Synthesis, Antibacterial Activity, and Docking Studies of 1,2,3-triazole-tagged Thieno[2,3-d]pyrimidinone Derivatives

Article abstract of DOI:10.1002/jhet.2995

Novel (1-(substituted phenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)acetate derivatives were synthesized. All the compounds showed significant antibacterial activity against Gram-negative (Escherichia coli and Klebsiella pneumonia) and Gram-positive (Bacillus subtilis and Bacillus cereus) bacteria. Particularly, (1-(3-nitrophenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-5,6,7,8-tetrahydrobenzo[1, 2]thieno[2,3-d]pyrimidin-3(4H)-yl)acetate was found to be most potent against E.?coli, K.?pneumonia, and B.?subtilis with MIC 25?μg/ml. Molecular docking was also performed on purine riboswitch of B.?subtilis and thiamine pyrophosphate riboswitch of E.?coli.

Full text of DOI:10.1002/jhet.2995

Month 2017
Synthesis, Antibacterial Activity, and Docking Studies of 1,2,3-triazole-
tagged Thieno[2,3-d]pyrimidinone Derivatives
M. Aruna Kumari, S. Triloknadh, N. Harikrishna, M. Vijjulatha, and C. Venkata Rao
a
a
a
b
a
*
aDepartment of Chemistry, Sri Venkateswara University, Tirupati 517 502, Andhra Pradesh, India
bDepartment of Chemistry, University College of Science, Osmania University, Telangana, India
*
E-mail: cvrsvu@gmail.com
Received March 6, 2017
DOI 10.1002/jhet.2995
Published online 00 Month 2017 in Wiley Online Library (wileyonlinelibrary.com).
Novel
(1-(substituted
phenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-5,6,7,8-tetrahydrobenzo[1,2]
thieno[2,3-d]pyrimidin-3(4H)-yl)acetate derivatives were synthesized. All the compounds showed
significant antibacterial activity against Gram-negative (Escherichia coli and Klebsiella pneumonia) and
Gram-positive (Bacillus subtilis and Bacillus cereus) bacteria. Particularly, (1-(3-nitrophenyl)-1H-1,2,3-
triazol-4-yl)methyl-2-(4-oxo-5,6,7,8-tetrahydrobenzo[1, 2]thieno[2,3-d]pyrimidin-3(4H)-yl)acetate was
found to be most potent against E. coli, K. pneumonia, and B. subtilis with MIC 25 μg/ml. Molecular
docking was also performed on purine riboswitch of B. subtilis and thiamine pyrophosphate riboswitch of
E. coli.
J. Heterocyclic Chem., 00, 00 (2017).
INTRODUCTION
activity of cyclopenteno and cyclohexeno[b]thieno
[2,3-d]-3,4-dihydropyrimidine-4-one derivatives (c) (Fig. 1).
Mulla et al. [11] synthesized triazolothienopyrimidine
derivatives (d) (Fig. 1) and studied their analgesic, anti-
inflammatory, and antibacterial activities.
Today, bacterial infection is one of the most complex
health issues globally. Because of increasing antibiotic
resistant organisms and also decreasing the discovery of
new antimicrobial agents, there is a pressing need of new
antimicrobial agents that are capable of treating those
strains [1]. As a result, increasing efforts have been made
to develop multidrug resistant (MDR) strategy [2] by
which each drug moiety is designed to bind independently
two different biological targets that offer the possibility to
overcome the current resistance [3].
Pyrimidine has always been an inimitable interesting
heterocyclic moiety, present in several biological systems
like DNA, RNA, vitamins, coenzymes, and purines.
Among fused pyrimidines, thieno[2,3-d]pyrimidines were
found to exhibit various biological activities like
In addition, 1,2,3-triazoles possess unique features that
make them important pharmacophore in curative
medicinal chemistry, and they act as bioisosters and
linkers [12]. They contain high aromatic stabilization and
large dipole moment (5D) [13] and participate actively in
hydrogen bonding and also easily soluble in water.
Moreover 1,2,3-triazole is an attractive linker unit between
two pharmacophore units to give an innovative
bifunctional drugs, which are useful and important in
constructing bioactive molecules [14]. They exhibit
several biological activities like antimicrobial [15],
antiallergic [16], antitubercular activity [17], antineoplastic
[18], antimalarial [19], antiviral [20], antiproliferative
[21], and anticancer activity [22]. Also, they are reliable
and quantitative to synthesize by using Huisgen’s copper
catalyzed azide-alkyne cyclo addition (CuAAC) under
mild conditions without protecting groups and purification
[23,24]. There are few 1,2,3-triazole bearing drugs
available in the market like Rufinamide, Tazobactum,
Cefatrizine, as shown in Figure 2.
antimicrobial,
analgesic,
anti-inflammatory,
and
anticonvulsant [4–6], and they also inhibit various protein
kinase such as CK2 involved in particular anticancer
activity [7]. Several thieno[2,3-d]pyrimidine derivatives
were synthesized and proved to be antibacterial agents.
Chambhare et al. [8] synthesized Schiff bases of thieno
[2,3-d]pyrimidine derivatives (a) (Fig. 1) and proved as
antimicrobial agents. Abbas et al. [9] studied antibacterial
and antitumor activity of tetrahydrobenzo[4,5]thieno
[2,3-d]pyrimidine derivatives (b) (Fig. 1), and EI-Sherbeny
et al. [10] have examined the antibacterial and antiviral
In literature, there are numerous fused 1,2,3-triazole
derivatives of thieno[2,3-d]pyrimidines, but 1,2,3-triazole-
linked thieno[2,3-d]pyrimidine derivatives are exceptional
© 2017 Wiley Periodicals, Inc.

