A reinvestigated mechanism of ribosylation of adenine under silylating conditions

Article abstract of DOI:10.1016/j.tet.2006.08.046

The mechanism of chemical synthesis of adenosine has been reinvestigated. Depending on the reaction conditions and the presence of N⁶-protecting groups, ribosylation of adenine proceeds via different kinetic products: 3-riboadenine in strongly acidic media, 7-ribosylated derivative in the silyl method, and 1-(β-d-ribofuranosyl)adenine when applying N⁶-acyladenine and silylating conditions.

Full text of DOI:10.1016/j.tet.2006.08.046

Tetrahedron 62 (2006) 10123–10129
A reinvestigated mechanism of ribosylation of adenine under
silylating conditions
Grzegorz Framski,a Zofia Gdaniec,a Maria Gdaniecb and Jerzy Boryskia,
*
aInstitute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, PL-61704 Poznan, Poland
bFaculty of Chemistry, Adam Mickiewicz University, Grunwaldzka 6, PL-60780 Poznan, Poland
Received 19 May 2006; revised 1 August 2006; accepted 11 August 2006
Available online 6 September 2006
Abstract—The mechanism of chemical synthesis of adenosine has been reinvestigated. Depending on the reaction conditions and the
presence of N6-protecting groups, ribosylation of adenine proceeds via different kinetic products: 3-riboadenine in strongly acidic media,
7-ribosylated derivative in the silyl method, and 1-(b-D-ribofuranosyl)adenine when applying N6-acyladenine and silylating conditions.
Ó 2006 Elsevier Ltd. All rights reserved.
1. Introduction
structures have been fully confirmed.2,3 In line with that ob-
servation, the following sequence of events has been estab-
lished: (i) initial ribosylation at N3, (ii) second ribosylation
at N9 with the formation of 3,9-bis-ribosyladenine, and (iii)
its decomposition to the stable 9-regioisomer (2). This mech-
anism has been extended for glycosylation reactions of all
purine bases and may be found in every handbook on nucleo-
side chemistry. More recently, however, it has been shown
that only N7 and N9 atoms can serve as glycosyl donors or
acceptors in glycosylation and transglycosylation reactions
in the guanine series.4,5
It has been known for a century that ribosylation of adenine
(1) in the presence of acidic catalysts gives 9-(b-D-ribo-
furanosyl)adenine, i.e., adenosine (2), one of the basic
components of ribonucleic acids (Scheme 1). However, the
literature data on the mechanism of ribosylation and the
structure of a kinetically controlled product of ribosylation
seem to be rather confusing. The first proposed mechanism1
postulates an initial formation of 3-(b-D-ribofuranosyl)ade-
nine (isoadenosine; 3), and in fact, compounds of the type
3 have been isolated from reaction mixtures and their
On the other hand, there are some literature reports on isola-
tion of protected derivatives of 7-(b-D-ribofuranosyl)adenine
(4) as kinetic products in the ribosylation of adenine.6–10 The
first synthesis of 7-riboadenine was presented in 1971: a
direct coupling of bis(trimethylsilyl)-N6-benzoyladenine and
1-bromo-2,3,5-tri-O-benzoyl-b-D-ribofuranose in the pres-
ence of HgBr2 reportedly gave the 7-isomer, in addition to
the predominant amount of N6-benzoyladenosine.6 Quite
similar results were obtained when SnCl4 was used as a cata-
lyst.7,8 More recently, it has been reported that glycosylation
of persilylated N6-benzoyladenine with 1-O-acetyl-2,3,5-tri-
O-benzoyl-b-L-ribofuranose in the presence of trimethylsilyl
triflate (TMSOTf) leads to a mixture of 7- and 9-regio-
isomers.9 Interestingly, 7-riboadenine (4) can be formed
not only under the Vorbruggen’s conditions10 of ribosylation:
€
compound 4 has been obtained in the fusion reaction of
N6-benzyladenine and 1-O-acetyl-2,3,5-tri-O-benzoyl-b-D-
ribofuranose performed in nitrophenols, and the kinetic
nature of the 7-regiosiomer has been demonstrated for the
first time.11 On the basis of those literature reports, we may
assume that there is an alternative mechanism of ribosylation
of adenine, different than the generally accepted 3/9
Scheme 1.
Keywords: Adenosine; 1-(b-D-Ribofuranosyl)adenine; 7-(b-D-Ribofurano-
syl)adenine; Ribosylation; Transglycosylation; Regioselectivity.
