67.5 (C-6), 55.9 (C-2'), 54.6 (C-2);
elemental analysis calcd (%) for C56H51N5O12 (986.05): C 68.21, H 5.21, N
7.10; found: C 68.09, H 5.15, N 7.02.
ˆ
Ph), 7.07 (d, J1,OH ˆ 6.5 Hz, 1H, OH-1), 6.95 (m, 1H, Ph), 5.60 (s, 1H, CH-
Ph), 5.48 (d, J2,OH ˆ 5.7 Hz, 1H, OH-2), 4.99 (dd, J2,3 ˆ J3,4 ˆ 9.5 Hz, 1H,
H-3), 4.63 (dd, J1,2 ˆ 7.5 Hz, 1H, H-1b), 4.22 (dd, Jgem ˆ 10.2 Hz, Jvic
ˆ
5.0 Hz, 1H, H-6a), 3.74 (dd, Jvic ˆ 10.2 Hz, 1H, H-6b), 3.64 (m, 1H, H-4),
3.53 (m, 1H, H-5), 3.28 (m, 1H, H-5); 13C NMR (125 MHz, [D6]DMSO):
2,4,6-Tri-O-acetyl-3-O-(N-phenylcarbamoyl)-a-d-glucopyranosylfluoride
(15): Acetate 14[32] (11.0 g, 23.5 mmol) was dissolved in HF/pyridine
(20 mL). After 20 h at ambient temperature the mixture was added to
dichloromethane (200 mL) and ice (200 mL) in a Teflon separatory funnel.
The organic phase was extracted with aq HCl (3 Â ), aq K2CO3 and dried
(MgSO4). After removal of the solvent the residue was purified by flash
chromatography (hexane/acetone 2.5:1) and furnished fluoride 15 (5.6 g,
55.7%). [a]2D3 ˆ ‡57.68 (c ˆ 1 in dichloromethane); Rf ˆ 0.38 (hexane/
acetone 1.5:1); 1H NMR (250 MHz, [D6]DMSO): d ˆ 9.8 (s, 1H, NH), 7.5
ˆ
ˆ
d ˆ 153.1 (C O urethane), 139.2 118.1 (C-Ar), 100.5 ( CH-Pha), 100.3
ˆ
( CH-Phb), 97.7 (C-1b), 93.3 (C-1a), 79.1 (C-4a), 78.6 (C-4b), 74.3 (C-3b),
73.7 (C-2b), 72.1 (C-3a), 70.9 (C-2a), 68.2 (C-6a), 67.9 (C-6b), 65.5 (C-5b),
62.1 (C-5a); FAB-MS (MB): m/z: calcd for C20H21NO7: 387.1; found: 388
‡
[M‡H] .
4,6-O-Benzylidene-2-O-chloroacetyl-3-O-(N-phenylcarbamoyl)-d-gluco-
pyranosyltrichloroacetimidate (20): Benzylidene acetal 19 (11.4 g,
29.4 mmol) and pyridine (20 mL) were dissolved in dichloromethane
(100 mL) and cooled to 08C. Chloroacetic acid anhydride (15 g, 87.7 mmol)
was added and the reaction was allowed to proceed at 08C until the starting
material disappeared (TLC: hexane/acetone 1.5:1). The solution was
diluted with dichloromethane (200 mL) and extracted with aq K2CO3, aq
HCl (3 Â ) and aq K2CO3. After drying (MgSO4) the solvent was
evaporated. The residue (14.6 g) was dissolved in DMF (21 mL) and
hydrazine acetate (2.4 g) was added. After complete reaction (TLC:
hexane/acetone 1.5:1) the mixture was diluted with dichloromethane
(400 mL) and extracted with water (2 Â ) and aq KHCO3. The organic
phase was dried (MgSO4) and concentrated. Subsequently, the hemiacetal
was dissolved in dry dichloromethane (100 mL) and cooled to 08C.
