From the ene reaction of nitrosocarbonyl intermediates with 3-methylbut-2-En-1-Ol, a new class of purine N,O-nucleoside analogues

Article abstract of DOI:10.1055/s-0032-1316916

Sterically encumbered nitrosocarbonyl derivatives of mesitylene or anthracene undergo ene reactions with 3-methylbut-2-en-1-ol to give the corresponding the 5-hydroxyisoxazolidine adducts in fair yields. According to the proposed mechanism, these heterocy

Full text of DOI:10.1055/s-0032-1316916

1414▌
PAPER
paper
From the Ene Reaction of Nitrosocarbonyl Intermediates with
3-Methylbut-2-en-1-ol, a New Class of Purine N,O-Nucleoside Analogues
Novel Purine N,O-Nucleoside Analogues
Paolo Quadrelli,* Mariella Mella, Serena Carosso, Bruna Bovio
Dipartimento di Chimica, Università degli Studi di Pavia, Viale Taramelli 12, 27100 Pavia, Italy
E-mail: paolo.quadrelli@unipv.it; Fax +39(0382)987323
Received: 25.01.2013; Accepted after revision: 15.03.2013
Applications of nitrosocarbonyl compounds in syntheses
Abstract: Sterically encumbered nitrosocarbonyl derivatives of
of biologically active molecules are well known.7 Al-
mesitylene or anthracene undergo ene reactions with 3-methylbut-
though nitrocarbonyl intermediates are powerful eno-
2-en-1-ol to give the corresponding the 5-hydroxyisoxazolidine ad-
ducts in fair yields. According to the proposed mechanism, these
philes,8 no applications of their ene reactions in syntheses
heterocycles are derived from the anti-Markovnikov orientation of of nucleosides have been reported. We found that nitroso-
the ene reaction. The isoxazolidin-5-yl acetate derivatives are syn-
thons of choice for the preparation of N,O-nucleoside analogues
containing purine rings by means of the Vorbrüggen protocol.
carbonyl intermediates undergo clean ene reactions with
trisubstituted olefins 8. Allylic hydrogens on the more-
congested side of the alkene are abstracted exclusively
(the cis-effect). The ene reaction of N-oxobenzamide (7P;
Key Words: nucleosides, ene reactions, heterocycles, cyclization
Ar = Ph) proceeds with a Markovnikov orientation,
whereas with the more-sterically encumbered 2,4,6-tri-
The synthesis of carbocyclic nucleosides1 through the methyl-N-oxobenzamide (7M; Ar = Mes), the
chemistry of nitrosocarbonyl intermediates has been ex- Markovnikov directing effect is eliminated and compara-
tensively investigated by many research groups.2 The gen- ble twix and lone abstractions are observed (Scheme 2).9
eration of nitrosocarbonyl intermediates 1 by mild
oxidation of nitrile oxides with tertiary amine N-oxides
Ph
C
N
OH
a R = H
b R = Me
c R = Et
offers an efficient alternative route to hetero-Diels–Alder
cycloadducts 2 with various substituents (Scheme 1).3 Cy-
cloadducts 2 are easily converted into 1,3-aminols 3,
which are useful synthons for the linear construction of
purine and pyrimidine nucleosides analogues 4.4 Some of
these nucleosides have been found to show moderate ac-
tivity against human herpes viruses (types 1 and 2) and
varicella viruses.5 Recently, anthryl derivatives have been
shown to be active against the human papilloma virus,
with no cellular toxicity at the doses tested.6
5
Cl
P Ar = Ph
M Ar = Mes
NMO, Et3N
CH2Cl2, r.t., 2 h
twix
CH2R
lone
8a–c
CH2R
CHOAr
HO
+
ArCONO
7P,M
N
R
twin
N
COAr
HO
NMO
CH2Cl2, r.t., 12 h
9P (%)
a 95
b 94
c 96
M
a 43
b 44
c 45
10P (%)
a
–
–
–
b
c
_
+
Mes
C N O
M
6
a
b
c
54
53
52
R
b
O
O
a
c
(
)
n
COR
N
N
Scheme 2
1,3-dipolar
cycloaddition
HDA
cycloaddition
O
1
2
In pursuing our researches on ene reactions, we extended
our studies to other trisubstituted olefins, particularly al-
lylic alcohols 8 (R = OH). We selected 3-methylbut-2-en-
1-ol (8d) as a model compound for our investigations of
the selectivity of the outcome of the ene reactions of nitro-
socarbonyl compounds bearing sterically demanding sub-
stituents. Unexpectedly, we obtained the isoxazolidines
11, derived from anti-Markovnikov addition to the alco-
hol 8. Isoxazolidines 11 are valuable synthons in the prep-
aration of new N,O-nucleoside pyrimidine analogues 12
through the Vorbrüggen protocol (Scheme 3).10
COR
detachment
HO
NH2
N–O bond
cleavage
a
c
b
3
linear
construction
of heterobases
HO
a
N
c
b
4
Scheme 1
To explore the reactivity of isoxazolidines 11, we extend-
ed the synthesis to some representative purine-type het-
erobases. 2,4,6-Trimethylbenzonitrile oxide (6) was
added to a stirred solution of N-methylmorpholine N-ox-
ide (1.1 equiv) in dichloromethane in the presence of an
SYNTHESIS 2013, 45, 1414–1420
Advanced online publication: 25.04.2013
0
0
3
9
-
7
8
8
1
1
4
3
7
-
2
1
0
X
DOI: 10.1055/s-0032-1316916; Art ID: SS-2013-Z0081-OP
© Georg Thieme Verlag Stuttgart · New York

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Novel Purine N,O-Nucleoside Analogues
1415
Ar
whereas the anti-Markovnikov route gives an enol inter-
mediate that readily converts into the corresponding alde-
hyde; this, in turn, cyclizes to give the required
isoxazolidine 11Md (Scheme 4).