* Corresponding author. Tel.: +48 61 852 8503; fax: +48 61 852 0532;
e-mail: jboryski@ibch.poznan.pl
0040–4020/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2006.08.046

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G. Framski et al. / Tetrahedron 62 (2006) 10123–10129
pathway of glycosylation. In this case, the kinetically con-
trolled 7-regiosiomer of adenine would be transformed to
adenosine (2) via a 7,9-bis-ribosyladenine intermediate, as
it has been documented for 6-oxopurine bases: hypoxan-
thine12 and guanine.4,5,13
contained 36% of the 7-isomer (5; isolated by column chro-
matography), along with a smaller amount (less than 10%) of
triacetyladenosine (6). When heating was continued for a
longer time, the 9-regioisomer 6 was the only remaining
product. Similarly, the isolated product 5 could be quantita-
tively isomerized to 6 in the presence of p-toluenesulfonic
acid on refluxing in chlorobenzene. This shows clearly that
7-riboadenine is a kinetic product in the ribosylation of ade-
nine. Most probably, the 7/9 transglycosylation proceeds
via a 7,9-bis-ribosyl intermediate, likewise in the guanine
series, but the reaction equilibrium is totally shifted toward
the 9-regioisomer (6). The product 5 was deprotected with
aqueous ammonia in methanol to give 7-(b-D-ribofuranosyl)-
adenine (7), identical in all respects with the sample obtained
in the presence of SnCl4.7
2. Results and discussion
In the course of our systematic reinvestigation on mecha-
nisms of the N-glycosylic bond formation, we performed a
series of experiments to establish the factors responsible for
either 3/9 or 7/9 mechanism in the ribosylation of ade-
nine. This time we focused our attention on the silyl method,
the most common synthetic procedure at present. Thus, ade-
nine (1) was silylated with hexamethyldisilazane (HMDS)
and then subjected to ribosylation with 1,2,3,5-tetra-O-ace-
tyl-b-D-ribofuranose in the presence of trimethylsilyl triflate
(TMSOTf) (Scheme 2). After 80 min the reaction mixture
Considering the above-mentioned result as well as the liter-
ature data,6,7 we could expect a similar initial 7-ribosylation
in the reaction of N6-acylated derivatives of adenine, per-
€
formed according to the Vorbruggen’s procedure. The reac-
tion sequence is shown in Scheme 3. N6-Isobutyryl (8a) and
N6-benzoyladenine (8b) were silylated with N,O-bis-tri-
methylsilylacetamide (BSA), and then subjected to ribosyl-
ation with 1,2,3,5-tetra-O-acetyl-b-D-ribofuranose in the
presence of TMSOTf. Surprisingly, the reaction gave, along
with the 9-ribo products (9a,b), new products of the structure
of 1-regioisomers (10a,b) in the yield of 33% and 30%, re-
spectively. In addition, a careful chromatographic separation
in the isobutyryl series allowed to isolate a minor reaction
product, the 1,9-bis-ribosyl derivative 11 (5%). The depro-
tection of 10a with methanolic ammonia gave 1-(b-D-ribo-
furanosyl)adenine (12), a new regioisomer of naturally
occurring adenosine.
Compounds 10a,b underwent isomerization to the respective
9-regioisomers (9a,b) after a prolonged reaction time, and
this proves the kinetic nature of the 1-regioisomers of
adenosine. Furthermore, the isolated N6-isobutyryl deriva-
tive (10a) underwent an almost quantitative conversion
to 9a under transglycosylation conditions (refluxing in
Scheme 2. Reagents and conditions: (a) (NH4)2SO4, HMDS, reflux, 2.5 h;
(b) CH3CN, 1,2,3,5-tetra-O-acetyl-b-D-ribofuranose, TMSOTf, rt, 80 min;
(c) p-TsOH, chlorobenzene, 150 ꢀC, 2.5 h; and (d) 25% NH4OH, MeOH,
rt, 30 min.
Scheme 3. Reagents and conditions: (a) BSA/CH3CN, Ar, 60 ꢀC, 30 min; (b) 1,2,3,5-tetra-O-acetyl-b-D-ribofuranose, TMSOTf, 60–75 ꢀC, 3 h; (c) p-TsOH,
chlorobenzene, 60–150 ꢀC, 10–150 min; (d) NH3/MeOH, 25 ꢀC, 24 h; and (e) AcOCH2CH2OCH2OAc, p-TsOH, chlorobenzene, reflux, 4 h.

G. Framski et al. / Tetrahedron 62 (2006) 10123–10129
10125
16
€
€
Vorbruggen and Hofle, but this has never been proved
experimentally.
chlorobenzene in the presence of p-toluenesulfonic acid).
Under the same conditions, the 1,9-bis-ribofuranosyl
intermediate (11) was decomposed to a 6:1 mixture of the
respective 9- and 1-regioisomers (9a and 10a). Finally, the
reaction of 10a with 2-acetoxyethyl acetoxymethyl ether14,15
resulted in the formation of acycloadenosine derivative 13,
and this is evidence for an intermolecular course of the
1/9 isomerization.
