Glycosylation of a newly functionalized orthoester derivative

Article abstract of DOI:10.3390/molecules19022602

Tandem glycosylation of the 6-O-Fmoc-substituted benzyl orthoester derivative 2a was carried out in moderate yields by electrogenerated acid (EGA). The Fmoc group was effectively removed under mild basic conditions, and the product was submitted to the subsequent glycosylation.

Full text of DOI:10.3390/molecules19022602

Molecules 2014, 19, 2602-2611; doi:10.3390/molecules19022602
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molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Article
Glycosylation of a Newly Functionalized Orthoester Derivative
Kohei Kawa, Tsuyoshi Saitoh, Eisuke Kaji and Shigeru Nishiyama *
Department of Chemistry, Faculty of Science and Technology, Keio University, Hiyoshi 3-14-1,
Kohoku-ku, Yokohama 223-8522, Japan
* Author to whom correspondence should be addressed; E-Mail: nisiyama@chem.keio.ac.jp;
Tel./Fax: +81-45-566-1717.
Received: 26 January 2014; in revised form: 20 February 2014 / Accepted: 21 February 2014 /
Published: 24 February 2014
Abstract: Tandem glycosylation of the 6-O-Fmoc-substituted benzyl orthoester derivative
2a was carried out in moderate yields by electrogenerated acid (EGA). The Fmoc group
was effectively removed under mild basic conditions, and the product was submitted to the
subsequent glycosylation.
Keywords: glycosylation; orthoester; electrogenerated acid (EGA); Fmoc group;
selective deprotection
1. Introduction
With the discovery of the numerous biologically important roles of sugar chains, a number of
glycosylation protocols have been reported [1,2]. Generally, the glycosylation process consists of
efficient generation of an oxocarbenium ion species as a glycosyl donor and modulation of its selective
nucleophilic attack on aglycons. Enhancement of β-selectivity is supported by the participation of
acyl-protected hydroxyl groups. In particular, construction of a cyclic carboxonium ion between the C1
and C2 positions is an effective method to obtain the corresponding β-glycosidic linkage of D-sugars,
although orthoesters are produced as side products when sterically hindered functional groups are used
in glycosyl donors and acceptors [3–6]. Orthoesters were known to be converted to the corresponding
glycosides under acidic conditions, and relatively stable derivatives were employed as glycoside
precursors by Kochetkov [7–11] and Kunz [12,13]. We reported the synthesis and reaction of the
orthoester 1 (Figure 1), which could be purified by silica gel column chromatography and kept for one
month at room temperature [14]. Upon glycosylation with appropriate alcohols in the presence of the

Molecules 2014, 19
2603
electrogenerated acid (EGA) [15–20], which is considered to be anhydrous HClO4 produced by anodic
oxidation of cyclohexanol and Bu4NClO4, the corresponding glycosides were produced in high
(primary OH) to moderate (secondary OH) yields, even on tertiary OH groups [14]. Upon comparison
with Lewis acids and Brønsted acids, EGA provided better results in glycosylation reactions. To
synthesize sugar chains by repeated glycosylation, devices providing effective activation of the
anomeric center and the subsequent selective deprotection of an appropriate hydroxyl group in sugar
units are required. The orthoester 1 was modified to examine its applicability to practical tandem
glycosylation. Here, we describe a synthesis of the orthoester derivative 2a carrying an Fmoc group at
the C-6 position, and its glycosylation reaction.
Figure 1. The benzyl orthoesters 1–4.
OMe
BzO
BzO
R
OMe
O
O
O
O
BzO
BzO
1: R= OBz
2a: R= OFmoc
3: R= OH
BzO
O
O
OH
OTBS
O
O
4
2. Results and Discussion
To examine the chemical properties of the orthoester 1 for the modulation of protecting groups, the
acyl protecting groups were removed under basic conditions to give an unstable triol. Although
removal of the TBS group in the benzylic region provided the stable phenol derivative 4 [14], the triol
was labile under standard work-up conditions, and only detected by ESI mass spectrum. These
observations prompted us to synthesize the orthoester derivative 2a, in which a protecting group at the
C-6 position can be selectively removed. After assessment of various protecting groups, we adopted
the Fmoc group, which is removed under mild basic conditions.
2.1. Synthesis of the Orthoester 2a
Synthesis of the C-6 modified orthoester 2a commenced with conversion of 6-O-trityl-1,2,3,4-tetra-
O-benzoyl-β-D-glucopyranose, which was synthesized by the standard procedure from D-glucose, into
6-O-Fmoc-2,3,4-tri-O-benzoyl-α-D-glucopyranosyl bromide (5) using a two-step manipulation,
followed by the anomeric bromination (Scheme 1). Glycosylation of 5 with the benzyl alcohol 6 under
Königs–Knorr reaction conditions provided the expected orthoester 2a (50%) [21], along with the
corresponding benzyl glycoside 2b (30%). Stability of 2a under standard work-up and chromatographic
conditions was similar to the previously reported derivative 1. The Fmoc group in 2a was selectively
removed under basic conditions to give 6-OH orthoester 3, which will be used as a glycosyl acceptor
under neutral glycosylation conditions.

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Scheme 1. Synthesis of the orthoester derivative 2a.
