Preparation of methyl 6-O-p-nitrobenzoyl-β-D-glucoside

Article abstract of DOI:10.1080/07328300802071245

Methyl 6-O-p-nitrobenzoyl-β-d-glucoside was synthesized by reacting methyl 4,6-O-p-nitrobenzylidine-β-d-glucoside with N-bromosuccinimide (NBS). First, methyl β-d-glucoside was converted into methyl 4,6-O-p-nitrobenzylidine-β-d-glucoside with p-nitrobenza

Full text of DOI:10.1080/07328300802071245

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Preparation of Methyl
6‐O‐p‐Nitrobenzoyl‐β‐d‐Glucoside
Özlem Akpınar a & Michael H. Penner b
a Department of Food Engineering , Gaziosmanpaşa University , Taşlıçiftlik ,
Tokat
b Department of Food Science and Technology , Oregon State University ,
Corvallis , OR , USA
Published online: 12 Jun 2008.
To cite this article: Özlem Akpınar & Michael H. Penner (2008) Preparation of Methyl
6‐O‐p‐Nitrobenzoyl‐β‐d‐Glucoside, Journal of Carbohydrate Chemistry, 27:3, 188-199, DOI:
10.1080/07328300802071245
To link to this article: http://dx.doi.org/10.1080/07328300802071245
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Journal of Carbohydrate Chemistry, 27:188–199, 2008
Copyright # Taylor & Francis Group, LLC
ISSN: 0732-8303 print 1532-2327 online
DOI: 10.1080/07328300802071245
Preparation of Methyl 6-O-p-
Nitrobenzoyl-b-D-Glucoside
1
2
¨
Ozlem Akpınar and Michael H. Penner
1Department of Food Engineering, Gaziosmanpas¸a University, Tas¸lıc¸iftlik, Tokat
2Department of Food Science and Technology, Oregon State University, Corvallis,
OR, USA
Methyl 6-O-p-nitrobenzoyl-b-D-glucoside was synthesized by reacting methyl 4,6-
O-p-nitrobenzylidine-b-D-glucoside with N-bromosuccinimide (NBS). First, methyl
b-D-glucoside was converted into methyl 4,6-O-p-nitrobenzylidine-b-D-glucoside
with p-nitrobenzaldehyde. Later, methyl 4,6-O-p-nitrobenzylidine-b-D-glucoside
was opened oxidatively with NBS to give methyl 6-O-p-nitrobenzoyl-b-D-glucoside.
Keywords Nonreducing end, NBS, Cyclic acetals, Oxidation, Nitrobenzoyl
INTRODUCTION
Cyclic acetals have been used in carbohydrate chemistry for protecting groups.
The main advantage of cyclic acetals is that they can block a pair of diols in one
step.[1] Generally an aldehyde and a sugar are mixed, either directly (benzal-
dehyde) or in solution by using a solvent (dioxane) in the presence of catalyst.
The catalyst can be either soluble acid (sulfuric acid, p-toluonesulfonic acid) or
an insoluble one (amberlyst resins).[2] Treatment of unprotected hexopyronoside
with aldehyde in the presence of catalytic acid gives selectively protected 4,6-O-
benzylide acetals since the least hindered hydroxyl group (the primary at C-6) is
more reactive than other hydroxyl groups, which leads to the formation of 4,6-
O-benzylide acetals.[3] Another alternative is p-nitrobenzylidene acetal (Fig. 1).
Benzylidene acetals can be opened selectively by acid-catalyzed reduction to
yield either 4-O or 6-O benzyl ether monohydroxyl derivatives. The selectivity of
the reduction is determined by the reducing agents (metal hydride), acid cata-
lysts, solvents, and steric effects.[3] Reduction of 4,6-O-benzylidene acetals with
lithium aluminium hydride-aluminium chloride gives 4-O-benzyl ethers. Lewis
acids like aluminium chloride coordinate with the less hindered 6-O to give a
Received December 4, 2008; accepted March 20, 2008.
¨
Address correspondence to Ozlem Akpınar, Department of Food Engineering, Gazios-
manpas¸a University, Tas¸lıc¸iftlik 60100, Tokat. E-mail: oakpinar@gop.edu.tr
188