M. Aruna Kumari, S. Triloknadh, N. Harikrishna, M. Vijjulatha, and C. Venkata Rao
Vol 000
Figure 1. Thieno[2,3-d]pyrimidine derivatives possessing antibacterial activity.
Figure 2. Biologically active drugs available in the market containing 1,2,3-triazole moiety.
in synthesis. In the light of the aforementioned specifics,
RESULTS AND DISCUSSION
considerable efforts have been made to develop novel
synthetic strategies by joining thieno[2,3-d]pyrimidinone
scaffold and 1,2,3-triazole moiety through ester linkage.
The synthesized compounds were screened for their
antibacterial activity to obtain novel candidates of
incredible activities with nominal side effects. In order to
establish the binding interactions, docking studies were
performed on B. subtilis purine riboswitch and E. coli TPP
riboswitch.
The synthesis of (1-(substituted phenyl)-1H-1,2,3-
triazol-4-yl)methyl2-(4-oxo-5,6,7,8-tetrahydrobenzo [4,5]
thieno[2,3-d]pyrimidin-3(4H)-yl)acetate derivatives (7a–j)
involved several steps as shown in Scheme 1. In the first
step,
2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-
carbonitrile (1) was prepared by multicomponent
condensation between elemental sulfur, cyclohexanone,
and activated malanonitrile in triethyl amine using Gewald
Scheme 1. Synthetic route for the compounds (7a–j).
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2017
Synthesis, Antibacterial Activity, and Docking Studies of 1,2,3-triazole-tagged
Thieno[2,3-d]pyrimidinone Derivatives
reaction conditions [25]. Cyclisation of amino and cyano
functionality of compound 1 on treatment with formic acid
and sulfuric acid at refluxing temperature 110°C resulted
in the formation of 5,6,7,8-tetrahydro[1]benzothieno[2,3-
d]pyrimidin-4(3H)-one (2). Compound 2 on treatment
with ethyl bromoacetate and potassium carbonate in
acetone gave ethyl 2-(4-oxo-5,6,7,8-tetrahydrobenzo[4,5]
thieno[2,3-d]pyrimidin-3(4H)-yl)acetate (3). In this step,
there is a possibility to yield two isomeric products
O-alkylated and N-alkylated thienopyrimidine derivatives.
The ratio of each isomer is based on the type of substituent
present in second position of thienopyrimidine derivative.
The O-alkylated product is formed in major when
substituent is aromatic, while N-alkylated compound is
major when substituent is aliphatic [26]. Here, N-alkylated
derivative 3 is formed in major amount because in
compound 2 substituent at the second position is
hydrogen. Next, compound 3 was hydrolyzed using
methanol and sodium hydroxide to prepare 2-(4-
oxo-5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d] pyrimidin-
3(4H)-yl)acetic acid (4). Further, propargylation of the
carboxyl group with potassium carbonate as a base in
dimethylformamide delivered the alkyne derivative 5. The
aryl azide derivatives 6a–j were prepared from their
respective anilines by diazotization with sodium nitrite in
acidic conditions followed by nucleophilic displacement
with sodium azide. Finally, compound 7 was obtained by
1,3-dipolar cycloaddition reaction of alkyne derivative 5
with aromatic azides 6a–j in the presence of CuSO4.5H2O
and reducing agent sodium ascorbate in THF:H2O (1:1)
mixture. This “click” reaction yields 1,4-disubstituted
triazolyl derivatives 7a–j regioselectively. The physical
and analytical data of these compounds (7a-j) was
mentioned in Table 1.
absence of a singlet peak at δ 12.32 ppm due to NH proton
confirms the formation of compound 3 as N-alkylated
derivative. Further, its IR spectrum showed an additional
frequency at 1736 cmÀ1 due to C=O for ester functional
group. The formation of compound 4 by the hydrolysis of
3 was supported by it mass spectrum, which revealed a
molecular ion peak at m/z 265.1 [M + H]+ corresponding
to molecular formula C14H16N2O3S and also by the
presence of a singlet at δ 12.9 ppm in its 1H NMR
spectrum corresponding to COOH proton. Compound 4
on treatment with propargyl bromide formed compound 5,
which showed a singlet at δ 2.53 ppm owing to acetylene
CH group. Further, its IR spectrum showed a peak at
3244 cmÀ1 due to ≡CH stretching. The conversion of
terminal alkyne 5 to triazole derivatives 7a–j on treatment
with aromatic azides 6a–j was confirmed by the
disappearance of acetylenic proton signal and appearance
1
of a singlet in H NMR at δ 7.85–8.33 ppm owing to the
resonance signal of C–H (5) of the triazole ring.
Antibacterial activity. All the synthesized compounds
7a–j were assayed for in vitro antibacterial activity
against two different strains of Gram-negative (E. coli
and K. pneumonia) and Gram-positive (B. subtilis and
B. cereus) bacteria by using Gentamicin as a standard
drug. The minimum inhibitory concentration (MIC) was
determined by broth dilution technique using Mueller-
Hinton nutrient broth medium. The MIC values of test
compounds are summarized in Table 2. The results
revealed that all the test compounds 7a–j exhibited
remarkable in vitro antibacterial activity against all tested
bacterial strains. The MIC values of these compounds are
in the range of 25–50 μg/mL. DMSO was taken as a
control, which showed no effect in the experiment. The
comparison of MIC values of the synthesized compounds
is shown in Figure 3.
