Synthesis of a structural analogue of the repeating unit from streptococcus pneumoniae 19F capsular polysaccharide based on the cross-metathesis- selenocyclization reaction sequence

Article abstract of DOI:10.1021/jo4001146

Pseudo-oligosaccharides have attracted much interest as scaffolds for the synthesis of sugar mimics endowed with very similar biological properties but structurally and synthetically simpler than their natural counterparts. Herein, the synthesis of pseudo-oligosaccharides using the cross-metathesis reaction between distinct sugar-olefins followed by intramolecular selenocyclization of the obtained heterodimer as key steps is first investigated. This methodology has been then applied to the preparation of structural analogues of the trisaccharide repeating unit from Streptococcus pneumoniae 19F. The inhibition abilities of the synthetic molecules were evaluated by a competitive ELISA assay using a rabbit polyclonal anti-19F serum.

Full text of DOI:10.1021/jo4001146

Article
pubs.acs.org/joc
Synthesis of a Structural Analogue of the Repeating Unit from
Streptococcus pneumoniae 19F Capsular Polysaccharide Based on the
Cross-Metathesis−Selenocyclization Reaction Sequence#
Paolo Ronchi,† Catalina Scarponi,‡ Matteo Salvi,† Silvia Fallarini,§ Laura Polito,∥ Enrico Caneva,⊥
Luana Bagnoli,*,‡ and Luigi Lay*,†
†Dipartimento di Chimica and ISTM-CNR, Universita
̀
degli Studi di Milano, via Golgi 19, I-20133 Milano, Italy
‡Dipartimento di Chimica e Tecnologia del Farmaco, Sezione di Chimica Organica, Universita
Perugia, Italy
̀
di Perugia, via del Liceo 1, I-06123
§DISCAFF, Universita
̀
del “Piemonte Orientale Amedeo Avogadro”, Via Bovio 6, I-28100 Novara, Italy
∥CNR-ISTM, via Fantoli 16/15, I-20138 Milano, Italy
⊥Centro Interdipartimentale Grandi Apparecchiature (CIGA), via Golgi 19, I-20133 Milano, Italy
S
* Supporting Information
ABSTRACT: Pseudo-oligosaccharides have attracted much interest
as scaffolds for the synthesis of sugar mimics endowed with very
similar biological properties but structurally and synthetically simpler
than their natural counterparts. Herein, the synthesis of pseudo-
oligosaccharides using the cross-metathesis reaction between distinct
sugar-olefins followed by intramolecular selenocyclization of the
obtained heterodimer as key steps is first investigated. This
methodology has been then applied to the preparation of structural
analogues of the trisaccharide repeating unit from Streptococcus
pneumoniae 19F. The inhibition abilities of the synthetic molecules
were evaluated by a competitive ELISA assay using a rabbit polyclonal
anti-19F serum.
carbon bonds.5 In particular, during the past decade, there has
INTRODUCTION
■
been an enormous growth of the metathesis area due to the
availability of robust and well-defined ruthenium alkylidene-
based precatalysts, endowed with high reactivity, air-stability,
and impressive functional group tolerance.5a,6
Carbohydrates feature an enormous structural diversity, which
makes them the most important mediators in the intercellular
interactions and with extracellular matrix components, such as
enzyme, hormones, toxins, bacteria, viruses, etc.1 It follows that
oligo- and polysaccharides play a fundamental role in signal
transduction and vital molecular recognition phenomena, thus
offering exciting new therapeutic opportunities in biomedical
fields.2 Nevertheless, the development of saccharide-based
drugs using classical carbohydrate synthesis can still be a
difficult task,3 despite the excellent progresses made by modern
carbohydrate chemistry.4 Sugar mimics endowed with very
similar biological properties, but structurally and synthetically
simpler than their natural counterparts, may offer a valuable
alternative. Thus, there is a strong demand for general and
efficient approaches to these structures that would allow
circumvention of the typical challenges associated with
oligosaccharide synthesis, such as laborious protecting group
manipulations and the lack of a general method for the
stereoselective formation of glycosidic linkages.
Although most applications of olefin metathesis employ the
more entropically favored ring-closing metathesis (RCM), the
advent of the new catalysts led to increasing applications of the
cross-metathesis (CM) reaction in synthetic organic chemistry.5
However, there are relatively few examples of selective cross-
metatheses applied to carbohydrate chemistry, and they are
mostly limited to the homodimerization of O-allyl and C-allyl
glycosides or their cross-linking with various types of aliphatic
and aromatic olefins.7 On the other hand, the selective cross-
coupling between unlike sugar olefins to provide the
corresponding heterodimer is a very attractive process, because
the diversity of accessible carbohydrate derivatives is much
higher compared to simple homodimerization. With the
purpose to further explore the potential of CM reaction in
the field of carbohydrate chemistry, we recently described a
preliminary investigation on the synthesis of pseudo-oligosac-
Among the most recent advances in synthetic organic
chemistry, transition metal-catalyzed olefin metathesis has
undoubtedly gained a prominent role as a highly promising
and valuable methodology for the construction of carbon−
Received: January 17, 2013
Published: May 8, 2013
© 2013 American Chemical Society
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Scheme 1. Synthetic Strategy
charides employing the cross-metathesis reaction between sugar
olefins followed by intramolecular cyclization.8
In our early investigation, the use of molecular iodine to
promote the electrophilic cyclization afforded iodocyclized type
D compounds (Scheme 1, E = I) with poor stereoselectivity
and as an inseparable mixture of diastereoisomers, thus
implying that the monosaccharide units do not exert a
significant asymmetric induction on the iodocyclization
reaction course.8 Moreover, the 1,2-elimination of iodine to
provide the endocyclic C−C double bond (compound E)
required very harsh reaction conditions (DBU in refluxing
toluene, 5−7 days), not applicable to substrates containing
base-sensitive functional groups.
In the quest for different promoters that could improve the
stereoselectivity of the cyclization and/or facilitate the
separation of the type D diastereoisomers (Scheme 1), we
considered the use of electrophilic organoselenium reagents.
The selenocyclization reaction has been frequently applied to
unsaturated alcohols9 and acids.10 Of particular importance is
the possibility of performing either reagent-controlled11 or
substrate-controlled12 asymmetric cyclization reactions from
which enantiomerically enriched or enantiopure heterocycles
can be obtained. In addition, once incorporated, the selenium
moiety can be easily converted into different functional groups
or eliminated.13 The best known and widely used syn
elimination of selenoxides14 would introduce a new endocyclic
C−C double bond that lends itself to further synthetic
transformations.
Figure 1. Structures of the trisaccharide repeating unit of S.
pneumoniae 19F and synthetic analogues 1a/1b.
RESULTS AND DISCUSSION
■
Synthesis of Pseudotrisaccharides by Selenocycliza-
tion. To the best of our knowledge, while the use of selenium
nucleophiles has been widely explored in carbohydrate
chemistry, only a few synthetic applications of selenium
electrophiles in this area have been reported.15 So our initial
effort was devoted to explore the behavior of different
electrophilic selenium reagents (compounds 2−4, Figure 2),
Figure 2. Structures of the selenium reagents employed in the present
work.
In the present paper, type E pseudo-oligosaccharides are
achieved by a strategy based on heterodimerization by CM
reaction of suitably elaborated monosaccharides A and B,
followed by intramolecular selenocyclization of intermediate C
and 1,2-elimination of the electrophile (Scheme 1).
in the selenocyclization reaction on heterodimers 5, 6, and 7
(Scheme 2), prepared as previously described and fully
characterized by assignment of the R configuration at C-78
(corresponding to S configuration at C-5 in tricyclic
compounds 8−19).
Because type E compounds can be seen as potentially useful
scaffolds for the synthesis of mimics of naturally occurring,
biologically important carbohydrates, we decided to check the
generality and feasibility of our methodology on more complex
and functionalized molecular target of biological relevance. We
therefore report herein on the synthesis of the pseudotrisac-
charides 1a/1b (Figure 1) as potential mimics of the
trisaccharide repeating unit of the capsular polysaccharide
(CPS) from the Gram+ bacterium Streptococcus pneumoniae
19F. Finally, the relative affinities of synthetic compounds 1a/
1b (differing in their configurations at C-3 and C-4; see Figure
1) were investigated in comparison with the natural
trisaccharide repeating unit and native 19F polysaccharide
(positive control) by a competitive ELISA assay using a rabbit
polyclonal anti-19F serum.
All the selenium reagents reported in Figure 2 were able to
promote the cyclization of our substrates, though with different
efficiencies.
Preliminary experiments were carried out on diastereoiso-
merically pure heterodimer 5 using phenylselenyl sulfate 2
(Figure 2) as a promoter, that was easily produced in situ by
the reaction of diphenyl diselenide with ammonium persulfate
in acetonitrile at 80 °C (Table 1, entry 1) or at room
temperature by the addition of trifluoromethanesulfonic acid to
a mixture of diphenyl diselenide and ammonium persulfate
(Table 1, entry 2).16 In both cases we obtained the two
diastereoisomers 8 and 9 (Scheme 2) in high to excellent yields
with a moderate diastereoisomeric ratio (1:2). The best
diastereoisomeric ratio was however achieved performing the
selenocyclization with N-(phenylseleno)phthalimide (N-PSP)
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a
Scheme 2. Synthesis of Pseudotrisaccharides by a Selenocyclization−Elimination Sequence
a
Key: (a) See Table 1; (b) H2O2, MeOH, rt, then NaHCO3, benzene, 80 °C.
a
strong dipolar interaction was observed between H-5 and H-2,
indicating the 5,2-syn relationship in the minor diaster-
eoisomer, while in pseudotrisaccharide 9 the dipolar interaction
between H-5 and H-3 confirmed their 5,2-anti relative
configuration in the major diastereoisomer. This allowed the
assignment of S configuration at C-2 of the major isomer 9, and
the R configuration at the same carbon of the minor isomer 8.
The selenocyclization of heterodimer 6 was carried out using
N-(phenylseleno)phthalimide, but an inseparable mixture of
diastereoisomers was obtained (Table 1, entry 5). The use of
(TIPPSe)2SO4 4 provided a mixture 1:1.9 of two diaster-
eoisomers 10 and 11 (Scheme 2, Ar = TIPP), that were easily
separated by column chromatography (Table 1, entry 6).
Although both diastereoisomers were fully characterized by
NMR, we could not determine the absolute configuration at C-
2, due to overlapping of the signals corresponding to H-5 and
H-2 protons (both in CDCl3 and C6D6). However, the
structure assignment for compounds 10 and 11 was later
deduced on the basis of the NMR characterization of the
corresponding elimination products 16 and 17 (see below).
