Synthesis of 2′-O-α-d-ribofuranosyladenosine, monomeric unit of poly(ADP-ribose)

Article abstract of DOI:10.1016/j.tet.2008.01.028

The first chemical synthesis of 2′-O-α-d-ribofuranosyladenosine, monomeric unit of poly(ADP-ribose), has been achieved starting from 3′,5′-O-bis protected 9-(2-O-α-d-arabinofuranosyl-β-d-ribofuranosyl)-adenine. Configurational inversion of 2′-hydroxyl gro

Full text of DOI:10.1016/j.tet.2008.01.028

Available online at www.sciencedirect.com
Tetrahedron 64 (2008) 2871e2876
www.elsevier.com/locate/tet
Synthesis of 20-O-a-D-ribofuranosyladenosine, monomeric
unit of poly(ADPeribose)
a
b
b
Sergey N. Mikhailov a, , Irina V. Kulikova , Koen Nauwelaerts , Piet Herdewijn
*
a Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov strasse 32, Moscow 119991, Russia
b Laboratory of Medicinal Chemistry, Rega Institute for Medical Research, K.U. Leuven, Minderbroedersstraat 10, B-3000 Leuven, Belgium
Received 14 November 2007; received in revised form 20 December 2007; accepted 7 January 2008
Available online 11 January 2008
Abstract
The first chemical synthesis of 20-O-a-D-ribofuranosyladenosine, monomeric unit of poly(ADPeribose), has been achieved starting from
30,50-O-bis protected 9-(2-O-a-D-arabinofuranosyl-b-D-ribofuranosyl)-adenine. Configurational inversion of 20-hydroxyl group of arabinose
moiety was performed by oxidationereduction sequence.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords: Poly(ADPeribose); Monomeric unit; Stereoselective synthesis
1. Introduction
problem and the first obvious step on this way is the synthesis
of 20-O-a-D-ribofuranosyladenosine (1).
Poly-ADP ribosylation is a posttranslational modification
of proteins in eukaryotic cells catalysed by poly(ADPeribose)
polymerases (PARPs). These enzymes are responsible for the
conversion of nicotinamide adenine dinucleotide (NADþ)
into a polymer with up to 200e300 residues in length, with si-
multaneous release of nicotinamide (Scheme 1, route a). The
polymer is covalently linked to nuclear proteins through glut-
amic and aspartic acids. The importance of poly(ADPeribose)
has been established in many cellular processes such as DNA
replication, recombination, and repair and cellular differentia-
tion.1,2 However, a clear and unified picture of its physiologi-
cal role still remains to be established.
The chemical structure of poly(ADPeribose) was deter-
mined using NMR spectroscopy and enzymatic hydrolysis of
the pyrophosphate bond by phosphodiesterase with subsequent
enzymatic dephosphorylation with alkaline phosphatase
(Scheme 1, routes b and c).3e5 The development of methods
for the synthesis of poly(ADPeribose) is still a challenging
NH2
N
N
O
N
O
P
O
P
OHHO
O
O
O
N
O
N
OH OH
H2N
C
O
HO
OH
a
O
protein
O
O
O
P
OH
O
P
OH
OHHO
Ade
O
O
O
O
HO O
O
O
P
O
P
OHHO
Ade
O
O
O
O
OH
b
OH
n
Ade
Ade
O
HO OH
H2O3PO
HO
O
c
HO O
O
HO O
O
OHHO
OHHO
OPO3H2
OH
1
Scheme 1. Structure, biosynthesis of poly(ADPeribose) and its enzymatic
degradation: (a) poly(ADPeribose) polymerase; (b) phosphodiesterase; (c)
alkaline phosphatase.
* Corresponding author. Tel.: þ07 499 1359733; fax: þ07 499 1351405.
E-mail address: smikh@eimb.ru (S.N. Mikhailov).
