Facile synthesis of a pentasaccharide mimic of a fragment of the capsular polysaccharide of Streptococcus pneumoniae type 15C

Article abstract of DOI:10.1016/j.carres.2007.12.002

A pentasaccharide mimic of a fragment of the capsular polysaccharide of Streptococcus pneumoniae type 15C β-d-Galp-(1→4)-β-d-Glcp-(1→6)-[α-d-Galp-(1→2)-β-d-Galp-(1→4)]-β-d-GlcpNAc-(1→OCH₂CH₂N₃) (1) was synthesized in a regio- and stereoselective manner. The 2-azidoethyl-spacered pentasaccharide mimic 1 can be used to construct a neoglycoconjugate antigen.{A figure is presented}.

Full text of DOI:10.1016/j.carres.2007.12.002

Available online at www.sciencedirect.com
Carbohydrate Research 343 (2008) 607–614
Facile synthesis of a pentasaccharide mimic of a fragment of
the capsular polysaccharide of Streptococcus pneumoniae type 15C
Chen Yong-Xiang, Zhao Wei, Zhao Lei, Zhao Yu-Fen and Li Yan-Mei,*
Key Laboratory of Bioorganic Phosphorous Chemistry and Chemical Biology (Ministry of Education),
Department of Chemistry, Tsinghua University, Beijing 100084, PR China
Received 12 October 2007; received in revised form 5 December 2007; accepted 6 December 2007
Available online 14 January 2008
Abstract—A pentasaccharide mimic of a fragment of the capsular polysaccharide of Streptococcus pneumoniae type 15C b-D-Galp-
(1?4)-b-D-Glcp-(1?6)-[a-D-Galp-(1?2)-b-D-Galp-(1?4)]-b-D-GlcpNAc-(1?OCH2CH2N3) (1) was synthesized in a regio- and
stereoselective manner. The 2-azidoethyl-spacered pentasaccharide mimic 1 can be used to construct a neoglycoconjugate antigen.
OH
O
OH
O
OH
HO
O
O
HO
OH
OH
OH
OH
O
O
O
O
HO
HO
1
N3
NHAc
O
O
HO
OH
HO
HO
Ó 2007 Elsevier Ltd. All rights reserved.
Keywords: Streptococcus pneumoniae; Oligosaccharide synthesis
1. Introduction
groups, such as infants, small children, immuno-com-
promised patients, and the elderly.3 Conjugation of
S. pneumoniae carbohydrate antigens to a protein carrier
results in a T-cell dependent neoglycoconjugate antigen
that gives an efficient immune response in the high-risk
groups.4
These neoglycoproteins were prepared by conjugation
of isolated capsular polysaccharides or a mixture of
polysaccharide-derived oligosaccharides to a protein
carrier.5,6 The studies with S. pneumoniae type 6B neo-
glycoproteins showed that a synthetic tetrasaccharide
fragment, that is, one repeating unit of the 6B CPS, cou-
pled to keyhole limpet hemocyanin (KLH) was sufficient
to generate a protective antibody response in mice.7
Similar results were shown for S. pneumoniae type 3 neo-
glycoproteins, consisting of di-, tri- and tetrasaccharides
The pathogenic bacterium Streptococcus pneumoniae,
which causes infections of the lung (pneumonia), middle
ear (otitis media) and meninges (meningitis) remains a
major cause of death all over the world.1 With the
increasing incidence of antibiotic resistance in this
organism, vaccination has become an effective means
of protection. A vaccine that consists of a mixture of
the capsular polysaccharides (CPSs) from 23 of the 90
serotypes of S. pneumoniae has been available for more
than 20 years.2 However, the T-cell-independent CPSs
vaccines are ineffective in the most important high-risk
*
Corresponding author. Fax: +86 10 62781695; e-mail: liym@
mail.tsinghua.edu.cn
0008-6215/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2007.12.002

608
Y.-X. Chen et al. / Carbohydrate Research 343 (2008) 607–614
tuted by an azido group from sodium azide in N,N-
OH
O
OH
O
OH
HO
dimethylformamide (DMF) (?4, 94%). After deacetyla-
tion (?5, 85%), product 5 was regioselectively tert-
butyldimethylsilylated at O-6 with tert-butyldimethyl-
silyl chloride in the presence of a catalytic amount of
4-dimethylaminopyridine (DMAP), pyridine and triethyl-
amine to give 6 in 78% yield. Subsequently, product 6
was regioselectively benzoylated at O-3 with a stoichio-
metric amount of benzoyl chloride in the presence of sil-
ver(I) oxide and a catalytic amount of potassium iodide
to give the acceptor 7 with a free hydroxyl group at O-4
in high regioselectivity and in good yield (81%). Ye’s
group18 has also reported the silver(I) oxide-mediated
selective monoprotection of 2,3-diols in pyranosides.
Here, silver(I) oxide was used to mediate selective mono-
protection of 3,4-diols in the pyranoside. The acceptor 7
O
O
HO
OH
OH
OH
OH
O
O
O
O
HO
HO
N3
NHAc
O
O
HO
OH
HO
HO
Figure 1. Overview of pentasaccharide mimic 1 with a 2-azidoethyl
spacer group, representing fragments of the repeating unit of the S.
pneumoniae type 15C capsular polysaccharide.
coupled to the cross-reacting material (CRM197) of
modified diphtheria toxin in different molar carbo-
hydrate–protein ratios.8 Kamerling’s group9–13 has
reported on the chemoenzymatic synthesis of spacered
oligosaccharide fragments of the CPS of S. pneumoniae
type 14. And the tetrasaccharide containing neoglyco-
conjugate (CRM197 as carrier protein) showed, particu-
larly, promising immunological data when tested in mice
models.14
In this paper, we report an original and efficient
approach proposed for the chemical synthesis of
2-azidoethyl-spacered pentasaccharide mimic 1 (Fig. 1)
(without a glycerol-2-phosphate substituent) of a frag-
ment of the CPS of S. pneumoniae type 15C,15 which
is suitable for the construction of a novel neoglycocon-
jugate antigen. The introduction of an azide group can
be useful to construct neoglycoconjugates through a
highly efficient coupling method: the copper (I)-cata-
lyzed 1,2,3-triazole formation from azides and terminal
acetylenes (click chemistry), developed by Sharpless
and co-workers.16
1
was verified by ESIMS and NMR spectroscopy (1D H
1
and 2D H COSY). The galactose trichloroacetimidate
donor 9 with regioselective protective groups was pre-
pared in the following way: Benzoylation of 1,2-O-ethyl-
idene-a-D-galactopyranoside (8),19 followed by sequent
de-ethylidenation, acetylation, 1-O-selective deacetyla-
tion and trichloroacetimidation afforded monosaccha-
ride donor 9 (35% over five steps). The lactose donor
1120 was easily prepared from fully protected lactose
10, in which the acetyl group at O-1 was selectively re-
moved by benzylamine in tetrahydrofuran (THF), fol-
lowed by activation to give the a-trichloroacetimidate
11 (50% over two steps). Allylation of isopropyl 4,6-O-
benzylidene-1-thio-b-D-galactopyranoside (12)21 with al-
lyl bromide in the presence of sodium hydride afforded
the thiogalactoside donor 13 in 80% yield.