Nonreducing End Modification
189
Figure 1: A: 4,6-O-Benzylide acetals. B: 4,6-O-p-Nitrobenzylidine acetals.
reactive complex that is reduced by lithium aluminium hydride to produce 4-O-
benzyl derivatives. Proton-catalyzed reduction (sodium cyano borohydride-
ethereal hydrogen chloride) gives 6-O-benzyl derivatives. This is because the
4-O oxygen is more basic than the 6-O oxygen and the proton is small enough to
reach the 4-O oxygen; thus, protonation of 4-O oxygen produces an intermediate
that reacts with sodium cyanoborohydride to give the 6-O benzyl derivative.[4]
However, p-nitrobenzylidene acetals are more resistant to acid-catalyzed
reduction than are simple benzylidene acetals. The nitro substituents on the
aromatic rings make them less reactive toward electrophilic attack. Electrons
can flow from the benzene ring to the substituent, thus leaving a positive
charge in the ring, destabilizing carbocation intermediates and thus making
the ring less susceptible to the electrophilic attack.[5] Another way to open ben-
zylidene acetals is by oxidative ring opening. Benzylidene acetals can be
oxidized with NBS (N-bromosuccinimide) in the presence of water. The mech-
anism of this reaction has been studied extensively for the synthesis of 4-O-
benzoyl-6-bromo-6-deoxy-a-D-glucopyranoside.[6–9] The purpose of this study
was to explore possible approaches for the opening 4,6-p-nitrobenzylidene
acetals with NBS oxidation. This reaction has not been tried for regioselective
ring opening for 4,6-p-nitrobenzylidene acetals in carbohydrate chemistry.
MATERIALS AND METHODS
General Method
All organic solutions were dried with anhydrous Na2SO4. The 1H NMR spectra
recorded at 400 MHz with a Bruker AM 400 spectrometer using tetramethylsilane
as an internal standard. Assignments were confirmed by 2D-COSY, HMBC, and
HSQC experiments. Spots on TLC were detected by exposure to UV light and by
spraying with p-anisaldehyde-sulfuric acid visualizing reagent.
Methyl 4,6-O-p-nitrobenzylidine-b-D-glucoside
Methyl-b-glucoside (2.04 g), p-nitrobenzaldehyde (1.46 g), and concentrated
sulfuric acid (0.8 mL) in p-dioxane (20 mL) were stirred together at rt for 2 h.