0040-4020/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2008.01.028

2872
S.N. Mikhailov et al. / Tetrahedron 64 (2008) 2871e2876
AdeBz
AdeBz
O
O
O
O
Ade
Si
HO
2
Si
O
O
O
f
a
Si O OH
+
Si O
O
HO
HO
O
BzO
9
4
BzO
O
HO
OAc
3
BzO
BzO
O
O
HO
BzO
BzO
b
AdeBz
O
AdeBz
O
AdeBz
AdeBz
O
O
Si
O
Si
O
Si
O
Si
O
O
O
O
O
e
f
c
d
Si
O O
Si
O O
Si O O
O
Si
O O
1
8
7
HO
HO
O
O
O
O
Si
Si
6
Si
5
O
Si
O
Si
O
Si
HO
O
HO
O
HO
O
O
O
O
Scheme 2. (a) SnCl4, 0 ꢀC, ClCH2CH2Cl, N2, 20 h, 62%; (b) 0.1 N MeONa/MeOH, 10 ꢀC, 40 min, 47%; (c) ClSi(i-Pr)2OSi(i-Pr)2Cl, Py, 35 ꢀC, 20 h, 69%; (d)
DMSO, Ac2O, 20 ꢀC, 24 h; (e) NaBH4, EtOH, 0 ꢀC, 1 h, 69% for two steps (d and e); (f) NH3/MeOH, 20 ꢀC, 2 days; Bu4NF$3H2O, THF, 20 ꢀC, 1 h, 93%.
2. Results and discussion
To verify the chemical structure of compounds 1, 9, and 10
and to derive their conformational properties, several NMR
spectra were recorded and all proton and carbon resonances
Here we report the first chemical synthesis of 1. The key
step is the formation of a(1/2) O-glycosidic bond between
ribofuranose residue and adenosine. The use of D-ribose as
starting compound defines the formation of 1,2-cis-substituted
products and usually results in formation of anomeric mixture.
We decided to use another approach consisting in the forma-
tion of 1,2-trans-substituted derivatives of D-arabinose with
the following configurational inversion of 20-hydroxyl group.
O-Glycosylation of nucleosides with fully acylated pento-
furanoses was recently developed.6e8 Condensation of a small
excess of 1-O-acetyl-2,3,5-tri-O-benzoyl-D-arabinofuranose
(3) with 30,50-O-protected adenosine 2 at 0 ꢀC in 1,2-dichloro-
ethane in the presence of tin tetrachloride results in disaccha-
ride nucleoside 4 (Scheme 2). O-Glycosylation proceeded
stereospecifically with formation of a a-glycosidic bond.
The presence of a participating 2-O-benzoyl group leads ex-
clusively to 1,2-trans-ribofuranosides.9 Selective O-deben-
zoylation was achieved using MeONa in methanol.
Simultaneous protection of 30,50-O-hydroxyl groups in 5
was achieved using Markiewicz blocking group.10 The reac-
tion was rather slow at room temperature and it was carried
at 35 ꢀC. Configurational inversion of 20-hydroxyl group of
arabinose moiety was performed by Robins oxidationereduc-
tion sequence.11 The reduction of the carbonyl function with
sodium borohydride in 7 gave the inverted 20-O-a-D-ribofura-
nosyl derivative 8 as the main product by attack at the less hin-
dered side of the sugar ring. After deprotection 1 was obtained
in good overall yield. The structure of 1 was unambiguously
proved by NMR, UV, and mass spectroscopies. For compari-
son 20-O-a-D-arabinofuranosyladenosine (9) was prepared af-
ter deblocking of disaccharide nucleoside 4. 1H NMR
spectra of 1, 9 and earlier prepared 20-O-b-D-ribofuranosyl-
adenosine (10)12 are presented in Table 1.
were assigned. NMR spectra of 1 are very close to that pre-
3e5
sented earlier.
The differences (Table 1) in the adenosine
part of the disaccharide nucleosides 1, 9, and 10 are rather
small, both in chemical shifts and coupling constants. This
translates to a high equivalence in conformation of the adeno-
sine parts of which the riboses mainly adopts a Southern con-
formation (22% N). In the additional sugar moieties, however,
several differences in chemical shift and scalar coupling can
be observed. The inversion of configuration at C-2 of the ad-
ditional sugar residue results in significant change of coupling
constants. The trans arrangements of H10eH20 and H20eH30
in a-D-arabinofuranose in 6 and 9 are changed to cis configu-
rations in 8 and 1 of a-D-ribofuranose moieties. This translates
to different conformations of the sugar moieties. In compound
10 the b-ribose moiety adopts a Northern conformation, In 9
the a-arabino moiety occurs as a mixture of Northern and
Southern and in 1, the a-ribose is in a Southern conformation.