FmocO
HO
TrO
BzO
a
O
b, c, d
BzO
O
O
OH
HO
HO
BzO
OH
OBz
BzO
BzO
Br
OBz
5
D
-glucose
OMe
RO
BzO
O
O
e
BzO
OMe
O
OTBS
2b
HO
O
2a: R= Fmoc
3: R= H
OTBS
6
f
Reagents and yields: (a) TrCl, Et3N, pyr.; BzCl. (b) H2Pd/C. (c) FmocCl, pyr.
(d) HBr-AcOH, Ac2O, AcOH (43% in four steps. (e) 6, Hg(CN)2, 4Å MS (2a:
50%, 2b: 30%). (f) pyr. (81%).
2.2. Glycosylation of the Orthoester 2a
When the orthoester 2a was submitted to reaction with cyclohexanol in the presence of EGA [14]
and 4Å MS in 1,2-DCM at 40 °C, the expected β-glycoside 7 was obtained in 62% yield (Scheme 2).
Next selective removal of the Fmoc group in 7 under basic conditions was carried out to give 8 in 95%
yield, without any acyl migration from C-4 to C-6 position or removal of the acyl group. Repeated
glycosylation of 8 with 2a under the same conditions as mentioned above, gave the disaccharide 9 in
38% yield. Alternatively, glycosylation of 10 with 2a provided the disaccharide 11 in 53% yield. After
removal of the Fmoc group, further glycosylation with 2a provided the corresponding trisaccharide 12 [22].
Scheme 2. Glycosylation reactions of the orthoester derivative 2a.
FmocO
O
BzO
HO
BzO
OMe
FmocO
a
BzO
b
O
O
O
O
BzO
BzO
OCy
OCy
BzO
BzO
BzO
O
OTBS
8
7
2a
c
HO
BzO
O
SPh
d
BzO
FmocO
BzO
10
O
BzO
BzO
O
BzO
BzO
BzO
O
OCy
9
BzO
FmocO
O
BzO
BzO
e, f
O
BzO
BzO
BzO
O
BzO
SPh
12
11
Reagents and yields: (a) cyclohexanol, EGA, 4Å MS (62%). (b) Et3N, pyr. (95%).
(c) 2a, EGA, 4Å MS (38%). (d) 10, EGA, 4Å MS (63%). (e) Et3N, pyr. (82%).
(f) 2a, EGA, 4Å MS (12%).

Molecules 2014, 19
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3. Experimental
General Information
All reactions were carried out under an argon atmosphere unless otherwise noted. When necessary,
solvents were dried prior to use. Dry THF, dry Et2O and dry CH2Cl2 were purchased from Kanto
Chemical Co., Inc. (Tokyo, Japan). Optical rotations were measured on a JASCO P-2200 digital
polarimeter with a sodium (D line) lamp. IR spectra were recorded on a JASCO Model A-202
1
spectrophotometer. H-NMR spectra and 13C-NMR spectra were obtained on JEOL JNM-GX400,
JNM-α400, JNM-AL400 or JNM-ECX400 spectrometers in deuterated solvent using tetramethylsilane
as an internal standard. Deuteriochloroform was used as a solvent, unless otherwise stated. High-
resolution mass spectra were obtained on a Waters LCT Premier XE (ESI) or JEOL JMS-700 (FAB).
Preparative and analytical TLC were carried out on silica gel plate (Kieselgel 60 F254, E. Merck AG.,
Darmstadt, Germany) using UV light, 1 M aq. sulfuric acid, and/or 5% molybdophosphoric acid in
ethanol for detection. Kanto silica 60N (spherical, neutral, 105–210 μm) was used for column
chromatography. All anodic oxidation was conducted using HA-151A (Hokuto Denko, Tokyo, Japan) as
a potentiostat and galvanostat, using glassy carbon plate as anodes, and platinum plate or wire as cathode.
2,3,4-Tri-O-benzoyl-6-O-9-fluorenylmethyloxycarboxyl-α-D-glucopyranosyl bromide (5). To
a
suspension of D-glucose (4.55 g, 0.025 mol) in pyridine (50 mL) were added TrCl (10.6 g, 0.038 mol)
and Et3N (18 mL, 0.13 mol), and the mixture was stirred at room temperature overnight. After the
addition of BzCl (23 mL, 0.20 mol) at the same temperature, stirring was continued overnight. The
reaction was quenched with sat. aq. NaHCO3, and the mixture was extracted with CHCl3. The organic
layer was dried (Na2SO4), and concentrated in vacuo. After removal of the high polarity byproducts by
silica gel short column (hexane/EtOAc = 5:1), a crude was used in the next step without further
purification. The crude product was solved in MeOH/EtOAc (1:1, 100 mL). After the addition of 5%
Pd-C (cat.), the mixture was stirred overnight under H2 atmosphere. The mixture was filtered and the
filtrate was concentrated in vacuo. After silica gel short column chromatography (hexane/EtOAc = 3:1),
a crude was used in the next step without further purification.
A mixture of the crude and FmocCl (9.80 g, 0.038 mol) in pyridine (100 mL) was stirred overnight.