Based on these building blocks, the target pentasac-
charide 1 was synthesized. Kamerling’s group13,11,12
has developed enzymic methods to connect galactose
residues to the internal N-acetyl-b-D-glucosamine resi-
dues with a b-(1?4) glycosidic linkage by using bovine
milk b-1,4-galactosyltransferase. Here, we took the
chemical coupling method. Condensation of the galact-
ose trichloroacetimidate donor 9 with the monoglucos-
amine acceptor 7 in DCM using TMSOTf as a catalyst
afforded the b-(1?4) linked disaccharide 14 in accept-
able yield (67%). Subsequent selective removal of 2-O-
acetyl and 6-O-tert-butyldimethylsilyl groups by meth-
anolysis with MeCOCl–MeOH–CH2Cl2 afforded the
disaccharide acceptor 15 (79%) with two free hydroxyl
groups at O-2 of the galactosyl residue and O-6 of Glc-
NAc. Then, a stoichiometric amount of lactose trichloro-
acetimidate donor 11 was regioselectively coupled
with the acceptor 15 at the primary hydroxyl group in
the presence of TMSOTf to give the b-(1?6) linked
tetrasaccharide 16 (50%) with a free hydroxyl group at
O-2, which was characterized by ESIMS and NMR
spectroscopy (1D 1H and 2D 1H COSY). Finally, conden-
sation of the tetrasaccharide 16 with isopropyl 2,3-di-
O-allyl-4,6-O-benzylidene-1-thio-b-D-galactopyranoside
2. Results and discussion
The CPS of S. pneumoniae type 15C is built up from the
pentasaccharide repeating unit?3)-b-D-Galp-(1?4)-b-
D-Glcp-(1?6)-[a-D-Galp-(1?2)-b-D-Galp(3-OR)-(1?4)]-
b-D-GlcpNAc-(1?(R is glycerol-2-phosphate substitu-
ent). Pentasaccharide 1 (Fig. 1) represents one repeating
unit. For the synthesis of 1, four building blocks were
designed: the glucosamine acceptor 7, the galactose tri-
chloroacetimidate donor 9, the lactose trichloroacetimi-
date donor 11 and the thiogalactoside donor 13 (Scheme
1).
The synthesis of monoglucosamine acceptor 7 in-
volved, as the first step, the coupling of 3,4,6-tri-O-acet-
yl-2-deoxy-2-phthalimido-D-glucopyranosyl trichloro-
acetimidate 217 to the spacer 2-bromoethan-1-ol in
dichloromethane (DCM) using trimethylsilyl trifluoro-
methanesulfonate (TMSOTf) as a catalyst (?3, 73%).
Then, the bromo group in the spacer was further substi-

Y.-X. Chen et al. / Carbohydrate Research 343 (2008) 607–614
609
OAc
O
OAc
O
OAc
O
c
b
a
AcO
AcO
AcO
AcO
AcO
AcO
O
O
NH
NPhth
Br
N3
NPhth
NPhth
O
CCl3
4
3
2
R1 R2 R3
OR3
O
H
H
H
5
6
7
R2O
R1O
O
d
N3
H
H
H
TBDMS
TBDMS
NPhth
BzO
BzO
OR1
O
i
OBz
O
e
Bz
O
O
Bz
N3
OR2
NPhth
OBz
OH
HO
OH
O
R1 R2
BzO
BzO
f
O
Ac
H
TBDMS
14
15
NH
CCl3
j
O
OAc
H
O
O
9
8
OAc
OAc
AcO
AcO
OAc
OAc
AcO
AcO
g
O
O
O
O
k
OAc
O
O
AcO
AcO
OAc
OAc
OAc
OAc
OC(NH)CCl3
11
10
Ph
OAc
Ph
O
OAc
O
AcO
AcO
O
O
h
O
O
O
O
AcO
O
OAc
OAc
S
O
OBz
O
BzO
BzO
S
HO
O
O
AllO
OH
O
OAll
N3
BzO
NPhth
OH
12
13
16
l
OR4
OR4
O
OR4
O
OAc
O
OAc
O
OAc
R4O
O
4
O
O
OR4
AcO
AcO
R O
O
OR4
OAc
OAc
OR3
OR3
OBz
m
OBz
O
O
O
O
O
O
O
NPhth
O
N3
R3O
R3O
N3
BzO
BzO
NR1R2
O
O
OR4
OAll
O
O
R4O
R4O
O
O
R1 R2 R3
R4
R4O
AllO
Ph
Phth
Bz
Ac
18
19
1
n
o
17
Ac
Ac
H
H
Ac
H
Ac
H
Scheme 1. Synthesis of pentasaccharide 1. Reagents and conditions: (a) CH2BrCH2OH, TMSOTf, CH2Cl2, À10 °C, 73%; (b) NaN3, DMF, 55 °C,
94%; (c) MeONa, MeOH, 85%; (d) TBDMSCl, DMAP, Pyr, Et3N, CH2Cl2, 78%; (e) BzCl, Ag2O, KI, CH2Cl2, 81%; (f) (1) BzCl/Pyr, (2) 80% HOAc,
(3) Ac2O/Pyr, (4) benzyl amine, THF, (5) CCl3CN, K2CO3, CH2Cl2; 35% over five steps; (g) (1) PhCH2NH2, THF, (2) CCl3CN, K2CO3, CH2Cl2;
50% over two steps; (h) AllBr/NaH, DMF, 80%; (i) TMSOTf, CH2Cl2, À10 °C, 67%; (j) AcCl, MeOH/CH2Cl2, 79%; (k) TMSOTf, CH2Cl2, À10 °C,
50%; (l) TMSOTf, NIS, CH2Cl2, À10 °C, 68%; (m) (1) F3CCO2H/H2O, CH2Cl2, (2) Ac2O/Pyr, (3) PdCl2, CH2Cl2/MeOH, (4) Ac2O/Pyr; 86% over
four steps; (n) (1) MeONa, MeOH, (2) NH2CH2CH2NH2, butanol, 80 °C, (3) Ac2O/Pyr; 77% over three steps; (o) MeONa, MeOH, 84%.
donor 13 using TMSOTf and N-iodosuccinimide (NIS)
as catalysts afforded the a-(1?2)-linked pentasaccharide
17 (68%).