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O. Akpınar and M. H. Penner
190
The solution was then diluted with dichloromethane (50 mL), neutralized with
solid sodium carbonate, filtered, and concentrated under reduced pressure. The
concentrate was dissolved in dichloromethane, washed with water, and then
dried. The solution was evaporated and crystallized from ethanol as a white
crystal (0.9635 g). 1H (Me2SO-d6) data: d 4.22 (s, 1H, J1,2 ¼ 7.7 Hz H-1), d 4.23
(dd, 1H, J6a,6b ¼ 10.3 Hz, J6a,5 ¼ 3.88 Hz H-6a), d 3.73 (dd, 1H, J6a,6b ¼ 10.3 Hz
H-6b), d 3.43–3.41 (m, 3H, H-3,4,5), d 3.05 (dd, 1H, J1,2 ¼ 7.7 Hz, J2.3 ¼ 13.1 Hz
H-2), d 5.37 (OH), d 5.35 (OH), d 3.39 (s, 3H-OCH3), d 5.73 (s PhCH), d 8.24 (d
Jmeta,ortho ¼ 8.82 Hmeta), d 7.71 (d Jmeta,ortho ¼ 8.82 Hortho). 13C: d 106 (C-1), d
74.7 (C-2), d 76.2 (C-3), d 82.6 (C-4), d 67.5 (C-5), d 70.1 (C-6), d 58.5 (OCH3), d
101 (PhC), d 130 (Ortho), d 125 (Meta).
Methyl 6-O-p-nitrobenzoyl-b-D-glucoside
Methyl 4,6-O-p-nitrobenzylidine-b-D-glucoside (30 mg) and NBS (214 mg) in
carbon tetrachloride containing 2 equiv of water (2 equiv) was refluxed for 2 h.
The solution was filtered and concentrated under diminished pressure. The con-
centrate was dissolved in ethyl acetate and washed with saturated NaHCO3 and
cold water, dried with Na2SO4, and evaporated. The target compound was
purified by silica gel chromatography (70–230 mesh, Sigma Chemical Co.,
St. Louis, MO), using ethyl acetate:ethanol:water (30:9.5:0.5). Column fractions
were monitored by thin layer chromatography using silica plates (LK6DF254
,
Whatman Inc., Clifton, NJ) with the same mobile phase used for column chrom-
atography. The compound of interest was identified on TLC plate by exposing to
UV light (phenyl group) and by spraying with p-anisaldehyde-sulfuric acid
visualizing reagent (carbohydrate). Fractions containing the target compound
were pooled, evaporated to dryness under reduced pressure, and then crystal-
lized from ethanol to yield methyl 6-O-p-nitrobenzoyl-b-D-glucoside (6.75 mg).
1H (Me2SO-d6) data: d 4.11 (d, 1H, J1,2 ¼ 7.7 Hz H-1), d 3.24–3.15 (m, 2H,
H-2,3), d 2.985 (1H H-4), d 3.50 (1H, H-5), d 4.37 (dd, 1H, J6a,6b ¼ 11.7,
J6a,5 ¼ 6.1 Hz H-6a), d 4.57 (dd, 1H, J6a,6b ¼ 11.7, J6b,5 ¼ 2.10 Hz H-6b), d
3.29d (s, 3H-OCH3), d 8.36 (d Hmeta), d 8.17 (d Hortho). 13C: d 104 (C-1), d 70.0
(C-2), d 76 (C-3), d 72.2 (C-4, C-5), d 60.1 (C-6), d 55.3 (OCH3), d 126 (Ortho), d
132 (Meta).
RESULTS AND DISCUSSIONS
Methyl 4,6-O-p-nitrobenzylidine-b-D-glucoside was prepared by selectively
protecting the hydroxyl groups at the C-4 and C-6 position with p-nitro-
benzaldehyde according to the method described by Collins and Oparaeche.[10]
Methyl b-D-glucoside was directly mixed with p-nitrobenzaldehyde in the
presence of an acid catalyst to give methyl 4,6-O-p-nitrobenzylidine-b-D-glu-
coside (Fig. 2). The 1H NMR spectra of this compound showed signals at

Nonreducing End Modification
191
Figure 2: Oxidation of methyl 4,6-O-p-nitrobenzylidine-b-D-glucoside with NBS. LN1: Methyl
4,6-O-p-nitrobenzylidine-b-D-glucoside. LN2: Reaction mixture. LN3: Methyl 6-O-p-
nitrobenzoyl-b-D-glucoside. LN4: Same as LN3.
d 8.24 (d Jmeta,ortho ¼ 8.82 Hz Hmeta) and d 7.71 (d Jmeta,ortho ¼ 8.82 Hz
Hortho) characteristic of the aromatic ring and at d 5.73 (s PhCH)
resulting from the proton belonging to the p-nitrobenzylidene (Fig. 1).
These assignments were confirmed with 2D-COSY, HMBC, and HSQC
experiments.
The last reaction was the regioselective opening of methyl 4,6-O-p-nitro-
benzylidine-b-D-glucoside with NBS (N-bromosuccinimide). When methyl
4,6-O-p-nitrobenzylidine-b-D-glucoside was oxidized with NBS (N-bromosucci-
nimide) in the presence of water, the resulting reaction mixture gave three
different products on TLC plate (Fig. 2). The first one was unreacted starting
material, the second one was an unknown side reaction product, and the
third one was methyl 6-O-p-nitrobenzoyl-b-D-glucoside, confirmed by NMR.
The 1H NMR spectra of this compound showed that the signal at d 5.73

