Evaluation of steric effects on the exocyclic conformations of 6-C-methyl-substituted 2-acetamido-2-deoxy-β-D-glucopyranosides

Article abstract of DOI:10.1016/S0008-6215(01)00295-6

Introduction of a stereodefined methyl group at the C-6 position of N-acetylglucosamine mono- and disaccharides creates a strong and predictable orientational bias on the geminal C-6 hydroxyl in solution, as determined by ¹H-¹H and <

Full text of DOI:10.1016/S0008-6215(01)00295-6

Carbohydrate Research 337 (2002) 83–86
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Evaluation of steric effects on the exocyclic conformations of
6-C-methyl-substituted 2-acetamido-2-deoxy-b- -glucopyranosides
D
Jihane Achkar, Isabel Sanchez-Larraza, Alexander Wei*
Department of Chemistry, Purdue Uni6ersity, 1393 Brown Building, West Lafayette, IN 47907-1393, USA
Received 1 October 2001; accepted 31 October 2001
Abstract
Introduction of a stereodefined methyl group at the C-6 position of N-acetylglucosamine mono- and disaccharides creates a
1
strong and predictable orientational bias on the geminal C-6 hydroxyl in solution, as determined by H–1H and 13C–1H NMR
coupling constants. The conformational directing effect is more pronounced in the disaccharides because of the greater steric
demand imposed by the neighboring glycosidic unit. © 2002 Elsevier Science Ltd. All rights reserved.
Keywords: Chitin; Glucosamine; Hydroxymethyl; Conformational analysis; NMR spectroscopy
The hydroxymethyl group plays a key role in the
structural and chemical biology of pyranoside carbohy-
drates whose interactions with receptor proteins and
other carbohydrate species are critical for cell–cell
recognition and other biological functions.1,2 The hy-
droxymethyl C-5–C-6 bond is conformationally mobile
and does not exhibit a strong preference for a single
staggered conformation.3 However, one or more ro-
tamers can be destabilized by introducing a small but
sterically demanding unit such as a methyl group at the
6R- or 6S-position. It is known that 1,3-diaxiallike
Me···OH interactions increase torsional strain energy
by over 2 kcal/mol;4 therefore, the exocyclic C-5–C-6
bond is expected to demonstrate a preference for stag-
gered conformers that avoid such interactions (Fig. 1).
Sterically driven conformational bias has been demon-
strated on 6-C-substituted gluco- and galactopyran-
osides,5,6 and has been used to probe the effect of
conformation on the recognition or enzymatic hydroly-
sis of 1,6-linked disaccharides.7–9
acetylglucosaminopyranosides (b-GlcNAc) and their
corresponding 1,4-linked disaccharides. We demon-
strate that the stereodefined methyl group at C-6 intro-
duces a strong and predictable conformational bias on
the C-5–C-6 bond, with a subsequent directing effect
on the C-6 hydroxyl groups. 6-C-Substituted glu-
cosamines are expected to be useful for investigating
conformational effects in the protein–carbohydrate and
carbohydrate–carbohydrate interactions of chitin10 and
the glycosaminoglycans.11,12
6-C-Substituted b-GlcNAc monosaccharides were
synthesized according to Scheme 1. All new compounds
were fully characterized by NMR spectroscopy and
elemental analysis or mass spectrometry. Protected
monosaccharide derivative 1 was prepared in multi-
gram quantities from glucosamine hydrochloride using
literature procedures.13–15 Conditions for the reductive
cleavage of the 4,6-O-p-methoxybenzylidene derivative
of 1 to the corresponding free O-6 hydroxyl group16
were found to be incompatible with the anomeric allyl
protecting group, but compound 2 could be obtained in
good yield by regioselectively protecting the primary
alcohol as an intermediate tert-butyldimethylsilyl ether.
Aldehyde 3 was obtained by Swern oxidation17 and was
readily reduced by NaBD4 to yield 6-C-monodeuter-
ated glucosamine derivative 4 as a 3:1 mixture of
Herein we report the stereoselective synthesis and
conformational
analysis
of
6-C-methyl
b-N-
* Corresponding author. Tel.: +1-765-4945257; fax: +1-
765-4940239.
E-mail address: alexwei@purdue.edu (A. Wei).
0008-6215/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(01)00295-6