Trichloroacetonitrile (8 mL, 54 mmol) and 1,8-diazabicyclo[5.4.0]undec-7-
ene (DBU) (775 mL, 7.7 mmol) were added. Upon complete consumption
of the starting material (TLC: hexane/acetone 1.5:1) the solution was
evaporated to dryness and the residue was purified by flash chromatog-
raphy (hexane/acetone 2:1) to give the imidate 20 (9.77 g, 54.6%). Rf bis-
chloroacetate ˆ 0.44 (hexane/acetone 1.5:1); Rf hemiacetal ˆ 0.33 (hexane/
acetone 1.5:1); Rf imidate ˆ 0.47 (hexane/acetone 1.5:1); 1H NMR
7.0 (m, 5H, Ph), 5.94 (dd, J1,F ˆ 52.9, J1,2 ˆ 2.5 Hz, 1H, H-1a), 5.32 (dd, J2,3
J3,4 ˆ 9.9 Hz, 1H, H-3), 5.15 (dd, J4,5 ˆ 8.8 Hz, 1H, H-4), 5.06 (ddd, J2,F
24.8 Hz, 1H, H-2), 4.28 4.2 (m, 2H, H-5, H-6a), 4.10 (dd, Jgem ˆ 14.4, Jvic
ˆ
ˆ
ˆ
4.5 Hz, 1H, H-6b), 2.04, 1.98 (2s, 9H, OAc); 13C NMR (125 MHz,
ˆ
ˆ
[D6]DMSO): d ˆ 169.9, 169.4, 169.1 (C O), 152.3 (C O urethane), 138.6
(C-i Ph), 128.7, 122.8, 118.4 (C-Ar), 104.0 (d, JC-1,F ˆ 226.4 Hz, C-1), 69.7 (C-
5), 69.4 (d, JC-2,F ˆ 24.2 Hz, C-2), 69.3 (C-3), 67.2 (C-4), 61.4 (C-6), 20.4, 20.2
(OAc); elemental analysis calcd (%) for C19H22FNO9 (427.38): C 53.4, H
5.19, N 3.28; found: C 53.51, H 5.10, N 3.44.
O-(2,4,6-Tri-O-acetyl-3-O-(N-phenylcarbamoyl)-b-d-glucopyranosyl)-
(1 ! 4)-O-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosyl)-
(1 ! 4)-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosylazide
(16): Disaccharide 13 (2.9 g, 2.9 mmol), fluoride 15 (1.95 g, 4.6 mmol) and
ground molecular sieves 4 ä (3.4 g) in dry dichloromethane (50 mL) were
stirred for 30 min at room temperature. BF3 ¥ OEt2 (500 mL, 4.1 mmol) was
added and the reaction was allowed to proceed for 7 h (TLC: hexane/
acetone 1.5:1). The suspension was filtered over Celite, washed with
dichloromethane followed by extraction with aq Na2CO3. The organic
phase was dried (MgSO4), concentrated and the residue was purified by
flash chromatography (hexane/acetone 2:1) to yield trisaccharide 16 (3.5 g,
85.4%). [a]2D3 ˆ À7.58 (c ˆ 0.5 in dichloromethane); Rf ˆ 0.28 (hexane/
