Synthesis of a mycobacterium tuberculosis tetra-acylated sulfolipid analogue and characterization of the chiral acyl chains using anisotropic NAD 2D-NMR spectroscopy

Article abstract of DOI:10.1021/jo4012255

Tetra-O-acylated sulfolipids are metabolites found in the cell wall of Mycobacterium tuberculosis, the causative agent of tuberculosis. Their role in pathogenesis remains, however, undefined. Here we describe a novel access to model tetra-O-acylated treha

Full text of DOI:10.1021/jo4012255

Article
pubs.acs.org/joc
Synthesis of a Mycobacterium tuberculosis Tetra-acylated Sulfolipid
Analogue and Characterization of the Chiral Acyl Chains Using
Anisotropic NAD 2D-NMR Spectroscopy
Aurelie Lemetais,†,‡ Yann Bourdreux,†,‡ Philippe Lesot,*,∥,‡ Jonathan Farjon,∥,‡ and Jean-Marie Beau*,†,‡,§
́
́
†Universite
́
Paris-Sud, Institut de Chimie Molec
́
ulaire et des Mater
́
iaux d’Orsay, UMR 8182, Laboratoire de Synthes
̀
e de
Biomolec
́
ules, F-91405 Orsay, France
‡CNRS, F-91405 Orsay, France
§Centre de Recherche de Gif, Institut de Chimie des Substances Naturelles du CNRS, Avenue de la Terrasse, F-91198 Gif-sur-Yvette,
France
∥Universite
́ ́ ́ ́
Paris-Sud, Institut de Chimie Moleculaire et des Materiaux d’Orsay, UMR 8182, Laboratoire de RMN en Milieu Oriente,
F-91405 Orsay, France
S
* Supporting Information
ABSTRACT: Tetra-O-acylated sulfolipids are metabolites
found in the cell wall of Mycobacterium tuberculosis, the
causative agent of tuberculosis. Their role in pathogenesis
remains, however, undefined. Here we describe a novel access
to model tetra-O-acylated trehalose sulfate derivatives having
simple acyl chains. The trehalose core was regioselectively
protected using a tandem procedure with catalytic iron(III)
chloride hexahydrate and further desymmetrized. Model chiral
fatty acids, prepared by a zinc-mediated cross-coupling, were
incorporated into the trehalose core. The enantiomeric excess
of the chiral fatty acids has been measured by natural
abundance deuterium (NAD) 2D-NMR spectroscopy in a
polypeptide based chiral liquid crystal. The synthetic approach
established for the model compounds can easily be developed for the preparation of other analogues and natural sulfolipids.
INTRODUCTION
RESULTS AND DISCUSSION
■
■
Tuberculosis is still responsible for approximately two million
deaths per year throughout the world, and especially in third
world countries.1,2 Sulfolipids I and IV are metabolites found in
the cell wall of Mycobacterium tuberculosis, the bacteria
responsible for this disease (Figure 1). Di-O-acylated sulfolipids
IV (SL-IV)3−6 were recently identified as new antigens able to
control mycobacterium infection,7−10 and tetra-O-acylated
derivatives I (SL-I) are the most abundant sulfolipids found
in the bacteria. These sulfolipids are O-acylated α,α-D-trehalose
derivatives11 bearing a sulfate moiety. More precisely, sulfolipid
I are 2′-O-sulfate trehalose cores, functionalized by two
hydroxyphthioceranoyl groups at the C-6 and C-6′ positions
and esterified by a palmitic or stearic acid at the C-2 position
and by a phthioceranoic acid at C-3. To the best of our
knowledge, only one access to a tetra-O-acylated sulfolipid
analogue starting from D-glucose was reported by Leigh and
Bertozzi.12 Herein we report our results for the synthesis of
tetra-O-acylated sulfolipid 1 related to SL-I starting from C2-
symmetric α,α-D-trehalose13 2. Our strategy is based on a
tandem regioselective protection of α,α-D-trehalose followed by
desymmetrization of this disaccharide.
Synthesis of the Tetra-acylated Sulfolipid 1 and 16.
We recently reported a tandem regioselective protection of
carbohydrates with catalytic Cu(OTf)2 or FeCl3·6H2O as Lewis
acids.14,15 The FeCl3·6H2O-catalyzed tandem procedure
applied to persilylated trehalose 3, obtained after treatment of
2 with an excess of TMSCl in pyridine, provided symmetrical
compound 4 in good yield,15 in which the Lewis acid catalyzed
two acetalizations and two regioselective reductive ether-
ifications (Scheme 1). Nonsymmetrical mono-O-palmitoylated
trehalose 7′, envisaged in a first approach as a key synthetic
intermediate to the tetra-O-acylated sulfolipid, was provided
directly by terminating this tandem procedure with a mono-O-
acylation step with palmitoyl chloride. This gave however a
complex reaction mixture from which mono-O-palmitoylated
derivative 7′ was isolated in a 27% yield, separated from diol 4
(17% yield). To overcome the modest efficiency of this one-pot
process, we chose the easy desymmetrization of diol 4 by
mono-O-silylation with TBSOTf at a low temperature to give 5
Received: June 6, 2013
© XXXX American Chemical Society
A
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Scheme 2. Syntheses of Enantiopure 10a and 10b via a Zinc-
a
Catalyzed Cross-Coupling
a
Reagents and conditions: (a) C8H17MgCl or C16H33MgCl, 1.5 equiv;
ZnCl2, 15 mol %; THF; 0 °C to rt; overnight; 9a: 98%, 9b: 81%; (b)
TFA, CH2Cl2; rt; 5 h; quantitative.
step with DCC and DMAP provided quantitatively bis-O-
palmitate derivative 11a (Scheme 3). The primary C-6 and C-
6′ positions were selectively deprotected by regioselective
reductive opening of the two 4,6-O-benzylidene acetals using
PhBCl2 in the presence of Et3SiH,21 affording diol 12a in a 93%
yield. Other known reductive conditions leading to the primary
alcohol, such as BH3·THF in the presence of Cu(OTf)2, AlCl3,
or TMSOTf as Lewis acids, were tested on the di-O-acetate
derivative of 4 as a model compound. These conditions led to
complex mixtures, and only the use of PhBCl2/Et3SiH was
effective. Acylation of diol 12a with excess palmitic acid gave
tetra-O-palmitate derivative 13a in a high yield (95%).
Desilylation and sulfation by the SO3·pyridine complex
provided sulfate 15a in 87% yield (two steps). Final
deprotection by hydrogenolysis of the three O-benzyl groups
of 15a furnished model tetra-O-acylated sulfolipids 16 in 86%
yields (Scheme 3). This optimized sequence of transformations
was subsequently applied to acids 10a and 10b, with an
expectation of a lower reactivity of these α-methylated
carboxylic acids. We indeed noticed a lower efficiency with
each step of the synthesis. Esterification of 7 with 10b to ester
11b (64% yield), regioselective reductive opening to diol 12b
(71% yield), and bis-esterification in the presence of an excess
of 10a led to 13a (83% yield, Scheme 3). Finally, desilylation
Figure 1. Structures of the major sulfolipids I and IV from
Mycobacterium tuberculosis and model sulfolipid 1.
in a 74% yield. During this step 3% of the starting material and
4% of the bis-O-silylated derivative were also isolated. The less
hindered O-benzyl group was then regioselectively removed
under catalytic hydrogen transfer conditions,16 providing diol 6
in 65% yield. Small amounts of side products resulting from bis-
3,3′-de-O-benzylation, mono-3′-de-O-benzylation, and hydro-
genolysis of the 4,6-O-benzylidene acetals were detected by
chromatography. Monoacylation using palmitic acid, DCC, and
DMAP finally led selectively to compound 7 in 85% yield
(Scheme 1).
Several approaches for the iterative syntheses of polydeoxy-
propionates have been developed.17 For the rapid syntheses of
the model enantiopure fatty acids 10a and 10b, we used the
zinc-catalyzed enantiospecific cross-coupling strategy reported
by Breit.18,19 The Grignard reagents were easily coupled in high
yields to methyl (L)-lactate triflate 8, and chiral esters 9a18 and
9b20 obtained were quantitatively converted to carboxylic acids
10a and 10b by treatment with trifluoroacetic acid (Scheme 2).
The synthetic route to a tetra-O-acyled trehalose sulfate
model was first tested with palmitic acid. The first acylation
a
Scheme 1. Preparation of Palmitoyled Trehalose Intermediate 7
a
Reagents and conditions: (a) TMSCl, 10 equiv; pyridine; 0 °C to rt; overnight; 96%; (b) PhCHO, 6 equiv; Et3SiH, 2.2 equiv; FeCl3·6H2O, 5 mol
%; CH2Cl2/CH3CN: 4/1; 0 °C to rt, 3 h; 61%; (c) TBSOTf, 1.1 equiv; 2,6-lutidine, 2 equiv; CH2Cl2, −78 °C, 3.5 h; 74%; (d) HCO2NH4, 20 equiv;
Pd/C, 10%; MeOH; 65 °C, 45 min; 65%; (e) C15H31CO2H, 1.1 equiv; DCC, 1.5 equiv; DMAP, 1.7 equiv; CH2Cl2; rt, 4 h; 85%; (f) step b then
C15H31COCl, 4 equiv; 50 °C overnight; 27%.
B
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a
Scheme 3. Completion of the Synthesis of Model Sulfolipids
a
Reagents and conditions: (a) for 11a: C15H31CO2H, 1.3 equiv; DCC, 1.3 equiv; DMAP, 1.9 equiv; CH2Cl2; rt; 6 h; quantitative; for 11b: 10b, 1.1
equiv; DCC, 1.1 equiv; DMAP, 1.1 equiv; CH2Cl2; rt; 6 h; 64%; (b) PhBCl2, 5 equiv; Et3SiH, 10 equiv; CH2Cl2; −78 to −20 °C; 6.5 h; 12a: 93%;
12b: 71%; (c) for 13a: C15H31CO2H, 3 equiv; DCC, 2.5 equiv; DMAP, 1 equiv; CH2Cl2; rt; overnight; 95%; for 13b: compound 10a, 3.2 equiv;
DCC, 3.2 equiv; DMAP, 3 equiv; CH2Cl2; rt; overnight; 83%; (d) TBAF, 2−3 equiv; THF; rt; 2 h; 14a: 91%; 14b: 85%; (e) [SO3:pyridine], 5 equiv;
pyridine; 90 °C; 2−3 h; then Amberlit IR120 Na; 15a: 96%; 15b: 93%; (f) H2, 1 atm; Pd/C 5%; MeOH/CH2Cl2: 1/1; rt; 2h; 16: 86%; 1: 56%.