The cyclization of heterodimer 7 with the selenium reagents
2, 3, and 4 (Figure 2) led in all cases to modest yields and
stereoselectivities. The best result was achieved using phenyl-
selenyl sulfate (Table 1, entry 9), that afforded a 1:2.3 ratio of
the cyclized products 12 and 13 in 45% yield (Scheme 2, Ar =
Ph). The stereochemical structures reported in Scheme 2 were
derived by a tentative attribution comparing the 13C NMR
chemical shifts of C-3 and C-4 with the corresponding values
measured for the same carbons in compounds 8 and 9, whose
configurations were unambiguously determined.18
All the selenocyclized pseudotrisaccharides 8−13 were next
subjected to the elimination reaction by oxidation with
hydrogen peroxide in methanol at room temperature (Scheme
2). The subsequent syn elimination occurred by refluxing the
obtained selenoxides in benzene containing 10% of sodium
hydrogencarbonate. In all cases the elimination products were
purified by column chromatography and fully characterized by
NMR spectroscopy. Following this procedure, compounds 14
and 15 were obtained from pseudotrisaccharides 8 and 9 in
77% and 96% yield, respectively. 1D NOE experiment carried
out in C6D6 on compound 14 showed a strong dipolar
interaction between H-5 and H-2, confirming their syn
relationship previously determined on 8. On the other hand,
NMR analysis by rotating-frame Overhauser spectroscopy (2D
ROESY experiment) of compounds 16 and 17, derived by
Table 1. Selenocyclization of Heterodimers 5, 6, and 7
b
Se
exptl
conditions
cyclic ether
(diast ratio)
b
c
d
entry heterodimer species
yield, %
82
1
2
3
4
5
6
7
8
9
5
5
5
5
6
6
7
7
7
2
2
3
4
3
4
3
4
2
method A,
15 min
8/9 (1/2)
8/9 (1/2)
8/9 (1/3)
8/9 (1/1.2)
method B,
30 min
92
79
70
60
79
23
45
45
method C,
1 h
method D,
5 h
e
method C,
1 h
10/11 (1:3.4)
10/11 (1:1.9)
method D,
1 h
method C,
1 h
12/13 (1/1.3)
12/13 (1/1.8)
12/13 (1/2.3)
method B,
1 h
method A,
1 h
a
Method A: (PhSe)2, (NH4)2S2O8, CH3CN, 80 °C; method B:
(PhSe)2 or (TIPPSe)2, (NH4)2S2O8, CF3SO3H, CH3CN, rt; method
C: N-PSP, BF3·OEt2, CH2Cl2, 0 °C to rt; method D: (TIPPSe)2,
(NH4)2S2O8, CF3SO3H, CH3CN, rt. See Scheme 2 for chemical
b
c
d
structures. See Figure 2 for Se species. Diasteroisomeric ratios were
calculated from the 1H NMR spectra of the crude mixture by
e
integration. Only in this case was an inseparable mixture of
diastereoisomers obtained.
3 (Figure 2) in the presence of BF3·OEt2 (Table 1, entry 3). On
the other hand, the use of the more sterically demanding (2,4,6-
triisopropylphenyl)selenium sulfate (TiPPSe)2SO4 4 (Figure
2), derived by addition of ammonium persulfate to the
corresponding diselenide17 in the presence of trifluorometha-
nesulfonic acid in acetonitrile, led to a reasonable yield but poor
diastereoselectivity (Table 1, entry 4). The obtained cyclic
ethers were separated by column chromatography and
examined by NMR spectroscopy using mono- and bidimen-
sional (H,H-COSY, HMQC) techniques. In all cases we
observed the exclusive formation of 2,3-anti diastereoisomers,
in agreement with the widely accepted mechanism of selenium
electrophilic addition to alkenes, i.e., the intramolecular
nucleophilic attack of the hydroxy group occurs by the back-
side at the bridged selenonium ion, giving overall anti addition.
In particular, the structures of tricyclic compounds 8 and 9 (Ar
= Ph, Scheme 2) were assigned on the basis of NOESY
experiments (see Supporting Information). In compound 8 a
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oxidative elimination of compounds 10 (90% yield) and 11
(98% yield), respectively, allowed the determination of the
absolute configuration at C-2, which was unfeasible for the
selenocyclized precursors (see Supporting Information for a
detailed NMR analysis).
Scheme 3. Retrosynthetic Strategy for the Synthesis of the
Pseudotrisaccharide 1
Finally, oxidative elimination of compounds 12 and 13
afforded pseudotrisaccharides 18 and 19 in 77% and 93% yield,
respectively (Scheme 2). An analogous trend with fully
characterized elimination products 14 and 15 was observed
1
in the H NMR analysis of compounds 18 and 19, further
supporting our previous attributions of the absolute config-
uration at C-2 indicated in Scheme 2 on selenocyclized
precursors 12 and 13.19
Altogether these results indicate that, in comparison to the
iodocyclization, the use of selenium electrophiles for the
cyclization reaction leads to higher, even if moderate,
stereoselectivity. Moreover, the possibility to change the R
group in the selenium electrophile (RSe+) permits in all cases
the separation of the two diastereoisomers. A second apparent
and more convincing advantage is the very mild conditions
required for the elimination step, which allows us to
considerably extend the feasibility of the selenium protocol
also on highly functionalized and delicate substrates.
a
Scheme 4. Synthesis of Alcohol 23
Synthesis of a Structural Analogue of the Trisacchar-
ide from S. pneumoniae 19F Capsular Polysaccharide.
The Gram-positive bacterium Streptococcus pneumoniae20 is a
major cause of death and disability throughout the world,21
causing meningitis, otitis, septicemia, and pneumonia, both in
developed and in developing countries.22 Although vaccination
against pneumococcal diseases has become a global health
priority,23 between 800 000 and one million children under five
years of age still die annually from Streptococcus pneumoniae
infections.24 Serotype 19F of Streptococcus pneumoniae, whose
CPS is composed of (→4)-β-D-ManpNAc-(1→4)-α-D-Glcp-
a
Key: (a) TrCl, TEA, CH2Cl2, 81%; (b) vinylMgBr, CuI, THF, −30
°C; (c) PMBBr, NaH, DMF; (d) PTSA, MeOH, 88% over three steps.
procedure reported by Mulzer and co-workers,29 alcohol 26
was obtained in enantiopure form. The 2-OH was then
protected as a p-methoxybenzyl (PMB) ether (compound 27),
and the subsequent hydrolysis of the trityl group with a
catalytic amount of p-toluensulfonic acid (PTSA) in methanol
cleanly furnished alcohol 23 in 88% yield over three steps.
Glycosylation of alcohol 23 with orthoester 24 was carried
out using TMSOTf as acidic promoter at −40 °C and afforded
a mixture of 2-OH and 2-OAc derivatives, which was subjected
to deacetylation under Zemplen conditions to provide
glucoside 28 in 66% yield over two steps (Scheme 5).
The 2-OH of 28 was then activated as a triflate with
trifluoromethanesulfonic anhydride in dichloromethane at −40
°C, followed by nucleophilic displacement with freshly
prepared tetrabutylammonium azide in toluene at 60 °C,
providing β-2-azidomannopyranoside 29. Finally, removal of
−
(1→2)-α-L-Rhap-(1-OPO3 →) repeating units (Figure 1), is
one of the most commonly isolated serotypes causing
pneumococcal disease. Some of us have a long established
experience on the synthesis of S. pneumoniae 19F CPS
fragments25 and structural analogues.26 We therefore consid-
ered the 19F trisaccharide repeating unit the ideal candidate as
a proof-of-concept for our synthetic methodology illustrated in
Scheme 1 to verify whether it could be a potentially useful
approach to target biologically important oligosaccharides.
As outlined in Scheme 3, we envisaged compound 20 as a
suitable precursor of our synthetic target 1. Synthesis of
pseudotrisaccharide 20 could be achieved by CM reaction
between sugar olefins 21 and 22 followed by electrophilic
cyclization and elimination.
The synthesis of the rahmnoside unit 22 was accomplished
by sequential silylation (thexyldimethylsilyl chloride and
imidazole in CH2Cl2) and 2-O-allylation (AllBr, NaH in
DMF) of 3,4-di-O-benzyl-L-rhamnopyranose, which in turn was
easily obtained from known 3,4-di-O-benzyl-1,2-O-(1-methox-
yethylidene)-β-L-rhamnopyranose.27
a
Scheme 5. Synthesis of Mannoside 21
The β-mannoside 21 can be derived by glycosylation with
the glucose-derived orthoester 2428 of the enantiopure alcohol
23, which in turn can be prepared starting from the
commercially available (R)-glycidol.
Our synthesis of alcohol 23 began with the protection of the
(R)-glycidol hydroxyl as triphenylmethyl (trityl, Tr) ether
followed by epoxide ring-opening with an organocopper
reagent generated in situ from vinylmagnesium bromide and
copper(I) iodide at −30 °C (Scheme 4). According to the
a
Key: (a) 23, TMSOTf, CH2Cl2, −40 °C ; (b) Na, MeOH, 66% over
two steps; (c) Tf2O, Py, CH2Cl2, −40 °C, then Bu4NN3, toluene, 60
°C; (d) DDQ, H2O:CH2Cl2 19:1, 55% over three steps.
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a
Scheme 6. Assembly of Pseudotrisaccharides 1a/1b
a
Key: (a) 5 mol % Hoveyda’s catalyst, refluxing CH2Cl2, 77%; (b) 10 mol % Hoveyda’s catalyst, CH2Cl2; 94% (c) PPh3, THF, then H2O, Ac2O,
MeOH, 80% over two steps; (d) Ph2Se2, (NH4)2S2O8, CH3CN, 80 °C, 75%; (e) 34, H2O2, MeOH, rt, then NaHCO3, C6H6, 80 °C, 88%; (f) OsO4,
NMO, H2O:acetone 1:1, 62%; (g) TBAF, THF; (h) H2 over 10% Pd/C: 60% (1a), 48% (1b), over two steps.
the PMB group with 2,3-dichloro-5,6-dicyano-1,4-benzoqui-
none (DDQ) afforded fragment 21 in 55% yield over three
steps.
However, the reaction did not occur, and unreacted substrate
20 was fully recovered. We conjectured that the presence of
bulky substituents on both allylic positions (C-2 and C-5) of
the five-membered ring might hamper the correct folding of the
substrate into the phathalazine pocket active site. On the other
hand, dihydroxylation with catalytic OsO4 and N-methylmor-
pholine-N-oxide (NMO) afforded syn-diols 35/36 in 62%
overall yield as a 3:2 mixture of diastereoisomers. The
challenging separation of the two stereoisomers was achieved
by HPLC on a chiral column (250 × 10 mm ID). The absolute
configuration at the newly formed stereogenic centers at C-3
and C-4 of 35 and 36 was determined for both compounds by
detailed NMR analysis (see Supporting Information).
Eventually, diols 35 and 36 were fully deprotected by
desilylation of the rhamnose unit with TBAF in THF followed
by hydrogenolysis of the benzyl ethers, affording pseudotri-
saccharides 1a and 1b in 60% and 48% yield over two steps,
respectively. Complete NMR characterization of compound 1a
confirmed absolutely our previous structural assignments
performed on protected precursor 35. Because compound 1b
was obtained from 36 in a very small amount (about 1 mg), it
was identified only by HRMS analysis (ESI source).
ELISA Assay. The ability of increasing concentrations (from
10−7mg/mL to 100 mg/mL) of the pseudotrisaccharides 1a and
1b to inhibit the binding between the 19F polysaccharide,
coated onto plates and used as positive control, and the anti-
19F human polyclonal antibody was evaluated by competitive
ELISA assay. The inhibition capacity of the synthetic
compounds was determined in comparison with the natural
trisaccharide repeating unit previously synthesized by our
group.25 Table 2 shows the relative efficacy of each compound
calculated by measuring the maximum effect elicited in this
system (relative efficacy), while the concentration that produces
50% of the maximum effect (EC50) was taken as indirect index
of its relative potency. As expected, the 19F native
polysaccharide exhibited both higher efficacy (100% of
With both sugar olefins 21 and 22 in our hands, the stage
was set for their condensation via cross-metathesis reaction.
The results of our previous study8 suggested that the two-step
procedure (self-metathesis followed by cross-metathesis, SM-
CM) developed by Grubbs et al.30 is a favorable approach over
the classical, straightforward CM reaction.31 Accordingly,
fragment 21 was first homodimerized using 5 mol % of
Hoveyda’s catalyst6c,d in refluxing dichloromethane, affording
homodimer 30 in 77% yield (3:1 E/Z ratio, Scheme 6). The
isomers mixture of 30 was then metathesized with fragment 22
using 10 mol % of Hoveyda’s catalyst in refluxing dichloro-
methane affording the heterodimer 31 in excellent yield (94%)
with 9:1 E/Z ratio. The major E stereoisomer was purified by
column chromatography and used in the following steps.
Reduction of the azide on 31 was carried out under
Staudinger’s condition32 followed by N-acetylation (MeOH,
Ac2O), to afford heterodimer 32 in 80% yield over two steps.
