Synthesis of rhamnogalacturonan I fragments by a modular design principle

Article abstract of DOI:10.1016/j.carres.2008.03.020

The improved syntheses of methyl 2-O-acetyl-3-O-benzyl-α-l-rhamnopyranoside (12) and 1,2-di-O-acetyl-3-O-benzyl-α-l-rhamnopyranose (15), which were used as glycosyl acceptor and donor, respectively, are described. Glycosylation of the O-4 position of both rhamnose derivatives with 2,3,4,6-tetra-O-benzoyl-α-d-galactopyranosyl bromide (26) provided disaccharides 27 and 29. After partial deprotection of 27 and coupling of the resulting 28 with disaccharide 19, tetrasaccharide 31 was obtained. Furthermore, transforming of 29 into the corresponding bromide 30 and coupling with galacturonates 16 and 32 provided trisaccharides 33 and 34, respectively, which could be regarded as building blocks of ramified rhamnogalacturonan fragments. The preparation of tetra- (21) and hexasaccharide (25) of rhamnogalacturonan I is reported to demonstrate the feasibility of the synthesis of larger pectin fragments using the modular design principle with this type of building blocks.

Full text of DOI:10.1016/j.carres.2008.03.020

Available online at www.sciencedirect.com
Carbohydrate Research 343 (2008) 1730–1742
Synthesis of rhamnogalacturonan I fragments
by a modular design principleI
Navid Nemati,a, Gnuni Karapetyan,a Birte Nolting,a,à Hans-Ulrich Endressb
and Christian Vogela,*
aUniversity of Rostock, Institute of Chemistry, Albert-Einstein-Strasse 3a, D-18059 Rostock, Germany
bHerbstreith & Fox, Pectin-Factory Neuenbuerg, Turnstrasse 37, D-75305 Neuenbuerg, Germany
Received 5 February 2008; received in revised form 3 March 2008; accepted 12 March 2008
Available online 18 March 2008
Abstract—The improved syntheses of methyl 2-O-acetyl-3-O-benzyl-a-L-rhamnopyranoside (12) and 1,2-di-O-acetyl-3-O-benzyl-a-
L-rhamnopyranose (15), which were used as glycosyl acceptor and donor, respectively, are described. Glycosylation of the O-4 posi-
tion of both rhamnose derivatives with 2,3,4,6-tetra-O-benzoyl-a-D-galactopyranosyl bromide (26) provided disaccharides 27 and
29. After partial deprotection of 27 and coupling of the resulting 28 with disaccharide 19, tetrasaccharide 31 was obtained.
Furthermore, transforming of 29 into the corresponding bromide 30 and coupling with galacturonates 16 and 32 provided trisac-
charides 33 and 34, respectively, which could be regarded as building blocks of ramified rhamnogalacturonan fragments. The prep-
aration of tetra- (21) and hexasaccharide (25) of rhamnogalacturonan I is reported to demonstrate the feasibility of the synthesis of
larger pectin fragments using the modular design principle with this type of building blocks.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords: D-Galacturonic acid; L-Rhamnose; D-Galactose; Oligosaccharides; Pectin fragments; Glycosylation; Building blocks
1. Introduction
arabinose.5 The enzymes responsible for the glycosyl-
ation of rhamnose residues with the first galactose or
arabinose molecule have not been identified at this
point.§,6,7 Therefore, classical synthetic carbohydrate
chemistry must be used to obtain these structures.8 Pur-
suing our programme directed towards the chemical syn-
thesis of well-defined pectin fragments for structure–
activity relationship investigations,9,10 we report herein
the preparation of a rhamnogalacturonan I (RG-I) tetra-
and hexasaccharide by means of a modular design prin-
ciple. Furthermore, the synthesis of building blocks for
D-galactose-ramified RG-I fragments is described.
Recent studies have shown that many plants used in
traditional medicine to treat various types of diseases
contain polysaccharides exhibiting biological activity of
different kinds. Pectins are an example for complex
polysaccharides possessing such an activity.1,2 Pectin
structures consist of a homogalacturonan backbone
interrupted by ramified rhamnogalacturonan regions.3,4
The latter, the so-called hairy region, contains a-(1?2)-
linked L-rhamnose units carrying neutral carbohydrate
side chains, typically composed of D-galactose and D-
q Part XV of the series ‘Galacturonic Acid Derivatives’, for part XIV
see Ref. 10.
2. Results and discussion
*
Corresponding author. Tel.: +49 381 498 6430; fax: +49 381 498
6412; e-mail: christian.vogel@uni-rostock.de
The backbone of the RG-I polymer is composed of
repeating units of the disaccharide ?2)-a-L-Rhap-
(1?4)-a-D-GalpA-(1?. In these oligomers, the first
Present address: Leibniz Institute for Catalysis at University Rostock,
Branch Berlin: Richard-Willstaetter-Strasse 12, D-12489 Berlin,
Germany.
à Present address: Wyeth Vaccine, 401 N. Middletown Rd., Pearl
River, NY 10965, USA.
§ Information from Henrik V. Scheller.
0008-6215/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2008.03.020

N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
1731
D-galactopyranose residue is b-glycosidically linked to
the O-4 position of a rhamnose moiety. Our strategy
adopted for the synthesis of larger RG-I oligomers, as
well as building blocks for D-galactose-ramified RG-I
fragments, involved in the preparation of L-rhamnose
intermediates that allowed stereoselective formation of
the required a-glycosidic bond to D-galacturonates, the
regioselective deblocking of a protected O-4 position,
the preparation of RG-I-modules and finally the cou-
pling of these modules by a modular design principle.
For the synthesis of suitable L-rhamnose derivatives,
we evaluated two routes as shown in Scheme 1. Methyl
a-L-rhamnopyranoside (1)11 was isopropylidenated
(2),12 allylated (3),13 and then hydrolyzed13 to give
intermediate 4. For the allylation reaction, N,N-dimeth-
ylformamide (DMF) was replaced by dry tetrahydrofu-
ran as the solvent to avoid the need for the tedious
removal of residual traces of the former solvent.
Treatment of 4 with triethylorthoacetate in CH2Cl2
followed by regioselective opening of the orthoester
structure in 614 led to the formation of an acetyl protect-
ing group at the O-2 position (10). This acetyl group was
involved in neighbouring participation in a later glycos-
ylation reaction and served as a temporary protecting
group during the block wise synthesis of higher pectin
fragments. The regioselective ring opening of orthoester
1
6 was proven by H NMR spectroscopy. As expected,
the acetyl substituent at O-2 position caused a consider-
able downfield shift of the H-2 signal from d 3.90 ppm in
the 1H NMR spectrum of compound 4 to d 5.01 ppm in
the spectrum of 10.
In a previous paper, we described an alternative route
targeting the preparation of the 4-O-benzoyl derivative
8.15 Here, the formation of the orthoester structure in
5 could only be achieved in a solvent mixture of DMF
and toluene. In multi-gram scale experiments, the use
of DMF as a solvent crucially prolonged work-up times
resulting in the migration of the acetyl groups in 8 as
well as in 10. Therefore, for scale-up, the pathway via
the isopropylidene derivative 2 was more convenient
because this route avoided DMF in every reaction step.
Benzylation of 8 and 10 using benzyl trichloroacetim-
idate and a catalytic amount of trifluoromethanesulfonic
acid16 provided 915 and 11 in 64% and 78% yield, respec-
tively. Acetolysis17 of 9 and 11 gave predominantly the
a-acetate 13a15 and compound 14 in 84% and 71%,
respectively. Small amounts of the b-acetate (14b) could
be isolated by flash chromatography. The stereochemis-
try at the anomeric centre was evident only from the
OH
O
HO
O
a
HO
RO
O
O
OMe
OMe
1
2 R = H
3 R = All
b
e
c
CH3
O
EtO
OH
HO
O
d
AllO
RO
O
O
OMe
OMe
5 R = H
6 R = All
7 R = Bz
4
f
g
h
OAc
R2O
OAc
BnO
R1O
l
RO
geminal 13C–1H coupling constants JC-1,H-1
. The
O
O
observed values of 173.5 Hz and 161.5 Hz for 14a and
14b, respectively, verified the proposed structures.18
Deallylation of compounds 11 and 14 with the aid of
palladium(II)chloride19 provided intermediates 12 and
15 in 63% and 67% yield, respectively, which served as
glycosyl acceptors for the introduction of a galactose
residue into the O-4 position.
The 4-O-benzoyl derivate 13 was employed as glycosyl
donor for the preparation of disaccharide module 17 by
coupling with the galacturonate acceptor 16 in the pres-
ence of trimethylsilyl trifluoromethanesulfonate
(Scheme 2). The disaccharide synthesis as well as the
transformation of module 17 into either glycosyl accep-
tor 20 by selective deacetylation of the O-20 position
with 0.28 M methanolic hydrochloric acid20 or into
glycosyl donor 19 by deallylation and the introduction
of the trichloroacetimidate function have been described
previously.15 Contrary to the results observed by
Reiffarth and Reimer21 who employed a similar block
synthetic approach,22 trimethylsilyl trifluoromethane-
OAc
OMe
8 R1 = Bz R2 = H
9 R1 = Bz R2 = Bn
10 R1 = All R2 = H
11 R1 = All R2 = Bn
12 R1 = H R2 = Bn
13 R = Bz
14 R = All
15 R = H
i
k
i
k
Scheme 1. Alternative pathways for the preparation of L-rhamnose
derivatives 12 and 15 used as glycosyl acceptors for the introduction of
a D-galactose residue at their O-4 positions. Reagents and conditions:
(a) anhyd CuSO4, dry acetone, 10 h, 20 °C (2 98%); (b) allyl bromide,
NaH, dry THF, 20 h, 20 °C, Ar atmosphere (3 75%); (c) H2SO4 (cat.),
EtOH–toluene (1:1), 2 h, reflux (4 86%); (d) CH(OEt)3, camphorsul-
fonic acid, dry CH2Cl2, 35 min, 20 °C, Ar atmosphere (6 98%); (e)
CH(OEt)3, camphorsulfonic acid, dry DMF, 40 min, 20 °C, Ar
atmosphere, (5 65%); (f) allyl bromide, NaH, dry DMF, 3 h,
0 °C?20 °C, Ar atmosphere (6 98%); (g) benzoyl chloride, dry
pyridine, 9 h, ꢀ20 °C?20 °C, Ar atmosphere (7 98%); (h) 60% aq
acetic acid, 30 min, 20 °C (8 90%, 10 91%); (i) benzyl 2,2,2-trichloro-
acetamide, CF3SO3H (cat.), dry CH2Cl2–heptane, 22 h,
ꢀ10 °C?20 °C, Ar atmosphere (9 64%, 11 78%); (k) PdCl2, AcOH,
NaOAc, H2O, 4–6 h, 40 °C (12 63%, 15 67%); (l) Ac2O, AcOH, H2SO4
(cat.), 3 h, ꢀ30 °C?0 °C (13 91%, 14 77%).

