Synthesis of Ribonucleosidic Dimers with an Amide Linkage from D-Xylose

Article abstract of DOI:10.1021/acs.joc.6b01822

An original and efficient stereocontrolled synthesis of ribonucleosidic homo- and heterodimers has been achieved from inexpensive d-xylose. This successful strategy involved the sequential introduction of nucleobases, using two stereocontrolled N-glycosidation reactions, from a common two-furanoside amide-linked scaffold offering the possibility of obtaining any given base sequence. The pertinence of this approach is illustrated through the preparation of the homodimers UU-34 and TT-35 in 18 steps with an excellent overall yield of more than 10% from d-xylose, while the heterodimer route led to UT-39 in 19 steps with around 10% overall yield.

Full text of DOI:10.1021/acs.joc.6b01822

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Article
Synthesis of ribonucleosidic dimers with an amide linkage from D-xylose.
Laurence Arzel, Didier Dubreuil, Fabrice Dénès, Virginie
Silvestre, Monique Mathé-Allainmat, and Jacques Lebreton
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.6b01822 • Publication Date (Web): 21 Oct 2016
Downloaded from http://pubs.acs.org on October 21, 2016
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Synthesis of ribonucleosidic dimers with an amide linkage from D-xylose.
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Laurence Arzel, Didier Dubreuil, Fabrice Dénès, Virginie Silvestre, Monique MathéꢀAllainmat* and
Jacques Lebreton*
Université de Nantes, CEISAMꢀUMR CNRS 6230, Faculté des Sciences et des Techniques, 2 rue de la
Houssinière, BP 92208, 44322 Nantes Cedex 3, France.
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Eꢀmail: jacques.lebreton@univꢀnantes.fr
Eꢀmail: Monique.Mathe@univꢀnantes.fr
Abstract: An original and efficient stereocontrolled synthesis of ribonucleosidic homoꢀ and heterodimers
has been achieved from inexpensive Dꢀxylose. This successful strategy involved the sequential introduction
of nucleobases, using two stereocontrolled Nꢀglycosidation reactions, from a common twoꢀfuranoside
amideꢀlinked scaffold offering the possibility of obtaining any given base sequence. The pertinence of this
approach is illustrated through the preparation of the homodimers UUꢀ34 and TTꢀ35 in 18 steps with an
excellent overall yield of more than 10% from
with around 10% overall yield.
Dꢀxylose while the heterodimer route led to UTꢀ39 in 19 steps
Key words: Dꢀxylose, glycosylation, azide reduction, amide formation, ribonucleosidic dimers, antisense,
siRNA.
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1
-Introduction
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1
Since the pioneering work of Zamecnik and Stephenson major advances have been made in the
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development of chemically modified oligonucleotides as a novel strategy for drug discovery in various gene
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4,5
silencing technologies, such as antisense, antigene and RNA interference . In 1998, the first antisense
TM
drug, the phosphorothioate oligonucleotide Vitravene (ISIS 2922), was approved for the treatment of
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AIDSꢀrelated cytomegalovirus (CMV) retinitis. Five years later, a second antisense phosphorothioate
TM
oligonucleotide drug Kynamro
(ISIS 301012) was approved to treat homozygous familial
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hypercholesterolemia (HoFH), a genetic defect that leads to high cholesterol levels. Currently, more than 40
antisense therapies are in clinical trials for diseases such as cancer, diabetes and neurodegenerative
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diseases.
Figure 1. Natural phosphodiester linkage in natural DNA A and RNA B strands and amide linkages C and
D proposed as a substitute for the phosphodiester in oligodeoxynucleotides and oligoribonucleotides,
respectively.
In the last two decades, a large variety of chemically modified oligonucleotides have been designed,
synthesized and evaluated in order to improve properties such as nuclease resistance to degradation in
biological fluids, biodistribution and pharmacokinetics, while keeping the affinity and specificity towards
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the targeted RNA messenger or RNA interference. Among these, the oligonucleotides modified with nonꢀ
ionic phosphate linkage mimics offered various advantages, especially an increase in hydrophobicity due to
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the reduced global charge. In this context, earlier studies showed that the C3’ꢀCH ꢀCOꢀNHꢀ C5’
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internucleoside amide linkage (C, Figure 1), first synthesized and patented by De Mesmaeker and coꢀ
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workers at CIBA in 1992, does not disturb the thermal stability of DNA/RNA duplexes.
In fact, with
oligodeoxynucleotides, this amide linkage improved the binding affinity by WatsonꢀCrick base pairing for
RNA complements, as measured by increased duplex stability in thermal denaturation experiments (ꢀtm per
modification: +0.4 to +0.9 °C). Moreover, these modified oligodeoxynucleotides displayed remarkable
resistance to enzymatic degradation relative to reference strands. It was also reported that this modified
internucleoside amide linkage, such as in C, with an additional methyl substituent at C5’ having the S
configuration improved the thermal stability of duplexes with RNA (ꢀtm per modification: +1.0 to +1.4 °C)
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compared to the R configuration (ꢀtm per modification: –3.6 to –4.9 °C). It should also be pointed out that
the other four possible structural isomers of amide dimers between the C3’ of the upper base and the C5’ of
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the lower base have been prepared and evaluated by the same group.
More recently, Rozners and
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Iwase showed that this same internucleoside amide linkage C3’ꢀCH ꢀCOꢀNHꢀ C5’, such as in dimer D in
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RNA strands, does not disturb the Aꢀtype structure of the corresponding RNA/RNA duplexes (ꢀtm per
modification: –0.2 to +0.7 °C). As previously mentioned for the DNA series, amideꢀlinked RNA is
remarkably resistant to nucleaseꢀmediated cleavage.
In the quest for ideal backbone replacements, various neutral linkages containing the amide function, such as
the phosphodiester mimic, have been extensively studied for potential nucleic acidꢀbased therapeutic
10,17
applications.
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Among them, the C3’ꢀCH ꢀCOꢀNHꢀ C5’ internucleoside amide linkage present in dimers C
and D (Figure 1) seems to be one of the most promising achiral nonꢀionic phosphate linkage mimic
candidates of chemically modified nucleosides developed so far. It is worth noting that in both deoxyriboꢀ
and riboꢀoligonucleotides with the C3’ꢀCH ꢀNHꢀCOꢀ C5’ internucleoside amide linkage, one of the other
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four possible structural isomers of amide dimers between the C3’ of the upper base and the C5’ of the lower
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base displayed very similar binding affinities compared to the C3’ꢀCH ꢀCOꢀNHꢀ C5’ internucleoside amide
linkage present in dimers C and D (see Figure 1).
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In general, the amideꢀlinked deoxyribonucleoside dimers, such as in C (see Figure 1), have been obtained
from the commercially available natural nucleosides. In this way, the carboxylic acid monomers were
efficiently synthesized by freeꢀradical allylation at C3’ with total αꢀdiastereoselectivity from 5′ꢀOꢀprotected
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2′ꢀdeoxynucleoside precursors. This key step was followed by successive oxidation reactions to provide
the desired acid monomer. This is in sharp contrast to the synthesis of amideꢀlinked ribonucleoside dimers
D, for which the reaction applied to 2’,5’ꢀdiꢀOꢀprotected ribonucleoside intermediates led to the desired
products contaminated by the other diastereoisomer arising from freeꢀradical allylation at C3’ on the βꢀface.
Consequently, in this series, the synthesis of these required carboxylic acid monomers has been largely
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reported from sugars, mainly
the required carboxylic acid monomers of amideꢀlinked ribonucleoside dimers D turned out to be more
efficient: from a protected αꢀ ꢀerythroꢀpentofuranosꢀ3ꢀulose intermediate, a sequence of the WittigꢀHorner
Dꢀxylose or Dꢀglucose. As previously mentioned, this sugar route to prepare
D
reaction followed by diastereoselective hydrogenation provided the desired glycosyl precursor, which was
then involved in an Nꢀglycosylation to introduce the desired nucleobase. At the final stage, these amine and
carboxylic acid monomers are assembled using well established peptide coupling chemistry and, after
protecting group manipulation, their protected phosphoramidite dimeric blocks are incorporated into the
desired amideꢀlinked RNA strands according to standard solidꢀphase oligomerization on RNA synthesizers.
To be complete, it should be noted that an original approach concerning the synthesis of RNA sequences
containing up to three consecutive internal amide linkages such as in C has been published by Rozners and
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coꢀworkers. From the 5’ꢀNꢀMMT (4ꢀmethoxytrityl) aminouridineꢀ3’ꢀphosphoramidite monomer, these
authors used a combination of standard solidꢀsupport automated phosphoramidite chemistry and manual
addition of 5′ꢀaminoꢀ3′ꢀcarboxymethyluridine derivative and 5′ꢀOꢀ MMT 3′ꢀcarboxymethyluridine
monomers, as their activated carboxylic ester derivatives formed separately under conventional peptide
coupling conditions.
2-Results and discussion
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As part of our interest in the synthesis of modified nucleosides, we initiated a new program for the efficient
and general preparation of amideꢀlinked ribonucleoside dimers D, with the aim of avoiding the fastidious
preparation of each of the four monomeric building block amine and carboxylic acid nucleoside precursors
required to access any given base sequence. To reach this goal, our retrosynthetic analysis of these protected
amideꢀlinked ribonucleoside dimers D, delineated in Scheme 1, relied on an original sequential introduction
of a nucleobase sequence from a common twoꢀfuranoside amideꢀlinked scaffold 4. It should be highlighted
that the laterꢀstage stereocontrolled implantation of nucleobases avoids the tedious manipulations of various
sensitive protected nucleoside derivatives, which greatly facilitates both the synthesis and the purification.
The crucial issue of this strategy was the stability of the isopropylidene protecting group in the 1,2 positions
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of the upper furanose in 4, under the Vorbrüggen Nꢀglycosylation conditions required to introduce the first
nucleobase. Then, it was also expected that further transformations, including acetolysis and a second
Vorbrüggen Nꢀglycosylation, should be compatible with the various chemical functionalities on this
advanced intermediate 3. The successful implementation of this strategy would provide amideꢀlinked
ribonucleoside dimers 1 with all possible combinations of nucleobases from a common scaffold 4.
Scheme 1. Retrosynthetic analysis of protected amideꢀlinked ribonucleoside dimers 1.
As a prelude to this approach, a straightforward synthesis of the amideꢀlinked ribonucleoside homodimers
1
2
such as 1 (Base =Base ) was investigated using a similar strategy from a tetraacetate scaffold 7 in which a
oneꢀpot double Vorbrüggen transglycosylation with a suitable single type of base provided the desired
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homodimer, as outlined in Scheme 5. It should be pointed out that the convergent synthesis of the common
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scaffold 4 and its related tetraacetate analogue 7, as outlined in Schemes 4 and 2 respectively, is based on
the same building blocks that are readily available from ꢀxylose.