In the deprotection sequence, the fully protected
pentasaccharide 17 was firstly O-debenzylidenated
and O-reacetylated, followed by O-deallylation and

610
Y.-X. Chen et al. / Carbohydrate Research 343 (2008) 607–614
O-reacetylation, to give the fully acylated pentasacha-
ride 18 (86% over four steps). Subsequently, 18 was O-
deacylated and N-dephthaloylated, followed by N,O-
reacetylation to yield the N-acetyl protected derivative
19 (77% over three steps). Complete conventional O-
deacetylation of 19 to give the final product 1 in 84%
yield required long reaction times (48 h), as indicated
by NMR spectroscopy.
4.3. 2-Bromoethyl 3,4,6-tri-O-acetyl-2-deoxy-2-phthal-
imido-b-D-glucopyranoside (3)
As described in the general procedure, 2 (1.31 g,
2.26 mmol) and 2-bromoethan-1-ol (185 lL, 2.60 mmol)
were coupled, and the product was purified by column
chromatography with 1:1 petroleum ether–EtOAc as
the eluent to give 3 (890 mg, 1.64 mmol, 73%) as a
foamy solid.
1H NMR (300 MHz, CDCl3): d 1.88 (s, 3H, OAc);
2.04 (s, 3H, OAc); 2.12 (s, 3H, OAc); 3.30–3.38 (m,
2H, BrCH2); 3.76 (m, 1H, OCH2); 3.88 (m, 1H, H-5);
4.12 (m, 1H, OCH2); 4.18 (dd, 1H, J6b,5 = 2.4 Hz,
J6b,6a = 12.4 Hz, H-6b); 4.28–4.38 (m, 2H, H-2, H-6a);
5.18 (dd, H-1, J4,3 = 9.0 Hz, J4,5 = 10.0 Hz, H-4); 5.42
3. Conclusion
In summary, based on readily accessible building blocks
7, 9, 11 and 13, a convergent and very efficient synthesis
of 2-azidoethyl-spacered pentasaccharide b-D-Galp-
(1?4)-b-D-Glcp-(1?6)-[a-D-Galp-(1?2)-b-D-Galp-(1?
4)]-b-D-GlcpNAc-(1?OCH2CH2N3) (1) was achieved in
a regio- and stereoselective manner. Conjugation of the
oligosaccharide 1 to peptides or proteins to form neoglyco-
conjugates and immunological studies are in progress.
(d, 1H, J1,2 = 8.6 Hz, H-1); 5.81 (dd, 1H, J3,
10.6 Hz, J3,4 = 9.0 Hz, H-3); 7.7–7.9 (m, 4H, Ph).
=
2
ESIMS: calcd for C22H24BrNNaO10 [M+Na]+
564.05, found 563.9.
4.4. 2-Azidoethyl 3,4,6-tri-O-acetyl-2-deoxy-2-phthal-
imido-b-D-glucopyranoside (4)
4. Experimental
4.1. General methods
To a solution of 3 (890 mg, 1.64 mmol) in 20 mL DMF
was added NaN3 (1.07 g, 16.4 mmol), and the mixture
was stirred at 55 °C overnight. After the solvent was
co-evaporated with toluene under reduced pressure,
the residue was dissolved in a mixture of water and
EtOAc. The organic phase was separated from the water
phase. The water phase was extracted two times with
EtOAc. The combined organic layers were washed with
water two times and then dried with Na2SO4. The sol-
vent was evaporated, and the crude product was purified
by silica gel flash chromatography (1:1 petroleum ether–
EtOAc) to give 4 (780 mg, 1.55 mmol, 94%).
1
1
1D H NMR and 2D H COSY NMR spectra were
recorded with Jeol ECA-600 and Jeol ECP-300 spectro-
meters for solutions in CDCl3 or D2O as indicated.
Chemical shifts are given in ppm downfield from internal
1
Me4Si. The H NMR resonance assignments have been
performed mainly according to the peak shapes, the cou-
pling constants and chemical shifts in combination of 2D
1H COSY. Mass spectra were recorded with Bruker Es-
quire-LC mass spectrometer in the ESI mode. High-reso-
lution mass spectra were recorded on a Thermo Electron
LTQ Orbitrap (ESI; source voltage 3.8 kV) spectrometer.
Thin-layer chromatography (TLC) was performed on sil-
ica gel HF254 with detection by charring with 30% (v/v)
H2SO4 in MeOH or in some cases by a UV detector. Col-
umn chromatography was conducted by elution of a col-
umn (16 Â 240 mm, 18 Â 300 mm, 35 Â 400 mm) of
silica gel (200–300 mesh) with EtOAc–petroleum ether
(bp 60–90 °C) as the eluent. Solutions were concentrated
at <50 °C under reduced pressure.
1H NMR (300 MHz, CDCl3): d 1.87 (s, 3H, OAc);
2.04 (s, 3H, OAc); 2.12 (s, 3H, OAc); 3.18 (m, 1H,
N3CH2); 3.40 (m, 1H, N3CH2); 3.66 (m, 1H, OCH2);
3.90 (m, 1H, H-5); 4.02 (m, 1H, OCH2); 4.21 (dd, 1H,
J6b,5 = 2.1 Hz, J6b,6a = 12.3 Hz, H-6b); 4.28–4.38 (m,
2H, H-2, H-6a); 5.19 (dd, H-1, J4,3 = 9.6 Hz, J4,5
=
9.6 Hz, H-4); 5.47 (d, 1H, J1,2 = 8.6 Hz, H-1); 5.79
(dd, 1H, J3,2 = 10.6 Hz, J3,4 = 9.6 Hz, H-3); 7.70–7.80
(m, 4H, Ph).
ESIMS: calcd for C22H24N4NaO10 [M+Na]+ 527.14,
found 527.2.