1732
N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
OAc
BnO
HO
BzO
OAc
CO2Me
BnO
O
O
a
BzO
+
OAll
O
BnO
O
CO2Me
BnO
OAc
O
13
16
OAll
BnO
BnO
17
b
d
OAc
O
OH
BnO
BnO
BzO
BzO
O
O
O
CO2Me
CO2Me
O
O
OAll
BnO
18 R = OH
BnO
BnO
BnO
R
20
c
19 R = OC(=NH)CCl3
e
OR1
O
R2O
R3O
O
4
CO2R
O
R2O
R2O
O
R2O
R3O
O
4
O
CO2R
O
OR5
R2O
R2O
21 R1 = Ac R2 = Bn R3 = Bz R4 = Me R5 = All
22 R1 = H R2 = Bn R3 = Bz R4 = Me R5 = All
f
g
h
23 R1 = Ac R2 = H R3 = Bz R4 = Me R5= Propyl
24 R1 = R2 = R3 = R4 = H R5= Propyl
Scheme 2. Synthesis of rhamnogalacturonan tetrasaccharide 21 by a modular design principle and its partial deprotection. Reagents and conditions:
(a) 13:16 molar ratio 1.3:1, CF3SO3SiMe3, mol. sieves, dry CH2Cl2, 18 h, 20 °C, Ar atmosphere, darkness (17, 88%); (b) PdCl2, AcOH, NaOAc, H2O,
4–6 h, 40 °C (18 75%); (c) trichloroacetonitrile, DBU (cat.), dry CH2Cl2, 2 h, ꢀ20 °C?20 °C, Ar atmosphere (19 69%); (d) 0.28 M methanolic HCl,
36 h, 20 °C, Ar atmosphere (20 98%); (e) CF3SO3SiMe3, mol. sieves, dry CH2Cl2, 21 h, ꢀ70 °C?20 °C, Ar atmosphere, darkness (21 60%); (f) 0.28 M
methanolic HCl, 48–72 h, 20 °C, Ar atmosphere (22 97%); (g) Pd/C, H2, EtOH, 18 h, 20 °C (23 97%); (h) LiOH, MeOH, H2O, 15 min, 20 °C (24
86%).
sulfonate-promoted glycosylation of acceptor 20 with
donor 19 in a molar ratio of 1:1, provided tetrasaccha-
ride 21 in 60% isolated yield after chromatographic puri-
fication (Scheme 2). In a model reaction, the benzyl
protecting groups of 21 were removed by hydrogenolysis
over Pd–C resulting in the propyl glycoside 23. The ano-
meric centre was not deprotected in these experiments to
maintain the configuration and obtain well-resolved
NMR spectra. Next, the cleavage of the ester linkages
was achieved in methanol and water in the presence of
lithium hydroxide at room temperature to provide the
nearly completely deprotected tetrasaccharide 24 in
excellent yield.
Selective deacetylation of tetrasaccharide 21 using
again 0.28 M methanolic hydrochloric acid yielded the
glycosyl acceptor 22, which was glycosylated (Scheme
3) with donor 19 to give hexasaccharide 25 in 59% yield
after HPLC purification. The a-configuration at the
newly generated stereogenic centre in the tetrasaccharide
as well as in the hexasaccharide was verified by 13C
NMR spectroscopy. For the completely protected
rhamnogalacturonate oligomers the a-linked anomeric

N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
1733
Furthermore, the coupling of the benzoylated galacto-
syl bromide 2623,24 prepared using the procedure by
Kochetkov and co-workers25 with either methyl rham-
noside 12 or diacetate 15 as glycosyl acceptors was
performed in the presence of silver trifluoromethane-
sulfonate (Scheme 4). After standard work-up and puri-
fication by HPLC, disaccharides 27 and 29 were
obtained in 66% and 68% yields, respectively. The pro-
tected disaccharide 27 was subjected to selective deacet-
ylation to give 58% of the glycosyl acceptor 28.
Alternatively, diacetate 29 was transformed in 90% yield
into the corresponding bromide 30 using the procedure
by Gillard and Israel.26 In an effort to evaluate the capa-
bility of the prepared galactosyl-rhamnosides to serve as
structural elements for the synthesis of ramified RG-1
fragments, acceptor 28 was glycosylated with rhamno-
galacturonate 19 to provide tetrasaccharide 31 in 62%
isolated yield. Again, traces (less than 5%) of a b-linked
isomer were observed in the 13C NMR spectra of the
crude reaction mixture.
OAc
O
BnO
BzO
O
CO2Me
a
O
22
19
+
BnO
BnO
BnO
O
BzO
O
2
O
CO2Me
O
OAll
BnO
BnO
25
Scheme 3. The extension of the synthesis of rhamnogalacturonan I
fragments by a modular design principle to furnish hexasaccharide 25.
Reagents and conditions: (a) CF3SO3SiMe3, mol. sieves, dry CH2Cl2,
21 h, ꢀ70 °C?20 °C, Ar atmosphere, darkness (25 59%).
carbons resonated between dC 97.60 and 99.77, while the
signals for the b-linked anomeric carbon (C-1) appeared
in the range of 102.72–102.84. All NMR data were in
accordance with the proposed structures. It should be
noted that in the crude glycosylation reaction mixtures,
traces (<5%) of b-linked oligosaccharides were observed
by NMR investigations. However, the total isolated
amounts of these isomers were too small for full
characterization.
Finally, the glycosylation of galacturonates 16 and
3227 with bromide 30 as donor gave trisaccharides 33
(68%) and 34 (74%), respectively (Scheme 4). Analytical
and spectral data of 33 and 34 confirmed their struc-
tures. To establish the 1,2-trans-glycosidic linkage
between L-rhamnose and D-galacturonate moieties,
the geminal 13C–1H coupling constants were again
BzO
BzO
OBz
OR2
OAc
BzO
BzO
OBz
BnO
BnO
O
a
O
HO
O
+
O
O
BzO
BzO
R1
Br
R
27 R1 = OMe R2 = Ac
28 R1 = OMe R2 = H
29 R1 = OAc R2 = Ac
30 R1 = Br R2 = Ac
26
12 R = OMe
15 R = OAc
b
c
+ 19
d
OAc
BnO
HO
CO2R
BzO
O
O
OAll
BnO
BnO
16 R = Me
32 R = Bn
O
CO2Me
O
e
BnO
BnO
BzO
BzO
OBz
O
BzO
BzO
OBz
OAc
O
BnO
BnO
O
O
O
O
O
BzO
BzO
OMe
O
CO2R
31
O
OAll
33 R = Me
34 R = Bn
BnO
BnO
Scheme 4. Synthesis of D-galactose-ramified rhamnogalacturonan fragments 31, 33 and 34 of which trisaccharides 33 and 34 represent potential
modules for larger oligomers. Reagents and conditions: (a) CF3SO3Ag, 2,6-tert-butyl-4-methyl-pyridine, mol. sieves, dry CH2Cl2, 20 h,
ꢀ65 °C?20 °C, Ar atmosphere, darkness (27 66%, 29 68%); (b) 0.28 M methanolic HCl, 12 h, 20 °C, Ar atmosphere (28 58%); (c) Me3SiBr, dry
CH2Cl2, 19 h, ꢀ10 °C?20 °C, Ar atmosphere (30 90%); (d) CF3SO3SiMe3, mol. sieves, dry CH2Cl2, 21 h, ꢀ70 °C?20 °C, Ar atmosphere, darkness
(31 62%); (e) CF3SO3Ag, 2,6-tert-butyl-4-methyl-pyridine, mol. sieves, dry CH2Cl2, 20 h, ꢀ65 °C?20 °C, Ar atmosphere, darkness (33 68%, 34 74%).