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D
The first common glycosyl donor 7 was prepared as depicted in Scheme 2. The synthesis of both amine 19
and acid 6 monomers started from the known TBDPSꢀprotected 3ꢀulose 9 derivative, which was readily
obtained according to a known procedure from
Dꢀxylose by a fourꢀstep sequence along with an optimized
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scalable oxidation step. Our synthesis commenced with acetalization of
D
ꢀxylose, followed by selective
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cleavage of the 3’,5’ꢀacetal as described by Bozo and coꢀworkers and finally selective protection of the
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resulting primary alcohol function with TBDPSCl to furnish the secondary alcohol 8 in 70% overall yield.
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The latter was oxidized into its corresponding ketone 9 in quantitative yield with a TEMPO/BAIB system
under mild conditions. It should be pointed out that this oxidation procedure was more efficient on a large
3 2
scale (up to 10 g) than conventional methods such as CrO /pyridine/Ac O, PCC, or Swern oxidation.
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Scheme 2. Conditions: (a) H
steps; (c) TBDPSCl, imidazole, DMF, rt, 48 h, 70%; (d) BAIB, TEMPO, DCM (0.6 M), rt, 40 h, quant. (e)
NaBH , EtOH/water (9:1), rt, 16 h, 95%, dr > 95:5; (f) NaH, BnBr, THF, rt, 16 h, 92%; (g) TBAF, THF, 4
C, 3 h then rt, 16 h, 80%; (h) MsCl, pyridine, rt, 3 h, quant.; (i) NaN , DMF, 90 °C, 3 h, 95%; (j) H (atm.
press.), Pd/C (5 wt.%), EtOH, rt, 4 h, 94%; (k) Ph P=CHCO Et, DCM, rt, 24 h, 95%, dr > 95:5; (l) TBAF,
2 4 4
SO , CuSO , acetone, rt, 40 h; (b) HCl, MeOH, 40 °C, 2 h, quant. for two
4
°
3
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3
2
THF, 4 °C, 3 h, then rt, 2 h, 92%; (m) H
BnBr, DMF, rt, 20 h, 84%; (o) LiOH, dioxane/water (1:1), rt, 22 h, quant.; (p) TBTU, HOBt, Et
°C, 30 min, then 19, Et N, 5 °C to rt, 16 h, quant.; (q) Ac O, CSA (0.05 eq.), AcOH, 85 °C, 1 h, then CSA
0.05 eq.), 85 °C, 3 h, 39%.
2
(atm. press.), Pd/C (5 wt.%), EtOH, rt, 24 h, quant.; (n) NaH,
3
N, CH CN,
3
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(
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At this juncture, having in hand this common ketone 9, we turned our attention toward the synthesis of the
carboxylic acid synthon 6. In accordance with our plan, the 5ꢀOꢀTBDPSꢀ1,2ꢀOꢀisopropylideneꢀαꢀ ꢀerythroꢀ
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D
pentofuranosꢀ3ꢀulose derivative 9 was treated with [(ethoxycarbonyl)ꢀmethylene]triphenylphosphorane in
DCM to provide stereoselectively the (Z)ꢀα,βꢀunsaturated ester 10 in 95% yield. Although of no
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consequence to the synthesis, the Z geometry of the ester was confirmed from the H and C NMR spectra,
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which were in agreement with the reported data. At this point, we envisaged that diastereoselective
reduction of the double bond could be carried out directly from the fully protected intermediate 10 to
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provide the desired 3α diastereoisomer. All attempts to reduce this α,βꢀunsaturated ester 10, either by
4
hydrogenation or with NaBH , led to a diastereomeric mixture. At this point, prompted by Robins’s
19c
reports,
the silyl group of 10 was cleaved with tetrabutylammonium fluoride (TBAF) and the
corresponding primary alcohol 11 was engaged in the reduction step. Classic catalytic hydrogenation of 11
under atmospheric pressure in the presence of 5% palladium on charcoal in EtOH at room temperature
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afforded the saturated ester 12 in 87% crude yield as the sole diastereoisomer according to H and C NMR
analyses. This result suggested that the alcohol function at C5 in intermediate 11 is able to bind to the
heterogeneous catalyst surface to exert stereochemical control of hydrogen addition to the olefin with a high
level of diastereoselectivity. We also anticipated that upon switching the protecting group from silylꢀ to
benzylꢀether would remedy the partial cleavage of the 5ꢀOꢀTBDPS group under the acidic conditions of the
acetolysis reaction (vide infra) as observed in our preliminary investigations. Finally, benzylation of the
primary alcohol 12 under classic conditions furnished the expected benzylether 13 in 84% yield, which upon
treatment with LiOH led quantitatively to the key acid synthon 6. With the synthesis of the acid 6 secured in
nine steps from Dꢀxylose with an excellent overall yield of 50%, we began to focus our attention on the
synthesis of the amine monomer 19. Preparation of the amine 19 began with a diastereoselective reduction
of the key ketone intermediate 9 and subsequent benzylation of the resulting alcohol function at C3 in 14,
followed by fluorideꢀmediated cleavage of the silylether. Finally, activation of the liberated alcohol as a
2
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mesylate led to compound 17 in 50% overall yield. Nucleophilic displacement of the 5ꢀOꢀmesylate group
of 17 with sodium azide gave the desired azido derivative 18, which was then reduced to provide the
corresponding amine monomer 19 in 90% overall yield for this twoꢀstep sequence. It is noteworthy that the
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palladium on a charcoal source had no influence on the chemoselectivity of this catalytic hydrogenation
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reaction, which was run at various scales with no problem. This key amine monomer 19 was synthesized in
six steps from the intermediate ketone 9 and in ten steps from Dꢀxylose with a very good yield of 45%. With
the synthesis of both amine 19 and acid 6 monomers secured, the stage was then set for the coupling reaction
to give the first amideꢀlinked glycosyl donor 7. The targeted amideꢀlinked pentose dimer 20 was isolated in
near quantitative yield upon treatment of acid 6 with amine 19 monomers with a coupling reagent cocktail of
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2ꢀ(1Hꢀbenzotriazolꢀ1ꢀyl)ꢀ1,1,3,3ꢀtetramethyluronium tetrafluoroborate (TBTU), 1ꢀhydroxybenzotriazole
3
0
(
3
HOBt) and Et N in DCM. Next, conversion of both 1,2ꢀOꢀisopropylidene protection groups of compound
20 into its corresponding peracetylated glycosyl donor proved to be problematic to provide our amideꢀlinked
dimer 7. Most of the common protocols to perform this latter transformation involve acidic cleavage
2 4 2
(AcOH, TFA, H SO ) of the acetonide followed by acetylation of the resulting 1,2ꢀdiol with Ac O in a oneꢀ
3
1
pot or sequential reaction. From dimer 20, extensive optimization of this crucial transformation in our plan,
32
33
34
2 4
with regard to the acid component (AcOH, TFA, H SO ), revealed that a oneꢀpot twoꢀstep sequence
3
5
with camphorsulfonic acid (CSA) in a mixture of AcOH and Ac O was the most efficient in our hands.
2
Obviously, acid additives significantly influenced the reaction. However, after silica gel column
chromatography purification, the yields of this acetolysis reaction leading to the tetraacetylated dimer 7 were
modest (39%) (vide infra). This tetraacetylated dimer 7 was finally obtained in 17 steps from Dꢀxylose and
with an 18% overall yield.
In addition to our acetolysis optimization, suspecting that the pendant amide moiety at C3 in dimer 20 could
36
interfere with the course of this transformation, we explored this process with model substrates 21 and 22,
as well as a more advanced model 23. Our more relevant findings are presented in Scheme 3.
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Scheme 3. Conditions: (a) AcOH aq (or TFA aq), 90 °C, 6 h
CSA (0.1 eq. 0.2 eq.), AcOH, 80 °C, 40 min 4 h.
2 2
−24 h then Ac O, pyridine, rt, 20 h; (b) Ac O,
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
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4
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5
5
5
5
5
5
5
5
6
−
−
2
Acetonide deprotection of the amide 21 with aqueous AcOH followed by acetylation with Ac O in pyridine
19c,37
resulted in a mixture of the lactone 24
in 45−50% yield, as well as a bridged heterotricyclic lactam 25,
arising from an intermolecular Nꢀglycosidation with the nitrogen atom of the amide function, in 25% yield
(Scheme 3, eq 1). To avoid the possibility of intramolecular amide cyclization at the anomeric position, the
Nꢀmethyl derivative 22 was subjected to a standard twoꢀstep acetolysis. Only the lactone 24 was isolated in
0
1
2
3
4
5
6
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8
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8
9
0
50% yield after purification (Scheme 3, eq 2). Under the same acetolysis conditions, the more advanced
model 23 led only to the formation of the lactam 27 in 32% yield (Scheme 3, eq 3). The formation of the
bridged heterotricyclic lactams 25 and 27 resulted from the intramolecular nucleophilic attack of the NꢀH of
the amide parents 21 and 23 on the furanose oxocarbenium ion formed by acidꢀpromoted cleavage of the
acetonide moiety and subsequent elimination of water. The structures of 24-25 and 27 were assigned by
extensive NMR study (see Supporting Information for details).
In the second set of experiments on these model substrates 21
−23, a oneꢀpot twoꢀstep acetolysis in the
_
presence of CSA afforded the desired 1,2ꢀdiꢀOꢀacetate adducts 26 and 28
−
29 (Scheme 3, eq 4 6) as an
anomeric mixture in moderate to good yields after purification. As will be seen, we later found that
substantial decomposition of these 1,2ꢀdiꢀOꢀacetate derivatives occurred during silica gel column
chromatography purification.
With an efficient preparation of our first glycosyl dimer 7 in hand, we next turned our efforts to the
production of the second key dimer 4 as depicted in Scheme 4. Based on our previous experience, as
discussed above, our optimized conditions of acetolysis were applied to the azido intermediate 18 to yield
1
the corresponding 1,2ꢀdiꢀOꢀacetate compound 30. Analysis of the H NMR spectrum of this latter crude
reaction mixture clearly indicated that the only product of this transformation was the required
azidodiacetate 30 (>95% crude yield, >95% purity). Surprisingly, silica gel chromatography purification
afforded only a modest 50% yield of the desired pure compound 30. To avoid loss of product during silica
gel purification, the organic layer was washed around ten times during the workup with an aqueous saturated
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solution of NaHCO
further purification. With this azidodiacetate 30 in hand, we explored its reduction under hydrogenation
conditions (1 atm. H , 5% palladium on charcoal, THF, rt) into the corresponding amine 5 required for the
3
to provide the azidodiacetate 30 in 87% crude yield, which was directly used without
1
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3
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9
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1
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3
4
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4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
2
amide coupling reaction to provide the dimeric amide framework 4. Unfortunately, under these conditions, a
mixture of products whose structures were tentatively assigned as amine 5 (M = 323) and acetamides 31 or
0
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0
38
39
3
1’ (M = 323) arising from the migration of acetate was obtained. Attempts to reduce the azide 30 using
the Staudinger reaction led to the formation of the corresponding unusual stable iminophosphorane
intermediate, which clearly could not be hydrolyzed into the amine 5. However, to circumvent the acetate
migration, we ultimately found that the most efficient route to the amideꢀlinked dimer 4 was the threeꢀ
40
component StaudingerꢀVilarrasa ligation from the carboxylic acid 6 and azide 30. A first experiment
41
employing a catalytic variant of this StaudingerꢀVilarrasa coupling reaction with PySeSePy as activator
40
afforded the targeted dimer 4 in a modest yield of 37%. In accordance with the work of Strazewski,
reaction between the azide 30 and trimethylphosphine provided the corresponding azaꢀylide intermediate,
which was then treated by addition of the corresponding activated ester of the parent acid 6 formed in situ by
exposure to DIC and HOBt in THF at room temperature. Under these optimized conditions, the amideꢀ
linked dimer 4 was isolated in 65% yield after purification on silica gel chromatography. Thus, synthesis of
the second glycosyl dimer 4 was achieved in 16 steps with an overall yield of 30% from Dꢀxylose.
