A one pot, two-step synthesis of 5-arylpyrrolo[2,3-d]pyrimidines and screening of their preliminary antibacterial properties

Article abstract of DOI:10.1016/j.bmcl.2015.12.047

A one pot two step methodology for the synthesis of ten derivatives of 5-arylpyrrolo[2,3-d]pyrimidine has been reported. The methodology exploits the strong reducing nature of alkaline Na₂S₂O₄solution coupled with favorabi

Full text of DOI:10.1016/j.bmcl.2015.12.047

Accepted Manuscript
A one pot, two-step synthesis of 5-arylpyrrolo[2,3-d]pyrimidines and screening
of their preliminary antibacterial properties
Lakhinath Saikia, P. Roudragouda, Ashim J. Thakur
PII:
S0960-894X(15)30360-7
DOI:
http://dx.doi.org/10.1016/j.bmcl.2015.12.047
BMCL 23408
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Bioorganic & Medicinal Chemistry Letters
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15 December 2015
Please cite this article as: Saikia, L., Roudragouda, P., Thakur, A.J., A one pot, two-step synthesis of 5-
arylpyrrolo[2,3-d]pyrimidines and screening of their preliminary antibacterial properties, Bioorganic & Medicinal
Chemistry Letters (2015), doi: http://dx.doi.org/10.1016/j.bmcl.2015.12.047
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arylpyrrolo[2,3-d]pyrimidines and screening
of their preliminary antibacterial properties
Lakhinath Saikia, P. Roudragouda and Ashim J. Thakur

Bioorganic & Medicinal Chemistry Letters
A one pot, two-step synthesis of 5-arylpyrrolo[2,3-d]pyrimidines and screening of
their preliminary antibacterial properties
Lakhinath Saikia,a,b P. Roudragoudac and Ashim J. Thakura
aDepartment of Chemical Sciences, Tezpur University (A Central University), Tezpur, Napaam 784028, Assam, India
bDepartment of Chemistry, Rajiv Gandhi University (A. Central University), Rono-Hills, Doimukh 791112, Arunachal Pradesh, India
cDepartment of Pharmaceutical Technology,Defence Research Laboratory, Tezpur, Post Bag no. 2, Solmara 784001, Assam, India
ARTICLE INFO
ABSTRACT
Article history:
Received
Revised
Accepted
Available online
A one pot two step methodology for the synthesis of ten derivatives of 5-arylpyrrolo[2,3-
d]pyrimidine has been reported. The methodology exploits the strong reducing nature of alkaline
Na2S2O4 solution coupled with favorability of Michael type addition reaction in alkaline
medium. The methodology demands attraction as it is non-catalytic, quite general for wide range
of nitrostyrenes and possesses comprehensive advantages over most of the earlier methods in
terms of reaction time as well as yield. The methodology enjoys additional advantage of
utilizing cheaper and easily available chemicals as reagent for the purpose.Some of the
synthesized compounds are found to possess remarkable activity againstsome of the tested
bacterial strains.
Keywords:
Pyrrolo[2,3-d]pyrimidine
Michael type addition
Na2S2O4
2009 Elsevier Ltd. All rights reserved.
NaOH
Reductive cyclisation
Among various fused heterocycles, pyrrolo[2,3-d]pyrimidines
has been considered as one of the most important classes of
organic compounds. Various compounds of this family have been
designed, synthesized and screened towards various biological
activities. A plentiful of pyrrolo[2,3-d]pyrimidines have been
established to possess diverse biological activities like antifolate,1
antitumour,2 antibacterial,3 antiviral,4 receptor tyrosine kinase
(RTK) inhibitor,5 firefly luciferase inhibitor,6 microtubule
inhibitor,7 affinity against -adrenosine receptor (-AR)8 etc.
Alimta (1, Fig. 1), a 5-substituted pyrrolo[2,3-d]pyrimidine has
been established as a unique antifolate with remarkable activity
against a broad spectrum of solid tumors.9 The scaffold draws
attention from the material chemists too due to its redox ability
and interesting photophysical properties.10 A plethora of reports
have been found reporting the synthesis and biological evaluation
of diversely substituted pyrrolo[2,3-d]pyrimidines.11However,
only a few reports have been found for the synthesis of 5-
arylpyrrolo[2,3-d]pyrimidine and to the best of our knowledge
not a single report has been found to evaluate the biological
significances of the synthesized compounds. Taylor et al. first
reported the synthesis of eight derivatives of 5-arylpyrrolo[2,3-
d]pyrimidine in a four-step process in modest yields12 and
another couples of reports have been published in subsequent
days.13Herein, we report the synthesis of ten derivatives of 5-
chemicals. A gentle attempt to carry out preliminary antibacterial
studyof the synthesized compounds has been made.
Figure 1. Structure of Alimta, an antifolate
Following the retrosynthetic analysis (Scheme 1) we sought to
design an efficient, one pot synthetic methodology for 5-
arylpyrrolo[2,3-d]pyrimidines (4) from aminopyrimidine(1)
andnitrostyrenes (2).We targeted to carry out the synthesis in a
one
arylpyrrolo[2,3-d]pyrimidines in
a
one pot, two-step
Scheme 1. Retrosynthetic analysis of 5-arylpyrrolo[2,3-
d]pyrimidine (4)
methodology using readily available and cheaper laboratory
———
 Corresponding author. Tel.: +91 275059; fax: +91 (3712) 267005/6; e-mail: ashim@tezu.ernet.in