1734
N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
determined. Because the observed values of 33 and 34
by elution from columns of slurry-packed Silica Gel 60
(Merck, 40–63 lm) and Nucleosil 100-7 (KNAUER,
7.0 lm), respectively. All solvents and reagents were
purified and dried according to the standard proce-
dures.28 After classical work-up of the reaction
mixtures, organic layers were dried over MgSO4 and
then concentrated under reduced pressure (rotary
evaporator).
0
0
are JC-1 ;H-1 ¼ 170 ꢁ 0:5 Hz, a comparison with the data
reported for other a-L-rhamnose examples verified the
proposed structure.18 Based on these results, com-
pounds 33 and 34 are considered to represent trisaccha-
ride modules that can be transformed into glycosyl
donor as well as glycosyl acceptors comparable to the
disaccharide module 17, procedure shown in Scheme 2.
In conclusion, simple conversion of disaccharide 17
into both glycosyl donor 19 and glycosyl acceptor 20,
and their successful coupling to tetrasaccharide 21 veri-
fied our hypothesis that rhamnogalacturonates pro-
tected permanently by benzyl groups and temporarily
by acetyl and allyl groups are suitable as modules for
the synthesis of larger RG-1 fragments by a modular
design principle. This concept was extended to the prep-
aration of hexasaccharide 25. Furthermore, complete
deprotection with exception of the reducing end of tetra-
saccharide 21 was easily achieved in two steps. For the
synthesis of b-galactosyl ramified RG-1 fragments,
modules 33 and 34 were prepared and will be used in
future experiments.
3.2. Methyl 4-O-allyl-a-L-rhamnopyranoside (4)13
24
Colourless syrup; ½aꢃD ꢀ86.1 (c 1.0, CHCl3), lit.13 [a]D
ꢀ85.3 (c 0.57, CHCl3); 1H NMR (250.13 MHz, CDCl3):
3
d 1.29 (d, 3H, J5,6 = 6.4 Hz, H-6), 2.69 (br d, 2H,
2 ꢄ OH), 3.20 (t, 1H, H-4), 3.32 (s, 3 H, OCH3), 3.62
3
(m, 1H, H-5), 3.81 (dd, 1H, J3,4 = 9.5 Hz, H-3), 3.90
(dd, 1H, 3J2,3 = 3.4 Hz, H-2), 4.19 (m, 2H,
3
CH2CH@CH2), 4.62 (d, 1H, J1,2 = 1.5 Hz, H-1), 5.12,
5.27 (2m, 2H, CH2CH@CH2), 5.91 (m, 1H,
CH2CH@CH2); 13C NMR (62.89 MHz, CDCl3): d
17.88 (C-6), 54.78 (OCH3), 67.03 (C-5), 71.01 (C-2),
71.34 (C-3), 73.84 (CH2CH@CH2), 81.35 (C-4), 100.38
(C-1), 117.28 (CH2CH@CH2), 134.77 (CH2CH@CH2).
3. Experimental
3.3. Methyl 2-O-acetyl-4-O-allyl-a-L-rhamnopyranoside
(10)
3.1. General methods
Via 5: Sodium hydride (288 mg, 7.2 mmol, 60% disper-
sion in oil) was successively added to a stirred solution
of methyl 2,3-O-methylorthoacetyl a-L-rhamnopyrano-
side (5)15 (523 mg, 2.0 mmol) in dry N,N-dimethylform-
amide (8 mL) and dry toluene (8 mL) at 0 °C under an
argon atmosphere. The solution was kept 30 min at that
temperature, then allyl bromide (1 mL, 12.0 mmol) was
added. The mixture was allowed to attain room temper-
ature and stirring was continued for another 2 h. After
complete reaction (6, TLC: Rf = 0.53, solvent A),
MeOH (1 mL) was added at 0 °C, and after stirring
for additional 20 min, the reaction mixture was concen-
trated. After dissolving the residue in 60% aq acetic acid
(25 mL), the resulting solution was stirred 30 min at
ambient temperature (monitored by TLC). The reaction
mixture was then concentrated and repeatedly co-con-
centrated with toluene. The crude product was purified
by flash chromatography (eluent gradient EtOAc
0%?30% in heptane) to give 10 (453 mg, 87%) as a
colourless syrup.
Via 4: Camphorsulfonic acid (21 mg, 0.09 mmol) and
triethylorthoacetate (0.41 mL, 2.2 mmol) were succes-
sively added to a solution of methyl 4-O-allyl-a-L-
rhamnopyranoside13 (4, 218 mg, 1.0 mmol) in dry
CH2Cl2 (8 mL). After stirring for 35 min at ambient
temperature under an argon atmosphere (6, TLC:
Rf = 0.53, solvent A), triethylamine (1 mL) was added,
and the mixture was concentrated. The residue was dis-
solved in chloroform (50 mL), and the organic solution
Melting points were determined with a Boetius micro-
heating plate BHMK 05 (Rapido, Dresden) and were
not corrected. Optical rotations were measured for solu-
tions in a 2-cm cell with an automatic polarimeter
GYROMAT (Dr. Kernchen Co.). 1H NMR spectra
(500.13 MHz, 300.13 MHz and 250.13 MHz) and 13C
NMR
spectra
(125.76 MHz,
75.47 MHz
and
62.89 MHz) were recorded using Bruker instruments
AVANCE 500, ARX 300 and AC 250, with CDCl3,
CD3OD or Me2SO-d6 as solvents. NMR spectra were
1
calibrated using solvent signals (CDCl3: d H 7.25, d
1
13C 77.0; CD3OD: d H 4.78, d 13C 49.0; Me2SO-d6: d
1H 2.50, d 13C 39.7). The 1H and 13C NMR signals were
1
assigned by DEPT and two-dimensional H,1H COSY,
and 1H,13C correlation spectra (HMBC and HSQC).
Elemental analysis was performed on a CHNS-Flash-
EA-1112 instrument (Thermoquest). All washing solu-
tions were cooled to ꢂ5 °C. The NaHCO3 solution
was saturated. Reactions were monitored by thin-layer
chromatography (TLC, Silica Gel 60, F254, 0.25 mm,
Merck KGaA). The following solvent systems (v/v) were
used: (A) 1:1, (B) 2:1, (C) 3:1 and (D) 1:2 heptane–ethyl
acetate, (E) 10:1, (F) 1.5:1 and (G) 1:2 chloroform–
MeOH. The spots were made visible by spraying with
ethanolic 10% H2SO4 solution and charring them for
3–5 min with a heat gun. Detection of benzyl derivatives
was effected by UV fluorescence. Preparative flash
chromatography, MPLC and HPLC were performed

N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
1735
was washed with ice water (2 ꢄ 20 mL), dried and con-
centrated. Exactly the same procedure as above was
used for the orthoester ring opening to yield 10
Anal. Calcd for C19H26O6 (350.41): C, 65.13; H, 7.48.
Found: C, 65.17; H, 7.37.
22
(237 mg, 91%) as a colourless syrup: ½aꢃD ꢀ21.7 (c 1.0,
3.5. Acetolysis of methyl 2-O-acetyl-4-O-allyl-3-O-
benzyl-a-L-rhamnopyranoside (11)
1
CHCl3); Rf = 0.34 (solvent A); H NMR (250.13 MHz,
CDCl3): d 1.31 (d, 3H, H-6), 2.12 (s, 3 H, OCOCH3),
2.35 (br d, 1H, OH), 3.19 (t, 1H, H-4), 3.31 (s, 3H,
Concd sulfuric acid (0.16 mL) dissolved in acetic anhy-
dride (3 mL) was added to a solution of compound 11
(1.84 g, 5.25 mmol) in acetic anhydride–acetic acid
(2.5:1, v/v, 20 mL) in three portions at ꢀ30 °C over a
period of 1 h. After stirring for 1.5 h at 0 °C (monitored
by TLC), cold aq 10% K2SO4 (200 mL) was slowly
added and the stirring was continued for 30 min at
0 °C. The reaction mixture was extracted with chloro-
form (3 ꢄ 50 mL), and the combined organic phases
were washed successively with cold aq NaHCO3
(3 ꢄ 50 mL), water (2 ꢄ 50 mL) dried and concentrated.
The residue was co-concentrated with toluene (3 times)
to remove the remaining traces of acetic acid. The resi-
due was then purified by flash chromatography (eluent
gradient ethyl acetate 0%?50% in heptane) to provide
14a (1.41 g, 71%) and 14b (119 mg, 6%) in a ratio of
12:1, respectively.
3
OCH3), 3.64 (m, 1H, J5,6 = 6.4 Hz, H-5), 3.99 (dd,
3
1H, J3,4 = 9.5 Hz, H-3), 4.12, 4.21 (2m, 2H,
3
CH2CH@CH2), 4.59 (d, 1H, J1,2 = 1.5 Hz, H-1), 5.01
3
(dd, 1H, J2,3 = 3.7 Hz, H-2), 5.11, 5.21 (2m, 2H,
CH2CH@CH2), 5.91 (m, 1H, CH2CH@CH2); 13C
NMR (62.89 MHz, CDCl3): d 17.91 (C-6), 21.04
(OCOCH3), 54.89 (OCH3), 67.22 (C-5), 69.96 (C-3),
72.60 (C-2), 73.99 (CH2CH@CH2), 81.50 (C-4), 98.34
(C-1), 117.12 (CH2CH@CH2), 134.78 (CH2CH@CH2),
170.78 (OCOCH3). Anal. Calcd for C12H20O6 (260.28):
C, 55.37; H, 7.74. Found: C, 55.45; H, 7.69.
3.4. Methyl 2-O-acetyl-4-O-allyl-3-O-benzyl-a-L-
rhamnopyranoside (11)
A catalytic amount of trifluoromethanesulfonic acid
(14 lL, 0.15 mmol) dissolved in dry CH2Cl2 (1 mL)
was added to a solution of compound 10 (350 mg,
1.3 mmol) and benzyl 2,2,2 trichloroacetimidate
(0.5 mL, 2.7 mmol) in a mixture of dry CH2Cl2 (4 mL)
and dry heptane (7 mL) at ꢀ10 °C under an argon atmo-
sphere. After stirring for 3.5 h at that temperature, the
mixture was allowed to attain room temperature and
stirring was continued for another 18 h (monitored by
TLC). Then, the mixture was passed through a layer
of alkaline alumina by elution with CH2Cl2–heptane
(2:3, v/v) and concentrated. After suspending the residue
in heptane–diethyl ether (6:1, v/v, 10 mL) the formed
carbohydrate-free crystals were filtered off, washed with
heptane–diethyl ether (6:1, v/v, 20 mL) and the com-
bined filtrate and washings were concentrated. The
crude product was purified by flash chromatography
(eluent solvent C) to give compound 11 (367 mg, 78%)
3.6. 1,2-Di-O-acetyl-4-O-allyl-3-O-benzyl-a-L-rhamno-
pyranose (14a)
21
Colourless foam; ½aꢃD ꢀ28.2 (c 1.0, chloroform);
1
Rf = 0.68 (solvent A); H NMR (300.13 MHz, CDCl3):
d 1.24 (d, 3H, H-6), 2.00, 2.08 (2s, 6H, 2 ꢄ OCOCH3),
3
3.25 (t, 1H, H-4), 3.66 (m, 1H, J5,6 = 6.3 Hz, H-5),
3
3.75 (dd, 1H, J3,4 = 9.5 Hz, H-3), 4.03, 4.29 (2m, 2H,
CH2CH@CH2), 4.44, 4.61 (2d, 2H, 2J = 11.3 Hz,
CH2C6H5), 5.08, 5.12 (2m, 2H, CH2CH@CH2), 5.24
(dd, 1H, 3J2,3 = 2.2 Hz, H-2), 5.84 (m, 1H,
3
CH2CH@CH2), 5.89 (d, 1H, J1,2 = 1.9 Hz, H-1),
7.20–7.31 (m, 5H, CH2C6H5); 13C NMR (75.47 MHz,
CDCl3): d 17.95 (C-6), 20.89, 20.93 (2 ꢄ OCOCH3),
67.87 (C-2), 70.02 (C-5), 71.92 (CH2C6H5), 74.21
(CH2CH@CH2), 76.54 (C-3), 79.85 (C-4), 91.13 (C-1),
117.03 (CH2CH@CH2), 127.80, 127.98, 128.40, 129.01,
137.81 (CH2C6H5, one signal is isochronic), 134.79
(CH2CH@CH2), 168.55, 170.03 (2 ꢄ OCOCH3). Anal.
Calcd for C20H26O7 (378.42): C, 63.48; H, 6.93. Found:
C, 63.54; H 6.84.
22
as a colourless foam: ½aꢃD ꢀ33.1 (c 1.0, CHCl3);
1
Rf = 0.65 (solvent A); H NMR (300.13 MHz, CDCl3):
d 1.26 (d, 3H, H-6), 2.06 (s, 3H, OCOCH3), 3.21 (t,
1H, H-4), 3.25 (s, 3H, OCH3), 3.60 (m, 1H,
3
3J5,6 = 6.1 Hz, H-5), 3.75 (dd, 1H, J3,4 = 9.5 Hz, H-3),
4.02, 4.29 (2m, 2H, CH2CH@CH2), 4.52 (d, 1H,
2
3J1,2 = 1.5 Hz, H-1), 4.43, 4.58 (2d, 2H, J = 11.3 Hz,
3.7. 1,2-Di-O-acetyl-4-O-allyl-3-O-benzyl-b-L-rhamno-
pyranose (14b)
CH2C6H5), 5.07, 5.17 (2m, 2H, CH2CH@CH2), 5.24
(dd, 1H, 3J2,3 = 3.4 Hz, H-2), 5.84 (m, 1H,
CH2CH@CH2), 7.19–7.30 (m, 5H, CH2C6H5); 13C
NMR (75.47 MHz, CDCl3): d 17.91 (C-6), 21.07
(OCOCH3), 54.80 (OCH3), 67.49 (C-5), 68.93 (C-2),
71.69 (CH2C6H5), 74.19 (CH2CH@CH2), 77.75 (C-3),
79.82 (C-4), 98.65 (C-1), 116.68 (CH2CH@CH2),
127.62, 127.78, 127.90, 128.32, 128.88, 138.09
(CH2C6H5), 135.01 (CH2CH@CH2), 170.40 (OCOCH3).
21
Colourless foam; ½aꢃD ꢀ7.1 (c 0.1, CHCl3); Rf = 0.65
(solvent A); 1H NMR (300.13 MHz, CDCl3): d 1.36
(d, 3H, H-6), 2.07, 2.12 (2s, 6H, 2 ꢄ OCOCH3), 3.28
3
(t, 1H, H-4), 3.49 (m, 1H, J5,6 = 6.1 Hz, H-5), 3.60
3
(dd, 1H, J3,4 = 9.5 Hz, H-3), 4.09, 4.36 (2m, 2H,
CH2CH@CH2), 4.47, 4.67 (2d, 2H, 2J = 11.3 Hz,
CH2C6H5), 5.15, 5.23 (2m, 2H, CH2CH@CH2), 5.55