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O. Akpınar and M. H. Penner
192
(s PhCH) arising from the proton belong to the p-nitrobenzylidene group
disappeared. The lowest field sugar proton resonance at d 4.57 (dd, 1H,
J6a,6b ¼ 11.7 Hz, J6b,5 ¼ 2.10 Hz H-6b) indicated the p-nitrobenzoate group at
position 6 of the glucose ring (Fig. 1). The other 2D-COSY, HMBC, and
HSQC experiments supported this conclusion.
As shown in the Figure 3, free radical bromination of the benzylidene acetal
carbon atoms gives a bromo derivative A, which generates stabilized carbo-
cation B. The carbocation intermediate is attacked by water to form very
reactive orthoacetal C, which may undergo ring opening by either of two
paths depending on the conditions used. Path “1” leads to the formation of
6-O-NO2Bz ester, whereas path “2” gives the 4-O-NO2Bz ester.
Although the origin of the regioselectivity of the ring opening has not been
fully explained, steric factors may affect this reaction. The nitro group plays an
important role in this reaction, since the selectivity is somewhat different from
that of other O-benzylidene acetals. Since the formation 4-O oxygen is more
basic than 6-O, enhancing the acidity of the reaction solution is expected to
increase the percentage of 4-O-Bz ester. However, the dominant product is
6-O-Bz ester. In addition, excess acid may lead to a general glycoside hydrolysis
reaction.[1]
Figure 3: Opening of benzylidene acetals selectively by NBS oxidation.

Nonreducing End Modification
193
The synthetic method described here can be used as a general method for
the modification of the nonreducing terminus of oligosaccharides. In this
method the anomeric carbon needs to be protected due to its high reactivity.
This was accomplished by blocking with a methyl group due to its stability
under various reaction conditions. Protection of 40-60-positions of methyl oligo-
saccharides with p-nitrobenzylidene acetals followed by regioselective ring
opening gives nonreducing end-modified oligosaccharides. This method is
simple because it does not require many protection and deprotection steps.
Such modified oligosaccharides might be used for testing of the chain end-
specificity of exoenzymes. The nitro group on the nonreducing end of the sub-
strate will have another advantage. Since the nonreducing end of the substrate
will be derivatized to give a p-nitro group, the p-nitro group can then be
reduced with catalytic hydrogenation under hydrogen atmosphere in the
presence of palladium on activated carbon, to give the nonreducing end of
the substrate a p-aminophenyl group. This compound can be immobilized to
the insoluble support by using CnBr- or NHS-activated agarose. Thus,
modified oligosaccharides can be used as affinity ligands for the separation of
exoenzymes.
SUPPORTING INFORMATION
1. NMR data for Methyl b-D-glucoside
1H (Me2SO-d6) data: d 4.02 (d, 1H, J1,2 ¼ 7.62 Hz H-1), d 3.66 (dd, 1H.
J ¼ 10.1 Hz, J ¼ 4.27 Hz), d 3.43 (1H, J ¼ 10.3 Hz), d 3.12–2.90 (m 4H), d
4.99 (OH), d 4.88 (OH), d 4.86 (OH), d 4.47 (OH), d 3.37 (s, 3H-OCH3).
(Figures S1–S9).
2. NMR data for Methyl 4-6-O-p-nitrobenzylidine-b-D-glucoside
1H (Me2SO-d6) data: d 4.22 (s, 1H, J1,2 ¼ 7.7 Hz H-1), d 4.23 (dd, 1H,
J6a,6b ¼ 10.3 Hz, J6a,5 ¼ 3.88 Hz H-6a), d 3.73 (dd, 1H, J6a,6b ¼ 10.3 Hz H-6b),
d 3.43–3.41 (m 3H, H-3,4,5), d 3.05 (dd, 1H, J1,2 ¼ 7.7 Hz, J2.3 ¼ 13.1 Hz
H-2), d 5.37 (OH), d 5.35 (OH), d 3.39 (s, 3H-OCH3), d 5.73 (s PhCH), d 8.24
(d Jmeta,ortho ¼ 8.82 Hmeta), d 7.71 (d Jmeta,ortho ¼ 8.82 Hortho). 13C: d 106 (C-
1), d 74.7 (C-2), d 76.2 (C-3), d 82.6 (C-4), d 67.5 (C-5), d 70.1 (C-6), d 58.5
(OCH3), d 101 (PhC), d 130 (Ortho), d 125 (Meta).
3. NMR data for Methyl 6-O-p-nitrobenzoyl-b-D-glucoside
1H (Me2SO-d6) data: d 4.11 (d, 1H, J1,2 ¼ 7.7 Hz H-1), d 3.24–3.15 (m, 2H,
H-2,3), d 2.985 (1H, H-4), d 3.50 (1H, H-5), d 4.37 (dd, 1H, J6a,6b ¼ 11.7,
J6a,5 ¼ 6.1 Hz H-6a), d 4.57 (dd, 1H, J6a,6b ¼ 11.7, J6b,5 ¼ 2.10 Hz H-6b), d
3.29d (s, 3H-OCH3), d 8.36 (d Hmeta), d 8.17 (d Hortho). 13C: d 104 (C-1), d
70.0 (C-2), d 76 (C-3), d 72.2 (C-4, C-5), d 60.1 (C-6), d 55.3 (OCH3), d 126
(Ortho), d 132 (Meta).