The reaction was quenched by sat. aq. NaHCO3, and the mixture was extracted with CHCl3. The
organic layer was dried (Na2SO4), and concentrated in vacuo. After purification by silica gel short
column chromatography (hexane/EtOAc = 10:1), a crude was used in the next step without
further purification. A mixture of the crude, HBr-AcOH (250 mL), AcOH (60 mL), and Ac2O (60 mL)
was stirred overnight. The mixture was extracted with CHCl3. The organic layer was dried (Na2SO4), and
concentrated in vacuo. The residue was purified by silica gel column chromatography (hexane/EtOAc =
5:1) to give 5 as a colorless oil (8.43 g, 43% in 4 steps): α23 +76.4 (c 1.00, CHCl3); IR (film) 1732,
D
1
1601, 1261 cm−1; H-NMR (400 MHz) δ 4.40 (5H, m, H-6a,6b, Fmoc), 4.67 (1H, m, H-5), 5.36 (1H,
dd, J = 9.8, 4.0 Hz, H-2), 5.78 (1H, t, J = 9.8 Hz, H-4), 6.28 (1H, t, J = 9.8 Hz, H-3), 6.89 (1H, d,
J = 4.0 Hz, H-4), 7.40 (11H, m, Ar), 7.54 (2H, m, Ar), 7.64 (2H, m, Ar), 7.78 (2H, m, Ar), 7.89 (2H,
m, Ar), 8.00 (4H, m, Ar); 13C-NMR (100 MHz) δ 46.5 (Fmoc), 64.9 (C-6), 67.7 (C-4), 70.4 (C-3), 70.5
(C-2), 71.3 (C-5), 72.5 (FmocCH2), 86.7 (C-1), 120.0 (Ar), 125.2 (Ar), 125.3 (Ar), 127.2 (Ar), 127.9

Molecules 2014, 19
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(Ar), 128.3 (Ar), 128.4 (Ar), 128.5 (Ar), 128.7 (Ar), 129.7 (Ar), 129.9 (Ar), 130.0 (Ar), 133.3 (Ar),
133.7 (Ar), 133.85 (Ar), 141.2 (Ar), 141.3 (Ar), 143.2 (Ar), 143.4 (Ar), 154.7 (C=O), 165.1 (C=O),
165.2 (C=O), 165.5 (C=O). ESI-MS: calcd for C42H33O10NaBr 799.1155 (M+Na)+, found m/z 799.1151.
(2R,5R,6R,7S)-5-(((((9H-Fluoren-9-yl)methoxy)carbonyl)oxy)methyl)-2-((2-((tert-butyldimethylsilyl)oxy)-
4-methoxybenzyl)oxy)-2-phenyltetrahydro-3aH-[1,3]dioxolo[4,5-b]pyran-6,7-diyl dibenzoate (2a) and
2-tert-Buthyldimethylsyloxy-4-methoxyphenylmethyl2,3,4-tetra-O-benzoyl-6-O-9-fluorenylmethyloxycarboxyl-
β-D-glucopyranoside (2b). To a solution of 5 (71 mg, 0.092 mmol) in anhydrous toluene (0.9 mL)
were added 6 (74 mg, 0.27 mmol), Hg(CN)2 (46 mg, 0.18 mmol), and MS 4A at 100 °C, and the
mixture was stirred overnight. The mixture was filtered through a Celite pad, and the filtrate was
washed with sat. aq. NaHCO3. The organic extracts were dried (Na2SO4), and concentrated in vacuo.
The residue was purified by silica gel column chromatography (hexane/acetone = 7:1) to give 2a
(45 mg, 50%) and 2b (26 mg, 30%) as oil: 2a: α23D +1.2 (c 1.00, CHCl3); IR (film) 1728, 1613, 1257
cm−1; 1H-NMR (400 MHz) δ 0.15 (6H, s, SiCH3), 0.93 (9H, s, tBu), 3.74 (3H, s, OCH3), 4.16 (2H, m,
Fmoc), 4.29 (6H, m, H-5,6a,6b, ArCH2, Fmoc), 4.78 (1H, m, H-2), 5.37 (1H, bd, H-4), 5.76 (1H, br,
H-3), 6.04 (1H, bd, H-1), 6.31 (1H, d, J = 2.2 Hz, Ar), 6.48 (1H, dd, J = 8.3, 2.2 Hz, Ar), 7.18 (1H, d,
J = 8.3 Hz, Ar), 7.41 (11H, m, Ar), 7.57 (4H, m, Ar), 7.75 (2H, m, Ar), 7.82 (2H, m, Ar), 7.97 (2H, m,
Ar), 8.11 (2H, m, Ar); 13C-NMR (100 MHz) δ −4.3 (SiCH3), 18.1 (tBu), 25.6 (tBu), 46.6 (Fmoc), 55.2
(OCH3), 61.3 (ArCH2), 67.3 (C-6), 68.3 (C-2), 69.2 (C-3), 70.1 (C-4), 72.3 (FmocCH2), 77.2 (C-5),
97.4 (C-1), 105.3 (Ar), 105.9 (Ar), 119.9 (Ar), 120.5 (Ar), 121.4 (Ar), 125.3 (orthoester-C), 126.5