1736
N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
(dd, 1H, 3J2,3 = 3.4 Hz, H-2), 5.69 (d, 1H,
3J1,2 = 1.2 Hz, H-1), 5.88 (m, 1H, CH2CH@CH2),
7.25–7.37 (m, 5H, CH2C6H5); 13C NMR (75.47 MHz,
CDCl3): d 17.83 (C-6), 20.82, 20.89 (2 ꢄ OCOCH3),
67.58 (C-2), 71.55 (CH2C6H5), 72.70 (C-5), 74.31
(CH2CH@CH2), 79.06 (C-4), 79.42 (C-3), 91.05 (C-1),
117.05 (CH2CH@CH2), 127.82, 127.99, 128.37, 128.44,
137.42 (CH2C6H5, one signal is isochronic), 134.72
(CH2CH@CH2), 168.80, 170.55 (2 ꢄ OCOCH3). Anal.
Calcd for C20H26O7 (378.42): C, 63.48; H 6.93. Found:
C, 63.51; H, 6.81.
1.9 Hz, H-1), 7.20–7.29 (m, 5H, CH2C6H5); 13C NMR
(75.47 MHz, CDCl3): d 17.75 (C-6), 20.85, 20.92
(2 ꢄ OCOCH3), 66.91 (C-2), 70.44 (C-5), 71.16 (C-4),
71.64 (CH2C6H5), 77.25 (C-3), 91.40 (C-1), 128.18,
128.21, 128.64, 137.28 (CH2C6H5, two signals are iso-
chronic), 170.04 (OCOCH3). Anal. Calcd for C17H22-
O7 (338.35): C, 60.35; H, 6.55. Found: C, 60.43; H 6.61.
3.11. Methyl 2-O-acetyl-4-O-benzoyl-3-O-benzyl-a-L-
rhamnopyranosyl-(1?4)-(methyl 2,3-di-O-benzyl-a-D-
galactopyranosyluronate)-(1?2)-4-O-benzoyl-3-O-
benzyl-a-L-rhamnopyranosyl-(1?4)-(allyl 2,3-di-O-benz-
yl-b-D-galactopyranosid)uronate (21)
3.8. Deallylation of compounds 11 and 14a
To a solution of compound 11 or 14a (0.8 mmol) in a
mixture of acetic acid and water (20:1, v/v, 25 mL)
sodium acetate (656 mg, 8.0 mmol) and palladium(II)chlo-
ride (563 g, 3.2 mmol) were added. After stirring for
4–6 h at 40 °C (monitored by TLC), the reaction mix-
ture was filtered and the filtrate was concentrated. The
residue was dissolved in heptane–chloroform (2:1 v/v,
100 mL), the organic layer was washed with water
(3 ꢄ 50 mL), dried and concentrated. The crude prod-
ucts were purified by column chromatography (eluent
solvent C) to provide 12 (248 mg, 63%) as colourless
foam and 15 (181 mg, 67%) as colourless crystals,
respectively.
Glycosyl acceptor 2020 (384 mg, 0.5 mmol), donor 1915
(458 mg, 0.5 mmol) and molecular sieves (4 A, ca.
˚
7.0 g) were dried under high vacuum for 1 h at ambient
temperature. Then, the solids were suspended in dry
CH2Cl2 (15 mL), and the reaction mixture was stirred
for 2 h at room temperature under an inert atmosphere
in the dark. After cooling to ꢀ70 °C, trimethylsilyl tri-
fluoromethanesulfonate (100 lL, 0.6 mmol) was added,
and the suspension was stirred 3 h at that temperature.
The reaction mixture was allowed to warm-up to room
temperature and stirring was continued for an addi-
tional 18 h. After complete reaction (monitored by
TLC), the mixture was passed through a layer of alka-
line alumina by elution with chloroform. Filtrate and
eluent (ca. 50 mL) were combined, dried and concen-
trated. Purification by HPLC (eluent gradient ethyl ace-
tate 0%?50% in heptane) provided tetrasaccharide 21
3.9. Methyl 2-O-acetyl-3-O-benzyl-a-L-rhamnopyran-
oside (12)
21
22
½aꢃD +11.1 (c 0.9, CHCl3); Rf = 0.46 (solvent A); 1H
(447 mg, 60%) as a colourless foam: ½aꢃD +65.9 (c 1.0,
3
1
NMR (250.13 MHz, CDCl3): d 1.25 (d, 3H, J5,6
=
CHCl3); Rf = 0.39 (solvent A); H NMR (500.13 MHz,
CDCl3): d 1.15 (d, 3H, 3J5
¼ 6:2 Hz, H-6000), 1.27
000;6000
6.1 Hz, H-6), 2.05 (s, 3H, OCOCH3), 2.20 (br d, 1H,
OH), 3.27 (s, 3H, OCH3), 3.41–3.66 (m, 3H, H-3, H-4,
(d, 3H, J5 ;6 ¼ 6:5 Hz, H-60), 2.07 (s, 3H, OCOCH3),
3
0
0
3
3
00
H-5), 4.56 (d, 1H, J1,2 = 1.5 Hz, H-1), 4.32, 4.63 (2d,
3.35 (s, 3H, OCH3 ), 3.52 (dd, 1H, J3,4 = 3.3 Hz, H-
3
3
2H, 2J = 11.3 Hz, CH2C6H5), 5.27 (dd, 1H, J2,3
=
3). 3.76 (dd, 1H, J2,3 = 9.7 Hz, H-2), 3.76 (m, 1H, H-
3.4 Hz, H-2), 7.15–7.32 (m, 5H, CH2C6H5); 13C NMR
(62.89 MHz, CDCl3): d 17.68 (C-6), 20.94 (OCOCH3),
54.89 (OCH3), 67.88 (C-2), 67.94 (C-5), 71.38
(CH2C6H5), 71.55 (C-4), 77.62 (C-3), 98.92 (C-1),
128.00, 128.11, 128.35, 128.43, 128.53, 137.57,
(CH2C6H5), 170.29 (OCOCH3). Anal. Calcd for
C16H22O6 (310.34): C, 61.92; H, 7.15. Found: C, 61.83;
H, 7.07.
5000), 3.80 (m, 1H, H-50), 3.80 (dd, 1H, J2 ;3 ¼ 9:8 Hz,
3
00 00
H-200), 3.80 (s, 3H, OCH3), 3.83 (dd, 1H, 3J3
¼
000;4000
9:9 Hz, H-3000), 3.99 (dd, 1H, J3 ;4 ¼ 9:9 Hz, H-30),
3
0
0
3
00 00
4.03 (d, 1H, H-5), 4.13 (dd, 1H, J3 ;4 ¼ 3:5 Hz,
H-300), 4.16 (m, 1H, CH2CH@CH2), 4.21–4.98 (s and
3
0
0
d, 12H, 6 ꢄ CH2C6H5), 4.30 (dd, 1H, J2 ;3 ¼ 3:1 Hz,
3
H-20), 4.38 (d, 1H, J1,2 = 7.6 Hz, H-1), 4.40 (dd, 1H,
3
3
00 00
J4,5= 0.9 Hz, H-4), 4.42 (dd, 1H, J4 ;5 ¼ 1:6 Hz,
H-400), 4.55 (m, 1H, CH2CH@CH2), 4.92 (d, 1H,
3
H-500), 5.00 (d, 1H, J1 ;2 ¼ 3:5 Hz, H-100), 5.07 (dd,
00 00
3.10. 1,2-Di-O-acetyl-3-O-benzyl-a-L-rhamnopyranose
(15)
1H, 3J1
¼ 1:4 Hz, H-1000), 5.18 (t, 1H, 3J4
¼
000;2000
000;5000
9:9 Hz, H-4000), 5.22, 5.34 (2m, 2H, CH2CH@CH2),
25
3
5.40 (d, 1H, J1 ;2 ¼ 1:4 Hz, H-10), 5.40 (t, 1H,
0
0
Mp 76 °C (from ethyl acetate–heptane); ½aꢃD +2.9 (c 1.1,
1
3
J4 ;5 ¼ 9:9 Hz, H-40), 5.54 (dd, 1H, 3J2
¼ 3:3 Hz,
0
0
000;3000
CHCl3); Rf = 0.52 (solvent A); H NMR (300.13 MHz,
3
CDCl3): d 1.24 (d, 3H, J5,6 = 6.1 Hz, H-6), 2.00, 2.08
H-2000), 5.96 (m, 1H, CH2CH@CH2), 7.00–7.37 (m,
30H, 6 ꢄ CH2C6H5), 7.42, 7.56, 7.97 (3m, 10H,
2 ꢄ OCOC6H5); 13C NMR (125.76 MHz, CDCl3): d
17.59 (C-6000), 17.64 (C-60), 21.08 (OCOCH3), 51.91
(2s, 6H, OCOCH3), 2.37 (br d, 1H, OH), 3.52 (t, 1H,
3
H-4), 3.64 (dd, 1H, J3,4 = 9.5 Hz, H-3), 3.68 (m, 1H,
2
H-5), 4.36, 4.66 (2d, 2H, J = 11.0 Hz, CH2C6H5), 5.26
3
3
000
0
00
(dd, 1H, J2,3 = 3.5 Hz, H-2), 5.94 (d, 1H, J1,2
=
ðOCH3 Þ, 52.43 (OCH3), 67.12 (C-5 ), 67.45 (C-5 ),