1
and sulfation (15b in a 79% yield) and deprotection provided
model sulfolipid 1 in a 56% yield.
to H, routine (9.4 T) or higher field NMR spectrometers,
equipped with classical or cryogenically cooled probes, allow
recording natural abundance deuterium (NAD) NMR spectra
in polypeptide CLC in about 15 h, with a satisfactory S/N ratio,
thus achieving the determination of enantiomeric purity for a
large range of chiral molecules. Combined with 2D-NMR
experiments able to simplify the analysis of complex NAD
spectra,25 this approach was successfully applied for discrim-
inating the R/S isomers of 3-methylhexane23 or the enantio-
isotopomers26 of saturated (prochiral) fatty acid methyl esters
(FAME), such as methyl linoleate or methyl stearate.27,28
Considering its analytical potentials, this approach was used
to evaluate the enantiopurity of 9a and 9b synthesized in
enantioenriched series. Anisotropic NAD 2D-NMR spectra
were recorded in the PBLG/pyridine chiral mesophase
w(PBLG)/w(total) ≈ 23%); this system was shown to be a
highly efficient CLC to discriminate enantiomers of weakly
polar compounds such as FAME.27,28 Although already
described elsewhere, the basics of anisotropic NAD 2D-NMR
spectroscopy, sample preparation, and tube compositions are
given in the Supporting Information. Figure 2 shows a zoom of
the tilted Q-COSY Fz 2D map of (rac)-9a centered on the
signals of the methyl groups recorded at 92.1 MHz equipped
Anisotropic Natural Abundance Deuterium 2D-NMR
Spectroscopy with Esters 9a and 9b. In order to determine
the enantiomeric excess of chiral compounds 9a and 9b, and as
a part of our work led by some of us on the spectral
enantiodiscrimination of chiral compounds coupled with the
analysis of the natural isotopic distribution of deuterium in fatty
acids, we turned to the analytical potential of anisotropic NMR
using polypeptide chiral liquid crystals (CLC) as enantio-
discriminating NMR solvents. Unlike isotropic solvents, all
solute molecules dissolved in a liquid-crystalline media adopt an
averaged orientational ordering. This ordering is generally
different for two enantiomers when a CLC is used; hence it can
be revealed by recording all magnetically active nuclei at a
22
natural abundance level (1H, 13C, H, ...). As the enantio-
discrimination mechanisms mainly depend on molecular-shape
recognition phenomena, no peculiar chemical groups of a chiral
guest are requested to discriminate between enantiomers.23,24
In contrast, the choice of order-sensitive NMR interaction, and
thereby the associated nuclei, is important to optimize the
magnitude of spectral discriminations. Among anisotropic
interactions, the quadrupolar interaction associated to nuclei
with I > 1/2 is the most sensitive one to molecular ordering
differences.23
2
2
with a 5-mm H cryoprobe. The series of NAD 1D subspectra
2
associated to the various H sites of 9a (and 9 b) are reported
As the second isotope of hydrogen, deuterium (I = 1) is
naturally present in all organic molecules and provides an
efficient nuclear spy to reveal chiral discrimination. Although
in the Supporting Information.
For a given 2H site, the presence of two quadrupolar
doublets, denoted ΔνQ(R or S), centered around the same
δ(2H) indicated that the spectral discrimination at this
particular site occurs. These kinds of spectral patterns were
2
the average natural abundance of H nuclei is equal to 1.55 ×
10−2 % (Vienna Standard Mean Ocean Water value) compared
C
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measured on the methyl groups are generally among the
smallest ones. This effect is due to the rotation of the methyl
group around the C−CH3 axis. Consequently, by calculating
the average orientation, the order parameter associated to a
methyl C−D bond appears equal to the order parameter of the
C−C bond times [3cos2(π − 2θm) − 1]/2 = −1/3, where θm is
the so-called magic angle. The same phenomenon occurs for
the three methyl groups of t-Bu, and hence the associated ΔνQ
is reduced again by a factor −1/3, leading to an averaged
quadrupolar splitting of 26 Hz (absolute value).
2
Although other H sites show enantiodiscriminations (C1,
C2a,b), the Me2 site is the most valuable position for a robust
evaluation of the e.e.: (i) the R- and S-signals have baseline
separation up to the peak’s base; (ii) three isotopomers
contribute to the NAD signal, thus increasing the S/N ratio by
a factor of about 3 compared to other sites. Figure 3a and 3b
Figure 2. Expansion of the NAD tilted Q-COSY Fz 2D map of (rac)-
9a (100 mg) recorded at 305 K in PBLG/pyridine (130 mg/365 mg)
showing the methyl groups (Me1, Me2, and Me9). The R/S assignment
is based on the results obtained with an enantioenriched series (see
text). The differences in peak intensity arise from the number of
isotopomers contributing to the NAD signal (9 for Me1, 3 for Me2 and
Me9) and whether the enantiodiscrimination occurs or not (Me2). The
horizontal scale has no spectral meaning. The structure of 9a is shown
above, with the methylene and methyl group numbering used for the
anisotropic NAD-NMR analysis.
Figure 3. Comparison of NAD signals of Me2 (columns extracted
from the 2D maps) of (R/S)-9a and (S)-9a (left panel, a and b) and
(R/S)-9b and (S)-9b (right panel, c and d). Spectra are plotted at the
same vertical scale. The difference of S/N ratios between a/b and c/d
spectra originates from the difference of MW of both solutes and the
mass used (50 mg for 9b).
typically observed on the 2D map of (rac)-9a at the methyl site
2 (see Figure 2), while no differentiation (a single doublet
observed) occurred for the terminal methyl (Me9) and the t-Bu
methyl groups (Me1). The absence of discrimination at these
2
latter H sites originates from different phenomena. For Me9,
the fast rotation of C−D directions in methyl combined with
the complex conformational dynamic of the alkyl chain and the
distance (≈ 10.4 Å) to the stereogenic center C-2 explains why
this group is not a good spy in terms of chiral recognition
mechanisms. In other words, for this specific position and from
an orientational viewpoint, the molecule formally behaves as if
it was of Cs symmetry on average instead of C1 symmetry.27,29
This qualitative interpretation is supported by three other
experimental facts:30 (i) the presence of two 2H doublets at the
CH2 groups 8 and 9 when 9a is dissolved in the CLC, while
four doublets are expected for diastereotopic directions in
methylene groups of a chiral flexible molecule; (ii) the presence
of a single doublet for methylene groups 8 and 9 when the
NAD spectra of 9a is recorded in the achiral liquid crystal (PBG
report the NAD signals of Me2 of 9a obtained in the racemic
series (a) and enantioenriched series (b). As seen a unique H
2
quadrupolar doublet is observed in Figure 3b. The total absence
2
of visible H signals (even on the 2D map) for the minor
enantiomer (R) indicates that the enantiopurity of the mixture
is at least over 98%. Taking into account the strategy to obtain
an enantiopure chiral lipid, the remaining doublets correspond
to the (S)-isomer. As both oriented samples containing (rac)-
9a and (S)-9a were prepared with the same mass composition,
the comparison of both NAD 1D-NMR subspectra allows
assigning the R/S doublet recorded in the racemic mixture.
Anisotropic NMR methodology has been applied to
determine the enantiopurity of 9b (see Scheme 2) using the
same NMR experimental conditions (except temperature: T =
315 K) and close to the sample compositions as those chosen
for analyzing 9a. Compared to 9a, 9b possesses eight additional
2
= 50% PBLG + 50% PBDG); (iii) the same number of H
doublets when the NAD spectrum of 9a in enantiopure series
(see below) is recorded in either the PBLG or PBG oriented
system. For Me1 of t-Bu, the distance to the C-2 remains
reasonable (≈ 4.7 Å) and chiral discrimination might be
expected. However the double rotation of C−D directions
around the C−CH3 and O−C(CH3) axis leads also to a
complex conformational dynamic that is incompatible with
efficient enantiorecognition at these sites.
2
methylene groups (namely an additional 16 inequivalent H
sites), both increasing its MW by ∼45%, and significantly
complicating the analysis of their anisotropic NAD spectra,
except for the methyl groups.31 Not surprisingly, the orienta-
tional behaviors of the t-Bu methyl groups (Me1), the methyl
group bonded to the asymmetric carbon (Me2), and the methyl
group at the terminal position (Me17 for 9b) in both chiral
solutes are very similar in terms of the magnitude of ΔνQ. Only
Finally it could be noted that compared to other 2H sites (see
the Supporting Information), the quadrupolar splittings
D
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1
mp 74−76 °C; [α]27 = +54 (c 0.6, CHCl3); H NMR (CDCl3, 400
2
the NAD signals of methyl group 2 show two H doublets,
D
2
MHz) δ (ppm): 7.50−7.27 (m, 20H, Har), 5.58 (s, 1H, CH−Ph), 5.57
(s, 1H, CH−Ph), 5.22 (d, J1′,2′ = 4.0 Hz, 1H, H1′), 5.05 (d, J1,2 = 3.5
Hz, 1H, H1), 5.01 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.96 (d, J = 11.2 Hz,
1H, CH2−Ph), 4.79 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.76 (d, J = 11.2
Hz, 1H, CH2−Ph), 4.32−4.23 (m, 2H, H6b, H6′b), 4.22−4.13 (m, 2H,
H5′, H5), 3.98 (t, J2′,3′ = J3′,4′ = 9.2 Hz, 1H, H3′), 3.92 (t, J2,3 = J3,4 = 9.5
Hz, 1H, H3), 3.82 (dd, J1,2 = 3.5 Hz, J2,3 = 9.5 Hz, 1H, H2), 3.81−3.68
(m, 4H, H2′, H6′a, H6a, H4′), 3.67 (t, J3,4 = J4,5 = 9.5 Hz, 1H, H4), 0.96
(s, 9H, SiC(CH3)3), 0.12 (s, 6H, Si(CH3)2); 13C NMR (CDCl3, 75
MHz) δ (ppm): 138.7, 138.3, (Cq-Ar), 137.4, (2C, Cq−Ar), 128.9,
128.8, 128.4, 128.2, 128.1, 127.9, 127.8, 126.4, 126.2, 126.0 (20C,
CH−Ar), 101.6 (CH−Ph), 101.2 (CH−Ph), 95.8 (C1), 94.4 (C1′),
82.5, 82.4 (C4, C4′), 78.6 (C3), 78.1 (C3′), 75.2 (CH2−Ph), 74.8
(CH2−Ph), 72.8 (C2), 71.7 (C2′), 69.0, 68.9 (C6, C6′), 63.3, 62.9 (C5,
C5′), 26.0 (SiC(CH3)3), 18.1 (SiC(CH3)3), −4.3, −4.7 (Si(CH3)2).
Anal. Calcd for C46H56O11Si: C, 67.96; H, 6.94. Found: C, 67.99; H,
7.05. ESI HRMS for C46H56O11Si [M+Na]+: Calcd 835.3484, Found
835.3453.
3′-O-Benzyl-4,6;4′,6′-di-O-benzylidene-2′-O-tert-butyldime-
thylsilyl-α,α-D-trehalose (6). A solution of compound 5 (398 mg,
0.490 mmol) in methanol (16 mL) containing HCOONH4 (618 mg,
9.80 mmol, 20 equiv) and 10% Pd/C (50 wt %, 230 mg) was stirred at
reflux for 45 min. After cooling to room temperature, the reaction
mixture was filtered through Celite and concentrated under reduced
pressure. The residue was purified by flash chromatography on silica
gel (cyclohexane/ethyl acetate: 90/10 then 80/20 then 75/25 then
70/30 then 60/40) to give the expected compound 6 (230 mg, 65%).
