Reconsidering glycosylations at high temperature: Precise microwave heating

Article abstract of DOI:10.1039/b511266d

Current methods for glycosylation of complex alcohols, e.g. with glycosyl trichloroacetimidates, generally occur in the presence of a strong Lewis acid 'promoter', and at sub-ambient temperatures. However, the older literature reports high-temperature gly

Full text of DOI:10.1039/b511266d

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A R T I C L E
Reconsidering glycosylations at high temperature: precise
microwave heating
Kim Larsen,a Kasper Worm-Leonhard,a Peter Olsen,a Andreas Hoelb and Knud J. Jensen*a
a Department of Natural Sciences, Section for Bioorganic Chemistry, Royal Veterinary and
Agricultural University, DK-1871, Frederiksberg, Denmark. E-mail: kjj@kvl.dk;
Fax: +45 3528 2398; Tel: +45 3528 2430
b Biotage AB, SE-75318, Uppsala, Sweden
Received 8th August 2005, Accepted 8th September 2005
First published as an Advance Article on the web 27th September 2005
Current methods for glycosylation of complex alcohols, e.g. with glycosyl trichloroacetimidates, generally occur in the
presence of a strong Lewis acid ‘promoter’, and at sub-ambient temperatures. However, the older literature reports
high-temperature glycosylations, especially of phenols. We have described an efficient method for glycosylation of
alcohols under neutral conditions, using as anomeric leaving group methyl 3,5-dinitrosalicylate (DISAL). Only a very
few reports have described the use of microwaves to promote glycosylations, mainly of simple alcohols. Here we
describe fast, high-temperature glycosylations using precise microwave heating in the synthesis of oligosaccharides,
with both DISAL and widely used trichloroacetimidate glycosyl donors in the absence of strong Lewis acids. Also,
we have applied microwave heating as a general protocol for evaluating new, potential glycosyl donors.
cosylation at 100 ◦C with n-pentenyl orthoesters in the presence
Introduction
of the strong Lewis acid N-iodosuccinimide (2.2 equiv).8f We
Glycoconjugates play crucial roles in the development, growth,
hypothesized that glycosylations under mild conditions, i.e.
in the absence of Lewis acid, could benefit from promotion
by elevated temperatures. We were encouraged by preliminary
results in which glycosylation of a secondary hydroxyl with a
and proper function of an organism.1 There are numerous
biologically important poly- and oligosaccharides, e.g., in gly-
cans of O- and N-glycopeptides and -proteins, tumor-associated
antigens, lipochitin nodulation factors, and amino glycoside an-
DISAL donor had proceeded with high b-selectivity by brief
tibiotics such as streptomycin. Synthesis of oligosaccharides of
microwave heating to 130 ◦C.5f
these glycoconjugates provides important tools for glycobiology.
Anomeric trichloroacetimidates are widely used glycosyl
Most glycosylation protocols rely on Lewis acid ‘promoters’
donors. They are conventionally activated by strong Lewis
for activation of an anomeric leaving group, and glycosylations
acids such as trimethylsilyl trifluoromethanesulfonate9 or boron
of aliphatic alcohols have generally been performed at below
trifluoride etherate.10 However, these promotors require special
ambient temperature.2 However, aryl glycosides can often be
prepared in the absence of a Lewis acid promoter, e.g., by
generation of a phenoxide nucleophile or by nucleophilic
attention regarding temperature and moisture. Efficient glyco-
sylations with trichloroacetimidates in the absence of strong
Lewis acids would be desirable. Waldmann,11 Lubineau12 and co-
aromatic substitution.3 Also, classical work by Helferich and
workers have observed that glycosylation with trichloroacetimi-
others described high-temperature glycosylations, especially of
dates could be promoted by lithium salts at ambient temperature,
phenols.4 Novel methods for glycosylation of aliphatic alcohols
albeit requring long reaction times. Our guiding question was
in the absence of strong Lewis acids, i.e. under mild conditions,
whether precise microwave heating can be used in general to
hold great promise.3 We have described a new, efficient method
promote and accelerate a-selective glycosylations with DISAL
for glycosylation under strictly neutral or mildly basic condi-
donors and with well-established glycosyl trichloroacetimidates.