acetone 1.5:1); 1H NMR (500 MHz, [D6]DMSO): d ˆ 9.7 (s, 1H, NH), 7.9
7.65 (m, 8H, Pht), 7.50 7.20 (m, 20H, Ar), 7.02 6.8 (m, 10H, Ar), 5.28 (d,
J1,2 ˆ 9.5 Hz, 1H, H-1b), 5.27 (m, 1H, H-3''), 5.25 (d, J1,2 ˆ 8.5 Hz, 1H,
H-1'b), 4.95 (dd, J3,4 ˆ J4,5 ˆ 9.6 Hz, 1H, H-4''), 4.90 4.86 (m, 2H, H-1'',
H-2''), 4.83 4.78 (m, 2H, CH2O), 4.63 (d, Jgem ˆ 12.3 Hz, 1H, CH2O), 4.57
(d, Jgem ˆ 12.3 Hz, 1H, CH2O), 4.41 (m, 3H, CH2O), 4.31 (d, Jgem ˆ 12.3 Hz,
ˆ
(500 MHz, [D6]DMSO): d ˆ 9.97 (s, 1H, C NH), 9.78 (s, 1H, NH
urethane), 7.49 6.96 (m, 10H, Ph), 6.53 (d, J1,2 ˆ 3.6 Hz, 1H, H-1a), 5.71
ˆ
(s, 1H, CH-Ph), 5.49(dd, J2,3 ˆ J3,4 ˆ 9.8 Hz, 1H, H-3), 5.30 (dd, 1H, H-2),
4.42 4.27 (m, 3H, CH2Cl, H-6a), 4.11 (dd, J4,5 ˆ 9.6 Hz, 1H, H-4), 4.01 (m,
1H, H-5), 3.89 (dd, Jgem ˆ Jvic ˆ 10.2 Hz, H-6b); 13C NMR (125 MHz,
ˆ
ˆ
ˆ
[D6]DMSO): d ˆ 166.3 (C O), 158.0 (C N), 152.3 (C O urethane), 138.7
ˆ
118.4 (C-Ar), 100.6 ( CH-Ph), 92.5 (C-1a), 77.2 (C-4), 71.4 (C-2), 68.8 (C-
3), 67.2 (C-6), 65.3 (C-5), 40.5 (CH2Cl); FAB-MS (MB): m/z: calcd for
C24H22Cl4N2O8: 606.0; found: 607 [M‡H] .
‡
1H, CH2O), 4.20 4.06 (m, 4H, H-3', H-3, H-4, H-6a''), 4.03 (dd, J2,3
10.5 Hz, 1H, H-2'), 3.94 3.89 (m, H-4', H-5'', H-6b''), 3.83 3.78 (m, 2H,
H-2, H-6a'), 3.61 3.58 (m, 2H, H-5, H-6b'), 3.50 (dd, Jvic < 1.0, Jgem
11.6 Hz, 1H, H-6a), 3.4 3.3 (m, 2H, H-5, H-6b); 13C NMR (125 MHz,
ˆ
4,6-O-Benzylidene-2-O-chloroacetyl-3-O-(N-phenylcarbamoyl)-b-d-glu-
copyranosylfluoride (21): Imidate 20 (2 g, 21.9 mmol) was dissolved in dry
dichloromethane (30 mL) in a polyethylene container and cooled to 08C.
To the stirred solution was added HF/pyridine complex (420 mL). The
reaction was complete after 5 min (TLC: hexane/acetone 2:1). Subse-
quently, the biphasic reaction was diluted with dichloromethane (130 mL),
transferred to a Teflon separaratory funnel and neutralized with cold aq
KHCO3. The organic phase was extracted with aq HCl (3 Â ) and aq
KHCO3. After concentration the residue was purified by flash chro-
matography (hexane/acetone 2:1) to yield the fluoride 21 (1.4 g, 91.4%).