For each compound the 20e100 glycosidic linkage of the sugar
moieties, could be confirmed using a HMBC spectrum in
which a clear coupling could be observed between C100 and
H20 and between H100and C20 (Fig. 1). The a configuration
of the extra ribose moiety in 1 was confirmed by a ROESY
spectrum in which clear crosspeaks could be identified be-
tween H100 and both H200 and H300, indicating cis orientation
of these protons. The b conformation of the nucleoside sugar
moiety of 1 was confirmed by a crosspeak between H10 and
H40 in the ROESY spectrum. No ROESY crosspeaks between
H8 of adenine and any of the sugar protons could be observed.
The conformation of 1 was evaluated by ROESY spectros-
copy. The intense crosspeak between H20 and H100, the cross-
peaks between H100 and H10/H30 and the absence of further
crosspeaks between protons of the different sugar rings,

S.N. Mikhailov et al. / Tetrahedron 64 (2008) 2871e2876
2873
Table 1
1H NMR (500 MHz) chemical shifts (ppm) and coupling constants (Hz) of 20-O-a-D-ribofuranosyladenosine (1) and related compounds in D2O at 27 ꢀC
Compound 10
Moieties
Ado
Compound 9
Compound 1
b-Rib
Ado
a-Ara
Ado
a-Rib
Chemical shifts
H-10
6.01 d
4.95 s
4.02 d
6.01 d
4.85 d
6.03 d
4.98 d
H-20
4.67 dd
4.44 dd
4.19 ddd
3.82 dd
3.75 dd
8.24 s
4.68 dd
4.40 dd
4.18 ddd
3.73 dd
3.82 dd
8.23 s
3.99 dd
3.80 dd
3.99 ddd
3.64 dd
3.53 dd
4.70 dd
4.41 dd
4.19 ddd
3.80 dd
3.72 dd
7.97 s
3.97 dd
3.93 dd
4.09 ddd
3.57 dd
3.50 dd
H-30
H-40
H-50a
H-50b
H-2
3.88 dd
3.71 ddd
3.21 dd
2.64 dd
H-8
8.40 s
8.39 s
8.17 s
Coupling constants
10,20
6.4
5.2
3.2
2.7
3.6
<0.5
4.6
6.3
5.1
3.1
3.5
2.7
1.6
3.5
6.0
3.3
5.8
6.2
5.1
3.1
2.7
3.5
4.2
6.2
2.7
3.5
4.7
20,30
30,40
7.5
3.7
40,50a
40,50b
50a,50b
6.8
ꢁ12.8
22
ꢁ12.4
92
ꢁ12.9
22
ꢁ12.4
51
ꢁ12.9
22
ꢁ12.4
26
% N
Based on scalar coupling constants the percentage North conformation of the sugar residues was calculated.
Figure 1. HSQC and HMBC spectrum of 1. The HMBC spectrum shows a clear coupling between C100and H20 and between H100 and C20 (both couplings are
boxed). This confirms the correct linkage of both rings.
indicate that the disaccharide nucleoside 1, may occur as an
extended molecule.
4. Experimental
4.1. General
3. Conclusion
NMR Spectra were recorded on a Bruker AMX-400 and
a Bruker Avance II 500 NMR spectrometers at 300 K; chem-
ical shifts d are indicated in parts per million relative to the
solvent signals (1H and 13C) or H3PO4 as external standard
(31P). The coupling constants (J) are given in hertz. The
signals of 4e6 and 8 were assigned using double resonance
techniques, COSY experiments and by comparison with
related disaccharide nucleosides. The signals of 1, 9, and 10
were assigned by HSQC, HMBC, COSY, and ROESY
(tmix: 300 ms) experiments. Analysis of the couplings was
performed with Pseurot 6.2.16 This program uses the
AltonaeSundaralingam formalism17,18 to describe the ring
puckering and a generalized Karplus equation developed by
The first chemical synthesis of 20-O-a-D-ribofuranosylade-
nosine opens new possibilities in search of new potential in-
hibitors of PARPs. Activation of PARP has been implicated
in the pathogenesis of stroke, myocardial ischemia, diabetes,
diabetes-associated cardiovascular dysfunction, shock, trau-
matic central nervous system injury, arthritis, colitis, allergic
encephalomyelitis and various other forms of inflammation.
Inhibition of PARP by pharmacological agents may prove use-
ful for the therapy of these diseases, as has been shown in pre-
clinical animal models.13,14 Moreover, PARP inhibitors may
have additional, potential utility as anticancer agents, radio-
sensitizers, and antiviral agents.13e15

2874
S.N. Mikhailov et al. / Tetrahedron 64 (2008) 2871e2876
Donders et al.19 to describe the relation between coupling and
dihedral angle. rms was minimized, using a NewtoneRaphson
minimization.