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J. Achkar et al. / Carbohydrate Research 337 (2002) 83–86
Scheme 1. Reagents and conditions: (a) (i) TBSCl, Et3N,
imidazole, CH2Cl2–THF (96%); (ii) dihydropyran, PPTS; (iii)
(n-Bu)4NF, THF (76% over two steps); (b) (COCl)2, DMSO,
CH2Cl2, −78 °C; Et3N, 0 °C (64%); (c) (i) NaBD4, CH2Cl2–
MeOH, 0 °C; (ii) p-TsOH, MeOH (70% isolated yield of 4
over two steps); (d) (i) AlMe3 (5 equiv), CuCN (1 equiv),
THF, −45 °C to rt; (ii) p-TsOH, MeOH (50% isolated yield
of 5 over two steps); (e) (i) AlMe3 (5 equiv), CuCN (1 equiv),
THF, −45 °C to rt; (ii) (COCl)2, DMSO, CH2Cl2, −78 °C;
Et3N, 0 °C; (iii) i-Bu2AlH, ZnCl2, THF, −78 °C; (iv) p-
TsOH, MeOH (37% isolated yield of 6 over four steps); (f) (i)
ethylenediamine, n-BuOH, 100 °C; (ii) Ac2O, C5H5N, 0 °C to
rt; (iii) NaOMe, MeOH–CH2Cl2, 0 °C to rt (87% over three
Fig. 1. Staggered conformations of the exocyclic C-5–C-6
bond in 6-C-methyl-substituted pyranosides.
steps).
All=allyl,
Phth=phthalimido,
TBS=tert-
butyldimethylsilyl, THP=tetrahydropyranyl.
diastereomers, providing reference compounds for the
NMR conformational studies.† Assignment of 6R- and
6S-stereochemistry was achieved via vicinal coupling
constant analysis of the corresponding 4,6-O-isopropy-
lidene or p-methoxybenzylidene ketal. Aldehyde 3 was
also subjected to chemoselective methylation conditions
but was found to be a surprisingly unreactive elec-
trophile. After extensive experimentation, it was deter-
mined that methylation of 3 could be achieved in good
yield with 6:1 6S:6R stereoselectivity using AlMe3 with
CuCN as an additive. It is noteworthy that the protect-
ing group at O-4 had a significant influence on the
efficiency and stereochemical outcome of the methyla-
tion by AlMe3. Replacing the tetrahydropyranyl (THP)
group with a benzyl ether lowered the stereoselectivity,
whereas a 2-methoxyethoxymethyl (MEM) ether re-
duced reactivity. Removal of the THP group enabled
separation of the diastereomers, affording 6-C-methyl-
substituted 5 and 6 in 58% combined yield after two
steps. Additional 6 could be obtained by Swern
oxidation17 of the THP-protected 6-C-methyl adduct to
the corresponding ketone, followed by reduction with
i-Bu2AlH in the presence of ZnCl2 (6:1 6R:6S ratio).
Conversion of the phthalimide group at C-2 into an
acetamide followed by global deprotection, yielded the
desired C-6-substituted b-GlcNAc monosaccharides 7–
9.
Scheme 2. Reagents and conditions: (a) (i) p-
,
MeO(C6H4)CH(OMe)2, camphorsulfonic acid, 4 A mol sieves,
,
toluene, 90 °C; (ii) NaBH3CN, HCl, 4 A mol sieves, THF–
Et2O, −30 °C (35–76% over two steps); (b) 2-deoxy-2-ph-
thalimido-3,4,6-O-triacetylglucosyl trichloroacetimidate (2
,
equiv), TMSOTf (0.2 equiv), 4 A mol sieves, CH2Cl2, −30 °C
(78–98%); (c) (i) DDQ, t-BuOH, pH 7 buffer, THF, 0 °C; (ii)
ethylenediamine, n-BuOH, 100 °C; (iii) Ac2O, C5H5N, 0 °C to
rt; (iv) NaOMe, MeOH–CH2Cl2, 0 °C to rt (64–76% over
four steps). All=allyl, Phth=phthalimido, PMB=p-
methoxybenzyl.
Disaccharides containing C-6-substituted b-GlcNAc
were also prepared to determine the relative influence of
a neighboring glycosidic unit on exocyclic conformation
(Scheme 2). Monosaccharides 4–6 were protected as
6-O-p-methoxybenzyl (PMB) ethers 10–12 via reduc-
tive cleavage of the corresponding 4,6-O-p-methoxy-
benzylidene acetals under acidic conditions. Glyco-
sylation of O-4 with 2-deoxy-2-phthalimido-3,4,6-O-
triacetylglucopyranose activated as a Schmidt trichloro-
† Diastereomeric monodeuteration greatly simplifies confor-
mational analysis of the C-5–C-6 bond by reducing the
H-5–H-6 coupling to a two-spin system. The diastereotopic
6R- and 6S-protons can thus be analyzed simultaneously.