The formation of compound 2 was confirmed by the
presence of singlets at δ 12.32 and 8.02 ppm due to NH
and CH protons of pyrimidine ring respectively in 1H
NMR spectrum. Apart from this, its IR spectrum showed a
peak at 1651 cmÀ1 corresponding to –NH–C=O
stretching. A signal at δ 4.84 ppm due to CH2 protons and
These results showed that compound 7g is most potent
against all bacterial strains with MIC 25 μg/mL.
Compounds 7a, 7d, 7f, 7i, and 7j showed good activity.
Compounds 7b, 7e, and 7h showed comparatively poor
activity against all the bacterial strains with MIC
Table 1
Physical and analytical data of compounds (7a–j).
Compound
7a
7b
7c
7d
7e
7f
R
Molecular formula (mol. wt.)
M.P (°C)
Yield (%)
Mass [M++1]
4-methyl
4-chloro
Hydrogen
C22H21N5O3S (435)
C21H18N5O3SCl (455)
C21H19N5O3S (421)
C21H16N5O3SCl3 (523)
C22H21N5O4S (451)
C21H18N5O3SF (439)
C21H18N6O5S (466)
C21H18N6O5S (466)
C22H19N6O6SCl (530)
C21H17N6O5SCl (500)
120–125
155–157
110–111
170–172
180–185
130–132
135–140
155–156
150–155
170–180
80
80
70
60
90
70
90
70
70
60
436.1
456.1
422
524
452.2
440
467.1
467.1
531.1
501.1
2,4,5-trichloro
4-methoxy
3-fluoro
3-nitro
4-nitro
7g
7h
7i
4-chloro, 2-methoxy,
6-nitro,2-chloro,5-nitro
7j
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

M. Aruna Kumari, S. Triloknadh, N. Harikrishna, M. Vijjulatha, and C. Venkata Rao
Vol 000
Table 2
Antibacterial activities of the synthesized compounds (7a–j).
MIC (μg/mL)
Gram-positive bacteria
Gram-negative bacteria
Escherichia coli Klebsiella pneumonia
Compound
Bacillus subtilis
Bacillus cereus
7a
7b
7c
7d
7e
7f
7g
7h
50
50
50
25
50
25
25
50
25
25
25
0
50
50
50
25
50
25
25
50
25
25
10
0
25
50
50
50
50
50
25
50
25
25
25
0
25
50
25
50
50
25
25
50
50
50
50
0
7i
7j
Gentamicin (standard)
DMSO (control)
Figure 3. Antibacterial activity of compounds 7a–j and standard drug Gentamicin against Gram-positive bacteria (B. subtilis and B. cereus) and Gram-
negative bacteria (E. coli and K. pneumonia) represented as MIC (μg/mL) values. [Color figure can be viewed at wileyonlinelibrary.com]
50 μg/mL. From these results, it was envisaged that the
position and nature of substituent influence the
antibacterial activity of the synthesized compounds.
Compound 7c exhibited good activity against
K. pneumonia with MIC 25 μg/mL, which is more potent
than Gentamicin. From these results, it is cleared that
compound 7g is worth for further studying to evaluate
better antibacterial agents in future.
Docking. In order to know the binding interactions of
these molecules docking studies were performed on
B. subtilis purine riboswitch and E. coli TPP riboswitch
using GLIDE 5.6. The ability of the riboswitches to
control key metabolic pathways in many bacterial species
like B. subtilis and E. coli makes them a promising target
for antibacterial drug therapy. The crystal structures were
downloaded from protein data bank (http://www.rcsb.
org). The dock score and hydrogen bond interactions are
used to predict the binding modes of synthesized
compounds at the active sites.
Compound 7g is most potent because of the cumulative
electron-withdrawing effect of the meta-substituted nitro
group. Compound 7h containing para-substituted nitro
group showed low activity compared with 7g. This is due
to conjugative effect of para nitro group. The same was
observed for compounds 7b and 7e containing para-
substituted chloro and methoxy groups, respectively.
Further, compounds 7i and 7j having electron-
withdrawing groups (chloro, nitro, and methoxy)
substituted benzene ring were found to be good active
with MIC 25 μg/mL against B. subtilis, B. cereus, and
E. coli. Moreover, compound 7f having fluoro group
showed MIC 25 μg/mL against B. subtilis, B. cereus, and
K. pneumonia. Compound 7d having trichloro-substituted
phenyl ring showed good activity against B. subtilis and
K. pneumonia with MIC values 25 and 50 μg/mL,
respectively, which are same as that of Gentamicin.
As shown from Table 3, the molecules showed good
dock score toward TPP riboswitch of E. coli ranging
from À6.738 to À3.959 Kcal/mol than purine
riboswitch of B. subtilis showing dock score ranging
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2017
Synthesis, Antibacterial Activity, and Docking Studies of 1,2,3-triazole-tagged
Thieno[2,3-d]pyrimidinone Derivatives
Table 3
from À6.091 to À3.074 Kcal/mol. Among all the
synthesized molecules, compound 7g, which is the most
active one, showed high dock scores of À6.738 and
À6.091 Kcal/mol against E. coli TPP riboswitch and
B. subtilis purine riboswitch, respectively.
Dock score of synthesized compounds (7a–j) from Glide Docking.