12.89, 12.84 and 12.57 (i-Pr). Anal. Calcd for
C55H63N5O13Si2: C, 62.42; H, 6.00; N, 6.62. Found: C,
62.25; H, 5.80; N, 6.42.
The UV spectra were recorded on a Cary 300 UV/VIS
spectrophotometer (Varian). Mass spectrometry and exact
mass measurements of the nucleoside intermediates were per-
formed on a quadrupole/orthogonal-acceleration time-of-flight
tandem mass spectrometer (Q-Tof-2, Micromass, Manchester,
UK) equipped with a standard electrospray ionization (ESI)
interface using positive ion mode. Column chromatography
was performed on silica gel (0.063e0.200 mm, Merck). TLC
was carried out on Alugram Sil G/UV 254 (Macherey-Nagel)
using UV-detection and the following solvent systems (com-
positions expressed as v/v): methylene chloride (A), methylene
chlorideemethanol 99:1 (B), methylene chlorideemethanol
98:2 (C), methylene chlorideemethanol 97:3 (D), methylene
chlorideemethanol 95:5 (E), methylene chlorideemethanol
9:1 (F), methylene chlorideemethanol 4:1 (G) and methylene
chlorideemethanol 1:1 (H).
4.3. N6-Benzoyl-9-[3,5-(1,1,3,3-tetraisopropyldisiloxane-1,3-
diyl)-2-O-(a-D-arabinofuranosyl)-b-D-ribofuranosyl]-
adenine (5)
To a solution of nucleoside 4 (2.12 g, 2 mmol) in dioxane
(21 ml) 0.3 M sodium methylate in dry methanol (21 ml)
was added and the solution was kept for 50 min at 10 ꢀC
and then neutralized by addition of 10% acetic acid in metha-
nol to pH 7.0. The resulting solution was concentrated in va-
cuo to dryness and the residue was partitioned between ethyl
acetate (100 ml) and water (40 ml), the organic layer was
washed with water (40 ml), dried over Na2SO4, and evapo-
rated to dryness. The residue was purified on a column with
silica gel (50 g). The column was washed with system C
(300 ml) and then eluted with system E to give 5 as a foam.
Yield 724 mg (47%). Rf 0.39 (system E). LSIMS:
(C34H51N5O10Si2þH) calcd 746.3253, found 746.3253. 1H
NMR (400 MHz, CDCl3): 9.73 (br s, 1H, NH), 8.72 (s, 1H,
H-8), 8.34 (s, 1H, H-2), 7.99 (d, 2H, J¼7.5 Hz, Bz), 7.51 (t,
1H, J¼7.5 Hz, Bz), 7.43 (t, 2H, J¼7.5 Hz, Bz), 6.16 (s, 1H,
4.2. N6-Benzoyl-9-[3,5-(1,1,3,3-tetraisopropyldisiloxane-1,3-
diyl)-2-O-(2,3,5-tri-O-benzoyl-a-D-arabinofuranosyl)-b-D-
ribofuranosyl]-adenine (4)
To a cool solution (0 ꢀC) of 1-O-acetyl-2,3,5-tri-O-ben-
zoyl-b-D-arabinofuranose (1.59 g, 3.2 mmol) in 1,2-dichloro-
ethane (25 ml), tin tetrachloride (0.52 ml, 4.42 mmol) was
added under nitrogen and the solution was kept at 0 ꢀC for
10 min. After addition of nucleoside 2 (1.23 g, 2 mmol) the
resulting solution was kept at 0 ꢀC for 30 h. Aqueous solution
of sodium bicarbonate (10%, 20 ml) was added and the sus-
pension was stirred at 0 ꢀC for 20 min. The suspension was
filtered through Hyflo Super Cel, organic layer was separated,
washed with water (20 ml), dried over Na2SO4, and evapo-
rated to dryness. The residue was purified by column chroma-
tography on silica gel (70 g). The column was washed with
system A (500 ml) and then eluted with system B to give 4
as a foam. Yield 1.31 g (62%). TLC: Rf 0.49 (system D).