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194
Figure S1: H NMR spectrum for methyl-b-D-glucoside (Me2SO-d6).
Figure S2: 1H NMR spectrum data for Methyl 4-6-O-p-nitrobenzylidine-b-D-glucoside (Me2SO-
d6).

Nonreducing End Modification
Figure S3: 2D COSY spectrum for methyl 4-6-O-p-nitrobenzylidine-b-D-glucoside (Me2SO-d6).
Figure S4: HSQC spectrum for methyl 4-6-O-p-nitrobenzylidine-b-D-glucoside (Me2SO-d6).
195

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196
Figure S5: HMBC spectrum for methyl 4-6-O-p-nitrobenzylidine-b-D-glucoside (Me2SO-d6).
Figure S6: 1H NMR spectrum for Methyl 6-O-p-nitrobenzoylidine-b-D-glucoside (Me2SO-d6).

Nonreducing End Modification
197
Figure S7: 2D COSY spectrum for methyl 6-O-p-nitrobenzoyl-b-D-glucoside (Me2SO-d6).
Figure S8: HSQC spectrum for methyl 6-O-p-nitrobenzoyl-b-D-glucoside (Me2SO-d6).

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O. Akpınar and M. H. Penner
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Figure S9: HMBC spectrum for methyl 6-O-p-nitrobenzoyl-b-D-glucoside (Me2SO-d6).
ACKNOWLEDGEMENT
This work was done at the Department of Food Science and Technology, Oregon
State University, Corvallis, OR, USA.
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[6] Hanessian, S. Reaction of O-benzylidene sugars with N-bromosuccinimide.
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[7] Hanessian, S.; Plessas, N.R.J. Reaction of O-benzylidene sugars with N-bromosuc-
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[8] Hanessian, S.; Plessas, N.R. Reaction of O-benzylidene sugars with N-bromosucci-
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[9] Hanessian, S. 6-Bromo-6-deoxy hexose derivatives by ring-opening of benzylidene
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