Dock score (Kcal/mol)
Bacillus subtilis Escherichia coli
The binding models of compound 7g and Gentamicin
with TPP riboswitch of E. coli and purine riboswitch of
B. subtilis were depicted in Figs 4 and 5, respectively.
Compound 7g showed two hydrogen bond interactions
with E. coli TPP riboswitch. One was between NO2
group of compound 7g and G40 active site. Another one
was between sulfur atom of thiophene ring with C58
active site. The standard drug Gentamicin showed
hydrogen bonds with C57, C58, G60, and C73 active
sites, and the dock score is À7.791 Kcal/mol. In
B. subtilis also, compound 7g forms two hydrogen bond
interactions. One hydrogen bond connects C=O group of
Compound
Pdb id: 2XNW
Pdb id: 4NYG
7a
7b
7c
7d
7e
7f
7g
7h
À4.169
À3.835
À3.810
À4.679
À4.391
À4.488
À6.091
À3.399
À3.074
À5.545
À7.687
À3.964
À4.455
À3.979
À4.477
À5.348
À3.959
À6.738
À4.439
À4.339
À5.202
À7.791
7i
7j
Gentamicin (standard)
Figure 4. Docked pose of 7g (a) and gentamicin (b) in the protein active site. Showing hydrogen bond interactions (yellow lines) with G40, C58 and C57,
C58, G60, and C73, respectively, in E. coli TPP riboswitch (pdb id: 4NYG). [Color figure can be viewed at wileyonlinelibrary.com]
Figure 5. Docked pose of 7g (a) and gentamicin (b) in the protein active site. Showing hydrogen bond interactions (yellow lines) with C18, A19 and A40,
G42, C53, C54, and G55, respectively, in Bacillus subtilis Purine riboswitch (pdb id: 2XNW). [Color figure can be viewed at wileyonlinelibrary.com]
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

M. Aruna Kumari, S. Triloknadh, N. Harikrishna, M. Vijjulatha, and C. Venkata Rao
Vol 000
pyrimidine with A19 active site. The second hydrogen
bond was formed between C=O group of ester with C18
active site. Gentamicin showed hydrogen bonds with
A40, G42, C53, C54, and G55 active sites with a dock
score of À7.687 Kcal/mol.
Synthesis
of
5,6,7,8-tetrahydro[3]benzothieno[2,3-d]
pyrimidin-4(3H)-one (2).
Concentrated sulfuric acid
(4 mL) at 0–5°C was added to a mixture of 2-amino-
4,5,6,7-tetrahydrobenzo[b]thiophene-3-carbonitrile
1
(1 g, 5.6 mmol) in formic acid (10 mL). The mixture
was refluxed for 2 h and allowed to cool at room
temperature (rt). Then, the mixture was poured into
crushed ice. The separated solid was filtered and
purified by recrystallization with ethanol.
CONCLUSIONS
In summary, a series of novel (1-(substituted phenyl)-1H-
1,2,3-triazol-4-yl)methyl2-(4-oxo-5,6,7,8-tetrahydrobenzo
Brown color solid. Yield 89%; mp 190–192°C; IR (KBr)
1
(νmax, cmÀ1): 2836 (ÀCH), 1651 (C=O), 1581 (C=N); H
[4,5]thieno[2,3-d]pyrimidin-3(4H)-yl)acetate
derivatives
NMR (400 MHz, DMSO-d6): δ 1.78–1.85 (m, 4H, CH2),
2.77 (t, J = 5.2 Hz, 2H, CH2), 2.91 (t, J = 5.6 Hz, 2H,
CH2), 8.02 (s, 1H, CH-pyrimidine), 12.32 (br s, 1H, NH);
13C NMR (100 MHz, DMSO-d6): δ 21.7, 22.4, 24.4,
25.2, 122.6, 130.7, 132.0, 144.7, 157.6, 162.3; LCMS
(negative ion mode) (m/z): 205 [MÀH]À for C10H10N2OS.
(7a–j) were synthesized and evaluated their in vitro
antibacterial activity by broth dilution method. All the
compounds showed promising activity with MIC values
between 25 and 50 μg/mL. Among these, compound 7g
containing meta-substituted nitro group displayed
promising activity against all bacterial strains with MIC
25 μg/mL except for B. cereus. It was found to be twofold
active than that of standard drug Gentamicin against
K. pneumonia. Moreover, it was same potent as that of
Gentamicin against B. subtilis and E. coli. Further, it was
supported by docking studies. Among all the compounds,
7g showed highest dock score in the range of À6.091 to
À6.738 Kcal/mol. The docking models indicated that
compound 7g could snugly occupy the active sites of TPP
riboswitch and Purine riboswitch. After comparing the
docking results and biological assay data, it could be
suggested that compound 7g was the most potential
antibacterial agent. These findings will guide us to develop
novel target specific antibacterial agents from the source of
thieno[2,3-d]pyrimidinone-tagged 1,2,3-triazole nucleoside
derivatives with improved efficacy and fewer side effects.
Synthesis of ethyl 2-(4-oxo-5,6,7,8-tetrahydrobenzo[1,2]
thieno[2,3-d]pyrimidin-3(4H)-yl)acetate (3). A solution of
compound 2 (2 g, 10 mmol), ethyl bromoacetate (1.3 mL,
12 mmol), and potassium carbonate (2 g, 15 mmol) in
20 mL acetone was added and stirred for 8 h. After
completion of reaction mixture, the acetone was
evaporated, and the precipitated solid was obtained,
washed with cold water, and recrystallized with ethanol.