The subsequent intramolecular cyclization was accomplished
using in situ-generated phenylselenyl sulfate 2 (Figure 2) and
furnished compounds 33/34 as a 1:2 mixture of diaster-
eoisomers in 75% yield. After careful separation by medium
pressure chromatography on silica gel, NOESY experiments
were performed on the two diastereoisomers. In the major
diastereoisomer 34 the dipolar interaction between H-2 and H-
4 confirmed the 2,4-syn and 2,5-anti relative configurations.
Only the major diastereoisomer 34 was subjected to the
oxidative elimination step to yield compound 20 in 88% yield
(Scheme 6).
The next stage of our endeavor has been the dihydroxylation
of the C−C double bond in the inner ring of pseudotri-
saccharide 20. In a first attempt, the dihydroxylation reaction
was carried out using either AD-α or AD-β mix according to the
Sharpless protocol33 to obtain a single pure diastereoisomer.
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warmed to 80 °C, and after 30 min heterodimer (1 equiv) was added
and the reaction was further stirred until its complete consumption
(TLC hexane:diethyl ether 1:1). The solution was poured into a
saturated solution of aq NaHCO3 and extracted twice with CH2Cl2.
The organic layers were collected, dried (Na2SO4) and evaporated to
dryness. The crude was purified by medium pressure silica gel column
chromatography affording the pure diastereoisomers.
Method B: To a stirred solution of diphenyl diselenide (0.5 equiv)
in acetonitrile were added ammonium persulfate (1 equiv) and a
catalytic amount of trifluoromethanesulfonic acid. After 30 min, the
heterodimer (1 equiv) was added and the reaction was further stirred
until its complete consumption (TLC hexane:diethyl ether 1:1). The
solution was poured into a saturated solution of aq NaHCO3 and
extracted twice with CH2Cl2. The organic layers were collected, dried
(Na2SO4), and evaporated to dryness. The crude was purified by
medium pressure silica gel column chromatography, affording the pure
diastereoisomers.
Method C: Heterodimer (1 equiv) was dissolved in dry CH2Cl2
(0.01 M) and cooled to 0 °C. N-Phenylselenylphthalimide (1.4 equiv)
and a catalytic amount of BF3·Et2O were added. The reaction was
slowly warmed to room temperature and stirred until complete
consumption of the heterodimer (TLC hexane:diethyl ether 1:1). The
solution was diluted in diethyl ether and washed twice with 5% NaOH
aq solution. The organic layer was dried (Na2SO4) and evaporated to
dryness. The crude was purified by medium pressure silica gel column
chromatography, affording the pure diastereoisomers.
Method D: To a stirred solution of bis(2,4,6-triisopropylphenyl)
diselenide [(TIPP)2Se2] (0.5 equiv) in acetonitrile were added
ammonium persulfate (1 equiv) and a catalytic amount of
trifluoromethanesulfonic acid. After 30 min, the heterodimer (1
equiv) was added and the reaction was further stirred until its
complete consumption (TLC hexane:diethyl ether 1:1). The solution
was poured into a saturated solution of aq NaHCO3 and extracted
twice with CH2Cl2. The organic layers were collected, dried (Na2SO4),
and evaporated to dryness. The crude was purified by medium
pressure silica gel column chromatography, affording the pure
diastereoisomers.
Table 2. Results of Competitive ELISA Assay
EC50
maximum inhibition
a
compound
19F polysaccharide
(mg/mL)
(%)
8.9 × 10−5
6.74 × 10−2
100
43
−
natural trisaccharide repeating unit
1a
1b
−
−
−
a
Measured at 1 mg/mL.
inhibition at 1 mg/mL) and affinity (EC50 = 8.9 × 10−5 mg/
mL) compared with that of the natural trisaccharide repeating
unit, confirming that high molecular weight polysaccharides
have a conformational specificity (conformational epitopes)
more suitable for antibody binding. On the contrary, both the
synthetic repeating unit analogues 1a and 1b did not exhibit
any inhibition of binding between 19F polysaccharide and the
specific antibody at all concentrations tested. This result
suggests that the replacement of the inner glucose moiety of the
trisaccharide with the five-membered ring strongly affects the
biological properties of the trisaccharide repeating unit.
CONCLUSION
■
Different pseudo-oligosaccharides were obtained by a strategy
based on the cross-metathesis reaction between distinct sugar-
olefins, followed by intramolecular cyclization of the obtained
heterodimer promoted by selenium electrophiles. The follow-
ing mild syn elimination of the selenocyclized products allowed
the introduction of a new endocyclic carbon−carbon double
bond, suitable for further functionalization and synthesis of
more elaborated molecular targets. The feasibility of our
methodology for target molecules of biological significance is
demonstrated by the synthesis of a structural analogue of the
trisaccharide repeating unit from the capsular polysaccharide of
Streptococcus pneumoniae 19F.
2-{Methyl-O-[2,3,4,6-tetra-O-benzyl-β-D-glucopyranosyl]}-5-
{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xylopyranosyl]}-3-
(phenylselenyl)tetrahydrofuran, 8/9. Compound 5 (1.07 g, 1.00
mmol) was cyclized following method C as described above, affording
a mixture of diastereoisomers in 1:3 dr. After workup, medium
pressure chromatography (hexane:diethyl ether 7:3 to 1:1 in gradient
elution) afforded 8 (242 mg) and 9 (726 mg) as pure diastereoisomers
in 79% overall yield.
Despite the poor stereoselectivity obtained in some trans-
formations, the described strategy is however a novel and
promising approach for future developments, as the use of
sugar partners featuring various structural motifs should allow
the introduction of further molecular diversity into the pseudo-
oligosaccharide products. Work is in progress in our
laboratories to further explore the scope and potential of this
methodology.
Minor Stereoisomer 8. Colorless oil; [α]25 = +15.2 (c 0.25,
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.30−7.07 (m, 40H), 4.93−
4.48 (m, 14H, OCH2Ph), 4.46 (d, J = 3.1 Hz, 1H), 4.27 (d, J = 7.8
Hz,1H), 4.19−4.14 (m, 1H), 4.03 (q, J = 4.8 Hz, 1H) 3.90 (dd, J = 4.6,
11.8 Hz, 1H), 3.86 (t, J = 9.2 Hz, 1H), 3.68−3.40 (m, 8H), 3.37−3.33
(m, 2H), 3.24 (t, 1H), 3.23 (s, 3H), 2.13−1.77 (m, 4H); 13C NMR
(100 MHz, CDCl3) δ 138.7, 138.6, 138.5, 138.4, 138.2, 138.1, 134.3,
129.1, 128.4−127.5, 103.9, 97.8, 84.5, 83.7, 82.0, 81.9, 81.7, 80.0, 75.7,
75.6, 75.5, 75.0, 74.9, 74.8, 74.6, 73.4, 73.3, 70.9, 68.8, 68.0, 55.1, 40.8,
39.5, 36.9; HRESI MS m/z calcd C73H78O12SeNa [M + Na]+
1249.4556, found 1249.4563, error 0.6 ppm.
EXPERIMENTAL SECTION
■
General Experimental Methods. All commercially available
reagents including dry solvents were used as received. Nonvolatile
materials were dried under high vacuum. Reactions were monitored by
thin-layer chromatography on precoated silica gel plates and visualized
by staining with a solution of cerium sulfate (1g) and ammonium
heptamolybdate tetrahydrate (27 g) in water (469 mL) and
concentrated sulfuric acid (31 mL). Flash chromatography was
performed on silica gel. NMR spectra were recorded at 300 K (unless
otherwise stated) on spectrometers operating at 400 and 500 MHz.
Proton chemical shifts are reported in ppm (δ) with the solvent
reference relative to tetramethylsilane (TMS) employed as the internal
standard (CDCl3 δ = 7.26 ppm). Carbon chemical shifts are reported
in ppm (δ) relative to TMS with the respective solvent resonance as
the internal standard (CDCl3, δ = 77.0 ppm). Optical rotations were
obtained on a polarimeter at 589 nm using a 5 mL cell with a length of
1 dm. High resolution mass spectra (HRMS) were performed with an
ESI source.
Major Stereoisomer 9. Colorless oil; [α]25 = −41.8 (c 0.25,
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 77.60−7.10 (m, 40H), 5.05−
4.55 (m, 14H, OCH2Ph), 4.46 (d, J = 7.9 Hz, 1H), 4.45 (d, J = 3.1 Hz,
1H), 4.25−4.17 (m, 1H), 4.15−4.10 (dt, J = 3.8, 7.9 Hz, 1H), 3.99
(dd, J = 3.8, 11.2 Hz, 1H), 3.93 (t, J = 9.4 Hz, 1H), 3.80−3.60 (m,
7H), 3.50−3.40 (m, 3H), 3.30 (s, 3H), 3.24 (t, J = 9.0 Hz, 1H), 2.48−
2.37 (m, 1H), 1.95−1.80 (m, 2H), 1.78−1.65 (m, 1H); 13C NMR
(100 MHz, CDCl3) δ 138.7, 138.6, 138.5, 138.3, 138.2, 138.1, 134.9,
129.1, 128.4−127.5, 103.6, 97.8, 84.7, 83.4, 82.2, 81.9, 81.8, 80.0, 76.2,
75.7, 75.6, 75.2, 74.9, 74.9, 74.7, 73.5, 73.3, 69.0, 68.8, 68.2, 55.2, 40.8,
39.6, 37.4; HRESI MS m/z calcd C73H78O12SeNa [M + Na]+
1249.4556, found 1249.4533, error 1.8 ppm.
Typical Procedures for Selenocyclization. Method A: To a
stirred solution of diphenyl diselenide (0.5 equiv) in acetonitrile was
added ammonium persulfate (1 equiv). The resulting solution was
2-{Methyl-[2,3,4,6-tetra-O-benzyl-1-deoxy-α-D-1-C-gluco-
pyranosyl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xylopyra-
5177
dx.doi.org/10.1021/jo4001146 | J. Org. Chem. 2013, 78, 5172−5183

The Journal of Organic Chemistry
Article
nosyl]}-3-(1,2-bis(2,4,6-triisopropylphenyl)selenyl)tetrahydro-
furan, 10/11. Compound 6 (1.05 g, 1.00 mmol) was cyclized
following method D as described above, affording a mixture of
diastereoisomers in 1:1.9 dr. After workup, medium pressure
chromatography (hexane:diethyl ether 7:3) afforded 10 (545 mg)
and 11 (182 mg) as pure diastereoisomers in 79% overall yield.
75.2, 73.6, 73.4, 73.3, 70.0, 68.6, 68.1, 55.2, 55.1, 40.8, 40.0, 37.6;
HRESI MS m/z calcd C67H74O12Se [M + Na]+ 1173.4243, found
1173.4222, error 1.8 ppm.
Typical Procedure for Elimination. Method E: Selenocyclized
compound (1 equiv) was dissolved in MeOH. Then H2O2 30% v/v (3
equiv) was added, and the mixture was stirred at room temperature.
The reaction was monitored by TLC (hexane:diethyl ether 6:4). When
the oxidation step was finished, the solvent was removed under
reduced pressure. The crude was dissolved in C6H6 (3 mL) and a
solution of NaHCO3 10% (m/m) was added (1 mL). The resulting
solution was heated at 80 °C. The reaction was monitored by TLC
(hexane:diethyl ether 6:4). After 3 h, the reaction was stopped and
poured into water. The organic phase was extracted with CH2Cl2,
dried (Na2SO4), and filtered, and the solvent was removed under
reduced pressure. The crude product was purified by medium pressure
flash chromatography (hexane:diethyl ether 6:4), affording elimination
compound.