(Ar), 127.1 (Ar), 127.2 (Ar), 127.8 (Ar), 128.3 (Ar), 1283.4 (Ar), 128.5 (Ar), 129.1 (Ar), 129.6 (Ar),
129.9 (Ar), 130.0 (Ar), 133.5 (Ar), 133.6 (Ar), 135.3 (Ar), 141.2 (Ar), 143.2 (Ar), 154.0 (C=O), 154.9
(C=O), 159.9 (C=O), 165.2 (C=O). ESI-MS: calcd for C56H56O13SiNa 987.3388 (M+Na)+, found m/z
1
987.3373. 2b: α23D −14.9 (c 1.00, CHCl3); IR (film) 1735, 1610, 1262 cm−1; H-NMR (400 MHz) δ
0.10 (3H, s, SiCH3), 0.16 (3H, s, SiCH3), 0.94 (9H, s, tBu), 3.72 (3H, s, OCH3), 4.06 (1H, ddd,
J = 9.8, 5.3, 3.2 Hz, H-5), 4.23 (1H, t, J = 7.6 Hz, Fmoc), 4.40 (4H, m, H-6a,6b, Fmoc), 4.76 (1H, d,
J = 12.4 Hz, ArCH2), 4.83 (1H, d, J = 12.4 Hz, ArCH2), 4.86 (1H, d, J = 8.0 Hz, H-1), 5.58 (1H, dd,
J = 9.8, 8.0 Hz, H-2), 5.60 (1H, t, J = 9.8 Hz,H-4), 5.83 (1H, t, J = 9.8 Hz, H-3), 6.27 (1H, dd, J = 8.7,
2.5 Hz, Ar), 6.30 (1H, d, J = 2.5 Hz, Ar), 7.12 (1H, d, J = 8.7 Hz, Ar), 7.35 (14H, m, Ar), 7.50 (2H, m,
13
Ar), 7.62 (2H, m, Ar), 7.76 (2H, m, Ar), 7.83 (4H, m, Ar), 7.92 (1H, m, Ar); C-NMR (100 MHz) δ
−4.4 (SiCH3), −4.3 (SiCH3), 18.1 (tBu), 25.6 (tBu), 29.7 (tBu), 46.6 (Fmoc), 55.2 OCH3), 65.6
(ArCH2), 66.3 (C-6), 69.6 (C-4), 70.2 (C-3), 71.6 (C-2), 72.0 (C-5), 72.9 (FmocCH2), 99.1 (C-1),
105.4 (Ar), 105.9 (Ar), 119.6 (Ar), 119.9 (Ar), 125.2 (Ar), 125.3 (Ar), 127.2 (Ar), 127.8 (Ar), 128.1
(Ar), 128.2 (Ar), 128.4 (Ar), 128.8 (Ar), 129.3 (Ar), 129.7 (Ar), 129.8 (Ar), 129.9 (Ar), 131.2 (Ar),
133.0 (Ar), 133.2 (Ar), 133.4 (Ar), 141.2 (Ar), 143.2 (Ar), 143.4 (Ar), 154.7 (C=O), 154.9 (C=O),
160.2 (C=O), 164.9 (C=O), 165.2 (C=O), 165.8 (C=O). ESI-MS: calcd for C56H56O13SiNa 987.3388
(M+Na)+, found m/z 987.3381.
(2R,5R,6R,7S)-2-((2-((tert-Butyldimethylsilyl)oxy)-4-methoxybenzyl)oxy)-5-(hydroxymethyl)-2-phenyl-
tetrahydro-3aH-[1,3]dioxolo[4,5-b]pyran-6,7-diyl dibenzoate (3). To a solution of 2a (30 mg,
0.032 mmol) in pyridine (0.5 mL) was added Et3N (13 µL, 0.095 mmol) at room temperature. After
being stirred overnight, the reaction mixture was concentrated in vacuo. The residue was purified by

Molecules 2014, 19
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silica gel column chromatography (hexane/acetone = 2:1) to give 3 as a clear oil (19 mg, 81%):
α23D +9.5 (c 1.00, CHCl3); IR (film) 3527, 1725, 1613, 1261 cm−1; 1H-NMR (400 MHz) δ 0.15 (6H,
s, SiCH3), 0.90 (9H, s, tBu), 3.65 (1H, m, H-6), 3.75 (3H, s, OCH3), 3.78 (1H, m, H-6'), 3.89 (1H, m,
H-5),4.29 (1H, d, J = 11.2 Hz, ArCH2), 4.36 (1H, d, J = 11.2 Hz, ArCH2), 4.75 (1H, ddd, J = 5.2, 2.9,
0.8 Hz, H-2), 5.37 (1H, m, H-4), 5.74 (1H, m, H-3), 6.03 (1H, bd, H-1), 6.31 (1H, d, J = 2.5 Hz, Ar),
6.48 (1H, dd, J = 8.5, 2.5 Hz, Ar), 7.18 (1H, d, J = 8.5 Hz, Ar), 7.44 (7H, m, Ar), 7.60 (2H, m, Ar),
7.81 (2H, m, Ar), 7.98 (2H, m, Ar), 8.05 (2H, m, Ar); 13C-NMR (100 MHz) δ −4.3 (SiCH3), 18.1
(tBu), 25.6 (tBu), 55.2 (OCH3), 61.2 (ArCH2), 62.4 (C-6), 68.3 (C-2), 69.4 (C-3), 69.8 (C-4), 72.6 (C-5),
97.5 (C-1), 105.3 (Ar), 105.9 (Ar), 120.5 (Ar), 121.1 (Ar), 126.4 (orthoester-C), 128.3 (Ar), 128.4 (Ar),
128.5 (Ar), 129.0 (Ar), 129.1 (Ar), 129.6 (Ar), 129.9 (Ar), 130.0 (Ar), 133.5 (Ar), 133.6 (Ar), 135.3
(Ar), 154.0 (C=O), 159.9 (C=O), 164.6 (C=O), 165.6 (C=O). ESI-MS: calcd for C41H46O11NaSi
765.2707 (M+Na)+, found m/z 765.2710.
