Efficient Synthesis of Ratiometric Fluorescent Nucleosides Featuring 3-Hydroxychromone Nucleobases

Article abstract of DOI:10.1016/j.tet.2009.07.021

The synthesis of a novel class of fluorescent nucleosides featuring 2-aryl-3-hydroxychromones (3-HC) as base analogues is described. Nucleoside 1a bearing the 2-thienyl-3-HC nucleobase was prepared using sequential aryl-aldol condensation/cycloetherificat

Full text of DOI:10.1016/j.tet.2009.07.021

Tetrahedron 65 (2009) 7809–7816
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Efficient Synthesis of Ratiometric Fluorescent Nucleosides Featuring
3-Hydroxychromone Nucleobases
Marie Spadafora a, Victoria Y. Postupalenko b, Volodymyr V. Shvadchak b, Andrey S. Klymchenko b,
b
a
,
a,
*
*
´
Yves Mely , Alain Burger , Rachid Benhida
a
´
ˆ
´
Laboratoire de Chimie des Molecules Bioactives et des Aromes, UMR 6001 CNRS, Institut de Chimie de Nice, Universite de Nice-Sophia Antipolis,
Parc Valrose, 06108 Nice Cedex 2, France
b Laboratoire de Biophotonique et Pharmacologie, UMR 7213 CNRS, Universite´ de Strasbourg, Faculte´ de Pharmacie 74, Route du Rhin,
BP 24, 67401 Illkirch cedex, France
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis of a novel class of fluorescent nucleosides featuring 2-aryl-3-hydroxychromones (3-HC) as
base analogues is described. Nucleoside 1a bearing the 2-thienyl-3-HC nucleobase was prepared using
sequential aryl–aldol condensation/cycloetherification or a Friedel–Crafts glycosylation as key steps. The
synthesis of the triazolyl derivative 1b was achieved using a convergent 1,3-dipolar cycloaddition
strategy. Fluorescence studies show that 3-HC-thienyl-nucleoside 1a displays high sensitivity of its dual
emission to polarity changes and therefore is highly promising for nucleic acid labelling.
Ó 2009 Elsevier Ltd. All rights reserved.
Received 16 June 2009
Received in revised form 2 July 2009
Accepted 4 July 2009
Available online 9 July 2009
1. Introduction
On the other hand, 3-hydroxychromones (3-HC) have been shown
to be powerful fluorescence probes for a large range of applications in
Due to its exquisite sensitivity, fluorescence is one of the most
sensitive techniques for investigating biological systems. In the
context of nucleic acids, the common strategy consists to substitute
a natural nucleoside by a fluorescent analogue because the ca-
nonical bases are poorly or non-fluorescent. Most of the fluorescent
analogues used for nucleic acid labelling are single band emitter
probes.1 Among them, 20-deoxyribosyl-2-aminopurine (2-AP),
a mimic of 20-deoxyadenosine, is the most popular fluorescent
base.2 However, 2-AP displays severe limitations related to the
dramatic drop of its quantum yield when incorporated in oligo-
nucleotides.3 Moreover, when protein binding to nucleic acids does
not induce a transition between stacked and unstacked confor-
mations, the interactions are difficult to monitor due to the limited
environment sensitivity of 2-AP. Recently, 8-vinyl-deoxyadenosine
(8-vdA), a new fluorescent analogue of 20-deoxyadenosine was
reported by us.4 Although 8-vdA displays improved sensitivity
compared to 2-AP, its domain of application is similar. Other fluo-
rescent nucleoside analogues have been developed but, when in-
corporated into ODNs, they are quenched, destabilizing or of
limited sensitivity to environmental change. As a consequence,
there is a strong demand for new fluorescent nucleoside analogues
with improved spectroscopic properties and this is the purpose of
intense research.1
model membranes, biomembranes and proteins.5 Due to an excited
state intramolecular proton transfer (ESIPT, Fig. 1),6 these fluo-
rophores exhibit two excited state forms: the initially excited normal
(N
*
) and the tautomeric (T
*
) forms.7 Since the ESIPT reaction in these
dyes is strongly sensitive to polarity, H-bonds8b,9 and electric
fields,10 the ratio of the two emission bands could be used to sensi-
tively monitor environmental changes. Moreover, the positions of the
absorption and emission bands as well as the fluorescence intensity
ratio in 3-HC dyes could be used to further characterize the properties
of the probe environment.8b,11 3-HC fluorophores bearing a small
heterocycle in 2-position are attractive analogues of the nucleic bases
8
H
+
H
O
O
O
O-
Ar
Ar
O
O
N*
T*
hνT*
hνN*
hνabs
H
+
H
O
O
O
O
O-
Ar
Ar
O
N
T
* Corresponding authors. Tel.: þ33 4 92 07 61 74; fax: þ33 4 92 07 61 51.
E-mail addresses: benhida@unice.fr (R. Benhida), burger@unice.fr (A. Burger).
Figure 1. ESIPT reaction in 3-hydroxychromones.
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.07.021

7810
M. Spadafora et al. / Tetrahedron 65 (2009) 7809–7816
because their size corresponds well to the size of the two comple-
mentary (A–T or G–C) bases. Moreover, according to our recent
studies, the 2-(2-furanyl)-3-hydroxychromone dye when conjugated
to spermine is able to intercalate within the base pairs of double-
stranded DNA, giving a strong variation of its dual emission and an
increase in its fluorescence intensity.12
starting from the aldehyde 2 (Path A)15 or the acetyl-deoxyribose
(3a or 3b, Path B) as illustrated in Scheme 2.
Thus, regioselective ortho-lithiation of 2-bromothiophene using
LDA in THF at ꢀ78 ꢁC followed by addition of the protected alde-
hyde 2 led to the alcohol 12 in 75% yield as a mixture of inseparable
R/S isomers (40:60). C-Nucleoside 13 (a/b mixture) was then
To explore the potentiality of 3-HC as fluorescent probes for
DNA labelling, we report here the first synthesis and preliminary
spectroscopic characterization of nucleoside analogues where a 3-
HC substitutes a natural base. The structures of the targeted 3-HC-
nucleosides 1a and 1b are shown in Figure 2. They were designed
obtained from 12 (R/S) following isopropylidene cleavage and
subsequent C40–C10 cycloetherification (85%). However, the sepa-
ration of a/b anomers required conversion of 13 to its acetyl-pro-
tected analogue 14. Interestingly and according to path B, the direct
glycosylation leading to 14 was cleanly achieved in one step using
the catalytic Friedel–Crafts reaction.16 Indeed, treatment of 2-bro-
mothiophene and acetyl-ribose 3a (or 3b) with SnCl4 in CH2Cl2
provided good yield and diastereoselectivity in favour of the de-
(i) to keep a 20-deoxy structure and a
b-anomeric configuration for
incorporation into DNA oligonucleotides and (ii) to have an electron
donating or an electron withdrawing group connected between the
sugar and chromone moieties in order to modulate the photo-
physical properties of the nucleoside analogues.
sired
anomeric configuration was clearly evidenced by the observed NOE
correlations between H10 and H40 for 14 , and between H10–H30
and H10–H5’ for 14
(Scheme 2). The synthesis of 1a was then
pursued from 14 following subsequent acetyl cleavage/TBDPS
b
-anomer 14
b
(73% yield,
b/a
¼70:30). Furthermore, the
b
1,3-dipolar cycloaddition
Stille coupling
a
C-glycosylation
b
HO
O
HO
O
protection to provide 15 (94%).16f Halogen–metal exchange/stan-
nylation followed by Pd-catalyzed Stille type coupling between 15
and 2-bromo-3-HC 9 led to the protected C-nucleoside 16 (72%),
which was quantitatively converted, after MEM and TBDPS cleav-
age, to the 2-thienyl-3-HC C-nucleoside 1a.