N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
1737
68.16 (C-2000), 70.42 (C-500), 70.78 (CH2CH@CH2), 70.92,
71.65, 71.85, 72.88, 72.96, 73.10, (6 ꢄ CH2C6H5), 72.70
(C-4000), 73.17 (C-40), 73.59 (C-5), 74.05 (C-400), 74.22
(C-200), 74.10 (C-3000), 74.61 (C-20), 76.02 (C-30), 76.72
(C-4), 77.20 (C-300), 78.76 (C-2), 79.95 (C-3), 97.60
(C-100), 99.10 (C-1000), 99.70 (C-10), 102.73 (C-1), 117.74
(CH2CH@CH2), 126.83, 127.14, 127.35, 127.38,
127.58, 127.75, 127.87, 127.89, 128.08, 128.11, 128.18,
128.22, 128.29, 128.32, 128.36, 128.40, 128.43, 128.51,
128.53, 128.92, 129.68, 129.75, 129.80, 129.83, 129.97,
133.10, 133.03, 137.50, 137.72, 137.83, 137.91, 138.33,
138.67 (6 ꢄ CH2C6H5, 2 ꢄ COC6H5, 15 signals are
isochronic), 133.74 (CH2CH@CH2), 165.69, 165.83
(2 ꢄ OCOC6H5), 168.16, 168.86, (C-6, C-600), 169.88
(OCOCH3). Anal. Calcd for C87H92O22 (1489.67): C,
70.15; H 6.23. Found: C, 70.18; H, 6.22.
51.92 ðOCH030Þ, 52.44 (OCH3), 66.83 (C-50), 67.43
(C-5000), 68.11 (C-2000), 70.51 (C-500), 70.76 (CH2CH@
CH2), 71.31, 71.65, 71.69, 72.86, 73.09, 73.15
(6 ꢄ CH2C6H5), 72.86 (C-4000), 73.15 (C-40), 73.57
(C-5), 73.94 (C-400), 74.26 (C-200), 76.17 (C-3000), 74.45
(C-20), 76.00 (C-30), 77.20 (C-4), 77.26 (C-300), 78.73
(C-2), 80.02 (C-3), 97.41 (C-100), 99.00 (C-1000), 101.29
(C-10), 102.72 (C-1), 117.74 (CH2CH@CH2), 126.86,
127.13, 127.26, 127.67, 127.70, 127.78, 128.01, 128.10,
128.13, 128.17, 128.25, 128.33, 128.39, 128.42, 128.50,
129.66, 129.75, 129.86, 129.95, 133.03, 133.12, 133.03,
133.09, 137.50, 137.72, 137.83, 137.91, 138.33, 138.67
(6 ꢄ CH2C6H5, 2 ꢄ OCOC6H5 19 signals are iso-
chronic), 133.74 (CH2CH@CH2), 165.73, 165.81
(2 ꢄ OCOC6H5), 168.16, 168.86, (C-6, C-600). Anal.
Calcd for C85H90O21 (1447.63): C, 70.52; H, 6.27.
Found: C, 70.54; H 6.23.
3.12. Methyl 4-O-benzoyl-3-O-benzyl-a-L-rhamnopyran-
osyl-(1?4)-(methyl 2,3-di-O-benzyl-a-D-galactopyran-
osyluronate)-(1?2)-4-O-benzoyl-3-O-benzyl-a-L-
rhamnopyranosyl-(1?4)-(allyl 2,3-di-O-benzyl-b-D-
galactopyranosid)uronate (22)
3.13. Methyl 2-O-acetyl-4-O-benzoyl-a-L-rhamnopyr-
anosyl-(1?4)-(methyl a-D-galactopyranosyluronate)-
(1?2)-4-O-benzoyl-a-L-rhamnopyranosyl-(1?4)-(propyl
b-D-galactopyranosid)uronate (23)
Compound 21 (149 mg, 0.1 mmol) was added under
stirring to a methanolic HCl solution [0.28 M, 10 mL,
prepared by adding of 0.2 mL acetyl chloride to 10 mL
ice-cold dry MeOH], and the mixture was kept under
an argon atmosphere for 48–72 h at room temperature
(monitored by TLC). The reaction solution was then
filtered through a layer of alkaline alumina by elution
with chloroform. The filtrate and eluent (ca. 40 mL)
were combined, dried and concentrated. The crude
product was purified by MPLC (eluent solvent B) to give
22 (140 mg, 97%) as a colourless foam: ½aꢃ2D0 +70.0 (c 1.0,
To a solution of compound 21 (149 mg, 0.1 mmol) in
ethanol (8 mL) 10% palladium-on-charcoal (ca. 20 mg)
was added. The suspension was stirred for 18 h at room
temperature under an atmosphere of hydrogen (moni-
tored by TLC). Then, the mixture was filtered over
Celite, eluted with ethanol and the combined filtrates
were concentrated. The residue was dissolved in water
and lyophilized to yield compound 23 (95 mg, 97%) as
a colourless powder. For NMR investigations the prod-
uct was evaporated several times together with CD3OD:
25
½aꢃD +1.2 (c 1.0, CHCl3); Rf = 0.34 (solvent E); 1H
1
CHCl3); Rf = 0.33 (solvent A); H NMR (500.13 MHz,
NMR (500.13 MHz, CDCl3): d 0.89 (t, 3H, CH2CH2-
CDCl3): d 1.15 (d, 3H, 3J5
¼ 6:1 Hz, H-6000), 1.28
CH3), 1.17 (d, 3H, 3J5
¼ 6:2 Hz, H-6000), 1.22 (d,
000;6000
000;6000
0
3
(d, 3H, H-6 ), 3.38 (s, 3H, OCH3 ), 3.51 (dd, 1H,
3H, J5 ;6 ¼ 6:4 Hz, H-60), 1.65 (dt, 2H, CH2CH2CH3),
00
0
0
3J3,4 = 3.1 Hz, H-3). 3.72 (dd, 1H, J2,3 = 9.5 Hz, H-2),
2.11 (s, 3H, OCOCH3), 3.44 (m, 2H, CH2CH2CH3),
3
3
3
3.77 (dd, 1H, J2 ;3 ¼ 10:1 Hz, H-200), 3.79 (m, 1H,
3.72 (s, 3H, OCH3 ), 3.77 (dd, 1H, J2,3 = 9.7 Hz,
00 00
00
3
H-5000), 3.80 (m, 1H, J5 ;6 ¼ 6:4 Hz, H-50), 3.80 (s, 3H,
H-2), 3.78 (s, 3H, OCH3), 3.86 (dd, 1H, J3,4 = 3.7 Hz,
H-3), 3.88 (2m, 2H, H-50, H-5000), 3.94 (dd, 1H,
3
0
0
OCH3), 3.81 (dd, 1H, 3J3
¼ 9:8 Hz, H-3000), 4.00
000;4000
(dd, 1H, J3 ;4 ¼ 10:1 Hz, H-30), 4.02 (d, 1H, H-5),
J3 ;4 ¼ 10:1 Hz, H-30), 4.09 (dd, 1H, J2 ;3 ¼ 3:1 Hz,
3
3
3
0
0
0
0
0
0
3
3
4.12 (dd, 1H, J3 ;4 ¼ 3:4 Hz, H-300), 4.15 (dd, 1H,
H-20), 4.12 (dd, 1H, J3 ;4 ¼ 3:6 Hz, H-300), 4.17 (dd,
00 00
00 00
3
3J2
¼ 3:4 Hz, H-2000), 4.17 (m, 1H, CH2CH@CH2),
1H, J2 ;3 ¼ 9:8 Hz, H-200), 4.25 (d, 1H, J1,2 = 7.3 Hz,
3
000;3000
00 00
H-1), 4.26 (dd, 1H, 3J3
¼ 9:8 Hz, H-3000), 4.22 (d,
000;4000
3
4.32–5.00 (s and d, 12H, 6 ꢄ CH2C6H5), 4.32 (dd, 1H,
3
J2 ;3 ¼ 3:1 Hz, H-20), 4.38 (d, 1H, J1,2 = 7.6 Hz,
1H, H-5), 4.43 (dd, 1H, J4,5 = 0.9 Hz, H-4), 4.50 (dd,
3
0
0
3
3
1H, J4 ;5 ¼ 0:9 Hz, H-400), 4.93 (d, 1H, H-500), 5.17 (d,
00 00
H-1), 4.40 (dd, 1H, J4,5 = 0.9 Hz, H-4), 4.43 (dd, 1H,
J4 ;5 ¼ 1:6 Hz, H-400), 4.53 (m, 1H, CH2CH@CH2),
1H, J1 ;2 ¼ 3:1 Hz, H-100), 5.43 (dd, 1H, 3J1
¼
3
3
00 00
00 00
000;2000
4.91 (d, 1H, H-500), 5.03 (d, 1H, J1 ;2 ¼ 3:1 Hz, H-100),
1:4 Hz, H-1000), 5.12 (t, 1H, 3J4
¼ 9:8 Hz, H-4000),
3
00 00
000;5000
5.14 (dd, 1H, 3J1
¼ 1:5 Hz, H-1000), 5.24 (t, 1H,
5.26 (d, 1H, J1 ;2 ¼ 1:4 Hz, H-10), 4.92 (t, 1H, J4 ;5
¼
3
3
000;2000
0
0
0
0
3J3
¼ 10:1 Hz, H-4000), 5.22, 5.34 (2m, 2H,
10:1 Hz, H-40), 5.37 (dd, 1H, J2
¼ 3:6 Hz, H-2000),
3
000;4000
000;3000
CH2CH@CH2), 5.39 (d, 1H, J1 ;2 ¼ 1:8 Hz, H-10),
7.39, 7.51, 7.96 (3m, 10H, 2 ꢄ OCOC6H5); 13C NMR
3
0
0
3
3
5.40 (t, 1H, J3 ;4 ¼ J4 ;5 ¼ 10:1 Hz, H-40), 5.97 (m,
1H, CH2CH@CH2), 7.00–7.36 (m, 30H, 6 ꢄ CH2C6H5),
7.46, 7.56, 7.97 (3m, 10H, 2 ꢄ OCOC6H5); 13C NMR
(125.76 MHz, CDCl3): d 17.49 (C-6000), 17.64 (C-60),
(125.76 MHz, CDCl3): d 10.80 (CH2CH2CH3), 18.02
0
0
0
0
(C-600), 18.07 (C-60), 20.94 (OCOCH3), 23.91 (CH2CH2-
000
00
CH3), 52.75 ðOCH3 Þ, 52.89 (OCH3), 68.20 (C-5 ),
68.31 (C-50), 68.43 (C-3000), 69.86 (C-200), 68.94 (C-20),