White solid; mp 101−103 °C; [α]27D = +48 (c 0.6, CHCl3); 1H NMR
(CDCl3, 300 MHz) δ (ppm): 7.51−7.28 (m, 15H, Har), 5.57 (s, 1H,
CH−Ph), 5.53 (s, 1H, CH−Ph), 5.22 (d, J1,2 = 3.9 Hz, 1H, H1), 5.08
(d, J1′,2′ = 3.9 Hz, 1H, H1′), 4.95 (d, J = 11.1 Hz, 1H, CH2−Ph), 4.77
(d, J = 11.1 Hz, 1H, CH2−Ph), 4.31 (dd, J5′,6′b = 4.8 Hz, J6′a,6′b = 10.2
Hz, 1H, H6′b), 4.26 (dd, J5,6b = 4.8 Hz, J6a,6b = 10.2 Hz, 1H, H6b),
4.16−4.06 (m, 2H, H5′, H5), 4.10 (t, J2,3 = J3,4 = 9.3 Hz, 1H, H3), 3.93
(t, J2′,3′ = J3′,4′ = 9.0 Hz, 1H, H3′), 3.84 (dd, J1′,2′ = 3.9 Hz, J2′,3′ = 9.0
Hz, 1H, H2′), 3.78−3.64 (m, 4H, H2, H4′, H6′a, H6a), 3.52 (t, J3,4 = J4,5
= 9.3 Hz, 1H, H4), 0.97 (s, 9H, SiC(CH3)3), 0.12, 0.11 (2s, 6H,
Si(CH3)2); 13C NMR (CDCl3, 75 MHz) δ (ppm): 138.7, 137.3, 137.1
(3C, Cq−Ar), 129.3, 128.9, 128.3, 128.2, 128.1, 127.9, 127.4, 126.5,
126.0 (15C, CH−Ar), 102.3 (CH−Ph), 101.3 (CH−Ph), 95.3 (C1′),
94.0 (C1), 82.6 (C4′), 81.2 (C4), 78.5 (C3′), 75.2 (CH2−Ph), 72.7
(C2′), 72.5 (C2), 71.2 (C3), 69.0, 68.9 (C6, C6′), 63.1, 63.0 (C5, C5′),
26.0 (SiC(CH3)3), 18.1 (SiC(CH3)3), −4.2, −4.7 (Si(CH3)2). Anal.
Calcd for C39H50O11Si: C, 64.80; H, 6.97. Found: C, 64.36; H, 7.13.
ESI HRMS for C39H50O11Si [M+Na]+: Calcd 745.3015, Found
745.2999.
indicating that this H site is spectrally enantiodiscriminated.
Due to their molecular similarities, the absence of chiral
discrimination of Me1 and Me17 for 9b originates for the same
reasons described for 9a.
The NAD signals of Me2 obtained with (rac)-9b and (S)-9b
in PBLG/pyridine are compared in Figure 3c and 3d. At first
glance, a single doublet is observed in the enantioenriched
mixture. A deeper examination of the NAD 1D subspectrum
associated to Me2 and the 2D maps32 shows however a residual
doublet weakly emerging from noise, corresponding to the
minor R-isomer (see Figure S12 in the Supporting
Information). Due to the very low S/N ratio, deconvolution
of the NAD signals of the minor isomer was not possible, but
we can safely estimate that the e.e. is over 95%.
CONCLUSION
■
This work highlights the usefulness of the FeCl3·6H2O-
mediated tandem regioselective protection of trehalose. We
have achieved an efficient synthesis of model tetra-O-acylated
sulfolipid 1 in 11 steps and in 4% overall yield starting from
α,α-D-trehalose. Interestingly, enantiomeric excesses of mono-
methylated fatty acids were measured for the first time by NAD
2D-NMR spectroscopy in a polypeptide chiral liquid crystal.
The analytical potential of this original approach and the quality
of the first experimental NMR results obtained for 9a and 9b
suggest that this tool should be efficient enough to analyze di-
or trimethylated fatty acid esters. To access active compounds
and natural metabolites from Mycobacterium tuberculosis, the
syntheses of enantiopure polydeoxypropionates are currently in
progress.
EXPERIMENTAL SECTION
■
General. All air sensitive reactions were carried out in oven-dried
glassware under a slight positive pressure of argon. Dichloromethane,
acetonitrile, methanol, and tetrahydrofuran were dried using a dry
solvent station. TLC plates (Silica Gel 60 F254) were visualized under
UV (254 nm) and by staining in a 5% ethanolic sulfuric acid solution.
Silica Gel 60 (40−63 μm) was used for column chromatography.
Melting points are uncorrected. NMR spectra were recorded at 250,
300, or 360 MHz. Chemical shifts (in ppm) were determined relative
to residual undeuterated solvent as an internal reference. Abbreviations
of multiplicity were as follows: s (singlet), d (doublet), t (triplet), m
(multiplet), b (broad). High-resolution mass spectra (positive or
negative mode ESI) were performed with a tandem Q-TOF analyzer.
For optical rotations, concentrations (c) are given in g/100 mL.
Elemental analyses were obtained at the Service de Microanalyse of
ICSN-CNRS (Gif-sur-Yvette).
3′-O-Benzyl-4,6;4′,6′-di-O-benzylidene-2′-O-tert-butyldime-
thylsilyl-2-O-palmitoyl-α,α-D-trehalose (7). To a solution of
compound 6 (205 mg, 0.283 mmol) and palmitic acid (80 mg,
0.312 mmol, 1.1 equiv) in dichloromethane (10 mL) were added DCC
(89.2 mg, 0.432 mmol, 1.5 equiv) and DMAP (53 mg, 0.432 mmol, 1.5
equiv). The mixture was stirred for 5 h at room temperature, filtered
through Celite, and rinsed with ice-cold dichloromethane. After
evaporation of the solvent, the residue was purified by flash
chromatography on silica gel (cyclohexane/ethyl acetate: 95/5 then
90/10) to afford the expected compound 7 (224 mg, 82%). White
solid; mp 89−91 °C; [α]27D = +48 (c 1.0, CHCl3); 1H NMR (CDCl3,
300 MHz) δ (ppm): 7.52−7.28 (m, 15H, Har), 5.56 (s, 1H, CH−Ph),
Compounds 3 and 4 were synthetized according to reported
procedures,15 and analytical data were in agreement with those
previously reported: 3,15,33 4.15,34 Compounds 8 and 9a were prepared
according to the procedure reported by Breit,18 and analytical data
were in agreement with those reported.
3,3′-Di-O-benzyl-4,6;4′,6′-di-O-benzylidene-2′-O-tert-butyl-
dimethylsilyl-α,α-D-trehalose (5). tert-Butyldimethylsilyl triflate
(540 μL, 2.34 mmol, 1.0 equiv) was added slowly at −78 °C to a
solution of compound 4 (1.63 g, 2.34 mmol) and 2,6-lutidine (550 μL,
4.72 mmol, 2.0 equiv) in dichloromethane (8.0 mL). The reaction
mixture was stirred for 3 h at −78 °C and then warmed to room
temperature for 1 h. Methanol (10 mL) was added and the resulted
mixture was diluted with ethyl acetate and with a saturated aqueous
NaHCO3 solution. The aqueous layer was extracted twice with ethyl
acetate. The combined organic layers were washed with brine, dried
over Na2SO4, and concentrated in vacuo. The residue was purified by
flash chromatography on silica gel (cyclohexane/ethyl acetate: 9/1 to
7/3) to afford the expected compound 5 (1.40 g, 74%). White solid;
5.54 (s, 1H, CH−Ph), 5.38 (d, J1,2 = 3.6 Hz, 1H, H1), 5.04 (d, J1′,2′
3.6 Hz, 1H, H1′), 4.94 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.93 (dd, J1,2
=
=
3.6 Hz, J2,3 = 9.6 Hz, 1H, H2), 4.77 (d, J = 11.0 Hz, 1H, CH2−Ph),
4.34 (t, J2,3 = J3,4 = 9.6 Hz, 1H, H3), 4.20−4.12 (m, 3H, H6b, H6′b, H5),
3.93 (t, J2′,3′ = J3′,4′ = 9.3 Hz, 1H, H3′), 3.91 (m, 1H, H5′), 3.81 (dd, J1′,2′
= 3.6 Hz, J2′,3′ = 9.3 Hz, 1H, H2′), 3.76 (t, J6,6′ = J5,6 = 9.6 Hz, 1H, H6a),
3.71 (t, J6′a,6′b = J5′,6′a = 9.6 Hz, 1H, H6′a), 3.66 (t, J3′,4′ = J4′,5′ = 9.3 Hz,
1H, H4′), 3.61 (t, J3,4 = J4,5 = 9.6 Hz, 1H, H4), 2.48 (m, 2H, CH2), 1.64
(m, 2H, CH2), 1.26−1.19 (m, 24H), 0.97 (s, 9H, SiC(CH3)3), 0.89 (t,
J = 7.0 Hz, 3H, CH3), 0.10 (s, 6H, Si(CH3)2); 13C NMR (CDCl3, 75
MHz) δ (ppm): 173.6 (CO), 138.7, 137.4, 137.0 (Cq-Ar), 129.3,
E
dx.doi.org/10.1021/jo4012255 | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
128.8, 128.3, 128.1, 128.0, 127.9, 127.4, 126.5, 126.0 (15C, CH−Ar),
102.4 (CH−Ph), 101.3 (CH−Ph), 95.3 (C1′), 91.8 (C1), 82.4 (C4′),
81.6 (C4), 78.6 (C3′), 75.2 (CH2−Ph), 72.9 (C2), 72.6 (C2′), 68.9,
68.8 (C6 and C6′), 68.5 (C3), 63.2 (C5′), 62.7 (C5), 34.1 (CH2), 31.9
(CH2), 29.7, 29.6, 29.5, 29.4, 29.2, 29.1 (10C, CH2), 26.0
(SiC(CH3)3), 24.7 (CH2), 22.7 (CH2), 18.1 (SiC(CH3)3), 14.1
(CH3), −4.2, −4.8 (Si(CH3)2). Anal. Calcd for C55H80O12Si: C, 68.72;
H, 8.39. Found: C, 68.59; H, 8.61. ESI HRMS for C55H80O12Si [M
+Na]+: Calcd 983.5311, Found 983.5280.
3,3′-Di-O-benzyl-4,6;4′,6′-di-O-benzylidene-2-O-palmitoyl-
α,α-D-trehalose (7′). To an ice-cold solution of the per-O-silylated
α,α-D-trehalose 3 (100 mg, 0.109 mmol) and benzaldehyde (66 μL,
0.654 mmol, 6 equiv) in dichloromethane (200 μL) were added
dropwise a solution of FeCl3·6H2O in acetonitrile (50 μL of a 111 mM
solution; 5 mol %) and triethylsilane (38 μL, 0.240 mmol, 2.2 equiv).
The reaction was stirred for 3 h at room temperature. Palmitoyl
chloride (133 μL, 0.136 mmol, 4 equiv) was then added. The solution
was stirred overnight at 50 °C and treated with TBAF (700 μL, 1 M
solution in THF). The crude product was purified by silica gel
chromatography (cyclohexane/ethyl acetate: 90/10 to 50/50) to give
(CDCl3, 75 MHz) δ (ppm): 182.2 (CO), 39.3 (CH), 33.5, 31.9,
29.5, 29.4, 29.2, 27.1, 22.7 (CH2), 16.8 (CH3), 14.1 (CH3); ESI
HRMS for C11H21O2 [M−H]−: Calcd 185.1547, Found 185.1544.