ˆ
ˆ
ˆ
[D6]DMSO): d ˆ 169.8, 169.2, 168.9, 168.1, 167.1 (C O), 152.42 (C O
urethane), 138.7, 138.31, 138.25, 138.1, 138.0 (C-i Ar), 134.8 (C-4/5 Pht),
130.6 (C-1/2 Pht), 128.8, 128.3, 128.2, 127.8, 127.7, 127.5, 127.4, 127.2, 127.0
(C-Ar), 123.5 (C-3/6 Pht), 122.8, 118.5 (C-Ar), 99.8 (C-1''), 96.3 (C-1'), 84.9
(C-1), 78.4 (C-4'), 76.8 (C-3'), 76.1 (C-3), 75.6 (C-5), 75.0 (C-4), 74.7 (C-5'),
73.8, 73.7 (CH2O), 72.4 (CH2O, C-3''), 71.7 (CH2O), 71.5 (C-2''), 70.6 (C-5''),
1374
¹ 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
0947-6539/03/0906-1374 $ 20.00+.50/0
Chem. Eur. J. 2003, 9, No. 6

Core Trisaccharide Building Blocks
1369 1376
[a]2D3 ˆ À32.28 (c ˆ 1 in dichloromethane); Rf ˆ 0.44 (hexane/acetone 2:1);
1H NMR (500 MHz, [D6]DMSO): d ˆ 9.80 (s, 1H, NH), 7.45 6.96 (m, 10H,
(1.395 g, 44.1%). [a]2D3 ˆ À8.28 (c ˆ 0.5 in dichloromethane); Rf ˆ 0.28
(hexane/acetone 1.5:1); 1H NMR (500 MHz, [D6]DMSO): d ˆ 9.6 (s, 1H,
NH), 7.95 7.65 (m, 8H, Pht), 7.50 7.20 (m, 20H, Ar), 7.00 6.75 (m, 10H,
ˆ
Ph), 5.79 (dd, J1,2 ˆ 6.5, J1,F ˆ 53.7 Hz, 1H, H-1b), 5.68 (s, 1H, CH-Ph),
ˆ
5.38 (dd, J2,3 ˆ J3,4 ˆ 8.9 Hz, 1H, H-3), 5.09 (m, 1H, H-2), 4.51, 4.41 (2d,
Jgem ˆ 15.3 Hz, 1H, CH2Cl), 4.34 (dd, Jgem ˆ 9.9, Jvic ˆ 4.5 Hz, 1H, H-6a),
Ar), 5.92 (d, J2,OH ˆ 6.0 Hz, 1H, OH-2''), 5.56 (s, 1H, CH-Ph), 5.28 (d,
J1,2 ˆ 9.2 Hz, 1H, H-1b), 5.27 (d, J1,2 ˆ 8.3 Hz, 1H, H-1'b), 5.01 (dd, J2,3
J3,4 ˆ 9.5 Hz, 1H, H-3''), 4.85 (d, Jgem ˆ 12.4 Hz, 1H, CH2O), 4.79 (d, Jgem
11.9 Hz, 1H, CH2O), 4.67 (d, J1,2 ˆ 6.9 Hz, 1H, H-1''b), 4.65 (d, Jgem
ˆ
ˆ
ˆ
4.03 (dd, J4,5 ˆ 9.4 Hz, 1H, H-4), 3.97 (m, 1H, H-5), (dd, Jgem ˆ 9.9, Jvic
ˆ
13
ˆ
10.1 Hz, 1H, H-6b); C NMR (125 MHz, [D6]DMSO): d ˆ 152.3 (C O
urethane), 138.6, 136.9 (C-i Ph), 129.4 118.5 (C-Ar), 106.3 (d, JC-1,F
ˆ
11.6 Hz, 1H, CH2O), 4.55 (d, Jgem ˆ 12.2 Hz, 1H, CH2O), 4.42 4.34 (m,
4H, CH2O), 4.27 (dd, J2,3 ˆ 10.4, J3,4 ˆ 8.9 Hz, 1H, H-3'), 4.14 3.96 (m, 6H,
H-3, H-4, H-6a', H-6a'', H-2', H-4'), 3.89 (dd, Jvic ˆ 4.2, Jgem ˆ 11.0 Hz, 1H,
H-6b'), 3.81 (dd, J2,3 ˆ 10.5 Hz, 1H, H-2), 3.67 3.60 (m, 3H, H-4'', H-6b'',
H-5), 3.54 (m, 1H, H-5'), 3.48 (dd, Jvic < 1.0, Jgem ˆ 10.6 Hz, 1H, H-6a),
3.45 3.32 (m, 3H, H-2'', H-5'', H-6b); 13C NMR (62.5 MHz, [D6]DMSO):
ˆ
214.1 Hz, C-1), 100.5 ( CH-Ph), 79.9 (C-4), 73.4 (d, JC-2,F ˆ 25.4 Hz, C-2),
70.8 (d, JC-3,F ˆ 9.5 Hz, C-3), 67.2 (C-6), 65.1 (C-5), 40.6 (CH2Cl); FAB-MS
‡
(MB): m/z: calcd for C22H21ClFNO7: 665.1; found: 666 [M‡H] .