All compounds were fully characterized by the H and 13C
1
NMR (1D & 2D, NOE) techniques and analytical methods.
Table 1 presents the first comparison of 13C NMR spectra
of adenosine and all its regioisomers. In particular, the
data for 7-riboadenine (7) are in good agreement with those
published for related compounds.17,18 Both isomeric nucleo-
sides, 1- and 7-(b-D-ribofuranosyl)adenine (12 and 7, re-
spectively) gave crystals suitable for X-ray diffraction and
their crystal structures have been determined (Fig. 1).19 In
the crystals of 12, there are two symmetry independent mole-
cules, denoted A and B, showing no significant differences
in their geometrical parameters. Interestingly, both regio-
isomeric nucleosides in their crystal structures adopt con-
formations, which enable the formation of a three-center
intramolecular N–H/O hydrogen bond between the amino
group of the base and O10 and O50 from the ribose moiety
(Fig. 2). To form such a bond, the isomers must adopt differ-
ent sugar conformations: 1-riboadenine (12) adopts an enve-
lope C20-endo form, while 7-riboadenosine (7) occurs in
crystal as a C20-endo-C10-exo conformer.
Taking into account these experimental facts, we can
propose a new mechanism of the ribosylation in the
N6-acyl series (Scheme 4), shown here for the N6-isobutyryl
derivatives. In the first step, 9,N6-bis-trimethylsilylated sub-
strate (14) reacts with the acyloxonium sugar cation (15),
generated from tetraacetylribose.10 The position N9, an ulti-
mate site of ribosylation, is already blocked by the TMS
group. Therefore, the initial ribosylation may take place
at any alternative site, e.g., N1, N3, or N7. As presented
above, the N1-position is ribosylated first, perhaps due to
a limited access to N7 in the presence of the two N6-substit-
uents, and this leads to the kinetically controlled 1-ribo-
nucleoside 16 (isolated as a desilylated derivative 10a).
Compound 16 then undergoes an intermolecular transglyco-
sylation to regain the thermodynamically preferred aro-
matic system, corresponding to the most stable N-9-H
tautomer of adenine. Thus, a second ribosylation at N9
gives the 1,9-bis-ribosyl intermediate 11, which after pro-
tonation (structure 17) undergoes a decomposition to the
final 9-regioisomer 9a, with liberation of the acyloxonium
cation 15. The 1/9 isomerization is irreversible. It is wor-
thy to note that quite a similar mechanism can be drawn for
an isomeric structure of the bis-trimethylsilyl substrate 14, in
which the second TMS group would be attached not to N6,
but to oxygen atom of the N6-acyl substituent, as it has been
proposed recently.10 In that case, explanation of the observed
regioselectivity would be even more convincing. Interest-
ingly, the formation of 1-(b-D-ribofuranosyl)adenine as
a possible kinetic intermediate has been anticipated by
3. Conclusion
The mechanism of ribosylation in the adenine series evi-
dently depends on reaction conditions. While application
of 1-halosugars in strongly acidic media favors the 3/9
pathway, the use of 1-O-acetylated sugars and Lewis acids
in the silyl approach results in either 7/9 or 1/9 glycosyl-
ation sequence. In the latter method, a comparison of the
data obtained in this work and those reported in the literature
allows us to formulate some general rules, which may be
useful in the synthesis of regioisomers of adenosine. The
6
7,8
use of Lewis acid catalysts like HgBr2 or SnCl4 results
in the formation of 7-(b-D-ribofuranosyl)adenine as
Scheme 4. A proposed mechanism for the 1/9 ribosylation pathway.

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G. Framski et al. / Tetrahedron 62 (2006) 10123–10129
Table 1. Comparison of the 13C NMR spectra of adenosine and its regioisomers (151 MHz, DMSO-d6, TMS)
Compounds
C2
C4
C5
C6
C8
C10
C20
C30
C40
C50
Adenosine
152.42
143.53
152.61
140.43
149.06
147.14
160.69
148.58
119.38
120.68
110.05
118.88
156.19
155.69
151.49
154.42
139.96
151.28
144.41
150.53
87.92
94.95
89.18
92.14
73.44
72.12
74.82
72.66
70.70
71.01
68.78
69.82
85.93
87.82
86.16
86.19
61.71
61.85
60.32
60.43
3-(b-D-Ribofuranosyl)adenine
7-(b-D-Ribofuranosyl)adenine (7)
1-(b-D-Ribofuranosyl)adenine (12)
Figure 2. A superposition of the two isomers (12 and 7) in the conforma-
tions adopted in their crystal structures.
in the synthesis of related analogs of potential biological
activity and in the study of base-pairing properties on the
oligonucleotide level.