Minor Stereoisomer 10. Colorless oil; [α]25 = +6.7 (c 0.65,
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.38−7.22 (m, 33H), 7.16−
7.11 (m, 2H), 7.03−7.01 (m, 2H), 5.00−4.76 (m, 7H, OCH2Ph),
4.68−4.57 (m, 5H, OCH2Ph), 4.48 (d, J = 3.8 Hz, 1H), 4.46 (d, J =
10.8 Hz, 1H), 4.43 (d, J = 10.8 Hz, 1H), 4.45−4.39 (m, 1H), 4.15−
4.07 (m, 2H), 3.92 (t, J = 9.3 Hz, 1H), 3.90 (sext, J = 6.7 Hz, 2H),
3.75−3.51 (m, 7H), 3.49 (dd, J = 3.5, 9.6 Hz, 1H), 3.35−3.29 (m,
1H), 3.27 (s, 3H), 3.25 (t, J = 9.3 Hz, 1H), 2.86 (sext, J = 6.9 Hz, 1H),
2.23 (ddd, J = 6.2, 6.6, 12.1 Hz,1H), 2.11 (dt, J = 4.2, 15.2 Hz, 1H),
2.02 (ddd, J = 5.5, 10.1, 15.2 Hz, 1H), 1.91−1.78 (m, 2H), 1.65−1.55
(m, 1H), 1.24 (d, J = 6.9 Hz, 6H), 1.21 (d, J = 6.8 Hz, 12H); 13C
NMR (100 MHz, CDCl3) δ 153.4, 149.8, 138.7, 138.4, 138.3, 138.1,
128.4−127.5, 125.6, 121.7, 97.8, 82.3, 82.0, 81.9, 81.7, 80.0, 79.7, 75.7,
75.3, 75.2, 75.1, 74.9, 73.4, 73.2, 72.5, 71.5, 71.4, 68.7, 68.1, 55.1, 44.6,
40.8, 37.6, 34.4, 34.1, 28.2, 24.5, 23.9; HRESI MS m/z calcd
C82H96O11SeNa [M + Na]+ 1359.6049, found 1359.6011, error 2.8
ppm.
(2R,5S)-2-{Methyl-O-[2,3,4,6-tetra-O-benzyl-β-D-glucopyra-
nosyl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xylopyrano-
syl]}-2H,5H-dihydrofuran, 14. Compound 8 (100 mg, 0.082
mmol) was subjected to the elimination step following the typical
procedure (method E) reported above, affording compound 14 (67
mg, 0.063 mmol) in 77% yield as a colorless oil. [α]25D = +66.3 (c 0.4,
1
Major Stereoisomer 11. Colorless oil; [α]25 = +23.2 (c 1.05,
CHCl3); H NMR (400 MHz, CDCl3) δ 7.30−7.01 (m, 35H), 5.82
D
CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.42−7.21 (m, 33H), 7.20−
7.15 (m, 2H), 7.03−6.99 (m, 2H), 4.98 (d, J = 10.8 Hz, 1H), 4.97−
4.70 (m, 6H, OCH2Ph), 4.73−4.49 (m, 7H, OCH2Ph), 4.50 (d, J = 3.8
Hz, 1H), 4.47−4.40 (m, 1H), 4.29−4.21 (m, 1H), 4.01−3.84 (m, 4H),
3.80−3.60 (m, 6H), 3.60−3.58 (m, 1H), 3.52 (dd, J = 3.5, 9.8 Hz,
1H), 3.29 (s, 3H), 3.28 (t, J = 9.3 Hz, 1H), 3.16−3.10 (m, 1H), 2.86
(sext, J = 6.8 Hz, 1H), 2.15−2.01 (m, 1H), 2.0−1.81 (m, 4H), 1.81−
1.70 (m, 1H), 1.24 (d, J = 6.9 Hz, 6H), 1.21 (d, J = 6.8 Hz, 12H); 13C
NMR (100 MHz, CDCl3) δ 153.0, 149.7, 138.8, 138.7, 138.4, 138.3,
138.2, 138.1, 128.4 −127.5, 126.4, 121.7, 97.8, 82.3, 82.1, 81.8, 80.7,
80.1, 79.5, 75.7, 75.2, 75.1, 74.9, 73.5, 73.3, 72.4, 71.4, 70.8, 68.9, 68.1,
55.1, 45.5, 39.8, 37.3, 34.4, 34.0, 29.9, 24.6, 24.5, 23.9, 23.8; Anal.
Calcd for C82H96O11Se: C, 73.69; H, 7.24; O, 13.17; found: C, 73.73;
H, 7.28; O, 13.15; HRESI MS m/z calcd C82H96O11SeNa [M + Na]+
1359.6049, found 1359.6028, error 1.6 ppm.
(dt, J = 1.0, 5.7 Hz, 1H), 5.77 (dt, J = 1.0, 5.7 Hz, 1H), 4.95−4.90 (m,
2H), 4.89−4.41 (m, 14H, OCH2Ph), 4.47 (d, J = 3.5 Hz, 1H), 4.32 (d,
J = 7.8 Hz, 1H), 3.87 (t, J = 9.3 Hz, 1H), 3.81 (dd, J = 5.0, 10.0 Hz,
1H), 3.66−3.44 (m, 6H), 3.43 (dd, J = 3.5, 9.3 Hz, 1H), 3.36−3.32
(m, 2H), 3.28 (s, 3H), 3.22 (t, J = 9.5 Hz, 1H), 1.89−1.79 (m, 2H);
13C NMR (100 MHz, CDCl3) δ 138.7, 138.6, 138.5, 138.4, 138.2,
138.1, 131.4, 128.4−127.6, 121.6, 104.2, 97.9, 84.9, 84.6, 83.2, 82.1,
82.0, 81.7, 80.1, 75.7, 75.6, 75.1, 74.9, 74.8, 74.6, 73.6, 73.4, 73.3, 68.8,
67.7, 55.4, 38.8; HRESI MS m/z calcd C67H72O12Na [M + Na]+
1091.4916, found 1091.4879, error 3.4 ppm.
(2S,5S)-2-{Methyl-O-[2,3,4,6-tetra-O-benzyl-β-D-glucopyra-
nosyl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xylopyrano-
syl]}-2H,5H-dihydrofuran, 15. Compound 9 (100 mg, 0.08
mmol) was subjected to the elimination step following the typical
procedure (method E) reported above, affording compound 15 (82
mg, 0.077 mmol) in 96% yield as a colorless oil. [α]25D = +58.2 (c 0.95,
2-{Methyl-O-[methyl 2,3,6-tri-O-benzyl-α-D-glucopyrano-
syl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xylopyranosyl]}-
3-phenylselenyltetrahydrofuran, 12/13. Compound 7 (0.995 g, 1
mmol) was cyclized following method A, affording a mixture of
distereoisomers in 1:2.3 dr. After workup, medium pressure
chromatography (hexane:diethyl ether 7:3 to 1:1 in gradient elution)
afforded 12 (157 mg) and 13 (360 mg) as pure diastereoisomers in
45% overall yield.
1
CHCl3); H NMR (400 MHz, CDCl3) δ 7.45−7.10 (m, 35H), 5.90
(dt, J = 1.0, 6.1 Hz, 1H), 5.86 (dt, J = 1.0, 6.1 Hz, 1H), 5.10−5.01 (m,
2H), 5.00−4.52 (m, 14H, OCH2Ph), 4.51 (d, J = 3.6 Hz, 1H), 4.43 (d,
J = 7.7 Hz, 1H), 4.0 (dd, J = 4.3, 10.5 Hz, 1H), 3.95 (t, J = 9.3 Hz, 1H)
3.76−3.64 (m, 3H), 3.63−3.56 (m, 3H), 3.51 (dd, J = 3.5, 9.7 Hz,
1H), 3.47−3.43 (m, 2H), 3.32 (t, J = 9.0 Hz, 1H), 3.31 (s, 3H), 1.96
(ddd, J = 2.5, 6.0, 13.6 Hz, 1H), 1.82 (ddd, J = 5.0, 8.3, 13.6 Hz, 1H);
13C NMR (100 MHz, CDCl3) δ 138.7, 138.6, 138.5, 138.4, 138.2,
Minor Stereoisomer 12. Colorless oil; [α]25 = +12.8 (c 0.25,
D
1
CHCl3); H NMR (400 MHz, CDCl3) δ 7.43−7.40 (m, 2H), 7.35−
138.1, 138.0, 131.6, 128.4−127.6, 127.4, 104.0, 98.0, 84.6, 84.4, 83.3,
82.1, 82.0, 81.7, 80.0, 75.7, 75.6, 75.1, 75.0, 74.9, 74.6, 73.4, 73.3, 71.9,
68.9, 67.9, 55.3, 37.7; HRESI MS m/z calcd C67H72O12Na [M + Na]+
1091.4916, found 1091.4905, error 1.0 ppm.
7.15 (m, 33H), 4.98 (d, J = 10.8 Hz, 1H), 4.90,4.84 (2 d, each 1 H, J =
10.8 Hz, OCH2Ph), 4.82−4.75 (m, 3H, OCH2Ph), 4.70−4.50 (m, 6H,
OCH2Ph), 4.59 (d, J = 3.5 Hz, 1H), 4.51 (d, J = 3.7 Hz, 1H), 4.22−
4.12 (m, 1H), 3.96−3.84 (m, 4H), 3.73−3.55 (m, 5H), 3.52−3.48 (m,
2H), 3.41 (t, J = 9.1 Hz, 1H), 3.40−3.38 (m, 1H), 3.37 (s, 3H), 3.29
(s, 3H), 3.21 (t, J = 9.3 Hz, 1H), 2.07−1.98 (m, 1H), 1.96−1.78 (m,
3H); 13C NMR (100 MHz, CDCl3) δ 138.8, 138.7, 138.3, 138.2,
138.1, 134.3, 129.1, 128.4−127.5, 98.2, 97.8, 83.7, 81.9, 81.8, 80.0,
79.5, 78.1, 75.8, 75.7, 75.5, 75.0, 73.9, 73.4, 73.3, 73.2, 69.9, 68.6, 68.0,
55.2, 55.1, 40.5, 39.8, 36.9; HRESI MS m/z calcd C67H74O12Se [M +
(2R,5S)-2-{Methyl-[2,3,4,6-tetra-O-benzyl-1-deoxy-α-D-1-C-
glucopyranosyl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xy-
lopyranosyl]}-2H,5H-dihydrofuran, 16. Compound 10 (100 mg,
0.082 mmol) was subjected to the elimination step following the
typical procedure (method E) reported above, affording compound 16
(78 mg, 0.074 mmol) in 90% yield as a colorless oil. [α]25D = +100.0 (c
1
0.4, CHCl3); H NMR (400 MHz, CDCl3) δ 7.42−7.22 (m, 35H),
Na]+ 1173.4243, found 1173.4240, error 0.25 ppm.
5.98−5.94 (m, 1H), 5.85−5.79 (m, 1H), 4.95−4.93 (m, 1H), 4.95−
4.45 (m, 14H, 7 OCH2Ph), 4.95−4.93 (m, 1H), 4.55 (d, J = 3.8 Hz,
1H), 4.38−4.30 (m, 1H), 3.96 (t, J = 8.9 Hz, 1H), 3.80−3.55 (m, 7H),
3.52 (dd, J = 3.5, 9.6 Hz, 1H), 3.39 (s, 3H), 3.28 (t, J = 9.0 Hz, 1H),
2.12−1.93 (m, 4H); 13C NMR (100 MHz, CDCl3) δ 138.8, 138.7,
138.3, 138.2, 138.1, 138.0, 130.7, 129.9, 128.4−127.5, 97.9, 83.6, 82.6,
82.3, 82.0, 81.4, 80.0, 79.5, 78.1, 75.7, 75.4, 75.1, 75.0, 73.5, 73.3, 72.6,
71.6, 71.4, 69.2, 67.9, 55.4, 39.1, 32.3; HRESI MS m/z calcd
C67H72O11Na [M + Na]+ 1075.4967, found 1075.4961, error 0.6 ppm.