N
N
N
O
O
O
O
HO
S
HO
HO
HO
1b
1a
The synthesis of compound 1b was achieved straightforwardly. As
shown in Scheme 3, after in situ cleavage of the TMS group, the 1,3-
dipolar cycloaddition between 10 and azido-sugar 317a was carried
out both in solution using AcOH as a co-catalyst and on SiO2 under
solvent-free and microwave activation, according to our recent
published work.17 Both procedures provided the cycloadduct 17 in
high yields (89–95%). Finally, methanolysis of 17 followed by catalytic
hydrogenolysis afforded the triazolyl-3-HC nucleoside 1b in 86%
yield (two steps). Following the same strategy, but using the 3-HC 11
rather than 10, the deprotection of the MEM group (HCl or TFA) failed
since a concomitant cleavage of the glycosidic bond was observed.
The photophysical properties of compounds 1a and 1b were
investigated in solvents of different polarity. To scale the polarity,
we used the ET(30) index. This empirical parameter accounts for the
dielectric constant of the solvent and its H-bond donor ability,18
both of which influence strongly the fluorescence properties of the
3-HC dyes.8b,11 The results are given in Table 1 and Figures 3 and 4.
In the studied solvents, compound 1a shows an absorption band
centred around 361–369 nm, whereas 1b exhibits a significantly
blue shifted absorption having two distinct maxima at 299–304
and 332–338 nm (Fig. 3). The position of the absorption maximum
slightly increases with ET(30) (Table 1). In all studied solvents,
compound 1a, featuring the electron-rich thienyl heterocycle,
presents a well-resolved dual emission (Fig. 4A), where the short-
and the long-wavelength bands could be assigned to the emission
Figure 2. Targeted 2-aryl-3-HC-nucleosides.
2. Results and discussions
For the synthesis of 1a and 1b we used two convergent strate-
gies based on C-glycosylation–Stille type coupling (for 1a), and
azide-alkyne 1,3-dipolar cycloaddition (for 1b) as key steps (Fig. 2).
The preparation of 3-HC key intermediates 8–11 required for the
synthesis of 1a and 1b is described in Scheme 1. Starting from 2-
hydroxy-acetophenone 5, condensation with N,N-dimethylforma-
mide dimethylacetal followed by HCl-mediated in situ cyclization
provided the chromone 6.13 Epoxidation of 6 using H2O2/NaOH in
CH2Cl2 followed by acid-mediated ring-opening afforded 7 in 82%
overall yield. The use of protic solvents (MeOH, EtOH) or other
oxidizing agents such as m-CPBA and 2-butanone peroxide only
gave low yields.14 Bromination of 7 followed by successive pro-
tection with benzyl or MEM group gave 8 and 9, respectively (83–
87%, two steps). Sonogashira coupling carried out on 8 and 9 using
TMS-acetylene provided the 3-HC 10 and 11, respectively (90–93%).
We found that the cleavage of the TMS group of compounds 10 and
11 resulted in unstable terminal alkynes. Therefore, the TMS-pro-
tected alkynes 10 and 11 were deprotected in situ and used without
purification.
With these key intermediates in hand, the thienyl-derived 3-
HC-C-nucleoside 1a bearing the
b
-C–C linkage was prepared
O
O
O
O
(i) DMF-DMA
μW
(i) H2O2, NaOH
CH2Cl2
(i) NBS,
OH
CH3CN/CCl4
OR
Br
(ii) aqHCl
(ii) HCl
90%
(Ref. 13)
(ii) BnBr or MEMCl
K2CO3, DMF
O
O
7
OH
O
82%
6
5
83-87%
8 : R = Bn
9 : R = MEM
O
TMS-acetylene
Pd(PPh3)2Cl2
OR
toluene, CuI
O
Et3N
TMS
92%
10 : R = Bn
11 : R = MEM
Scheme 1. Synthesis of protected 3-HC 10 and 11.

M. Spadafora et al. / Tetrahedron 65 (2009) 7809–7816
7811
HO
O
Path A
O
p-TSA, toluene
Br
TBDPSO
S
TBDPSO
S
(85%)
(92%)
Br
O
HO
LDA /THF / -78 °C
12 (R/S = 2/3)
13 (α/β = 2/3)
then 2
(75%)
(i) Bu4NF/THF
(ii) Ac2O, Py
S
Br
(73%)
3a, SnCl4, CH2Cl2
Path B
O
O
(i) K2CO3/MeOH
Br
S
Br
TBDPSO
TBDPSO
AcO
S
(ii) TPDPSCl, Imid
(94%)
AcO
14
α/β = 3/7 from 3a
α/β = 2/3 from 13
15 (from 14β)
(i) BuLi, Et2O/-78 °C
Bu3SnCl
(72%)
(ii) 9, Pd(PPh3)4
CuI, toluene
MEM
O
O
(ii) aqHCl, CH2Cl2
(85%)
O
1a
S
TBDPSO
TBDPSO
O
16
Scheme 2. Synthesis of 3-HC C-nucleoside 1a.
Scheme 3. Synthesis of 3-HC nucleoside 1b.
Table 1
Spectroscopic data of nucleosides 1a and 1b in different solventsa
Nucl.
Solv.
ET(30)
labs
3
lN*
lT*
N
*
/T
*
QY
1a
Buffer
MeOH
EtOH
BuOH
DMSO
MeCN
Acetone
CH2Cl2
EtOAc
63.1
55.4
51.9
49.7
45.1
45.6
42.2
40.7
38.1
367
368
369
369
368
362
362
363
361
13,000
12,500
12,300
d
440
434
433
431
431
421
421
418
415
515
545
548
546
554
546
552
543
551
1.72
1.34
0.68
0.52
0.30
0.13
0.22
0.04
0.09
0.046
0.075
0.077
0.091
0.197
0.179
0.053
0.141
0.100
d
d
d
d
d
1b
Buffer
MeOH
EtOH
BuOH
DMSO
MeCN
Acetone
CH2Cl2
EtOAc
63.1
55.4
51.9
49.7
45.1
45.6
42.2
40.7
38.1
338
336
335
333
341
332
334
333
332
11,500
12,000
12,000
d
401
398
397
393
390
387
387
d
500
516
517
518
520
511
515
506
512
0.06
0.17
0.13
0.16
0.06
0.02
0.07
0.022
0.024
0.027
0.033
0.052
0.189
0.037
0.183
0.074
d
d
d
d
<0.01
0.03
d
387
a
labs is the position of the absorption maxima, lN* and lT* are the positions of the fluorescence maxima of the N
*
and T
*
bands, respectively; N
*
/T
*
is the intensity ratio of the
(Mꢀ1 cmꢀ1) is the
two emission bands at their maxima; QY is the fluorescence quantum yield calculated using quinine sulfate (QY¼0.577 in 0.5 M H2SO4) as a reference;
3
extinction coefficient at the maximum of absorption; 10 mM phosphate buffer (pH 6.5) was used.

7812
M. Spadafora et al. / Tetrahedron 65 (2009) 7809–7816
A
A
1.0
0.8
0.6
0.4
0.2
0.0
1.0
0.8
0.6
0.4
0.2
0.0
Buffer
MeOH
EtOH
1.6
1.2
0.8
0.4
0.0
DMSO
MeCN
B
B
1.0
Buffer
MeOH
EtOH
0.8
buffer pH 6.5
MeOH
DMSO
MeCN
0.6
EtOH
DMSO
0.4
MeCN
0.2
0.0
400
450
500
Wavelength, nm
550
600
650
300
350
Wavelength, nm
400
450
Figure 4. Fluorescence spectra of 1a (A) and 1b (B) in different solvents. The spectra
were normalized at the long-wavelength maximum. Excitation wavelength was 360
and 340 nm for 1a and 1b, respectively.