O
AM
O
N
base
O
O
ArCO
OH ArCO
M
N
N
H
H
H
OH
The acetyl derivative 13, when prepared by standard
procedures11 in nearly quantitative yield, was found to be
identical to an authentic sample.10 The Vorbrüggen proto-
col can be applied to silylated heterobases or to commer-
cially available heterobases in the presence of a silylating
agent.12 We synthesized purine derivatives from commer-
cially available heterobases in the presence of trimethyl-
silyl N-(trimethylsilyl)ethanecarboximidate (BSA) as a
silylating agent and trimethylsilyl triflate as a promoter.13
The acetylated isoxazolidine 13 was added under nitrogen
at room temperature to a solution of the selected hetero-
base (2 equiv) and BSA (2 equiv) in dichloromethane, and
the solution became clear after boiling for two hours. The
mixture was then cooled to 0 °C on ice, trimethylsilyl tri-
flate was added, and the mixture was refluxed overnight
(Scheme 5). Purification by column chromatography gave
the desired compound 14a–c as white solids.
11
12 (48–99%)
O
N
NH2
N
Ar
Ph
Mes
M path AM path
R
0
25
52
a R = H
60
50
27
NH
b R = F
c R = Cl
d R = Br
e R = I
base =
Anthra
O
N
O
f R = Me
g
Scheme 3
excess (5 equiv) of 3-methylbut-2-en-1-ol (8d) at room
temperature. After stirring the mixture overnight and col-
umn chromatographic separation, we isolated the ene ad-
ducts 9Md and 11Md in 50% and 25% yield,
respectively.10 Adduct 9Md is formed by addition of the
nitrosocarbonyl 7M at the less-substituted carbon atom of
the allylic alcohol 8d by the Markovnikov pathway,
The nucleoside analogues 14a–c were isolated as racemic
mixtures in fair yields (32–42%), and their structures were
confirmed by analysis and spectroscopic studies. Each of
the reactions gave a single product, except in the case of
6-chloropurine, which gave an approximately 1:1 mixture
of the isomeric products 14a and 14a′ through coupling at
either the N7 or the N9 nitrogen atom of the purine ring.
This behavior of the 6-chloropurine is frequently dis-
played in coupling reactions that involve the nucleophilic
properties of this heterocyclic ring.14
Mes
7M
O
AM
O
OH
OH
MesCO
MesCO
OH
O
N
N
N
M
H
H
H
OH
enol intermediate
aldehyde
intermediate
8d
In the 1H NMR spectra of nucleosides 14a–c, signals cor-
responding to the N-(mesitylcarbonyl)isoxazolidine moi-
ety were easily detectable and they were found in the
expected ranges, as previously described.10 Substitution
of the acetate group with a heterocyclic ring in the 5-posi-
tion showed a smooth effect on the corresponding chemi-
cal shifts. The new representative signals correspond to
the purine moieties. The results obtained with 6-chloropu-
HO
O
O
MesCO
MesCO
OH
OAc
N
COMes
N
N
AcCl, Py
CH2Cl2, r.t., 24 h
CH2OH
9Md
13b (99%)
11Md (25%)
Scheme 4 (M = Markovnikov path, AM = anti-Markovnikov path)
Cl
N
N
N
N
N
N
N
O
O
MesCO
MesCO
N
N
N
Cl
14a (42%)
14a' (40%)
O
MesCO
OAc
purine-type heterobase
N
BSA, TMSOTf
CH2Cl2, 24 h
O
O
N
13
NH
N
NH
O
MesCO
N
N
NH2
O
MesCO
N
N
N
N
14b (40%)
14c (32%)
Scheme 5
© Georg Thieme Verlag Stuttgart · New York
Synthesis 2013, 45, 1414–1420

1416
P. Quadrelli et al.
PAPER
rine deserve special comment, because two isomeric prod- group is highly deshielded at δ = 10.25. In the hypoxan-
ucts, 14a and 14a′, were obtained in which the purine ring thine adduct 14c, the heterobase CH protons appear as sin-
is linked to the isoxazolidine unit at the N7 or N9 atom, glets at δ = 8.05 and 8.28, whereas the NH appears as a
respectively. The 1H NMR spectra confirmed the presence broad singlet at δ = 12.42.
of the 6-chloropurine ring in both products. In 14a, signals
for the N=CH singlets of the purine ring occur at δ = 8.60
and 8.73, whereas in 14a′, the same signals occur at δ =
Products containing fluorescent chromophores can be
useful as chemical probes for various biological targets,
and the field of imaging has received a great deal of atten-
8.78 and 8.91, respectively. However, the spectroscopic
tion in relation to its applications in achieving a better un-
1
pattern is very similar for the two regioisomers, and H–
derstanding of in vivo mechanisms.15 As a result, new
13C long-range correlation NMR experiments were unable
fluorophores and their applications are constantly being
to distinguish between the two isomers.14 A definitive
developed.16 Moreover, fluorescent polyaromatic groups
characterization was achieved by means of X-ray analysis
can be active through their ability to establish π–π stack-
of isomer 14a, in which the purine ring is linked to the
isoxazolidine moiety through the N7 atom (Figure 1; for
ing interactions with themselves17 or with DNA intercala-
tors;18 these mechanisms occur in various cases,
full details, see the Supporting Information). As a conse-
depending on the biological target.15b,19
quence, the structure of 14a′ was defined as shown in
We have previously reported an ene reaction of another
Scheme 5.
typical stable nitrile oxide,20 anthracene-9-carbonitrile ox-
ide, prepared from the anthracene-9-carbaldehyde oxime
by the procedure reported in literature.21 The solid nitrile
oxide was added in a portionwise manner to a stirred so-
lution of allylic alcohol 8d (5 equiv) in dichloromethane
containing N--methylmorpholine N-oxide (2 equiv), and
the mixture was stirred for 48 hours at room temperature.