4.2. General procedure for the glycosylations
The mixture of donor and acceptor was dried under high
vacuum for 2 h, then dissolved in dry CH2Cl2. TMSOTf
(0.05 equiv) was added dropwise at À10 °C under an
argon atmosphere. The reaction mixture was stirred for
3 h, during which time the temperature was gradually
raised to ambient temperature. Then the mixture was
neutralized with Et3N. Concentration of the reaction
mixture, followed by purification on a silica gel column,
gave the desired product.
4.5. 2-Azidoethyl 2-deoxy-2-phthalimido-6-O-tert-butyl-
dimethylsilyl-b-D-glucopyranoside (6)
To a solution of 4 (780 mg, 1.55 mmol) in 15 mL of
methanol was added MeONa (pH 10). The mixture
was stirred for 0.5 h at room temperature, then neutral-
ized with Dowex 50WX8 (H+), filtered and concen-
trated. The residue was eluted on a Sephadex LH-20
column with MeOH to give 5 (500 mg, 1.32 mmol,

Y.-X. Chen et al. / Carbohydrate Research 343 (2008) 607–614
611
85%) as a white solid. To a solution of 5 (385 mg,
1.02 mmol) in 15 mL of dry CH2Cl2 was added tert-
butyldimethylsilyl chloride (TBDMSCl) (216 mg,
1.43 mmol), 658 lL pyridine, a catalytic amount of
DMAP and 187 lL Et3N. The mixture was stirred over-
night, and 15 mL of cold water was then added. The
organic phase was separated from the water phase.
The water phase was extracted two times with CH2Cl2.
The combined organic layers were dried with Na2SO4.
The solvent was evaporated, and the residue was puri-
fied by silica gel flash chromatography (1:1 petroleum
ether–EtOAc) to give a product 6 (390 mg, 0.792 mmol,
78%).
separated from the water phase. The water phase was
extracted two times with EtOAc. The combined organic
layers were dried with Na2SO4. The solvent was evapo-
rated, and the residue was purified by silica gel flash
chromatography (1:2 petroleum ether–EtOAc) to give
a pure product, which was dissolved in 15 mL of pyri-
dine. After adding 10 mL of Ac2O at 0 °C, the mixture
was stirred at room temperature overnight. The solvent
was evaporated, and the residue was purified by silica
gel flash chromatography (3:2–1:1 petroleum ether–
EtOAc) to give 1,2-di-O-acetyl-3,4,6-tri-O-benzoyl-b-D-
galactopyranoside, which was dissolved in 20 mL dry
THF, and benzylamine (3 mL, 27.4 mmol) was added.
The mixture was stirred at rt for 8 h and then neutral-
ized with 2 N HCl. After the solvent was removed, the
residue was dissolved in a mixture of EtOAc and water.
The organic phase was separated from the water
phase. The water phase was extracted two times with
EtOAc. The combined organic layers were dried with
Na2SO4. The solvent was evaporated, and the residue
was purified by silica gel flash chromatography (1:1–
1:2 petroleum ether–EtOAc) to give 2-O-acetyl-3,4,6-
tri-O-benzoyl-b-D-galactopyranoside, which was dis-
solved in 15 mL of CH2Cl2. Then, 3 mL of trichloroace-
tonitrile and 3 g K2CO3 were added to the solvent, and
the mixture was stirred overnight. After filtration, the
mixture was concentrated. The residue was purified by
silica gel flash chromatography (1:1–2:3 petroleum
ether–EtOAc) to give product 9 (296 mg, 0.42 mmol).
The yield over five steps was 35%.
1H NMR (300 MHz, CDCl3): d 0.12 (s, 3H, SiCH3);
0.13 (s, 3H, SiCH3); 0.92 (s, 3 Â 3H, SiC(CH3)3); 3.16
(m, 1H, N3CH2); 3.36 (m, 1H, N3CH2); 3.51–3.68 (m,
3H, H-5, OCH2, H-4); 3.85–4.03 (m, 3H, OCH2, H-6b,
H-6a); 4.15 (dd, 1H, J2,1 = 8.5 Hz, J2,3 = 11.0 Hz, H-
2); 4.35 (dd, 1H, J3,2 = 11.0 Hz, J3,4 = 8.2 Hz, H-3);
5.31 (d, 1H, J1,2 = 8.5 Hz, H-1); 7.70–7.85 (m, 4H,
Ph). ESIMS: calcd for C22H32N4NaO7Si [M+Na]+
515.19, found 515.1.
4.6. 2-Azidoethyl 3-O-benzoyl-2-deoxy-2-phthalimido-6-
O-tert-butyldimethylsilyl-b-D-glucopyranoside (7)
To a solution of 6 (354 mg, 0.719 mmol) in 15 mL of dry
CH2Cl2 was added benzoyl chloride (92 lL,
0.791 mmol) in 5 mL of dry CH2Cl2, Ag2O (250 mg,
1.08 mmol), KI (25.8 mg, 0.155 mmol) and 390 lL of
pyridine. The mixture was stirred overnight, and then
the solvent was removed. The residue was purified by sil-
ica gel flash chromatography (1:1 petroleum ether–
EtOAc) to give product 7 (350 mg, 0.587 mmol, 81%).
1H NMR (300 MHz, CDCl3): d 0.12 (s, 3H, SiCH3);
0.13 (s, 3H, SiCH3); 0.92 (s, 3 Â 3H, SiC(CH3)3); 3.19
(m, 1H, N3CH2); 3.37 (m, 1H, N3CH2); 3.63–3.75 (m,
2H, H-5, OCH2); 3.91–4.05 (m, 4H, H-4, OCH2, H-6b,
H-6a); 4.43 (dd, 1H, J2,1 = 8.2 Hz, J2,3 = 10.6 Hz,
H-2); 5.51 (d, 1H, J1,2 = 8.2 Hz, H-1); 5.90 (dd, 1H,
J3,2 = 10.6 Hz, J3,4 = 8.6 Hz, H-3); 7.30–7.90 (m, 4H,
Ph). ESIMS: calcd for C29H36N4NaO8Si [M+Na+]
619.22, found 619.2.
1H NMR (300 MHz, CDCl3): d 1.98 (s, 1H, OAc);
4.42 (dd, 1H, J6b,6a = 11.3 Hz, J6b,5 = 6.9 Hz, H-6b);
4.57 (dd, 1H, J6a,6b = 11.3 Hz, J6a,5 = 6.9 Hz, H-6a);
4.80 (t, J5,6a = J5,6b = 6.9 Hz, H-5); 5.72 (dd, 1H,
J2,3 = 10.6 Hz, J2,1 = 3.4 Hz, H-2); 5.85 (dd, 1H,
J3,2 = 10.6 Hz, J3,4 = 3.4 Hz, H-3); 6.10 (d, 1H, J4,3
=
2.7 Hz, H-4); 6.78 (d, 1H, J1,2 = 3.4 Hz, H-1); 7.28–
8.09 (m, 15H, PhCO); 8.71 (s, 1H, C@NH).