Figure 3. Normalized absorption spectra of nucleosides 1a (A) and 1b (B) in different
solvents.
of the N
*
and T
*
species, respectively. The N
*
*
/T intensity ratio of
1.8
1.5
1.2
0.9
0.6
0.3
0.0
-0.3
this nucleoside increases gradually with increasing ET(30) values
from 0.04 to 0.09 in dichloromethane and ethyl acetate up to 1.72 in
water (Table 1 and Fig. 4A). Figure 5 shows that the N
correlates well with the ET(30) values. This correlation suggests that
the ESIPT reaction and thus, the formation of the T form in 1a is
*
/T
*
ratio
*
hampered especially in polar protic solvents, as for its analogue 2-
(2-furyl)-3-hydroxychromone.11a The hampering of the ESIPT re-
action by protic solvents in 3-HC is connected with the formation of
intermolecular H-bonds that weaken or even disrupt the intra-
molecular H-bond, needed for the ESIPT reaction.8b,9 Moreover, the
increase in ET(30) leads to a large blue shift of the T
cially in water (Table 1), due to its high H-bond donor ability.19 In
contrast, the N band maximum shifts to the red on ET(30) increase,
*
band, espe-
*
similarly to the absorption band. The fluorescence quantum yield of
1a varies considerably with solvent, showing the highest value in
polar aprotic DMSO (0.197) and the lowest one in polar protic water
(0.046).
35
40
45
50
ET(30)
55
60
65
For derivative 1b bearing the triazole electron-deficient system,
the emission maxima are blue shifted in respect to 1a (Table 1,
Figure 5. Correlation between the N
*
/T intensity ratio of 1a and the ET(30) polarity
*
index of the different solvents used in this study (see Table 1). The line represents the
linear fit to the experimental data.
Fig. 4B). The N
for 1a, suggesting a faster ESIPT reaction that results in a pre-
dominant emission of the T form. This observation is in line with
the fast ESIPT previously reported for 3-HC dyes bearing 2-aryl
group with low electron donor ability.20 Remarkably, the N
/T ratio
*
/T
*
band ratio in all the solvents is much lower than
properties and sensitivity to solvent polarity. The strong variation
*
of the N
*
/T ratio of 1a, especially in polar solvents makes it at-
tractive for applications in DNA research, where the local envi-
ronment is expected to be relatively polar.
*
*
*
is very low in buffer (0.06), probably due to specific solvation of the
triazole ring. Moreover, compound 1b exhibits a relatively low
fluorescence quantum yield in most solvents, except in dichloro-
methane and acetonitrile (Table 1).
3. Conclusion
From the comparison of these two nucleosides, it is clear that 1a
is much more advantageous than 1b in terms of spectroscopic
In conclusion, we developed an efficient synthesis of un-
precedented
environment-sensitive
ratiometric
fluorescent

M. Spadafora et al. / Tetrahedron 65 (2009) 7809–7816
7813
nucleosides bearing 2-aryl-3-HC nucleobases. The synthesis of 1a
involved as key steps an aryl–aldol condensation followed by regio-
controlled C40–C1’ cycloetherification or Friedel–Crafts type glyco-
sylation. In the case of 1b we used the azide-alkyne 1,3-dipolar
cycloaddition, conveniently achieved under the cooperative effects of
microwave activation and Cu(I)/SiO2 catalysis. Fluorescence studies
showed that 1a and to a lesser extent 1b retain the dual emission
highly sensitive to the environment of the 3-HC fluorophore. Thus,
HCl (4 mL). The resulting mixture was refluxed for 1 h. After cool-
ing, the mixture was extracted with methylene chloride (3ꢂ40 mL).
The combined organic layers were washed with saturated NaHCO3
solution, then with brine, dried over MgSO4, filtered and concen-
trated to afford chromone 6 (red crystals, 700 mg, 90%). Mp
(ether)¼52–54 ꢁC (lit. 52 ꢁC, Ref. 1). Rf¼0.45 (cyclohexane/ethyl
acetate: 60:40). 1H NMR (CDCl3, 200 MHz)
d¼6.31 (d, 1H, J¼6.0 Hz),
7.33–7.44 (m, 2H), 7.63 (ddd, 1H, J¼1.8, 7.1 and 8.4 Hz), 7.83 (d, 1H,
a decrease of solvent polarity induces a significant change of the N
/T
* *
J¼6.0 Hz), 8.18 (dd, 1H, J¼1.5 and 7.9 Hz). 13C NMR (CDCl3, 50 MHz)
ratio in 1a, together with red and blue shifts of the T
*
and N
*
bands,
d
¼113.1, 118.3, 125.0, 125.3, 125.9, 133.8, 155.4, 156.6, 177.7. MS (ESI,
respectively.
MeOH) m/z: 168.9 [MþNa]þ.
Due to the large polarity differences between the interior and the
surface of the DNA helix, incorporation of 1a in DNA should allow
monitoring the microenvironmental changes and DNA dynamics at
4.2.3. 2,3-Epoxy-chromone (intermediate)
To a solution of chromone 6 (740 mg, 5 mmol) in methylene
chloride (7.5 mL), was slowly added hydrogen peroxide (2 equiv,
35% solution, 1.4 mL) and NaOH (1.5 equiv, 407 mg) at 0 ꢁC. The
mixture was stirred 3 h at 0 ꢁC, quenched with water and extracted
with methylene chloride (3ꢂ20 mL). The combined organic layers
were dried (Na2SO4) and evaporated under reduced pressure to
give a crude amorphous solid (668 mg, 82%), which was used in the
next step without purification. Mp (ether)¼64–66 ꢁC. Rf¼0.8 (cy-
the probe site. Moreover, the N
*
and T bands of 1a at 440 and
*
515 nm, respectively, are well separated from each other and from
the absorption/emission of the natural nucleobases and amino
acids. Therefore, 1a can be selectively excited and studied even in
the presence of other nucleic acids and proteins. Incorporation of
such ratiometric nucleosides into synthetic oligonucleotides is un-
der way to explore their applications as nucleic acid labels.
clohexane/ethyl acetate: 60:40). 1H NMR (CDCl3, 200 MHz)
d¼3.69
4. Experimental section
4.1. General
(d, 1H, J¼2.4 Hz), 5.66 (d, 1H, J¼2.5 Hz), 7.05 (dd, 1H, J¼0.7 and
8.5 Hz), 7.14 (td, 1H, J¼1.0 and 8.0 Hz), 7.55 (ddd, 1H, J¼1.8, 7.2 and
8.5 Hz), 7.85 (dd, 1H, J¼1.8 and 8 Hz). 13C NMR (CDCl3, 50 MHz)
d
¼55.4, 77.3, 118.1, 119.9, 123.4, 127.2, 136.4, 155.5, 188.2. MS (ESI,
All reactions were run under nitrogen atmosphere in dried
glassware. Solvents were dried and distilled by standard pro-
cedures. Toluene was purchased from commercial sources and
dried over 4 Å molecular sieves before use. Reagents were pur-
chased and used without further purification. All reactions were
monitored by thin layer chromatography (TLC) plates (0.2 mm,
silica gel 60 with fluorescent indicator UV254). Flash chromatogra-
phy was performed using silica gel (60, 0.040–0.063 mm). 1H NMR
and 13C NMR were recorded on 200 and 500 instruments (200 and
MeOH) m/z: 160.7 [MꢀH]ꢀ.
4.2.4. 3-Hydroxy-chromone 7
To the epoxide previously obtained (570 mg, 3.5 mmol) was
added concd HCl (20 mL) and the resulting mixture was heated at
70 ꢁC for 1 h. After cooling, water was added (20 mL) and the
mixture was extracted with methylene chloride (3ꢂ30 mL). The
combined organic layers were washed with saturated aqueous
NaHCO3 solution, dried over MgSO4, filtered and concentrated to
afford 3-hydroxychromone 7 (brown powder, 568 mg, 100%). Mp
(ether)¼179–181 ꢁC. Rf¼0.5 (cyclohexane/ethyl acetate: 50:50). 1H
500 MHz for 1H, 50 and 125 MHz for 13C). Chemical shifts (
d) were
reported in parts per million and the coupling constants were
reported in hertz (Hz). Analytic High Performance Liquid Chroma-
NMR (CDCl3, 200 MHz)
d
¼6.33 (s, 1H), 7.41 (ddd, 1H, J¼1.1, 7.0 and
tography (HPLC) was recorded using a RP-C18 column (300A, 5
m
m
8.0 Hz), 7.50 (dd, 1H, J¼0.6 and 8.6 Hz), 7.69 (ddd, 1H, J¼1.8 and 7.0
particle size). Absorption and fluorescence spectra were recorded
on Cary 4 spectrophotometer (Varian) and FluoroMax 3.0 spec-
trofluorometer (Jobin Yvon, Horiba), respectively. Fluorescence
quantum yields were determined by taking quinine sulfate in 0.5 M
sulfuric acid (quantum yield, QY ¼ 0.577)21 as a reference.
and 8.6 Hz), 8.01 (s, 1H), 8.26 (dd, 1H, J¼1.3 and 8 Hz). 13C NMR
(CDCl3, 50 MHz)
d
¼117.3, 123.4, 124.3, 132.3, 137.2, 140.5, 147.9,
155.1, 172.2. MS (ESI, MeOH) m/z: 184.9 [MþNa]þ. HRMS (ESI) calcd
for C9H7O3 [MþH]þ, 163.0395; found, 163.0389.