The two expected ene products 9Ad and 11Ad (Ar = 9-
anthryl) were isolated in 28% and 53% yield, respectively
(Scheme 6). Compound 9Ad is formed by addition of the
nitrosocarbonyl 7A through the Markovnikov pathway,
whereas the isoxazolidine 11Ad is derived from the pre-
ferred anti-Markovnikov pathway. The synthesis was op-
timized, and crystal structures of both the ene adducts
were obtained. The structure of the isoxazolidine 11Ad
has already been reported,10 and X-ray analysis of ene
compound 9Ad confirmed the spectroscopic structural as-
signments (for the ORTEP view, see the Supporting Infor-
mation).
Figure 1 ORTEP plot of the compound 14a with atom labeling (el-
lipsoids at 25% probability). Hydrogen atoms are omitted for clarity.
The acetyl derivative 15 was prepared according to stan-
dard procedures11 in nearly quantitative yield, and it was
found to be identical to an authentic sample previously
prepared.10 The compound was subjected to a Vorbrüggen-
type reaction with adenine by means of the protocol de-
scribed above. When acetylated isoxazolidine 15 was add-
The guanine nucleoside 14b is characterized by a single
proton attributable to the guanine ring at δ = 7.88, whereas
that of the NH2 group is found at δ = 6.47 and the NH
Anthryl
O
AM
7A
O
M
OH
OH
AnthrylCO
AnthrylCO
OH
O
N
N
N
H
H
H
OH
8d
enol intermediate
aldehyde intermediate
HO
O
O
AnthrylCO
OH
AnthrylCO
OAc
N
N
COAnthryl
N
AcCl, Py
CH2Cl2, r.t., 24 h
CH2OH
9Ad
15 (99%)
11Ad (53%)
Scheme 6
Synthesis 2013, 45, 1414–1420
© Georg Thieme Verlag Stuttgart · New York

PAPER
Novel Purine N,O-Nucleoside Analogues
1417
ed to a solution of adenine (2 equiv) and BSA (2 equiv) in of the allylic alcohol 8d. From the point of view of the
dichloromethane under nitrogen at room temperature, the Markovnikov approach, it seems quite clear that the hy-
solution became clear after boiling for two hours. The droxy group does not play any role in directing the addi-
mixture was then cooled to 0 °C on ice, trimethylsilyl tri- tion of the nitrosocarbonyl intermediates in the transition
flate was added, and the mixture was refluxed overnight state (M path, top view). The steric clashes that operate in
(Scheme 7). Simple column chromatographic purification the Markovnikov case shift the addition to the anti-
gave the desired compound 16 as a white solid in 36% Markovnikov path that leads to enol formation and subse-
yield.
quent cyclization to an isoxazolidine structure.10
H3C
H3C
NH2
O
AnthrylCO
OAc
N
N
N
N
purine-type heterobase
BSA, TMSOTf
CH2Cl2, r.t., 24 h
CH3
CH3
O
AnthrylCO
N
CH3
H
CH3
H3C
O
O
N
CH3
H
15
16 (36%)
N
N
O
HOH2C
O
CH3
H3C
CH2OH
Scheme 7
M path (top view)
AM path (top view)
Figure 3 (M = Markovnikov, AM = anti-Markovnikov)
Adduct 16 is a high-melting compound (mp > 205 °C) and
its IR spectrum showed the presence of an NH2 group
1
(νNH2 = 3375, 3321 cm–1). Its H NMR spectrum con-
The structures of the 5-hydroxyisozaxolidines 11M and
11Ad resemble those that are obtained through addition of
nitrones to vinyl ether derivatives, and they constitute use-
ful synthons for the preparation of N,O-nucleoside ana-
logues. By adapting the Vorbrüggen protocol,22 as well as
other known procedures for similar compounds,23 we
have reported syntheses of a selection of uracil deriva-
tives, which we obtained in good-to-excellent yields.10
However, the yields of the purine adducts were somewhat
lower than expected on the basis of previous observations,
and were generally below 50%. In view of planned future
applications of this synthetic method, it is important to
elucidate the limitations of the protocol beyond the scope
of the current research.
firmed the presence of the adenine ring, the N=CH sin-
glets of the purine ring being found at δ = 8.22 and 8.46.
These results show that isoxazolidine substrates, properly
derivatized as their acetates, can be used in syntheses of a
variety of N,O-nucleoside analogues. The novel isoxa-
zolidine synthons can be easily prepared through ene re-
actions of nitrosocarbonyl intermediates bearing sterically
encumbered aromatic substituents that drive ene additions
to allylic alcohols along the anti-Markovnikov path (Fig-
ure 2). Because the addition of the nitrosocarbonyl moiety
occurs on the more-substituted carbon atom of the allylic
alcohol 8d, the primary adducts are enolic forms that
evolve into a nonisolable aldehydes that, in turn, cyclize
to form the corresponding hemiacetals 11 (Scheme 4 and
6) as final and isolable compounds.
One possible explanation of the low yields that were ob-
tained when purine heterobases were used comes from the
proposed mechanism for the Vorbrüggen-type coupling
of these heterocycles with the isoxazolidine intermediate
in comparison to the case when uracil is used. As soon as
uracil is silylated by the BSA, the triflate anion triggers
nucleophilic attack on the stabilized isoxazolidine cation
generated from compounds 11 by loss of acetate, which is
a good leaving group. In the case of 6-chloropurine, BSA
silylation is somewhat less efficient and more complex,
because many silylated species can be present in the reac-
tion mixture.