Pale yellow color solid. Yield: 83%; mp 192–194°C; IR
(KBr) (νmax, cmÀ1): 3115 (=CH), 1736 (ester C=O), 1668
(amide C=O), 1609 (C=N); 1H NMR (400 MHz,
DMSO-d6): δ 1.28 (t, J = 8.0 Hz, 3H, CH2CH3), 1.81–
1.87 (m, 4H, 2CH2), 2.82 (t, J = 5.6 Hz, 2H, CH2), 2.93
(t, J = 6.0 Hz, 2H, CH2), 4.24 (q, J = 8.0 Hz, 2H,
CH2CH3), 4.84 (s, 2H, CH2CO), 8.36 (s, 1H, CH-
pyrimidine); 13C NMR (100 MHz, DMSO-d6): δ 13.0,
21.1, 21.8, 24.2, 24.5, 45.9, 61.1, 121.5, 130.6, 133.5,
144.3, 156.4, 161.1, 166.3; LCMS (m/z): 293 [M + H]+,
315 [M + Na]+ for C14H16N2O3S.
EXPERIMENTAL
Synthesis of 2-(4-oxo-5,6,7,8-tetrahydrobenzo[1,2]thieno
All the solvents and reagents were obtained from
commercial sources and were used without further
purification. Melting points were determined in open
capillaries on a MelTemp apparatus and are uncorrected.
All the reactions were monitored by thin layer
chromatography (TLC) on precoated silica gel 60 F254
(mesh); spots were visualized with UV light. Merck silica
gel (60–120 mesh) was used for column chromatography.
The IR spectra were recorded on a PerkinElmer BX1
FTIR Spectrophotometer as KBr pellets, and the wave
[2,3-d]pyrimidin-3(4H)-yl)acetic acid (4). To a solution of
compound 3 (2 g, 6.8 mmol) in methanol, sodium
hydroxide (0.68 g, 17 mmol) solution in water was added
and subjected to stirring at room temperature overnight.
After completion of reaction, the reaction mixture was
poured into crushed ice, and con. HCl was added till it
becomes acidified. The formed solid was filtered and
purified by recrystallization with ethanol.
White solid yield: 80%; mp 130–133°C; IR (KBr) (νmax,
cmÀ1): 3615 (OH), 3115 (=CH), 1716 (ester C=O), 1668
1
1
numbers were given in cmÀ1. H NMR (400 MHz) and
(amide C=O), 1558 (C=N); H NMR (400 MHz, CDCl3):
13C NMR (100 MHz) spectra were recorded on a Bruker
AMX 400 MHz NMR spectrometer in CDCl3/DMSO-d6
solution using TMS as an internal standard. All chemical
shifts are reported in δ (ppm) using TMS as an internal
standard. The mass spectra were recorded on Agilent
1100 LC/MSD instrument with method API-ES at 70 eV.
δ 1.74–1.81 (m, 4H, 2CH2), 2.71 (t, J = 4.7 Hz, 2H,
CH2), 2.86 (t, J = 5.8 Hz, 2H, CH2), 4.7 (s, 2H, CH2),
8.06 (s, 1H, pyrimidine-CH), 12.90 (s, 1H, COOH); 13C
NMR (100 MHz, CDCl3): δ 20.6, 21.3, 23.3, 24.0, 55.7,
116.6, 123.4, 144.0, 145.5, 151.3, 163.7, 166.7; LCMS
(m/z): 265 [M + H]+ for C12H12N2O3S.
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DOI 10.1002/jhet

Month 2017
Synthesis, Antibacterial Activity, and Docking Studies of 1,2,3-triazole-tagged
Thieno[2,3-d]pyrimidinone Derivatives
Synthesis of prop-2-yn-1-yl-2-(4-oxo-5,6,7,8-tetrahydro-
benzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)acetate (5). To a
(1-(p-Tolyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-5,6,7,8-
tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)acetate
(7a). Light brown solid. Yield 83%, mp 120–125°C. IR
solution of compound 4 (2 g, 7.6 mmol) in dry acetone
(20 mL) were added anhydrous potassium carbonate
(2.12 g, 11.4 mmol) and 3-bromoprop-1-yne (1.04 mL,
11.4 mmol) successively. The resultant mixture was
stirred overnight at room temperature. The reaction was
monitored by TLC. Upon completion of reaction, the
reaction mixture was poured into crushed ice and stirred
continuously. The obtained solid was filtered and purified
by column chromatography using ethyl acetate and
hexane as eluent.