(2S,5S)-2-{Methyl-[2,3,4,6-tetra-O-benzyl-1-deoxy-α-D-1-C-
glucopyranosyl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xy-
25
Major Stereoisomer 13. Colorless oil; [α]
= +4.3 (c 0.25,
D
1
CHCl3); H NMR (400 MHz, CDCl3) δ 7.60−7.20 (m, 35H), 4.95
(d, J = 10.8 Hz, 1H), 4.88−4.73 (m, 5H, OCH2Ph), 4.70−4.46 (m,
6H, OCH2Ph), 4.57 (d, J = 3.5 Hz, 1H), 4.45 (d, J = 3.2 Hz, 1H),
4.11−4.03 (m, 1H), 4.03−3.95 (m, 1H), 3.94−3.80 (m, 3H), 3.70−
3.52 (m, 4H), 3.50−3.37 (m, 4H), 3.34 (s, 3H), 3.31−3.28 (m, 1H),
3.26 (s, 3H), 3.21 (t, J = 9.3 Hz, 1H), 2.36 (dt, J = 6.2, 12.5 Hz, 1H),
1.90−1.84 (m, 2H), 1.67 (dt, J = 9.4, 12.5 Hz, 1H); 13C NMR (100
MHz, CDCl3) δ 138.8, 138.7, 138.3, 138.2, 138.1, 134.9, 129.1, 128.5−
127.5, 98.2, 97.9, 82.9, 81.9, 81.6, 80.0, 79.5, 78.0, 75.8, 75.7, 75.6,
5178
dx.doi.org/10.1021/jo4001146 | J. Org. Chem. 2013, 78, 5172−5183

The Journal of Organic Chemistry
Article
lopyranosyl]}-2H,5H-dihydrofuran, 17. Compound 11 (150 mg,
0.123 mmol) was subjected to the elimination step following the
typical procedure (method E) reported above, affording compound 17
127.9, 127.2, 117.6, 70.3, 67.1, 38.2; HRESI MS m/z calcd C24H2402Na
[M + Na]+ 367.1669, found 367.1668, error 0.27 ppm.
(2S)-1-Trityloxy-2-(4-methoxybenzyloxy)-4-pentene, 27.
Compound 26 (14.46 g, 41.98 mmol) was dissolved in dry DMF
(50 mL) under nitrogen atmosphere. Under stirring, PMBBr (7.20
mL, 50.40 mmol) was added and then 95% NaH (1.16 g, 45.95 mmol)
in small amounts, and the stirring was continued overnight. The
reaction was monitored by TLC (hexane:ethyl acetate 95:5). The
reaction was quenched with MeOH after 18 h. DMF was evaporated
under reduced pressure, and the crude was dissolved in CH2Cl2 (100
mL) and washed with brine (2 × 50 mL). The organic layers were
collected, and dried (over Na2SO4), and the solvent was removed
under reduced pressure. An analytical sample of 27 was obtained by
(126 mg, 0.12 mmol) in 98% yield as a colorless oil. [α]25 = +9.3 (c
D
1
0.4, CHCl3); H NMR (400 MHz, CDCl3) δ 7.40−7.26 (m, 33H),
7.14−7.01 (m, 2H), 5.89−5.85 (m, 2H), 5.09−5.02 (m, 2H), 5.01 (d, J
= 10.8 Hz, 1H, 1/2 OCH2Ph), 4.95, 4.90 (2 d, each 1H, J = 10.8 Hz,
OCH2Ph), 4.86−4.78 (m, 4H, OCH2Ph), 4.71−4.60 (m, 5H,
OCH2Ph), 4.51 (d, J = 3.5 Hz, 1H), 4.45−4.40 (2 d, each 1H, J =
10.8 Hz, OCH2Ph), 4.24−4.18 (m, 1H), 3.97 (t, J = 9.3 Hz, 1H),
3.80−3.58 (7H), 3.54 (dd, J = 3.6, 9.7 Hz, 1H), 3.37 (s, 3H), 3.36 (t, J
= 9.4 Hz, 1H), 2.15−2.05 (m, 1H), 1.97 (ddd, J = 2.9, 5.9, 13.9 Hz,
1H), 1.91 (ddd, J = 3.1, 6.8, 14.8 Hz, 1H), 1.90−1.79 (m, 1H); 13C
NMR (100 MHz, CDCl3) δ 138.8, 138.7, 138.4, 138.3, 138.2, 138.0,
130.1, 129.6, 128.4−127.5, 98.0, 82.8, 82.6, 82.4, 82.1, 82.0, 80.0, 80.3,
78,4, 75.7, 75.4, 75.1, 75.0, 73.4, 73.3, 72.9, 71.4, 71.3, 68.8, 68.0, 55.3,
37.7, 31.0; HRESI MS m/z calcd C67H72O11Na [M + Na]+ 1075.4967,
found 1075.4970, error 0.3 ppm.
flash chromatography (hexane:ethyl acetate 95:5). [α]25 = −55.0 (c
D
1
1.0, CHCl3); H NMR (400 MHz, CDCl3) δ 7.41−7.39 (m, 6H),
7.23−7.14 (m, 11H), 6.79 (d, J = 8.6 Hz, 2H), 5.66 (ddt, J = 17.2,
10.2, 7.2 Hz, 1H), 4.98−4.89 (m, 2H), 4.52 (d, J = 11.3 Hz, 1H), 4.42
(d, J = 11.3 Hz, 1H), 3.73 (s, 3H), 3.55−3.48 (m, 1H), 3.13 (dd, J =
9.8, 3.9 Hz, 1H), 3.13 (dd, J = 9.3, 6.9 Hz, 1H),2.29−2.26 (m, 2H);
13C NMR (100 MHz, CDCl3) δ 159.2, 144.2, 134.8, 131.3, 129.4,
128.7, 127.7, 126.9, 116.9, 113.7, 86.6, 71.6, 65.5, 55.3, 36.6; HRESI
MS m/z calcd C32H32O3Na [M + Na]+ 487.2244, found 487.2239,
error 1.0 ppm.
(2R,5S)-2-{Methyl-O-[methyl 2,3,6-tri-O-benzyl-α-D-gluco-
pyranosyl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xylopyra-
nosyl]}-2H,5H-dihydrofuran, 18. Compound 12 (200 mg, 0.173
mmol) was subjected to the elimination step following the typical
procedure (method E) reported above, affording compound 18 (133
mg, 0.134 mmol) in 77% yield as a colorless oil. [α]25D =+8.1 (c 0.65,
(2S)-2-(4-Methoxybenzyloxy)-4-pentenol, 23. To a solution of
crude compound 27 in MeOH (100 mL) was added PTSA in a
catalytic amount. The reaction was monitored by TLC (hexane:ethyl
acetate 8:2). The reaction was stopped after 3 h by addition of solid
NaHCO3 and filtered over a short pad of Celite. The solvent was
removed under reduced pressure. The crude was purified by flash
chromatography (hexane:ethyl acetate 8:2), affording the alcohol 23
1
CHCl3); H NMR (400 MHz, CDCl3) δ 7.38−7.21 (m, 30H), 5.88
(d, J = 5.6 Hz, 1H), 5.65 (d, J = 5.6 Hz, 1H), 5.03−4.95 (m, 2H),
4.88−4.75 (m, 7H), 4.68−4.50 (m, 7H), 3.93−3.82 (m, 3H), 4.76−
4.64 (m, 3H), 4.60−4.62 (m, 1H), 3.51−3.43 (m, 4H), 3.36 (s, 3H),
3.35 (s, 3H), 3.22 (t, J = 9.4 Hz, 1H), 1.90−1.86 (m, 2H); 13C NMR
(100 MHz, CDCl3) δ 138.8, 138.7, 138.3, 138.2, 138.0, 131.4, 128.4−
127.5, 127.4, 98.2, 97.9, 84.6, 83.2, 82.0, 81.8, 81.7, 80.0, 79.6, 78.0,
76.2, 75.6, 75.5, 75.0, 73.4, 73.3, 73.2, 70.0, 68.5, 67.8, 55.4, 55.1, 38.9;
HRESI MS m/z calcd C61H68O12Na [M + Na]+ 1015.4608, found
1015.4626, error 1.8 ppm.
(7.83 g, 88% over three steps) as a colorless liquid. [α]25 = −4.5 (c
D
0.95, CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.29 (d, J=8.7 Hz, 2H),
6.90 (d, J = 9.0 Hz, 2H), 5.83 (ddt, J=15.5, 10.7, 7.2 Hz, 1H), 5.14 (d,
J= 15.5 Hz, 1H), 5.11 (d, J = 10.7 Hz, 1H), 4.62 (d, J= 11.1 Hz, 1H),
4.49 (d, J = 11.1, 1H), 3.89 (s, 3H), 3.69−3.67 (m, 1H), 3.59−3.52
(m, 2H), 2.44−2.32 (m, 2H); 2.10 (bs, 1H); 13C NMR (100 MHz,
CDCl3) δ 160.0, 134.8, 131.1, 130.1, 118.2, 114.6, 79.5, 71.9, 64.7,
55.9, 36.1. HRESI MS m/z calcd C13H1803Na [M + Na]+ 245.1148,
found 245.1140, error 3.3 ppm.
(2S,5S)-2-{Methyl-O-[methyl 2,3,6-tri-O-benzyl-α-D-gluco-
pyranosyl]}-5-{methyl-[methyl 2,3,4-tri-O-benzyl-α-D-xylopyra-
nosyl]}-2H,5H-dihydrofuran, 19. Compound 13 (250 mg, 0.217
mmol) was subjected to the elimination step following the typical
procedure (method E) reported above, affording compound 19 (200
mg, 0.202 mmol) in 93% yield as a colorless oil. [α]25D = +28.5 (c 1.35,
(2S)-2-(4-Methoxybenzyloxy)-1-(3,4,6-tri-O-benzyl-β-D-glu-
copyranosyl)-4-pentene, 28. Compounds 2428 (1.42 g, 2.82 mmol)
and 23 (2.97 g, 13.36 mmol) were dissolved with an equal amount of
dry CH2Cl2 (10 mL), under N2 atmosphere, and mixed together. Then
powdered 4 Å molecular sieves (4.4 g) were added to the resulting
solution and stirred at rt for 1 h. Then the mixture was cooled at −40
°C, and a solution of TMSOTf 0.17 M in CH2Cl2 (5 mL, 0.85 mmol)
was slowly added. The reaction was monitored by TLC (hexane:ethyl
acetate 7:3), and after 1.5 h, the reaction was neutralized by adding
TEA and the filtered through a short pad of Celite. The organic phase
was washed with brine and dried (over Na2SO4). Removal of the
solvent afforded a crude product as a mixture of 2-hydroxy and 2-O-
acetylated derivatives. This crude product was dissolved in dry MeOH
(30 mL), under N2 atmosphere, and a catalytic amount of sodium was
added. The reaction was monitored by TLC (hexane:ethyl acetate
4:1). The deacetylation reaction was stopped by neutralization with
Amberlite IR-120 resin (H+) form. The reaction was filtered, and the
solvent was removed under reduced pressure. Because the complete
chromatographic separation of 28 from unreacted 23 was unfeasible,
the crude product was subjected to tritylation (regioselective for
primary alcohols) using the same protocol employed for the synthesis
of 2529 to easily remove the excess of alcohol 23. In this way, the crude
product was purified by flash chromatography (hexane:ethyl acetate
4:1), affording the tritylated alcohol (27) and the glucoside 28 (1.21 g,
66% yield) as a colorless oil. [α]25D = +10.1 (c 0.95, CHCl3); 1H NMR
(400 MHz, CDCl3) δ 7.41−7.18 (m, 17H), 6.81 (d, J = 8.5 Hz, 2H),
5.85 (ddt, J= 17.2, 10.1, 7.0 Hz, 1H), 5.18−5.08 (m, 2H), 4.96 (d, J =
11.2 Hz, 1H), 4.87−4.82 (m, 2H), 4.64−4.51 (m, 5H), 4.32 (d, J = 7.1
Hz, 1H), 3.99 (dd, J = 3.5, 10.6 Hz, 1H), 3.81 (s, 3H), 3.77−3.69 (m,
3H), 3.65−3.57 (m, 4H), 3.51−3.46 (m, 1H), 2.37−2.33 (m, 3H); 13C
NMR (100 MHz, CDCl3) δ 159.2, 138.7, 138.1, 134.2, 130.5, 129.5,
1
CHCl3); H NMR (400 MHz, CDCl3) δ 7.25−7.18 (m, 30H), 5.77
(d, J = 6.0 Hz, 1H), 5.49 (d, J = 6.0 Hz, 1H), 4.95−4.87 (m, 1H), 4.82,
4.79 (2 d, each 1H, J = 11.0 Hz, OCH2Ph), 4.73−4.66 (m, 5H), 4.57−
4.47 (m, 5H), 4.42−4.38 (m, 1H), 3.83 (t, J = 9.0 Hz, 1H), 3.79 (t, J =
9.0 Hz, 1H), 3.72−3.53 (m, 5H), 3.43−3.31 (m, 5H), 3.28−3.23 (m,
7H), 3.20 (t, J = 9.2 Hz, 1H), 1.86 (ddd, J = 2.4, 7.0, 13.9 Hz, 1H),
1.74 (ddd, J = 5.6, 8.6, 14.0 Hz, 1H); 13C NMR (100 MHz, CDCl3) δ
138.9, 138.8, 138.3, 138.2, 138.1, 138.0, 131.6, 128.4−127.5, 126.9,
98.2, 98.0, 84.8, 82.9, 82.0, 81.9, 81.8, 80.0, 79.6, 78.2, 75.7, 75.6, 75.5,
75.1, 73.5, 73.4, 73.3, 70.0, 68.4, 67.8, 55.3, 55.1, 37.7; HRESI MS m/z
calcd C61H68O12Na [M + Na]+ 1015.4608, found 1015.4615, error 0.7
ppm.