4.2.5. 2-Bromo-3-hydroxy-chromone (intermediate)
4.2. Synthesis of the ethynyl-3-hydroxy-chromone 10 and 11
To a solution of 3-HC 7 (540 mg, 3.34 mmol, 0.65 equiv) in
acetonitrile (10 mL) was added NBS (1 equiv, 940 mg) and 2,20-
azobis(2-methylpropionitrile) (vazoÒ 67, 0.1 equiv, 100 mg), and
the mixture was refluxed for 8 h (0.1 equiv of vazoÒ 67 was added
each 2 h). The mixture was cooled and the solvent evaporated in
vacuo. The crude product was purified on silica gel (cyclohexane/
ethyl acetate: 90:10) to give the corresponding 2-Br-3-HC de-
rivative (colourless powder, 730 mg, 90%). Mp (ether)¼178–180 ꢁC.
Rf¼0.57 (cyclohexane/ethyl acetate: 50:50). 1H NMR (MeOD,
4.2.1. (E)-3-(Dimethylamino)-1-(2-hydroxyphenyl)-prop-2-en-1-
one (enamide intermediate)
A mixture of 20-hydroxyacetophenone (5 mmol, 0.6 mL) and
N,N-dimethylformamide-dimethylacetal (1 equiv, 0.66 mL) was ir-
radiated under microwave for 15 s (300 W max, T¼115 ꢁC). The
resulting mixture was cooled at room temperature and crystallized
in pentane to give the enamine intermediate (red crystals, 955 mg,
100%). Mp (ether)¼130–131 ꢁC (lit. 132–134, Ref. 1). Rf¼0.2 (cyclo-
200 MHz)
and 8.6 Hz), 7.66 (ddd, 1H, J¼1.6, 7.0 and 8.6 Hz), 8.06 (dd, 1H, J¼1.7
and 8.0 Hz). 13C NMR (CDCl3, 50 MHz)
d
¼7.37 (ddd, 1H, J¼1.0, 7.1 and 8.1 Hz), 7.47 (dd, 1H, J¼0.6
hexane/ethyl acetate: 60:40). 1H NMR (CDCl3, 200 MHz)
d
¼2.95 (s,
3H, NMe), 3.17 (s, 3H, NMe), 5.76 (d, 1H, J¼12.1 Hz), 6.81 (dt, 1H,
J¼1.0 and 7.1 Hz), 6.92 (dd, 1H, J¼0.9 and 8.4 Hz), 7.30 (dt, 1H, J¼1.5
and 8.4 Hz), 7.69 (dd, 1H, J¼1.4 and 8.0 Hz), 7.87 (d, 1H, J¼12.0 Hz),
d¼118.1, 121.0, 125.4, 126.0,
129.8, 134.0, 140.3, 156.7, 171.3. MS (ESI, MeOH) m/z: 238.9–240.9
[MꢀH]ꢀ. HRMS (ESI) calcd for C9H4BrO3 [MꢀH]ꢀ, 238.9349; found,
238.9353.
13.99 (s, 1H, OH). 13C NMR (CDCl3, 50 MHz)
118.3, 120.4, 128.3, 134.0, 154.9, 163.0, 191.6. MS (ESI, MeOH) m/z:
213.70 [MþNa]þ.
d
¼37.5, 45.5, 90.1, 118.1,
4.2.6. 2-Bromo-3-benzyloxy-chromone 8
To a solution of 2-bromo-3-HC previously obtained (222 mg,
0.92 mmol) in DMF (3 mL) was added K2CO3 (2 equiv, 254 mg) and
benzylbromide (2 equiv, 0.22 mL). The mixture was stirred over-
night under N2 atmosphere, then quenched by addition of water
4.2.2. Chromone 6
To
a solution of the enamide previously obtained (1 g,
5.24 mmol) in methylene chloride (40 mL) was added concentrated

7814
M. Spadafora et al. / Tetrahedron 65 (2009) 7809–7816
and extracted with methylene chloride. The organic layer was dried
(MgSO4), concentrated and the residue was purified by silica gel
column chromatography using 10% of ethyl acetate in cyclohexane
to afford compound 8 (yellow oil, 275 mg, 90%). Rf¼0.37 (cyclo-
completion of the reaction (TLC monitoring), the mixture was
quenched with saturated aq NH4Cl solution, extracted with meth-
ylene chloride, dried over MgSO4, filtered and concentrated in
vacuo. The obtained residue was purified by flash chromatography
on silica gel (eluting with cyclohexane/ethyl acetate: 90:10) to give
compound 12 (yellow oil, R/S¼40:60, 75%). Rf¼0.53 (cyclohexane/
ethyl acetate: 70:30). Major isomer: 1H NMR (CDCl3, 200 MHz)
hexane/ethyl acetate: 90:10). 1H NMR (CDCl3, 200 MHz)
d¼5.26 (s,
2H, CH2), 7.34–7.55 (m, 7H), 7.65 (ddd, 1H, J¼1.8, 7.1 and 8.6 Hz),
8.25 (dd, 1H, J¼1.8 and 8.0 Hz). 13C NMR (CDCl3, 50 MHz)
¼74.6,
d
117.8,124.1,125.7,126.4, 128.5,129.1, 133.9,136.4,142.4,156.2,172.9.
MS (ESI, MeOH) m/z: 352.8–354.8 [MþNa]þ. HRMS (ESI) calcd for
C16H12O3Br [MþH]þ, 330.9969; found, 330.9975.
d
¼1.08 (s, 9H, t-Bu), 1.35 (s, 3H, CH3), 1.41 (s, 3H, CH3), 2.14–2.20 (m,
2H, H-20), 3.41 (br d, 1H, J¼6.3 Hz, OH), 3.70–3.74 (m, 2H, 2H-50),
4.26 (dd, 1H, J¼5.9 and 12.8 Hz, H-40), 4.53 (dd, 1H, J¼6.0 and
13.1 Hz, H-30), 5.12 (dd, 1H, J¼5.5 and 11.1 Hz, H-10), 6.68 (dd, J¼0.9
and 3.7 Hz, H-thiophene), 6.91–6.93 (d, J¼3.7 Hz, H-thiophene),
7.39–7.43 (m, 6H), 7.63–7.69 (m, 4H). 13C NMR (CDCl3, 50 MHz)
4.2.7. 2-Bromo-3-(2-methoxyethoxy)methoxy-chromone 9
To a solution of 2-bromo-3-HC previously obtained (566 mg,
2.33 mmol) in DMF (6 mL) was successively added K2CO3 (2 equiv,
644 mg) and MEMCl (2 equiv, 0.53 mL). The mixture was stirred
overnight under N2 atmosphere, then quenched by addition of
water and extracted three times with methylene chloride. The or-
ganic layer was dried (MgSO4), concentrated and the residue was
purified by silica gel column chromatography using 30% of ethyl
acetate in cyclohexane to give compound 9 (white resin, 725 mg,
95%). Rf¼0.47 (cyclohexane/ethyl acetate: 50:50). 1H NMR (CDCl3,
d
¼19.3, 25.6, 27.0, 28.1, 29.8, 30.3, 37.6, 39.0, 62.4, 68.1, 70.4, 74.2,
76.8, 77.5, 108.4, 111.1, 123.3, 127.9, 129.6, 130.0, 133.0, 133.1, 135.6,
135.7, 150.8. MS (ESI, MeOH) m/z: 596.7–598.7 [MþNa]þ.
