Regioselective nitration of purine nucleosides: Synthesis of 2- nitroadenosine and 2-nitroinosine

Article abstract of DOI:10.1016/S0040-4039(99)02271-6

Nitration reactions of 6-substituted purine nucleosides with tetrabutylammonium nitrate/trifluoroacetic anhydride (TBAN/TFAA) have been studied. This nitrating mixture selectively nitrates C-6 substituted purines at the 2-position, but the method is limited to substrates without NH or OH substituents. Acetylated 6-chloropurine riboside was cleanly nitrated (DCM, 0°C, 71%) and converted to nitro substituted adenosine and inosine in a few simple steps. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4039(99)02271-6

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 41 (2000) 1291–1295
Regioselective nitration of purine nucleosides: synthesis of
2-nitroadenosine and 2-nitroinosine
Paymaneh Y. F. Deghati, Martin J. Wanner and Gerrit-Jan Koomen ∗
Laboratory of Organic Chemistry, Institute of Molecular Chemistry, University of Amsterdam, Nieuwe Achtergracht 129,
NL-1018 WS Amsterdam, The Netherlands
Received 2 November 1999; accepted 7 December 1999
Abstract
Nitration reactions of 6-substituted purine nucleosides with tetrabutylammonium nitrate/trifluoroacetic anhy-
dride (TBAN/TFAA) have been studied. This nitrating mixture selectively nitrates C-6 substituted purines at the
2-position, but the method is limited to substrates without NH or OH substituents. Acetylated 6-chloropurine
riboside was cleanly nitrated (DCM, 0°C, 71%) and converted to nitro substituted adenosine and inosine in a few
simple steps. © 2000 Elsevier Science Ltd. All rights reserved.
Keywords: nitration; adenosine; inosine.
Adenosine analogs substituted at the 2-position show interesting activity in several biological systems.
For instance adenosines containing a halogen atom at the 2-position display cytotoxic activity, and
especially 2-chloro-20-deoxyadenosine (cladribine), which is a potent inhibitor of DNA synthesis, is
currently used against leukaemia and in the treatment of chronic lymphoid malignancies.1 The apoptosis
inducing properties of some 2-halo-substituted adenosine analogs were recently published.2 In addition,
introduction of carbon, amino or oxygen substituents at the adenosine 2-position increases the selectivity
of binding to adenosine receptors.3,4
Most of the procedures towards the synthesis of 2-substituted adenosines are based on 2,6-
dichloropurine or use guanosine as starting material.5 To improve the availability of disubstituted purine
systems we describe here our studies to functionalize 6-substituted nucleosides at the 2-position.
During our investigations towards selective derivatization of 1-deazapurine6 and 1-deaza-2-azapurine
ribosides,7 we introduced the tetrabutylammonium nitrate/trifluoroacetic anhydride (TBAN/TFAA)
system8 for the nitration of electron deficient and acid labile nucleosides. The surprising mildness and
selectivity of this reagent stimulated us to apply this nitration to the purine series using adenosine and
inosine as starting materials.
∗
Corresponding author.
0040-4039/00/$ - see front matter © 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(99)02271-6
tetl 16199