(S)-1-Trityloxypent-4-en-2-ol, 26. Vinylmagnesium bromide (1
M) (63.30 mL, 63.30 mmol) in THF was added dropwise to a slurry
of CuI (1.21 g, 6.33 mmol) in dry THF (50 mL) at −40 °C. The
mixture was stirred for 30 min, and then a solution of 2529 (6.67 g,
21.08 mmol) in dry THF (50 mL) was added dropwise over 30 min.
The reaction was stirred at −40 to −30 °C for 1 h and then poured
into a cold mixture of sat. aq NH4Cl (100 mL), NH4OH (100 mL),
and Et2O (100 mL) with vigorous stirring. The phases were separated,
and the aqueous phase was further extracted with Et2O (3 × 100 mL).
The organic solution was washed with brine, dried (NaSO4), and
filtered through a short pad of silica gel. The solvent was removed
under reduced pressure. An analytical sample of 26 was obtained by
flash chromatography (hexane:ethyl acetate 9:1). [α]25 = +79.5 (c
D
1
0.55, CHCl3); H NMR (400 MHz, CDCl3) δ 7.47−7.25 (m, 15H),
5.77 (dd, J = 13.1, 8.8 Hz, 1H), 5.11−5.05 (m, 2H), 3.87−3.85 (m,
1H), 3.20 (dd, J =9.4 J 3.9 Hz, 1H), 3.11 (dd, J = 9.4 J 6.9, 1H), 2.28−
2.24 (m, 3H); 13C NMR (100 MHz, CDCl3) δ 143.9, 134.4, 128.7,
5179
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The Journal of Organic Chemistry
Article
128.4−127.7, 117.5, 113.8, 103.4, 84.4, 75.2, 75.1, 74.9, 73.5, 73.1,
71.9, 71.7, 69.7, 68.9, 55.3, 35.9; HRESI MS m/z calcd C40H4608Na
[M + Na]+ 677.3084, found 677.3078, error 0.9 ppm.
evaporated under reduced pressure, and the crude was dissolved in
CH2Cl2 (100 mL) and washed with brine (2 × 50 mL). The organic
phase was dried (over Na2SO4) and the solvent evaporated under
reduced pressure. The crude was purified by flash chromatography
(2S)-2-(4-Methoxybenzyloxy)-1-(2-azido-3,4,6-tri-O-benzyl-
2-deoxy-β-D-mannopyranosyl)-4-pentene, 29. To a solution of
compound 28 (0.500 g, 0.76 mmol) in dry dichloromethane (18 mL)
under a nitrogen atmosphere was added pyridine (250 μL, 3.05
mmol). The mixture was cooled to −40 °C, and after 15 min, triflic
anhydride (376 μL, 2.34 mmol) was slowly added dropwise. After
being stirred 6 h, the solvent was removed under vacuum without
heating. The crude 2-O-triflate intermediate was dissolved under a
(hexane:ethyl acetate 95:5), affording 22 (850 mg, 77% yield) as a
1
colorless oil. [α]25 = −37.2 (c 1.0, CHCl3); H NMR (400 MHz,
D
CDCl3) δ 7.41−7.29 (m, 10H), 5.96−5.87 (m, 1H), 5.29 (dd, J =18.0,
1.6, 1H), 5.20 (dd, J= 10.8, 1.8, 1H), 4.90 (1d, J= 10.9, 1H), 4.76 (d, J
=11.7 Hz,1H), 4.68−4.62 (m, 3H), 4.20−4.16 (m, 1H), 4.10 (bt, J=
2.3 Hz, 1H), 4.00−3.95 (m, 1H), 3.83(dd, J = 9.3, 2.7 Hz, 1H), 3.75−
3.68 (m, 1H), 3.57 (t, J = 9.4 Hz, 1H), 1.67 (hept, J= 6.9 Hz, 1H),
1.29 (d, J= 6.1 Hz, 3H), 0.95 (bt, J = 6.1 Hz, 6H), 0.89 (s, 6H), 0.14
(s, 3H), 0.14 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 134.1, 128.3−
127.3, 117.0, 99.6, 80.1, 80.1, 75.2, 72.1, 69.8, 68.4, 67.6, 34.3, 25.0,
20.5, 20.2, 18.8, 18.6, 18.1, −2.7, −2.8; HRESI MS m/z calcd
C31H46O5SiNa [M + Na]+ 549.3012, found 549.3005, error 1.3 ppm.
1-O-(2-Azido-3,4,6-tri-O-benzyl-2-deoxy-β-D-mannopyrano-
syl)-6-O-(thexyldimethylsilyl-3,4-di-O-benzyl-β-D-rhamnopyra-
nosyl)-2-hydroxy-4-hexenediol, 31. Homodimer 30 (400 mg, 0.37
mmol) was dissolved under argon atmosphere in dry dichloromethane
(10 mL), and the solution was warmed to 40 °C. Argon was bubbled
through the solution for 10 min. Hoveyda’s catalyst (11 mg, 0.018
mmol) was added. After 20 min, a solution of allyl derivative 22 (96
mg, 0.18 mmol) in dry dichloromethane (2 mL) was slowly added,
and the reaction was monitored by TLC (hexane:ethyl acetate 7:3).
After 6 h, DMSO (20 μL) was added and stirring was continued
overnight. The solvent was removed under reduced pressure, and the
crude was purified by flash chromatography (toluene:acetone 95:5),
−
nitrogen atmosphere in dry toluene (8 mL), a solution of Bu4N+N3
(1.085 g, 3.81 mmol) in dry toluene (5 mL) was added quickly, and
the reaction was stirred at 55 °C (hexane:ethyl acetate 7:3). After 24 h,
the reaction mixture was concentrated with a high vacuum pump. An
analytical sample of 29 was purified by flash chromatography. [α]25
=
D
1
+2.6 (c 1.2, CHCl3); H NMR (400 MHz, CDCl3) δ 7.31−7.19 (m,
15H), 7.11−7.09 (m, 2H), 6.79 (d, J = 8.5 Hz, 2H), 5.74 (ddt, J =
17.1, 10.1, 7.1 Hz, 1H), 5.03−4.97 (m, 2H), 4.77 (d, J = 10.8 Hz, 1H),
4.61−4.41 (m, 8H), 3.88 (dd, J = 10.6, 3.3 Hz, 1H), 3.89 (bd, J = 3.3
Hz, 1H), 3.70 (s, 3H), 3.69−3.59 (m, 4H), 3.51−3.42 (m, 2H), 3.31−
3.28 (m, 1H), 2.21 (bt, J=6.5 Hz, 2H); 13C NMR (100 MHz, CDCl3)
δ 138.2, 134.1, 137.6, 134.1, 130.9, 129.3, 128.6−127.6, 117.5, 113.8,
99.9, 80.9, 77.8, 75.7, 75.3, 74.4, 73.6, 72.6, 72.2, 69.1, 62.0, 55.3, 36.1;
HRESI MS m/z calcd C40H45N307Na [M + Na]+ 702.3150, found
702.3144, error 0.8 ppm.
(2S)-1-(2-Azido-3,4,6-tri-O-benzyl-2-deoxy-β-D-mannopyra-
nosyl)-4-penten-2-ol, 21. Crude compound 29 was dissolved in
CH2Cl2/H2O (12 mL, 19:1), and DDQ (190 mg, 0.84 mmol) was
added. The reaction was monitored by TLC (hexane:ethyl acetate
1:1), and after 2 h, the reaction was diluted with CH2Cl2, and the
organic layer was washed with brine and dried over Na2SO4. The
solvent was evaporated under reduced pressure, and the crude was
purified by flash chromatography (hexane:ethyl acetate 6:4), affording
affording compound 31 (180 mg, 94%) as a colorless oil. [α]25
=
D
+79.5 (c 0.55, CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.40−7.20 (m,
25H), 5.75 (dt, J = 15.9, 6.6 Hz, 1H), 5.67 (dt, J = 15.9, 5.4 Hz, 1H),
4.90 (bt, J = 10.1, 10.0 Hz, 2H), 4.78−4.71 (m, 3H), 4.67−4.54 (m,
7H), 4.13 (dd, J = 12.2, 5.1 Hz, 1H), 4.09 (m, 1H), 4.01 (d, J = 1.5 Hz,
1H), 3.94−3.81−3.61 (m, 10H), 3.58 (t, J = 9.4 Hz, 1H,), 3.48−3.44
(m, 1H), 2.77 (bs, 1H), 2.34−2.30 (m, 2H), 1.68 (hept, J = 6.9 Hz,
1H), 1.28 (d, J = 6.1 Hz, 3H), 0.96 (bt, J = 6.3 Hz, 6H), 0.89 (s, 6H),
0.13 (s, 3H), 0.11 (s, 3H); 13C NMR (100 MHz, CDCl3) δ 138.8,
138.6, 138.0, 137.9, 137.4, 128.6, 128.4−127.3, 100.3, 99.6, 80.8, 80.2,
75.6, 75.3, 75.2, 74.3, 73.6, 72.3, 72.1, 69.8, 61.8, 36.3, 34.4, 25.0, 20.5,
20.3, 18.2, 18.6, 18.1, −2.7, −2.8; HRESI MS m/z calcd
C61H79N3O11SiNa [M + Na]+ 1080.5376, found 1080.5360, error
1.5 ppm.
compound 21 (220 mg, 55%) as pale yellow solid. [α]25 = +52.0 (c
D
1
1.0, CHCl3); H NMR (400 MHz, CDCl3) δ 7.31−7.11 (m, 15H),
5.78 (ddt, J =17.2, 10.1, 7.1 Hz, 1H), 5.10−5.03 (m, 2H), 4.80 (d,
J=10.8 Hz, 1H), 4.67 (d, J =11.8 Hz, 1H), 4.62 (d, J =11.8 Hz, 1H),
4.56−4.45 (m, 4H), 3.93 (d, J= 3.0 Hz, 1H), 3.84−3.79 (m, 2H),
3.69−3.64 (m, 2H), 3.61−3.50 (m, 3H), 3.39−3.35 (m, 1H), 2.23 (d, J
= 4.9, 1H), 2.32 (bt, J = 6.8, 2H); 13C NMR (100 MHz, CDCl3) δ
138.0, 137.9, 137.4, 134.2, 128.4−127.9, 117.8, 100.3, 80.9, 75.6, 75.3,
74.4, 73.6, 72.3, 69.9, 69.0, 61.8, 37.8; HRESI MS m/z calcd
C32H37N306Na [M + Na]+ 582.2574, found 582.2562, error 2.0 ppm.