Cyclohexyl 2,3,4-tri-O-benzoyl-6-O-9-fluorenylmethyloxycarboxyl-β-D-glucopyranoside (7). A 0.1 M
solution of cyclohexanol in 1,2-DCE (10 mL) containing 4Å MS and Bu4NClO4 (3.42 g, 1 M) as a
supporting salt, was electrolyzed by constant current electrolysis (C.C.E., 6 mA/cm2) at 40 °C, using a
glassy carbon plate (1.5 cm × 1.5 cm) as an anode and a Pt plate (1.8 cm × 1.8 cm) as a cathode. After
the electrolysis, the reaction mixture (3.4 mL, 0.1 M EGA solution, 3 equiv.) was added to a solution
of 2a (0.11 g, 0.11 mmol), cyclohexanol (0.4 mL, 0.34 mmol), and 4Å MS in 1,2-DCE (1.1 mL). After
being stirred 20 min, the mixture was filtered through a Celite pad and the filtrate was washed with sat.
aq. NaHCO3. The organic extracts were dried (Na2SO4), and concentrated in vacuo. The residue was
purified by preparative TLC (hexane/acetone = 5:1) to give 7 as an oil (0.056 g, 62%): α23 −4.10
D
1
(c 1.00, CHCl3); IR (film) 1732, 1601, 1262 cm−1; H-NMR (400 MHz) δ 1.22 (5H, m, cyclohexyl),
1.46 (2H, m, cyclohexyl), 1.70 (2H, m, cyclohexyl), 1.88 (1H, m, cyclohexyl), 3.72 (1H, m,
cyclohexyl), 4.09 (1H, m, Fmoc), 4.22 (1H, m, Fmoc), 4.36 (3H, m, H-5,6a, Fmoc), 4.46 (1H, d,
J = 11.4, 6.5 Hz, H-6b), 4.93 (1H, d, J = 8.0 Hz, H-1), 5.51 (1H, dd, J = 9.6, 8.0 Hz, H-2), 5.55 (1H, t,
J = 9.6 Hz, H-4), 5.90 (1H, t, J = 9.6 Hz, H-3), 7.30 (11H, m, Ar), 7.51 (2H, m, Ar), 7.60 (2H, m, Ar),
7.77 (2H, m, Ar), 7.84 (2H, m, Ar), 7.95 (4H, m, Ar); 13C-NMR (100 MHz) δ 23.4 (cyclohexyl), 23.6
(cyclohexyl), 25.3 (cyclohexyl), 31.5 (cyclohexyl), 33.1 (cyclohexyl), 46.6 (Fmoc), 66.5 (C-6), 69.8
(C-4), 70.2 (C-2,3), 72.0 (FmocCH2), 72.9 (C-5), 78.2 (cyclohexyl), 99.7 (C-1), 120.0 (Ar), 125.2 (Ar),
127.2 (Ar), 127.9 (Ar), 128.3 (Ar), 128.4 (Ar), 128.7 (Ar), 128.8 (Ar), 129.5 (Ar), 129.7 (Ar), 129.8
(Ar), 133.1 (Ar), 133.2 (Ar), 133.5 (Ar), 141.2 (Ar), 143.2 (Ar), 123.3 (Ar), 154.8 (C=O), 164.9
(C=O), 165.3 (C=O), 165.8 (C=O). ESI-MS: calcd for C48H45O117997.2962 (M+H)+, found m/z 797.2936.
Cyclohexyl 2,3,4-tri-O-benzoyl-β-D-glucopyranoside (8). To a solution of 7 (54 mg, 0.068 mmol) in
pyridine (1 mL) was added Et3N (19 µL, 0.14 mmol) at room temperature, and the mixture was stirred
overnight. The mixture was concentrated in vacuo. The residue was purified by silica gel column
chromatography (hexane/acetone = 2:1) to give 8 as an oil (37 mg, 95%): α23D−11.5 (c 1.00, CHCl3);
IR (film) 3523, 1731, 1601, 1263 cm−1; 1H-NMR (400 MHz) δ 1.05 (3H, m, cyclohexyl), 1.48 (6H, m,
cyclohexyl), 1.77 (1H, m, cyclohexyl), 2.41 (1H, bs, OH), 3.68 (4H, m, cyclohexyl, H-5,6a,6b), 4.38
(1H, d, J = 8.0 Hz, H-1), 5.37 (2H, bt, H-2,4), 5.82 (1H, t, J = 9.6 Hz, H-3), 7.18 (3H, m, Ar), 7.31
(4H, m, Ar), 7.41 (2H, m, Ar), 7.74 (2H, m, Ar), 7.85 (4H, m, Ar); 13C-NMR (100 MHz) δ 23.4

Molecules 2014, 19
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(cyclohexyl), 23.6 (cyclohexyl), 25.4 (cyclohexyl), 31.5 (cyclohexyl), 33.2 (cyclohexyl), 61.5 (C-6),
69.7 (C-4,4',6'), 72.1 (C-2,3,3'), 72.9 (C-2',5'), 74.5 (C-5), 78.1 (cyclohexyl), 99.6 (C-1,1'), 128.2 (Ar),
128.3 (Ar), 128.5 (Ar), 128.6 (Ar), 128.9 (Ar), 129.5 (Ar), 129.6 (Ar), 129.7 (Ar), 129.9 (Ar), 133.1
(Ar), 133.2 (Ar), 133.6 (Ar), 164.9 (C=O), 165.8 (C=O), 165.9 (C=O). ESI-MS: calcd for C33H34O9Na
597.2101 (M+Na)+, found m/z 597.2112.