(KBr) ν, cmÀ1: 3122 (=CH), 2922 (ÀCH), 1735 (ester
C=O), 1676 (amide C=O), 1611 (C=N). 1H NMR
(400 MHz, CDCl3): δ 1.81–1.89 (m, 4H, 2CH2), 2.43 (s,
3H, CH3), 2.78 (t, J = 5.5 Hz, 2H, CH2), 2.98 (t,
J = 5.5 Hz, 2H, CH2), 4.71 (s, 2H, CH2), 5.44 (s, 2H,
CH2-triazole), 7.31 (d, J = 8.0 Hz, 2H, Ar–H), 7.61 (d,
J = 8.0 Hz, 2H, Ar–H), 7.90 (s, 1H, triazole-CH), 8.11 (s,
1H, pyrimidine-CH); 13C NMR (100 MHz, CDCl3): δ
22.1, 30.2, 30.8, 33.2, 33.6, 54.9, 70.1, 112.5, 116.9,
123.9, 130.6, 142.6, 145.2, 148.0, 153.3, 159.0, 161.4,
165.5, 170.2, 174.9; LCMS (m/z): 436.1 [M + H]+, 458.2
Yellow solid yield: 80%; mp 110–112°C; IR (KBr)
(νmax, cmÀ1): 3244 (≡CH), 3125 (=CH), 2124 (C≡C),
[M
+
Na]+ for C22H21N5O3S. Elem. Anal. for
1
1719 (ester C=O), 1645 (amide C=O), 1618 (C=N); H
C22H21N5O3S: (calcd) C, 60.68; H, 4.86; N, 16.08; S,
NMR (400 MHz, CDCl3): δ 1.83–1.88 (m, 4H, 2CH2),
2.53 (s, 1H, ≡CH), 2.81 (t, J = 6.1 Hz, 2H, CH2), 3.00 (t,
J = 5.6 Hz, 2H, CH2), 4.72 (s, 2H, CH2–C≡CH), 4.81 (s,
2H, N–CH2–C=O), 7.85 (s, 1H, pyrimidine-CH); 13C
NMR (100 MHz, CDCl3): δ 22.2, 22.8, 25.2, 25.5, 46.8,
53.3, 75.8, 76.4, 128.8, 131.7, 145.0, 149.2, 156.7,
157.4, 166.6; LCMS (m/z): 303.1 [M + H]+, 325
[M + Na]+ for C15H14N2O3S.
7.36. Found: C, 60.45; H, 4.52; N, 15.75; S, 7.62.
(1-(4-Chlorophenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-
5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)
acetate (7b). Light brown solid. Yield: 80%; mp 155–
157°C; IR (KBr) (νmax, cmÀ1): 3107 (=CH), 2924
(ÀCH), 1740 (ester C=O), 1669 (amide C=O), 1605
(C=N); 1H NMR (400 MHz, CDCl3): δ 1.81–1.89 (m,
4H, 2CH2), 2.78 (t, J = 6.0 Hz, 2H, CH2), 2.99 (t,
J = 6.0 Hz, 2H, CH2), 4.71 (s, 2H, CH2), 5.43 (s, 2H,
CH2-triazole), 7.45 (d, J = 8.1 Hz, 2H, Ar–H), 7.65 (d,
J = 8.1 Hz, 2H, Ar–H), 7.89 (s, 1H, triazole-CH), 8.01 (s,
1H, pyrimidine-CH); 13C NMR (100 MHz, CDCl3): δ
26.9, 27.5, 29.9, 30.2, 52.1, 63.7, 125.2, 126.9, 127.2,
133.6, 134.4, 136.2, 139.5, 141.5, 147.5, 149.9, 162.2,
167.0, 172.1. LCMS (m/z): 456.1[M + H]+, 478.1
Synthesis of aryl azides (6a–j).
Aniline derivative
(9.1 mmol) was dissolved in con. HCl (91 mmol) at room
temperature and cooled up to 0°C, followed by addition of
a solution of sodium nitrite (45.5 mmol). The reaction
mixture was stirred for 10 min at 0–5°C. Sodium azide
(27.3 mmol) was added, and the mixture was further
stirred for 2 h at room temperature. After completion of
reaction, the mixture was extracted with hexane. The
organic layer was dried over anhydrous sodium sulfate,
and after evaporation of the solvent, the desired azide
derivative was obtained.
[M
+
Na]+ for C21H18N5O3SCl. Elem. Anal. for
C21H18N5O3SCl: (calcd) C, 55.32; H, 3.98; N, 15.36; S,
7.03. Found: C, 54.95; H, 3.58; N, 15.50; S, 6.87.
(1-Phenyl-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-5,6,7,8-
tetrahydrobenzo[1,2]thieno[2,3-d] pyrimidin-3(4H)-yl) acetate
General procedure for the synthesis of (1-(substituted
phenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-5,6,7,8-
tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)acetate
derivatives (7a–j). Compound 5 (0.2 g, 0.7 mmol) was
(7c).
Brown solid. Yield: 70%; mp 110–111°C; IR
(KBr) (νmax, cmÀ1): 3116 (=CH), 2921 (ÀCH), 1735
1
(ester C=O), 1665 (amide C=O), 1615 (C=N); H NMR
dissolved in 2 mL of THF : H2O (1:1) solvent in a
50 mL round bottom flask. To this, CuSO.45H2O
(0.17 g, 0.7 mmol) was added, and the reaction mixture
was stirred for 5 min. To this, light blue color reaction
mixture sodium ascorbate (0.2 g, 1.05 mmol) was
added and stirred for 15 min. The reaction mixture
becomes dark yellow. Then aryl azide (6a–j)
(1.05 mmol) was added and allowed to stir for 30 min
to 1 h. After completion of reaction monitored by TLC,
the reaction mixture was extracted with ethyl acetate by
adding excess water. The combined organic layer was
washed with brine solution and dried over anhydrous
sodium sulfate. The solvent was evaporated under
reduced pressure, and the crude product was purified by
silica gel column chromatography using hexane ethyl
acetate as eluent.