4. Experimental
4.1. General
3-(b-D-Ribofuranosyl)adenine for comparative study was
obtained according to Leonard and Laursen.2 N6-Benzoyl-
adenine (8b) was prepared according to the published proce-
dure.20 Melting points were determined on a Laboratory
Devices Mel-Temp II micromelting points apparatus and
are uncorrected. UV spectra were measured on a Beckman
DU-65 spectrophotometer. The optical rotations were mea-
sured with a Perkin–Elmer 243B polarimeter. The infrared
spectra were determined in KBr with a Bruker IFS
66v/s spectrophotometer. 1H (300 MHz) and 13C NMR
(75.5 MHz) spectra were recorded on a Varian Unity 300
FT NMR 300 MHz spectrometer with tetramethylsilane as
an internal standard, and chemical shifts are reported in
d-values (ppm). 2D 1H and 13C NMR (151 MHz) were
recorded on a Bruker Avance 600 MHz spectrometer. The
following NMR techniques were applied for structural
assignment of the obtained compounds: COSY, HMQC,
HMBC, TOCSY, and NOE. Mass spectra were taken on an
AMD-604 spectrometer using the LSIMS technique (Cs+,
12 keV; in NBA). Elemental analyses were performed on a
Perkin–Elmer 240 Elemental Analyzer. TLC was conducted
Figure 1. Ortep drawings of (a) 12, molecule A, and (b) 7 at 50% probability
level with atom numbering. Intramolecular hydrogen bonds are shown as
dashed lines.
a kinetically controlled product, no matter whether the N6-
exocyclic amino group is protected or not. However, when
€
the Vorbruggen catalyst (TMSOTf) is applied, the presence
of N6-protection is crucial for the course of reaction: the
ribosylation of persilylated derivatives of adenine without
the N6-acyl protection leads to the formation of 7-(b-D-ribo-
furanosyl)adenine, while N6-acylated substrates are directly
ribosylated in the position N1. To our knowledge, this is the
first synthesis of the so far unknown 1-(b-D-ribofuranosyl)
adenine, and that compound can now be obtained in a reason-
able yield. The approach presented here should prove useful

G. Framski et al. / Tetrahedron 62 (2006) 10123–10129
10127
on Merck silica gel F254 60 plates using the following solvent
systems (measured by volume): A, chloroform/methanol
(9:1); B, toluene/ethanol (4:1); C, isopropanol/concd
NH4OH/water (7:1:2). For preparative short-column chro-
matography Merck TLC gel H 60 was used.
was crystallized from MeOH (60 mg, 72%): mp 207–
210 ꢀC (lit. 211–2127 and 24623); Rf 0.57 (C; adenosine
0.65); [a]2D0 ꢁ93.6 (c 0.25, H2O; lit. ꢁ1007); lmax (MeOH)
245 (sh), 270 nm; nmax 3500–2600 (br), 3443, 3345, 3246,
1636, 1593, 1561, 1520, 1489, 1447, 1408, 1303, 1116, 1057,
990, 881 cmꢁ1; 1H NMR (DMSO-d6): 3.68 (m, 2H), 3.99 (q,
1H, J¼5.0 Hz), 4.07 (q, 1H, J¼6.0 Hz), 4.12 (m, 1H), 5.31
(d, 1H, J¼4.2 Hz), 5.36 (t, 1H, J¼4.8 Hz), 5.62 (d, 1H,
J¼6.0 Hz), 5.82 (d, 1H, J¼7.2 Hz), 6.99 (s, 2H), 8.22 (s,
1H), 8.52 (s, 1H). Anal. calcd for C10H13N5O4 (267.25): C,
44.94; H, 4.90; N, 26.21. Found: C, 44.86; H, 4.79; N, 26.17.
4.2. Crystal data
The diffraction data were collected at 130 K with a Kuma-
CCD diffractometer, CrysAlis CCD, and CrysAlis RED.
Version 1.171. Oxford Diffraction, using graphite mono-
chromated Mo Ka radiation. The structures were solved
by direct methods with the program SHELXS-9721 and
refined by full-matrix least-squares method on F2 with
SHELXL-97.22
4.5. N6-Isobutyryladenine (8a)
Adenine (6.7 g, 49.58 mmol) (vacuum dried) was stirred in
isobutyric anhydride (160 mL) at 70 ꢀC (oil bath tempera-
ture) for 4 h. TLC analysis showed the presence of two prod-
ucts, mono- and disubstituted ones. The mixture was then
refluxed in absolute methanol (170 mL) until the disubsti-
tuted product was completely decomposed (2 h). The solvent
was evaporated, and the resulting syrup was crystallized from
ethanol. The product was recrystallized from boiling ethanol
to give 9.15 g of white crystals (90%): mp 231–233 ꢀC; Rf
0.38(A), 0.23(B); lmax (MeOH) 281, 291 nm; 1H NMR
(DMSO-d6): 1.17 (d, 6H, J¼6.9 Hz), 2.92 (septet, 1H,
J¼6.9 Hz), 8.39 (s, 1H), 8.63 (s, 1H), 11.15 (s, 1H), 12.21
(s, 1H); HRMS: calcd for C9H12N5O (M+H): m/z
206.1041, found: 206.1032.