1738
N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
71.13 (C-300), 71.75 (C-2), 71.76 (C-500), 72.21 (C-30),
72.24 (CH2CH2CH3), 72.95 (C-2000), 73.86 (C-5, C-3,
one signal is isochronic), 74.50 (C-4000), 74.95 (C-40),
78.51 (C-4), 79.26 (C-400), 99.73 (C-100), 100.65 (C-1000),
101.03 (C-10), 104.70 (C-10), 129.60, 129.72, 130.66,
130.69, 131.27, 131.33, 134.40, 134.42 (2 ꢄ OCOC6H5,
four signals are isochronic), 167.50, 167.72
(2 ꢄ COC6H5), 170.51, 171.37, (C-6, C-600), 171.97
(OCOCH3). Anal. Calcd for C45H58O24 (982.94): C,
54.99; H 5.95. Found: C, 54.89; H 5.98.
were dried 1 h at ambient temperature under high vac-
uum. Then, the solids were suspended in dry CH2Cl2
(8 mL), and the reaction mixture was stirred for 2 h at
room temperature under an inert atmosphere in the dark.
After cooling to ꢀ70 °C, trimethylsilyl trifluorometh-
anesulfonate (83 lL, 0.5 mmol) was added, and stirring
was continued for 3 h at that temperature. Subse-
quently, the reaction mixture was allowed to warm-up
to room temperature and stirring was continued for an
additional 18 h. After complete reaction (monitored by
TLC), the mixture was passed through a layer of alka-
line alumina by elution with chloroform. Filtrate and
eluent (ca. 30 mL) were combined, dried and concen-
trated. The crude product was purified by HPLC (eluent
gradient ethyl acetate 0%?50% in heptane) to afford
hexasaccharide 25 (133 mg, 59%) as a colourless foam:
3.14. Propyl a-L-rhamnopyranosyl-(1?4)-a-D-galacto-
pyranosyluronic acid-(1?2)-a-L-rhamnopyranosyl-
(1?4)-b-D-galactopyranosiduronic acid (24)
To a suspension of compound 23 (49 mg, 0.05 mmol) in
MeOH and water (2:1, v/v, 10 mL) lithium hydroxide
(24 mg, 0.1 mmol) was added. After stirring for 15 min
at room temperature (monitored by TLC), the solution
was passed through a column of Dowex-50 (H+) resin.
In the following, MeOH was removed from the filtrate
by evaporation in vacuo, and the residue was repeatedly
dissolved in water and lyophilized to give compound 24
(30 mg, 86%) as a colourless powder. For NMR investi-
gations the product was evaporated several times
20
1
½aꢃD +77.7 (c 1.0, CHCl3); Rf = 0.32 (solvent A); H
NMR (500.13 MHz, CDCl3): d 1.17, 1.20, 1.21 (3d,
9H, H-60, H-600, H-600000), 2.09 (s, 3H, OCOCH3), 3.37,
00
0000
3.39 (2s, 6H, OCH3 , OCH3 ), 3.79 (s, 3H, OCH3),
3
3.53 (dd, 1H, J2,3 = 10.1 Hz, H-2), 3.72 (dd, 1H,
3J3,4 = 3.6 Hz, H-3), 3.76 (m, 1H, H-500000), 3.87 (dd,
3
1H, J3
¼ 9:8 Hz, H-30000), 3.70–5.40 (m, H-20, H-30,
0000 ;40000
H-40, H-50, H-200, H-300, H-400, H-500, H-2000, H-3000, H-4000,
H-5000, H-20000, H-30000, H-40000, H-50000, CH2CH@CH2,
CH2CH@CH2, 9 ꢄ CH2C6H5), 4.04 (d, 1H, H-5), 4.27
25
together with CH3OD: ½aꢃD +2.3 (c 0.5, Me2SO);
1
3
3
Rf = 0.21 (solvent G); H NMR (500.13 MHz, Me2SO-
(dd, 1H, J4,5 = 0.9 Hz, H-4), 4.36 (d, 1H, J1,2 =
d6): d 0.83 (t, 3H, CH2CH2CH3), 1.14, 1.16 (2d, 6H,
H-60, H-6000), 1.53 (dt, 2H, CH2CH2CH3), 3.24 (m, 2H,
7.5 Hz, 1-H), 4.91, 5.02, 5.05, 5.29 (4d, 4H, H-10, H-100,
H-1000, H-10000), 5.21 (t, 1H, J3
¼ 10:1 Hz, H-400000),
3
00000;400000
3
5.40 (d, 1H, 3J1
¼ 1:9 Hz, H-100000), 5.56 (dd, 1H,
00000;200000
CH2CH2CH3), 3.32 (dd, 1H, J2,3 = 9.7 Hz, H-2),
3.40–4.02 (4m, 11H, H-3, H-20, H-30, H-40, H-50, H-200,
H-300, H-2000, H-3000, H-4000, H-5000), 4.26 (d, 1H,
3J1,2 = 7.3 Hz, H-1), 3.92 (d, 1H, H-5), 4.22 (dd, 1H,
3J2
¼ 3:1, H-200000), 5.96 (m, 1H, CH2CH@CH2),
00000;300000
7.05–7.17 (m, 15H, 3 ꢄ CH2C6H5Rha), 7.18–7.39 (m,
30H, 6 ꢄ CH2C6H5GalA), 7.42, 7.56, 8.00 (3m, 15H,
3 ꢄ OCOC6H5); 13C NMR (125.76 MHz, CDCl3): d
17.59, 17.67, 17.69 (C-60, C-6000, C-600000), 22.63 (OCH3),
51.74, 51.76 ðOCH00; OCH0000 Þ, 52.28 (OCH3), 67.18,
3
3
00 00
J4,5 = 0.9 Hz, H-4), 4.32 (dd, 1H, J4 ;5 ¼ 0:9 Hz,
H-400), 4.33 (d, 1H, H-500), 4.94 (d, 1H, J1 ;2 ¼ 3:1 Hz,
3
00 00
H-100), 5.13 (dd, 1H, J1
¼ 1:4 Hz, H-1000), 5.15 (d,
3
000;2000
3
3
1H, J1 ;2 ¼ 1:4 Hz, H-10); 13C NMR (125.76, Me2SO-
d6): d 10.83 (CH2CH2CH3), 18.10, 18.17 (C-60, C-6000),
23.89 (CH2CH2CH3), 70.30, 70.95, 71.49, 71.82, 72.01,
72.14, 73.03, 73.80, 74.52, 75.04, 78.11, 78.74, 79.18
(C-2, C-3, C-4, C-5, C-20, C-30, C-40, C-50, C-200, C-300,
C-400, C-500, CH2CH2CH3), 99.49 (C-100), 99.65 (C-1000),
100.10 (C-10), 103.41 (C-1), 170.38, 171.32 (C-6, C-600).
Anal. Calcd for C27H44O21 (704.63): C, 46.02; H, 6.29.
Found: C: 45.99; H, 6.35.
67.41, 67.52 (C-50, C-5000, C-500000), 68.34 (C-200000), 70.41,
70.57, 70.73, 70.99, 71.43, 71.75, 71.77, 71.91, 72.45,
72.51, 72.88, 73.04, 73.27, 73.31, 73.43, 73.64, 73.83,
74.33, 74.38, 74.47, 74.57, 75.08, 75.22, 75.50, 75.99,
76.19, 76.79, 77.20 (C-4, C-5, C-20, C-30, C-40, C-200,
C-300, C-400, C-500, C-2000, C-3000, C-4000, C-20000, C-30000,
C-40000, C-50000, C-300000, C-400000, 9 ꢄ CH2C6H5, CH2CH@
CH2), 78.79 (C-2), 80.44 (C-3), 97.05, 97.42 (C-100,
C-10000), 98.81, 99.77 (C-10, C-1000), 98.93 (C-100000), 102.84
(C-1), 117.60 (CH2CH@CH2), 126.83, 126.95, 127.08,
127.13, 127.27, 127.35, 127.42, 127.53, 127.68, 127.73,
127.82, 127.96, 128.08, 128.19, 128.28, 128.34, 128.50,
128.61, 128.86, 129.67, 129.78, 130.00, 130.14, 132.94,
132.98, 137.56, 137.66, 137.77, 137.83, 137.89, 137.97,
138.05, 138.42, 138.70, 138.75, 138.89 (9 ꢄ CH2C6H5,
3 ꢄ OCOC6H5),133.84 (CH2CH@CH2), 165.68, 165.70,
165.72 (3 ꢄ OCOC6H5), 168.09 (C-6), 168.81, 168.89
(C-600, C-60000), 169.79 (COCH3). Anal. Calcd for
3
0
0
3.15. Methyl 2-O-acetyl-4-O-benzoyl-3-O-benzyl-a-L-
rhamnopyranosyl-(1?4)-(methyl 2,3-di-O-benzyl-a-D-
galactopyranosyluronate)-(1?2)-4-O-benzoyl-3-O-benz-
yl-a-L-rhamnopyranosyl-(1?4)-(methyl 2,3-di-O-benz-
yl-a-D-galactopyranosyluronate)-(1?2)-4-O-benzoyl-3-
O-benzyl-a-L-rhamnopyranosyl-(1?4)-(allyl 2,3-di-O-
benzyl-b-D-galactopyranosid)uronate (25)
Glycosyl acceptor 22 (145 mg, 0.1 mmol), donor 1915
C
129H134O35 (2244.46): C, 69.03; H, 6.02. Found: C,
˚
(92 mg, 0.1 mmol) and molecular sieves (4 A, 800 mg)
69.10; H, 6.00.