tions, or in the presence of lithium salts.5 In this glycosylation
Thus, we studied the synthesis of oligosaccharides in the
technique, the anomeric leaving group on benzyl- or benzoyl-
absence of added Lewis acids with microwave heating, aim-
protected donors is methyl 3,5-dinitrosalicylate (DISAL) or
ing to determine general conditions for microwave-promoted
its para regioisomer. The potential of DISAL glycosyl donors
glycosylations. We investigated glycosylations with DISAL and
was demonstrated in their successful application to solution-
trichloroacetimidate donors, as well as reactions with glycosyl
phase5a,d and solid-phase5b oligosaccharide synthesis, glycosy-
donors carrying a modified DISAL leaving group. Microwave
lation of natural product analogs (phenazines), as well as
heating was performed in closed (septum) vessels in a Biotage
intramolecular glycosylation via a novel 1,9-glycosyl shift.5c
Initiator instrument. Reaction times were generally 5–40 min
DISAL glycosyl donors are prepared by a convenient and robust
and the temperature in the 100–150 ◦C range. Reaction times
nucleophilic aromatic substitution protocol. These glycosyla-
tions are operationally simple and can be carried out in standard
plastic vials. However, to further optimize this procedure we
without microwave heating had been up to 31 h.
needed to shorten reaction times and improve yields.
Results and discussion
Microwave heating can dramatically increase reaction rates
in organic chemistry.6 Modern microwave instruments allow
First, can glycosylations with a DISAL donor be accelerated by
precise control of temperature and pressure in sealed reaction
tubes. However, changing the reaction temperature may alter
the distribution between the main reaction and side-reactions,
hence, while precise microwave heating can promote many
reactions, not all reactions will benefit.7 This technique has not
been employed widely in glycosylations, with few exceptions.8
Fraser-Reid and co-workers reported microwave-promoted gly-
precise microwave heating? To answer this question we repeated
previous experiments but now under microwave conditions
(Scheme 1, Table 1). First, the reaction time for (solvolytic)
glycosylation of methanol with DISAL donor 1 was reduced
◦
from 6 h at 30 C5a to 5 min at 100 ◦C (Table 1, entry 1).
Importantly, the reaction proceeded with inversion, as before.
Similarly, glycosylation of cyclohexanol (5 equiv) in NMP also
3 9 6 6
O r g . B i o m o l . C h e m . , 2 0 0 5 , 3 , 3 9 6 6 – 3 9 7 0
T h i s j o u r n a l i s
T h e R o y a l S o c i e t y o f C h e m i s t r y 2 0 0 5
©

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Table 1 Glycosylations with DISAL donor 1 (a/b 4.7 : 1)
Entry
Acceptor
Additive, solvent
Microwave
Product, yield (a/b ratio)
1
2
3
4
5
MeOH
Cyclohexanol
4
4
6
MeOH
NMP
NMP
LiClO4, (CH2Cl)2
LiClO4, (CH2Cl)2
100 ◦C, 5 min
100 ◦C, 5 min
130 ◦C, 40 min
100 ◦C, 30 min
100 ◦C, 40 min
2, 91% (1 : 4.4)
3, 71% (2.1 : 1)
5, 60% (1.9 : 1)
5, 97% (2 : 1)
7, 72% (4 : 1)
Scheme 1 Microwave promoted a-selective 1,6- and 1,4-glycosylation
with DISAL donor 1. Conditions and results in Table 1.
proceeded in 5 min at 100 ◦C (entry 2) (previously 2 h at 40 ◦C
in NMP).5a It was gratifying to see that microwave heating
at 130 ◦C accelerated the glycosylation of monosaccharide
4 to give disaccharide 5 in 60% yield (entry 3). Performing
the same reaction in (CH2Cl)2 with LiClO4 improved the
yield to an impresive 97% (entry 4). The 4-OH in GlcNAc
derivatives is considered one of the most challenging hydroxyls
to glycosylate.13 It was very rewarding to see that the 4-OH in
GlcNAc derivative 65b was glycosylated in a high yield of 72%
(a/b 4 : 1) (entry 5).
Next, can the reactivity of the DISAL donor be ‘tuned’ by
substituting its methyl moiety for a sterically or electronically dif-
ferent group? To test this, the 2-propyl (9) and 2-(4-nitrophenyl)
(10) ester analogs of DISAL were prepared and subjected to
the above conditions for microwave glycosylation (Scheme 2).