4.2.1. 7-(b-D-Ribofuranosyl)adenine 7 (CCDC 297135).
C10H13N5O4, orthorhombic, space group P21212,
˚
a¼8.2006(7), b¼17.8347(14), c¼7.7064(7) A, V¼
1346.04(19) A , Z¼4, dx¼1.575 g cmꢁ3, T¼130 K. Data
3
˚
were collected for a crystal with dimensions 0.2ꢂ
0.2ꢂ0.02 mm3. Final R indices for 1021 reflections with
I>2s(I) and 177 refined parameters are: R1¼0.0346,
wR2¼0.0765 (R1¼0.0442, wR2¼0.0799 for all 1177 data).
4.2.2. 1-(b-D-Ribofuranosyl)adenine 12 (CCDC 297134).
C10H13N5O4, monoclinic, space group P21, a¼6.7737(4),
b¼16.7201(8), c¼9.7134(4) A, b¼90.895(4)ꢀ, V¼
˚
1099.98(10) A , Z¼4, dx¼1.614 g cmꢁ3, T¼130 K. Data
3
˚
were collected for a crystal with dimensions 0.5ꢂ0.2ꢂ
0.2 mm3. Final R indices for 2232 reflections with I>2s(I)
and 369 refined parameters are: R1¼0.0240, wR2¼0.0620
(R1¼0.0251, wR2¼0.0620 for all 2314 data).
4.6. N6-Isobutyryl-9-(20,30,50-tri-O-acetyl-b-D-ribofura-
nosyl)adenine (9a), N6-isobutyryl-1-(2,3,5-tri-O-acetyl-
b-D-ribofuranosyl)adenine (10a), and N6-isobutyryl-1,9-
bis-(20,30,50-tri-O-acetyl-b-D-ribofuranosyl)adenine (11)
4.3. 7-(2,3,5-Tri-O-acetyl-b-D-ribofuranosyl)adenine (5)
and 9-(2,3,5-tri-O-acetyl-b-D-ribofuranosyl)adenine (6)
An anhydrous suspension of 8a (0.90 g, 4.38 mmol) and
1,2,3,5-tetra-O-acetyl-b-D-ribofuranose (1.79 g, 5.63 mmol)
in dry acetonitrile (20 mL) was washed with argon for
30 min, then BSA (1.64 g, 8.09 mmol) was added. The mix-
ture was stirred at 75 ꢀC for 30 min until a clear solution was
obtained. TMSOTf (0.48 g, 2.11 mmol) was then added and
the mixturewas stirred at 75 ꢀC for 3 h. After cooling down to
rt the obtained solution was diluted with CH2Cl2 (180 mL)
and extracted with cold saturated solution of NaHCO3
(150 mL). The organic layer was dried over Na2SO4, evapo-
rated to a yellow solid foam and then chromatographed on
a SiO2 column in a CH3Cl/CH3CN gradient (from 2:1 to
1:1) to give (in order of elution): compound 11, an oil:
0.16 g, 5%; Rf 0.80(A), 0.50(B); [a]2D0 +20.7 (c 0.31,
MeOH); lmax (MeOH) 293 nm; nmax 3600–2850 (br), 1755,
1685, 1629, 1547, 1531, 1508, 1464, 1430, 1374, 1229
To a stirred suspension of adenine (1; 200 mg, 1.48 mmol) in
HMDS (6 mL) was added (NH4)2SO4 (10 mg, 0.087 mmol)
and the mixture was refluxed under argon for 2.5 h. The
resulting solution was concentrated to an oil, dissolved in
dry acetonitrile (6 mL), and treated with tetraacetylribose
(470 mg, 1.47 mmol) and TMSOTf (267 mL, 1.47 mmol).