H
H
H
H
H
Ar
7P
7M
O
O
AM
N
M
7A
H
H
H
H
OH
The coupling reaction might be more difficult when other
functional groups are present on the purine rings, as in the
cases of guanine and adenine. An amino group can be eas-
ily involved in other tautomeric equilibria or silylation
processes. Aware of these facts, we tried to modify our
coupling protocol by increasing or decreasing the propor-
tions of BSA or the triflate, but these experiments proved
unsuccessful. Changing the reaction temperature or time
did not greatly affect the reaction yields, and the reported
conditions are, indeed, optimal for the Vorbrüggen glyco-
sidation reactions of our isoxazolidine derivatives.
yields of AM
products (11)
0
25
52
7P
7M 50
7A 27
60
yields of M
products (9)
Figure 2
We have recently reported that crowding around the nitro-
so group slows the Markovnikov approach of 2,4,6-tri-
methylbenzonitrile oxide (7M) to a trisubstituted ethene
and compensates somewhat for the Markovnikov elec-
tronic bias. Figure 3 shows the same features in the case
© Georg Thieme Verlag Stuttgart · New York
Synthesis 2013, 45, 1414–1420

1418
P. Quadrelli et al.
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Ene Reactions of Nitrosocarbonyls 7M and 7A with 3-Methyl-
but-2-en-1-ol (8d); General Procedure
NMO (1.8 g, 1.3 equiv) was added an ice-cooled stirred soln of enol
8d (99%; 5 equiv) in CH2Cl2 (200 mL). A soln of 2,4,6-trimethyl-
benzonitrile oxide (2.0 g, 12.4 mmol) or anthracene-9-carbonitrile
Future developments in synthetic methods for the prepa-
ration of N,O-nucleoside analogues are currently being
pursued by varying both the allylic alcohol substrate and
the substituents on the nitrosocarbonyl compound to per-
mit the introduction of other functionalities onto the isox- oxide (2.0 g, 9 mmol) in CH2Cl2 (100 mL) was added dropwise, and
the mixture was stirred at r.t. for 24 h. After dilution with an equal
volume of CH2Cl2, the organic phase was washed with H2O (2 × 50
azolidine ring.
The Vorbrüggen protocol has been shown to provide a ro-
bust method for the attachment of heterobases to activated
isoxazolidine substrates. We believe that its application,
with minor experimental adjustments, might be successful
with other heterocyclic rings in various specific biological
targets.
mL), dried (Na2SO4), filtered, and concentrated. The residue was
purified by column chromatography to give the corresponding ene
adduct.10
2-Acyl-3,3-dimethyl-1,2-isoxazolidin-5-yl Acetates 13 and 15;
General Procedure
AcCl (2.2 equiv), DMAP (0.34 equiv), and Et3N (2.2 equiv) were
added to an ice-cooled stirred soln of isoxazolidinol 11 (4.78 mmol)
in anhyd CH2Cl2 (50 mL), and the mixture was stirred at r.t. for 24
h. The mixture was then diluted with CH2Cl2 (50 mL), and the or-
ganic phase was washed with sat. aq NaHCO3 (25 mL), dried
(Na2SO4), and concentrated to give a solid product.10
We are currently planning biological assays to test the in-
hibitory activity of our products against some viruses and
to elucidate the apoptotic behavior of our N,O-nucleoside
analogues. On request, the purine derivatives 14a–c and
16 were sent to the Southern Research Institute (Birming-
ham, AL) as part of the Tuberculosis Antimicrobial Ac-
quisition and Coordinating Facility program and they
were tested against the Mycobacterium tuberculosis
H37Rv in BACTEC 12B medium by using the microplate
Alamar Blue assay.24 The minimum inhibition concentra-
tions were found greater than 6.25 μg/mL for all the com-
pounds, with inhibition indices (%Inh) in the range 22–25.
Nucleoside Derivatives 14; General Procedure
A soln of the appropriate heterobase (2 equiv) and TMS-
N=C(Me)OTMS (2 equiv) in anhyd CH2Cl2 (50 mL) was refluxed
under N2 for 15–20 min until it became clear. It was then cooled to
r.t. and a soln of isoxazolidine 13 (0.20 g, 0.65 mmol) in CH2Cl2 (10
mL) was added dropwise. The mixture was cooled to 0 °C and
TMSOTf (0.12 mL, 1 equiv) was added. The resulting mixture was
refluxed and stirred overnight, and then the reaction was quenched
with sat. aq NaHCO3 (pH 7). The mixture was diluted with an equal
volume of CH2Cl2, washed with H2O (2 × 20 mL), dried (Na2SO4),
filtered, and concentrated to give a residue that was purified by col-
umn chromatography.
In conclusion, we have shown that sterically encumbered
nitrosocarbonyl derivatives of mesitylene and anthracene
undergo ene reactions with 3-methylbut-2-en-1-ol to give
the corresponding 5-hydroxyisoxazolidine adducts in
fair yields. These heterocycles derive from the anti-
Markovnikov orientation of the ene reaction, according to
the proposed mechanism. The isoxazolidin-5-yl acetates
are synthons of choice for the preparation, by means of the
Vorbrüggen protocol, of N,O-nucleoside analogues con-
taining purine rings.
6-Chloro-9-[2-(mesitylcarbonyl)-3,3-dimethylisoxazolidin-5-
yl]-9H-purine (14a)
Pale-yellow solid; yield: 0.11 g (42%); mp 192–194 °C (EtOAc).
IR (Nujol): 1630, 1589, 1537, 1210, 1078, 979, 722 cm–1.
1H NMR (300 MHz, acetone-d6): δ = 1.79 and 1.92 (s, 6 H, 2CH3),
1.99, 2.17, and 2.26 (s, 9 H, 3CH3–Ar), 3.17 (dd, J = 14 and 8 Hz,
1 H, CH2), 3.52 (dd, J = 14 and 5 Hz, 1 H, CH2), 6.65 (s, 1 H, Ph),
6.70 (dd, J = 7 and 4 Hz, 1 H, CH), 6.78 (s, 1 H, Ph), 8.60 and 8.73
(s, 2 H, CH purine ring).
13C NMR (75 MHz, acetone-d6): δ = 19.1, 19.5, 21.4, 25.8, 26.8,
48.5, 64.2, 83.6, 121.3, 128.8, 128.9, 133.3, 134.1, 134.9, 135.3,
139.0, 151.6, 153.1, 168.2.