(400 MHz, DMSO): δ 1.72–1.80 (m, 4H, 2CH2), 2.75 (t,
J = 5.5 Hz, 2H, CH2), 2.83 (t, J = 5.5 Hz, 2H, CH2),
4.85 (s, 2H, CH2), 5.36 (s, 2H, CH2-triazole), 7.49–7.63
(m, 3H, Ar–H), 7.89 (d, J = 7.9 Hz, 2H, Ar–H), 8.10 (s,
1H, triazole-CH), 8.59 (s, 1H, pyrimidine-CH); 13C NMR
(100 MHz, CDCl3): δ 22.1, 22.7, 25.1, 25.5, 47.3, 59.0,
120.4, 122.1, 122.4, 128.8, 129.7, 131.4, 134.7, 136.7,
142.8, 145.1, 157.4, 162.2, 167.3; LCMS (m/z): 422
[M
+
H]+ for C21H19N5O3S. Elem. Anal. for
C21H19N5O3S: (calcd) C, 59.84; H, 4.54; N, 16.62; S,
7.61. Found: C, 61.18; H, 4.55; N, 16.67; S, 7.40.
(1-(2,4,5-Trichlorophenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-
oxo-5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-
yl)acetate (7d). Yellow solid. Yield: 60%; mp 170–172°C;
IR (KBr) (νmax, cmÀ1): 3115.3 (=CH), 2915 (ÀCH), 1732
1
(ester C=O), 1664 (amide C=O), 1598 (C=N); H NMR
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

M. Aruna Kumari, S. Triloknadh, N. Harikrishna, M. Vijjulatha, and C. Venkata Rao
Vol 000
(400 MHz, CDCl3): δ 1.81–1.89 (m, 4H, 2CH2), 2.79 (t,
J = 5.5 Hz, 2H, CH2), 2.95 (t, J = 6.1 Hz, 2H, CH2),
4.71 (s, 2H, CH2), 5.46 (s, 2H, CH2-triazole), 7.71 (s,
1H, Ar–H), 7.81 (s, 1H, Ar–H), 7.92 (s, 1H, triazole-
CH), 8.14 (s, 1H, pyrimidine-CH); 13C NMR (100 MHz,
CDCl3): δ 22.7, 23.5, 23.9, 34.6, 42.5, 62.7, 117.6,
119.1, 125.4, 128.2, 131.8, 131.9, 132.8, 135.2, 139.4,
142.7, 144.4, 147.6, 155.5, 161.6, 169.5; LCMS (m/z):
524 [M + H]+ for C21H16N5O3SCl3. Elem. Anal. for
C21H16N5O3SCl3: (calcd) C, 48.06; H, 3.07; N, 13.35; S,
1H, triazole-CH), 8.19–8.21 (m, 1H), 8.31–8.33 (m, 2H),
8.64 (s, 1H, pyrimidine-CH); 13C NMR (100 MHz,
CDCl3): δ 22.6, 23.2, 25.6, 25.9, 47.3, 62.5, 106.3,
108.8, 112.3, 116.3, 123.0, 123.2, 132.1, 134.9, 137.5,
145.7, 157.8, 162.6, 167.7, 173.7, 175.1; LCMS (m/z):
467.1 [M + H]+, 489.2 [M + Na]+ for C21H18N6O5S.
Elem. Anal. for C21H18N6O5S: (calcd) C, 54.07; H, 3.89;
N, 18.02; S, 6.87. Found: C, 54.10; H, 4.06; N, 17.72; S,
6.46.
(1-(4-Nitrophenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-
5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)
acetate (7h). Yellow solid. Yield: 70%; mp 155–156°C;
6.11. Found: C, 47.84; H, 2.76; N, 13.18; S, 6.28.
(1-(4-Methoxyphenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-
5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)
IR (KBr) (νmax, cmÀ1): 3112.8 (=CH), 2939 (ÀCH),
1
acetate (7e).
Pale yellow solid. Yield: 90%; mp 180–
1738 (ester C=O), 1675 (amide C=O), 1606 (C=N); H
185°C; IR (KBr) (νmax, cmÀ1): 3111 (=CH), 2926
(ÀCH), 1741 (ester C=O), 1672 (amide C=O), 1609
(C=N); 1H NMR (400 MHz, CDCl3): δ 1.81–1.89 (m,
4H, 2CH2), 2.78 (t, J = 6.0 Hz, 2H, CH2), 2.98 (t,
J = 6.0 Hz, 2H, CH2), 3.89 (s, 3H, ÀOCH3), 4.70 (s,
2H, CH2), 5.43 (s, 2H, CH2-triazole), 7.02 (d, J = 9.0,
2H, Ar–H), 7.64 (d, J = 9.0 Hz, 2H, Ar–H), 7.86 (s, 1H,
triazole-CH), 8.03 (s, 1H, pyrimidine-CH); 13C NMR
(100 MHz, CDCl3): δ 24.1, 24.7, 27.1, 27.4, 32.8, 49.3,
60.9, 122.4, 124.1, 124.4, 130.8, 131.6, 133.4, 136.7,
138.7, 144.7, 147.1, 159.4, 164.2, 169.3. LCMS (m/z):
452.2 [M + H]+, 474.1 [M + Na]+ for C22H21N5O4S.
Elem. Anal. for C22H21N5O4S: (calcd) C, 58.53; H, 4.69;
N, 15.51; S, 7.10. Found: C, 58.16; H, 4.90; N, 15.17;
NMR (400 MHz, CDCl3): δ 1.80–1.90 (m, 4H, 2CH2),
2.79 (t, J = 6.0 Hz, 2H, CH2), 2.99 (t, J = 6.0 Hz, 2H,
CH2), 4.70 (s, 2H, CH2), 5.44 (s, 2H, CH2-triazole), 7.02
(d, J = 9.0 Hz, 2H, Ar–H), 7.64 (d, J = 9.0 Hz, 2H, Ar–
H), 7.86 (s, 1H, triazole-CH), 8.04 (s, 1H, pyrimidine-
CH); 13C NMR (100 MHz, CDCl3): δ 21.7, 23.1, 23.5,
27.5, 41.5, 60.5, 117.6, 119.1, 120.9, 123.9, 125.4, 139.4,
142.9, 144.4, 147.6, 148.2, 155.5, 161.6, 169.5; LCMS
(m/z): 467.1 [M
C21H18N6O5S. Elem. Anal. for C21H18N6O5S: (calcd) C,
54.07; H, 3.89; N, 18.02; S, 6.87. Found: C, 54.37; H,
4.17; N, 17.75; S, 6.48.