J. Achkar et al. / Carbohydrate Research 337 (2002) 83–86
85
Table 1
1H–1H and 13C–1H coupling constants of 6-C-substituted glucopyranosides a
3JC-7,H-5
C-5–C-6 conformational preferences c
b
b
b
b-GlcNAc derivative
3J5,6
2JC-6,H-5
d
(6S/6R)-C-d Monosaccharide (7)
(6S/6R)-C-d Disaccharide (16)
(6S)-C-Methyl monosaccharide (8)
(6S)-C-Methyl disaccharide (17)
(6R)-C-Methyl monosaccharide (9)
(6R)-C-Methyl disaccharide (18)
6.2/2.6
4.9/2.0
1.8
1.5
3.9
–
–
2.9
2.3
4.2
4.5
–
–
1.6
1.2
3.4
3.8
gt#gg\tg
d
gt#gg\tg
e
gg\tg, gt
gg\tg, gt
e
f
gt\tg\gg
gtꢀtg\gg
f
2.4
a 1H NMR spectra were obtained using a 600 MHz Varian spectrometer in MeOH-d4 at 298 K. Coupled 13C NMR spectra were
obtained using a 500 MHz Bru¨ker spectrometer in methanol-d4 at 298 K.
3
2,3
b In Hz (90.25 Hz for J5,6, 90.3 Hz for
J
).
C,H
c Order of conformational preferences was established by J5,6 coupling constant analysis and correlated with JC,H and JC,H
3
2
3
values.
d Theoretical 3J5,6R/3J5,6S values for the staggered conformers of 1,2-dialkoxypropane are: 10.7/3.1 for gt; 5.0/10.7 for tg; 0.9/2.8
for gg.3,22,23
e Theoretical J5,6 values for the staggered conformers of 2,3-(S,S)-dialkoxybutane (calculated using empirical parameters in
3
Ref. 19) are: 9.2 for gt; 3.9 for tg; 0.7 for gg.
3
f Theoretical J5,6 values for the staggered conformers of 2,3-(S,R)-dialkoxybutane are: 2.3 for gt; 9.2 for tg; 2.3 for gg.
3
acetimidate18 produced 1,4-b-linked disaccharides 13–
15, and was followed by global deprotection to give the
desired 6-C-substituted disaccharides 16–18 in high
overall yields.
(6S)-C-Methyl b-GlcNAc 8 at 298 K has a small J5,6
value of 1.8 Hz, indicating a strong preference for the
gg conformation (3J5,6(theor.) 0.7 Hz) relative to the tg
or gt conformers (3J5,6(theor.) 3.9 and 9.2 Hz, respec-
tively). This conformational assignment is supported by
The conformational preferences of the C-5–C-6
bonds of 7–9 were evaluated as a function of C-6
substitution using vicinal 1H–1H coupling constants
(3J5,6) from nuclear magnetic resonance (NMR) spec-
troscopy, supported by 13C–1H coupling constants
(Table 1).‡ Changes in conformational preference as a
function of C-6 substitution were evaluated to the first
3
a small JC-7,H-5 constant of 1.6 Hz. In comparison,
(6R)-C-methyl b-GlcNAc 9 has a relatively large
2JC-6,H-5 value of 4.2 Hz, which correlates with a deple-
tion of the gg conformer.20,21 Evaluation of the remain-
3
ing two staggered conformations using the J5,6
constant (3.9 Hz) and the appropriately parameterized
Karplus equation indicates that the gt conformer is
strongly favored over the tg conformer (3J5,6(theor.) 2.3
and 9.2 Hz, respectively).
3
degree of approximation by correlating J5,6 coupling
constants with values derived from Karplus equations