pot two-step process by exploiting two well known facts: (i) the
favorability of Michael addition reaction in alkaline medium and
(ii) the strong reducing capability of alkaline Na2S2O4 solution.
These two facts led us to the idea that there could be a useful
synthetic route for pyrrolo[2,3-d]pyrimidine starting from
aminopyrimidine (1) and nitrostyrene (2) by carrying out the
Michael type addition (first step) in alkaline medium followed by
reductive cyclisation of the Michael type adduct (second step)
using alkaline Na2S2O4 solution (Scheme 2). The complete
process was expected to be carried out in a single pot, if both the
steps would become successful in a common solvent.
requirement for complete consumption of 1 to 1 hour only. Work
up was carried out to isolate the crude 3a, which was then
subjected to reductive cyclisation using alkaline Na2S2O4 solution
Scheme 2. Proposed route for pyrrolo[2,3-d]pyrimidine
To start with, 6-amino-1,3-dimethylpyrimidine-2,4(1H,3H)-
dione (1) was treated with (E)-(2-nitrovinyl)benzene (2a) in
equimolar ratio in water at room temperature (Scheme 3). The
starting materials were observed to be consumed completely
within 6 hours resulting a single product which was later on
confirmed as the Michael type adduct (3a) by single crystal X-
ray analysis (Fig. 2).14 The same reaction was then tested in
presence of 1 equivalent of aqueous NaOHsolution and to our
delight, it worked sufficiently well to reduce the time required for
complete consumption of the substrates from 6 hrs to 2 hrs. Other
Figure 3. Bar diagram showing the effect of solvent-base combination
on Scheme 3. Here, A=H2O; B=EtOH; C=CH3CN; D=DMF;
E=CH3COCH3; F=CHCl3; P=No base; Q=NaOH; R=Na2CO3;
S=CH3COONa; T=Et3N; U=Ph3P (% conversions were calculated from
1H NMR spectra)
Scheme 3. Michael type addition between1 &2a
Scheme 4. Reductive cyclisation of Michael type adduct (3a)
(Scheme 4). Effect of solvent, stoichiometry of NaOH as base
and temperature on the yield of product (4a) were also studied
and the results are summarized in Table 1. Solvents like DMF,
DMSO were not considered due to their high boiling point. Table
1 clearly indicates the suitability of EtOH and (CH3)2CO for the
purpose. However, EtOH became the first choice considering the
fact that it was the most suitable solvent for the Scheme 3 too,
which would offer the opportunity to carry out both Schemes 3
and 4 in a common solvent. The Table 1 also establishes 1:3:4 as
the best stoichiometry for 3a, Na2S2O4 and NaOH as well as 60
0C as the optimum temperature for the reaction. After optimizing
the reaction conditions for both Schemes 3and 4, an attempt was
made to carry out the synthesis of pyrrolo[2,3-d]pyrimidine via a
single pot two-steps route without isolating the intermediate
Michael type product. Optimization for this overall route was
also carried out which concluded with a reduction of the amount
of NaOH added in the second step from 4 equivalent to 3.5
equivalent. The overall process has been expressed as Scheme 5.
Figure 2. ORTEP diagram of 3a
solvent-base combinations were also examined under identical
conditions and the results are summarized in a bar diagram (Fig.
3). Fig. 3 clearly signifies the suitability of polar protic solvents
over polar aprotic as well as non-polar solvents and EtOH-NaOH
is established as the best solvent-base combination for the
purpose. EtOH is found to be more effective than H2O (A-Q and
B-Q in Fig. 3), which may be attributed to the greater solubility
of starting materials in EtOH than in H2O. Optimization of NaOH
added was carried out and an amount of 1.5 equivalent was
established as the most suitable as it reduced the time
The final product (4a) was confirmed as 1,3-dimethyl-5-
phenyl-1H-pyrrolo[2,3-d]pyrimidine-2,4(3H,7H)-dione by single
crystal X-ray analysis (Fig. 4).14 Senda et al. reported the
synthesis of the same compound (4a) in 1974 via a different
route.15 However, the compound synthesized by them might
probably be some kind of structural isomer of 4a, as we have
observed clear mp difference (reported 287 0C; found 191 0C for