All melting points are uncorrected. Elemental analyses were per-
formed on a Carlo Erba 1106 elemental analyzer, available in our
department. IR spectra (Nujol mulls for solid compounds, neat for
1
oils) were recorded on a PerkinElmer RX-1 FTIR instrument. H
Anal. Calcd for C20H22ClN5O2 (399.87): C, 60.07; H, 5.55; N,
17.51. Found: C, 59.89; H, 5.51; N, 17.49.
and 13C NMR spectra were recorded on a Bruker AVANCE 300
spectrometer in the specified deuterated solvents. Chemical shifts
are expressed in ppm (δ) relative to TMS as internal standard. Col-
umn chromatography and TLC were performed on silica gel 60
(0.063–0.200 mm) (Merck) with cyclohexane–EtOAc (9:1 to 5:5)
as the eluent. Medium-pressure LC was performed on a Biotage
FMP apparatus equipped with KP-SIL columns; the eluent was cy-
clohexane–EtOAc (9:1 to 7:3). The identity of samples from differ-
ent experiments was confirmed by mixed-mp studies and by
superimposition of IR spectra. X-ray crystallographic analysis was
conducted at the Centro Grandi Strumenti of the University of Pavia
by using an Enraf Nonius CAD4 diffractometer with graphite-
monochromated Mo Kα radiation (see Supporting Information for
details).
6-Chloro-7-[2-(mesitylcarbonyl)-3,3-dimethylisoxazolidin-5-
yl]-7H-purine (14a′)
Pale-yellow solid; yield: 0.10 g (40%); mp 217–219 °C (EtOAc–
EtOH).
IR (Nujol): 1631, 1588, 1537, 1211, 1079, 979, 722 cm–1.
1H NMR (300 MHz, acetone-d6): δ = 1.62 and 1.78 (s, 6 H, 2CH3),
1.96, 2.18, and 2.26 (s, 9 H, 3CH3–Ar), 3.24 (dd, J = 14 and 7 Hz,
1 H, CH2), 3.45 (dd, J = 14 and 4 Hz, 1 H, CH2), 6.44 and 6.79 (s, 2
H, Ph), 6.90 (dd, J = 7 and 4 Hz, 1 H, CH), 8.78 and 8.91 (s, 2 H,
CH purine ring).
13C NMR (75 MHz, acetone-d6): δ = 18.6, 19.5, 21.4, 26.6, 26.9,
48.5, 63.6, 85.2, 122.5, 128.4, 128.8, 133.7, 134.9, 139.0, 148.0,
153.4, 160.1, 164.2.
3-Methylbut-2-en-1-ol (8d, 99%) was purchased from chemical
suppliers. 2,4,6-Trimethylbenzonitrile oxide (6) was prepared by
oxidation of 2,4,6-trimethylbenzaldoxime with bromine.22 Anthra-
cene-9-carbaldehyde oxime was purchased from chemical suppli-
ers.
Anal. Calcd for C20H22ClN5O2 (399.87): C, 60.07; H, 5.55; N,
17.51. Found: C, 60.10; H, 5.51; N, 17.49.
2-Amino-9-[2-(mesitylcarbonyl)-3,3-dimethylisoxazolidin-5-
yl]-1,9-dihydro-6H-purin-6-one (14b)
Pale-yellow solid; yield: 0.10 g (40%); mp 213–217 °C (EtOAc).
Synthesis 2013, 45, 1414–1420
© Georg Thieme Verlag Stuttgart · New York

PAPER
Novel Purine N,O-Nucleoside Analogues
1419
IR (Nujol): 3313, 3150, 1691, 1626, 1529, 1223, 1094, 973, 778
cm–1.
to be ‘Active’ in terms of antitubercular activity. For further details,
see http://www.nih.gov.
1H NMR (300 MHz, DMSO): δ = 1.63 and 1.86 (s, 6 H, 2CH3), 1.93
and 2.16 (s, 9 H, 3CH3-Ar), 2.89 and 3.08 (dd, J = 14 and 6 Hz, 2
H, CH2), 6.20 (t, J = 6 Hz, 1 H, CH), 6.47 (br s, 2 H, exch. D2O),
NH2], 6.70 and 6.78 (s, 2 H, Ph), 7.88 (s, 1 H, CH), 10.25 (br s, 1 H,
NH, exch. D2O).
13C NMR (75 MHz, DMSO): δ = 18.0, 18.5, 20.6, 24.9, 25.8, 46.3,
62.7, 80.3, 116.8, 127.5, 132.7, 133.0, 133.8, 135.1, 137.3, 151.2,
153.7, 156.6, 165.6.
X-ray Crystallography
The crystal data, data collection, and structural refinements for
compounds 9Ad and 14a are summarized in Table S3 of the Sup-
porting Information. The structures were solved by direct methods.
Nonhydrogen atoms were refined anisotropically, and hydrogen at-
oms, located from the difference Fourier synthesis, were refined
isotropically.25,26
Crystallographic data for compounds 9Ad (CCDC 921636) and 14a
(CCDC 921635) have been deposited and can be obtained free of
charge from the Cambridge Crystallographic Data Centre, 12 Union
Road, Cambridge CB2 1EZ, UK; Fax: +44(1223)336033; E-mail:
deposit@ccdc.cam.ac.uk; Web site: www.ccdc.cam.ac.uk/conts/re-
trieving.html.
Anal. Calcd for C20H24N6O3 (396.44): C, 60.59; H, 6.10; N, 21.20.
Found: C, 60.53; H, 5.99; N, 20.99.
9-[2-(Mesitylcarbonyl)-3,3-dimethylisoxazolidin-5-yl]-1,9-di-
hydro-6H-purin-6-one (14c)
Pale-yellow solid; yield: 0.08 g (32%); mp 175–177 °C (MeOH).
IR (Nujol): 3425, 1686, 1635, 1545, 1210, 1090, 953, 777 cm–1.