(2S,7S)-1,8-Bis(2-azido-3,4,6-tri-O-benzyl-2-deoxy-β-D-man-
nopyranosyl)-2,7-dihydroxyoctanediol, 30. Compound 21
(0.372 g, 0.667 mmmol) was dissolved in dry dichloromethane (15
mL), and argon was bubbled through the resulting solution for 10 min.
Hoveyda’s catalyst (20 mg, 0.033 mmol) was added, and the solution
was refluxed and monitored by TLC (hexane:ethyl acetate 2:3). After
3 h, DMSO was added and the stirring was continued overnight at
room temperature. The solvent was evaporated, and the crude product
was purified by flash chromatography (hexane:ethyl acetate 2:3),
1-O-(2-Acetammido-3,4,6-tri-O-benzyl-2-deoxy-β-D-manno-
pyranosyl)-6-O-(thexyldimethylsilyl 3,4-di-O-benzyl-β-D-rham-
nopyranosyl)-2-hydroxy-4-hexenediol, 32. To a solution of
compound 31 (220 mg, 0.21 mmol) in dry THF (3 mL) was added
PPh3 (163 mg, 0.62 mmol), and the solution was stirred at 60 °C. The
reaction was monitored by TLC (toluene:acetone 9:1). After 4 h, H2O
(100 μL) was added and the solution was stirred overnight. The
solvent was evaporated under reduced pressure, the crude amine was
dissolved in dry MeOH (3 mL), and Ac2O (77 μL, 0.83 mmol) was
added, stirred for 1 h, and then concentrated to dryness. The crude
was purified by flash chromatography (toluene acetone 95:5),
affording compound 32 (178 mg, 80% yield) as a colorless oil.
[α]25D = +64.1 (c 0.55, CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.40−
7.20 (m, 25H); 5.87 (d, J = 9.7 Hz, 1H), 5.74 (dt, J = 15.5, 8.2 Hz,
1H), 5.64 (dt, J = 15.5, 5.9 Hz, 1H), 4.92−4.84 (m, 4H), 4.73 (d, J =
11.7 Hz, 1H), 4.66−4.59 (m, 4H), 4.54−4.48 (m, 4H), 4.18−4.07 (m,
2H), 3.93−3.86 (m, 2H), 3.82−3.79 (m, 2H), 3.74−3.63 (m, 5H),
3.58−3.46 (m, 3H), 3.04 (bs, 1H), 2.29−2.23 (m, 2H), 2.08 (s, 3H),
1.66 (hept, J = 6.8 Hz, 1H), 1.27 (d, J = 6.2 Hz, 3H), 0.94 (bt, J = 6.3
Hz, 6H), 0.88 (s, 6H), 0.13 (s, 3H), 0.10 (s, 3H); 13C NMR (100
MHz, CDCl3) δ 171.5, 139.2, 139.0, 138.5, 138.2, 138.0, 128.8−127.5,
101.0, 100.0, 80.6, 75.6, 75.5, 75.1, 74.4, 76.0, 72.5, 71.6, 70.2, 69.1,
68.7, 67.7, 49.8, 36.5, 34.8, 25.3, 24.0, 20.7, 20.6, 19.1, 19.0, −2.3,
−2.4; HRESI MS m/z calcd C63H83NO12SiNa [M + Na]+ 1096.5577,
found 1096.5561, error 1.4 ppm.
1
affording 30 (E/Z ratio 3:1, 0.280 g, 77% yield) as yellow pale oil. H
NMR (400 MHz, CDCl3, E/Z mixture) δ 7.42−7.20 (m, 30H), 5.56−
5.58 (m, 2H), 4.88 (d, J = 10.8 Hz, 2H), 4.76 (d, J = 11.8 Hz, 2H),
4.71 (d, J = 11.8 Hz, 2H), 4.63−4.53 (m, J = 12.1 Hz, 8H), 4.02−4.01
(m, 2H), 3.92−3.64 (m, 12 H), 3.59 (dd, J = 6.7, 10.3 Hz, 2H), 3.47−
3.43 (m, 2H), 2.78 (bs, 2H), 2.28−2.24 (m, 4H); 13C NMR (100
MHz, CDCl3) δ 138.1, 138.0, 137.5, 129.0, 128.6−127.7, 100.2, 100.2,
80.9, 75.6, 75.3, 74.4, 73.5, 72.2, 70.1, 69.0, 61.8, 36.7; HRESI MS m/z
calcd C62H70N6012Na [M + Na]+ 1113.4944, found 1113.4928, error
1.4 ppm.
Thexyldimethylsilyl 2-O-Allyl-3,4-di-O-benzyl-β-D-rhamno-
pyranoside, 22. Thexyldimethylsilyl-3,4-di-O-benzylrahmnopyrano-
side27 (1.01 g, 2.08 mmol) was dissolved in dry DMF (10 mL) under
nitrogen atmosphere. Then AllBr (210 μL, 2.5 mmol) and NaH 95%
(60 mg, 2.3 mmol) were added. The resulting solution was stirred
overnight and monitored by TLC (hexane:ethyl acetate 95:5). The
reaction was quenched by addition of 100 μL of MeOH. DMF was
2-[Methyl-O-(2-acetammido-3,4,6-tri-O-benzyl-2-deoxy-β-D-
mannopyranosyl)]-5-methyl-O-(thexyldimethylsilyl 3,4-di-O-
benzyl-β-D-rhamnopyranosyl)-4-selenylphenyltetrahydrofur-
5180
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1
an, 33/34. To a stirred solution of 1,2-bis(phenyl) diselenide (32 mg,
0.10 mmol) in CH3CN (2 mL) at 80 °C was added (NH4)2S2O8 (22.8
mg, 0.10 mmol). After 30 min, compound 32 (214 mg, 0.17 mmol)
was added and the reaction mixture was stirred at 80 °C. The reaction
was monitored by TLC (hexane:diethyl ether 7:3). The reaction was
stopped after 1 h. The reaction mixture was poured into a saturated
aqueous solution of NaHCO3, and the organic phase was extracted
with CH2Cl2. The organic layer was dried (Na2SO4) and filtered, and
the solvent was removed under reduced pressure. The crude was
purified by medium pressure flash chromatography (hexane:diethyl
ether 7:3 to 1:1), affording selenocyclized compounds 33 (53 mg,
0.044 mmol) and 34 (106 mg, 0.087 mmol) in 75% overall yield and
1:2 dr as a colorless oil.
Compound 35. [α]25 = +15.3 (c 0.6, CHCl3); H NMR (500
D
MHz, CDCl3) δ 7.39−7.28 (m, 25H); 6.09 (d, J = 8.3 Hz, 1H), 5.10
(d, J = 8.0 Hz, 1H), 4.91−4.84 (m, 3H), 4.78 (d, J = 11.1 Hz, 1H),
4.73 (d, J = 11.7 Hz, 1H), 4.69 (d, J = 1.7 Hz, 1H), 4.65−4.60 (m,
3H), 4.55−4.57 (bs, 1H), 4.51−4.49 (m, 3H), 4.17 (m, 1H), 4.12−
4.08 (m, 3H), 4.01−3.99 (m, 1H), 3.93 (dd, J = 10.8, 4.6 Hz, 1H),
3.90 (dd, J = 10.5, 6.5 Hz, 1H), 3.80−3.67 (m, 8H), 3.65 (dd, J = 10.8,
6.7 Hz, 1H), 3.56 (t, J = 9.5 Hz, 1H), 3.46−3.44 (m, 1H), 2.07 (s,
3H), 1.66 (hept, J = 5.6 Hz, 1H), 1.27 (d, J = 6.2 Hz, 3H), 0.95−0.94
(bt, J = 6.3 Hz, 6H), 0.87 (s, 6H), 0.11 (s, 3H), 0.10 (s, 3H); 13C
NMR (125 MHz, CDCl3) δ 171.8, 138.8, 138.5, 138.1, 137.7, 137.4,
128.5−124.4, 100.9, 100.1, 80.1, 80.0, 79.7, 75.1, 74.7, 73.9, 73.8, 73.6,
72.1, 71.9, 71.2, 70.5, 69.4, 68.7, 68.5, 66.5, 49.4, 34.4, 24.9, 23.5, 20.4,
20.2, 19.8, 18.8, 18.6, 18.1, −2.7, −2.8; HRESI MS m/z calcd
C63H83NO14SiNa [M + Na]+ 1128.5475, found 1128.5507, error 2.8
Minor Stereoisomer 33. [α]25D = −25.5 (c 0.25, CHCl3); 1H NMR
(400 MHz, CDCl3) δ 7.56−7.54 (m, 2H), 7.30−7.26 (m, 25H), 7.18−
7.16 (m, 2H), 5.92 (d, J = 5.9 Hz, 1H), 4.90−4.83 (m, 4H), 4.67−4.43
(m, 9H), 4.23−4.20 (m, 1H), 4.03−3.98 (m, 2H), 3.79−3.42 (m,
13H), 2.03−1.98 (s, 3H), 1.64 (q, J = 6.4 Hz, 1H), 1.26 (s, 3H), 1.21
(d, J = 4.8 Hz, 3H), 0.92 (t, J = 3.6 Hz, 6H), 0.85 (s, 6H), 0.11 (s,
3H), 0.08 (s, 3H); HRESI MS m/z calcd C69H87NO12SeSiNa [M +
ppm.
1
Compound 36. [α]25 = −17.5 (c 0.3, CHCl3); H NMR (500
D
MHz, CDCl3, T = 27 °C) δ 7.39−7.28 (m, 25H), 5.97 (d, J = 8.3 Hz,
1H), 4.91−4.81 (m, 4H), 4.77−4.45 (m, 8H), 4.23−4.22 (m, 1H),
4.09−3.95 (m, 3H), 3.90−3.83 (m, 2H), 3.77−3.65 (m, 5H), 3.57−
3.54 (m, 2H), 3.45−3.41 (m, 1H), 2.05 (s, 3H), 1.66 (q, J = 4.5 Hz,
6H), 0.88 (s, 6H), 0.13 (s, 3H), 0.10 (s, 3H). 13C NMR (125 MHz,
CDCl3) δ 170.9, 138.8, 138.6, 138.0, 137.5, 128.8−127.6, 100.5, 100.5,
82.0, 80.2, 79.8, 79.7, 78.7, 75.1, 74.9, 74.8, 74.0, 73.7, 72.1, 71.4, 69.7,
68.5, 68.0, 67.9, 49.2, 34.4, 25.0, 23.7, 20.4, 20.2, 18.7, 18.6, 18.1, −2.6,
−2.8; HRESI MS m/z calcd C63H83NO14SiNa [M + Na]+ 1128.5475,
found 1128.5497, error 1.9 ppm.
Na]+ 1252.5088, found 1252.5116, error 2.2 ppm.
1
Major Stereoisomer 34. [α]25 = +1.9 (c 0.25, CHCl3); H NMR
D
(400 MHz, CDCl3) δ 7.57 (d, J = 8.0 Hz, 2H), 7.38−7.28 (m, 25H),
7.21 (d, J = 8.0 Hz, 2H), 5.99 (d, J = 9.4 Hz, 1H), 4.92−4.85 (m, 3H),
4.77 (d, J = 11.7 Hz, 1H), 4.72 (d, J = 11.7 Hz, 1H), 4.65−4.46 (m,
7H), 4.19−4.12 (m, 2H), 4.08−4.05 (m, 2H), 4.00−3.99 (m, 1H),
3.95−3.89 (m, 2H), 3.83−3.32 (m, 11H), 2.46 (q, J = 6.4 Hz, 1H),
2.04 (s, 3H), 1.91−1.83 (m, 1H), 1.65 (q, J = 6.8 Hz, 1H), 1.24 (d, J =
6.4 Hz, 3H), 0.93 (bt, J = 6.7 Hz, 6H), 0.87 (s, 6H), 0.12 (s, 3H), 0.09
(s, 3H); 13C NMR (100 MHz, CDCl3) 170.8, 138.8, 138.5, 138.2,
137.8, 134.8, 129.2, 128.4−127.4, 100.4, 99.6, 83.8, 80.5, 80.0, 79.8,
77.2, 75.2, 75.2, 75.0, 73.5, 72.0, 71.3, 69.6, 68.4, 67.7, 49.3, 40.2, 37.2,
34.3, 24.9, 23.6, 20.4, 20.2, 18.8, 18.6, 18.1, −2.3, −2.4; HRESI MS m/
z calcd C69H87NO12SeSiNa [M + Na]+ 1252.5088, found 1252.5059,
error 2.3 ppm.