After stirring at rt for 80 min, the reaction mixturewas evapo-
rated to a white solid foam. The products were isolated by
SiO2 column chromatography in a gradient of CHCl3/
MeOH (from 95:5 to 9:1) to give (in order of elution) the
9-isomer 6 (31 mg, 5.3%); Rf 0.48(A), 0.32(B) and the 7-
isomer 5 (208 mg, 36%) as a solid foam: Rf 0.37(A),
0.10(B); lmax (MeOH) 245 (sh), 274 nm; 1H NMR (CDCl3):
2.04, 2.10, 2.16 (3s, 3ꢂ3H), 4.36 (m, 3H), 5.47 (dd, 1H, J¼
4.5, 6.4 Hz), 5.59 (t, 1H, J¼6.5 Hz), 5.70 (br s, 2H), 6.02
(d, 1H, J¼6.6 Hz), 8.14 (s, 1H), 8.54 (s, 1H); 13C NMR
(DMSO-d6): 20.11, 20.32, 20.39, 62.43, 68.74, 72.49,
79.65, 86.41, 110.19, 143.94, 151.30, 152.89, 160.43,
169.10, 169.48, 169.99; HRMS: calcd for C16H20N5O7
(M+H): m/z 394.1362, found: 394.1347.
1
(br), 1097, 1047, 1017, 904, 757 cmꢁ1; H NMR (CDCl3):
1.22, 1.24 (2d, 6H, J¼6.9 Hz), 2.08, 2.09, 2.10, 2.12, 2.14,
2.20 (6s, 6ꢂ3H), 2.75 (septet, 1H, J¼6.9 Hz), 4.41–4.45
(m, 6H), 5.33 (dd, 1H, J¼5.4, 7.8 Hz), 5.60 (m, 2H), 5.88
(t, 1H, J¼5.4 Hz), 6.03 (d, 1H, J¼5.4 Hz), 6.27 (d, 1H,
J¼2.4 Hz), 7.81 (s, 1H), 8.35 (s, 1H); 13C NMR (CDCl3):
18.97, 19.06, 20.21, 20.31, 20.35, 20.44, 20.49, 20.73, 37.41,
62.43, 62.76, 68.96, 69.88, 72.23, 72.42, 78.76, 79.54, 85.83,
90.04, 120.84, 139.87, 141.73, 143.35, 146.22, 169.13,
169.26, 169.40, 169.43, 170.23, 170.30, 187.60; HRMS:
calcd for C31H40N5O15 (M+H): m/z 722.2521, found:
722.2538; the 9-isomer 9a, an oil: 0.21 g, 10%; Rf 0.70(A),
0.46(B); lmax (MeOH) 272 nm; the 1-isomer 10a, a white
4.4. 7-(b-D-Ribofuranosyl)adenine (7)
A solution of 5 (122 mg, 0.310 mmol) in methanol (4 mL)
was treated with 25% NH4OH (2 mL). After 30 min at rt
the solvent was evaporated to obtain a white solid, which

10128
G. Framski et al. / Tetrahedron 62 (2006) 10123–10129
solid: 0.66 g, 33%; mp 159–161 ꢀC (40% EtOH); Rf 0.54(A),
0.45(B); [a]2D0 +86.5 (c 0.27, MeOH); lmax (MeOH) 313 nm;
nmax 3500–2850 (br), 1750, 1651, 1599, 1504, 1427, 1375,
1365, 1243, 1231, 1215, 1115, 1099, 1063, 1018 cmꢁ1; 1H
NMR (CDCl3): 1.17 (d, 3H, J¼7.0 Hz), 1.19 (d, 3H,
J¼7.0 Hz), 2.06, 2.18, 2.24 (3s, 3ꢂ3H), 2.65 (septet, 1H,
J¼7.0 Hz), 4.42 (dd, 1H, J¼3.0, 12.6 Hz), 4.51–4.55 (m,
2H), 5.34 (dd, 1H, J¼4.8, 8.4 Hz), 5.63 (d, 1H, J¼4.8 Hz),
6.65 (s, 1H), 8.13 (s, 1H), 8.82 (s, 1H), 12.49 (br s, 1H);
13C NMR (CDCl3): 19.55, 19.84, 20.26, 20.30, 20.72,
39.51, 61.05, 67.67, 74.51, 78.86, 90.31, 114.27, 141.53,
142.17, 148.02, 157.05, 168.93, 169.20, 170.23, 188.87;
HRMS: calcd for C20H26N5O8 (M+H): m/z 464.1781, found:
464.1809. Anal. calcd for C20H25N5O8 (463.45): C, 51.83; H,
5.44; N, 15.11. Found: C, 51.75; H, 5.23; N, 15.01.