4.3.2.
a
and
b
-1-(5-Bromothiophen-2-yl)-3-hydroxy-5-O-tert-
-ribofuranose 13
butyldiphenylsilyl-2-deoxy-
D
To a solution of 12 (R/S¼40:60) (1.51 mmol) in toluene (40 mL)
was added p-toluenesulfonic acid (0.2 equiv, 0.3 mmol). The mix-
ture was stirred at 50 ꢁC for 1 h then quenched with a saturated
solution of sodium hydrogencarbonate and extracted with methy-
lene chloride. The combined organic layers were dried (MgSO4) and
evaporated under reduced pressure to give a crude oil. Silica gel
column chromatography purification (cyclohexane/ethyl acetate:
200 MHz)
d
¼3.37 (s, 3H, CH3), 3.57–3.62 (m, 2H, CH2), 4.00–4.06
(m, 2H, CH2), 5.39 (s, 2H, CH2), 7.38–7.49 (m, 2H), 7.68 (ddd, 1H,
J¼1.8, 7.1 and 8.6 Hz), 8.20 (dd, 1H, J¼1.5 and 8.0 Hz). 13C NMR
(CDCl3, 50 MHz)
d¼59.2, 69.5, 71.8, 96.5, 117.7, 124.0, 125.7, 126.4,
134.0, 142.2, 172.6. MS (ESI, MeOH) m/z: 351–353 [MþNa]þ. HRMS
(ESI) calcd for C13H14O5Br [MþH]þ, 329.0024; found, 329.0020.
90:10 to 50:50) afforded 13 as a yellow oil (740 mg, 85%,
a/
b
¼40:60). Rf¼0.35 (cyclohexane/ethyl acetate: 70:30). 1H NMR
4.2.8. 3-Benzyloxy-2-(trimethysilyl-ethynyl)-chromone 10
(CDCl3, 200 MHz)
d
¼1.02 (s, 9H, t-Bu), 2.00–2.32 (m, 2H, 2H-20),
Note that the cleavage of TMS group of pure compounds 10 and
11 was observed. In addition, the corresponding terminal alkynes of
10 and 11 were unstable and underwent slow degradation in so-
lution. Therefore, these compounds were stored at low tempera-
ture under N2 atmosphere and deprotected and used in situ in the
next coupling step.
To a solution of compound 8 (245 mg, 0.74 mmol), trime-
thylsilyl-acetylene (0.615 mL, 6 equiv), TEA (2 mL, 13.86 mmol,
20 equiv) in toluene (12 mL) was added PdCl2(PPh3)2 (52 mg,
0.1 equiv), CuI (26 mg, 0.2 equiv) under N2 atmosphere. The mix-
ture was stirred at 120 ꢁC for 1 h, filtered through Celite and
washed several times with ethyl acetate. The filtrate was evapo-
rated and the obtained residue was purified by flash chromatog-
raphy on silica gel, using 5% of ethyl acetate in cyclohexane, to
afford 10 as yellow oil (235 mg, 92%). Compound 11 was prepared in
similar manner. Analysis of compound 10 and the corresponding
terminal alkyne (partial TMS-cleavage in solution). Rf¼0.26 (cy-
clohexane/ethyl acetate: 90:10). 1H NMR of a mixture of 10 and its
3.63–4.08 (m, 3H, 2H-50and H-40), 4.53–4.60 (m, 1H, H-30), 5.26–
5.34 (m, 1H, H-10), 6.72 (dd, J¼0.8 and 3.7 Hz, H-thiophene), 6.87 (d,
J¼3.7 Hz, H-thiophene), 7.35–4.10 (m, 6H), 7.67–7.80 (m, 4H). 13C
NMR (CDCl3, 50 MHz)
d¼19.4, 27.0, 43.0, 43.7, 64.7, 64.9, 74.4, 74.6,
76.2, 85.6, 87.4, 111.7 (C–Br), 124.5, 124.7, 127.9, 127.9, 127.9, 129.4,
129.6, 129.9, 130.0, 133.2, 134.9, 135.6, 135.7, 135.8, 147.1. MS (ESI,
MeOH) m/z: 539.2–541.2 [MþNa]þ. HRMS (ESI) calcd for
C25H28BrO3SSi [MꢀH]ꢀ, 515.0717; found, 515.0723.
Path B. Friedel–Crafts glycosylation.
4.3.3.
b-1-(5-Bromothiophen-2-yl)-3,5-di-O-acetyl-2-deoxy-D-
ribofuranose 14
b
To a stirred solution of acetyl-deoxyribose 3a or 3b (8.6 mmol)
in CH2Cl2 (50 mL) and 2-bromothiophene (2 equiv) was added
dropwise SnCl4 (1 equiv) at 0 ꢁC. The mixture was stirred 30 min,
then quenched with saturated aqueous NaHCO3 solution and
extracted with methylene chloride (3ꢂ50 mL). The combined or-
ganic layers were dried over MgSO4, concentrated and the crude
product was purified on silica gel chromatography (cyclohexane/
acetylenic derivative (CDCl3, 200 MHz)
d
¼0.31 (s, 5H, TMS), 3.79 (s,
0.3H, H-alkyne), 5.32 and 5.34 (2s, 2H, CH2), 7.32–7.54 (m, 7H),
7.61–7.70 (m, 1H), 8.20 (dd, 1H, J¼1.0 and 8.0 Hz). 13C NMR (CDCl3,
ethyl acetate: 95:5 to 80:20) to give 14 (2.78 g,
combined yield). Compound 14-
etate: 50:50). 1H NMR (CDCl3, 200 MHz)
a
/
b
¼30:70, 73%
b
: Rf¼0.55 (cyclohexane/ethyl ac-
50 MHz)
d
¼0.0, 74.8, 89.8, 93.9, 109.8, 118.1, 124.7, 124.9, 125.1,
d
¼2.09 (s, 3H, CH3), 2.10 (s,
125.9, 126.0, 128.3, 128.4, 128.5, 128.8, 129.0, 130.6, 130.8, 132.1,
132.3, 133.8, 134.0, 134.3, 134.6, 136.6, 136.9, 155.6, 174.3. MS (ESI,
MeOH) m/z: 370.8 [MþNa]þ.
0
3H, CH3), 2.17 (ddd, 1H, J¼13.8, 10.7 and 6.0 Hz, H2 ), 2.36 (dd, 1H,
0
0
0
J¼13.8, and 5.8 Hz, H2 ), 4.18 (m, 2H, H4 and H5 ), 4.32 (dd, 1H,
0
0
J¼13.2 and 5.2 Hz, H5 ), 5.22 (d, J¼6.0 Hz, 1H, H3 ), 5.26 (dd, 1H,
0
J¼10.7 and 5.2 Hz, H1 ), 6.75 (dd, 1H, J¼3.7 and 0.6 Hz, H-thio-
4.3. Synthesis of the thienyl 3-hydroxychromone
nucleoside 1a
phene), 6.89 (d, 1H, J¼3.7 Hz, H-thiophene). 13C NMR (CDCl3,
50 MHz)
d
¼21.0, 21.2, 41.5, 64.3, 76.5, 76.7, 82.8, 112.3, 125.3, 129.5,
145.7, 170.6, 170.8. MS (ESI, MeOH) m/z: 384.9–386.9 [MþNa]þ,
400.8–402.8 [MþK]þ. HRMS (ESI) calcd for C13H14BrO5S [MꢀH]ꢀ,
360.9750; found, 360.9740.
Path A. Aryl–aldol condensation.