(S)-2-Methyloctadecanoic Acid (10b). To a solution of 9b (501
mg, 1.41 mmol) in dichloromethane (10 mL) at room temperature
was added dropwise TFA (1.54 g, 13.4 mmol, 9.5 equiv). The solution
was stirred for 5 h at room temperature, and the mixture was
evaporated to dryness. The residue was coevaporated with toluene and
dichloromethane to afford the expected compound 10b (421 mg,
99%) as a white solid. [α]20D = +7 (c 1.0, CHCl3); mp 49−51 °C; 1H
NMR (CDCl3, 300 MHz) δ (ppm): 10.96 (bs, COOH), 2.47 (m, 1H,
CH), 1.68 (m, 1H, CH2), 1.43 (m, 1H, CH2), 1.37−1.21 (m, 28H,
CH2), 1.18 (d, 3H, J = 6.9 Hz, CH3), 0.89 (t, 3H, J = 6.9 Hz, CH3);
13C NMR (CDCl3, 75 MHz) δ (ppm): 182.2 (CO), 39.2 (CH),
33.5, 31.9, 29.7, 29.6, 29.5, 29.4, 27.1, 22.7 (14C, CH2), 16.8 (CH3),
14.1 (CH3); ESI HRMS for C19H37O2 [M−H]−: Calcd 297.2799,
Found 297.2796.
3′-O-Benzyl-4,6;4′,6′-di-O-benzylidene-2′-O-tert-butyldime-
thylsilyl-2,3-di-O-palmitoyl-α,α-D-trehalose (11a). To a solution
of the monoacylated derivative 7 (164 mg, 0.171 mmol) and palmitic
acid (60 mg, 0.232 mmol, 1.4 equiv) in dichloromethane (6 mL) were
added DCC (54 mg, 0.263 mmol, 1.5 equiv) and DMAP (31 mg,
0.254 mmol, 1.5 equiv). The reaction mixture was stirred overnight at
room temperature, filtered through Celite, and rinsed with ice-cold
dichloromethane. After evaporation of the solvent, the residue was
purified by flash chromatography on silica gel (cyclohexane/ethyl
acetate: 95/5 then 90/10 then 80/20). The expected compound 11a
the expected product 7′ as a colorless syrup (28 mg, 27%). [α]27
=
D
1
+61 (c 0.1, CHCl3); H NMR (CDCl3, 360 MHz) δ (ppm): 7.55−
7.28 (m, 20H, Har), 5.63 (s, 1H, CH−Ph), 5.60 (s, 1H, CH−Ph), 5.37
(d, J1,2 = 3.6 Hz, 1H, H1), 5.17 (d, J1′,2′ = 3.6 Hz, 1H, H1′), 5.06 (d, J =
11.5 Hz, 1H, CH2−Ph), 5.01 (dd, J1,2 = 3.6 Hz, J2,3 = 9.7 Hz, 1H, H2),
4.97 (d, J = 11.9 Hz, 1H, CH2−Ph), 4.79 (d, J = 11.5 Hz, 1H, CH2−
Ph), 4.77 (d, J = 11.9 Hz, 1H, CH2−Ph), 4.33 (dd, J5,6b = 4.5 Hz, J6a,6b
(224 mg) was obtained quantitatively as a colorless syrup. [α]20
=
= 10.0 Hz, 1H, H6b), 4.26−4.16 (m, 2H, H5, H6′b), 4.12 (t, J2,3 = J3,4
=
D
1
9.7 Hz, 1H, H3), 3.95 (m, 1H, H5′), 3.94 (t, J2′,3′ = J3′,4′ = 9.4 Hz, 1H,
H3′), 3.82−3.67 (m, 5H, H2′, H6a, H6′a, H4, H4′), 2.40 (t, J = 7.2 Hz,
2H, CH2), 1.64 (m, 2H, CH2), 1.42−1.12 (m, 24H, CH2), 0.91 (t, J =
6.8 Hz, 3H, CH3); 13C NMR (CDCl3, 75 MHz) δ (ppm): 173.0 (C
O), 138.5, 138.3, 137.3 (4C, Cq−Ar), 128.9, 128.5, 128.3, 128.2, 128.0,
127.9, 127.6, 127.5, 126.0, 125.9 (20C, CH−Ar), 101.2 (2C, CH−Ph),
95.1 (C1′), 93.1 (C1), 82.1, 81.9 (C4, C4′), 78.7 (C3′), 76.3 (C3), 75.0,
74.9 (CH2−Ph), 72.4 (C2), 71.5 (C2′), 68.8 (2C, C6, C6′), 63.5 (C5′),
63.0 (C5), 34.1 (CH2), 31.9 (CH2), 29.7, 29.6, 29.5, 29.3, 29.2 (10C,
CH2), 24.8 (CH2), 22.7 (CH2), 14.1 (CH3). Anal. Calcd for
C56H72O12: C, 71.77, H, 7.74. Found: C, 71.33, H, 7.78. ESI HRMS
for C56H72O12 [M+Na]+: Calcd 959.4916, Found 959.4884.
+38 (c 1.0, CHCl3); H NMR (CDCl3, 360 MHz) δ (ppm): 7.45−
7.27 (m, 15H, Har), 5.75 (t, J2,3 = J3,4 = 9.7 Hz, 1H, H3), 5.52 (s, 1H,
CH−Ph), 5.50 (s, 1H, CH−Ph), 5.38 (d, J1,2 = 4.0 Hz, 1H, H1), 5.06
(dd, J1,2 = 4.0 Hz, J2,3 = 9.7 Hz, 1H, H2), 5.05 (d, J1′,2′ = 3.6 Hz, 1H,
H1′), 4.96 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.81 (d, J = 11.0 Hz, 1H,
CH2−Ph), 4.31−4.23 (m, 2H, H6b, H5), 4.13 (dd, J5′,6′b = 4.9 Hz, J6′a,6′b
= 10.2 Hz, 1H, H6′b), 4.01 (t, J2′,3′ = J3′,4′ = 9.0 Hz, 1H, H3′), 3.91 (m,
1H, H5′), 3.81 (dd, J1′,2′ = 3.6 Hz, J2′,3′ = 9.0 Hz, 1H, H2′), 3.76−3.63
(m, 4H, H6a, H6′a, H4, H4′), 2.42−2.36 (m, 2H), 2.34−2.26 (m, 2H),
1.62−1.55 (m, 4H), 1.36−1.16 (m, 48H), 0.94 (s, 9H, SiC(CH3)3),
0.90 (t, J = 7.2 Hz, 6H, CH3), 0.10, 0.09 (s, 6H, Si(CH3)2); 13C NMR
(CDCl3, 90 MHz) δ (ppm): 173.2, 172.5 (CO), 138.9, 137.4, 137.1
(Cq−Ar), 129.0, 128.8, 128.1, 128.0, 127.3, 126.4, 126.1 (15C, CH−
Ar), 102.0 (CH−Ph), 101.4 (CH−Ph), 95.7 (C1′), 92.1 (C1), 82.6
(C4′), 79.6 (C4), 78.5 (C3′), 75.3 (CH2−Ph), 72.7 (C2′), 71.0 (C2),
68.9, 68.8 (C6, C6′), 68.7 (C3), 63.2, 63.0 (C5′, C5), 34.3, 34.1 (CH2),
31.9 (2C, CH2), 29.7, 29.5, 29.4, 29.3, 29.1, 29.0 (20C, CH2), 26.0
(SiC(CH3)3), 25.1, 24.8 (CH2), 22.7 (2C, CH2), 18.2 (SiC(CH3)3),
14.1 (2C, CH3), −4.3, −4.7 (Si(CH3)2). Anal. Calcd for C71H110O13Si:
C, 71.08; H, 9.24. Found: C, 71.04; H, 9.46. ESI HRMS for
C71H110O13Si [M+Na]+: Calcd 1221.7608, Found 1221.7543.
3′-O-Benzyl-4,6;4′,6′-di-O-benzylidene-2′-O-tert-butyldime-
thylsilyl-3-O-[(S)-2-methyloctadecanoyl]-2-O-palmitoyl-α,α-D-
trehalose (11b). To a solution of compound 7 (389 mg, 0.405
mmol) and (S)-2-methyloctadecanoic acid 10b (133 mg, 0.446 mmol,
1.1 equiv), in dichloromethane (17 mL) were added DCC (92 mg,
0.446 mmol, 1.1 equiv) and DMAP (54 mg, 0.446 mmol, 1.1 equiv).
The reaction was stirred overnight at room temperature and
concentrated under vacuum. The residue was purified by flash
chromatography on silica gel (cyclohexane/ethyl acetate: 90/10). The
(S)-tert-Butyl 2-Methyloctadecanoate (9b). To a solution of
compound 8 (1.22 g, 4.39 mmol) in dry THF (11 mL) under an argon
atmosphere, were successively added at 0 °C a solution of anhydrous
ZnCl2 in dry THF (1 mL of a 0.65 M solution, 15 mol %) and
C16H33MgCl35 (6 mL of a 1.1 M solution, 1.5 equiv). The reaction
mixture was stirred for 3 h at 0 °C and diluted with n-pentane. A
saturated aqueous NH4Cl solution was added, the aqueous layer was
extracted with n-pentane, and the combined organic layers were
carefully concentrated. The crude product was purified by silica gel
chromatography (n-pentane/Et2O: 50/1). The solvents were removed
under atmospheric pressure to give the expected compound 9b as a
colorless oil (1.26 g, 81%) (ee > 95%, see above). [α]20 = +8 (c 1.3,
D
1
CHCl3); H NMR (CDCl3, 300 MHz) δ (ppm): 2.29 (m, 1H, CH),
1.65−1.55 (m, 1H, CH2), 1.44 (s, 9H, C(CH3)3), 1.35−1.20 (m, 29H,
CH2), 1.08 (d, J = 6.9 Hz, 3H, CH3), 0.88 (t, J = 6.9 Hz, 3H, CH3);
13C NMR (CDCl3, 75 MHz) δ (ppm): 176.3 (CO), 79.6
(C(CH3)3), 40.6 (CH), 33.9 (CH2), 31.9, 29.7, 29.6, 29.5, 29.4,
28.1, 27.2, 22.7 (14C, CH2), 28.1 (C(CH3)3), 17.1 (CH3), 14.1
(CH3). ESI HRMS for C23H46NaO2 [M+Na]+: Calcd 377.3390,
Found 377.3359.