2-acetoxyethyl acetoxymethyl ether14 (253 mL, 1.62 mmol),
and p-toluenesulfonic acid monohydrate (6.2 mg,
0.032 mmol) in chlorobenzene (7.5 mL) was refluxed for
4 h. The solvent was removed in vacuo and the residue was
subjected to SiO2 column chromatography in a CH3Cl/
CH3OH gradient (from 98:2 to 9:1) to yield 13 as an oil,
57 mg (55%); Rf 0.60(A), 0.34(B); lmax (MeOH) 273 nm;
nmax 3400–2800 (br), 3262, 1736, 1729, 1721, 1686, 1605,
1
1589, 1580, 1454, 1266, 1237, 1220, 1050, 761 cmꢁ1; H
NMR (DMSO-d6): 1.13 (d, 6H, J¼6.9 Hz), 1.92 (s, 3H),
2.94 (septet, 1H, J¼6.9 Hz), 3.74 (m, 2H), 4.07 (m, 2H),
5.68 (s, 2H), 8.61 (s, 1H), 8.68 (s, 1H), 10.67 (s, 1H);
HRMS: calcd for C14H20N5O4 (M+H): m/z 322.1515, found:
322.1510.
4.9.2. Isomerization of 5 to 6. A suspension of protected
7-isomer 5 (4.2 mg, 0.01 mmol) and p-toluenesulfonic acid
monohydrate (0.4 mg, 0.002 mmol) in chlorobenzene
(0.4 mL) was stirred at 150 ꢀC for 2.5 h to give a product
identical with an authentic sample of 6 (>90%; TLC, H
NMR).
4.7. N6-Benzoyl-9-(20,30,50-tri-O-acetyl-b-D-ribofurano-
syl)adenine (9b) and N6-benzoyl-1-(2,3,5-tri-O-acetyl-b-
D-ribofuranosyl)adenine (10b)
1
A similar experimental procedure as described for synthesis
of 10a was applied in the N6-benzoyl series. Reaction
of 8b (0.60 g, 2.50 mmol), tetraacetylribose (0.955 g,
3.00 mmol), BSA (1.006 g, 4.90 mmol), and TMSOTf
(0.280 g, 1.30 mmol) was carried out at 60 ꢀC for 3 h to
give, after chromatographic separation, the 9-isomer 9b, an
oil: 0.099 g, 8%; Rf 0.80(A), 0.50(B); lmax (MeOH) 232,
280 nm, and the 1-isomer 10b, an oil: 0.377 g, 30%; mp
202–205 ꢀC (EtOH); Rf 0.62(A), 0.49(B); lmax (MeOH)
228, 332 nm; nmax 3600–2900 (br), 1749, 1745, 1641,
1599, 1557, 1500, 1487, 1423, 1373, 1315, 1287, 1230, 1118,
4.9.3. Isomerization of 10a to 9a. A sample of 10a (6.0 mg,
0.013 mmol) was stirred with p-toluenesulfonic acid
(0.25 mg, 0.0013 mmol) in chlorobenzene (1 mL) at 60 ꢀC
for 2 h. TLC analysis showed the formation of 9a (ca.
90%), and traces of 8a.
4.9.4. Decomposition of 11. A sample of 1,9-bis-ribofura-
nosyl derivative 11 (90.0 mg, 0.12 mmol) was refluxed
with p-toluenesulfonic acid (2.2 mg, 0.012 mmol) in chloro-
benzene (5 mL) for 10 min. After this time TLC analysis
showed a mixture of 9a and 10a in a ratio 6:1, respectively.
The structure of products was confirmed after their chroma-
tographic separation (TLC, 1H NMR, UV).
1
1095, 1058 cmꢁ1; H NMR (CDCl3): 2.07, 2.08, 2.25 (3s,
3ꢂ3H), 4.41–4.60 (m, 3H), 5.38 (dd, 1H, J¼5.1, 8.1 Hz),
5.64 (dd, 1H, J¼2.1, 5.1 Hz), 6.70 (d, 1H, J¼2.1 Hz), 7.42
(m, 2H), 7.52 (m, 1H), 8.17 (s, 1H), 8.22 (m, 2H), 8.90 (s,
1H), 12.65 (br s, 1H); 13C NMR (CDCl3): 20.31, 20.41,
20.80, 61.26, 68.09, 74.90, 79.29, 89.93, 114.61, 128.03,
128.17, 129.75, 137.25, 141.95, 142.20, 148.81, 157.60,
168.88, 169.21, 170.24, 175.18; HRMS: calcd for
C23H24N5O8 (M+H): m/z 498.1625, found: 498.1655.
References and notes
1. Watanabe, K. A.; Hollenberg, D. H.; Fox, J. J. J. Carbohydr.
Nucl. Nucl. 1974, 1, 1–37 and references cited therein.
2. Leonard, N. J.; Laursen, R. A. Biochemistry 1965, 4, 354–364.
3. Shimizu, B.; Miyaki, M. Chem. Pharm. Bull. 1970, 18,
732–740; Chem. Pharm. Bull. 1970, 18, 1446–1456.
4. Boryski, J. Nucleosides Nucleotides 1996, 15, 771–791 and
references cited therein.