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Nitration of protected inosine using a reagent prepared from TFAA and ammonium nitrate is described
in the literature and gives the N1-nitrated nucleoside (Scheme 1).9 Application of the TBAN/TFAA
nitration on triacetyl protected inosine 3 or adenosine 4 was, in our hands, also ineffective for C-nitration.
We were however pleased to find that triacetyl protected 6-chloropurine riboside 2 was cleanly nitrated
using TBAN/TFAA. Of the two available carbon positions in the purine ring only the 2-position was
reactive, and crystalline 2-nitro-6-chloropurine riboside 10 was readily obtained in good yield.10 To
investigate the scope of this reaction we returned to triacetyl adenosine, which was completely acetylated
using acetic anhydride and DMAP at 130°C to remove all the acidic hydrogen atoms. Nitration of N,N-
diacetyladenosine triacetate 6 was now possible to give 55% of the corresponding 2-nitro derivative 14.
Extension of the nitration method was accomplished using N6-cyclopentyladenosine (CPA) as a substrate.
Peracetylation (Ac2O/DMAP, 120°C) gave 7 which was subsequently nitrated to give 15 as the only
product.
Scheme 1.
This TBAN/TFAA nitration proved to be a strongly substrate dependent process, since nucleosides
such as adenosine tetraacetate 5 or nebularine triacetate 1 did not give any of the expected nitrated
products. Formation of polar side products as a result of N-nitration and/or glycosidic bond cleavage
was observed. Nebularine N-1-oxide 8, prepared by DMDO oxidation of 1 was also not suitable as a
substrate although the introduction of N-oxides in the 1-deazapurine riboside series was shown to be
very successful for the synthesis of nitro compound 17 (Fig. 1).6 It seems likely however that no suitable
position for C-nitration in 8 is available since under these conditions N-oxides have strong meta-directing
properties. We have observed before that 3,5-dinitro derivative 18 was formed from pyridine N-oxide.
Pyridine itself produced no C-nitrated products under these conditions since N-nitration is prevailing.11
Fig. 1. R=2,3,5-tri-O-acetyl-β-D-ribofuranosyl
To summarize, for successful nitration the following substrate requirements can be deduced from these
studies:
– no acidic protons (e.g. NH or OH);
– no nucleophilic nitrogen atoms (as in pyridine);
– radical stabilizing substituent (Cl, NR2, N-oxide) is required.

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1. Mechanism of the nitration
Since the conventional ‘nitronium-ion’ nitration mechanism was introduced12 numerous alternative
processes have been observed, all as a result of the many forms NOx can adopt. In a recent publication,
Ridd13 reviewed a group of unconventional nitration pathways, most of them based on radical species
and/or electron transfer processes. Only a few examples are known in which electron deficient substrates
are nitrated at room temperature and from these, the nitration reactions of chloro-nitrobenzenes using
14
N2O5/HNO3 give a clear indication of radical addition. The nitro-nitrate-addition products, formed as
intermediates, were observed by 15N CIDNP NMR and support a radical addition mechanism, although
electrophilic processes catalyzed by HNO3 seem to dominate the formation of the end products. NO2
is not reactive enough for addition to the aromatic ring and therefore a more reactive species such as
NO3, which is formed in an equilibrium from N2O5, initiates the substitution reaction. Comparable me-
chanisms were suggested to explain unusual selectivity during Kyodai nitration with NO2/O3, although
electron transfer from electron rich substrates to NO3 was preferred as the initiating step.15
In the TBAN/TFAA system, trifluoroacetyl nitrate presumably splits homolytically into NO2 and the
trifluoroacetate radical (Scheme 2).16 It should be noted that in theory N2O5 and consequently NO3
can be formed during the TBAN/TFAA nitrations. Addition of the reactive trifluoroacetyl radical to the
imidazole C-8 in e.g. 2 gives a highly delocalized radical that is stabilized by the substituent at C-6. In
the next step, combination of the radical with NO2 takes place at C-2, which is the only available carbon
atom. Elimination of trifluoroacetic acid from the unstable intermediate affords the product. In view of
the high oxidation potential of purines the alternative mechanism via electron transfer to NO3 seems
unlikely.
Scheme 2.
Any concurrent electrophilic processes during TBAN/TFAA nitration were excluded by a control
experiment using nitronium tetrafluoroborate. No nitration was observed and the starting material was
recovered. In addition, Evans16 and Njoroge8 have already shown that radical capture by adding up to
4 equiv. of TEMPO (2,2,6,6-tetramethyl-1-piperidinyloxy) to the TBAN/TFAA nitrating mixture almost
completely inhibited the reaction.
Although additional studies are necessary to clarify the exact mechanism, similarities between N2O5
and the TBAN/TFAA combination are obvious. The ease of handling the TBAN/TFAA mixture, in
combination with the relatively weakly acidic reaction conditions17 makes this reagent preferred over
several other nitrating agents.
2. Synthesis of 2-nitroadenosine and 2-nitroinosine
The versatility of 2-nitro-6-chloropurine riboside 10 was demonstrated by its conversion to the
biologically interesting 2-nitroadenosine 20 and 2-nitroinosine 19. Only 2-nitroinosine is described in