Cyclohexyl 2,3,4-tri-O-benzoyl-β-D-glucopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-6-O-9-fluorenyl-
methyloxycarboxyl-β-D-glucopyranoside (9). A solution of cyclohexanol in 1,2-DCE (10 mL, 0.1 M)
containing 4Å MS and Bu4NClO4 (3.42 g, 1 M) as a supporting salt, was electrolyzed by constant
current electrolysis (C.C.E., 6 mA/cm2) at 40 °C, using a glassy carbon plate (1.5 cm × 1.5 cm) as an
anode and a Pt plate (1.8 cm × 1.8 cm) as a cathode. The reaction mixture (0.6 mL, 0.1 M EGA
solution, 3 equiv.) was added to a solution of 2a (31 mg, 0.032 mmol) and 8 (37 mg, 0.064 mmol), and
MS 4A in 1,2-DCE (0.5 mL). After being stirred 20 min, the mixture was filtered through a Celite pad,
and the filtrate was washed with sat. aq. NaHCO3. The organic extract was dried (Na2SO4), and
concentrated in vacuo. The residue was purified by preparative TLC (hexane/acetone = 2:1) to give 9
1
as an oil (16 mg, 38%): α23 −9.8 (c 1.00, CHCl3); IR (film) 1734, 1601, 1261 cm−1; H-NMR
D
(400 MHz) δ 1.41 (10H, m, cyclohexyl), 3.62 (1H, m, cyclohexyl), 4.00 (4H, m, Fmoc, H-5,5'), 4.29
(5H, m, H-6a,6b,6a',6b'), 4.78 (1H, d, J = 7.8 Hz, H-1'), 5.04 (1H, d, J = 7.8 Hz, H-1), 5.34 (1H, t,
J = 9.8 Hz, H-4'), 5.41 (1H, dd, J = 9.8, 7.8 Hz, H-2'), 5.48 (1H, dd, J = 9.8, 7.8 Hz, H-2), 5.52 (1H, t,
J = 9.8 Hz, H-4), 5.80 (1H, t, J = 9.8 Hz, H-3'), 5.83 (1H, t, J = 9.8 Hz, H-3), 7.41 (22H, m, Ar), 7.62
(2H, m, Ar), 7.78 (6H, m, Ar), 7.92 (8H, m, Ar); 13C-NMR (100 MHz) δ 22.9 (cyclohexyl), 23.2
(cyclohexyl), 25.4 (cyclohexyl), 31.2 (cyclohexyl), 32.9 (cyclohexyl), 46.6 (Fmoc), 66.0 (C-6), 68.3
(C-6'), 69.3(C-4), 69.9 (C-4'), 70.3 (C-3), 71.7 (C-2,3'), 71.9 (C-2'), 72.1 C-5'), 72.8 (C-5), 72.9
(cyclohexyl), 74.1 (FmocCH2), 99.4 (C-1'), 100.8 (C-1), 119.9 (Ar), 125.3 (Ar), 125.4 (Ar), 127.2
(Ar), 127.8 (Ar), 128.2 (Ar), 128.3 (Ar), 128.4 (Ar), 128.7 (Ar), 128.9 (Ar), 129.3 (Ar), 129.5 (Ar),
129.6 (Ar), 129.7 (Ar), 129.8 (Ar), 133.0 (Ar), 133.1 (Ar), 133.2 (Ar), 133.4 (Ar), 133.5 (Ar), 141.2
(Ar), 143.3 (Ar), 143.4 (Ar), 154.8 (C=O), 164.9 (C=O), 165.1 (C=O), 165.2 (C=O), 165.4 (C=O),
165.6 (C=O), 165.7 (C=O). ESI-MS: calcd for C75H66O19Na 1293.4096 (M+Na)+, found m/z 1293.4078.