Thus, the aryl fluoride 2-fluoro-3,5-dinitrobenzoic acid (8) was
converted to the 2-propyl ester 9 by treatment with oxalyl
chloride (cat. DMF) followed by 2-propanol. The 2-propyl
DISAL glycoside 11 was synthesized using the standard protocol
for the preparation of DISAL glycosides. The nitrophenyl
DISAL donor 12 was prepared in the same way.
Scheme 2 Synthesis of DISAL donors 9 and 10 and subsequent glyco-
sylations. Reagents and conditions: (a) i) oxalyl chloride, DMF, DCM,
r.t.; ii) 2-propanol, 78%; (b) i) SOCl2, THF, 80 ◦C; ii) 4-nitrophenol,
THF, pyridine, 0 ◦C, 92%; (c) Li2CO3, DMAP, CH2Cl2, r.t., 20 min; (d)
see Table 2.
crowave heating under very mild conditions. Thus, 2,3,4,6-tetra-
O-benzyl-D-glucopyranosyl trichloroacetimidate 1314 was used
to glycosylate cyclohexanol, the galactose derivative 4, and the
glucose derivative 14 (Scheme 3 and Table 3).
Solvolytic glycosylation of MeOH with 2-propyl DISAL
donor 11 proceeded in 5 min at 100 ◦C (Table 2, entry 1).
Glycosylation of cyclohexanol (5 equiv) with conventional
◦
heating in NMP at 60 C required 31 h (Table 2, entry 2). The
same reaction but with microwave heating at 100 ◦C took only
5 min (Table 2, entry 3; somewhat lower yield but with improved
a-selectivity). In these experiments the 2-propyl DISAL donor
showed no real advantages compared to the original DISAL
leaving group. Somewhat surprisingly, the nitrophenyl DISAL
donor 12 proved less reactive (Table 2, entry 4). However, these
microwave conditions provided a fast method for the evaluation
of these modified DISAL glycosyl donors.
Scheme 3 Microwave-promoted disaccharide synthesis using trichloro-
acetimidate donor 13 (Table 3).
Waldmann11 reported that glycosylation of cyclohexanol with
13 in 1 M solutions of LiClO4 in either Et2O or DCM gave the
cyclohexyl derivative 3 in 50–63% yield after 24 h. We now report
that the same reaction in (CH2Cl)2 with 0.09 M LiClO4 under
microwave irradiation for 5 min at 100 ◦C increased the yield to
80% (Table 3, entry 1).
Finally, we wanted to study whether glycosylations with
widely used trichloroacetimidates could be accelerated by mi-
Table 2 Glycosylation with donors 11 and 12
Entry
Donor (a/b ratio)
Conventional
Microwave
Product, yield (a/b ratio)
1
2
3
4
11 (>49 : 1)
11 (>1 : 49)
11 (>1 : 49)
12 (>1 : 49)
—
100 ◦C MeOH, 5 min
—
2, 73% (1 : 17)
3, 82% (1 : 1)
3, 64% (6.5 : 1)
2, 54% (2 : 1)
60 ◦C, cyclohexanol, NMP, 31 h
—
—
100 ◦C, cyclohexanol, NMP, 5 min
150 ◦C, MeOH, 2 × 10 min
O r g . B i o m o l . C h e m . , 2 0 0 5 , 3 , 3 9 6 6 – 3 9 7 0
3 9 6 7

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Table 3 Microwave glycosylation with donor 13 in (CH2Cl)2 in the presence of LiClO4
Entry
Donor
Acceptor
Reaction time
Product, yield (a/b ratio)
1
2
3
13
13
13
Cyclohexanol
4
14
5 min
25 min
30 min
3, 80% (2 : 1)
5, 80% (1 : 1)
15, 72% (1.5 : 1)
The glycosylation of 4 with 13 under microwave conditions
increased the yield from its previous value of 45–47% to 80%.