ESIMS: calcd for C31H26Cl3NNaO10 [M+Na]+
700.05, found 700.2.
4.8. Isopropyl 2,3-di-O-allyl-4,6-O-benzylidene-1-thio-b-
D-galactopyranoside (13)
4.7. 2-O-Acetyl-3,4,6-tri-O-benzoyl-a-D-galactopyranosyl
trichloroacetimidate (9)
To a solution of isopropyl 4,6-O-benzylidene-1-thio-b-
D-galactopyranoside (12, 500 mg, 1.53 mmol) in 10 mL
DMF was added allyl bromide (135 lL, 1.56 mmol)
twice and NaH (74 mg, 3.07 mmol) at 0 °C. The mixture
was stirred at 0 °C for 0.5 h, and then at rt for 4 h. The
solvent was evaporated under reduced pressure, and the
residue was dissolved in a mixture of EtOAc and water.
The organic phase was separated from the water phase.
The water phase was extracted two times with EtOAc.
The combined organic layers were dried with Na2SO4.
The solvent was evaporated and the residue was purified
To a solution of 1,2-O-ethylidene-a-D-galactopyrano-
side (8, 250 mg, 1.21 mmol) in 15 mL of pyridine was
added 3 mL of benzoyl chloride at 0 °C, and then the
mixture was stirred at room temperature overnight.
After the solvent was evaporated, the residue was
dissolved in 80% CH3CO2H, and stirred for one day,
followed by neutralization with Et3N. After the solvent
was removed, the residue was redissolved in a certain
amount of EtOAc and water. The organic phase was

612
Y.-X. Chen et al. / Carbohydrate Research 343 (2008) 607–614
by silica gel flash chromatography (2:1–3:2 petroleum
ether–EtOAc) to give 13 (527 mg, 1.23 mmol, 80%).
1H NMR (300 MHz, CDCl3): d 1.32 (d, 3H,
J = 6.9 Hz, CH3); 1.37 (d, 3H, J = 6.9 Hz, CH3); 3.28
(m, 1H, SCH); 3.38 (d, 1H, J4,3 = 3.5 Hz, H-4); 3.47
(dd, 1H, J3,2 = 9.3 Hz, J3,4 = 3.5 Hz, H-3); 3.68 (dd,
1H, J2,1 = 9.3 Hz, J2,3 = 9.3 Hz, H-2); 4.00 (dd, 1H,
J6b,5 = 2.1 Hz, J6b,6a = 12.4 Hz, H-6b); 4.19–4.39 (m,
1H NMR (300 MHz, CDCl3): d 3.21 (m, 1H, N3CH2);
3.38 (m, 1H, N3CH2); 3.60–3.74 (m, 3H, HI-6b, OCH2,
HI-6a); 3.81–3.93 (m, 2H, HI-5, HII-5); 4.01–4.19 (m,
4H, HII-2, OCH2, HII-6b, HII-6a); 4.33 (dd, 1H,
J4,3 = 9.0 Hz, J4,5 = 9.6 Hz, HI-4); 4.50 (dd, 1H, J2,1
8.6 Hz, J2,3 = 10.6 Hz, HI-2); 4.82 (d, 1H, J1,2
7.9 Hz, HII-1); 5.34 (dd, 1H, J3,2 = 10.3 Hz, J3,4
=
=
=
3.0 Hz, HII-3); 5.61 (d, 1H, J1,2 = 8.6 Hz, HI-1); 5.67
6H, H-6b, H-5, 2 Â C@CCH2); 4.45 (d, 1H, J1,2
=
(d, 1H, J4,3 = 3.0 Hz, HII-4); 6.24 (dd, 1H, J3,2
=
9.3 Hz, H-1); 5.17 (m, 2H, 2 Â CHb@CC); 5.30 (m,
2H, 2 Â CHa@CC); 5.51 (s, 1H, PhCH); 5.96 (m, 2H,
2 Â C@CHC); 7.31–7.56 (m, 5H, PhC).
ESIMS: calcd for C22H30O5NaS [M+Na]+ 429.17,
found 429.3.
10.6 Hz, J3,4 = 9.0 Hz, HI-3); 7.10–8.10 (m, 24H, Ph).
ESIMS: calcd for C50H44N4NaO16 [M+Na]+ 979.27,
found 979.4.
4.11. 2-Azidoethyl 2,3,4,6-tetra-O-acetyl-b-D-galacto-
pyranosyl-(1?4)-2,3,6-tri-O-acetyl-b-D-glucopyranosyl-
(1?6)-[3,4,6-tri-O-benzoyl-b-D-galactopyranosyl-(1?4)]-
3-O-benzoyl-2-deoxy-2-phthalimido-b-D-glucopyranoside
(16)
4.9. 2-Azidoethyl 2-O-acetyl-3,4,6-tri-O-benzoyl-b-D-
galactopyranosyl-(1?4)-3-O-benzoyl-2-deoxy-2-phthal-
imido-6-O-tert-butyldimethylsilyl-b-D-glucopyranoside (14)
As described in the general procedure, 7 (74 mg, 0.124
mmol) and 9 (92.6 mg, 0.136 mmol) were coupled,
and the product was purified by column chromatogra-
phy with 2:1 petroleum ether–EtOAc as the eluent
to give 14 (102 mg, 0.092 mmol, 67%) as a foamy
solid.
As described in the general procedure, 15 (237 mg,
0.248 mmol) and 11 (203 mg, 0.260 mmol) were coupled,
and the product was purified by column chromato-
graphy with 1:1–1:2 petroleum ether–EtOAc as the
eluent to give 16 (197 mg, 0.125 mmol, 50%).