LSIMS: (C55H63N5O13Si2þH) calcd 1058.4039, found
1058.3985. 1H NMR (400 MHz, CDCl3): 8.96 (br s, 1H,
NH), 8.73 (s, 1H, H-8), 8.37 (s, 1H, H-2), 8.12e7.95 (m,
8H, Bz), 7.64e7.41 (m, 12H, Bz), 6.25 (s, 1H, H-10, Ado),
H-10 Ado), 5.60 (s, 1H, H-10 Ara), 4.68 (dd, 1H, J3 ,2 ¼5.0 Hz,
0
0
0
0
0
0
J3 ,4 ¼9.3, H-3 Ado), 4.62 (d, 1H, H-2 Ado), 4.18 (br d, 1H,
J5 a,5 b¼ꢁ13.7, H-50a Ado), 4.12 (m, 2H, H-20,40 Ara), 4.05 (d,
0
0
0
0
0
0
1H, J3 ,4 ¼9.3, H-3 Ara), 3.97 (m, 1H, H-4 Ado), 3.92 (dd,
0
0
0
0
0
1H, J5 b,4 ¼2.5, H-5 b Ado), 3.81 (dd, 1H, J5 a,4 ¼2.5,
J5 a,5 b¼ꢁ11.8, H-50a Ara), 3.76 (dd, 1H, J5 b,4 ¼1.9, H-5 b
Ara), 1.08e0.88 (m, 28H, i-Pr). 13C NMR (CDCl3): 165.20
(C]O), 152.59 (C-2), 150.58 (C-6), 149.52 (C-4), 140.94
(C-8), 132.67, 128.64, and 128.01 (Bz), 123.33 (C-5),
105.67 (C-10, Ara), 88.46 (C-10, Ado), 87.73 (C-40, Ara),
81.94 (C-40, Ado), 78.68 (C-20, Ado), 77.91 (C-20, Ara), 76.11
(C-30, Ara), 67.74 (C-30, Ado), 61.52 (C-50, Ara), 59.43 (C-50,
Ado), 17.36, 17.27, 17.19, 16.95, 16.84, 13.78, 13.22, 13.18
and 12.96 (i-Pr). Anal. Calcd for C34H51N5O10Si2: C, 62.42;
H, 6.00; N, 6.62. Found: C, 62.24; H, 6.07; N, 6.83.
0
0
0
0
0
4.4. N6-Benzoyl-9-[3,5-(1,1,3,3-tetraisopropyldisiloxane-1,3-
diyl)-2-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl-a-D-
arabinofuranosyl)-b-D-ribofuranosyl]-adenine (6)
5.83 (s, 1H, H-10 Ara), 5.72 (s, 1H, H-20 Ara), 5.63 (d, 1H,
0
0
0
0
0
J3 ,4 ¼5.0 Hz, H-3 , Ara), 4.87 (ddd, 1H, J4 ,5 a¼3.4 Hz,
J4 ,5 b¼4.7 Hz, H-40 Ara), 4.79 (dd, 1H, J5 a,5 b=ꢁ12.5 Hz,
To a solution of nucleoside 5 (540 mg, 0.72 mmol) in
pyridine (7 ml) 1,3-dichloro-1,1,3,3-tetraisopropyldisiloxane
(0.30 ml, 0.94 mmol) was added and the solution was kept for
24 h at 35 ꢀC, evaporated to dryness and evaporated with tolu-
ene (10 ml). The residue was partitioned between methylene
chloride (100 ml) and water (40 ml), the organic layer was
washed successively with water (40 ml), 10% aqueous solution
of sodium bicarbonate (40 ml), water (40 ml), dried over
Na2SO4, evaporated to dryness, and evaporated with toluene
(2ꢂ10 ml). The residue was purified on a column with
silica gel (30 g). The column was washed with system A
(200 ml) and then eluted with system B to give 6 as a foam.