4.3.1. 1-(5-Bromothiophen-2-yl)-2-((4R,5R)-5-((tert-
butyldiphenylsilyloxy)methyl)-2,2-dimethyl-1,3-dioxolan-4-
yl)ethanol 12
4.3.4.
b
-1-(5-Bromothiophen-2-yl)-2-deoxy-
D
-ribofuranose (diol
intermediate)
To a solution of 2-bromothiophene (4 mmol), in THF (30 mL)
was slowly added LDA (6 mmol) at ꢀ78 ꢁC. After 45 min, the al-
dehyde 2 was added dropwise (1.5 mmol in 2 mL of THF). After
To a solution of 14-
added K2CO3 (3 equiv, 953 mg) and the mixture was stirred at room
b (835 mg, 2.3 mmol) in MeOH (12 mL) was

M. Spadafora et al. / Tetrahedron 65 (2009) 7809–7816
7815
temperature for 1 h. The solution was evaporated in vacuo and the
residue was purified by flash chromatography (methylene chloride/
methanol: 90:10) to give the diol as a foam (630 mg, 98%). The
spectral data of this product are in accordance with those recently
reported by M. Hocek et al. (Ref. 2) (see 1H and 13C spectral data).
HRMS (ESI) calcd for C9H10BrO3S [MꢀH]ꢀ, 276.9539; found,
276.9528.
4.3.8.
ribofuranose 1a
b-1-(5-(3-Hydroxy-chromone-2)-thiophen-2-yl)-2-deoxy-D-
To a solution of MEM-protected nucleoside 16 previously
obtained (140 mg, 0.15 mmol) in dioxane (3 mL) was added HCl 6 N
(1.5 mL). The mixture was stirred at room temperature overnight
then neutralized by saturated aqueous NaHCO3 solution and
extracted with ethyl acetate (3ꢂ10 mL). The organic layers were
evaporated in vacuo and the residue was purified by silica gel col-
umn chromatography (methylene chloride/methanol: 95:5 to
90:10) to give the free 3-HC-nucleoside 1a as a pale resin (46 mg,
85%). Rf¼0.26 (9:1 CH2Cl2/MeOH). 1H NMR (DMSO-d6, 500 MHz)
0
4.3.5.
b-1-(5-Bromothiophen-2-yl)-3,5-di-O-tert-
butyldiphenylsilyl-2-deoxy-
D-ribofuranose 15
To a solution of diol previously obtained (610 mg, 2.19 mmol) in
dry DMF (11 mL) were successively added imidazole (3.5 equiv) and
TBDPSCl (3.5 equiv,1.97 mL) under N2 atmosphere. After stirring for
24 h, the reaction mixture was quenched with a saturated solution
of NH4Cl and then extracted three times with methylene chloride.
The combined organic layers were dried over MgSO4, filtered and
evaporated. The residue was purified by silica gel column (cyclo-
hexane/ethyl acetate: 90:10) to afford 15 as a colourless oil (1.57 g,
95%). Rf¼0.8 (cyclohexane/ethyl acetate: 80:20). 1H NMR (CDCl3,
d
¼1.85 (ddd, 1H, J¼2.1, 11.3 and 13.7 Hz, H2 ), 2.12 (ddd, 1H, J¼2.1
0
0
and 3.8 and 13.7 Hz, H2 ), 3.30–3.62 (m, 1H, H5 ), 3.65–3.67 (m, 2H,
0
0
0
H5 and H4 ), 3.99 (br m, 1H, H3 ), 4.97 (dd, 1H, J¼2.1 Hz and 11.3 Hz,
0
H1 ), 7.15 (d, 1H, J¼3.9 Hz, H-thiophene), 7.47 (t, 1H, J¼7.4 Hz, H-6),
7.71 (d, 1H, J¼8.3 Hz, H-8), 7.81 (m, 1H, H-7), 7.82 (d, 1H, J¼3.9 Hz,
H-thiophene), 8.11 (dd, 1H, J¼1.2 and 7.4 Hz, H-5). 13C NMR (DMSO-
d6, 125 MHz)
d
¼42.4, 67.8, 68.2, 68.7, 71.0, 120.0, 123.7, 126.0, 126.5,
126.7, 129.9, 132.8, 135.5, 138.5, 145.2, 152.8, 156.1, 173.9. MS (ESI,
200 MHz)
d
¼0.85 (s, 9H, t-Bu), 1.01 (s, 9H, t-Bu), 1.81 (ddd, 1H,
MeOH) m/z: 358.9 [MꢀH]ꢀ. HRMS (ESI) calcd for C18H17O6S
J¼12.6, 10.9 and 5.2 Hz, H2’), 1.81 (dd, 1H, J¼11.4 and 5.1 Hz, H2 ),
[MþH]þ, 361.0745; found, 361.0741. IR (KBr)
n .
: 3432, 1651 cmꢀ1
0
0
3.24 (dd, 1H, J¼11.0 and 4.0 Hz, H5 ), 3.40 (dd, 1H, J¼11.0 and 4.0 Hz,
0
0
0
H5 ), 4.00 (dt,1H, J¼3.7 and 1.1 Hz, H4 ), 4.32 (br d,1H, J¼5.0 Hz, H3 ),
4.4. Synthesis of the triazolyl-3-hydroxy-chromone 1b
0
4.96 (dd, 1H, J¼10.9 and 5.1 Hz, 1H, H1 ), 6.64 (dd, 1H, J¼3.7 and
0.6 Hz, H-thiophene), 6.77 (d, 1H, J¼3.7, H-thiophene), 7.115–7.65
4.4.1.
tolouyl-2-deoxy-D
b
-1-(4-(3-Benzyloxy-chromone-2)-triazol-1-yl)-3,5-di-O-
-ribofuranose 17
(m, 20H, Ar). 13C NMR (CDCl3, 50 MHz)
d
¼19.2, 26.9, 27.1, 44.7, 64.3,
75.6, 88.5, 127.7, 127.7, 127.8, 127.9, 129.3, 129.8, 130.0, 133.2, 133.7,
133.8, 135.9, 147.2. MS (ESI, MeOH) m/z: 777.4–779.4 [MþNa]þ.
HRMS (ESI) calcd for C41H46BrO3SSi2 [MꢀH]ꢀ, 753.1895; found,
753.1884.
Method A. To a stirred solution of azido-sugar 4 (1 mmol), alkyne
10 (1.1 equiv) and n-Bu4NF (1.1 equiv) in methylene chloride (5 mL)
were successively added CuI (164 mg, 2 equiv), DIEA (0.37 mL,
5 equiv) and acetic acid (1 equiv). The mixture was stirred 4 h at
room temperature, filtered and the solvent removed in vacuo. The
crude product was purified by flash chromatography (cyclohexane/
ethyl acetate: 80:20) to give 17 (598 mg, 89%).
4.3.6.
b-1-(5-Tributylstannylthiophen-2-yl)-3,5-di-O-tert-
butyldiphenylsilyl-2-deoxy-
D-ribofuranose (intermediate)
To a solution of 15 (2.08 mmol) in dry ether (8 mL) was added
dropwise n-BuLi (1.1 equiv, 1.6 M solution) at 0 ꢁC, under N2 at-
mosphere. After 1 h, the mixture was cooled to ꢀ78 ꢁC and tribu-
tyltin chloride (1.1 equiv) was slowly added and the mixture was
stirred overnight. The reaction mixture was quenched with satu-
rated NH4Cl solution and extracted with ether. The combined or-
ganic layers were dried over MgSO4, filtered and concentrated in
vacuo. The obtained crude product was used in the next step
without further purification.