expected compound 11b (322 mg, 64%) was obtained as a colorless
1
syrup. [α]20 = +37(c 1.1, CHCl3); H NMR (CDCl3, 300 MHz) δ
D
(ppm): 7.46−7.28 (m, 15H, Har), 5.78 (t, J2,3 = J3,4 = 10.0 Hz, 1H, H3),
(S)-2-Methyldecanoic Acid (10a). To a solution of 9a (312 mg,
1.29 mmol) in dichloromethane (10 mL) at room temperature was
added dropwise TFA (1.54 g, 13.5 mmol, 10.5 equiv). The solution
was stirred for 5 h at room temperature, and the mixture was
evaporated to dryness. The residue was coevaporated with toluene and
dichloromethane to afford the expected compound 10a36 (236 mg,
95%) as a yellow oil. [α]20D = +12 (c 1.2, CHCl3); 1H NMR (CDCl3,
300 MHz) δ (ppm): 11.24 (bs, COOH), 2.47 (m, 1H, CH), 1.69 (m,
1H, CH2), 1.45 (m, 1H, CH2), 1.37−1.22 (m, 12H, CH2), 1.18 (d,
3H, J = 6.9 Hz, CH3), 0.88 (t, 3H, J = 6.9 Hz, CH3); 13C NMR
5.54 (s, 1H, CH−Ph), 5.52 (s, 1H, CH−Ph), 5.40 (d, J1,2 = 3.9 Hz,
1H, H1), 5.09 (dd, J1,2 = 3.9 Hz, J2,3 = 10.0 Hz, 1H, H2), 5.06 (d, J1′,2′
=
3.3 Hz, 1H, H1′), 4.98 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.82 (d, J = 11.0
Hz, 1H, CH2−Ph), 4.34−4.22 (m, 2H, H6b, H5), 4.14 (dd, J5′,6′b = 4.5
Hz, J6′a,6′b = 10.2 Hz, 1H, H6′b), 4.02 (t, J2′,3′ = J3′,4′ = 9.0 Hz, 1H, H3′),
3.92 (m, 1H, H5′), 3.82 (dd, J1′,2′ = 3.3 Hz, J2′,3′ = 9.0 Hz, 1H, H2′),
3.78−3.61 (m, 4H, H6a, H6′a, H4, H4′), 2.51−2.30 (m, 3H), 1.67−1.54
(m, 3H), 1.46−1.14 (m, 53H), 1.12 (d, J = 7.2 Hz, 3H, CH3), 0.95 (s,
9H, SiC(CH3)3), 0.91 (t, J = 7.2 Hz, 6H, CH3), 0.10, 0.09 (2s, 6H,
F
dx.doi.org/10.1021/jo4012255 | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
Si(CH3)2); 13C NMR (CDCl3, 75 MHz) δ (ppm): 175.5, 173.2 (C
O), 138.8, 137.4, 137.0 (Cq−Ar), 128.9, 128.8, 128.1, 128.0, 127.3,
126.1, 126.0 (15C, CH−Ar), 101.8 (CH−Ph), 101.4 (CH−Ph), 95.6
(C1′), 92.0 (C1), 82.5 (C4′), 79.6 (C4), 78.5 (C3′), 75.3 (CH2−Ph),
72.7 (C2′), 70.8 (C2), 68.9, 68.8 (C6, C6′), 68.4 (C3), 63.2 (C5′), 62.9
(C5), 39.9 (CH), 34.0, 33.7, 31.9, 29.7, 29.6, 29.5, 29.4, 29.3, 29.2, 27.3
(27C, CH2), 26.0 (SiC(CH3)3), 24.7, 22.8 (CH2), 18.2 (SiC(CH3)3),
17.3 (CH3), 14.1 (2C, CH3), −4.2, −4.6 (Si(CH3)2); ESI HRMS for
C74H116O13Si [M+Na]+: Calcd 1263.8077, Found 1263.8090.
−4.8 (Si(CH3)2). ESI HRMS for C74H120O13Si [M+Na]+: Calcd
1267.8390, Found 1267.8375.
3′,4,4′-Tri-O-benzyl-2′-O-tert-butyldimethylsilyl-2,3,6,6′-
tetra-O-palmitoyl-α,α-D-trehalose (13a). To a solution of
compound 12a (33.0 mg, 0.027 mmol) and palmitic acid (21 mg,
0.081 mmol, 3.0 equiv) in dichloromethane (1.4 mL) were added
DCC (14 mg, 0.068 mmol, 2.5 equiv) and DMAP (5.0 mg, 0.041
mmol, 1.5 equiv). The reaction mixture was stirred overnight at room
temperature and then concentrated. The residue was purified by flash
chromatography on silica gel (cyclohexane/ethyl acetate: 95/5 then
90/10). The expected tetra-O-acylated derivative 13a (43.7 mg, 95%)
was obtained as a colorless syrup. [α]20D +47 (c 1.0, CHCl3); 1H NMR
(CDCl3, 300 MHz) δ (ppm): 7.37−7.15 (m, 15H, Har), 5.72 (dd, J2,3
= 10.2 Hz, J3,4 = 9.4 Hz, 1H, H3), 5.24 (d, J1,2 = 4.0 Hz, 1H, H1), 5.01
(d, J1′,2′ = 3.2 Hz, 1H, H1′), 5.00 (dd, J1,2 = 4.0 Hz, J2,3 = 10.2 Hz, 1H,
H2), 4.99 (d, J = 11.5 Hz, 1H, CH2−Ph), 4.89 (d, J = 11.5 Hz, 1H,
CH2−Ph), 4.82 (d, J = 11.2 Hz, 1H, CH2−Ph), 4.62 (d, J = 11.5 Hz,
1H, CH2−Ph), 4.57 (d, J = 11.5 Hz, 1H, CH2−Ph), 4.52 (d, J = 11.2
Hz, 1H, CH2−Ph), 4.29−4.16 (m, 5H, H5, H6b, H6a, H6′b, H6′a), 3.97
3′,4,4′-Tri-O-benzyl-2′-O-tert-butyldimethylsilyl-2,3-di-O-
palmitoyl-α,α-D-trehalose (12a). To a solution of compound 11a
(82 mg, 0.068 mmol) in dry dichloromethane (2.5 mL) containing 4 Å
molecular sieves at −78 °C were added dichlorophenylborane (45 μL,
0.343 mmol, 5 equiv) and triethylsilane (110 μL, 0.689 mmol, 10
equiv). The mixture was stirred for 6 h at −78 °C and then warmed to
−20 °C for 1 h. The reaction mixture was quenched with triethylamine
(2.5 mL) and methanol (2.5 mL) and concentrated under vacuum.
The residue was purified by flash chromatography on silica gel
(cyclohexane/ethyl acetate: 80/20 to 70/30) to afford the title
compound 12a (77 mg, 93%) as a colorless syrup. [α]20D = +65 (c 0.3,
CHCl3); 1H NMR (CDCl3, 300 MHz) δ (ppm): 7.39−7.19 (m, 15H,
Har), 5.71 (t, J2,3 = J3,4 = 9.8 Hz, 1H, H3), 5.27 (d, J1,2 = 4.0 Hz, 1H,
H1), 4.97 (d, J1′,2′ = 3.5 Hz, 1H, H1′), 4.97 (d, J = 11.7 Hz, 1H, CH2−
Ph), 4.90 (dd, J1,2 = 4.0 Hz, J2,3 = 9.8 Hz, 1H, H2), 4.89 (d, J = 11.7 Hz,
1H, CH2−Ph), 4.81 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.62 (m, 2H,
CH2−Ph), 4.59 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.09 (m, 1H, H5), 3.94
(t, J2′,3′ = J3′,4′ = 9.0 Hz, 1H, H3′), 3.86 (m, 1H, H5′), 3.75 (dd, J1′,2′
=
3.2 Hz, J2′,3′ = 9.0 Hz, 1H, H2′), 3.70 (t, J3,4 = J4,5 = 9.4 Hz, 1H, H4),
3.44 (t, J3′,4′ = 9.0 Hz, J4′,5′ = 10.0 Hz, 1H, H4′), 2.36−2.18 (m, 8H,
CH2), 1.68−1.52 (m, 8H, CH2), 1.35−1.19 (m, 96H, CH2), 0.93−
0.88 (m, 21H, SiC(CH3)3, 4CH2), 0.09, 0.07 (2s, 6H, Si(CH3)2); 13
C
NMR (CDCl3, 90 MHz) δ (ppm): 173.4, 173.3, 172.8, 172.5 (CO),
138.8, 137.9, 137.5 (Cq−Ar), 128.4, 128.3, 128.2, 127.8, 127.7, 127.5,
127.2 (15C, CH−Ar), 94.5 (C1′), 91.1 (C1), 81.7 (C3′), 78.1 (C4′),
76.1 (C4), 75.5 (CH2−Ph), 74.9 (CH2−Ph), 74.4 (CH2−Ph), 73.0
(C2′), 71.9 (C3), 70.3 (C2), 69.6 (C5′), 68.9 (C5), 62.7, 62.3 (C6′, C6),
34.3, 34.1, 34.0 (3C, CH2), 31.9 (4C, CH2), 29.7, 29.6, 29.5, 29.4,
29.3, 29.2, 29.1 (40C, CH2), 26.0 (SiC(CH3)3), 24.9, 24.8, 24.7 (4C,
CH2), 22.7 (4C, CH2), 18.0 (SiC(CH3)3), 14.1 (4C, CH3), −4.1, −4.8
(Si(CH3)2); ESI HRMS for C103H174O15Si [M+Na]+: Calcd
1702.2514, Found 1702.2474.
3′,4,4′-Tri-O-benzyl-2′-O-tert-butyldimethylsilyl-3-O-[(S)-2-
methyloctadecanoyl]-6,6′-di-O-[(S)-2-methyldecanoyl]-2-O-
palmitoyl-α,α-D-trehalose (13b). To a solution of compound 12b
(100 mg, 0.080 mmol) and (S)-2-methyldecanoic acid 10a (48 mg,
0.256 mmol, 3.2 equiv) in dichloromethane (1 mL) were added DCC
(53 mg, 0.256 mmol, 3.2 equiv) and DMAP (29 mg, 0.240 mmol, 3
equiv). The reaction was stirred overnight at room temperature and
concentrated under vacuum. The residue was purified by flash
chromatography on silica gel (cyclohexane/ethyl acetate: 90/10). The
(t, J2′,3′ = J3′,4′ = 9.5 Hz, 1H, H3′), 3.82−3.60 (m, 7H, H6b, H6a, H6′b
,
H6′a, H4, H5′, H2′), 3.52 (t, J3′,4′ = J4′,5′ = 9.5 Hz, 1H, H4′), 2.29−2.15
(m, 4H, CH2), 1.70−1.49 (m, 4H, CH2), 1.40−1.10 (m, 48H, CH2),
0.95−0.83 (m, 15H, SiC(CH3)3, 2CH3), 0.07, 0.06 (2s, 6H,
Si(CH3)2); 13C NMR (CDCl3, 90 MHz) δ (ppm): 173.2, 172.6
(CO), 139.0, 138.2, 137.9 (Cq−Ar), 128.4, 128.3, 128.2, 127.8,
127.7, 127.6, 127.5, 127.1 (15C, CH−Ar), 95.1 (C1′), 91.6 (C1), 81.5
(C3′), 77.8 (C4′), 75.5 (C4), 75.3 (CH2−Ph), 74.8 (CH2−Ph), 74.4
(CH2−Ph), 73.1 (C5′), 71.7, 71.6 (C3, C2′), 71.1 (C5), 70.8 (C2), 61.7
(C6′), 61.2 (C6), 34.3, 34.1 (CH2), 31.9 (2C, CH2), 29.7, 29.6, 29.5,
29.4, 29.3, 29.2, 29.1 (20C, CH2), 26.0 (SiC(CH3)3), 24.9, 24.7
(CH2), 22.7 (2C, CH2), 18.0 (SiC(CH3)3), 14.1 (2C, CH3), −4.2,
−4.8 (Si(CH3)2); ESI HRMS for C71H114O13Si [M+Na]+: Calcd
1225.7921, Found 1225.7860.