In this case, the time for microwave irradiation was increased
to 25 min to ensure complete conversion to the disaccharide
5 (Table 3, entry 2). Finally, the glycosylation of acceptor 14
proceeded in 72% yield after 30 min of microwave irradiation
at 100 ◦C (entry 3). This compares favorably with the results
obtained by Lubineau11 using LiOTf (0.5 equiv), which gave
77% yield after 86 h. These glycosylations gave a/b ratios
comparable to those obtained by Waldmann11 and Lubineau.12
During our microwave-promoted glycosylations, no hydrolysis
or rearrangement of the trichloroacetimidate was detected.
5.0% C, 95.0% D; 9.00 min: 1.00 ml min−1, 95.0% C, 5.0% D;
15.00 min: 1.00 ml min−1, 95.0% C, 5.0% D.
Characteristic absorption maxima were: Bn: 256–257 nm,
DISAL-OH: 220 nm and 286 nm.
Preparative HPLC was performed on a Waters 600 system
with Waters 996 diode array detector and three consecutive
columns (40 × 100 mm prep. NOVA Pak HR C18 6 lm
˚
60 A units). Linear gradients of CH3CN (D) and water (MilliQ)
(C) were used.
Program A: 0.00 min: 0.00 ml min−1, 95.0% C, 5.0% D;
1.00 min: 20.00 ml min−1, 95.0% C, 5.0% D; 20.00 min:
20.00 ml min−1, 50.0% C, 50.0% D; 60.00 min: 20.00 ml min−1,
5.0% C, 95.0% D; 75.00 min: 20.00 ml min−1, 0.0% C, 100.0%
D; 81.00 min: 0.00 ml min−1, 0.0% C, 100.0% D.
Summary
1H, 13C, gHSQC, HMBC and H,H-COSY NMR spectra were
recorded on a Bruker Avance 300 spectrometer. The chemical
shifts are referred to the residual solvent signal. Chemical shifts
(d) values are in ppm, coupling constants (J) are in Hz. Mass
determination (high-resolution MS, HR-MS) was performed
on a Micromass LCT instrument with an ESI probe. For
TLC, Merck TLC aluminum sheets coated in silica gel 60 F254
were used. Compounds containing UV-absorbing groups were
visualized under UV light (254 nm), and carbohydrates were
developed with 2 M H2SO4 followed by charring with a heat
gun. Vacuum liquid chromatography (VLC) was performed on
columns of Merck 60H silica packed under vacuum. The crude
product was dissolved in CH2Cl2, the equivalent amount of silica
added, concentrated, placed on the column and finally covered
with acid-rinsed sea sand. Chromatography was thereafter run
with the appropriate eluents until the product was collected.
Microwave experiments was carried out in a Biotage Initiator
(Biotage, Sweden). The reaction times are specified as the ramp
time, and the hold times at the final temperature. Temperatures
are measured by IR, and pressure via the septum. The new
compounds described in this paper are all fully characterized.
All known compounds have been verified by NMR and ESI-MS.
In conclusion, microwave heating to 100–130 ◦C efficiently
promoted O-glycosylations with DISAL donors in either NMP
or (CH2Cl)2 with LiClO4. Even the difficult 1,4-glycosylation of
a GlcNAc derivative was achieved. Glycosylation with widely
used trichloroacetimidates in the presence of LiClO4 were also
promoted by precise microwave heating. These glycosylations
are operationally simple. The obtained a/b ratios were compa-
rable to those previously obtained at 20–60 ◦C; glycosylation of
sterically demanding alcohols with DISAL donors proceeded
with a-selectivity. The key to this success appears to be the
absence of strong Lewis acids. These conditions have also been
applied in the screening of new, potential glycosyl donors.
Evaluation of two new DISAL donor analogs under microwave
conditions showed that they could O-glycosylate, but that they
had no improved properties compared to the original DISAL
donor. We have thus demonstrated that heating to temperatures
hitherto not generally applied can be used to accelerate slow
glycosylations in oligosaccharide synthesis. Here, we have fo-
cused on a-selective glycosylations; very recently we reported
a b-selective glycosylation under comparable conditions.5f We
believe that the simplicity and efficiency of these microwave-
promoted glycosylations in the absence of strong Lewis acids
should make these protocols widely applicable.
Isopropyl 2-fluoro-3,5-dinitrobenzoate (9)
In a dry, round-bottomed flask, 2-fluoro-3,5-dinitrobenzoate
(8) (305 mg; 1.3 mmol) was dissolved in dry DCM (9 mL).