1294
the literature via formation in small amounts as a side product during deamination of guanosine with
nitrous acid.18
In a first approach to 2-nitroadenosine, the acetate protective groups were removed from 2-
nitroadenosine pentaacetate 14. However, under mild conditions such as KCN in methanol, replacement
of the nitro group occurred as a side reaction, affording 2-methoxyadenosine 21 in 47% yield and only
10% 2-nitroadenosine 20. In particular, removal of the second N-acetyl group was rather slow, allowing
substitution of the nitro group to give 21.
Alternatively 2-nitro-6-chloropurine riboside 10 was converted to 2-nitroadenosine triacetate 12
(Scheme 3) by replacement of the chloro substituent with sodium azide followed by conversion of the
azide to the corresponding amine. This triacetate was deacetylated to 2-nitroadenosine 2019 in satisfactory
yield using KCN in methanol. Finally the nitro substituent in 20 was hydrogenated to give the known
2-aminoadenosine 22.
Scheme 3. R=2,3,5-tri-O-acetyl-β-D-ribofuranosyl (i) NaOAc, EtOH, 81%; (ii) NH3, MeOH, 52%; (iii) NaN3, DMF; (iv) PPh3;
(v) HOAc, H2O, 64%, three steps; (vi) KCN, MeOH, 68%; (vii) H2, Raney Ni, 80%
2-Nitroinosine 19 was obtained by reaction of 10 with sodium acetate followed by aminolysis of the
acetates. Loss of the nitro group during the deprotection is prevented by deprotonation of the 6-OH by
ammonia.
The position of the nitro group in these nucleosides was unequivocally established using NMR
spectroscopy: long range correlation between C-10 and H-8 and between H-10 and C-8 was observed.
These structural assignments were confirmed by comparison of the spectroscopic data of diamine 22
with literature values.
In conclusion, a mild and selective nitration method for purine ribosides has been developed, which
has given access to a series of new purine nucleosides. The synthesis of further derivatives and studies of
their biological activity are currently in progress.
Acknowledgements
We wish to thank Dr. Brigitte E. A. Burm for stimulating discussions.
References
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1295
4. Examples of functionalization of the adenosine 2-position: (a) Nair, V.; Sells, T. B. Tetrahedron Lett. 1990, 31, 807; (b)
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10. A nitrating mixture prepared from TBAN (16 mmol) and TFAA (16 mmol) in DCM at 0°C was added to a solution of 2 (10
mmol) in DCM at 0°C. After 2.5 h the reaction mixture was poured into aqueous NaHCO3 and diluted with ether. Workup
of the organic layer produced 10 in 71% yield after trituration with methanol: mp 170–172°C; 1H NMR (400 MHz, CDCl3):
δ 8.58 (s, 1H), 6.30 (d 1H, J=5.3 Hz), 5.76 (dd, 1H, J=5.3 and 5.3 Hz), 5.57 (m, 1H), 4.52 (m, 1H), 4.44 (m, 2H), 2.16,
2.09, 2.06 (3×s, 9H); 13C NMR (100 MHz, CDCl3): δ 170.0, 169.4, 169.4, 153.1, 152.7, 151.3, 147.0, 134.8, 87.13, 81.16,
73.53, 70.53, 62.82; 20.56, 20.35, 20.14; obs. mass 458.0712, calcd mass for C16H17N5O9Cl (M+1) 458.0719.
11. An interesting β-nitration of pyridines was recently developed by treatment of N-nitropyridinium salts with sulfite: Bakke,
J. M.; Ranes, E.; Riha, J.; Svenson, H. Acta Chem. Scand. 1999, 53, 141.
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R. J. Chem. Soc., Perkin Trans. 2 1991, 645; (c) Titov, A. I. Tetrahedron 1963, 19, 557.
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19. Data for 20: mp 218–220°C; H NMR (DMSO-d6): δ 8.67 (s, 1H, H2), 8.31 (s, 2H, NH2), 5.93 (d, 1H, J=5.8 Hz, H1 ),
5.53, 5.27, 5.02 (3×m, OH), 4.61 (m, 1H, H2 ), 4.19 (m, 1H, H3 ), 3.99 (m, 1H, H4 ), 3.73–3.56 (m, 2H, H5 ); 13C NMR
(DMSO-d6): δ 158.9, 157.6, 151.4, 146.8, 124.5, 91.60, 88.80, 76.80, 73.36, 64.36; obs. mass 313.0895, calcd mass for
C10H13N6O6 (M+1) 313.0901.
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