Phenyl 2,3,4-tri-O-benzoyl-6-O-9-fluorenylmethyloxycarboxyl-β-D-glucopyranosyl-(1→6)-2,3,4-tri-O-
benzoyl-1-thio-β-D-glucopyranoside (11). A solution of cyclohexanol in 1,2-DCE (10 mL, 0.1 M)
containing 4Å MS and Bu4NClO4 (3.42 g, 1 M) as a supporting salt was electrolyzed by constant
current electrolysis (C.C.E., 6 mA/cm2) at 40 °C, using a glassy carbon plate (1.5 cm × 1.5 cm) as an
anode and a Pt plate (1.8 cm × 1.8 cm) as a cathode. After the electrolysis, the reaction mixture
(1.3 mL, 0.1 M EGA solution, 3 equiv.) was added to a solution of 2a (34 mg, 0.036 mmol), 10 (42 mg,
0.072 mmol), m and 4Å MS in 1,2-DCE (0.5 mL). After being stirred for 20 min, the reaction mixture
was filtered through a Celite pad, and the filtrate was washed with sat. aq. NaHCO3. The organic
extracts were dried (Na2SO4), and concentrated in vacuo. The residue was purified by preparative TLC
(hexane/acetone = 2:1) to give 11 as an oil (29 mg, 63%): α23 +8.9 (c 1.00, CHCl3); IR (film) 1733,
D
1
1601, 1261 cm−1; H-NMR (400 MHz) δ 3.99 (4H, m, Fmoc, H-5,5'), 4.24 (1H, m, Fmoc), 4.38 (4H,
m, H-6a,6b,6a',6b'), 4.92 (1H, d, J = 9.8 Hz, H-1'), 4.97 (1H, d, J = 8.0 Hz, H-1), 5.31 (1H, t,
J = 9.8 Hz, H-4'), 5.34 (1H, t, J = 9.8 Hz, H-3'), 5.51 (2H, m, H-2,4), 5.81 (1H, t, J = 9.8 Hz, H-3'),

Molecules 2014, 19
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5.85 (1H, t, J = 9.8 Hz, H-3), 7.23 (2H, m, Ar), 7.39 (25H, m, Ar), 7.61 (2H, m, Ar), 7.74 (4H, m, Ar),
7.85 (4H, m, Ar), 7.93 (6H, m, Ar); 13C-NMR (100 MHz) δ 46.6 (Fmoc), 65.9 (C-6), 68.4 (C-6'), 69.4
(C-4'), 69.5 (C-4'), 70.2 (C-3'), 70.4 (C-3), 71.6 (C-2), 72.0 (C-2'), 72.8 (C-5'),74.0 (FmocCH2), 78.3
(C-5), 85.9 (C-1'), 101.0 (C-1), 119.9 (Ar), 125.3 (Ar), 125.4 (Ar), 127.2 (Ar), 128.2 (Ar), 128.3 (Ar),
128.4 (Ar), 128.5 (Ar), 128.6 (Ar), 128.7 (Ar), 128.8 (Ar), 129.1 (Ar), 129.2 (Ar), 129.7 (Ar), 129.8
(Ar), 131.7 (Ar), 133.2 (Ar), 133.4 (Ar), 133.5 (Ar), 141.2 (Ar), 141.3 (Ar), 143.3 (Ar), 143.4 (Ar),
154.8 (C=O), 164.9 (C=O), 165.1 (C=O), 165.2 (C=O), 165.3 (C=O), 165.6 (C=O), 165.7 (C=O).
ESI-MS: calcd for C75H60O18NaS 1303.3398 (M+Na)+, found m/z 1303.3385.
Phenyl 2,3,4-tri-O-benzoyl-6-O-9-fluorenylmethyloxycarboxyl-β-D-glucopyranosyl-(1→6)-2,3,4-tri-O-
benzoyl-β-D-glucopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-1-thio-β-D-glucopyranoside (12). To
a
solution of 11 (29 mg, 0.022 mmol) in pyridine (0.5 mL) was added Et3N (6.3 µL, 0.045 mmol) at
room temperature. After being stirred overnight, the mixture was concentrated in vacuo. The residue
was purified by silica gel column chromatography (hexane/acetone = 2:1) to give an alcohol as an oil
1
(19 mg, 82%): α23 −2.1 (c 1.00, CHCl3); IR (film) 3062, 1731, 1601, 1262 cm−1; H-NMR
D
(400 MHz) δ 2.81 (1H, dd, J = 8.0, 5.6 Hz, OH), 3.60 (1H, m, H-5), 3.74 (2H, m, H-5',6a'), 3.95 (1H,
dd, J = 10.5, 5.6 Hz, H-6b'), 4.05 (2H, m, H-6a,6b), 4.92 (2H, bt, H-1,1'), 5.23 (1H, t, J = 9.6 Hz,
H-4'), 5.40 (3H, m, H-2,2'), 5.81 (1H, t, J = 9.6 Hz, H-3'), 5.85 (1H, t, J = 9.6 Hz, H-3), 7.38 (19H, m,
13
Ar), 7.53 (4H, m, Ar), 7.76 (2H, m, Ar), 7.81 (2H, m, Ar), 7.93 (8H, m, Ar); C-NMR (100 MHz) δ
61.2 (C-6), 67.9 (C-6'), 69.4 (C-4'), 69.9 (C-4), 70.4 (C-3'), 71.5 (C-3), 72.8 (C-2), 73.9 (C-2'), 74.6
(C-5'), 77.8 (C-5), 86.0 (C-1'), 100.5 (C-1), 128.2 (Ar), 128.3 (Ar), 128.4 (Ar), 128.5 (Ar), 128.7 (Ar),
128.8 (Ar), 129.1 (Ar), 129.2 (Ar), 129.7 (Ar), 129.8 (Ar), 129.9 (Ar), 131.5 (Ar), 133.2 (Ar), 133.3
(Ar), 133.5 (Ar), 133.6 (Ar), 164.9 (C=O), 165.1 (C=O), 165.6 (C=O), 165.7 (C=O), 165.9 (C=O).