Acknowledgement
1H NMR (300 MHz, DMSO): δ = 1.65 and 1.84 (s, 6 H, 2CH3),
1.87, 2.15, and 2.17 (s, 9 H, 3CH3-Ar), 2.94 and 3.22 (dd, J = 14 and
7 Hz, 2 H, CH2), 6.45 (t, J = 7 Hz, 1 H, CH), 6.66 and 6.77 (s, 2 H,
Ph), 8.05 and 8.28 (s, 2 H, CH hypox ring), 12.42 (br s, 1 H, NH).
13C NMR (75 MHz, DMSO): δ = 17.9, 18.5, 20.6, 24.9, 25.8, 46.4,
62.7, 81.2, 114.7, 127.5, 132.6, 133.1, 137.3, 138.5, 144.9, 148.3,
156.4, 165.7.
Financial support by the University of Pavia and MIUR (PRIN
2011, CUP: F11J12000210001) is gratefully acknowledged. The
authors wish to thank Steroid S.p.A. for a research grant. Thanks are
due Dr. Massimo Boiocchi (CGS) for performing the X-ray data
collection and analysis.
Anal. Calcd for C20H23N5O3 (381.42): C, 62.98; H, 6.08; N, 18.36.
Found: C, 63.10; H, 6.11; N, 18.40.
Supporting Information for this article is available online at
http://www.thieme-connect.com/ejournals/toc/synthesis. Included
are crystal structure analyses for compounds 9Ad and 14a and co-
9-[2-(9-Anthrylcarbonyl)-3,3-dimethylisoxazolidin-5-yl]-9H-
purin-6-amine (16)
1
pies of the H and 13C NMR spectra of all the synthetized com-
pounds.SupngIopif
omitrSatnuIpofomnipgrairtt
n
A soln of adenine (2 equiv) and TMSN=C(Me)OTMS (2 equiv) in
anhyd CH2Cl2 (50 mL) was refluxed under N2 for 15–20 min until
it became clear. The mixture was cooled to r.t. and a soln of isoxa-
zolidine 15 (0.20 g, 0.65 mmol) in CH2Cl2 (10 mL) was added drop-
wise. The resulting soln was cooled to 0 °C and TMSOTf (0.12 mL,
1 equiv) was added. The mixture was refluxed with stirring over-
night and then the reaction was quenched with sat. aq NaHCO3
(pH 7). The resulting mixture was diluted with an equal volume of
CH2Cl2, washed with H2O (2 × 20 mL), dried (Na2SO4), filtered,
and concentrated. The residue was purified by column chromatog-
raphy to give a yellowish solid; yield: 0.10 g (36%); mp > 205 °C
(EtOH).
References
(1) (a) Bodnar, B. S.; Miller, M. J. Angew. Chem. Int. Ed. 2011,
50, 5630. (b) Romeo, G.; Chiacchio, U.; Corsaro, A.;
Merino, P. Chem. Rev. 2010, 110, 3337. (c) Vogt, P. F.;
Miller, M. J. Tetrahedron 1998, 54, 1317. (d) Boger, D. L.;
Weinreb, S. M. Hetero Diels–Alder Methodology in Organic
Synthesis; Academic Press: San Diego, 1987. (e) Herdewijn,
P. Drug Discovery Today 1997, 2, 235. (f) Wang, P.; Hong,
J. H.; Cooperwood, J. S.; Chu, C. K. Antiviral Res. 1998, 40,
19. (g) De Clercq, E. Biochim. Biophys. Acta 2002, 1587,
258. (h) De Clercq, E. Med. Res. Rev. 2002, 22, 531.
(i) Desgranges, C.; Razaka, G.; Rabaud, M.; Bricaud, H.;
Balzarini, J.; De Clercq, E. Biochem. Pharmacol. 1983, 32,
3583.
(2) For the generation of nitrosocarbonyl compounds, see:
(a) Surman, M. D.; Mulvihill, M. J.; Miller, M. J. J. Org.
Chem. 2002, 67, 4115. (b) Surman, M. D.; Mulvihill, M. J.;
Miller, M. J. Tetrahedron Lett. 2002, 43, 1131. (c) Ware, R.
W. Jr.; Day, C. S.; King, S. B. J. Org. Chem. 2002, 67, 6174.
(d) Chow, C. P.; Shea, K. J.; Sparks, S. M. Org. Lett. 2002,
4, 2637. (e) Momiyama, N.; Yamamoto, H. Org. Lett. 2002,
4, 3579. (f) Sato, T.; Aoyagi, S.; Kibayashi, C. Org. Lett.
2003, 5, 3839. (g) Jiang, M. X.-W.; Warshakoon, N. C.;
Miller, M. J. J. Org. Chem. 2005, 70, 2824. (h) Bodnar, B.
S.; Miller, M. J. J. Org. Chem. 2007, 72, 3929. (i) Iwasa, S.;
Tajima, K.; Tsushima, S.; Nishiyama, H. Tetrahedron Lett.
2001, 42, 5897. (j) Flower, K. R.; Lightfoot, A. P.; Wan, H.;
Whiting, A. J. Chem. Soc., Perkin Trans. 1 2002, 2058.
(k) Iwasa, S.; Fakhruddin, A.; Tsukamoto, Y.; Kameyama,
M.; Nishiyama, H. Tetrahedron Lett. 2002, 43, 6159.
(l) Pulacchini, S.; Sibbons, K. F.; Shastri, K.; Motevalli, M.;
Watkinson, M.; Wan, H.; Whiting, A.; Lightfoot, A. P.
Dalton Trans. 2003, 2043. (m) Adamo, M. F. A.; Bruschi, S.
J. Org. Chem. 2007, 72, 2666. (n) Adam, W.; Bottke, N.;
IR (Nujol): 3375, 3321, 1664, 1596, 1560, 1354, 975, 885, 730
cm–1.