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The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Scheme 4. Conditions: (a) Ac
2
O, CSA (0.05 eq.), AcOH, 85 °C, 1 h, then CSA (0.05 eq.), 85 °C, 1 h, 87%
(
α:β = 15/85); (b) H (atm. press.), Pd/C (5 wt.%), THF, 3 h, total conversion; (c) Me P then 6, DIC, HOBt,
2
3
THF, rt, 4 h, 65%.
Then, we proceeded to evaluate our plan for a oneꢀpot and sequential introduction of the nucleobases as
outlined in Scheme 5. In the first instance, we examined the preparation of ribonucleosidic homodimers
from the amideꢀlinked sugar dimer precursor 7.
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1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
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3
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3
4
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8
9
0
Scheme 5. Conditions: (a) base, BSA, CH
3 3
CN, 60 °C, 1 h, then TMSOTf, CH CN, reflux, 2 h, 62% for
thymine, 56% for uracil; (b) H (atm. press.), Pd/C (30 wt%), EtOH, 24 h, 93% for 34, 71% for 35; (c)
2
thymine, BSA, CH CN, 60 °C, 1 h, then TMSOTf, CH CN, rt, 24 h, 89%; (d) Ac O, CSA (0.05 eq.), AcOH,
3
3
2
85 °C, 1 h, then CSA (0.05 eq.), 85 °C, 3 h; (e) uracil, BSA, CH
3
3
CN, 60 °C, 1 h, then TMSOTf, CH CN, rt,
2
6 h, 83%, for 2 steps. (f) K CO , MeOH, rt, 4 h, quant.
2
3
To this end, the tetraacetate glycosyl donor 7 was submitted to a double oneꢀpot Vorbrüggen Nꢀ
2
1
glycosylation with uridine. According to standard Vorbrüggen Nꢀglycosylation protocols, the glycosyl
donor 7 was reacted with bis(trimethylsilyl)uracil, formed in situ by silylation of uracil with
bis(trimethylsilyl)acetamide (BSA), and trimethylsilyltriflate (TMSOTf) as a promoter, in freshly distilled
acetonitrile at 60 °C, to give stereoselectively the βꢀconfigured protected homodimer UUꢀ32 in 56% yield
after purification. Importantly, the anchimeric assistance of the 2ꢀOꢀacetyl group in this Vorbrüggen type
coupling prevented the formation of an anomeric mixture and led to the exclusive formation of the amideꢀ
backbone uracil protected ribodinucleoside 32 as the βꢀanomer. The same transformation using thymine
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The Journal of Organic Chemistry
resulted in essentially identical efficiency with a 62% yield of the homodimer TTꢀ33. To finish, cleavage of
the benzyl protecting groups in UUꢀ32 and TTꢀ33 under classic hydrogenolysis conditions proceeded
smoothly to furnish the target homodimers UUꢀ34 and TTꢀ35 in 93% and 71% yields, respectively. It should
be noted that no cleavage by ethanolysis or migration of the acetyl group at C3’ in both nucleosides, due to
the slightly acidic nature of the palladium on charcoal catalyst, was observed during this transformation.
Following the successful synthesis of the homodimers UUꢀ34 and TTꢀ35, our efforts were focused on our
ultimate goal, which was the preparation of a heterodimer to validate our strategy. At this point, the
Vorbrüggen Nꢀglycosylation with thymine, using the previous conditions, was performed on the glycosyl
donor 4 to furnish the intermediate 36 in an excellent yield of 89% after purification. Then, this latter
glycosyl donor 36 was subjected to previously optimized acetolysis conditions and the resulting diacetate
intermediate 37 was directly engaged without further purification in the last Vorbrüggen Nꢀglycosylation
step with uracil to afford stereoselectively the desired heterodimer UTꢀ38 in 83% overall yield, after silica
gel chromatography. Cleavage of the acetate groups in dimer 38 by classic methanolysis furnished the
benzyl protected dimer 39 in quantitative yield. It should be pointed out that orthogonal deprotection of
dimers 32, 33 and 38 afforded efficiently the corresponding partially protected dimers 34, 35 and 39 in good
yields.
1
2
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0
3
-Conclusion
To the best of our knowledge, the work reported here is the first example of the synthesis of nucleosidic
heterodimers with an internucleoside amide linkage, such as dimers D, via the sequential introduction of
pyrimidine bases using two stereocontrolled Nꢀglycosidation reactions on a twoꢀfuranoside platform
precursor 4. From Dꢀxylose, homodimers UUꢀ34 and TTꢀ35 were obtained in 18 steps with excellent overall
yields of more than 10% while the heterodimer route led to UTꢀ39 in 19 steps with an overall yield of
around 10%.
The present preliminary investigation demonstrates the feasibility of our original strategy to introduce all
four natural nucleobases into both top and bottom sugars of the dimer platform 4, potentially leading to the
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1
6 possible combinations. The extension of this strategy to the preparation of dimers bearing various or
1
2
3
4
5
6
7
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9
1
1
1
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1
1
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4
4
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4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
original bases other than thymine and uracil is underway.
Experimental Section
General information: All solvents were reagent grade. Solvent used for chromatography
0
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1
2
3
4
5
6
7
8
9
0
(Dichloromethane, ethyl acetate and petroleum ether) were distilled on a rotavapor. Acetonitrile was freshly
distilled from calcium hydride under argon.
Purifications were performed with flash column chromatography on silica gel or with flash chromatography
instrument. TLC was performed on silica gel 60 F 254 analytical plates, compounds were detected by UV (λ
=
254 nm).
1
13
H and C NMR spectra were recorded with 300 MHz spectrometer fitted with a 5 mm i.d. BBO probe
1
13
carefully tuned to the recording frequency of 300.13 MHz (for H) and 75.47 MHz (for C), the temperature
of the probe was set at room temperature (around 293–294 K), on the spectrometer fitted with a 5 mm i.d.
1
13
BBFO+ probe carefully tuned to the recording frequency of 400.13 MHz (for H) and 100.61 MHz (for C),
13
1
the temperature of the probe was set at 303 K, on the 500 MHz spectrometer fitted with a 5mm i.d. C/ H
1
13
cryoprobe carefully tuned to the recording frequency of 500.13 MHz (for H) and 125.76 MHz (for C), the
temperature of the probe was set at 303 K. The spectra are referenced to the solvent in which they were run
1
13
1
13
(7.26 ppm for H CDCl
3
and 77.16 ppm for C CDCl
3
, 2.50 ppm for H DMSOꢀd
6
and 39.52 ppm for C
1
13
1
13
DMSOꢀd
6
, 3.31 ppm for H CD
3
OD and 49.00 ppm for C, 7.16 ppm for H C
6
D
6
and 128.06 ppm for C
C D ). Chemical shifts (δ) are given in parts per million (ppm), and coupling constants (J) are given in Hz.
6
6
Multiplicity of signals is indicated as following: s (singlet), d (doublet), t (triplet), q (quadruplet), m
multiplet), brs (broad singlet), dd (doublet of doublet), dt (doublet of triplet). All assignments were
(
confirmed with the aid of twoꢀdimensional experiments both homonuclear 1Hꢀ1H (COSY) and
heteronuclear 1Hꢀ13C (HSQC and HMBC) correlation spectroscopy , standard pulse programs were used. In
case of dimer, we used the symbol A for the upper unit and B for the lower unit.
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The Journal of Organic Chemistry
Low resolution mass spectrometry (MS) were recorded on a quadripolar spectrometer (coupled with a
1
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7
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1
1
1
1
1
1
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1
1
2
2
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2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
TracUltra GC apparatus) for Chemical Ionization (CI), on a TOF spectrometer for ElectroSpray Ionisation
(ESI).
High resolution mass spectrometry (HRMS) were recorded on an electroꢀmagnetic spectrometer (for CI), on
a TOF spectrometer (for ESI+ and MALDI+).
0
1
2
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Optical rotation data were obtained on a polarimeter, in a 100 mm cell, under Na lamp radiation at 20 °C.
1,2-O-isopropylidene-3-deoxy-3-(1,2-O-acetyl-3-O-benzyl-5-deoxy-5-acetamido-
β-ribosyl)-5-O-benzyl-
α-D-ribofuranose (4): Acid 6 (1.3 eq., 0.74 mmol, 239 mg) and HOBt.H O (1.3 eq., 0.74 mmol, 100 mg)
2
were coꢀevaporated with toluene and then diluted with THF (7 mL), cooled down to 0 °C under N for 10
2
min. DIC (1.25 eq., 0.7 mmol, 88 mg) was added and the reaction mixture was stirred at 0 °C for 15 min and
then 10 min at room temperature. A 1 M solution of trimethylphosphine in THF (2 eq., 1.14 mmol, 1.14 mL)
was added to azide 30 (1 eq, 0.57 mmol, 200 mg) in THF (7 mL) and the reaction mixture was stirred at
room temperature for 5 min. The acid mixture was added to the iminophosphorane solution and stirred for 4
h at room temperature. The mixture was concentrated under reduced pressure, diluted with toluene (20 mL)
2 4
and washed with water (2 x 20 mL) to eliminate urea. The organic layer was dried over Na SO , filtered,
concentrated under reduced pressure. The crude was purified by flash column chromatography (DCM/iꢀ
PrOH: 99.5/0.5 to 95/5) to give 4 as a beige sticky oil (225 mg, 0.36 mmol, 65%). TLC (DCM/iꢀ
1
PrOH:95/5): R
f
= 0.40. H NMR (400 MHz, CDCl
3
) of two anomers
α/β
in a 20/80 ratio: δ (ppm) = 7.36–
7
.28 (m, 10H, Har), 6.30 (d, J1B,2B = 4.5 Hz, 0.2H, H1Bα), 6.11 (s, 0.8H, H1Bβ), 5.89 (m, 1H, NHamide), 5.80
d, J = 3.9 Hz, 1H, H ), 5.27 (d, J
(
= 4.4 Hz, 1H, H2Bβ), 5.07 (dd, J2B,3B = 6.5 Hz and J1B,2B = 4.5 Hz,
2B,3B
1A
1H, H2Bα), 4.69 (t, J = 3.7 Hz, 1H, H2A), 4.59 (d, Jgem = 11.1 Hz, 1H, CH
2
PhAa), 4.58–4.56 (m, 2H, CH
2 B
Ph ),
4
.46 (d, Jgem = 11.1 Hz, 1H, CH Ph ), 4.15–4.10 (m, 1H, H ), 4.01 (d, J
= 4.4 Hz, 1H, H ), 3.96–3.91
2
Ab
4B
2B,3B
3B
(m, 1H, H4A), 3.65–3.41 (m, 4H, H5A and H5B), 2.45–2.37 (m, 2H, H3A and CHH'CO), 2.31–2.24 (m, 1H,
13
CHH'CO), 2.12 (s, 3H, MeAc), 2.08 (s, 3H, MeAc), 1.49 (s, 3H, Meacetonide), 1.31 (s, 3H, Meacetonide).