Dry DMF (30 lL) was added and oxalyl chloride (125 lL;
1.45 mmol) was added to the solution. Moderate gas evolution
from the solution was seen after addition of oxalyl chloride.
After 90 min dry 2-propanol (0.71 mL) was added, and after an
additional 60 min the reaction mixture was concentrated under
vacuum, yielding a yellowish-white powder. Recrystallization
fro◦m toluene–hexane gave white crystals (284 mg; 78%), mp 92–
93 C. 1H NMR (300 MHz, CDCl3): 9.05 (s, 2H, Ha + Hb), 5.35
(m, 1H, (CH3)2CH), 1.46 (s, 3H, CH3), 1.44 (s, 3H, CH3). 13C
NMR (75 MHz, CDCl3): 160.2, 157.7 (d, 1C, JC–F = 287 Hz,
C–F), 142.7, 138.7, 131.6, 125.6, 124.0, 71.5, 21.8, 21.8.
Experimental
˚
Molecular sieves (4 A) were activated under high vacuum at
◦
150 C for 24 h. CH3NO2 was purified by vacuum distillation
to remove the water–CH3NO2 azeotrope, and stored in dark
˚
bottles over 4 A molecular sieves. The water content (<20 ppm)
was measured by Karl-Fischer titration. All other solvents were
˚
˚
distilled and/or stored over 3 A or 4 A molecular sieves as
appropriate. Analytical HPLC was performed on a Waters 600
system with a 996 diode array detector and a 717 Autosampler
˚
equipped with a 3.9 × 50 mm Nova-Pak C18 4 lm 60 A column.
The following solvents were used: 0.1% TFA–H2O (A); 0.1%
TFA–CH3CN (B); H2O (C); CH3CN (D). Analytical HPLC
was performed on microfiltered or centrifuged 0.1% solutions
in MeCN (for more hydrophilic compounds, solubility was
improved by addition of water). The following programs were
used:
4-Nitrophenyl 2-fluoro-3,5-dinitrobenzoate (10)
To 8 (2.3 g; 10.0 mmol) in a dry round-bottomed flask was
added thionyl chloride (40 mL) under Ar. The reaction mixture
was refluxed for 2 h 45 min, followed by the removal of thionyl
chloride under reduced pressure. The residue was further dried
under high vacuum, resolubilized in dry THF (40 mL) and
cooled to 0 ◦C. 4-Nitrophenol (1.4 g; 10.0 mmol) with dry
pyridine (810 lL) in dry THF (10 mL) was added dropwise
Program A: 0.00 min: 1.00 ml min−1, 95.0% A, 5.0% B; 7.00
min: 1.00 ml min−1, 5.0% A, 95.0% B; 8.50 min: 1.00 ml min−1,
5.0% A, 95.0% B; 9.00 min: 1.00 ml min−1, 95.0% A, 5.0% B;
15.00 min: 1.00 ml min−1, 95.0% A, 5.0% B.
Program B: 0.00 min: 1.00 ml min−1, 95.0% C, 5.0% D; 7.00
min: 1.00 ml min−1, 5.0% C, 95.0% D; 8.50 min: 1.00 ml min−1,
3 9 6 8
O r g . B i o m o l . C h e m . , 2 0 0 5 , 3 , 3 9 6 6 – 3 9 7 0

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˚
under Ar over 5 min. After 15 min, the cold bath was removed
and the reaction mixture was filtered over a layer of silica gel,
which was subsequently washed with THF (3 × 20 mL). The
organic fractions was pooled and concentrated to dryness under
vacuum, and crystallized from toluen◦e to yield slightly yellowish
microcentrifuge tube (Plasticbrand), crushed 3 A molecular
sieves were added, and for the glycosylation of cyclohexa◦nol
(5 equiv.), NMP was added. The mixture was stirred at 60 C,
during which time the progress of the reaction was monitored
by HPLC and TLC (30% EtOAc in hexane). Evaporation
of the solvent was followed by purification by either VLC
chromatography (hexane–EtOAc 18 : 1 → 1 : 1) or preparative
HPLC to give the products indicated in Table 2.