1H NMR (300 MHz, CDCl3): δ = 2.05 (s, 3 H, CH3), 2.06 (s, 3 H,
CH3), 3.00 (dd, J = 14 and 7 Hz, 1 H, H–CH), 3.47 (dd, J = 14 and
3 Hz, 1 H, HC–H), 5.62 (br s, 2 H, NH2), 6.10 (dd, J = 7 and 3 Hz,
1 H, O–CH–NH), 7.45 (m, 4 H, arom.), 7.86 (d, J = 9 Hz, 1 H,
arom.), 8.01 (m, 4 H, arom.), 8.22 (s, 1 H, CH=N), 8.46 (s, 1 H,
CH=N).
13C NMR (75 MHz, CDCl3): δ = 26.3, 30.8, 40.7, 63.1, 82.5, 119.9,
124.0, 124.4, 125.3, 125.4, 126.5, 127.1, 127.3, 127.5, 128.1, 128.6,
130.9, 137.8, 152.9, 155.1, 165.8.
Anal. Calcd. for C25H22N6O2 (438.47): C, 68.48; H, 5.06; N, 19.17.
Found: C, 68.49; H, 5.05; N, 19.20.
Primary Screen (Dose Response): Determination of 90% Inhib-
itory Concentration
The initial screen was conducted against Mycobacterium tuberculo-
sis H37Rv (ATCC 27294) in BACTEC 12B medium using the mi-
croplate Alamar Blue assay. The compounds were tested in ten
twofold dilutions, typically from 100 μg/mL to 0.19 μg/mL. The
90% inhibitory concentration (IC90) is defined as the concentration
that produces a 90% reduction in fluorescence relative to controls.
This value is determined from the dose–response curve by using a
curve-fitting program. An IC90 value of ≤10 μg/mL is considered
© Georg Thieme Verlag Stuttgart · New York
Synthesis 2013, 45, 1414–1420

1420
P. Quadrelli et al.
PAPER
Krebs, O.; Saha-Möller, C. R. Eur. J. Org. Chem. 1999,
1963.
(14) Quadrelli, P.; Mella, M.; Vazquez Martinez, N.; Piccanello,
A. ARKIVOC 2009, (ii), 183.
(3) (a) Quadrelli, P.; Gamba Invernizzi, A.; Caramella, P.
Tetrahedron Lett. 1996, 37, 1909. (b) Quadrelli, P.; Mella,
M.; Gamba Invernizzi, A.; Caramella, P. Tetrahedron 1999,
55, 10497.
(15) (a) Shin, D.; Sinkeldam, R. W.; Tor, Y. J. Am. Chem. Soc.
2011, 133, 14912. (b) Hernández, A. R.; Kool, E. T. Org.
Lett. 2011, 13, 676. (c) Wilson, J. N.; Kool, E. T. Org.
Biomol. Chem. 2006, 4, 4265.
(4) (a) Quadrelli, P.; Mella, M.; Paganoni, P.; Caramella, P. Eur.
J. Org. Chem. 2000, 2613. (b) Quadrelli, P.; Fassardi, V.;
Cardarelli, A.; Caramella, P. Eur. J. Org. Chem. 2002, 2058.
(c) Quadrelli, P.; Mella, M.; Carosso, S.; Bovio, B.;
Caramella, P. Eur. J. Org. Chem. 2007, 6003. (d) Quadrelli,
P.; Mella, M.; Assanelli, G.; Piccanello, A. Tetrahedron
2008, 64, 7312.
(5) (a) Quadrelli, P.; Scrocchi, R.; Caramella, P.; Rescifina, A.;
Piperno, A. Tetrahedron 2004, 60, 3643. (b) Quadrelli, P.;
Piccanello, A.; Vazquez Martinez, N.; Bovio, B.; Mella, M.;
Caramella, P. Tetrahedron 2006, 62, 7370. (c) Savion, M.;
Memeo, M. G.; Bovio, B.; Grazioso, G.; Legnani, L.;
Quadrelli, P. Tetrahedron 2012, 68, 1845.
(16) Okamoto, A.; Tainaka, K.; Nishiza, K.; Saito, I. J. Am.
Chem. Soc. 2005, 127, 13128.
(17) Sekine, M.; Oeda, Y.; Iijima, Y.; Taguchi, H.; Ohkubo, A.;
Seio, K. Org. Biomol. Chem. 2011, 9, 210.
(18) Bair, K. W.; Andrews, C. W.; Tuttle, R. L.; Knick, V. C.;
Cory, M.; McKee, D. D. J. Med. Chem. 1991, 34, 1983.
(19) (a) Chiacchio, U.; Saita, M. G.; Crispino, L.; Gumina, G.;
Mangiafico, S.; Pistarà, V.; Romeo, G.; Piperno, A.; De
Clercq, E. Tetrahedron 2006, 62, 1171. (b) Leonard, N. J.;
Sprecker, M. A.; Morrice, A. G. J. Am. Chem. Soc. 1976, 98,
3987. (c) Charubala, R.; Maurinsh, J.; Rösler, A.; Melguizo,
M.; Jungmann, O.; Gottlieb, M.; Lehbauer, J.; Hawkins, M.;
Pfleiderer, W. Nucleosides Nucleotides 1997, 16, 1369.
(20) (a) Caramella, P.; Grünanger, P. In 1,3-Dipolar
Cycloaddition Chemistry, Vol. 1; Padwa, A., Ed.; Wiley:
New York, 1984, 291. (b) Grundman, C.; Datta, S. K. J. Org.
Chem. 1969, 34, 2016; and references cited therein.
(21) (a) Grundman, C.; Grünanger, P. The Nitrile Oxides:
Versatile Tools of Theoretical and Preparative Chemistry;
Springer: Heidelberg, 1971. (b) Grundmann, C.; Dean, J. M.
J. Org. Chem. 1965, 30, 2809.
(6) Moggio, Y.; Legnani, L.; Bovio, B.; Memeo, M. G.;
Quadrelli, P. Tetrahedron 2012, 68, 1384.