C
NMR (100 MHz, CDCl ): δ (ppm) = 171.2 (COamide), 169.8 (COAc), 168.9 (COAc), 138.0 (CIV(ar)), 137.2
3
(C
IV(ar)), 128.6 (Car), 128.5 (Car), 128.1 (Car), 127.8 (Car), 111.5 (CIV(acetonide)), 105.0 (C1A), 98.6 (C1Bβ), 94.3
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1Bα) 81.2 (C2A), 80.9 (C4B), 80.1 (C4A), 77.8 (C3B), 73.6 (CH Ph), 73.6 (C2Bβ), 73.6 (CH
0.0 (C5A), 42.4 (C3A), 41.4 (C5B), 32.0 (CH CO), 26.8 (Meacetonide), 26.4 (Meacetonide), 21.1 (MeAc), 20.7
Page 18 of 50
(
C
2
2
Ph), 70.9 (C2Bα),
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
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3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
7
2
+
+
+
+
(
MeAc). MS (CI ): m/z = 628.4 Da [MH] , 570.3 Da [MHꢀacetone] . HRMS (MALDI : DHB, PEG 600):
calcd for C H N O Na = 650.2572 Da, found 650.2577 Da.
33
41
1
11
1
,2-O-isopropylidene-3-deoxy-3-C-carboxymethyl-5-O-benzyl-α-D-ribofuranose (6): To a solution of 13
(1 eq, 1.86 mmol, 650 mg) in a 1:1 mixture dioxane/water (40 mL) was added LiOH (20 eq, 37.2 mmol,
.56 g). The mixture was stirred at room temperature for 48 h, then diluted with DCM (40 mL) and acidified
0
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3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
with HCl 2N (16.8 mL) until pH=2. The aqueous layer was extracted with DCM (3 x 20 mL), the combined
organic layers were dried over Na SO , filtered and concentrated under reduced pressure to give the desired
2
4
1
compound as a brown oil (600 mg, 1.86 mmol, quantitative). TLC (DCM): R
CDCl3):
f
= 0.09. H NMR (300 MHz,
δ
(ppm) = 7.38–7.33 (m, 5H, H ), 5.85 (d, J = 3.8 Hz, 1H, H ), 4.80 (t, J = J2,3 = 3.8 Hz, 1H, H2
ar 1,2 1 1,2
)
, 4.61 (d, Jgem = 12.2 Hz, 1H, CH
2
Ph), 4.53 (d, Jgem = 12.2 Hz, 1H, CH
2
Ph), 3.94 (dt, J3,4 = 10.1 Hz and
J
4,5a = J4,5b = 4.2 Hz, 1H, H
4
), 3.66 (dd, Jgem = 10.8 Hz and J4,5a = 3.8 Hz, 1H, H5a), 3.56 (dd, Jgem = 10.8 Hz
(CH CO)), 2.47–2.36 (m,
): δ
and J4,5b = 4.2 Hz, 1H, H5b), 2.71 (dd, Jgem = 17.8 Hz and J3,Ha = 10.8 Hz, 1H, H
a
2
1
3
2
H, H
b
(CH
2 3
CO) and H ), 1.49 (s, 3H, Meacetonide), 1.32 (s, 3H, Meacetonide). C NMR (75 MHz, CDCl
3
(
ppm) = 177.2 (CO), 138.0 (CIV(ar)), 128.6 (Car), 127.9 (Car), 111.8 (CIV(acetonide)), 105.0 (C
1
), 81.0 (C
2
), 79.6
+
(
C
4
), 73.7 (CH
2
Ph), 69.4 (C
5
), 41.6 (C
3
), 29.6 (CH
2
CO
2
H), 26.8 (Meacetonide), 26.5 (Meacetonide). MS (ESI ):
+
+
20
D
m/z = 345.1 Da [MNa] . HRMS (ESI ): calcd for C17
H
22
O
6
Na = 345.1314 Da, found = 345.1314 Da. [α]
=
3
+33.3 (CHCl , c = 1 g/100mL).
1
,2-O-acetyl-3-deoxy-3-(1,2-O-acetyl-3-O-benzyl-5-deoxy-5-acetamido-β-D-ribosyl)-5-O-benzyl-β-D-
ribofuranose (7): To a solution of 20 (1 eq, 0.41 mmol, 240 mg) in acetic acid (18 mL) was added acetic
anhydride (80 eq, 32.8 mmol, 3 mL), camphor sulfonic acid (0.05 eq, 0.02 mmol, 4.7 mg). The mixture was
stirred for 4 h at 80–85 °C, then another portion of camphor sulfonic acid (5% weight, 0.02 mmol, 4.7 mg)
was added and stirring was pursued for 1 h. The reaction mixture was diluted with DCM (15 mL), washed
3
with water (3 x 20 mL) and a saturated aqueous solution of NaHCO (3 x 20 mL). The organic layer was
dried over Na
2
SO
4
, filtered, concentrated under reduced pressure and coꢀevaporated with toluene (20 mL)
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The Journal of Organic Chemistry
then Et
2
2
O (20 mL). The crude was purified by flash column chromatography (Et O/Petroleum Ether: 90/10)
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
to give the desired compound as a single diastereoisomer as a beige oil (110 mg, 0.16 mmol, 39%). TLC
1
(Et
2
O): R
f
= 0.18. H NMR (400 MHz, CDCl
3
):
1
δ (ppm) = 7.35–7.30 (m, 10H, Har), 6.10 (s, 1H, H ), 6.07
(s, 1H, H ), 5.78 (t, J
= 5.7 Hz, 1H, NHamide), 5.29–5.27 (m, 2H, H ), 4.60–4.54 (m, 4H, 2CH Ph), 4.13–
1
NH,5B
2
2
4
.08 (m, 2H, H
4
), 3.97 (dd, J = 8.1 Hz and J = 4.4 Hz, 1H, H3B), 3.67–3.58 (m, 3H, 2H5A and H5B), 3.32–
CO), 2.09 (s, 3H, MeAc), 2.07 (s, 6H,
(ppm) = 170.4 (COamide), 169.88 (COAc), 169.84
(CO ), 169.5 (CO ), 169.1 (CO ), 138.1 (CIV(ar)), 137.2 (CIV(ar)), 128.7 (C ), 128.6 (C ), 128.6 (C ),
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
3.26 (m, 1H, H5B), 2.96–2.88 (m, 1H, H3A), 2.37–2.31 (m, 2H, CH
2
13
3
MeAc), 1.95 (s, 3H, MeAc). C NMR (100 MHz, CDCl ): δ
Ac
Ac
Ac
ar
ar
ar
128.5 (C ), 128.4 (C ), 128.2 (C ), 128.1 (C ), 127.8 (C ), 127.8 (C ), 127.7 (C ), 98.8 (C ), 98.7 (C ),
ar ar ar ar ar ar ar 1 1
8
3.6 (C4A), 80.8 (C4B), 78.2 (C3B), 77.9 (C2A), 73.62 (CH
5B), 39.0 (C3A), 32.4 (CH
2 2
Ph), 73.58 (C2B), 73.5 (CH Ph), 71.9 (C5A), 42.2
+
(
[
[
C
2
CO), 21.2 (2MeAc), 20.8 (2MeAc). MS (MALDI : DHB): m/z = 694.2 Da
+
+
41 1
MNa] . HRMS (MALDI : DHB, PEG 600): calcd for C34H N O13Na = 694.2470 Da, found 694.2450 Da.
2
D
0
α]
3
= +8.7 (CHCl , c = 1.5 g/100mL).
1,2-O-isopropylidene-5-O-tert-butyldiphenylsilyl-α-
isopropylideneꢀαꢀ ꢀxylofuranose, was obtained as followed. To 350 mL of reagent acetone were added
successively: sulfuric acid concentrated (2 mL), copper sulfate (1.88 eq, 250 mmol, 40 g) and xylose (1 eq,
33 mmol, 20 g). The reaction was stirred at room temperature for 48 h. The mixture was filtered,
NH OHconc. (25%, 6.4 mL) was added, the mixture was filtered, dried over Na SO , filtered and
D-xylofuranose (8): The precursor of 8, the 1,2ꢀOꢀ
D
1
4
2
4
concentrated under reduced pressure. The residue was diluted with methanol (150 mL) and HCl 0.1 N (44
mL) was added. The reaction was stirred at 40 °C for 2 h and quenched with NaHCO (6.3 g until neutral
pH). The mixture was filtered and concentrated under reduced pressure. The residue was coꢀevaporated with
3
EtOH/toluene, was diluted with DCM, dried over Na
to obtain the desired 1,2ꢀOꢀisopropylideneꢀαꢀ
300 MHz, CDCl ): (ppm) = 5.96 (d, J1,2 = 3.6 Hz, 1H, H
), 4.15 (q, J4,5a = J4,5b = J4,3 = 3.2 Hz, 1H, H ), 4.08 (d, J5a,5b = 12.2 Hz, 1H, H5a), 3.50 (d, J5a,5b = 12.2 Hz,
2
SO
4
, filtered and concentrated under reduced pressure
1
D
ꢀxylofuranose (25.3 g, 133 mmol, quantitative). H NMR
(
3
δ
1 2
), 4.51 (d, J1,2 = 3.6 Hz, 1H, H ), 3.34 (br, 1H,
H
3
4
1
3
3
1H, H5b), 1.47 (s, 3 H, Meacetonide), 1.31 (s, 3 H, Meacetonide). C NMR (75 MHz, CDCl ): δ (ppm) = 112.1
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+
(C
IV), 105.6 (C
1
), 85.8 (C
2
), 83.8 (C
4
3
), 79.00 (C ), 66.3 (C
5
), 27.00 (Meacetonide), 26.3 (Meacetonide). MS (CI ):
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
+
+
+
+
+
m/z = 208 Da [MNH
4
] , 191 Da [MH] . MS (ESI ): m/z = 213.1 Da [MNa] . HRMS (ESI ): calcd for
8 14 5
C H O Na = 213.0739 Da, found = 213.0739 Da.
To a cooled solution of 1,2ꢀOꢀisopropylideneꢀαꢀ
1 mmol, 5.5 g) in DMF (70 mL) was added tertꢀbutyldiphenylsilyl chloride (1 eq, 36.8 mmol, 9.5 mL).