parameterized for 1,2-dialkoxypropanes or 2,3-di-
alkoxybutanes.19 Two- and three-bond 13C–1H cou-
Conformational analysis of 16–18 at 298 K in
MeOH-d4 indicates that the C-4 glycoside reinforces the
conformational preference of the exocyclic C-5–C-6
bond (Table 1). In the case of (6S)-C-methyl substi-
tuted 17, the preference for the gg conformation is
increased (3J5,6 1.5 Hz); in the case of (6R)-C-methyl
substituted 18, gt is more strongly favored (3J5,6 2.4
Hz). These observations suggest that the neighboring
glycosidic unit enhances the directing effect of the
6-C-methyl group by increasing steric demand. In-
tramolecular hydrogen bonding, if any, does not appear
to have any significant influence on the exocyclic con-
formation of 16–18 under these conditions. This is in
accord with previous solution conformation studies on
C-glycosides, whose secondary structures are deter-
mined essentially by local steric effects on torsional
strain.24
pling constants (2JC,H
,
3JC,H) were obtained from
coupled 13C spectra and correlated with empirical val-
ues reported by Serianni20 and Murata.21 The JH,H
3
coupling constants in the pyranose rings of 8 and 9
describe stable chair conformations which are essen-
tially unaffected by C-6 methyl substitution.
3J5,6 coupling constant analysis of 6-C-monodeuter-
ated b-GlcNAc 7 indicates an approximately equal
mixture of gt and gg conformations at 298 K in MeOH-
d4, in general agreement with earlier reports on hydrox-
ymethyl conformation (Table 1).3,22,23 Introducing a
stereodefined 6-C-methyl group dramatically changes
the conformational preference of the C-5–C-6 bond.
‡ It is important to note that NMR coupling constant
analyses in solution are derived from time-averaged confor-
mational distributions and cannot precisely measure the per-
centage of individual conformers in a general sense. However,
such analyses are useful for probing relative changes in con-
formational preference.
In conclusion, we have demonstrated that stereose-
lective methylation at C-6 can be used to direct the
conformational preference of the exocyclic O-6 hydrox-

86
J. Achkar et al. / Carbohydrate Research 337 (2002) 83–86
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This work was supported by the American Chemical
Society Petroleum Research Foundation (33341-G4,
36069-AC1), the American Heart Association Midwest
Affiliates (30399Z), the American Cancer Society (IRG-
58-006-41), and the Purdue Research Foundation in the
form of a fellowship to J.A. The authors gratefully
acknowledge Dr Klaas Hallenga and Dr Edwin Rivera
for NMR assistance, and Fabien Boulineau for con-
tributing toward the chemoselective methylation study.
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