(2S,5S)-2-[Methyl-O-(2-acetammido-3,4,6-tri-O-benzyl-2-
deoxy-β-D-mannopyranosyl)]-5-methyl-O-(thexyldimethylsilyl
3,4-di-O-benyl-β-D-rhamnopyranosyl)-2H,5H dihydrofuran, 20.
Compound 34 (100 mg, 0.081 mmol) was subjected to the
elimination step following method E reported above, affording
compound 20 (76 mg, 0.063 mmol) in 88% yield as a colorless oil.
[α]25D = +32.1 (c 0.55, CHCl3); 1H NMR (400 MHz, CDCl3) δ 7.39−
7.21 (m, 25H), 5.89−5.84 (m, 3H), 5.02−5.01 (m, 1H), 4.99−4.95
(m, 1H), 4.96−4.86 (m, 4H), 4.73 (d, J = 11.7 Hz, 1H), 4.66−4.59
(m, 6H), 4.53−4.46 (m, 3H), 4.14−4.12 (m, 1H), 3.85−3.75 (m, 4H),
3.72−3.53 (m, 6H), 3.48−3.40 (m, 2H), 2.06 (s, 3H), 1.67 (q, J = 6.8
Hz, 1H), 1.26 (d, J = 6.2 Hz, 3H), 0.94 (bt, J = 6.8 Hz, 6H), 0.87 (s,
6H), 0.12 (s, 3H), 0.10 (s, 3H); 13C NMR (100 MHz, CDCl3) δ
170.7, 138.8, 138.5, 138.2, 137.9, 137.8, 128.8−127.2, 100.5, 99.7, 85.4,
85.1, 80.5, 80.1, 80.0, 75.2, 75.0, 74.9, 74.0, 73.5, 72.0, 71.5, 71.3, 69.6,
69.5, 68.8, 68.4, 60.4, 49.3, 34.4, 29.7, 24.9, 23.6, 20.4, 20.2, 18.7, 18.6,
18.1, −2.3, −2.4; HRESI MS m/z calcd C63H81NO12SiNa [M + Na]+
1094.5420, found 1094.5439, error 1.7 ppm.
2-[Methyl-O-(2-acetammido-3,4,6-tri-O-benzyl-2-deoxy-β-D-
mannopyranosyl)]-5-methyl-O-(thexyldimethylsilyl 3,4-di-O-
benzyl-β-D-rhamnopyranosyl)-3,4-dihydroxytetrahydrofuran,
35/36. Compound 20 (40 mg, 0.037 mmol) was dissolved in
H2O:acetone (1.6 mL, 1:10), and the resulting solution was cooled to
0 °C. N-Methylmorpholine N-oxide (6 mg, 0.055 mmol) and a
solution of 2.5% OsO4 in tert-butyl alcohol (47 μL, 0.0037 mmol)
were slowly added dropwise, and the reaction was monitored by TLC
(toluene:acetone 7:3). After 3 h, the reaction was diluted with CH2Cl2
(5 mL) and washed with sat. aq NaHSO3 (5 mL). The organic layer
was dried over Na2SO4, and the solvent was evaporated under reduced
pressure to dryness, affording a mixture of diastereoisomers in 1:2 dr.
The diastereoisomers were completely separated by semipreparative
HPLC (chiral column 250 × 10 mm ID, hexane:2-propanol 7:3, flow 2
mL\min, λ 210 nm), affording compound 35 (12 mg, 0.011 mmol) as
a major diastereoisomer together with compound 36 (6 mg, 0.0054
mmol) in 62% overall yield.
(2S,3S,4R,5R)-2-[Methyl-O-(2-acetammido-2-deoxy-β-D-man-
nopyranosyl)]-5-methyl-O-(2-D-rhamnopyranosyl)-3,4-dihy-
droxytetrahydrofuran, 1a. Compound 35 (11.9 mg, 0.011 mmol)
was dissolved in dry THF (0.6 mL) under nitrogen atmosphere, in the
presence of 4 Å molecular sieves, and the resulting solution was cooled
to 0 °C. A 1 M TBAF solution in THF (0.16 mL, 0.16 mmol) was
slowly added, and the reaction mixture was stirred at room
temperature. The reaction was monitored by TLC (toluene:acetone
1:1). After the complete consumption of starting compound, 4 mL of
brine was added and the solution was extracted (3 × 10 mL) with
ethyl acetate. The organic layers were collected and dried (over
Na2SO4), and the solvent was evaporated under reduced pressure. The
crude product was purified by flash column chromatography on silica
gel (toluene:acetone 8:2 to 1:1 in gradient elution), affording
desilylated compound as an amorphous white solid.
This compound was dissolved in a MeOH/H2O mixture (0.7:0.7
mL), Pd/C catalyst (10%, 5 mg) was added, and the reaction mixture
was vigorously stirred under hydrogen at room temperature. After 48
h, a second portion of the catalyst (5 mg) was added and the mixture
was stirred under hydrogen for an additional 12 h. The reaction
mixture was diluted with MeOH/H2O, filtered over a Celite pad,
concentrated under reduced pressure to remove MeOH, and finally
lyophilized to give 1a as an amorphous solid (3.3 mg, 60%). 1H NMR
(500 MHz, D2O, T = 30 °C) δ 4.82 (d, J = 1.5 Hz, 1H), 4.80 (d, J =
1.7 Hz, 1H), 4.52 (dd, J = 1.5, 4.5 Hz, 1H), 4.23−4.21 (m, 2H), 4.17−
4.15 (m, 1H), 4.08 (dd, J = 2.5, 11.3 Hz, 1H), 4.02 (m, 1H), 3.96−
3.94 (m, 2H), 3.91 (J = 12.4, 2.3 Hz, 1H), 3.81 (dd, J = 4.6, 3.8 Hz,
1H), 3.80 (dd, J = 12.4, 5.9 Hz, 1H) 3.74 (dd, J = 9.7, 3.5 Hz, 1H),
3.73−3.68 (m, 2H), 3.59 (dd, J = 8.0, 10.6 Hz, 1H), 3.53 (t, J = 9.8 Hz,
1H), 3.43 (t, J = 9.7 Hz, 1H), 3.42−3.38 (m, 1H), 2.06 (s, 3H), 1.28
(d, J = 6.5 Hz, 3H); 13C NMR (125 MHz, D2O, T = 30 °C) δ 100.3,
99.7, 80.1, 79.6, 76.4, 72.0, 72.0, 71.8, 71.6, 70.0, 70.0, 69.5, 68.7, 66.8,
66.5, 60.5, 53.1, 22.1, 16.6; HRESI MS m/z calcd C20H35NO14Na [M
+ Na]+ 536.1950, found 536.1948, error 0.4 ppm.
(2S,3R,4S,5R)-2-[Methyl-O-(2-acetammido-2-deoxy-β-D-man-
nopyranosyl)]-5-methyl-O-(2-D-rhamnopyranosyl)-3,4-dihy-
droxytetrahydrofuran, 1b. Compound 36 (5.8 mg, 0.0052 mmol)
was fully deprotected as described for major diastereoisomer 35,
affording isomer 1b as an amorphous solid (1.3 mg, 48%). HRESI MS
m/z calcd C20H35NO14Na [M + Na]+ 536.1950, found 536.1956, error
1.1 ppm.
Competitive ELISA Assay. Flat-bottomed plates (96-well) were
incubated overnight at 4−8 °C with a mixture of S. pneumoniae 19F
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Article
CPS (1 mg/mL) and methylated human serum albumin (1 mg/mL).
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applied to the plates for blocking of nonspecific binding sites. The
plates were incubated overnight at 4−8 °C with a solution (1:200) of
rabbit anti-19F, used as reference serum. When compounds 1a and 1b
were tested, they were added to each well immediately before the
addition of the reference serum. The plates were then incubated with
alkaline phosphatase conjugate goat antirabbit IgG, stained with p-
nitrophenyl phosphate, and the absorbance was measured at 405 nm.
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ASSOCIATED CONTENT
■
(9) Tiecco, M.; Testaferri, L.; Tingoli, M.; Bartoli, D.; Balducci, R. J.
Org. Chem. 1990, 55, 429−434.
S
* Supporting Information
1H and 13C NMR spectra for all new compounds. This material
is available free of charge via the Internet at http://pubs.acs.org.
(10) Tiecco, M.; Testaferri, L.; Tingoli, M.; Bagnoli, L.; Santi, C.
Synlett 1993, 10, 798−799.
(11) (a) Tiecco, M.; Testaferri, L.; Bagnoli, L.; Marini, F.; Santi, C.;
Temperini, A.; Scarponi, C.; Sternativo, S.; Terlizzi, R.; Tomassini, C.
ARKIVOC (Gainesville, FL, U. S.) 2006, 186−206. (b) Tiecco, M.;
Testaferri, L.; Marini, F.; Sternativo, S.; Bagnoli, L.; Santi, C.;
Temperini, A. Tetrahedron: Asymmetry 2001, 12, 1493−1502.
(c) Tiecco, M.; Testaferri, L.; Bagnoli, L.; Scarponi, C.; Purgatorio,
V.; Temperini, A.; Marini, F.; Santi, C. Tetrahedron: Asymmetry 2005,
16, 2429−2435. (d) Tiecco, M.; Testaferri, L.; Bagnoli, L.; Scarponi,
C.; Temperini, A.; Marini, F.; Santi, C. Tetrahedron: Asymmetry 2006,
17, 2768−2774. (e) Tiecco, M.; Testaferri, L.; Bagnoli, L.; Purgatorio,
V.; Temperini, A.; Marini, F.; Santi, C. Tetrahedron: Asymmetry 2001,
12, 3297−3304.
(12) (a) Tiecco, M.; Testaferri, L.; Bagnoli, L.; Purgatorio, V.;
Temperini, A.; Marini, F.; Santi, C. Tetrahedron: Asymmetry 2004, 15,
405−412. (b) Tiecco, M.; Testaferri, L.; Bagnoli, L.; Terlizzi, R.;
Temperini, A.; Marini, F.; Santi, C.; Scarponi, C. Tetrahedron:
Asymmetry 2004, 15, 1949−1955. (c) Tiecco, M.; Testaferri, L.;
Bagnoli, L.; Scarponi, C.; Temperini, A.; Marini, F.; Santi, C.
Tetrahedron: Asymmetry 2007, 18, 2758−2767. (d) Tiecco, M.;
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2008, 19, 2411−2416.
AUTHOR INFORMATION
Corresponding Author
*E-mail: luigi.lay@unimi.it; bagnoli@unipg.it.
Notes
■
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We gratefully acknowledge the financial support from M.I.U.R.
(Ministero Italiano Universita
̀
e Ricerca), National Projects
PRIN2007 (prot. N. 2007FJC4SF_001) and PRIN2008 (prot.
N. 2008CZ3NP3), Fondazione Cassa di Risparmio: Project
2012.0114.021 “Processi ecosostenibili per la produzione e
l’analisi di molecole di interesse farmaceutico”, and Consorzio
CINMPIS.
DEDICATION
#Dedicated to the memory of Prof. Marcello Tiecco.
■
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