After stirring at room temperature for 48 h, the reaction was quenched with Et O (70 mL) and water (70
mL). The aqueous layer was extracted with Et O (3 x 70 mL) and the combined organic layers were washed
with brine (5 x 70 mL), dried over Na SO , filtered and concentrated under reduced pressure. The crude was
Dꢀxylofuranose (1 eq, 36.8 mmol, 7g) and imidazole (2.2 eq,
8
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
2
2
4
poured into pentane, after 1 h at –18 °C the colorless solid was filtered to obtain the desired compound (11.3
g, 25 mmol, 70%). Spectroscopic data of this compound 8 were consistent with those reported in the
2
5
1
literature . Mp: 91ꢀ92 °C. TLC (DCM): R
H, Har), 7.48–7.37 (m, 6H, Har), 6.02 (d, J1,2 = 3.7 Hz, 1H, H
2.0 Hz, 1H, H ), 4.14–4.10 (m, 3H, H , H5a, H5b), 4.07 (d, JOH,3 = 3.3 Hz, 1H, OH), 1.47 (s, 3H, Meacetonide),
1.33 (s, 3H, Meacetonide), 1.05 (s, 9 H, MetBu). C NMR (75 MHz, CDCl ): δ (ppm) = 135.8 (Car), 135.6
f
= 0.35. H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 7.73–7.66 (m,
4
1
), 4.55 (d, J1,2 = 3.7 Hz, 1H, H
2
), 4.38 (d, J3,4
=
4
3
13
3
(
C
C
ar), 132.6 (CIV(ar)), 132.0 (CIV(ar)), 130.2 (Car), 128.1 (Car), 111.7 (CIV(acetonide)), 105.1 (C
1
2
), 85.6 (C ), 78.5
+
(
3
), 77.1 (C
4
), 63.0 (C
5
), 26.9 (Meacetonide), 26.8 (MetBu), 26.3 (Meacetonide), 19.2 (CIV(tBu)). MS (ESI ): m/z =
+
+
4
51.2 Da [MNa] . HRMS (ESI ): calcd for C24
H
32
O
5
SiNa = 451.1917 Da, found = 451.1923 Da.
1
,2-O-isopropylidene-3-deoxy-3-keto-5-O-tert-butyldiphenylsilyl-α-
D-xylofuranose (9): To a solution of
1
,2ꢀOꢀisopropylideneꢀ5ꢀOꢀtertꢀbutyldiphenylsilylꢀαꢀ ꢀxylofuranose 8 (1 eq, 23.3 mmol, 10 g) in DCM (40
D
mL) were added BAIB (1.1 eq, 25.6 mmol, 8.3 g) and TEMPO (0.1 eq, 2.3 mmol, 350 mg). After 36 h, the
mixture was diluted with DCM (20 mL), quenched with Na (20 mL) and extracted with DCM (4 x 20
mL). The combined organic layers were dried over Na , filtered and concentrated under reduced
2 2 3
S O
2
SO
4
pressure. The crude was purified by flash column chromatography (Petroleum Ether/Et O, 1/0 to 4/1) to
2
obtain the desired compound as a beige oil (10 g, 23.4 mmol, quantitative). Spectroscopic data of this
25
1
f
compound 9 were consistent with those reported in the literature . TLC (DCM): R = 0.35. H NMR (300
MHz, CDCl
3
):
δ
(ppm) = 7.69–7.66 (m, 2H, Har), 7.62–7.59 (m, 2H, Har), 7.43–7.39 (m, 6H, Har), 6.26 (d,
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The Journal of Organic Chemistry
), 4.43 (dd, J1,2 = 4.5 Hz and J2,4 = 1.0 Hz, 1H, H
gem = 11 Hz and J4,5a = 1.8 Hz, 1 H, H5a), 3.85 (dd, Jgem = 11 Hz and J4,5b = 2.0 Hz, 2H, H5b), 1.48 (s, 3H,
J
J
1,2 = 4.5 Hz, 1H, H
1
2 4
), 4.40–4.35 (m, 1H, H ), 3.92 (dd,
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
1
3
3
Meacetonide), 1.47 (s, 3H, Meacetonide), 1.00 (s, 9 H, MetBu). C NMR (75 MHz, CDCl ): δ (ppm) = 211.1
(CO), 135.7 (C ), 135.6 (C ), 132.5 (CIV(ar)), 132.3 (CIV(ar)), 130.1 (C ), 130.1 (C ), 128.0 (C ), 114.4
ar
ar
ar
ar
ar
(C
IV(acetonide)), 103.9 (C
1
), 81.7 (C
4
), 77.3 (C
2
5
), 64.6 (C ), 27.9 (Meacetonide), 27.4 (Meacetonide), 26.8 (MetBu),
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
+
+
+
1
9.2 (CIV(tBu)). MS (CI ): m/z = 444.0 Da [MNH
4
] , MS (ESI ): m/z = 481.2 Da [M+MeOH+Na] . HRMS
+
20
D
(
ESI ): calcd for C25
H
34
O
6
SiNa = 481.2022 Da, found = 481.2028 Da. [α]
3
= +76.9 (CHCl , c = 1.7
g/100mL).
1
,2-O-isopropylidene-3-deoxy-3-C-(Z)-ethoxycarbonylmethylene-5-O-tert-butyldiphenylsilyl-α-
xylofuranose (10): To a solution of 9 (1eq, 10.5 mmol, 4.5g) in DCM (30 mL) was added
carbethoxymethylene) triphenylphosphorane (1.15 eq, 12.07 mmol, 4.2 g). After stirring under argon at
room temperature for 30 min the mixture became pink. After stirring for 24 h the mixture was concentrated
and the residue was diluted with Et O (100 mL) and stirred for 30 min. Triphenylphosphine oxide was
filtered, the organic layer was dried over Na SO , filtered and concentrated under reduced pressure. The
D-
(
2
2
4
crude was purified by flash column chromatography (DCM) to obtain the desired compound as a yellow oil
1
(
5.0 g, 10 mmol, 95%). TLC (DCM): R = 0.64. H NMR (300 MHz, CDCl ):
δ
(ppm) = 7.67–7.61 (m, 4H,
f
3
H
ar), 7.44–7.37 (m, 6H, Har), 5.99 (d, J1,2 = 4.1 Hz, 1H, H
1
), 5.97 (t, JCH,2 = JCH,4 = 1.7 Hz, 1H, CH=C), 5.71
(dt, J1,2 = 4.1 Hz and J2,4 = JCH,2 = 1.7 Hz, 1H, H ), 4.90 (tt, J = J4,5b = 3.5, J4,2 = J4,CH = 1.7, 1 H, H4),
2
4,5a
4.27 (q, J = 7.1 Hz, 1H, CH
2
CH
3
), 4.26 (q, J = 7.1 Hz, 1H, CH
2
3
CH ), 3.86 (dd, Jgem = 10.9 Hz and J4,5a = 3.8
Hz, 1 H, H5a), 3.71 (dd, Jgem = 10.9 Hz and J4,5b = 3.2 Hz, 1H, H5b), 1.49 (s, 3H, Meacetonide), 1.45 (s, 3H,
1
3
Meacetonide), 1.33 (t, J = 7.1 Hz, 3H, CH
2
CH
3 3
), 1.03 (s, 9H, tBu). C NMR (75 MHz, CDCl ): δ (ppm) =
165.1 (CO), 156.8 (C ), 135.74 (Car(o)), 135.70 (Car(o)), 132.9 (C ), 130.0 (Car(m)), 129.8 (Car(m)), 127.9
3
IV
(
C
ar(p)), 116.9 (CHCO
2
Et), 113.1 (CIV(acetonide)), 105.8 (C
1
), 81.5 (C
4
), 79.1 (C
CH
2 5 2 3
), 66.2 (C ), 60.8 (CH CH ),
+
2
7.7 (Meacetonide), 27.4 (Meacetonide), 26.9 (MetBu), 19.3 (CIV(tBu)), 14.3 (CH
2
3
). MS (ESI ): m/z = 519.2 Da
+
+
20
D
[
MNa] . HRMS (ESI ): calcd for C H O SiNa = 519.2178 Da, found = 519.2179 Da. [α]
= +47
28 36
6
3
(CHCl , c = 1 g/100mL).
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1
,2-O-isopropylidene-3-deoxy-3-C-ethoxycarbonylmethylene-α-
D-xylofuranose (11): To a solution of 10
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
(1 eq, 12.9 mmol, 6.4 g) in THF (100 mL) at 0 º C was added TBAF (1.2 eq, 15.5 mmol, 1M solution in
THF). The brown reaction mixture was stirred under argon at 4 °C for 3 h. The cold bath was removed and
the reaction mixture was stirred at room temperature for 2 h. The reaction was quenched with a saturated
aqueous solution of NaHCO (100 mL), extracted with DCM (3 x 80 mL), dried over Na SO , filtered and
3
2
4
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
concentrated under reduced pressure. The crude was purified by flash column chromatography
(
DCM/Petroleum Ether: 9/1 then DCM/MeOH: 100/0 to 96/4) to give the desired compound as a yellow oil
3.06 g, 11.86 mmol, 92%). Spectroscopic data of this compound were consistent with those reported in the
(
2
7
1
literature . TLC (DCM/MeOH:99/1): R
Hz, 1H, H ), 5.90 (t, JCH,2 = JCH,4 = 1.8 Hz, 1H, CH=C), 5.75–5.68 (m, 1H, H
q, J = 7.1 Hz, 2H, CH CH ), 3.92 (dd, Jgem = 12.2 Hz and J4,5a = 3.3 Hz, 1H, H5a), 3.72 (dd, Jgem = 12.2 Hz
and J4,5b = 4.3 Hz, 1H, H5b), 1.50 (s, 3H, Meacetonide), 1.42 (s, 3H, Meacetonide), 1.30 (t, J = 7.1 Hz, 3H,
f
= 0.18. H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 5.92 (d, J1,2 = 4.2
1
2
), 4.92–4.86 (m, 1H, H
4
), 4.23
(
2
3
1
3
CH
2
CH
3 3 3 2
). C NMR (75 MHz, CDCl ): δ (ppm) = 164.9 (CO), 154.6 (C ), 116.9 (CHCO Et), 113.0
(CIV(acetonide)), 105.0 (C ), 80.5 (C ), 78.5 (C ), 63.1 (C ), 61.0 (CH CH ), 27.5 (Meacetonide), 27.2 (Meacetonide),
1
4
2
5
2
3
+
+
+
1
2 3 4 18 6
4.3 (CH CH ). MS (ESI ): m/z = 281.1 Da [MNH ] . HRMS (ESI ): calcd for C12H O Na = 281.1001
Da, found = 281.1005 Da.
,2-O-isopropylidene-3-deoxy-3-C-[(ethoxycarbonyl)methyl]-α-
8.5 mmol, 2.2 g) in EtOH (35 mL) was added 10% Pd/C (5% weigth, 110 mg). The reaction was stirred
1
D-ribofuranose (12): To a solution of 11
(
under an atmosphere of hydrogen (1 bar) at room temperature for 24 h. The mixture was filtered and
concentrated under reduced pressure to give 12 as a single product (2.2 g, 8.5 mmol, quantitative).
28b
Spectroscopic data of this compound were consistent with those reported in the literature.