Table 1. Effect of reaction condition on Scheme 4
Scheme 5. One pot two- step synthesis of 4a
rt
6
32
32
1:1:1
1:2:3
1:3:4
60
reflux
rt
60
reflux
6
6
6
6
6
42
48
45
60
64
40
45
43
60
57
1
2
3
4
H2O
rt
60
6
6
6
6
6
6
6
6
6
6
4
4
6
1
1
6
1
1
6
6
6
6
6
6
6
6
6
6
6
6
6
6
6
6
4
4
6
1
0.5
6
1
0.5
46
72
45
68
72
46
69
67
30
43
52
40
68
70
58
92
84
56
92
86
25
42
53
48
60
62
50
85
82
52
84
85
32
44
53
36
68
70
63
90
86
63
91
79
reflux
rt
80
48
1:3:5
1:1:1
1:2:3
1:3:4
1:3:5
1:1:1
1:2:3
1:3:4
1:3:5
1:1:1
1:2:3
1:3:4
1:3:5
60
70
reflux
rt
82
30
60
45
Figure 4. ORTEP diagram of 4a
reflux
rt
58
43
in many case, but we have not used any organic solvent for the
reaction except in the work up procedure and purification.
60
70
reflux
rt
75
1
EtOH
The complete transformation can be studied by H NMR
60
spectra as shown in Fig. 5. To generalize the methodology, it was
applied to a series of nitrostyrene derivatives (Scheme 6) and the
results are summarized in Table 2.16 Table 2 clearly reflects that
the methodology is applicable to a wide variety of nitrostyrenes
resulting in excellent to moderate yield. Presence of weakly
electron withdrawing & electron donating groups (Entries b, c, d
& h, Table 2) in the aromatic ring of the nitrostyrene derivatives
60
100
100
62
reflux
rt
60
100
100
25
reflux
rt
60
45
reflux
rt
55
50
60
62
reflux
rt
63
CH3CN
52
60
88
reflux
rt
88
54
60
86
reflux
rt
90
35
60
45
reflux
rt
55
40
60
70
reflux
rt
75
(CH3)2CO
65
60
100
100
65
reflux
rt
60
100
100
reflux
aCalculated from 1H NMR spectra.
bIsolated yield.
entry a, table 2 for example) between the two and our structure
for 4a has been supported by single crystal X-ray analysis.
Contrary to our method, here, the 6-position of pyrimidine was
targeted and then cyclisation. Although our method is a two-step
process, still the same can be effected in a single pot. But,
methodology adopted by Senda et el although is a two step
process, but it is not one pot. Senda et al used tetralin as solvent
Figure 5. Comparison of 1H NMR spectra of substrates, intermediate
and product