3′,4,4′-Tri-O-benzyl-2′-O-tert-butyldimethylsilyl-3-O-[(S)-2-
methyloctadecanoyl]-2-O-palmitoyl-α,α-D-trehalose (12b). To
a solution of compound 11b (161 mg, 0.130 mmol) in dry
dichloromethane (2 mL) containing 4 Å molecular sieves at −78 °C
were added dichlorophenylborane (85 μL, 0.650 mmol, 5 equiv) and
triethylsilane (207 μL, 1.30 mmol, 10 equiv). The mixture was stirred
for 6 h at −78 °C and then warmed to −20 °C for 1 h. The reaction
mixture was quenched with triethylamine (5 mL) and methanol (5
mL) and concentrated under vacuum. The residue was purified by
flash chromatography on silica gel (cyclohexane/ethyl acetate: 80/20
expected compound 13b (105 mg, 83%) was obtained as a colorless
1
syrup. [α]20 = +60 (c 1.0, CHCl3); H NMR (CDCl3, 360 MHz) δ
D
(ppm): 7.37−7.14 (m, 15H, Har), 5.75 (dd, J2,3 = 10.1 Hz, J3,4 = 9.4
Hz, 1H, H3), 5.25 (d, J1,2 = 3.6 Hz, 1H, H1), 5.00 (dd, J1,2 = 3.6 Hz, J2,3
= 10.1 Hz, 1H, H2), 4.99 (d, J1′,2′ = 3.6 Hz, 1H, H1′), 4.98 (d, J = 11.5
Hz, 1H, CH2−Ph), 4.88 (d, J = 11.5 Hz, 1H, CH2−Ph), 4.82 (d, J =
11.2 Hz, 1H, CH2−Ph), 4.63 (d, J = 11.5 Hz, 1H, CH2−Ph), 4.54 (d, J
= 11.5 Hz, 1H, CH2−Ph), 4.51 (d, J = 11.2 Hz, 1H, CH2−Ph), 4.35−
4.23 (m, 3H, H5, H6b, H6a), 4.21−4.16 (m, 2H, H6′b, H6′a), 3.97 (t, J2′,3′
= J3′,4′ = 9.4 Hz, 1H, H3′), 3.85 (m, 1H, H5′), 3.73 (dd, J1′,2′ = 3.6 Hz,
to 70/30) to afford the title compound 12b (115 mg, 71%) as a
1
colorless syrup. [α]20 = +68 (c 1.0, CHCl3); H NMR (CDCl3, 300
D
MHz) δ (ppm): 7.41−7.22 (m, 15H, Har), 5.78 (t, J2,3 = J3,4 = 9.9 Hz,
1H, H3), 5.37 (d, J1,2 = 3.6 Hz, 1H, H1), 5.03 (d, J1′,2′ = 3.0 Hz, 1H,
J
2′,3′ = 9.4 Hz, 1H, H2′), 3.72 (t, J3,4 = J4,5 = 9.4 Hz, 1H, H4), 3.45 (dd,
H1′), 5.01 (d, J = 11.5 Hz, 1H, CH2−Ph), 4.96 (dd, J1,2 = 3.6 Hz, J2,3
=
J3′,4′ = 9.4 Hz, J4′,5′ = 9.0 Hz, 1H, H4′), 2.53−2.43 (m, 2H), 2.35 (m,
1H), 2.27−2.22 (m, 2H,), 1.72−1.60 (m, 3H), 1.54−1.49 (m, 2H),
1.35−1.19 (m, 79H), 1.18 (d, J = 7.2 Hz, 3H, CH3), 1.14 (d, J = 7.2
Hz, 3H, CH3), 1.08 (d, J = 6.8 Hz, 3H, CH3), 0.91−0.85 (m, 21H,
SiC(CH3) and 4 CH3), 0.09, 0.07 (2s, 6H, Si(CH3)2); 13C NMR
(CDCl3, 90 MHz) δ (ppm): 176.6, 176.5, 175.5, 172.7 (CO), 138.8,
137.9, 137.5 (Cq-Ar), 128.4, 128.3, 128.2, 127.7, 127.6, 127.5, 127.2,
127.1, 127.0 (15C, CH−Ar), 94.3 (C1′), 91.0 (C1), 81.6 (C3′), 78.3
(C4′), 76.3 (C4), 75.5 (CH2−Ph), 75.0 (CH2−Ph), 74.4 (CH2−Ph),
73.0 (C2′), 71.7 (C3), 70.4 (C2), 69.5 (C5′), 68.9 (C5), 62.6 (C6′), 62.1
(C6), 39.5, 39.4, 39.3 (CH), 34.2, 34.0, 33.8, 33.7, 31.9, 3.8, 29.9, 29.8,
29.7, 29.6, 29.5, 29.4, 29.3, 29.2, 27.3, 27.2, (40C, CH2), 26.0
(SiC(CH3)3), 24.6, 22.7, 22.6 (CH2), 18.0 (SiC(CH3)3), 17.1, 17.0,
16.5 (CH3), 14.1, 14.0 (4C, CH3), −4.1, −4.8 (Si(CH3)2); ESI HRMS
for C96H160O15Si [M+Na]+: Calcd 1604.1419, Found 1604.1457.
9.9 Hz, 1H, H2), 4.93 (d, J = 11.5 Hz, 1H, CH2−Ph), 4.85 (d, J = 11.1
Hz, 1H, CH2−Ph), 4.71 (s, 2H, CH2−Ph), 4.63 (d, J = 11.1 Hz, 1H,
CH2−Ph), 4.14 (m, 1H, H5), 3.98 (t, J2′,3′ = J3′,4′ = 9.1 Hz, 1H, H3′),
3.85 (t, J3,4 = J4,5 = 9.9 Hz, 1H, H4), 3.84−3.60 (m, 6H, H6b, H6a, H6′b
,
H6′a, H5′, H2′), 3.54 (t, J4′,3′ = J4′,5′ = 9.1 Hz, 1H, H4′), 2.45−2.34 (m,
1H), 2.34−2.26 (m, 2H), 1.75−1.50 (m, 3H), 1.47−1.16 (m, 53H),
1.12 (d, J = 7.2 Hz, 3H, CH3), 0.95−0.88 (m, 15H, SiC(CH3)3 and 2
CH3), 0.11, 0.09 (2s, 6H, Si(CH3)2); 13C NMR (CDCl3, 75 MHz) δ
(ppm): 175.5, 173.3 (CO), 138.9, 138.1, 137.9 (Cq−Ar), 128.2,
128.1, 127.7, 127.6, 127.5, 127.3, 127.2, 127.1 (15C, CH−Ar), 94.8
(C1′), 91.2 (C1), 81.5 (C3′), 77.8 (C4′), 75.5 (C4), 75.3 (CH2−Ph),
74.8 (CH2−Ph), 74.3 (CH2−Ph), 72.9 (C2′), 71.9 (C5′), 71.4 (C3),
71.2 (C5), 70.9 (C2), 61.5 (C6), 61.0 (C6′), 39.5 (CH), 34.0, 33.7,
31.9, 29.7, 29.6, 29.5, 29.3, 27.2 (27C, CH2), 25.9 (SiC(CH3)3), 24.5,
22.8 (CH2), 17.9 (SiC(CH3)3), 16.6 (CH3), 14.1 (2C, CH3), −4.3,
G
dx.doi.org/10.1021/jo4012255 | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
3′,4,4′-Tri-O-benzyl-2,3,6,6′-tetra-O-palmitoyl-α,α-D-treha-
lose (14a). To a solution of compound 13a (72 mg, 0.043 mmol) in
THF (1.0 mL) was added a 1 M solution of tetrabutylammonium
fluoride in THF (130 μL, 0.130 mmol, 3 equiv). The reaction mixture
was stirred for 1.5 h at room temperature, diluted with ethyl acetate,
and hydrolyzed with a saturated aqueous NaHCO3 solution. The
aqueous layer was extracted twice with ethyl acetate, and the combined
organic layers were washed with brine, dried over Na2SO4, and
concentrated under vacuum. The residue was purified by flash
chromatography on silica gel (cyclohexane/ethyl acetate: 90/10 then
coevaporated with toluene and dichloromethane. The residue was
subjected to flash chromatography on silica gel (CH2Cl2/MeOH: 90/
10), and the product-containing fractions were passed through an
Amberlite IR-120 column (Na+ form) with the use of the same solvent
system before to be concentrated. The expected O-sulfated compound
15a (22.4 mg, 96%) was obtained as a colorless syrup. 1H NMR
(CDCl3/CD3OD: 2/1, 300 MHz) δ (ppm): 7.42−7.14 (m, 15H, Har),
5.60 (t, J2,3 = J3,4 = 10.0 Hz, 1H, H3), 5.46 (d, J1′,2′ = 3.6 Hz, 1H, H1′),
5.17 (d, J1,2 = 3.6 Hz, 1H, H1), 5.11 (d, J = 10.2 Hz, 1H, CH2−Ph),
4.91 (dd, J1,2 = 3.6 Hz, J2,3 = 10.0 Hz, 1H, H2), 4.82 (d, J = 11.1 Hz,
1H, CH2−Ph), 4.70 (d, J = 10.2 Hz, 1H, CH2−Ph), 4.55 (s, 2H,
CH2−Ph), 4.49 (d, J = 11.1 Hz, 1H, CH2−Ph), 4.51−4.27 (m, 4H,
H6b, H6a, H5, H2′), 4.13 (m, 2H, H6′a, H6′b), 4.04 (t, J2′,3′ = J3′,4′ = 9.5
Hz, 1H, H3′), 3.86 (m, 1H, H5′), 3.72 (t, J3,4 = J4,5 = 10.0 Hz, 1H, H4),
3.43 (t, J3′,4′ = J4′,5′ = 9.5 Hz, 1H, H4′), 2.34−2.13 (m, 8H, CH2), 1.66−
1.46 (m, 8H, CH2), 1.34−1.12 (m, 96H, CH2), 0.84 (m, 12H, CH3);
13C NMR (CDCl3/CD3OD: 2/1, 75 MHz) δ (ppm): 174.0, 173.6,
80/20) to afford the expected compound 14a (61 mg, 91%) as a
1
colorless syrup. [α]20 = +63 (c 1.0, CHCl3); H NMR (CDCl3, 300
D
MHz) δ (ppm): 7.41−7.24 (m, 15H, Har), 5.62 (t, J2,3 = J3,4 = 9.5 Hz,
1H, H3), 5.21 (d, J1,2 = 3.9 Hz, 1H, H1), 5.07 (d, J1′,2′ = 3.3 Hz, 1H,
H1′), 4.96 (dd, J1,2 = 3.9 Hz, J2,3 = 9.5 Hz, 1H, H2), 4.93 (d, J = 11.0
Hz, 1H, CH2−Ph), 4.87 (d, J = 10.8 Hz, 1H, CH2−Ph), 4.86 (d, J =
11.0 Hz, 1H, CH2−Ph), 4.59 (d, J = 10.8 Hz, 1H, CH2−Ph), 4.59 (d, J
= 10.8 Hz, 1H, CH2−Ph), 4.53 (d, J = 10.8 Hz, 1H, CH2−Ph), 4.34−
4.14 (m, 5H, H5, H6b, H6a, H6′b, H6′a), 3.90 (t, J2′,3′ = J3′,4′ = 9.0 Hz, 1H,
H3′), 3.88 (m, 1H, H5′), 3.66 (m, 1H, H2′), 3.65 (t, J3,4 = J4,5 = 9.5 Hz,
1H, H4), 3.47 (t, J3′,4′ = J4′,5′ = 9.0 Hz, 1H, H4′), 2.35−2.21 (m, 8H,
CH2), 1.88 (d, J = 5.7 Hz, 1H, OH), 1.67−1.56 (m, 8H, CH2), 1.30−
1.20 (m, 96H, CH2), 0.89 (t, J = 7.0 Hz, 12H, CH3); 13C NMR
(CDCl3, 75 MHz) δ (ppm): 173.4, 173.3, 172.7, 172.5 (CO), 138.3,
137.7, 137.3 (Cq−Ar), 128.7, 128.5, 128.4, 128.1, 128.0, 127.9 (15C,
CH−Ar), 94.3 (C1′), 91.9 (C1), 82.1 (C3′), 77.7 (C4′), 76.1 (C4), 75.6
(CH2−Ph), 75.0 (CH2−Ph), 74.6 (CH2−Ph), 71.9 (C2′), 71.7 (C3),
70.3 (C2), 69.9 (C5′), 69.0 (C5), 62.5, 62.2 (C6, C6′), 34.4, 34.2, 34.1
(4C, CH2), 31.9 (4C, CH2), 29.7, 29.6, 29.5, 29.4, 29.3, 29.2, (40C,
CH2), 24.9, 24.8 (4C, CH2), 22.7 (4C, CH2), 14.1 (4C, CH3); ESI
HRMS for C97H160O15 [M+Na]+: Calcd 1588.1649, Found 1588.1641.