TLC
1
(DCM/Et
2
O:9/1): R
f
= 0.40. H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 5.82 (d, J1,2 = 3.9 Hz, 1H, H
1
), 4.78 (t,
J1,2 = J2,3 = 3.9 Hz, 1H, H ), 4.16 (q, J = 7.1 Hz, 2H, CH CH ), 3.89–3.85 (m, 2H, H and H ), 3.56 (dd,
2
2
3
4
5a
J
gem = 13.2 Hz and J4,5b = 4.2 Hz, 1H, H5b), 2.70 (dd, Jgem = 16.3 Hz and J = 7.9 Hz, 1H, CH
2
CO), 2.49–2.34
(m, 2H, CH CO, H ), 1.98 (brs, 1H, OH), 1.49 (s, 3H, Meacetonide), 1.32 (s, 3H, Meacetonide), 1.27 (t, J = 7.1
2
3
1
3
Hz, 3H, CH
2
CH
3
). C NMR (75 MHz, CDCl
3
):
δ
1
(ppm) = 172.5 (CO), 111.9 (CIV(acetonide)), 104.9 (C ), 81.8
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(
C
2
), 81.5 (C
4.3 (CH CH
found = 283.1160 Da. [α]
,2-O-isopropylidene-3-deoxy-3-C-[(ethoxycarbonyl)methyl]-5-O-benzyl-α-
4
), 61.5 (C
5
), 61.0 (CH
2
CH
3
), 39.8 (C
3 2 2
), 30.0 (CH CO Et), 26.8 (Meacetonide), 26.4 (Meacetonide),
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
+
+
+
1
2
3
). MS (ESI ): m/z = 283.1 [MNa] . HRMS (ESI ): calcd for C12
H
20
O
6
Na = 283.1158 Da,
2
D
0
3
= +59.4 (CHCl , c = 1 g/100mL).
1
D-ribofuranose (13): To a
solution of 12 (1 eq, 3.8 mmol, 1 g) in dry DMF (30 mL) at 0 ºC was added slowly NaH (1.2 eq, 4.61 mmol,
185 mg of a 60% suspension in oil). The mixture was stirred under argon at 5 °C for 10 min and then was
added BnBr (1.5 eq, 5.8 mmol, 0.70 mL). The cold bath was removed and the reaction mixture was stirred at
room temperature until completion (TLC monitoring) and then cooled to 0 °C. The reaction mixture was
quenched with NH Cl (30 mL) and extracted with Et O (3 x 30 mL). The combined organic layers were
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
4
2
washed with brine (5 x 30 mL), dried over Na
2
SO
4
, filtered and concentrated under reduced pressure. The
crude was purified by flash column chromatography (DCM/Petroleum Ether: 9/1 to 9/0) to give the desired
1
compound as a yellow oil (1.1 g, 3.14 mmol, 84%). TLC (DCM/Et
CDCl ): (ppm) = 7.37–7.28 (m, 5H, Har), 5.84 (d, J1,2 = 3.9 Hz, 1H, H
3.9 Hz, 1H, H ), 4.60 (d, Jgem = 12.2 Hz, 1H, CH Ph), 4.53 (d, Jgem = 12.2 Hz, 1H, CH
Hz, 2H, CH CH
2
O:9/1): R
f
= 0.31. H NMR (300 MHz,
3
δ
1
), 4.78 (dd, J1,2 = 3.9 Hz and J2,3
=
2
2
2
Ph), 4.13 (q, J = 7.1
2
3
4
), 3.97–3.92 (m, 1H, H ), 3.65 (dd, Jgem = 10.8 Hz and J4,5a = 3.5 Hz, 1H, H5a), 3.53 (dd,
3
J
gem = 10.8 Hz and J4,5b = 4.4 Hz, 1H, H5b), 2.64 (dd, Jgem = 15.9 Hz and J = 9.4 Hz, 1H, CH
2
CO), 2.44–
2
.31 (m, 2H, CH
2
CO, H
3
), 1.49 (s, 3H, Meacetonide), 1.31 (s, 3H, Meacetonide), 1.26 (t, J = 7.1 Hz, 3H,
(ppm) = 172.2 (CO), 138.1 (CIV(ar)), 128.7 (Car), 128.5 (Car), 127.8
13
CH
2
CH
3
). C NMR (75 MHz, CDCl ): δ
3
(C
ar), 127.1 (Car), 111.7 (CIV(acetonide)), 105.0 (C
1
), 81.2 (C
2
4
), 79.8 (C ), 73.6 (CH
2
Ph), 69.5 (C
5
), 60.7
+
(
CH
373.1 Da [MNa] . HRMS (ESI+): calcd for C19
+31.6 (CHCl , c = 1 g/100mL).
2
CH
3
), 41.7 (C
3
), 29.9 (CH
2
CO
2
Et), 26.8 (Meacetonide), 26.5 (Meacetonide), 14.3 (CH
2
CH
3
). MS (ESI ): m/z
+
20
D
=
H
26
O
6
Na = 373.1627 Da, found = 373.1631 Da. [α]
=
3
1,2-O-isopropylidene-5-O-tert-butyldiphenylsilyl-α-
mmol, 950 mg) in a mixture of EtOH/water 9/1 (13.5 mL/1.5 mL) at 0 ºC was added NaBH
mmol, 317 mg) portionꢀwise. The reaction mixture was stirred overnight at room temperature. The reaction
D-ribofuranose (14): To a solution of 9 (1 eq, 2.2
4
(3.6 eq, 8.3
4
mixture was concentrated under reduced pressure, diluted in mixture DCM/NH Cl (25 mL/25 mL) and
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The Journal of Organic Chemistry
Page 24 of 50
stirred for 30 min. The aqueous layer was extracted with DCM (25 mL), the combined organic layers were
washed with NH Cl, water and brine, then dried over Na SO , filtered and concentrated under reduced
pressure to give the desired compound as a pink oil (900 mg, 2.1 mmol, 95%). Except for the Hꢀ3 and Hꢀ4
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
4
2
4
42
attribution, spectroscopic data of this compound were consistent with those reported in the literature. TLC
1
(DCM/PE:9/1): R
f
= 0.40. H NMR (300 MHz, CDCl
3
):
), 4.61 (dd, J2,3 = 5.0 Hz and J1,2 = 3.9 Hz, 1H, H
), 3.98–3.93 (m, 1H, H5a), 3.88–3.82 (m, 2H, H5b and H
δ
(ppm) = 7.71–7.67 (m, 4H, Har), 7.41–7.37 (m,
), 4.14 (dd, J3,4
), 2.30 (brs, 1H,
): δ
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
6
H, Har), 5.85 (d, J1,2 = 3.9 Hz, 1H, H
.1 Hz and J2,3 = 5.0 Hz, 1H, H
1
2
=
8
3
4
13
OH), 1.56 (s, 3H, Meacetonide), 1.38 (s, 3H, Meacetonide), 1.05 (s, 9H, MetBu). C NMR (75 MHz, CDCl
3
(
ppm) = 135.8 (C ), 135.7 (C ), 133.5 (CIV(ar)), 133.3 (CIV(ar)), 129.8 (C ), 127.8 (C ), 112.7 (CIV(acetonide)),
ar
ar
ar
ar
1
04.3 (C
1
), 81.4 (C
2
), 78.9 (C
3
), 71.4 (C
4
), 62.5 (C
5
), 26.9 (Meacetonide), 26.9 (MetBu), 26.8 (Meacetonide), 19.4
+
+
+
(C
IV(tBu)). MS (ESI ): m/z = 451.2 Da [MNa] . HRMS (ESI ): calcd for C24
H
32
O
5
SiNa = 451.1911 Da,
2
D
0
found = 451.1904 Da. [α]
3
= +28.2 (CHCl , c = 1 g/100mL).
1,2-O-isopropylidene-3-O-benzyl-5-O-tert-butyldiphenylsilyl-α-D-ribofuranose (15): To a solution of 14
(1 eq, 2.1 mmol, 900 mg) in dry THF (30 mL) at 4 ºC was added NaH (1.2 eq, 2.52 mmol, 101 mg of a 60%
suspension in oil) under argon and portionꢀwise. Benzyl bromide (1 eq, 2.1 mmol, 900 mL) was then added
slowly. The reaction mixture was stirred overnight at room temperature. The mixture was diluted with Et
2
O,
washed with NH Cl, water and brine then dried over Na SO , filtered and concentrated under reduced
4
2
4
pressure. The crude was purified by flash column chromatography (Petroleum Ether/AcOEt: 9/1) to give the
1
desired compound as a colorless oil (1 g, 1.93 mmol, 92%). TLC (PE/AcOEt: 9/1): R
MHz, CDCl3):
H, H ), 4.78 (d, Jgem = 12.1 Hz, 1H, CHH'Ph), 4.60 (d, Jgem = 12.1 Hz, 1H, CHH'Ph), 4.61 (t, J1,2 = J2,3
4.0 Hz, 1H, H ), 4.14 (dt, J = 8.8 Hz and J4,5 = 2.3 Hz, 1H, H ), 4.06 (dd, J = 8.8 Hz and J2,3 = 4.1 Hz,
f
= 0.21. H NMR (300
δ
(ppm) = 7.68–7.65 (m, 4H, Har(Ph)), 7.43–7.30 (m, 11H, Har(Ph, Bn)), 5.77 (d, J1,2 = 3.6 Hz,
1
1
=
2
3,4
4
3,4
1H, H
3
), 3.98 (dd, Jgem = 11.8 Hz and J4,5a = 1.6 Hz, 1H, H5a), 3.80 (dd, Jgem = 11.8 Hz and J4,5b = 2.7 Hz,
13
1
H, H5b), 1.60 (s, 3H, Meacetonide), 1.38 (s, 3H, Meacetonide), 1.01 (s, 9H, MetBu). C NMR (75 MHz, CDCl
3
):
δ
(ppm) = 137.9 (CIV(Bn)), 135.8 (C ), 135.7 (C ), 133.7 (CIV(ar)), 133.3 (C
), 129.77ꢀ127.77 (C ), 113.0
ar
ar
IV(ar)
ar
(CIV(acetonide)), 104.3 (C
1
), 79.7 (C
4
), 77.8 (C
2
), 76.8 (C
3
), 72.4 (CH
2
Ph), 61.9 (C
5
), 27.1 (Meacetonide), 26.9
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The Journal of Organic Chemistry
+
+
+
(
[
(
MetBu), 26.7 (Meacetonide), 19.5 (CIV(tBu)). MS (CI ): m/z = 536.2 Da [MNH
4
] ; (ESI ): m/z = 541.2 Da
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
+
+
20
D
MNa] . HRMS (ESI ): calcd for C H O SiNa = 541.2386 Da, found = 541.2390 Da. [α]
= +8.9
31
38
5
3
CHCl , c = 1 g/100mL).