5. Boryski, J. J. Chem. Soc., Perkin Trans. 2 1997, 649–652.
6. Ryan, K. J.; Acton, E. M.; Goodman, L. J. Org. Chem. 1971, 36,
2646–2657.
7. Akhrem, A. A.; Adarich, A. K.; Kulinkovich, N. L.;
Mikhailopulo, I. A.; Posshasteva, E. B.; Timoshchuk, V. A.
Dokl. Akad. Nauk. SSSR 1974, 219, 99–102.
8. Itoh, T.; Mizuno, Y. Heterocycles 1976, 5, 285–292.
9. Moyroud, E.; Strazewski, P. Tetrahedron 1999, 55, 1277–1284.
10. Vorbr€uggen, H. Acta Biochim. Polon. 1996, 43, 25–36 and
references cited therein.
4.8. 1-(b-D-Ribofuranosyl)adenine (12)
A solution of 10a (0.222 g, 0.48 mmol) in saturated metha-
nolic ammonia (10 mL) was stirred at 25 ꢀC for 24 h. The
solvent was evaporated to a white solid, which was stirred
in CHCl3/MeOH (1:1, 5 mL) for 2 h. The precipitate was fil-
tered off to give 0.120 g (93%) of 12. An analytical sample
was crystallized from water, mp>178 ꢀC (decomp.); Rf
0.53 (C); [a]D20 ꢁ23.5 (c 0.14, H2O); lmax (MeOH) 228,
275 nm; nmax 3550–2300 (br), 3407, 3339, 3189, 1679,
1623, 1561, 1557, 1476, 1450, 1358, 1310, 1117, 1071, 907,
863 cmꢁ1; 1H NMR (D2O): 4.04 (dd, 1H, J¼3.6, 12.6 Hz),
4.07 (dd, 1H, J¼2.4, 12.6 Hz), 4.49 (dd, 1H, J¼3.6,
51 Hz), 4.79 (t, 1H, J¼6.0 Hz), 6.15 (d, 1H, J¼6.0 Hz),
8.21 (s, 1H), 8.60 (s, 1H). Anal. calcd for C10H13N5O4
(267.25): C, 44.94; H, 4.90; N, 26.21. Found: C, 44.74; H,
4.69; N, 26.15.
11. Nakazaki, N.; Sekiya, M.; Yoshino, T.; Ishido, Y. Bull. Chem.
Soc. Jpn. 1973, 46, 3858–3863.
12. Dudycz, L. W.; Wright, G. E. Nucleosides Nucleotides 1984, 3,
33–44.
4.9. Transglycosylation reactions
13. Manikowski, A.; Boryski, J. Nucleosides Nucleotides 1999, 18,
1057–1059.
14. Rosovsky, A.; Kim, S.-H.; Wick, M. J. Med. Chem. 1981, 24,
1177.
4.9.1. N6-Isobutyryl-9-[(2-acetoxyethoxy)methyl]adenine
(13). An anhydrous solution of 10a (150 mg, 0.32 mmol),

G. Framski et al. / Tetrahedron 62 (2006) 10123–10129
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15. Framski, G.; Manikowski, A.; Zandecki, T.; Boryski, J. Nucl.
Acids Res. Suppl. 2003, 3, 11–12.
336033 or e-mail: deposit@ccdc.cam.ac.uk], quoting the depo-
sition numbers.
€
€
16. Vorbruggen, H.; Hofle, G. Chem. Ber. 1981, 114, 1256–1268.
17. Akhrem, A. A.; Mikhailopulo, I. A.; Abramov, A. F. Org.
Magn. Reson. 1979, 12, 247–253.
20. Ness, N. K. Synthetic Procedures in Nucleic Acid Chemistry;
Zorbach, W. W., Tipson, R. S., Eds.; Interscience: New York,
NY, 1968; Vol. 1, pp 183–187.
18. Seela, F.; Winter, H. Bioorg. Med. Chem. Lett. 1993, 3,
273–276.
21. Sheldrick, G. M. SHELXL-97: Program for a Crystal Structure
€
€
Solution; University of Gottingen: Gottingen, Germany, 1997.
22. Sheldrick, G. M. SHELXL-97: Program for the Refinement of
a Crystal Structure from Diffraction Data; University of
19. Crystallographic data for compounds 12 and 7 have been
deposited with Cambridge Crystallographic Data Centre
(CCDC deposition numbers 297134 and 297135, respectively).
Copies of the data can be obtained upon request from CCDC,
12 Union Road, Cambridge CB2 1EZ, UK [fax: +44 1223
€
€
Gottingen: Gottingen, Germany, 1997.
23. Montgomery, J. A.; Thomas, H. J. J. Am. Chem. Soc. 1965, 87,
5442–5447.