(7) (a) Miller M. J.; In Proceedings of the International
Symposium on Advances in Synthetic, Combinatorial and
Medicinal Chemistry Held in Moscow, May 5–8, 2004; LD
Organisation: Louvain-la-Neuve, 2004; T14 and P140.
(b) Kitade, Y.; Kojima, H.; Zulfiqur, F.; Kim, H.-S.; Wataya,
Y. Bioorg. Med. Chem. Lett. 2003, 13, 3963. (c) Ramesh, N.
G.; Klunder, A. J. H.; Zwanenburg, B. J. Org. Chem. 1999,
64, 3635. (d) Wu, J.; Schneller, S. W.; Seley, K. L.; De
Clerq, E. Heterocycles 1998, 47, 757. (e) Siddiqi, S. M.;
Raissian, M.; Schneller, S. W.; Ikeda, S.; Snoeck, R.;
Andrei, G.; Balzarini, J.; De Clerq, E. Bioorg. Med. Chem.
Lett. 1993, 3, 663. (f) Da Silva, A. D.; Coimbra, E. S.;
Fourrey, J. L.; Machado, A. S.; Robert-Géro, M.
Tetrahedron Lett. 1993, 34, 6745. (g) Koga, M.; Schneller,
S. W. Tetrahedron Lett. 1990, 31, 5861.
(8) (a) Quadrelli, P.; Mella, M.; Caramella, P. Tetrahedron Lett.
1998, 39, 3233. (b) Quadrelli, P.; Mella, M.; Caramella, P.
Tetrahedron Lett. 1999, 40, 797.
(9) (a) Quadrelli, P.; Mella, M.; Piccanello, A.; Romano, S.;
Caramella, P. J. Org. Chem. 2007, 72, 1807. (b) Quadrelli,
P.; Romano, S.; Piccanello, A.; Caramella, P. J. Org. Chem.
2009, 74, 2301. (c) Adam, W.; Krebs, O. Chem. Rev. 2003,
103, 4131.
(22) (a) Vorbrüggen, H.; Krolikiewicz, K.; Bennua-Skalmowski,
B. US 5750676, 1998. (b) Vorbrüggen, H.; Krolikiewicz,
K.; Bennua, B. Chem. Ber. 1981, 114, 1234. (c) Vorbrüggen,
H.; Höfle, G. Chem. Ber. 1981, 114, 1256. (d) Vorbrüggen,
H.; Bennua, B. Chem. Ber. 1981, 114, 1279.
(23) (a) Pistarà, V.; Corsaro, A.; Chiacchio, M. A.; Greco, G.;
Quadrelli, P. ARKIVOC 2011, (vi), 270. (b) Chiacchio, U.;
Genovese, F.; Iannazzo, D.; Piperno, A.; Quadrelli, P.;
Corsaro, A.; Romeo, R.; Valveri, V.; Mastino, A. Bioorg.
Med. Chem. 2004, 12, 3903. (c) Chiacchio, U.; Genovese,
F.; Iannazzo, D.; Librando, V.; Merino, P.; Rescifina, A.;
Romeo, R.; Propopio, A.; Romeo, G. Tetrahedron 2004, 60,
441. (d) Saita, M. G.; Chiacchio, U.; Iannazzo, D.; Merino,
P.; Piperno, A.; Previtera, T.; Rescifina, A.; Romeo, G.;
Romeo, R. Nucleosides, Nucleotides Nucleic Acids 2003, 22,
739. (e) Chiacchio, U.; Corsaro, A.; Iannazzo, D.; Piperno,
A.; Pistarà, V.; Rescifina, A.; Romeo, A.; Valveri, V.;
Mastino, A.; Romeo, G. J. Med. Chem. 2003, 46, 3696.
(f) Chiacchio, U.; Corsaro, A.; Iannazzo, D.; Piperno, A.;
Pistarà, V.; Procopio, A.; Rescifina, A.; Romeo, G.; Romeo,
R.; Siciliano, M. C. R.; Valveri, E. ARKIVOC 2002, (ix),
159. (g) Gotkowska, J.; Balzarini, J.; Piotrowska, D. G.
Tetrahedron Lett. 2012, 53, 7097.
(10) Quadrelli, P.; Mella, M.; Carosso, S.; Bovio, B. J. Org.
Chem. 2013, 78, 516.
(11) For acetylation of isoxazolidin-5-ols, see: (a) Xiang, Y.;
Gong, Y.; Zhao, K. Tetrahedron Lett. 1996, 37, 4877.
(b) Zhang, X.; Qing, F.-L.; Yu, Y. J. Org. Chem. 2000, 65,
7075. (c) Zhang, X.; Xia, H.; Dong, X.; Jin, J.; Meng, W.-D.;
Qing, F.-L. J. Org. Chem. 2003, 68, 9026.
(12) Yoshimura, Y.; Kitano, K.; Yamada, K.; Satoh, H.;
Watanabe, M.; Miura, S.; Sakata, S.; Sasaki, T.; Matsuda, A.
J. Org. Chem. 1997, 62, 3140.
(24) Collins, L. A.; Franzblau, S. G. Antimicrob. Agents
Chemother. 1997, 41, 1004.
(25) North, A. C. T.; Phillips, D. C.; Mathews, F. S. Acta
Crystallogr. Sect. A 1968, 24, 351.
(13) Caddell, J. M.; Chapman, A. M.; Cooley, B. E.; Downey, B.
P.; LeBlanc, M. P.; Jackson, M. M.; O’Connell, T. M.;
Phung, H.-M.; Roper, T. D.; Xie, S. J. Org. Chem. 2004, 69,
3212.
(26) (a) Sheldrick, G. M. SHELXL–93: Programs for Crystal
Structure Analysis; Institüt für Anorganische Chemie der
Universität Göttingen: Germany, 1998. (b) Johnson, C. K.
ORTEP Report ORNL-3793; Oak Ridge National
Laboratory: USA, 1965.
Synthesis 2013, 45, 1414–1420
© Georg Thieme Verlag Stuttgart · New York