don’t alter the yield of 4 to a great extent. However, the
consumption of 1 as well as the yield of 4 is observed to be
comparatively low in presence of strongly electron donating
groups (Entries I & k, Table 2). Presence of heteroaryl units in
the nitrostyrene derivatives also seemed to lower the yield of the
final product 4 (Entries e, f & j, Table 2).
Table 3. Michael type addition of 1 with electron deficient
olefins via 1s t step of Scheme 6
a
b
c
d
e
C6H5-
1
100
100
100
100
100
95
92
92
88
64
202
154
126
242
195
p-ClC6H4-
o-HOC6H4-
p-MeOC6H4-
1.5
1.5
2
Scheme 6. General reaction scheme for synthesis of 4
1.5
Although detailed mechanistic studies were not performed, a
plausible mechanism has been suggested based on our
observation towards Michael type addition reaction as well as
literature reports available for reduction using alkaline Na2S2O4
(Scheme 7). Once 7 is formed, it eliminatesone molecule of
f
1.5
1.5
100
100
62
88
191
137
g
Table 2. Synthesis of pyrrolo[2,3-d]pyrimidine derivatives
via Scheme 6
h
i
p-MeC6H4-
1.5
4
100
55
86
38
202
216
j
4
4
64
62
54
54
160
195
a
b
c
d
e
C6H5-
1
100
100
100
100
100
1
92
87
82
72
48
191
227
278
246
262
p-ClC6H4-
o-HOC6H4-
p-MeOC6H4-
1.5
1.5
2
1.2
2
k
1.5
1.5
l
1.5
6
6
--
--
--
--
--
--
m
f
1.5
1.5
100
100
1.5
1
53
83
251
276
n
o
PhCH=CHCOOH
PhCH=CHCOPh
6
6
--
--
--
--
--
--
g
aCalculated from 1H NMR spectra.
h
i
p-MeC6H4-
1.5
4
100
55
1
3
82
--
237
--
bIsolated yield
Table 4. Bacterial zones of inhibition (in mm) by synthesized
pyrrolo[2,3-d]pyrimidines
Entry Compounds
Microorganisms
Gram positive Gram negative
B.
j
4
4
64
62
2
2
42
46
230
218
B.
subtilis mycoides
E. coli
P.
aeruginosa
k
1
4a
-
-
-
-
-
2
4b
-
-
-
aCalculated from 1H NMR spectra. bIsolated yield
3
4c
-
-
-
-
4
4d
-
-
-
-
5
4e
20
20
18
20
20
15
-
12
13
14
15
12
11
18
20
20
20
20
20
6
4f
13
17
12
-
7
4g
8
4h
9
4j
10
4k
12
Scheme 7. Plausible mechanism for the formation of 4
-, no inhibition zone

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aromatic counterpart 4.
3.
4.
Being benefitted by the fastness of the methodology, we
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by stopping the reaction at the first step in a separate set of
reactions. A few compounds of these series (3a, 3d & 3k) have
already been reported,17 however our methodology is found to be
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dry methanol is required in our case) time and product yield. An
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acrylic acid, cinnamic acid and chalcone as electron deficient
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5.
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7.
8.
The synthesized pyrrolo[2,3-d]pyrimidines (4a-h, 4j & 4k) were
screened for their in vitro antibacterial property against both
Gram positive (Bacillus subtilis and Bacillus mycoides) and
Gram negative (E. coli. and Pseudomonous aeruginosa) bacterial
strains at a concentration of 5mg/ml by taking ampicillin (5
mg/mL) as reference compound.16 The bacterial zones of
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method and the results are summarized in Table 4. It is well
noticed that some of the synthesized compounds (4e-4h, 4j&4k)
showed remarkable activity against P. aeruginosa and B. subtilis.
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In conclusion, a convenient one-pot, two-step synthesis of 5-
arylpyrrolo[2,3-d]pyrimidines has been accomplished, which
involves Michael type addition followed by reductive cyclisation
of the adduct. The methodology is non-catalytic, however still
attractive as it is quite general and covers a wide spectrum of
nitrostyrenes. The methodology enjoys additional advantage of
utilizing cheaper and easily available reagent system. From
biological assay studies, it is found that six of the synthesized
compounds show satisfactory activity against P. aeruginosa and
B. subtilis.
Acknowledgment
LS and AJT are indebted to the Council of Scientific and
Industrial Research (CSIR), New Delhi, India, for financial
supportto theproject CSIR (01(2147)/07/EMR-II).
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16. Supplementary
data
(synthetic
procedures,
structural
characterization as well as bioassay tests) associated with this
article can be found, in the online version.
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