Microwave heating: The glycosyl donor (typically 0.05 mmol)
were dissolved in either dry methanol or NMP (500 lL), and in
the case of NMP, cyclohexanol (0.25 mmol) was added followed
by mixing. The reaction mixture was subjected to microwave
radiation for 5 min at 100 or 150 ◦C (see Tables 1 and 2), a
sample (12 lL) was diluted in acetonitrile (0.7 mL) and analyzed
by analytical HPLC. From a series of test reactions, the product
was purified by preparative HPLC and characterized by NMR
and ES-MS.
1
crystals (3.2 g; 92%), mp 146–147 C. H NMR (300 MHz,
CDCl3): 9.25 (dd, 1H, JH = 5.3 Hz, JH
= 2.8 Hz, Hb),
–F
b
–H
b
a
9.19 (dd, 1H, JH = 5.8 Hz, JH
= 3.0 Hz, Ha), 8.9 (d, 2H,
–F
–H
b
a
a
ꢀ
ꢀ
ꢀ
ꢀ
ꢀ
a
ꢀ
–H
b
JH
= 9.2 Hz, Ha ), 7.51 (d, 2H, JH
= 9.0 Hz, Hb ).
–H
a
b
13C NMR (75 MHz, CDCl3): 158.4, 158.3, 158.1 (d, 1C, JC–F
=
289 Hz, C–F), 154.2, 146.2, 142.9, 132.0, 132.0, 126.2, 125.6,
125.6, 122.9, 122.9.
Synthesis of DISAL and DISAL-derived donors 1, 11, and 12
The hemiacetal (0.75 mmol), Li2CO3 (110 mg) and DISAL or
the DISAL derivative (0.9 mmol) were suspended in dry CH2Cl2
(15 mL). A solution of DMAP (28 mg) in dry CH2Cl2 (2 mL) was
added dropwise over 20 min at RT. This changed the color of the
mixture from colorless to yellow to orange. After a total reaction
time of 30 min the crude reaction mixture was transferred to a
packed VLC column. Elution was performed using CH2Cl2–
Et2O (95 : 5).
Methyl 2,3,4,6-tetra-O-benzyl-a,b-D-glucopyranoside (2a/b)
The observed NMR data were identical with literature values.15
Cyclohexyl 2,3,4,6-tetra-O-benzyl-a,b-D-glucopyranoside (3a/b)
The observed NMR data were identical with literature values.16
2,4-Dinitro-6-(methoxycarbonyl)phenyl
2,3,4,6-tetra-O-benzyl-a,b-D-glucopyranoside (1a/b)
The observed NMR data were identical with literature
values.5a,b,f
General procedure for the microwave-assisted glycosylation
reactions (Table 1–3)
The glycosyl donor (0.075 mmol, 1.5 equiv), the glycosyl
acceptor (0.05 mmol, 1 equiv), activators and additives (if
required – see Tables 2 and 3), crushed 3 A molecular sieves,
2,4-Dinitro-6-(isopropoxycarbonyl)phenyl
2,3,4,6-tetra-O-benzyl-a,b-D-glucopyranoside (11)
˚
Obtained as a slightly yellow foam (569 mg; 97%, a/b 3 : 1)
after evaporation of solvents from column chromatography. 1H
NMR: (300 MHz, CDCl3): 8.78 (d, 1H, Ha + Hb), 8.74 (d, 1H,
Ha + Hb), 8.64 (d, 1H, Ha + Hb), 8.54 (d, 1H, Ha + Hb), 7.47–
7.20 (m, 40H, Harom), 5.63 (d, 1H, JH1–H2 = 3.3 Hz, H-1a), 5.31
(m, 2H, (CH3)2CH, both anomers), 5.20 (d, 1H, JH1–H2 = 7.2 Hz,
H-1b), 5.04 (m, 3H, Ph–CH), 4.80 (d, 1H, Ph–CH), 4.70 (d, 1H,
Ph–CH), 4.60 (d, 1H, Ph–CH), 4.59 (m, 2H, Ph–CH,), 4.50 (d,
1H, Ph–CH), 4.46 (d, 1H, Ph–CH), 4.39 (m, 2H, Ph–CH), 4.14
(1H, JH2–H3 = 10.0 Hz, H-3a), 4.02 (ddd, 1H, JH4–H5 = 10 Hz,
JH5–H6 = 2.2 Hz, H-5a), 3.79–3.71 (m, 5H, H-4a, H-3a, H-2b,
H-6b), 3.68–3.57 (m, 3H, H-2a, H-6a, H-3b), 3.60 (dd, 1H, H-
4b), 3.34 (ddd, 1H, H-5b), 1.43 (m, 12H, (CH3)2CH). 13C NMR:
(75 MHz, CDCl3): for both anomers 162.7, 162.3, 153.8, 151.3,
144.7–122.4, 104.6, 103.4, 84.1, 82.1, 81.4, 80.9, 76.5, 75.8, 75.6,
75.1, 75.1, 75.0, 74.1, 73.9, 73.5, 71.2, 71.0, 68.7, 67.9, 53.4, 29.7,
21.8, 21.7, 21.6. ES-MS: m/z calcd. for C44H48N2O12Na [M +
NH4]+ 810.32; found 810.34.