O-(4,6-O-Benzylidene-2-O-chloroacetyl-3-O-(N-phenylcarbamoyl)-b-d-
glucopyranosyl)-(1 ! 4)-O-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-
glucopyranosyl)-(1 ! 4)-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glu-
copyranosylazide (22): a) Startingfrom 21: Disaccharide 13 (85 mg,
86 mmol), fluoride 21 (120 mg, 258 mmol) and ground molecular sieves
4 ä (320 mg) were suspended in dry dichloromethane (3 mL) and stirred
for 30 min at 08C. BF3 ¥ OEt2 (40 mL, 326 mmol) was added and the reaction
was allowed to proceed at ambient temperature for 24 h (TLC: hexane/
acetone 1.5:1). Subsequently, the suspension was filtered over celite,
washed with dichloromethane and extracted with aq K2CO3. The organic
phase was dried (MgSO4), concentrated and the residue was purified by
flash chromatography (hexane/acetone 2:1) to furnish the title trisacchar-
ide 22 (56 mg, 45.5%).
ˆ
ˆ
d ˆ 168.1, 167.0 (C O), 152.9 (C O urethane), 139.1, 138.6, 138.1, 138.0,
137.9, 137.3 (C-i Ar), 134.8 (C-4/5 Pht), 130.6 (C-1/2 Pht), 128.7 126.1 (C-
ˆ
Ar), 123.4 (C-3/6 Pht), 122.3, 118.2 (C-Ar), 103.2 (C-1''), 100.2 ( CH-Ph),
96.5 (C-1'), 84.8 (C-1), 78.1 (C-4''), 77.9 (C-4'), 76.6 (C-3'), 76.2 (C-3), 75.7
(C-5), 75.0 (C-4), 74.7 (C-5'), 74.0 (C-3''), 73.7 (CH2O), 72.7 (C-2''), 72.1,
71.6 (CH2O), 67.5 (C-6), 67.5 (C-6', C-6''), 65.7 (C-5''), 56.0 (C-2'), 54.6 (C-
2); elemental analysis calcd (%) for C76H70N6O18 (1355.42): C 67.35, H 5.21,
N 6.20; found: C 67.32, H 5.15, N 6.00.
O-(4,6-O-Benzylidene-b-d-mannopyranosyl)-(1 ! 4)-O-(3,6-di-O-benzyl-
2-deoxy-2-phthalimido-b-d-glucopyranosyl)-(1 ! 4)-3,6-di-O-benzyl-2-de-
oxy-2-phthalimido-b-d-glucopyranosylazide (2): Trisaccharide 17 (4.2 g,
3.1 mmol) and pyridine (2 mL) were dissolved in dry dichloromethane
(150 mL). The stirred solution was cooled to À408C and trifluorometha-
nesulfonic acid anhydride (700 mL, 4.16 mmol) was added. Upon warming
to 08C the reaction went to completion (TLC: hexane/acetone 1.5:1). The
mixture was concentrated in a water bath at ambient temperature followed
by evaporation to drynesss in high vacuo. Subsequently, dry DMF (20 mL)
and pyridine (2 mL) were added prior to heating to 658C. After 2 h the
reaction was complete (TLC: dichloromethane/methanol 50:1) and con-
centrated in high vacuo. The residue was dissolved in dichloromethane
(250 mL) and extracted with aq K2CO3. The organic phase was dried
(MgSO4), concentrated to dryness and subsequently dissolved in a mixture
of dioxane (20 mL), acetic acid (6 mL) and water (4 mL). After complete
hydrolysis of the iminocarbonate (TLC: dichloromethane/methanol 50:1)
the solvents were removed at ambient temperature in a rotary evaporator.