3′,4,4′-Tri-O-benzyl-3-O-[(S)-2-methyloctadecanoyl]-6,6′-di-
O-[(S)-2-methyldecanoyl]-2-O-palmitoyl-α,α-D-trehalose (14b).
To a solution of compound 13b (94 mg, 0.06 mmol) in THF (2 mL)
was added a 1 M solution of tetrabutylammonium fluoride in THF
(120 μL, 0.120 mmol, 2 equiv). The reaction mixture was stirred for 2
h at room temperature, diluted with ethyl acetate, and hydrolyzed with
a saturated aqueous NaHCO3 solution. The aqueous layer was
extracted twice with ethyl acetate, and the combined organic layers
were washed with brine, dried over Na2SO4, and concentrated under
vacuum. The residue was purified by flash chromatography on silica gel
(cyclohexane/ethyl acetate: 90/10 then 80/20) to afford the expected
compound 14b (74 mg, 85%) as a colorless syrup. [α]23D = +74 (c 1.0,
CHCl3); 1H NMR (CDCl3, 300 MHz) δ (ppm): 7.41−7.20 (m, 15H,
Har), 5.65 (t, J2,3 = J3,4 = 9.9 Hz, 1H, H3), 5.23 (d, J1,2 = 3.6 Hz, 1H,
H1), 5.09 (d, J1′,2′ = 3.6 Hz, 1H, H1′), 4.98 (dd, J1,2 = 3.6 Hz, J2,3 = 10.0
Hz, 1H, H2), 4.92 (d, J = 11.1 Hz, 1H, CH2−Ph), 4.88 (d, J = 11.0 Hz,
1H, CH2−Ph), 4.85 (d, J = 11.1 Hz, 1H, CH2−Ph), 4.62 (d, J = 10.5
Hz, 1H, CH2−Ph), 4.59 (d, J = 11.0 Hz, 1H, CH2−Ph), 4.52 (d, J =
10.5 Hz, 1H, CH2−Ph), 4.37−4.14 (m, 5H, H5, H6b, H6a, H6′b, H6′a),
3.91 (t, J2′,3′ = J3′,4′ = 9.6 Hz, 1H, H3′), 3.88 (m, 1H, H5′), 3.70−3.62
(m, 2H, H2′, H4), 3.50 (t, J3′,4′ = J4′,5′ = 9.6 Hz, 1H, H4′), 2.54−2.35
(m, 3H), 2.27 (m, 2H), 1.89 (bs, 1H, OH), 1.74−1.49 (m, 5H), 1.49−
1.09 (m, 88H), 0.92−0.85 (m, 12H, 4 CH3); 13C NMR (CDCl3, 75
MHz) δ (ppm): 176.5, 176.4, 175.6, 172.7 (CO), 138.2, 137.7,
137.2 (Cq−Ar), 128.7, 128.5, 128.2, 128.0, 127.9, 127.8 (15C, CH-Ar),
94.2 (C1′), 91.8 (C1), 82.0 (C3′), 77.9 (C4′), 76.3 (C4), 75.7 (CH2−
Ph), 75.1 (CH2−Ph), 74.6 (CH2−Ph), 71.9 (C2′), 71.4 (C3), 70.5
(C2), 70.0 (C5′), 69.0 (C5), 62.3, 62.2 (C6, C6′), 39.6, 39.5, 39.4 (CH),
34.0, 33.8, 33.7, 33.6, 31.9, 31.8, 29.7, 29.6, 29.5, 29.4, 29.3, 29.2, 27.3,
27.2, 24.7, 22.7, 22.6 (43C, CH2), 17.1, 17.0, 16.6 (CH3), 14.1 (4 C,
CH3); ESI HRMS for C90H146O15 [M+Na]+: Calcd 1490.0554, Found
1490.0468.
172.7, 172.6 (CO), 138.0, 137.5, 137.1 (Cq−Ar), 128.3, 128.0,
127.9, 127.7, 127.6, 127.4, 127.2 (15C, CH−Ar), 92.9 (C1′), 91.8 (C1),
79.5 (C3′), 76.8, 76.7 (C4′, C2′), 75.3 (C4), 75.2 (CH2−Ph), 74.6
(CH2−Ph), 74.0 (CH2−Ph), 71.5 (C3), 70.3 (C2), 69.0 (C5′), 68.4
(C5), 62.5 (C6′), 61.9 (C6), 33.9, 33.7, 33.6 (4C, CH2), 31.6 (4C,
CH2), 29.3, 29.2, 29.1, 29.0, 28.9, 28.8 (40C, CH2), 24.6, 24.5 (4C,
CH2), 22.3 (4C, CH2), 13.5 (4C, CH3); ESI HRMS for
C97H159NaO18S [M-Na]−: Calcd 1644.1253, Found 1644.1158.
Sodium 3′,4,4′-Tri-O-benzyl-3-O-[(S)-2-methyloctadecano-
yl]-6,6′-di-O-[(S)-2-methyldecanoyl]-2-O-palmitoyl-2′-O-sul-
fate-α,α-D-trehalose (15b). To a solution of the desilylated
compound 14b (55 mg, 0.038 mmol) in dry pyridine (1 mL) was
added [SO3:pyridine] (61 mg, 0.190 mmol, 5 equiv). The reaction
mixture was stirred for 3 h at 90 °C and then cooled to room
temperature. The reaction was quenched with methanol (1.0 mL) and
stirred until the precipitate disappeared. The mixture was concentrated
under vacuum and coevaporated with toluene and dichloromethane.
The residue was subjected to flash chromatography on silica gel
(CH2Cl2/MeOH: 90/10), and the product-containing fractions were
passed through an Amberlite IR-120 column (Na+ form) with the use
of the same solvent system before being concentrated. The expected
O-sulfated compound 15b (55 mg, 93%) was obtained as a colorless
syrup. [α]20D = +57 (c 0.9, CHCl3/CH3OH: 1/1); 1H NMR (CDCl3/
CD3OD: 3/2, 360 MHz) δ (ppm): 7.42−7.17 (m, 15H, Har), 5.64 (t,
J2,3 = J3,4 = 9.7 Hz, 1H, H3), 5.50 (d, J1′,2′ = 3.0 Hz, 1H, H1′), 5.21 (d,
J1,2 = 3.8 Hz, 1H, H1), 5.12 (d, J = 10.4 Hz, 1H, CH2−Ph), 4.94 (dd,
J1,2 = 3.8 Hz, J2,3 = 9.7 Hz, 1H, H2), 4.84 (d, J = 11.2 Hz, 1H, CH2−
Ph), 4.70 (d, J = 10.4 Hz, 1H, CH2−Ph), 4.61−4.30 (m, 7H, CH2−Ph
(3H), H2′, H6b, H5, H6a), 4.19−4.16 (m, 2H, H6′b, H6′a), 4.06 (t, J2′,3′
=
J
3′,4′ = 9.4 Hz, 1H, H3′), 3.88 (m, 1H, H5′), 3.75 (t, J3,4 = J4,5 = 9.7 Hz,
1H, H4), 3.48 (t, J3′,4′ = J4′,5′ = 9.4 Hz, 1H, H4′), 2.55−2.41 (m, 1H),
2.40−2.28 (m, 2H), 2.27−2.19 (m, 2H), 1.71−1.04 (m, 93H), 0.89−
0.78 (m, 12H); 13C NMR (CDCl3/CD3OD: 3/2, 90 MHz) δ (ppm):
177.8, 177.6, 176.5, 173.3 (CO), 138.9, 138.3, 137.9 (Cq−Ar),
129.0, 128.7, 128.6, 128.2, 128.1, 128.0 (15C, CH−Ar), 93.5 (C1′),
92.4 (C1), 80.3 (C3′), 77.7 (C4′), 77.6 (C2′), 76.4 (C4), 75.9 (CH2−
Ph), 75.5 (CH2−Ph), 74.7 (CH2−Ph), 72.1 (C3), 71.3 (C2), 69.8
(C5′), 69.2 (C5), 63.2 (C6′), 62.7 (C6), 40.1, 40.0, 39.9 (CH), 34.4,
34.3, 34.2, 34.1, 32.3, 32.2, 30.1, 30.0, 29.9, 29.8, 29.7, 29.6, 27.7, 27.6,
27.5, 25.1, 23.0 (43C, CH2), 17.4, 17.1, 16.8 (CH3), 14.3 (4C, CH3);
ESI HRMS for C90H145Na2O18S [M+Na]+: Calcd 1591.9942, Found
1591.9963.
Sodium 2,3,6,6′-Tetra-O-palmitoyl-2′-O-sulfate-α,α-D-treha-
lose (16). To a solution of compound 15a (20.7 mg, 0.012 mmol) in a
1/1 mixture of CH2Cl2/MeOH (1.0 mL) were added 32 mg of 5%
Pd/C. The suspension was vigorously stirred under 1 atm of H2 for 2
h. The resulted mixture was then filtered through Celite, rinsed with
methanol, and concentrated under reduced pressure. The residue was
subjected to flash chromatography (CH2Cl2/MeOH: 90/10 then 80/
20), and the product-containing fractions were passed through an
Amberlite IR-120 column (Na+ form) with the use of the same solvent
system before being concentrated. The expected sulfolipid 16 (15.0
Sodium 3′,4,4′-Tri-O-benzyl-2,3,6,6′-tetra-O-palmitoyl-2′-O-
sulfate-α,α-D-trehalose (15a). To a solution of compound 14a
(21.9 mg, 0.014 mmol) in dry pyridine (0.3 mL) was added
[SO3:pyridine] (23 mg, 0.070 mmol, 5 equiv). The reaction mixture
was stirred for 3 h at 90 °C and then cooled to room temperature. The
reaction was quenched with methanol (1 mL) and stirred until the
precipitate disappeared. The mixture was concentrated in vacuo and
mg, 86%) was obtained as a syrup. [α]24 = +39 (c 0.8, CHCl3/
D
H
dx.doi.org/10.1021/jo4012255 | J. Org. Chem. XXXX, XXX, XXX−XXX

The Journal of Organic Chemistry
Article
CH3OH: 50/50); 1H NMR (CDCl3/CD3OD: 1/1, 250 MHz) δ
(ppm): 5.15 (d, J1′,2′ = 3.6 Hz, 1H, H1′), 5.14 (dd, J2,3 = 10.2, J3,4 = 9.9
Hz, 1H, H3), 4.94 (d, J1,2 = 3.7 Hz, 1H, H1), 4.60 (dd, J1,2 = 3.7 Hz, J2,3
(4) Goren, M. B. Biochim. Biophys. Acta 1970, 210, 127−138.