Yield 496 mg (69%). Rf 0.54 (system C). LSIMS:
0
0
0
00
H-50a Ara), 4.72 (d, 1H, J2 ,3 ¼4.7 Hz, H-2 Ado), 4.69
0
0
(dd, 1H, H-50b Ara), 4.64 (dd, 1H, J3 ,4 ¼9.3 Hz, H-3 Ado),
0
0
0
4.36e4.23 (m, 2H, H-40,50a Ado), 4.01 (dd, 1H,
J5 b,4 ¼2.5 Hz, J5 a,5 b¼ꢁ13.4 Hz, 50b Ado), 1.13e0.85 (m,
28H, i-Pr). 13C NMR (CDCl3): 165.90, 165.77, 165.63 and
164.66 (C]O), 153.01 (C-2), 150.81 (C-6), 149.54 (C-4),
140.90 (C-8), 133.95, 133.76, 133.18, 132.83, 130.09,
129.88, 129.16, 128.83 and 128.56 (Bz), 123.69
(C-5), 104.96 (C-10, Ara), 89.16 (C-10, Ado), 82.21 (C-40,
Ara), 81.93 (C-40, Ado), 81.47 (C-20, Ara), 79.05 (C-20,
Ado), 78.16 (C-30, Ara), 67.92 (C-30, Ado), 63.63 (C-50,
Ara), 59.67 (C-50, Ado), 17.59, 17.42, 17.04, 16.86, 13.34,
0
0
0
0

S.N. Mikhailov et al. / Tetrahedron 64 (2008) 2871e2876
2875
(C46H77N5O11Si4þH) calcd 988.4775, found 988.4737. 1H
4.6. 9-(2-O-a-D-Ribofuranosyl-b-D-ribofuranosyl)-adenine (1)
NMR (400 MHz, CDCl3): 8.78 (s, 1H, H-8), 8.41 (s, 1H,
H-2), 8.06 (d, 2H, J¼7.5 Hz, Bz), 7.60 (t, 1H, J¼7.5 Hz, Bz),
A solution of nucleoside 8 (309 mg, 0.31 mmol) in 5 M am-
monia in methanol (15 ml) was kept for 2 days at 20 ꢀC and
then concentrated in vacuo to dryness. The residue was parti-
tioned between methylene chloride (60 ml) and water (20 ml),
and the organic layer was washed with water (2ꢂ20 ml), dried
over Na2SO4, and evaporated to dryness. The residue was dis-
solved in 0.5 M tetrabutylammonium fluoride trihydrate in tet-
rahydrofuran (3.8 ml), kept for 1 h at 20 ꢀC. Water (5 ml) and
Dowex-50 (Naþ form, 3 ml) were added, and the suspension
was stirred for 20 min. The resin was filtered and washed
with water. The combined filtrates were washed with methy-
lene chloride (2ꢂ5 ml), evaporated to dryness, and evaporated
with ethanol (2ꢂ5 ml). The residue was purified by column
chromatography on silica gel (25 g, dry application). The col-
umn was washed with system F (200 ml) and then eluted with
system G. The fractions containing 1 were combined and
evaporated in vacuo to dryness. The residue was dissolved in
methanol (1 ml), filtered, and evaporated in vacuo to dryness
to give 1 as a foam. Yield 117 mg (92%). TLC: Rf 0.32 (sys-
tem H). UV (pH 7e13): lmax 261 nm (3 14,300); (pH 1): lmax
258 nm (3 13,800). LSIMS: (C15H21N5O8þH) calcd 400.1468,
7.53 (t, 2H, J¼7.5 Hz, Bz), 6.18 (s, 1H, H-10 Ado), 5.34 (d,
0
0
0
0
0
1H, J1 ,2 ¼1.9 Hz, H-1 Ara), 4.58 (dd, 1H, J3 ,2 ¼4.7 Hz,
0
0
0
0
J3 ,4 ¼9.3, H-3 Ado), 4.51 (d, 1H, H-2 Ado), 4.41 (dd, 1H,
0
0
0
0
0
J2 ,3 ¼5.6 Hz, H-2 Ara), 4.29e4.18 (m, 3H, H-5 a, 5 b Ado,
H-30 Ara), 4.07e4.01 (m, 2H, H-40Ado, H-40Ara), 3.94 (dd,
1H, J5 a,4 ¼2.8, J5 a,5 b¼ꢁ12.5, H-50a Ara), 3.89 (dd, 1H,
0
0
0
0
J5 b,4 ¼4.4, H-5 b Ara), 1.13e0.95 (m, 56H, i-Pr). 13C NMR
(CDCl3): 164.69 (C]O), 152.95 (C-2), 150.69 (C-6), 149.70
(C-4), 140.97 (C-8), 132.87, 129.00, and 128.05 (Bz), 123.62
(C-5), 106.79 (C-10, Ara), 89.26 (C-10, Ado), 83.38 (C-40,
Ara), 82.01 (C-40, Ado), 81.21 (C-20, Ara), 78.94 (C-20, Ado),
78.28 (C-30, Ara), 67.82 (C-30, Ado), 61.87 (C-50, Ara),
59.93 (C-50, Ado), 17.64, 17.59, 17.56, 17.49, 17.43, 17.29,
17.27, 17.24, 17.21, 17.09, 13.65, 13.56, 13.24, 13.11,
13.07, 12.94, 12.85 and 12.79 (i-Pr). Anal. Calcd for
C46H77N5O11Si4: C, 55.89; H, 7.85; N, 7.09. Found: C, 55.78;
H, 7.81; N, 7.19.