Method B. A mixture of azido-sugar 4 (1 mmol), alkyne 10
(1.1 equiv), n-Bu4NF (1.1 equiv), CuI (164 mg, 2 equiv), DIEA
(0.37 mL, 5 equiv) was adsorbed on silica gel (1 g) using methylene
chloride. After evaporation, the resulting yellow powder was
placed into a microwave and irradiated for 2 min. The mixture was
eluted twice with ethyl acetate and the solvent evaporated under
reduced pressure to give a crude product, which was subjected to
a simple filtration over silica gel (cyclohexane/ethyl acetate: 80:20)
to give 17 (638 mg, 95%). Mp (methylene chloride/ether)¼183–
185 ꢁC. Rf¼0.56 (cyclohexane/ethyl acetate: 50:50). 1H NMR (CDCl3,
4.3.7.
b-1-(5-(3-Hydroxy-chromone-2)-thiophen-2-yl)-3,5-di-O-
200 MHz)
d¼2.25 (s, 3H, CH3), 2.44 (s, 3H, CH3), 2.82–2.92 (m, 1H,
0
0
0
tert-butyldiphenylsilyl-2-deoxy-D-ribofuranose 16
H2 ), 3.11–3.24 (m, 1H, H2 ), 4.50 (d, 2H, J¼4.4 Hz, 2H5 ), 4.63–4.67
0
To a stirred solution of 2-bromo-3-HC 9 (284 mg, 0.86 mmol)
and tin derivative (2 mmol) in toluene (10 mL) under N2 atmo-
sphere were successively added Pd(PPh3)4 (50 mg) and CuI
(16 mg). The mixture was stirred at 120 ꢁC for 2 h, filtered
through Celite and concentrated in vacuo. The crude residue was
then purified by flash chromatography on silica gel (eluting with
cyclohexane/ethyl acetate: 90:10) to afford 16 as a yellow oil
(573 mg, 72%). Rf¼0.30 (cyclohexane/ethyl acetate: 70:30). 1H
(m, 1H, H4 ), 5.21 and 5.32 (2d, 2H, J¼11.0 Hz, CH2), 5.69–5.74 (m,
0
0
1H, H3 ), 6.42 (t, 1H, J¼5.7 Hz, H1 ), 7.13 (d, 2H, J¼8.0 Hz), 7.30–7.47
(m, 8H), 7.75 (m, 2H), 7.90 (d, 2H, J¼8.2 Hz), 7.95 (d, 2H, J¼8.2 Hz),
8.30 (d, 2H, J¼8.0 Hz). 13C NMR (CDCl3, 50 MHz)
¼21.7, 21.9, 38.4,
d
63.7, 74.4, 74.6, 83.9, 89.2, 118.7, 124.5, 125.0, 125.8, 126.5, 128.7,
129.0, 129.3, 129.4, 129.8, 129.9, 133.8, 136.8, 139.2, 144.2, 144.7,
155.3, 165.9, 166.2, 174.4. MS (ESI, MeOH) m/z¼709.8 [MþK]þ.
HRMS (ESI) calcd for C39H33N3O8 [MþH]þ, 672.2345; found,
672.2341.
NMR (acetone-d6, 200 MHz)
d
¼0.87 (s, 9H, t-Bu), 1.02 (s, 9H, t-
0
0
Bu), 1.97 (m, 1H, 2H2 ), 2.29 (dd, 1H, J¼12.6 and 5.0 Hz, H2 ), 3.02
(s, 3H, CH3), 3.19 (t, 2H, J¼4.6 Hz, CH2 MEM), 3.30 (dd, 1H, J¼11.0
4.4.2.
b-1-(4-(3-Benzyloxy-chromone-2)-triazol-1-yl)-2-deoxy-D-
0
0
and 3.7 Hz, H5 ), 3.47 (dd, 1H, J¼11.1 and 3.8 Hz, H5 ), 3.62 (t, 2H,
ribofuranose (intermediate)
0
J¼4.6 Hz, CH2 MEM), 4.05 (dt, 1H, J¼3.5 and 1.3 Hz, H4 ), 4.58 (d,
By the same procedure as described above for the synthesis of
14, compound 17 (100 mg, 0.15 mmol) was deprotected using
K2CO3 in MeOH to afford the pure product (62 mg, 95%). Mp
(methylene chloride/ether)¼94–96 ꢁC. Rf¼0.24 (9:1 CH2Cl2/MeOH).
0
0
1H, J¼5.1 Hz, H3 ), 5.37 (s, 2H, OCH2O), 5.47 (dd, 1H, J¼10.4 and
0
5.0 Hz, H1 ), 7.08 (d, 1H, J¼3.9, H-thiophene), 7.15–7.70 (m, 23H,
Ar), 7.85 (d, 1H, J¼3.9 Hz, H-thiophene), 8.02 (dd, 1H, J¼7.8 and
1.4 Hz, H-chromone). 13C NMR (CDCl3, 50 MHz)
d
¼20.0, 20.1, 27.6,
1H NMR (MeOD, 200 MHz)
d
¼2.47–2.78 (m, 2H, 2H2 ), 3.60 (dd, 1H,
0
0
27.8, 46.2, 59.2, 65.5, 71.0, 72.7, 76.9, 77.7, 89.8, 97.0, 119.1, 125.3,
126.0, 126.0, 126.5, 129.0, 129.1, 129.2, 131.0, 131.1, 131.3, 131.5,
132.0, 134.2, 134.3, 134.8, 134.9, 136.4, 136.7, 136.8, 137.0, 151.7,
154.8, 158.1, 174.0. MS (ESI, MeOH) m/z: 947.4 [MþNa]þ.
J¼4.6 and 11.9 Hz, 1H5 ), 3.70 (dd, 1H, J¼3.8 and 11.9 Hz, 1H5 ), 4.03
0
(dd, 1H, J¼4.2 and 8.2 Hz, H4 ), 4.48–4.56 (dd, 1H, J¼5.5 and 10.0 Hz,
0
0
H3 ), 5.28 (s, 2H, CH2), 6.42 (t, 1H, J¼5.9 Hz, H1 ), 7.20–7.35 (m, 4H),
7.44 (td, 1H, J¼1.1 and 8.1 Hz), 7.64 (d, 1H, J¼8.4 Hz), 7.75 (ddd, 1H,

7816
M. Spadafora et al. / Tetrahedron 65 (2009) 7809–7816
Chem. B 2007, 111, 5757; (d) DeLucia, A. M.; Grindley, N. D. F.; Joyce, C. M. Bio-
J¼1.5, 7.0 and 8.4 Hz), 7.85 (d, 1H, J¼8.2 Hz), 8.15 (dd, 1H, J¼1.3 and
8.1 Hz), 8.71 (s, 1H, H-5). 13C NMR (MeOD, 50 MHz)
63.0, 72.1, 75.0, 89.9, 90.7, 119.5, 125.1, 126.2, 126.4, 127.3, 129.4,
129.7, 130.0, 130.5, 130.7, 135.4, 137.6, 139.2, 139.5, 156.5, 176.1. MS
(ESI, MeOH) m/z¼457.9 [MþNa]þ, 473.8 [MþK]þ. HRMS (ESI) calcd
for C23H22N3O6 [MþH]þ, 436.1508; found, 436.1509.
´
chemistry 2007, 46, 10790; (e) Lenz, T.; Bonnist, E. Y. M.; Pljevaljic, G.; Neely,
d
¼42.0, 56.0,
R. K.; Dryden, D. T.; Scheidig, A. J.; Jones, A. C.; Weinhold, E. J. Am. Chem.
Soc. 2007, 129, 6240; (f) Reha, D.; Hocek, M.; Hobza, P. Chem. Eur. J. 2006, 12,
3587; (g) Dupradeau, F.-Y.; Case, D. A.; Yu, C.; Jimenez, R.; Romesberg, F. E. J.
Am. Chem. Soc. 2005, 127, 15612. See also Ref. 1.
3. (a) Nordlund, T. M.; Andersson, S.; Nilsson, L.; Rigler, R.; Graslund, A.;
McLaughlin, L. W. Biochemistry 1989, 28, 9095; (b) Guest, C. R.; Hochstrasser, R.
A.; Dupuy, C. G.; Allen, D. J.; Benkovic, S. J.; Millar, D. P. Biochemistry 1991, 30,
8759.