(5) Goren, M. B.; Brokl, O.; Das, B. C.; Lederer, E. Biochemistry
1971, 10, 72−81.
= 10.2 Hz, 1H, H2), 4.09 (m, 2H, H6b, H6a), 4.04−3.86 (m, 4H, H6′b
,
(6) Goren, M. B.; Brokl, O.; Roller, P.; Fales, H. M.; Das, B. C.
Biochemistry 1976, 15, 2728−2735.
(7) Gilleron, M.; Stenger, S.; Mazorra, Z.; Wittke, F.; Mariotti, S.;
H6′a, H5, H2′), 3.61 (t, J2′,3′ = J3′,4′ = 9.0 Hz, 1H, H3′), 3.52 (m, 1H,
H5′), 3.33 (t, J3,4 = J4,5 = 9.9 Hz, 1H, H4), 3.06 (t, J3′,4′ = J4′,5′ = 9.0 Hz,
1H, H4′), 2.10−1.98 (m, 8H, CH2), 1.35−1.24 (m, 8H, CH2), 1.10−
0.90 (m, 96H, CH2), 0.57 (t, J = 6.8 Hz, 12H, CH3); 13C NMR
(CDCl3/CD3OD: 1/1, 90 MHz) δ (ppm): 174.1, 173.8, 173.3, 172.4
(CO), 91.6 (C1′), 90.9 (C1), 76.0 (C2′), 71.9 (C3), 71.2 (C3′), 70.2
(2C, C2, C4′), 69.8 (C5′), 69.4 (C5), 68.0 (C4), 63.1 (C6′), 61.7 (C6),
33.8, 33.6, 33.5, 33.4 (CH2), 31.4 (4C, CH2), 29.1, 29.0, 28.9, 28.8,
28.7, 28.6, 28.5 (40C, CH2), 24.5, 24.4, 24.3 (4C, CH2), 22.1 (4C,
CH2), 13.2 (4C, CH3); ESI HRMS for C76H141NaO18S [M+Na]+:
Calcd 1419.9629, Found 1419.9536.
Bohmer, G.; Prandi, J.; Mori, L.; Puzo, G.; De Libero, G. J. Exp. Med.
̈
2004, 199, 649−659.
(8) Guiard, J.; Collmann, A.; Gilleron, M.; Mori, L.; De Libero, G.;
Prandi, J.; Puzo, G. Angew. Chem., Int. Ed. 2008, 47, 9734−9738.
(9) Layre, E.; Cala-De Paepe, D.; Larrouy-Maumus, G.; Vaubourgeix,
J.; Mundayoor, S.; Lindner, B.; Puzo, G.; Gilleron, M. J. Lipid Res.
2011, 52, 1098−1110.
(10) Geerdink, D.; ter Horst, B.; Lepore, M.; Mori, L.; Puzo, G.;
Hirsch, A. K. H.; Gilleron, M.; de Libero, G.; Minnaard, A. J. Chem. Sci.
2013, 4, 709−716.
(11) For a recent review on trehalose-based glycolipids, see: Khan, A.
A.; Stocker, B. L.; Timmer, M. S. M. Carbohydr. Res. 2012, 356, 25−
36.
Sodium 3-O-[(S)-2-Methyloctadecanoyl]-6,6′-di-O-[(S)-2-
methyldecanoyl]-2-O-palmitoyl-2′-O-sulfate-α,α-D-trehalose
(1). To a solution of compound 15b (43 mg, 0.027 mmol) in a 1/1
mixture of CH2Cl2/MeOH (2 mL) were added 66 mg of 5% Pd/C.
The suspension was vigorously stirred under 1 atm of H2 for 2 h. The
resulted mixture was then filtered through Celite, rinsed with MeOH,
and concentrated under reduced pressure. The residue was subjected
to flash chromatography (dichloromethane/methanol: 90/10 then 80/
20), and the product-containing fractions were passed through an
Amberlite IR-120 column (Na+ form) with the use of the same solvent
system before being concentrated. The expected sulfolipid 1 (20 mg,
56%) was obtained as a syrup [α]24D = +57 (c 1.0, CHCl3/CH3OH: 1/
1); 1H NMR (CDCl3/CD3OD: 1/1, 360 MHz) δ (ppm): 5.42 (d, J1′,2′
= 3.6 Hz, 1H, H1′), 5.41 (dd, J2,3 = 10.1 Hz, J3,4 = 9.7 Hz, 1H, H3), 5.21
(d, J1,2 = 3.6 Hz, 1H, H1), 4.90 (dd, J1,2 = 3.6 Hz, J2,3 = 10.1 Hz, 1H,
H2), 4.37 (m, 2H, H6b, H6a), 4.28 (m, 1H, H5), 4.25−4.18 (m, 3H,
H6′b, H6′a, H2′), 3.91 (t, J2′,3′ = J3′,4′ = 9.0 Hz, 1H, H3′), 3.81 (m, 1H,
H5′), 3.58 (t, J3,4 = J4,5 = 9.7 Hz, 1H, H4), 5.36 (dd, J3′,4′ = 9.0, Hz, J4′,5′
= 9.7 Hz, 1H, H4′), 2.53−2.39 (m, 3H), 2.32−2.24 (m, 2H), 1.69−
1.49 (m, 5H), 1.44−1.04 (m, 88H), 0.85 (t, J = 7.0 Hz, 12H, CH3);
13C NMR (CDCl3/CD3OD: 1/1, 90 MHz) δ (ppm): 178.3, 178.0,
(12) Leigh, C. D.; Bertozzi, C. R. J. Org. Chem. 2008, 73, 1008−1017.
(13) For some recent synthetic studies on substituted trehalose
derivatives, see for example: (a) Patel, M. K.; Davis, B. G. Org. Biomol.
Chem. 2010, 8, 4232−4235. (b) Sarpe, V. A.; Kulkarni, S. S. J. Org.
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̈
Kurzchalia, T. V.; Knolker, H.-J. Synlett. 2011, 2482−2486. (d) Backus,
̈
K. M.; Boshoff, H. I.; Barry, C. S.; Boutureira, O.; Patel, M. K.;
D’Hooge, F.; Lee, S. S.; Via, L. E.; Tahlan, K.; Barry, C. E.; Davis, B. G.
Nat. Chem. Biol. 2011, 7, 228−235. (e) Paul, N. K.; Twibanire, J.-d. A.
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(15) Bourdreux, Y.; Lemetais, A.; Urban, D.; Beau, J.-M. Chem.
Commun. 2011, 47, 2146−2148.
(16) For the selective removal of benzyl groups in the presence of
benzylidene acetals using ammonium formate as a hydrogen donor,
see: (a) Bieg, T.; Szeja, W. Synthesis 1985, 76−77. (b) Beaupere, D.;
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155.
177.6, 173.4 (CO), 92.6 (C1′), 91.9 (C1), 77.0 (C2′), 72.7 (C3), 72.2
(C3′), 71.3 (C4′), 71.2 (C2), 70.7 (C5′), 70.5 (C5), 69.2 (C4), 64.1
(C6′), 62.7 (C6), 40.2, 40.1, 40.0 (CHCH3), 34.4, 34.3, 32.4, 30.3, 30.2,
30.1, 30.0, 29.9, 29.8, 27.8, 27.7, 25.2, 23.1 (43C, CH2), 17.3, 17.2,
17.1 (3C, CH3), 14.3 (4C, CH3); ESI HRMS for C69H127O18S [M-
Na]−: Calcd 1275.8749, Found 1275.8764.
(17) For a recent review, see: ter Horst, B.; Feringa, B. L.; Minnaard,
A. J. Chem. Commun. 2010, 46, 2535−2547.
(18) Studte, C.; Breit, B. Angew. Chem., Int. Ed. 2008, 47, 5451−5455.
In this work, the enantiomeric excess of tBu ester 9a was determined
by chiral GC after conversion of the reduced ester to the acetate.
(19) Brand, G. J.; Studte, C.; Breit, B. Org. Lett. 2009, 11, 4668−
4670.
(20) For the enantiomeric excesses measurements, the same strategy
was applied to the racemic lactate triflate (rac-8) leading to racemic
compounds (rac)-9a and (rac)-9b.
(21) Sakagami, M.; Hamana, H. Tetrahedron Lett. 2000, 41, 5547−
5551.
ASSOCIATED CONTENT
■
S
* Supporting Information
Copies of 1H and 13C NMR spectra for new compounds, details
on anisotropic NAD NMR, composition of oriented sample,
further NAD spectra, and extra discussion. This material is
available free of charge via the Internet at http://pubs.acs.org.
AUTHOR INFORMATION
Corresponding Author
*E-mail: philippe.lesot@u-psud.fr; jean-marie.beau@u-psud.fr.
Notes
(22) Sarfati, M.; Lesot, P.; Merlet, D.; Courtieu, J. Chem. Commun.
2000, 2069−2081 and references therein.
■
(23) Lesot, P.; Lafon, O.; Courtieu, J.; Berdague,
́
P. Chem.Eur. J.
2004, 10, 3741−3746.
(24) Ziani, L.; Lesot, P.; Meddour, A.; Courtieu, J. Chem. Commun.
2007, 4737−4739.
The authors declare no competing financial interest.
(25) Lesot, P.; Courtieu, J. Prog. Nucl. Magn. Reson. Spectrosc. 2009,
55, 128−159.
ACKNOWLEDGMENTS
■
(26) Chiral entities due to the natural D/H isotope substitution.
(27) Lesot, P.; V. Baillif, V.; Billault, I. Anal. Chem. 2008, 80, 2963−
2972.
The authors thank the Minister
grant to AL, the CNRS and the Institut Universitaire de France
̀
e de la Recherche for a Ph.D.
(IUF) for continuous financial support.
(28) Serhan, Z.; I. Billault, I.; Borgogno, A.; Ferrarini, A.; Lesot, P.
Chem.Eur. J. 2012, 18, 117−126.
REFERENCES
(29) Lesot, P.; Z. Serhan, Z.; C. Aroulanda, C.; Billault, I. Magn.
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■
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(30) See Figures S3 to S6 in the Supporting Information.
(31) In the Supporting Information are provided various anisotropic
NAD 1D/2D spectra of 9b recorded under the same conditions as 9a.
(32) See Figure S12 in the Supporting Information.
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