1H NMR (600 MHz, CDCl3): d 1.96 (s, 3H, OAc);
2.01 (s, 3H, OAc); 2.03 (s, 3H, OAc); 2.04 (s, 3H,
OAc); 2.08 (s, 3H, OAc); 2.10 (s, 3H, OAc); 2.12 (s,
3H, OAc); 2.14 (s, 3H, OAc); 3.22 (m, 1H, N3CH2);
3.40 (m, 1H, N3CH2); 3.53 (dd, 1H, J6b,5 = 7.4 Hz,
J6b,6a = 11.8 Hz; HII-6b); 3.62–3.74 (m, 3H, HIII-5,
HII-5, OCH2); 3.76-4.17 (m, 2H, HII-6a, HIII-4, HIV-5,
HI-5, HI-6b, OCH2, HII-2, HI-4, HIII-6b, HIV-6b, HIV-
6a); 4.29 (dd, 1H, J6a,5 = 1.2 Hz, J6a,6b = 9.6 Hz, HI-
6a); 4.48 (dd, 1H, J2,1 = 8.9 Hz, J2,3 = 11.0 Hz; HI-2);
1H NMR (300 MHz, CDCl3): d 0.14 (s, 6H,
2 Â SiCH3); 0.91 (s, 3 Â 3H, SiC(CH3)3); 1.98 (s, 3H,
OAc); 3.21 (m, 1H, N3CH2); 3.38 (m, 1H, N3CH2);
3.60–3.74 (m, 3H, HI-5, OCH2, HII-6b); 3.84–4.04 (m,
5H, HII-6a, HII-5, HI-6b, HI-6a, OCH2); 4.24 (dd, 1H,
J4,3 = 9.0 Hz, J4,5 = 9.6 Hz, HI-4); 4.43 (dd, 1H, J2,1
8.6 Hz, J2,3 = 10.6 Hz, HI-2); 4.95 (d, 1H, J1,2
7.9 Hz, HII-1); 5.22 (dd, 1H, J3,2 = 10.3 Hz, J3,4
3.0 Hz, HII-3); 5.38 (d, 1H, J2,1 = 7.9 Hz, J2,3
=
=
=
=
10.3 Hz, HII-2); 5.51 (d, 1H, J1,2 = 8.6 Hz, HI-1); 5.71
4.53 (d, 1H, J1,2 = 8.3 Hz, HIV-1); 4.59 (d, 1H, J1,2
7.6 Hz, HII-1); 4.64 (dd, 1H, J6a,5 = 1.2 Hz, J6a,6b
=
=
(d, 1H, J4,3 = 3.0 Hz, HII-4); 6.13 (dd, 1H, J3,2
=
10.6 Hz, J3,4 = 9.0 Hz, HI-3); 7.10–8.10 (m, 24H,
Ph). ESIMS: calcd for C58H60N4NaO17Si [M+Na]+
1135.36, found 1135.1.
10.3 Hz, HIII-6a); 4.95–5.00 (m, 2H, HIV-3, HIII-2);
5.11 (dd, 1H, J2,1 = 8.3 Hz, J2,3 = 10.3 Hz, HIV-2);
5.21 (dd, 1H, J3,2 = 8.9 Hz, J3,4 = 8.9 Hz, HIII-3); 5.27
(dd, 1H, J3,2 = 10.4 Hz, J3,4 = 3.5 Hz, HII-3); 5.35 (d,
1H, J4,3 = 3.4 Hz, HIV-4); 5.52 (d, 1H, J1,2 = 8.2 Hz,
HI-1); 5.65 (d, 1H, J4,3 = 2.8 Hz, HII-4); 6.15 (dd, 1H,
J3,2 = 11.0 Hz, J3,4 = 8.9 Hz, HI-3); 7.10-8.10 (m, 24H,
Ph).
4.10. 2-Azidoethyl 3,4,6-tri-O-benzoyl-b-D-galactopyr-
anosyl-(1?4)-3-O-benzoyl-2-deoxy-2-phthalimido-b-D-
glucopyranoside (15)
To a solution of 14 (350 mg, 0.314 mmol) in 10 mL of
1:1 MeOH–CH2Cl2 was added 500 lL of AcCl at 0 °C.
The mixture was stirred at rt for 24 h and then neutral-
ized with Et3N. The solvent was evaporated, and the res-
idue was dissolved in the mixture of EtOAc and water.
The organic phase was separated from the water
phase. The water phase was extracted two times with
EtOAc. The combined organic layers were dried with
Na2SO4. The solvent was evaporated, and the residue
was purified by silica gel flash chromatography (1:1–
1:2 petroleum ether–EtOAc) to give 15 (237 mg,
0.248 mmol, 79%).
ESIMS: calcd for C76H79N4O33 [M+H]+ 1597.44,
found 1597.8.
4.12. 2-Azidoethyl 2,3,4,6-tetra-O-acetyl-b-D-galactopyr-
anosyl-(1?4)-2,3,6-tri-O-acetyl-b-D-glucopyranosyl-
(1?6)-[2,3-di-O-allyl-4,6-O-benzylidene-a-D-galactopyr-
anosyl-(1?2)-3,4,6-tri-O-benzoyl-b-D-galactopyranosyl-
(1?4)]-3-O-benzoyl-2-deoxy-2-phthalimido-b-D-gluco-
pyranoside (17)
To a solution of the donor 13 (36 mg, 0.086 mmol) and
the tetrasaccharide acceptor 16 (90 mg, 0.057 mmol) in

Y.-X. Chen et al. / Carbohydrate Research 343 (2008) 607–614
613
˚
10 mL dry CH2Cl2 was added 50 mg of dry 4 A molec-
ular sieves under argon. The mixture was stirred at rt
for 0.5 h, followed by the addition of 10 lL of TMSOTf
and NIS (24 mg, 0.107 mmol) at À10 °C. The mixture
was stirred at rt for 2 h, neutralized with Et3N and fil-
tered. The filtrate was concentrated. The residue was
purified by silica gel flash chromatography (1:1–1:2
petroleum ether–EtOAc) to give 17 (75 mg, 0.039 mmol,
68%).
catalyst, the filtrate was concentrated. The residue was
dissolved in 5 mL of pyridine, followed by the addition
of 5 mL of Ac2O at 0 °C. The mixture was stirred at rt
overnight and then concentrated. The residue was
purified by silica gel flash chromatography (1:1–1:2
petroleum ether–ethyl acetate) to give 18 (61 mg,
0.032 mmol). The total yield was 86%.