4.4.3.
b-1-(4-(3-Hydroxy-chromone-2)-triazol-1-yl)-2-deoxy-D-
4. (a) Ben Gaied, N.; Glasser, N.; Ramalanjaona, N.; Beltz, H.; Wolff, P.; Marquet, R.;
Burger, A.; Mely, Y. Nucleic Acids Res. 2005, 33, 1031; (b) Kenfack, C.; Burger, A.;
Mely, Y. J. Phys. Chem. B 2006, 110, 26327; (c) Kenfack, C.; Piemont, E.; Ben
ribofuranose 1b
To a degassed solution of the above triazolyl-compound in-
termediate (0.3 mmol) in THF (10 mL) was added Pd/C (10% molar).
Hydrogenolysis was then conducted in a high pressure autoclave
(Parr apparatus, 3 bar) for 10 h. The catalyst was filtered and washed
with THF. The filtrate was concentrated in vacuo and the obtained
residue was purified by flash chromatography to afford 1b as a white
powder (34 mg, 90%). Rf¼0.32 (8:2 CH2Cl2/MeOH). 1H NMR (DMSO-
0
´
Gaied, N.; Burger, A.; Mely, Y. J. Phys. Chem. B 2008, 112, 9736.
5. (a) Shynkar, V. V.; Klymchenko, A. S.; Kunzelmann, C.; Duportail, G.; Muller, C.
D.; Demchenko, A. P.; Freyssinet, J. M.; Mely, Y. J. Am. Chem. Soc. 2007, 129, 2187;
(b) Shynkar, V. V.; Klymchenko, A. S.; Duportail, G.; Demchenko, A. P.; Mely, Y.
Biochim. Biophys. Acta 2005, 1712, 128; (c) Klymchenko, A. S.; Duportail, G.;
Mely, Y.; Demchenko, A. P. Proc. Natl. Acad. Sci. U.S.A. 2003, 100, 11219; (d)
Enander, K.; Choulier, L.; Olsson, A. L.; Yushchenko, D. A.; Kanmert, D.; Klym-
chenko, A. S.; Demchenko, A. P.; Mely, Y.; Altschuh, D. Bioconjugate Chem. 2008,
19, 1864.
6. Formosinho, J. S.; Arnaut, G. L. J. Photochem. Photobiol. A: Chem. 1993, 75, 21.
7. Sengupta, P. K.; Kasha, M. Chem. Phys. Lett. 1979, 68, 382.
8. (a) Chou, P. T.; Martinez, M. L.; Clements, J. H. J. Phys. Chem. 1993, 97, 2618; (b)
Klymchenko, A. S.; Demchenko, A. P. Phys. Chem. Chem. Phys. 2003, 5, 461.
9. McMorrow, D.; Kasha, M. J. Phys. Chem. 1984, 88, 2235.
d6, 500 MHz)
d
¼2.42–2.47 (m, 1H, H2 ), 2.73 (ddd, 1H, J¼5.8 and
0
0
13.5 Hz, H2 ), 3.45 (dd,1H, J¼4.8 and 11.7 Hz, H5 ), 3.55 (dd,1H, J¼4.3
0
0
and 11.7 Hz, H5 ), 3.91 (dd, 1H, J¼4.8 and 8.8 Hz, H4 ), 4.43–4.44 (m,
0
0
0
1H, H3 ), 4.87 (br s, 1H, OH5 ), 5.36 (br s, 1H, OH3 ), 6.52 (t, 1H,
0
J¼5.9 Hz, H1 ), 7.48 (ddd, 1H, J¼1.0, 7.0 and 8.0 Hz), 7.73 (d, 1H,
10. Klymchenko, A. S.; Demchenko, A. P. J. Am. Chem. Soc. 2002, 124, 12372.
11. Klymchenko, A. S.; Kenfack, C.; Duportail, G.; Me´ly, Y. J. Chem. Sci. 2007, 119, 83.
12. Klymchenko, A. S.; Shvadchak, V. V.; Yushchenko, D. A.; Jain, N.; Me
Chem. B 2008, 112, 12050.
J¼8.2 Hz), 7.81 (ddd,1H, J¼1.6, 7.0 and 8.6 Hz), 8.13 (dd,1H, J¼1.6 and
8.0 Hz), 8.81 (s, 1H, H-5), 10.00 (s, 1H, OH). 13C NMR (DMSO-d6,
´ly, Y. J. Phys.
125 MHz)
d
¼40.0, 61.5, 70.4, 88.4, 88.5, 118.4, 122.0, 124.7, 124.9,
13. Pleier, A.-K.; Glas, H.; Grosche, M.; Sirsch, P.; Thiel, W. R. Synthesis 2001, 1, 55.
14. Bernini, R.; Mincione, E.; Coratti, A.; Fabrizi, G.; Battistuzzi, G. Tetrahedron 2004,
60, 967.
15. (a) Spadafora, M.; Burger, A.; Benhida, R. Synlett 2008, 1225; (b) Peyron, C.;
Benhida, R. Synlett 2009, 472.
125.3, 133.8, 137.9, 138.0, 140.3, 154.5, 172.1. MS (ESI, MeOH) m/z:
343.7 [MꢀH]ꢀ, 368.0 [MþNa]þ. HRMS (ESI) calcd for C16H16N3O6
[MþH]þ, 346.1039; found, 346.1042. IR (KBr)
n .
: 3392, 1621 cmꢀ1
16. The Friedel–Craft C-glycosylation approach was published simultaneously by
some of us (Ref. 16a) and by M. Hocek group (Ref. 16b): (a) Spadafora, M.;
Mehiri, M.; Burger, A.; Benhida, R. Tetrahedron Lett. 2008, 49, 3967; (b) Ba´rta, J.;
Pohl, R.; Klepeta´rova´, B.; Ernsting, N. P.; Hocek, M. J. Org. Chem. 2008, 73, 3798;
(c) Hainke, S.; Arndt, S.; Seitz, O. Org. Biomol. Chem. 2005, 3, 4233; For other
contributions see: (d) Joubert, N.; Pohl, R.; Klepeta´rova´, B.; Hocek, M. J. Org.
Acknowledgements
The authors thank J-M. Guigonis IFR 50 for HRMS spectra, ANR,
`
CNRS and Ministere de l’Education Nationale et de la Recherche for
´
´
Chem. 2007, 72, 6797; (e) Hocek, M.; Pohl, R.; Klepetarova, B. J. Eur. J. Org. Chem.
2005, 4525; (f) Guianvarc’h, D.; Fourrey, J.-L.; Maurisse, R.; Sun, J.-S.; Benhida, R.
Org. Lett. 2002, 4, 4209; (g) Guianvarc’h, D.; Fourrey, J.-L.; TranHuuDau, M.-E.;
Gue´rineau, V.; Benhida, R. J. Org. Chem. 2002, 67, 3724; (h) The spectral data of
these analogues are in accordance with those reported by M. Hocek, Ref. 16b
(see Supplementary data).
financial support and for the doctoral fellowships of M.S. (MENRT)
and V.Y.P. (ANR). V.V.S. is supported by an Eiffel doctoral fellowship.
Supplementary data
17. (a) Guezguez, R.; Bougrin, K.; EL Akri, K.; Benhida, R. Tetrahedron Lett. 2006, 47,
4807; (b) EL Akri, K.; Bougrin, K.; Balzarini, J.; Faraj, A.; Benhida, R. Bioorg. Med.
Chem. Lett. 2007, 17, 6656; (c) Broggi, J.; Joubert, N.; Dı´ez-Gonza´lez, S.; Berteina-
Raboin, S.; Zevaco, T.; Nolan, S. P.; Agrofoglio, L. A. Tetrahedron 2009, 65, 4525;
(d) Broggi, J.; Kumamoto, H.; Berteina-Raboin, S.; Nolan, S. P.; Agrofoglio, L. A.
Eur. J. Org. Chem. 2009, 1880; (e) Pradere, U.; Roy, V.; McBrayer, T. M.; Schinazi,
R. F.; Agrofoglio, L. A. Tetrahedron 2008, 64, 9044.
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.tet.2009.07.021.
References and notes
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1. For recent reviews see: (a) Tor, I. Tetrahedron 2007, 63, 3415 (special issue, see
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2006, 4, 4265; (c) Rist, M. J.; Marino, J. P. Curr. Org. Chem. 2002, 6, 775.
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