N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
1739
3.16. Glycosylation of the rhamnose derivatives 12 and 15
with 2,3,4,6-tetra-O-benzoyl-a-D-galactopyranosyl
bromide (26)
3.18. 2,3,4,6-Tetra-O-benzoyl-b-D-galactopyranosyl-
(1?4)-1,2-di-O-acetyl-3-O-benzyl-a-L-rhamnopyranose
(29)
21
Rhamnose acceptor 12 or 15 (0.5 mmol), bromide
½aꢃD +45.2 (c 1.0, CHCl3); Rf = 0.44 (solvent A); 1H
3
2623–25 (330 mg, 0.5 mmol) and molecular sieves (4 A,
NMR (300.13 MHz, CDCl3): d 1.41 (d, 3H, J5,6
=
˚
4.0 g) were dried for 1 h at ambient temperature under
high vacuum in the dark. Then, the solids were sus-
pended in dry CH2Cl2 (10 mL), 2,6-di-tert-butyl-4-
methyl-pyridine (123 mg, 0.6 mmol) was added and the
suspension was stirred 30 min at room temperature
under an argon atmosphere in the dark. After chilling
to ꢀ65 °C, silver trifluoromethansulfonate (141 mg,
0.55 mmol) was added, and the reaction mixture was
stirred for 2 h at that temperature. Subsequently, the
mixture was allowed to attain room temperature and
stirring was continued for further 18 h. At completion
(monitored by TLC), the reaction mixture was passed
through a layer of alkaline alumina by elution with chlo-
roform. The combined filtrate and eluent (ca. 50 mL)
were extracted with cold aq 3% hydrochloric acid
(2 ꢄ 20 mL), ice water (20 mL), cold aq NaHCO3
(2 ꢄ 20 mL), ice water (2 ꢄ 20 mL), dried and concen-
trated. Finally, the residue was purified by HPLC
(eluent solvent B) to give the desired disaccharides 27
(293 mg, 66%) and 29 (312 mg, 68%), respectively, both
as colourless foam.
5.8 Hz, H-6), 1.96, 2.05 (2s, 6H, 2 ꢄ OCOCH3), 3.65
3
(dd, 1H, J3,4 = 9.5 Hz, H-3), 3.76 (m, 2H, H-4, H-5),
2
4.16, 4.36 (2d, 2H, J = 11 Hz, CH2C6H5), 4.25 (t, 1H,
3
J5 ;6 a ¼ 6:7 Hz, J5 ;6 b 6:6 Hz, H-50), 4.35 (d, 1H,
0
0
0
0
2
J6 a;6 b ¼ 11 Hz, H-60a), 4.60 (dd, 1H, H-60b), 5.09 (dd,
0
0
3
3
0
0
1H, J2,3 = 3.6 Hz, H-2), 5.24 (d, 1H, J1 ;2 ¼ 7:9 Hz,
H-10), 5.53 (dd, 1H, J3 ;4 ¼ 3:4 Hz, H-30), 5.73 (dd,
3
0
0
3
1H, J2 ;3 ¼ 10:4 Hz, H-20), 5.88 (d, 1H, J1,2 = 1.8 Hz,
H-1), 5.91 (d, 1H, H-40), 7.09–8.04 (m, 25H,
4 ꢄ OCOC6H5, CH2C6H5); 13C NMR (75.47 MHz,
CDCl3): d 18.25 (C-6), 21.03, 21.16 (2 ꢄ OCOCH3),
62.04 (C-60), 67.62 (C-2), 68.46 (C-40), 69.50 (C-5),
70.55 (C-20), 71.12 (C-50), 71.97 (C-30), 71.97
(CH2C6H5), 76.72 (C-4), 77.72 (C-3), 91.09 (C-1),
101.27 (C-10), 127.62, 128.11, 128.50, 128.64, 128.70,
128.89, 129.33, 129.58, 129.90, 129.93, 129.96, 130.14,
133.53, 133.82 (4 ꢄ OCOC6H5, CH2C6H5, 16 signals
are isochronic), 168.76, 170.14 (2 ꢄ OCOCH3). Anal.
Calcd for C51H48O16 (916.92): C, 66.80; H 5.28. Found:
C, 66.73; H 5.20.
3
0
0
3.19. Methyl 2,3,4,6-tetra-O-benzoyl-b-D-galactopyran-
osyl-(1?4)-3-O-benzyl-a-L-rhamnopyranoside (28)
3.17. Methyl 2,3,4,6-tetra-O-benzoyl-b-D-galactopyran-
osyl-(1?4)-2-O-acetyl-3-O-benzyl-a-L-rhamnopyrano-
side (27)
Compound 27 (149 mg, 0.1 mmol) was dissolved in
methanolic HCl (0.28 M, 10 mL, prepared by adding
0.2 mL acetyl chloride to 10 mL ice-cold dry MeOH),
and the solution was stirred for 12 h at room tempera-
ture under an argon atmosphere (monitored by TLC).
The solution was neutralized by filtration through a
layer of alkaline alumina using chloroform as an eluent.
Filtrate and eluent (ca. 40 mL) were combined, concen-
trated and the crude product was purified by HPLC
(eluent solvent C) to yield 28 (98 mg, 58%) as a colour-
23
1
½aꢃD +12.5 (c 1.0, CHCl3); Rf = 0.42 (solvent A); H
NMR (300.13 MHz, CDCl3):
d
1.46 (d, 3H,
3J5,6 = 5.7 Hz, H-6), 2.05 (s, 3H, OCOCH3), 3.34 (s,
3H, OCH3), 3.77 (m, 3H, H-3, H-4, H-5), 4.21, 4.41
(2d, 2H, 2J = 11.4 Hz, CH2C6H5), 4.34 (t, 1H,
J5 ;6 a ¼ J5 ;6 b ¼ 6:6 Hz, H-50), 4.44, 4.71 (2dd, 2H,
3
3
0
0
0
0
3
J6 a;6 b ¼ 11:2 Hz, H-60), 4.60 (d, 1H, J1,2 = 1.6 Hz,
2
0
0
3
24
H-1), 5.20 (dd, 1H, J2,3 = 3.3 Hz, H-2), 5.35 (d,
less foam: ½aꢃD +29.1 (c 1.0, CHCl3); Rf = 0.42 (solvent
3
A); 1H NMR (300.13 MHz, CDCl3): d 1.40 (s, 3H,
3J5,6 = 6.0 Hz, H-6), 1.85 (br d, 1H, OH), 3.30 (s, 3H,
0
0
1H,
J1 ;2 ¼ 8:2 Hz,
H-10),
5.63
(dd,
1H,
J3 ;4 ¼ 3:6 Hz, H-30), 5.82 (dd, 1H, J2 ;3 ¼ 10:4 Hz,
H-20), 6.05 (d, 1H, H-40), 7.21–8.12 (m, 25H,
4 ꢄ OCOC6H5, CH2C6H5); 13C NMR (75.47 MHz,
3
3
0
0
0
0
3
OCH3), 3.61 (dd, 1H, J3,4 = 9.1 Hz, H-3), 3.72 (m,
1H, H-5), 3.76–3.82 (m, 2H, H-2, H-4), 4.27 (m, 1H,
2
J5 ;6 ¼ 6:6 Hz, H-50), 4.29, 4.34 (2d, 2H, J = 11.7 Hz,
CDCl3):
d
17.98 (C-6), 20.92 (OCOCH3), 54.84
3
0
0
(OCH3), 61.79 (C-60), 66.68 (C-5), 68.25 (C-40), 68.32
(C-2), 70.23 (C-20), 70.76 (C-50), 71.51 (CH2C6H5),
71.67 (C-30), 76.92 (C-4), 77.77 (C-3), 98.40 (C-1),
101.00 (C-10), 127.32, 127.70, 128.23, 128.29, 128.32,
128.40, 128.61, 129.12, 129.62, 129.67, 129.70, 129.88,
133.17, 133.22, 133.22, 133.52 (4 ꢄ OCOC6H5,
CH2C6H5, 14 signals are isochronic) 165.39, 165.47,
165.53 (4 ꢄ OCOC6H5), 170,23 (OCOCH3). Anal. Calcd
for C50H48O15 (888.91): C, 67.56; H 5.44. Found: C,
67.52; H 5.37.
3
0
0
CH2C6H5), 4.39, 4.60 (2m, 2H, J6 a;6 b ¼ 11:0 Hz,
3
H-60a, H-60b), 4.63 (d, 1H, J1,2 = 1.9 Hz, H-1), 5.27
(d, 1H, J1 ;2 ¼ 8:2 Hz, H-10), 5.61 (dd, 1H, J3 ;4
¼
3
3
0
0
0
0
3:5 Hz, H-30), 5.77 (dd, 1H, J2 ;3 ¼ 10:4 Hz, H-20),
5.96 (dd, 1H, H-40), 7.11–8.09 (m, 25H, 4 ꢄ OCOC6H5,
CH2C6H5); 13C NMR (75.47 MHz, CDCl3): d 17.94
(C-6), 54.81 (OCH3), 61.88 (C-60), 66.40 (C-5), 67.90
(C-4), 68.25 (C-40), 70.28 (C-20), 70.99 (C-50), 71.72
(C-30), 71.84 (CH2C6H5), 77.15 (C-2), 80.13 (C-3),
99.89 (C-1), 101.29 (C-10), 127.18, 127.96, 128.00,
3
0
0