1H NMR (600 MHz, CDCl3): d 1.80 (s, 3H, OAc);
1.969 (s, 3H, OAc); 1.973 (s, 3H, OAc); 2.04 (s, 3H,
OAc); 2.061 (s, 3H, OAc); 2.065 (s, 3H, OAc); 2.07 (s,
3H, OAc); 2.16 (s, 6H, 2 Â OAc); 2.18 (s, 3H, OAc);
2.28 (s, 3H, OAc); 3.25 (m, 1H, N3CH2); 3.41 (m, 1H,
J6b,5 = 8.2 Hz, J6b,6a = 11 Hz, N3CH2); 3.66–3.70 (m,
3H, HIII-5, OCH2, HI-6b); 3.76–3.79 (m, 2H, HI-5,
HV-5); 3.87–3.96 (m, 3H, HII-3, HIV-5, HIII-4); 4.02–
4.18 (m, 9H, HII-6a, HI-6a, OCH2, HIV-6b, HIV-6a,
1H NMR (600 MHz, CDCl3): d 1.97 (s, 3H, OAc);
2.05 (s, 6H, 2 Â OAc); 2.06 (s, 3H, OAc); 2.15 (s, 6H,
2 Â OAc); 2.17 (s, 3H, OAc); 3.16 (d, 1H,
J6b,6a = 12.6 Hz, HV-6b); 3.27 (m, 1H, N3CH2); 3.38–
3.44 (m, 2H, HV-5, N3CH2); 3.56–3.64 (m, 2H, HIII-5,
OCH2) 3.75 (d, 1H, J4,3 = 3.5 Hz, HV-4); 3.86–3.94 (m,
4H, HIII-4, HI-6b, HIV-5 HV-2); 4.00–4.35 (m, 14H,
HII-6a, HI-5, HII-5, HIII-6b, 2 Â C@CCH2, OCH2,
HII-2, HI-4, HI-6a, HIV-6b, HIV-6a); 4.47 (dd, 1H,
HV-6b, HV-6a, HIII-6b, HII-5); 4.28 (t, 1H, J4,3
=
J4,5 = 9 Hz, HI-4); 4.48 (dd, 1H, J2,1 = 8.2 Hz,
J2,3 = 11.0 Hz, HI-2); 4.56 (d, 1H, J1,2 = 7.6 Hz, HIV-
1); 4.66 (dd, 1H, J6a,5b = 2.0 Hz, J6a,6b = 12.4 Hz, HIII-
6a); 4.78 (d, 2H, J1,2 = 7.6 Hz, HII-1, HIII-1); 4.97–5.02
(m, 2H, HIII-2, HIV-3); 5.06 (dd, 1H, J3,2 = 11.0 Hz,
J3,4 = 3.4 Hz, HV-3); 5.12–5.16 (m, 2H, HIV-2, HV-1);
5.18 (dd, J2,1 = 2.8 Hz, J2,3 = 11.0 Hz, 1H, HV-2); 5.24
J1 = 8.9 Hz, J2,3 = 11.0 Hz, HI-2); 4.52 (d, 1H, J1,2
=
8.1 Hz, HIV-1); 4.58 (d, 1H, J6a,6b = 11.6 Hz, HIII-6a);
4.69 (d, 1H, J1,2 = 8.06 Hz, HII-1); 4.79 (d, 1H,
J1,2 = 8.2 Hz, HIII-1); 4.96–5.00 (m, 2H, HIV-3, HIII-2);
5.10–5.14 (m, 2H, HIV-2, CH@CC); 5.19 (s, 1H, PhCH);
5.20–5.26 (m, 2H, HIII-3, CH@CC); 5.32–5.42 (m, 4H,
CH@CC, HIV-4, CH@CH, HV-1); 5.47–5.53 (m, 2H,
HI-1, HII-3); 5.67 (d, 1H, J4,3 = 3.5 Hz, HII-4); 5.84–
(t, 1H, J3,2 = J3,4 = 9.0 Hz, HIII-3); 5.35 (d, 1H, J4,3
=
3.4 Hz, HV-4); 5.36 (d, 1H, J4,3 = 2.8 Hz, HIV-4); 5.47
6.02 (m, 2H, 2 Â C@CHC); 6.05 (dd, 1H, J3,2
=
(d, 1H, J1,2 = 8.2 Hz, HI-1); 5.49 (dd, 1H, J3,2
10.3 Hz, J3,4 = 3.5 Hz, HII-3); 5.68 (d, 1H, J4,3
=
=
=
11.0 Hz, J3,4 = 9.1 Hz, HI-3); 7.16–8.03 (m, 29H, Ph).
ESIMS: calcd for C95H100N4NaO38 [M+Na]+ 1927.59,
found 1927.6.
3.5 Hz, HII-4); 6.08 (dd, 1H, J3,2 = 11.0 Hz, J3,
4
9.0 Hz, HI-3); 7.16–8.03 (m, 29H, Ph). ESIMS:
calcd for C90H96N4NaO42 [M+Na]+ 1927.54, found
1927.7.
4.13. 2-Azidoethyl 2,3,4,6-tetra-O-acetyl-b-D-galactopyr-
anosyl-(1?4)-2,3,6-tri-O-acetyl-b-D-glucopyranosyl-
(1?6)-[2,3,4,6-tetra-O-acetyl-a-D-galactopyranosyl-
(1?2)-3,4,6-tri-O-benzoyl-b-D-galactopyranosyl-(1?4)]-
3-O-benzoyl-2-deoxy-2-phthalimido-b-D-glucopyranoside
(18)
4.14. 2-Azidoethyl 2,3,4,6-tetra-O-acetyl-b-D-galactopyr-
anosyl-(1?4)-2,3,6-tri-O-acetyl-b-D-glucopyranosyl-
(1?6)-[2,3,4,6-tetra-O-acetyl-a-D-galactopyranosyl-
(1?2)-3,4,6-tri-O-acetyl-b-D-galactopyranosyl-(1?4)]-2-
acetamido-3-O-acetyl-2-deoxy-b-D-glucopyranoside (19)
To a solution of pentasaccharide 17 (70 mg, 0.037
mmol) in 5 mL of CH2Cl2 were added 400 lL of
CF3CO2H and 250 lL of water. The mixture was stirred
at rt for 24 h, followed by neutralization with Et3N.
After the solvent was evaporated, the residue was dis-
solved in the mixture of EtOAc and water. The organic
phase was separated from the water phase. The water
phase was extracted two times with EtOAc. The com-
bined organic layers were dried with Na2SO4. The sol-
vent was evaporated, and the crude product was
dissolved in 5 mL of pyridine, followed by the addition
of 5 mL of Ac2O at 0 °C. The mixture was stirred at rt
overnight and then concentrated. The residue was puri-
fied by silica gel flash chromatography (1:1–1:2 petro-
leum ether–EtOAc) to give an intermediate product,
which was dissolved in 10 mL of 1:1 MeOH–CH2Cl2.