The residue was taken up in dichloromethane (250 mL), extracted with aq
HCl and aq K2CO3, dried (MgSO4) and concentrated. The crude product
(carbonate) was dissolved in dry dichloromethane (150 mL) and a freshly
prepared solution (5 mL) of sodium (100 mg) in dry methanol (100 mL)
was added. After complete reaction (TLC: hexane/acetone 1.5:1) the
solution was neutralized with acetic acid, concentrated and purified by flash
chromatography (hexane/acetone 1.5:1) to give trisaccharide 2 (2.69 g,
70.2%). [a]2D3 ˆ À3.38 (c ˆ 0.5 in dichloromethane); Rf phenylurethane
17 ˆ 0.28 (hexane/acetone 1.5:1); Rf triflate ˆ 0.34 (hexane/acetone 1.5:1);
Rf triflate 17a ˆ 0.67 (dichloromethane/methanol 50:1); Rf iminocarbonate
17b ˆ 0.62 (dichloromethane/methanol 50:1); Rf carbonate 17c ˆ 0.64
(dichloromethane/methanol 50:1); Rf carbonate 17c ˆ 0.29 (hexane/ace-
tone 1.5:1); Rf diol 2 ˆ 0.21 (hexane/acetone 1.5:1); 1H NMR (500 MHz,
[D6]DMSO): d ˆ 7.95 7.65 (m, 8H, Pht), 7.41 7.22 (m, 15H, Ar), 6.96
b) Startingfrom 20: Disaccharide 14 (5.8 g, 5.9 mmol), imidate 20 (6.65 g,
10.9 mmol) and ground molecular sieves 4 ä (18 g) were suspended in dry
dichloromethane (50 mL) and stirred for 20 min at 08C. Trifluorometha-
nesulfonic acid trimethylsilylester (600 mL, 3.2 mmol) was added and the
reaction was monitored for 3 h (TLC: hexane/acetone 1.5:1). Subsequently,
the solids were filtered off over Celite, washed with dichloromethane and
the organic phase was extracted with dilute K2CO3. The dried solution
(MgSO4) was concentrated and the residue was purified by flash
chromatography (hexane/acetone 2:1) to give trisaccharide 22 (5.26 g,
62.4%). [a]2D3 ˆ À14.18 (c ˆ 0.5 in dichloromethane); Rf ˆ 0.33 (hexane/
acetone 1.5:1); 1H NMR (500 MHz, [D6]DMSO): d ˆ 9.71 (s, 1H, NH),
7.95 7.65 (m, 8H, Pht), 7.50 7.20 (m, 20H, Ar), 7.00 6.78 (m, 10H, Ar),
ˆ
5.61 (s, 1H, CH-Ph), 5.30 5.24 (m, 3H, H-1, H-1', H-3''), 4.95 (m, 2H,
H-1'', H-2''), 4.83 (d, Jgem ˆ 12.4 Hz, 1H, CH2O), 4.79 (d, Jgem ˆ 11.9 Hz, 1H,
CH2O), 4.63 (d, Jgem ˆ 11.6 Hz, 1H, CH2O), 4.58 (d, Jgem ˆ 12.2 Hz, 1H,
CH2O), 4.43 4.37 (m, 4H, CH2O), 4.32 (m, 2H, CH2Cl), 4.20 (dd, J2,3
ˆ
J3,4 ˆ 9.8 Hz, 1H, H-3'), 4.15 (dd, Jvic ˆ 4.7 Hz, Jgem ˆ 9.8 Hz, 1H, H-6a''),