0
0
0
4.5. N6-Benzoyl-9-[3,5-(1,1,3,3-tetraisopropyldisiloxane-1,3-
diyl)-2-O-(1,1,3,3-tetraisopropyldisiloxane-1,3-diyl-a-D-
ribofuranosyl)-b-D-ribofuranosyl]-adenine (8)
1
found 400.1459. H NMR (600 MHz, see Table 1). 13C NMR
(D2O): 154.96 (C-6), 151.99 (C-2), 147.74 (C-4), 140.11
(C-8), 118.41 (C-5), 101.49 (C-10, Rib), 86.41 (C-10, Ado),
85.68 (C-40, Ado), 85.05 (C-40, Rib), 78.71 (C-20, Ado),
71.14 (C-20, Rib), 70.43 (C-30, Ado), 69.32 (C-30, Rib),
61.04 (C-50, Rib), 57.66 (C-50, Ado). Anal. Calcd for
C15H21N5O8: C, 45.11; H, 5.30; N, 17.54. Found: C, 45.01;
H, 5.15; N, 17.34.
A solution of nucleoside 6 (471 mg, 0.48 mmol) in di-
methyl sulfoxide (3.4 ml) was treated with acetic anhydride
(0.22 ml, 2.38 mmol) and the mixture was kept at 20 ꢀC for
22 h. Ethanol (8 ml) was added, the mixture was cooled to
0 ꢀC, and treated with sodium borohydride (72 mg). The mix-
ture was stirred at 0 ꢀC for 1 h and then acetone (1 ml) and sat-
urated aqueous solution of sodium chloride (20 ml) were
added. The mixture was extracted with ethyl acetate (50 ml)
and the organic layer was washed successively with saturated
aqueous solution of sodium chloride (2ꢂ20 ml) and water
(2ꢂ20 ml), dried over Na2SO4, and evaporated to dryness.
The residue was purified on a column with silica gel (30 g).
The column was washed with system A (300 ml) and then
eluted with system B to give 8 as a foam. Yield 327 mg
(69%). Rf 0.60 (system C). LSIMS: (C46H77N5O11Si4þH) calcd
988.4775, found 988.4758. 1H NMR (400 MHz, CDCl3): 9.23
(br s, 1H, NH), 8.78 (s, 1H, H-8), 8.37 (s, 1H, H-2), 8.03 (d,
2H, J¼7.5 Hz, Bz), 7.59 (t, J¼7.5 Hz, 1H, Bz), 7.51 (t,
J¼7.5 Hz, 2H, Bz), 6.18 (s, 1H, H-10 Ado), 5.47 (d, 1H,
4.7. 9-(2-O-a-D-Arabinofuranosyl-b-D-ribofuranosyl)-
adenine (9)
A solution of nucleoside 4 (417 mg, 0.39 mmol) in 5 M am-
monia in methanol (15 ml) was kept for 2 days at 20 ꢀC and
then concentrated in vacuo to dryness. The residue was parti-
tioned between methylene chloride (60 ml) and water (20 ml),
and the organic layer was washed with water (2ꢂ20 ml), dried
over Na2SO4, and evaporated to dryness. The residue was dis-
solved in 0.5 M tetrabutylammonium fluoride trihydrate in tet-
rahydrofuran (2.4 ml) and kept for 1 h at 20 ꢀC. Water (5 ml)
and Dowex-50 (Naþ form, 3 ml) were added, and the suspen-
sion was stirred for 20 min. The resin was filtered and washed
with water. The combined filtrates were washed with methyl-
ene chloride (2ꢂ5 ml), evaporated to dryness, and evaporated
with ethanol (2ꢂ5 ml). The residue was purified by column
chromatography on silica gel (25 g, dry application). The col-
umn was washed with system F (200 ml) and then eluted with
system G. The fractions containing 9 were combined and