After the addition a catalytic amount of PdCl2, the mix-
ture was stirred at rt for 8 h. After the filtration of the
To a solution of 18 (50 mg, 0.026 mmol) in 5 mL of
MeOH was added NaOMe (pH 10), and the mixture
was stirred for 24 h. After neutralization with Dowex
50WX8 (H+) and filtration, the mixture was concen-
trated. To a solution of the residue in 10 mL of 1-BuOH
was added 2 mL of 1,2-diaminoethane, and the mixture
was stirred overnight at 80 °C, then, co-concentrated
with toluene. A solution of the residue in 10 mL of pyr-
idine and 10 mL of Ac2O was stirred overnight, then co-
concentrated with toluene. The residue was purified by
silica gel flash chromatography (1:1–1:2 petroleum
ether–EtOAc) to give 19 (32 mg, 0.020 mmol). The total
yield was 77%.
1H NMR (600 MHz, CDCl3): d 1.95 (s, 3H, NHAc);
1.97 (s, 3H, OAc); 1.987 (s, 3H, OAc); 1.991 (s, 3H,
OAc); 2.047 (s, 6H, 2 Â OAc); 2.055 (s, 3H, OAc); 2.06
(s, 9H, 3 Â OAc); 2.09 (s, 3H, OAc); 2.13 (s, 3H,

614
Y.-X. Chen et al. / Carbohydrate Research 343 (2008) 607–614
OAc); 2.14 (s, 6H, 2 Â OAc); 2.15 (s, 3H, OAc); 2.16 (s,
3H, OAc); 3.32 (m, 1H, N3CH2); 3.46–3.53 (m, 2H,
in the online version, at doi:10.1016/j.carres.
2007.12.002.
N3CH2, HIII-5); 3.63–3.69 (m, 3H, OCH2, HII-2, HIII
-
4, OCH2); 3.78 (m, 1H, HI-2); 3.88–4.18 (m, 14H, HII-
6b, HII-5, HI-6b, HI-5, OCH2, HIV-6b, HIV-6a, HIV-5,
HV-6b, HV-6a, HV-5, HIII-6b, HIII-6a, HI-4); 4.28 (m,
1H, HII-6a); 4.52–4.57 (m, 3H, HIV-1, HII-1, HI-6a);
References
1. Fine, M. J.; Smith, M. A.; Carson, C. A.; Mutha, S. S.;
Sankey, S. S.; Weissfeld, L. A.; Kapoor, W. N. J. Am.
Med. Assoc. 1996, 275, 134–141.
4.68 (d, 1H, J1,2 = 6.8 Hz, HIII-1); 4.71 (d, 1H, J1,2
7.6 Hz, HI-1); 4.70 (dd, 1H, J2,1 = 6.8 Hz, J2,3
=
=
8.1 Hz, HIII-2); 4.97 (dd, 1H, J3,2 = 10.3 Hz,
J3,4 = 3.4 Hz, HIV-3); 5.05 (dd, 1H, J3,2 = 9.7 Hz,
J3,4 = 4.1 Hz, HII-3); 5.07 (dd, 1H, J2,1 = 3.4 Hz,
J2,3 = 10.3 Hz, HV-2); 5.12 (dd, 1H, J2,3 = 10.3 Hz,
J2,1 = 8.2 Hz, HIV-2); 5.16–5.24 (m, 3H, HIII-3, HV-3,
HI-3); 5.35–5.32 (m, 3H, HIV-4, HII-4, HV-1); 5.61
(d, 1H, JNH,HI-2 = 8.3 Hz). ESIMS: calcd for
C64H88N4NaO41 [M+Na]+ 1591.48, found 1591.9.
2. Robbins, J. B.; Austrian, R.; Lee, C.-J.; Rastogi, S. C.;
Schiffman, G.; Henrichsen, J.; Makela, P. H.; Broome, C.
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4.15. 2-Azidoethyl b-D-galactopyranosyl-(1?4)-b-D-
glucopyranosyl-(1?6)-[a-D-galactopyranosyl-(1?2)-
b-D-galactopyranosyl-(1?4)]-2-acetamido-2-deoxy-
b-D-glucopyranoside (1)
To a solution of 19 (30 mg, 0.019 mmol) in 5 mL of
MeOH was added NaOMe (pH 10), and the mixture
was stirred for 48 h. After neutralization with Dowex
50WX8 (H+) and filtration, the mixture was concen-
trated. The residue was eluted on a Sephadex LH-20 col-
umn with MeOH to give 1 (15 mg, 0.016 mmol, 84%) as
a white solid.
8. Benaissa-Trouw, B.; Lefeber, D. J.; Kamerling, J. P.;
Vliegenthart, J. F. G.; Kraaijeveld, K.; Snippe, H. Infect.
Immun. 2001, 69, 4698–4701.
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Bioorg. Med. Chem. 1998, 6, 1605–1612.
10. Niggemann, J.; Kamerling, J. P.; Vliegenthart, J. F. G.
J. Chem. Soc., Perkin Trans. 1 1998, 3011–3020.
11. Joosten, J. A. F.; Kamerling, J. P.; Vliegenthart, J. F. G.
Carbohydr. Res. 2003, 338, 2611–2627.
12. Joosten, J. A. F.; Lazet, B. J.; Kamerling, J. P.; Vliegent-
hart, J. F. G. Carbohydr. Res. 2003, 338, 2629–2651.
13. Michalik, D.; Vliegenthart, J. F. G.; Kamerling, J. P. J.
Chem. Soc., Perkin Trans. 1 2002, 1973–1981.
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Kamerling, J. P.; Vliegenthart, J. F. G. Infect. Immun.
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1H NMR (300 MHz, D2O): d 1.95 (s, 3H, NHAc);
3.25–4.04 (m, 20H); 4.17–4.24 (m, 2H); 4.38 (d, 1H,
J = 7.6 Hz); 4.46 (d, 1H, J = 8.2 Hz); 4.50 (d, 1H, J =
7.9 Hz); 4.60–4.80 (m, H2O); 5.32 (d, 1H, J = 3.7 Hz).
ESIMS: calcd for C34H58N4NaO26 [M+Na]+ 961.32,
found 961.5.
HRESIMS: calcd for C34H59N4O26 [M+H]+
939.34120, found 939.34215.
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409.
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Acknowledgements
This work was supported by The National Natural
Science Foundation of China (Projects 20472041 and
20532020).
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Supplementary data
NMR spectra of 7, 14, 16, 17, 18, 19 and 1. Supple-
mentary data associated with this article can be found,