and a magnetic stirrer bar were placed in a 5 mL microwave
reaction vial and fitted with a septum, which was then pierced
with a needle. The closed vial was then evacuated under high
vacuum; Ar was let in followed by re-evacuation. This cycle
was repeated twice and the mixture left to dry for 1–2 h. Ar
was let in, the needle was removed, dry solvent (500 lL) was
added under argon and the reaction mixture was subjected to
microwave radiation for 5 min at 100 ◦C (unless stated othervise).
The reaction mixture was transferred to a 15 mL Falcon plastic
tube, centrifuged for 3 min at 4000 rpm and the supernatant was
transferred to a new 15 mL Falcon plastic tube and concentrated
to dryness under a stream of air. The residue was loaded onto a
preparative HPLC column with 2 mL of CH3CN and purified,
followed by evaporation of appropriate fractions.
1,2;3,4-Di-O-isopropylidene-6-O-(2,3,4,6-tetra-O-benzyl-a,b-D-
glucopyranosyl)-a-D-galactopyranoside (5a/b)
The observed NMR data were identical with literature values.17
2,4-Dinitro-6-(4-nitrophenoxycarbonyl)phenyl
2,3,4,6-tetra-O-benzyl-a,b-D-glucopyranoside (12)
Methyl 2-acetamido-3,6-di-O-benzyl-2-deoxy-4-O-(2,3,4,6-tetra-
O-benzyl-a,b-D-glucopyranosyl)-a-D-glucopyranoside (7a/b)
The observed NMR data were identical with literature values.5b
Obtained as a white foam (200 mg; quant.; a/b 2 : 1) after
evaporation of solvents from column chromatography. 1H
NMR:(300 MHz, CDCl3): 8.98 (d, 1H, JH
= 3.0Hz, Hb), 8.87
–H
a
b
a
ꢀ
ꢀ
ꢀ
(d, 1H, JH
= 2.6 Hz, Ha), 8.18 (d, 2H, JH
= 9.6 Hz, Ha ),
–H
–H
a
8.09 (d, 2H, bJH
= 9.2 Hz, Hb ), 7.35–6.79 (m, 40H, Harom),
b
ꢀ
ꢀ
ꢀ
O-(2,3,4,6-Tetra-O-benzyl-b-D-
glucopranosyl)trichloroacetimidate (13)
–H
a
b
6.52 (d, 1H, JH1–H2 = 3.6 Hz, H-1a), 5.73 (d, 1H, JH1–H2 = 7.7 Hz,
H-1b), 4.85–4.44 (m, 16H, PhCH), 3.82–3.48 (m, 12H, H-2, H-
3, H-4, H-5, H-6ab, from both anomers). 13C NMR: (75 MHz,
CDCl3): 161.8–159.0, 150.7, 150.1, 144.3–137.2, 131.1–124.8,
116.1, 115.7, 95.2, 93.6, 84.7, 81.3, 80.7, 78.6, 76.4, 75.7, 75.5,
75.3, 75.0, 74.8, 73.8, 73.6, 73.6, 73.5, 68.0, 67.8. ESI-MS: m/z
calcd. for C47H45N3O14Na [M + NH4]+ 889.29; found 889.30.