1
,2-O-isopropylidene-3-O-benzyl-α-
D
-ribofuranose (16): To a solution of 15 (1 eq, 1.6 mmol, 800 mg) in
THF (20 mL) at 0 ºC was added TBAF (1.1 eq, 1.8 mmol, 1.8 mL of a 1M solution in THF). The yellow
reaction mixture was stirred at 4 °C for 3 h, then at room temperature overnight. The reaction mixture was
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
quenched with a saturated aqueous solution of NaHCO
3
(15 mL), extracted with DCM (3 x 15 mL), dried
over Na SO , filtered and concentrated under reduced pressure. The crude was purified by flash column
2
4
chromatography (DCM/ MeOH: 98/2 to 97/3) to give the desired compound as an orange oil (360 mg, 1.28
1
mmol, 80%). TLC (DCM/MeOH:98/2): R
H, Har), 5.72 (d, J1,2 = 3.6 Hz, 1H, H
CHH'Ph), 4.57 (dd, J2,3 = 4.3 Hz and J1,2 = 3.6 Hz, 1H, H
f
= 0.35. H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 7.38–7.30 (m,
5
1
), 4.77 (d, Jgem = 11.9 Hz, 1H, CHH'Ph), 4.59 (d, Jgem = 11.9 Hz, 1H,
), 4.11 (dt, J3,4 = 9.1 Hz and J4,5a = J4,5b = 2.7 Hz,
), 3.91 (dd, Jgem = 12.5 Hz and J4,5a = 2.4 Hz, 1H, H5a), 3.84 (dd, J3,4 = 9.1 Hz and J2,3 = 4.3 Hz, 1H,
3
), 3.64 (dd, Jgem = 12.5 Hz and J4,5b = 3.0 Hz, 1H, H5b), 1.73 (brs, 1H, OH), 1.60 (s, 3H, Meacetonide), 1.36
2
1
H, H
4
H
13
(s, 3H, Meacetonide). C NMR (75 MHz, CDCl
3
):
δ (ppm) = 137.7 (CIV(ar)), 128.6 (Car), 128.2 (Car(p)), 128.1
(C
ar), 113.2 (CIV(acetonide)), 104.2 (C
1
), 78.9 (C
4
), 77.7 (C
2
), 76.7 (C
3
2 5
), 72.5 (CH Ph), 60.7 (C ), 27.0
+
+
+
+
(Meacetonide), 26.6 (Meacetonide). MS (ESI ): m/z = 303.1 Da [MNa] , 583.2 Da [2MNa] . HRMS (ESI ): calcd
2
D
0
for C15
H
20
O
5
Na = 303.1208 Da, found 303.1210 Da. [α]
3
= +54 (CHCl , c = 1 g/100mL).
1,2-O-isopropylidene-3-O-benzyl-5-O-mesyl-α-D-ribofuranose (17): To a solution of 16 (1 eq, 4.5 mmol,
.27 g) in pyridine (18 mL) at 0 ºC was added dropwise mesylchloride (2 eq, 9 mmol, 0.7 mL). The mixture
1
was stirred overnight under argon. The reaction mixture was quenched with water (2 mL), concentrated
under reduced pressure and coꢀevaporated with toluene (20 mL). The residue was diluted with DCM (30
mL), washed with a saturated aqueous solution of NaHCO
Na SO , filtered and concentrated under reduced pressure to obtain the desired compound as a brown oil
1.64 g, 4.5 mmol, quantitative). TLC (PE/AcOEt: 9/1): R = 0.25. H NMR (300 MHz, CDCl ):
3
(3 x 20 mL), brine (1 x 20 mL) then dried over
2
4
1
(
δ
(ppm) =
f
3
7.38ꢀ7.33 (m, 5 H, Har), 5.73 (d, J1,2 = 3.6 Hz, 1H, H
1
), 4.77 (d, Jgem = 11.8 Hz, 1H, CHH'Ph), 4.58 (d, Jgem =
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The Journal of Organic Chemistry
1.8 Hz, 1H, CHH'Ph), 4.57 (t, J1,2 = J2,3 = 4.0 Hz, 1H, H ), 4.48–4.43 (m, 1H, H5a), 4.27 (dd, Jgem = 13.3
Hz and J4,5b = 4.0 Hz, 1 H, H5b), 4.24 (ddd, J3,4 = 9.0 Hz, J4,5b = 4.0 Hz and J4,5a = 1.8 Hz, 1H, H ), 3.76 (dd,
3,4 = 9.0 Hz and J2,3 = 4.0 Hz, 1H, H ), 2.98 (s, 3H, MeSO ), 1.59 (s, 3H, Meacetonide), 1.37 (s, 3H,
(ppm) = 137.2 (CIV(ar)), 128.8 (Car), 128.4 (Car), 128.3 (Car),
Page 26 of 50
1
2
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
4
J
3
2
1
3
Meacetonide). C NMR (75 MHz, CDCl
3
):
δ
1
13.5 (CIV(acetonide)), 104.3 (C
1
), 77.4 (C
2
), 77.0 (C
3
), 76.4 (C
4
), 72.6 (CH
2
Ph), 67.8 (C
5
), 37.7 (MeSO
2
), 26.9
Na
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
+
+
(Meacetonide), 26.6 (Meacetonide). MS (ESI ): m/z = 381.1 Da [MNa] . HRMS (ESI ): calcd for C16
H
22
O
7
S
1
20
D
=
381.0978 Da, found 381.0977 Da. [α]
3
= +8.1 (CHCl , c = 1 g/100mL).
1,2-O-isopropylidene-3-O-benzyl-5-azido-α-D-ribofuranose (18): To a solution of 17 (1 eq, 3.2 mmol,
1.16 g) in DMF (30 mL) was added sodium azide (1.5 eq, 4.8 mmol, 312 mg) at 0 ºC. The reaction mixture
was stirred 3 h at 90 °C. The mixture was quenched with water (30 mL), extracted with Et
2
O (3 x 30 mL).
The combined organic layers were washed with brine, then dried over Na SO , filtered and concentrated
2
4
under reduced pressure to give the desired compound as a yellow oil (940 mg, 3.07 mmol, 95%). TLC
1
(
DCM/PE: 7/3): R
3.6 Hz, 1H, H
1.9 Hz, 1H, CHH'Ph), 4.22–4.16 (m, 1H, H
gem = 13.5 Hz and J4,5a = 2.6 Hz, 1H, H5a), 3.24 (dd, Jgem = 13.5 Hz and J4,5b = 3.9 Hz, 1H, H5b), 1.59 (s, 3H,
f
= 0.54. H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 7.38–7.32 (m, 5H, Har), 5.75 (d, J1,2
), 4.55 (d, Jgem
), 3.77 (dd, J3,4 = 8.9 Hz and J2,3 = 4.2 Hz, 1H, H
=
1
), 4.77 (d, Jgem = 11.9 Hz, 1H, CHH'Ph), 4.58 (t, J2,3 = J1,2 = 3.9 Hz, 1H, H
2
=
1
4
3
), 3.66 (dd,
J
13
Meacetonide), 1.36 (s, 3H, Meacetonide). C NMR (75 MHz, CDCl
3
):
δ (ppm) = 137.4 (CIV(ar)), 128.7 (Car),
128.4 (Car(p)), 128.2 (Car), 113.4 (CIV(acetonide)), 104.2 (C
1
), 77.9 (C
3
), 77.5 (C
4
), 77.4 (C
2
2
), 72.5 (CH Ph), 50.6
+
+
+
(C
5
), 26.9 (Meacetonide), 26.6 (Meacetonide). MS (ESI ): m/z = 328.1 Da [MNa] . HRMS (ESI ): calcd for
20
C
15
H
19
N
3
O
4
Na = 328.1268 Da, found 328.1264 Da. [α]
D
3
= +158.3 (CHCl , c = 1 g/100mL).
1,2-O-isopropylidene-3-O-benzyl-5-amino-α-D-ribofuranose (19): The compound 18 (1.1 mmol, 340 mg)
was hydrogenated in the presence of 10% Pd/C (5% weigth, 17 mg) in EtOH (5 mL) under a hydrogen
atmosphere. After stirring at room temperature for 4 h, the TLC analysis (DCM) showed disappearance of
the starting material. The mixture was filtered and concentrated under reduced pressure to give the desired
1
compound as a light brown oil (290 mg, 1.04 mmol, 94%). H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 7.35–
7.28 (m, 5H, H ), 5.72 (d, J = 3.7 Hz, 1H, H ), 4.80 (d, J
= 12.0 Hz, 1H, CHH'Ph), 4.60 (t, J1,2 = J2,3
=
gem
ar
1,2
1
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The Journal of Organic Chemistry
4
.1 Hz, 1H, H
2
), 4.54 (d, Jgem = 12.0 Hz, 1H, CHH'Ph), 4.05 (ddd, J3,4 = 9.0 Hz, J4,5b = 5.4 Hz and J4,5a = 3.2
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
4 3
Hz, 1H, H ), 3.64 (dd, J3,4 = 9.0 Hz and J2,3 = 4.3 Hz, 1H, H ), 3.04 (dd, Jgem = 13.8 Hz and J4,5a = 3.4 Hz,
1H, H5a), 2.73 (dd, Jgem = 13.8 Hz and J4,5b = 5.4 Hz, 1H, H5b), 1.60 (s, 3H, Meacetonide), 1.36 (s, 3H,
1
3
Meacetonide). C NMR (75 MHz, CDCl
3
): δ (ppm) = 137.7 (CIV(ar)), 128.6 (Car), 128.5 (Car), 128.14 (Car),
1
28.06 (Car), 128.0 (Car), 113.0 (CIV(acetonide)), 104.1 (C
1
), 79.5 (C
3
), 78.2 (C
4
), 77.5 (C
2
2
), 72.1 (CH Ph), 42.5
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
+
+
(C
5
), 26.8 (Meacetonide), 26.6 (Meacetonide). MS (ESI ): m/z = 280.2 Da [MH] . HRMS (ESI ): calcd for
2
D
0
C
15
H
22
N
1
O
4
= 280.1549 Da, found 280.1546 Da. [α]
3
= +53.8 (CHCl , c = 1 g/100mL).
1
,2-O-isopropylidene-3-(1,2-O-isopropylidene-3-O-benzyl-5-deoxy-5-acetamido-α-
benzyl-α- -ribofuranose (20): To a solution of 6 (1 eq, 1.5 mmol, 483 mg) in DCM (18 mL) at 0 ºC was
added TBTU (1.4 eq, 2.1 mmol, 690 mg), HOBt.H O (0.5 eq, 0.75 mmol, 100 mg) and triethylamine (1.1
D-ribosyl)-5-O-
D
2
eq, 1.65 mmol, 230 µL). The mixture was stirred under argon for 30 min at 5 °C. The amine 19 was then
added (1 eq, 1.5 mmol, 419 mg) followed by triethylamine (1.1 eq, 1.65 mmol, 230 µL), and the reaction
mixture was stirred overnight at room temperature. The reaction mixture was quenched with water (35 mL),
extracted with DCM (3 x 35 mL) and the combined organic layers were washed with brine (2 x 30 mL),
dried over Na
2
SO
4
, filtered and concentrated under reduced pressure. The crude was diluted with toluene
SO , filtered and
and washed with water (3 x 30 mL) to eliminate urea. The organic layer was dried over Na
2
4
concentrated under reduced pressure. The crude was purified by flash column chromatography (DCM to
DCM/MeOH: 99/1) to give the desired compound as a beige sticky oil (880 mg, 1.5 mmol, quantitative).