evaporated in vacuo to dryness. The residue was dissolved in
methanol (1 ml), filtered, and evaporated in vacuo to dryness
to give 9 as a foam. Yield 125 mg (80%). TLC: Rf 0.49 (sys-
tem H). UV (pH 7e13): lmax 261 nm (3 14,300); (pH 1): lmax
258 nm (3 13,800). LSIMS: (C15H21N5O8þH) calcd 400.1468,
0
0
0
0
0
0
J1 ,2 ¼4.4 Hz, H-1 Rib), 4.59 (dd, 1H, J2 ,3 ¼4.7 Hz, H-3
Ado), 4.56(dd, 1H, H-20 Ado), 4.28(brd, 1H, J5 a,5 b¼ꢁ13.4 Hz,
50a Ado), 4.24e4.15 (m, 5H, H-20, 30, 40 Rib, H-40, 50b Ado), 4.02
(m, 2H, H-50a, 50b Rib), 1.16e0.88 (m, 56H, i-Pr). 13C NMR
(CDCl3): 165.26 (C]O), 152.95 (C-2), 150.55 (C-6), 149.96
(C-4), 140.78 (C-8), 132.72, 128.77, and 128.26 (Bz), 114.14
(C-5), 101.92 (C-10, Rib), 89.47 (C-10, Ado), 84.10 (C-40, Rib),
81.91 (C-40, Ado), 80.52 (C-20, Ado), 71.22 (C-20, 30 Rib),
67.74 (C-30, Ado), 64.69 (C-50, Rib), 59.67 (C-50, Ado), 17.55,
17.49, 17.44, 17.39, 17.32, 17.26, 17.18, 17.11, 17.02, 16.93,
14.16, 13.83, 13.62, 13.57, 13.42, 13.22, 12.81, and 12.78
(i-Pr). Anal. Calcd for C46H77N5O11Si4: C, 55.89; H, 7.85; N,
7.09. Found: C, 55.69; H, 7.68; N, 6.83.
0
0
1
found 400.1459. H NMR (600 MHz, see Table 1). 13C NMR

2876
S.N. Mikhailov et al. / Tetrahedron 64 (2008) 2871e2876
5. Ferro, A. M.; Oppenheimer, N. J. Proc. Natl. Acad. Sci. U.S.A. 1978, 75,
809e813.
(D2O): 154.96 (C-6), 151.97 (C-2), 147.69 (C-4), 140.04
(C-8), 118.42 (C-5), 106.92 (C-10, Ara), 86.43 (C-10, Ado),
85.75 (C-40, Ado), 84.02 (C-40, Ara), 80.30 (C-20, Ara),
78.22 (C-20, Ado), 75.78 (C-30, Ara), 70.24 (C-30, Ado),
61.04 (C-50, Ado), 60.62 (C-50, Ara). Anal. Calcd for
C15H21N5O8: C, 45.11; H, 5.30; N, 17.54. Found: C, 44.93;
H, 5.18; N, 17.29.
6. Nauwelaerts, K.; Efimtseva, E. V.; Mikhailov, S. N.; Herdewijn, P.
Disaccharide Nucleosides, an Important Group of Natural Compounds.
In Frontiers in Nucleosides and Nucleic Acids; Schinazi, R. F., Liotta,
D. C., Eds.; IHL: Tucker, GA, 2004; pp 187e220.
7. Efimtseva, E. V.; Mikhailov, S. N. Russ. Chem. Rev. 2004, 73, 401e414.
8. Efimtseva, E. V.; Kulikova, I. V.; Mikhailov, S. N. Curr. Org. Chem. 2007,
11, 337e354.
9. Efimtseva, E. V.; Bobkov, G. V.; Mikhailov, S. N.; Van Aerschot, A.;
Schepers, G.; Busson, R.; Rozenski, J.; Herdewijn, P. Helv. Chim. Acta
2001, 84, 2387e2397.
Acknowledgements
10. Markiewicz, W. T. J. Chem. Res., Synop. 1979, 24e25.
11. Hansske, F.; Madej, D.; Robins, M. J. Tetrahedron 1984, 40, 125e135.
12. Mikhailov, S. N.; Efimtseva, E. V.; Gurskaya, G. V.; Zavodnik, V. E.; De
Bruyn, A.; Rozenski, J.; Herdewijn, P. J. Carbohydr. Chem. 1997, 16,
75e92.
Financial support of the Russian Foundation for Basic Re-
search, INTAS, and KUL Research Council is acknowledged.
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