A solution of 2,3,4,6-tetra-O-benzyl-D-glucopyranose (1 g;
1.8 mmol) and trichloroacetonitrile (16 mL) in dry CH2Cl2
(65 mL) was stirred vigorously with anhydrous K2CO3 (6.5 g)
over night at room temperature under Ar atmosphere. The
mixture was diluted with dry Et2O (30 mL) and filtered through
a layer of sand and silica gel. The silica gel layer was washed
several times with Et2O and the combined filtrates were evapo-
rated to give the trichloroacetimidate in a chromatographically
pure form as a colorless syrup (quantitative) as the b anomer.
The observed NMR data were identical with literature values.18
Optimization of glycosylation conditions (Table 1 + 2)
Conventional heating: The glycosyl donor (0.027 to 0.05 mmol)
was dissolved in dry methanol or dry NMP (1 mL) in a
O r g . B i o m o l . C h e m . , 2 0 0 5 , 3 , 3 9 6 6 – 3 9 7 0
3 9 6 9

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1,2;5,6-Di-O-isopropylidene-3-O-(2,3,4,6-tetra-O-benzyl-a,b-D-
glucopyranosyl)-a-D-glucofuranoside (15a/b)
The observed NMR data were identical with literature values.17
acyl protecting groups on the donor, additional side-reactions are
trapping of products as orthoesters and O-acyl transfer to the
aglycon.
8 Microwave-promoted glycosylations: Fischer glycosylation: (a) C.
Limousin, J. Cleophax, A. Petit, A. Loupy and G. Lukacs, J. Car-
bohydr. Chem., 1997, 16, 327; L. F. Bornaghi and S.-A. Poulsen,
Tetrahedron Lett., 2005, 46, 3485; (b) Microwave-induced Ferrier
rearrangement: B. Shanmugasundaram, A. K. Bose and K. K.
Balasubramanian, Tetrahedron Lett., 2002, 43, 6795; H.-C. Lin,
C.-C. Chang, J.-Y. Chen and C.-H. Lin, Tetrahedron: Asymmetry,
2005, 16, 297; (c) Activation of unreactive oxazoline donors: H.
Mohan, E. Gemma, K. Ruda and S. Oscarson, Synlett., 2003,
1255; (d) Transglycosylations: Y. Yoshimura, H. Shimizu, H. Hinou
and S.-I. Nishimura, Tetrahedron Lett., 2005, 46, 4701; (e) Trans-
glycosylation and glycosylation of 3-methylpentan-1-ol: H. Ohrui,
R. Kato, T. Kodaira, H. Shimizu, K. Akasaka and T. Kitahara,
Biosci., Biotechnol., Biochem., 2005, 5, 1054; (f) Pentenyl orthoesters:
F. Mathew, K. N. Jayaprakash, B. Fraser-Reid, J. Mathew and J.
Scicinski, Tetrahedron Lett., 2003, 44, 9051.
9 R. R. Schmidt, Angew. Chem., Int. Ed. Engl., 1986, 25, 212.
10 R. R. Schmidt and J. Michel, Angew. Chem., 1980, 92, 763.
11 G. Bo¨hm and H. Waldmann, Liebigs Ann., 1996, 613.
12 A. Lubineau and B. Drouillat, J. Carbohydr. Chem., 1997, 16,
1179.
13 D. Crich and V. Dudkin, J. Am. Chem. Soc., 2001, 123, 6819.
14 R. R. Schmidt, J. Michel and M. Roos, Liebigs Ann., 1984,
1343.
Acknowledgements
We thank Dr Lars Petersen for the preparation of 4-nitrophenyl
2ꢀ-fluoro-3ꢀ,5ꢀ-dinitrobenzoate. Financial support fom the Lund-
beck Foundation and the Hede-Nielsen Foundation is acknowl-
edged.
References and notes
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6 C. O. Kappe, Angew. Chem., Int. Ed., 2004, 43, 6250.
7 The most common side reactions in O-glycosylations are hydrol-
ysis of the donor and elimination to form the glycal; with 2-
17 S. Houdier and J. A. Vottero, Carbohydr. Res., 1992, 232, 349.
18 R. R. Schmidt, J. Michel and M. Roos, Liebigs Ann. Chem., 1984,
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3 9 7 0
O r g . B i o m o l . C h e m . , 2 0 0 5 , 3 , 3 9 6 6 – 3 9 7 0