ESI-MS: calcd for C60H50O16NaS 1081.2717 (M+Na)+, found m/z 1081.2732.
A solution of cyclohexanol in 1,2-DCE (10 mL, 0.1 M) containing 4Å MS and Bu4NClO4 (3.42 g, 1 M)
as a supporting salt, was electrolyzed by constant current electrolysis (C.C.E., 6 mA/cm2) at 40 °C,
using a glassy carbon plate (1.5 cm × 1.5 cm) as an anode and a Pt plate (1.8 cm × 1.8 cm) as a
cathode. The reaction mixture (0.6 mL, 0.1 M EGA solution, 3 equiv.) was added to a solution of 2a
(37 mg, 0.038 mmol), the alcohol (20 mg, 0.019 mmol), and 4Å MS in 1,2-DCE (0.5 mL). After being
stirred at 40 °C for 20 min, the reaction mixture was filtered through a Celite pad, and the filtrate was
washed with saturated aqueous NaHCO3. The organic extracts were dried (Na2SO4), and concentrated
in vacuo. The residue was purified by preparative TLC (Et2O/hexane = 2:1) to give 12 as a clear oil
1
(4.1 mg, 12%): α23D −5.4 (c 1.00, CHCl3); IR (film) 1733, 1601, 1261 cm−1; H-NMR (400 MHz) δ
3.62 (1H, dd, J = 11.2, 5.0 Hz, H-6a), 3.84 (3H, m, Fmoc, H-5',6), 4.01 (2H, m, H-5,5''), 4.31 (6H, m,
H-6a,6b,6a',6b'), 4.61 (1H, d, J = 8.0 Hz, H-1'), 4.99 (1H, d, J = 10.0 Hz, H-1''), 5.08 (1H, d,
J = 8.0 Hz, H-1), 5.10 (1H, t, J = 10.0 Hz, H-4), 5.21 (1H, dd, J = 9.8, 8.0 Hz, H-2''), 5.51 (4H, m,
H-2,2',4',4''), 5.68 (1H, t, J = 9.8 Hz, H-3'), 5.84 (1H, t, J = 9.8 Hz, H-3''), 6.06 (1H, t, J = 9.8 Hz, H-3),
7.18 (2H, m, Ar), 7.36 (34H, m, Ar), 7.58 (2H, m, Ar), 7.76 (2H, m, Ar), 7.92 (14H, m, Ar); 13C-NMR
(100 MHz) δ 46.6 (Fmoc), 66.2 (C-6), 68.3 (C-6''), 69.4 (C-6'), 69.7 (C-4''), 70.1 (C-4), 70.2 (C-4'),
70.5 (C-3''), 71.7 (C-3), 71.9 (C-2,2',3'), 72.0 (C-2''), 72.6 (C-5'), 72.7 (C-5''), 73.9 (C-5), 74.1
((FmocCH2), 86.3 (C-1''), 100.5 (C-1), 101.2 (C-1'), 119.9 (Ar), 120.0 (Ar), 125.3 (Ar), 125.4 (Ar),

Molecules 2014, 19
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127.2 (Ar), 127.8 (Ar), 128.1 (Ar), 128.2 (Ar), 128.3 (Ar), 128.4 (Ar), 128.5 (Ar), 128.6 (Ar), 128.7
(Ar), 128.8 (Ar), 129.0 (Ar), 129.1 (Ar), 129.3 (Ar), 129.4 (Ar), 129.7 (Ar), 129.8 (Ar), 130.0 (Ar),
132.1 (Ar), 132.8 (Ar), 133.0 (Ar), 133.2 (Ar), 133.4 (Ar), 141.2 (Ar), 143.3 (Ar), 143.4 (Ar), 154.8
(C=O), 164.9 (C=O), 165.2 (C=O), 165.3 (C=O), 165.7 (C=O). ESI-MS: calcd for C102H82O26NaS
1777.4713 (M+Na)+, found m/z 1777.4736.
4. Conclusions
The orthoester 2a bearing the selectively removable Fmoc group at the C-6 position was
synthesized, and its utility as a glycosyl donor and acceptor after deprotection of the Fmoc group
was demonstrated.
Acknowledgments
We thanked for MEXT-Supported Programs for the Strategic Research Foundation at Private
Universities. 2009-2013, 2012-2016; Scientific Research (C) from MEXT; Sasakawa Research Grant
for Financial Support (to KK).
Author Contributions
The listed authors contributed to this work as described in the following. Kohei Kawa synthesized
the sugar derivatives. Tsuyoshi Saitoh made technical supports of the chemical and electrochemical
procedures. Eisuke Kaji contributed with discussions of carbohydrate chemistry and the research
direction. Shigeru Nishiyama conducted the research progress. All authors helped preparing
manuscript and approved the final version.
Conflicts of Interest
The authors declare no conflict of interest.
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21. When the glycosylation using Ag2CO3, the yield of 2a was reduced to 17%.
22. The reaction condition has not yet optimized.
Sample Availability: Samples are available from the authors.
© 2014 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
(http://creativecommons.org/licenses/by/3.0/).