1
TLC (DCM/MeOH: 98/2): R
f
= 0.21. H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 7.41–7.29 (m, 10H, Har),
5
.83 (dd, JNH,5Ba = 5.6 Hz and JNH,5Bb = 4.3 Hz, 1H, NHamide), 5.74 (d, J1A,2A = 3.8 Hz, 1H, H1A), 5.67 (d,
1B,2B = 3.7 Hz, 1H, H1B), 4.78 (d, Jgem = 12.3 Hz, 1H, CHH'Ph ), 4.60 (d, Jgem = 12.3 Hz, 1H, CHH'Ph
4.59 (d, Jgem = 12.3 Hz, 1H, CHH'Ph ), 4.54 (d, Jgem = 12.3 Hz, 1H, CHH'Ph
Hz, 1H, H ), 4.36 (t, J = J2A,3A = 3.8 Hz, 1H, H ), 4.16–4.10 (m, 1H, H ), 3.93–3.88 (m, 1H, H ),
J
B
B
),
A
A
), 4.50 (t, J1B,2B = J2B,3B = 3.9
2
B
1A,2A
2A
4B
4A
3
.70–3.61 (m, 2H, H5Aa and H5Ba), 3.55–3.45 (m, 3H, H5Ab,
H5Bb and H3B), 2.42–2.23 (m, 2H, H3A, CHH'CO),
2
.26–2.20 (m, 1H, CHH'CO), 1.59 (s, 3H, Meacetonide), 1.46 (s, 3H, Meacetonide), 1.36 (s, 3H, Meacetonide), 1.23
1
3
(s, 3H, Meacetonide). C NMR (100 MHz, CDCl
3
):
δ
(ppm) = 171.1 (COamide), 138.1 (CIV(ar)), 137.6 (CIV(ar)),
27
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Page 28 of 50
1
28.7 (Car), 128.6 (Car), 128.2 (Car), 128.1 (Car), 127.9 (Car),127.9 (Car), 113.4 (CIV(acetonide)), 111.6
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
(
(
(
CIV(acetonide)), 105.2 (C1A), 104.2 (C1B), 81.2 (C2A), 80.3 (C4A), 78.5 (C3B), 77.7 (C2B), 77.4 (C4B), 73.7
CH
2
Ph
A
), 72.4 (CH
2
Ph
B
2
), 70.0 (C5A), 42.7 (C3A), 39.7 (C5B), 32.5 (CH CO), 26.91 (Meacetonide), 26.88
+
+
+
Meacetonide), 26.63 (Meacetonide), 26.61 (Meacetonide). MS (CI ): m/z = 584.4 Da [MH] , 601.4 Da [MNH ] .
4
+
20
HRMS (MALDI : DHB, PEG 600): calcd for C32
25 (CHCl , c = 1 g/100mL).
,2-O-isopropylidene-3-deoxy-3-propylacetamido-5-O-benzyl-α-D-ribofuranose (21): To a cooled
H
41
N
1
O
9
Na = 606.2674 Da, found 606.2662 Da. [α]
D
=
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
+
3
1
solution (0 °C) of 6 (1 eq, 1.86 mmol, 600 mg) in DCM (30 mL) was added TBTU (1.4 eq, 2.6 mmol, 840
mg), HOBt.H O (0.5 eq, 0.9 mmol, 126 mg) and triethylamine (1.1 eq, 2 mmol, 285 µL). The mixture was
2
stirred for 30 min at 5 °C. Then was added propylamine (1.5 eq, 2.8 mmol, 230 µL) and triethylamine (1.1
eq, 1.24 mmol, 285 µL), the mixture was stirred overnight at room temperature. The reaction was quenched
with water (40 mL), extracted with DCM (3 x 40 mL) and the combined organic layers were washed with a
saturated aqueous solution of NaHCO
concentrated under reduced pressure. If necessary, the crude was diluted with toluene and washed with
water (3 x 40 mL) to eliminate urea. The organic layer was dried over Na SO , filtered and concentrated
3 2 4
(2 x 40 mL) and brine (2 x 40 mL), dried over Na SO , filtered and
2
4
under reduced pressure to obtain the desired compound as a yellow oil (570 mg, 1.578 mmol, 74%). TLC
1
(DCM): R
f
= 0.84. H NMR (300 MHz, CDCl
3
):
δ
(ppm) = 7.33–7.32 (m, 5H, Har), 5.83 (d, J1,2 = 3.7 Hz,
1H, H ), 5.71 (brs, 1H, NHamide), 4.69 (d, J = 3.7 Hz, 1H, H ), 4.56 (s, 2H, CH Ph), 3.95 (ddd, J = 8.9
1 1,2 2 2 3,4
Hz and J4,5a = J4,5b = 4.4 Hz, 1H, H
4
), 3.64 (dd, Jgem = 10.7 Hz and J4,5a = 3.5 Hz, 1H, H5a), 3.53 (dd, Jgem
=
=
10.7 Hz and J4,5b = 4.8 Hz, 1H, H5b), 3.25 (dq, Jgem = 18.0 Hz and JHa(CH2NH),Ha(CH2CH3) = JHa(CH2NH),Hb(CH2CH3)
=
JHa(CH2NH),NH = JHa(CH2NH),NH = 6.6 Hz, 1H, Ha(CH2NH)), 3.14 (dq, Jgem = 18.0 Hz and JHb(CH2NH),Ha(CH2CH3)
J
Hb(CH2NH),Hb(CH2CH3) = JHb(CH2NH),NH = JHb(CH2NH),NH = 6.6 Hz, 1H, Hb(CH2NH)), 2.44–2.37 (m, 2H, Ha(CH2CO) and
H ), 2.28–2.18 (m, 1H, Hb(CH2CO)), 1.50 (q, JCH2,CH3 = 7.4 Hz, 2H, CH CH ), 1.50 (s, 3H, Meacetonide), 1.31 (s,
3
2
3
13
3
H, Meacetonide), 0.91 (t, JCH2,CH3 = 7.4 Hz, 3H, CH
2
CH
3
). C NMR (75 MHz, CDCl
3
):
δ
(ppm) = 170.1
),
(COamide), 137.0 (CIV(ar)), 128.9 (Car), 127.5 (Car), 126.9 (Car), 126.8 (Car), 110.7 (CIV(acetonide)), 104.0 (C
1
8
2 4 2 5 3 2 2
0.3 (C ), 79.3 (C ), 72.7 (CH Ph), 69.0 (C ), 41.8 (C ), 40.4 (NHꢀCH ), 31.6 (CH
CO), 25.8 (Meacetonide),
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The Journal of Organic Chemistry
+
+
+
2
5.5 (Meacetonide), 21.9 (CH
2
CH
3
), 10.4 (CH
2
CH
3
). MS (ESI ): m/z = 386.2 Da [MNa] . HRMS (ESI ): calcd
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
for C20
H
29
N
1
O
5
Na = 386.1943 Da, found = 386.1945 Da.
1,2-O-isopropylidene-3-deoxy-3-propyl-N-methyl-acetamido-5-O-benzyl-α-D-ribofuranose (22): To a
cooled solution of 6 (1 eq, 1.55 mmol, 500 mg) in DCM (30 mL) was added TBTU (1.4 eq, 2.17 mmol, 600
mg), HOBt.H O (0.5 eq, 0.78 mmol, 90 mg) and triethylamine (1.1 eq, 1.7 mmol, 200 µL). The mixture was
2
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
stirred for 30 min about 5 °C. Then was added methylpropylamine (1.5 eq, 2.32 mmol, 210 µL) and
triethylamine (1.1 eq, 1.7 mmol, 200 µL), the mixture was stirred overnight at room temperature. The
reaction was quenched with water (30 mL), extracted with DCM (3 x 30 mL) and the combined organic
layers were washed with a saturated aqueous solution of NaHCO
over Na SO , filtered and concentrated under reduced pressure. The crude was purified by flash column
chromatography (DCM/Et
3
(2 x 40 mL) and brine (2 x 30 mL), dried
2
4
2
O, 99/1) to obtain the desired compound as a colorles oil (375 mg, 1 mmol,
1
65%). TLC (DCM/Et O: 9/1): R = 0.34. H NMR (400 MHz, CDCl ) of two rotamers in 1/1 ratio:
δ
(ppm)
2
f
3
=
7.37–7.29 (m, 5H, Har), 5.82 (d, J1,2 = 3.8 Hz, 0.5H, H
1
1
), 5.81 (d, J1,2 = 3.8 Hz, 0.5H, H ), 4.83 (t, J1,2 = J2,3
=
4.2 Hz, 1H, H ), 4.56–4.55 (m, 2H, CH Ph), 4.02–3.96 (m, 1H, H ), 3.65–3.56 (m, 2H, H ), 3.41–3.16 (m,
2
2
4
5
2H, CH
2
NMe), 2.92 (s, 1.5H, NMe), 2.90 (s, 1.5H, NMe), 2.70–2.59 (m, 2H, Ha(CH2CO), H
3
), 2.34 (dd, Jgem =
1
5.9 Hz and JCH2,H3 = 3.5 Hz, 0.5H, Hb(CH2CO)), 2.29 (dd, Jgem = 15.9 Hz and JCH2,H3 = 3.5 Hz, 0.5H,
H
2 3
b(CH2CO)), 1.60–1.49 (m, 2H, CH CH ), 1.48 (s, 1.5H, Meacetonide), 1.47 (s, 1.5H, Meacetonide), 1.29 (s, 3H,
1
3
Meacetonide), 0.88 (t, JCH2,CH3 = 7.4 Hz, 1.5H, CH
(ppm) = 170.99 (COamide), 170.98 (COamide), 138.3 (CIV(ar)), 138.2 (CIV(ar)), 128.6 (Car),
28.5 (Car), 128.5 (Car), 127.9 (Car), 127.8 (Car), 127.7 (Car), 127.1 (Car), 111.4 (CIV(acetonide)), 111.4
2 3 2 3
CH ), 0.87 (t, JCH2,CH3 = 7.4 Hz, 1.5H, CH CH ). C NMR
3
(100 MHz, CDCl ): δ
1
(C
IV(acetonide)), 105.2 (C
1
), 105.1 (C
1
), 81.5 (C
2
), 80.0 (C
4
), 79.9 (C
4
), 73.7 (CH
2
Ph), 73.6 (CH
2
Ph), 70.7 (C
5
),
7
0.5 (C
5
), 51.6 (CH
2
NMe), 49.6 (CH
2
NMe), 42.9 (C
3
), 42.7 (C
3
), 35.4 (CH
2
2
CO), 33.5 (CH CO), 29.0
(NMe), 28.5 (NMe), 26.88 (Meacetonide), 26.86 (Meacetonide), 26.48 (Meacetonide), 26.46 (Meacetonide), 21.5
+
+
(
[
(
CH
2
CH
3
), 20.7 (CH
2
CH
3
), 11.3 (CH
2
CH
3
), 11.2 (CH
2
CH
3
). MS (ESI ): m/z = 400.2 Da [MNa] , 777.4 Da
+
+
20
D
2MNa] . HRMS (ESI ): calcd for C21
H
31
N
1
O
5
Na = 400.2094 Da, found = 400.2103 Da. [α]
= +13.6
3
CHCl , c = 1 g/100mL).
29
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