4.12 4.08 (m, 2H, H-3, H-4), 4.03 (m, 1H, H-2'), 3.98 (dd, J4,5 ˆ 8.4 Hz, 1H,
H-4'), 3.84 3.80 (m, 3H, H-2, H-4'', H-6a'), 3.68 3.60 (m, 3H, H5, H-6b',
H-6b''), 3.54 (m, 1H, H-5''), 3.49 (dd, Jvic < 1.0, Jgem ˆ 10.8 Hz, 1H, H-6a),
3.41 (m, 1H, H-5'), 3.36 (dd, Jvic ˆ 3.1 Hz, 1H, H-6b); 13C NMR (62.5 MHz,
ˆ
ˆ
[D6]DMSO): d ˆ 167.1, 166.1 (C O), 152.5 (C O urethane), 138.7, 138.3,
138.0, 137.0 (C-i Ar), 134.8 (C-4/5 Pht), 130.6 (C-1/2 Pht), 129.0 126.1 (C-
ˆ
Ar), 123.4 (C-3/6 Pht), 122.3, 118.6 (C-Ar), 100.4 ( CH-Ph), 99.6 (C-1''),
96.4 (C-1'), 84.8 (C-1), 77.6 (C-4'', C-4'), 76.5 (C-3'), 76.2 (C-3), 75.7 (C-5),
75.1 (C-4), 74.5 (C-5'), 74.2 (C-2''), 73.9 (CH2O), 73.7 (CH2O), 72.3
(CH2O), 71.7 (C-2'', CH2O), 67.6 (C-6, C-6'), 67.4 (C-6''), 65.5 (C-5''), 55.8
(C-2'), 54.6 (C-2), 40.6 (CH2Cl); ESI-MS: m/z: calcd for C78H71ClN6O19
1430.45; found: 1453.50 [M‡Na] .
:
‡
ˆ
6.75 (m, 10H, Ar), 5.51 (s, 1H, CH-Ph), 5.28 (d, J1,2 ˆ 9.5 Hz, 1H, H-1b),
O-(4,6-O-Benzylidene-3-O-(N-phenylcarbamoyl)-b-d-glucopyranosyl)-
(1 ! 4)-O-(3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosyl)-
(1 ! 4)-3,6-di-O-benzyl-2-deoxy-2-phthalimido-b-d-glucopyranosylazide
5.25 (d, J1,2 ˆ 8.5 Hz, 1H, H-1'b), 4.96 (d, J2,OH ˆ 4.3 Hz, 1H, OH-2''), 4.92
(d, J3,OH ˆ 6.9 Hz, 1H, OH-3''), 4.82 (d, Jgem ˆ 12.1 Hz, 2H, CH2O), 4.62 (d,
J1,2 < 1.0 Hz, 1H, H-1''b), 4.60 (d, Jgem ˆ 12.2 Hz, 1H, CH2O), 4.55 (d,
Jgem ˆ 12.2 Hz, 1H, CH2O), 4.43 4.37 (m, 4H, CH2O), 4.21 (dd, J2,3 ˆ 10.4,
J3,4 ˆ 8.8 Hz, 1H, H-3'), 4.14 3.96 (m, 5H, H-3, H-4, H-6a'', H-2', H-4'),
3.80 3.76 (m, 3H, H-2, H-2'', H-6a'), 3.71 (dd, J3,4 ˆ J4,5 ˆ 9.5 Hz, 1H,
H-4''), 3.66 3.47 (m, 5H, H-5, H-6a, H-6b', H-3'', H-6b''), 3.41 (m, 1H,
H-5'), 3.36 (m,1H, H-6b), 3.10 (m, 1H, H-5''); 13C NMR (62.5 MHz,
(17): a) Startingfrom 22: Trisaccharide 22 (14 g, 9.78 mmol) was dissolved
in methanol/dichloromethane 1:2 (120 mL). To the stirred solution was
added K2CO3 (300 mg). After complete deacylation (TLC: hexane/acetone
1.5:1) the solids were filtered off, followed by neutralization with acetic
acid. The solution was concentrated and purified by flash chromatography
(hexane/acetone 1.5:1) furnishing the trisaccharide 17 (11.82 g, 89.2%).