1740
N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
128.26, 128.34, 128.40, 128.47, 128.64, 129.14, 129.22,
129.35, 129.66, 129.72, 129.75, 129.93, 129.96, 133.20,
133.26, 133.54, 133.77 (CH2C6H5, 4 ꢄ OCOC6H5, 10
signals are isochronic), 165.32, 165.50, 165.58, 166.03
(4 ꢄ OCOC6H5). Anal. Calcd for C48H46O14 (846.87):
C, 68.08.; H 5.47. Found: C, 68.14; H 5.52.
1H NMR (500.13 MHz, CDCl3): d 1.35 (d, 3H,
3
3
00 00
J5,6 = 6.0 Hz, H-6), 1.45 (d, 3H, J5 ;6 ¼ 6:0 Hz,
H-600), 2.05 (s, 3H, OCOCH3), 3.22 (s, 3H, OCH3),
3.30 (s, 3H, OCH03), 3.35–3.90 (m, 7H, H-20, H-30, H-3,
H-4, H-5, H-300, H-500), 3.91–5.00 (m, 13H, 4 ꢄ
CH2C6H5, H-40, H-50, H-5000, H-6000a, H-6000b), 4.65 (d,
3
3
0
0
1H, J1,2 = 2.5 Hz, H-1), 4.93 (d, 1H, J1 ;2 ¼ 3:5 Hz,
H-10), 5.00–5.80 (m, 3H, H-2, H-200, H-400), 5.07 (d,
3.20. 2,3,4,6-Tetra-O-benzoyl-b-D-galactopyranosyl-
(1?4)-2-O-acetyl-3-O-benzyl-a-L-rhamnopyranosyl
bromide (30)
3
1H, J1 ;2 ¼ 1:8 Hz, H-100), 5.38 (d, 1H, 3J1
¼
00 00
000;2000
7:9 Hz, H-1000), 5.55 (dd, 1H, 3J3
¼ 2:8 Hz, H-3000),
000;4000
3
000;3000
5.77 (dd, 1H, J2
¼ 10:4 Hz, H-2000), 5.99 (dd, 1H,
H-4000), 7.00–8.10 (m, 45H, 4 ꢄ CH2C6H5, 5 ꢄ OCOC6-
H5); 13C NMR (125.76 MHz, CDCl3): d 17.40 (C-6),
17.58 (C-600), 21.03 (OCOCH3), 51.99 (OCH3), 54.79
(OCH03), 61.70 (C-6000), 72.89, 73.02, 73.10, 73.56
(4 ꢄ CH2C6H5), 67.10, 67.36, 68.39, 68.81, 70.24,
71.43, 71.99, 74.37, 74.47, 74.65, 75.54, 76.39, 78.06,
78.15, 79.24, 79.35 (C-2, C-3, C-4, C-5, C-2000, C-3000,
C-4000, C-5000, C-20, C-30, C-40, C-50, C-200, C-300, C-400,
C-500), 98.09 (C-1), 98.36 (C-10), 99.72 (C-100), 101.22
(C-1000), 125.56, 126.38, 127.04, 127.17, 127.37, 127.47,
127.63, 127.83, 127.93, 128.09, 128.13, 128.31, 128.44,
128.63, 128.69, 128.82, 128.88, 129.00, 129.15, 129.21,
129.27, 129.37, 129.47, 129.70, 129.80, 132.89, 133.03,
133.19, 133.41, 133.51, 137.71, 137.92, 138.01, 138.32
(4 ꢄ CH2C6H5, 5 ꢄ OCOC6H5, 20 signals are isochro-
nic), 165.34, 165.48, 165.51, 165.64, 166.14 (5 ꢄ
OCOC6H5), 168.45 (C-60), 170.14 (OCOCH3). Anal.
Calcd for C91H90O26 (1599.67): C, 68.32; H 5.67. Found:
C, 68.14; H 5.53.
Trimethylsilyl bromide (31 lL, 0.24 mmol) was added to
a stirred solution of 29 (147 mg, 0.16 mmol) in dry
CH2Cl2 (3 mL) at ꢀ10 °C under an argon atmosphere.
After stirring for 1 h at that temperature, the mixture
was allowed to attain room temperature and stirring
was continued for further 24 h (TLC solvent A). Then,
the reaction mixture was concentrated and dried to yield
30 (135 mg, 90%) as a colourless foam: Rf = 0.48
(solvent A); 1H NMR (250.13 MHz, CDCl3): d 1.45
3
(d, 3H, J5,6 = 6.1 Hz, H-6), 2.03 (s, 3H, OCOCH3),
3.77–4.06 (m, 2H, H-4, H-5), 4.11 (dd, 1H,
3J3,4 = 3.4 Hz, H-3), 4.18–4.45 (m, 5H, H-50, H-60a,
3
H-60b, CH2C6H5), 5.28 (dd, 1H, J2,3 = 9.5 Hz, H-2),
5.32 (d, 1H, J1 ;2 ¼ 7:6 Hz, H-10), 5.59 (dd, 1H,
3
0
0
3
3
J3 ;4 ¼ 3:4 Hz, H-30), 5.81 (dd, 1H, J2 ;3 ¼ 10:7 Hz,
0
0
0
3
0
H-20), 5.97 (d, 1H, H-40), 6.03 (d, 1H, J1,2 = 1.2 Hz,
H-1), 7.14–8.12 (m, 25H, 4 ꢄ OCOC6H5, CH2-
C6H5).
3.21. Methyl 2-O-acetyl-4-O-benzoyl-3-O-benzyl-a-L-
rhamnopyranosyl-(1?4)-(methyl 2,3-di-O-benzyl-a-D-
galactopyranosyluronate)-(1?2)-[2,3,4,6-tetra-O-
benzoyl-b-D-galactopyranosyl)-(1?4)]-3-O-benzyl-a-L-
rhamnopyranoside (31)
3.22. Glycosylation of acceptors 16 and 32 with donor 30
Acceptor 16 or 32 (0.3 mmol), bromide 30 (281 mg,
˚
0.3 mmol) and molecular sieves (4 A, 2.4 g) were dried
for 1 h at ambient temperature under high vacuum in
the dark. The solids were suspended in dry CH2Cl2
(8 mL), 2,6-di-tert-butyl-4-methyl-pyridine (74 mg,
0.36 mmol) was added, and the suspension was stirred
for 30 min at room temperature under an argon atmo-
sphere in the dark. After chilling to ꢀ65 °C, silver tri-
fluoromethansulfonate (85 mg, 0.33 mmol) was added,
and the reaction mixture was stirred 2 h at that temper-
ature. At this point, the mixture was allowed to attain
room temperature and stirring was continued for further
18 h. At completion (monitored by TLC), the reaction
mixture was passed through a layer of alkaline alumina
by elution with chloroform. The filtrate and eluent (ca.
50 mL) were extracted with cold aq 3% hydrochloric
acid (2 ꢄ 20 mL), ice water (20 mL), cold aq NaHCO3
(2 ꢄ 20 mL), ice water (2 ꢄ 20 mL), dried and concen-
trated. Finally, the residue was purified by HPLC (elu-
ent solvent B) to give the desired trisaccharides 33
(262 mg, 68%) and 34 (302 mg, 74%), respectively, both
as colourless foam.
Glycosyl acceptor 28 (95 mg, 0.11 mmol), donor 1915
(103 mg, 0.11 mmol) and molecular sieves (4 A,
˚
900 mg) were dried 1 h at ambient temperature under
high vacuum. Then, the solids were suspended in dry
CH2Cl2 (8 mL), and the reaction mixture was stirred
for 2 h at room temperature under an inert atmosphere
in the dark. After cooling to ꢀ70 °C, trimethylsilyl tri-
fluoromethanesulfonate (36 lL, 0.2 mmol) was added
and stirring was continued for 3 h at that temperature.
At that point, the reaction mixture was allowed to
warm-up to room temperature and stirring was contin-
ued for an additional 18 h. At completion (monitored
by TLC), the reaction mixture was passed through a
layer of alkaline alumina by elution with chloroform.
Filtrate and eluent (ca. 30 mL) were combined, dried
and concentrated. The crude product was purified by
HPLC (eluent gradient EtOAc 0%?50% in heptane)
to give tetrasaccharide 31 (109 mg, 62%) as a colourless
22
foam: ½aꢃD +61.9 (c 0.53, CHCl3); Rf = 0.45 (solvent A);

N. Nemati et al. / Carbohydrate Research 343 (2008) 1730–1742
1741
3
00 00
3.23. Methyl 2,3,4,6-tetra-O-benzoyl-b-D-galactopyrano-
syl-(1?4)-2-O-acetyl-3-O-benzyl-a-L-rhamnopyranosyl-
(1?4)-(allyl 2,3-di-O-benzyl-b-D-galactopyranosid)uro-
nate (33)
0:9 Hz, H-10), 5.56 (dd, 1H, J3 ;4 ¼ 3:4 Hz, H-300),
5.80 (dd, 1H, J2 ;3 ¼ 10:4 Hz, H-200), 5.85–6.00 (m,
3
00 00
1H, CH2CH@CH2), 5.96 (d, 1H, H-400), 7.15–8.08 (m,
40 H, 4 ꢄ CH2C6H5, 4 ꢄ OCOC6H5); 13C NMR
(125.76 MHz, CDCl3): d 18.08 (C-60), 20.91 (OCOCH3),
61.94 (C-600), 67.35, 70.69, 71.52, 73.09 (4 ꢄ CH2C6H5),
67.48 (C-50), 68.20 (C-20), 68.45 (C-400), 70.11 (C-200),
70.92 (C-500), 71.83 (C-300), 72.00 (C-4), 73.61 (C-5),
75.24 (CH2CH@CH2), 76.74 (C-40), 77.43 (C-30), 78.15
(C-2), 81.27 (C-3), 98.19 (C-10), 101.13 (C-100), 102.73
(C-1), 117.53 (CH2CH@CH2), 127.51, 127.57, 127.61,
127.64, 127.73, 128.23, 128.25, 128.31, 128.33, 128.36,
128.43, 128.58, 128.63, 128.84, 129.27, 129.36, 129.52,
129.69, 129.71, 129.76, 129.92, 133.03, 133.19, 133.25,
133.46, 135.36, 137.84, 138.08, 138.35 (4 ꢄ CH2C6H5,
4 ꢄ OCOC6H5, 19 signals are isochronic), 133.87
24
1
½aꢃD +43.6 (c 1.0, CHCl3); Rf = 0.46 (solvent A); H
3
0
0
NMR (500.13 MHz; CDCl3): d 1.40 (d, 3H, J5 ;6
¼
6:3 Hz, H-60), 1.92 (s, 3H, OCOCH3), 3.52 (dd, 1H,
J3 ;4 ¼ 9:8 Hz, H-30), 3.55 (m, 1H, H-50), 3.68–3.74
(m, 3H, H-40, H-2, H-3), 3.76 (s, 3H, OCH3), 4.03 (s,
1H, H-5), 4.08–4.49 (m, 7H, 2 ꢄ CH2C6H5,
3
0
0
3
CH2CH@CH2, H-4), 4.34 (d, 1H, J1,2 = 7.6 Hz, H-1),
4.41 (dd, 1H, J5 ;6 ¼ 6:3 Hz, H-600a), 4.44 (s, 1H,
3
00 00
H-500), 4.63 (dd, 1H, J6 a;6 b ¼ 11:0 Hz, H-600b), 4.70 (s,
2H, CH2C6H5), 4.71, 4.86 (2d, 2H, 2J = 10.7 Hz,
CH2C6H5), 5.18, 5.30 (2m, 2H, CH2CH@CH2), 5.28
2
00
00
(d, 1H, J1 ;2 ¼ 7:9 Hz, H-100), 5.32 (d, 1H, J1 ;2
¼
(CH2CH@CH2),
165.41,
165.56,
166.03
(4 ꢄ
3
3
00 00
0
0
1:9 Hz, H-10), 5.39 (dd, 1H, J2 ;3 ¼ 2:8 Hz, H-20), 5.52
OCOC6H5), 167.02 (C-6), 169.65 (OCOCH3). Anal.
Calcd for C79H76O21 (1361.44): C, 69.69; H 5.63. Found:
C, 69.63; H 5.69.
3
0
0
(dd, 1H, J3 ;4 ¼ 3:5 Hz, H-300), 5.79 (dd, 1H, J2 ;3
¼
3
3
00 00
00 00
10:4 Hz, H-200), 5.94 (dd, 1H, H-400), 5.91 (m, 1H,
CH2CH@CH2), 7.15–8.10 (m, 35H, 3 ꢄ CH2C6H5,
4 ꢄ OCOC6H5); 13C NMR (125.76 MHz, CDCl3): d
17.89 (C-60), 20.90 (OCOCH3), 52.78 (OCH3), 61.90
(C-600), 67.36 (C-50), 68.13 (C-20), 68.34 (C-4), 70.20
(C-200), 70.76, 73.06, 75.22 (3 ꢄ CH2C6H5), 70.87
(C-400), 71.56 (CH2CH@CH2), 71.85 (C-500), 71.90
(C-300), 73.87 (C-5), 76.33 (C-40), 77.50 (C-3), 78.15
(C-2), 81.11 (C-30), 98.21 (C-10), 100.89 (C-100), 102.71
(C-1), 117.62 (CH2CH@CH2), 132.50 (CH2CH@CH2),
127.37, 127.42, 127.48, 127.61, 127.64, 128.21, 128.23,
128.26, 128.33, 128.39, 128.41, 128.44, 128.50, 128.65,
128.80, 128.84, 129.26, 129.30, 129.49, 129.67, 129.71,
129.76, 129.92, 130.83, 133.17, 133.20, 133.48, 133.79,
137.80, 138.07, 138.35 (3 ꢄ CH2C6H5, 4 ꢄ OCOC6H5,
11 signals are isochronic), 165.35, 165.53, 166.02,
167.71, (4 ꢄ OCOC6H5), 168.28 (C-6), 169.64
(OCOCH3). Anal. Calcd for C73H72O21 (1285.34): C,
68.21; H 5.65. Found: C, 68.30; H 5.72.
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21
1
½aꢃD +54.3 (c 1.0, CHCl3); Rf = 0.48 (solvent A); H
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3
00 00
0
0

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