ˆ
[D6]DMSO): d ˆ 168.0, 167.1, 167.0 (C O), 138.4, 138.3, 138.1, 137.9 (C-i
b) Startingfrom 16: Trisaccharide 16 (3.25 g, 2.3 mmol) was dissolved in
dioxane/methanol 1:1 (80 mL) and cooled to 48C. K2CO3 (400 mg) was
added and the suspension was stirred for 36 h. Subsequently, the solids were
removed by filtration. The solution was neutralized with Amberlyst 15 ion-
Ar), 134.8 (C-4/5 Pht), 130.7, 130.5 (C-1/2 Pht), 128.7 126.3 (C-Ar), 123.4
ˆ
(C-3/6 Pht), 100.9 ( CH-Ph), 100.4 (C-1''b, JC-1,H-1 ˆ 158.6 Hz from a
coupled HMQC spectrum), 96.5 (C-1'b, JC-1,H-1 ˆ 169.0 Hz see above), 84.8
(C-1b, JC-1,H-1 ˆ 166.9 Hz see above), 78.3 (C-4''), 77.3 (C-4'), 76.2 (C-3), 76.0
(C-3'), 75.6 (C-5), 74.9 (C-4), 74.5 (C-5'), 73.7, 73.6, 72.1, 71.5 (CH2O), 70.9
(C-2''),69.9 (C-3''), 67.8 (C-6', C-6''), 67.5 (C-6), 66.8 (C-5''), 56.0 (C-2'), 54.7
(C-2); elemental analysis calcd (%) for C69H65N5O17 (1236.30): C 67.04, H
5.30, N 5.66; found: C 66.70, H 5.18, N 5.52. ESI-MS: m/z: calcd for
‡
exchange resin (H ) and concentrated. The residue was dissolved in
acetonitrile (50 mL) and benzaldehyde dimethylacetal (10 mL) and p-
toluenesulfonic acid (100 mg) were added. After 1 h the reaction was
stopped with triethylamine (150 mL), concentrated and purified by flash
chromatography (hexane/acetone 2.5:1) to give the trisaccharide 17
‡
C69H65N5O17: 1235.44); found: 1258.45 [M‡Na] .
Chem. Eur. J. 2003, 9, No. 6
¹ 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
0947-6539/03/0906-1375 $ 20.00+.50/0
1375

C. Unverzagt
FULL PAPER
[22] C. Unverzagt, Angew. Chem. 1994, 106, 1170 1173; Angew. Chem.
Int. Ed. Engl. 1994, 33, 1102 1104.
Acknowledgements
[23] H. Kunz, C. Unverzagt, Angew. Chem. 1988, 100, 1763 1765; Angew.
Chem. Int. Ed. Engl. 1988, 27, 1697 1699.
This work was supported by the Deutsche Forschungsgemeinschaft and the
Leonhard-Lorenz-Stiftung. Special gratitude is expressed to Prof. Dr. Horst
Kessler for his generous support. The chemicals supplied by Hoechst AG
(Aventis) are highly appreciated.
[24] H. Kunz, C. Unverzagt, J. Prakt. Chem. 1992, 334, 579 583.
[25] W. Brˆder, H. Kunz, Carbohydr. Res. 1993, 249, 221 241.
[26] W. Brˆder, H. Kunz, Bioorg. Med. Chem. 1997, 5, 1 19.
[27] H. Herzner, T. Reipen, M. Schultz, H. Kunz, Chem. Rev. 2000, 100,
4495 4538.
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Received: September 06, 2002 [F4398]
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