+ +
H]+, 489.2 [M Na]+ for
(1-(4-Chloro-2-methoxy-6-nitrophenyl)-1H-1,2,3-triazol-4-
yl)methyl-2-(4-oxo-5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]
pyrimidin-3(4H)-yl)acetate (7i). Brown solid. Yield: 70%;
S, 6.87.
mp 150–155°C; IR (KBr) (νmax, cmÀ1): 3114 (=CH),
2939 (ÀCH), 1730 (ester C=O), 1672 (amide C=O),
1610 (C=N); 1H NMR (400 MHz, CDCl3): δ 1.79–
1.89 (m, 4H, 2CH2), 2.77 (t, J = 5.8 Hz, 2H, CH2),
2.96 (t, J = 5.9 Hz, 2H, CH2), 3.93 (s, 3H, ÀOCH3),
4.72 (s, 2H, CH2), 5.44 (s, 2H, CH2-triazole), 7.14 (s,
1H, Ar–H), 7.49 (s, 1H, Ar–H), 7.86 (s, 1H, triazole-
CH), 8.26 (s, 1H, pyrimidine-CH); 13C NMR
(100 MHz, CDCl3): δ 22.5, 23.0, 23.7, 25.4, 42.5,
55.8, 71.5, 107, 116.5, 117.6, 119.1, 121.2, 125.4,
136.2, 139.4, 144.4, 147.6, 149.1, 155.5, 156.4, 161.6,
(1-(3-Fluorophenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-
5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)
acetate (7f).
Light brown solid. Yield: 70%; mp 130–
132°C; IR (KBr) (νmax, cmÀ1): 3120 (=CH), 2920 (ÀCH),
1
1734 (ester C=O), 1679 (amide C=O), 1614 (C=N); H
NMR (400 MHz, DMSO): δ 1.72–1.80 (m, 4H, 2CH2),
2.74 (t, J = 5.5 Hz, 2H, CH2), 2.82 (t, J = 5.2 Hz, 2H,
CH2), 4.85 (s, 2H, CH2), 5.35 (s, 2H, CH2-triazole), 7.61–
7.69 (m, 1H, Ar–H), 7.78–7.86 (m, 2H, Ar–H), 8.07 (s,
1H, Ar–H), 8.33 (s, 1H, triazole-CH), 8.60 (s, 1H,
pyrimidine-CH); 13C NMR (100 MHz, CDCl3): δ 23.8,
24.5, 26.8, 27.2, 48.5, 63.7, 110.1, 113.4, 117.5, 124.2,
124.4, 129.6, 133.3, 136.2, 138.7, 146.2, 146.9, 159.1,
163.8, 168.4, 168.9; LCMS (m/z): 440 [M + H]+ for
C21H18N5O3SF. Elem. Anal. for C21H18N5O3SF.: (calcd)
C, 57.39; H, 4.13; N, 15.94; S, 7.30. Found: C, 57.75; H,
169.5; LCMS (m/z): 531.1 [M
+
H]+ for
C22H19N6O6SCl. Elem. Anal. for C22H19N6O6SCl:
(calcd) C, 49.77; H, 3.61; N, 15.83; S, 6.04. Found: C,
50.04; H, 3.98; N, 15.63; S, 5.99.
(1-(2-Chloro-5-nitrophenyl)-1H-1,2,3-triazol-4-yl)methyl-2-
(4-oxo-5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-
3(4H)-yl)acetate (7j). Brown solid. Yield: 60%; mp 170–
4.04; N, 15.98; S, 7.21.
(1-(3-Nitrophenyl)-1H-1,2,3-triazol-4-yl)methyl-2-(4-oxo-
5,6,7,8-tetrahydrobenzo[1,2]thieno[2,3-d]pyrimidin-3(4H)-yl)
acetate (7g). Brown solid. Yield: 90%; mp 135–140°C;
180°C; IR (KBr) (νmax, cmÀ1): 3108.6 (=CH), 2938
(ÀCH), 1751 (ester C=O), 1676 (amide C=O), 1610
(C=N); 1H NMR (400 MHz, CDCl3): δ 1.79–1.89 (m,
4H, 2CH2), 2.78 (t, J = 6.1 Hz, 2H, CH2), 2.95 (t,
J = 6.1 Hz, 2H, CH2), 4.71 (s, 2H, CH2), 5.48 (s, 2H,
CH2-triazole), 7.81 (d, J = 8.8 Hz, 1H, Ar–H), 7.87 (s,
1H, triazole-CH), 8.21 (s, 1H, pyrimidine-CH), 8.35 (d,
J = 6.2 Hz, 1H, Ar–H), 8.57 (d, J = 2.6 Hz, 1H, Ar–H);
IR (KBr) (νmax, cmÀ1): 3113 (=CH), 2932 (ÀCH), 1731
(ester C=O), 1670 (amide C=O), 1608 (C=N), 1527
(NO2); 1H NMR (400 MHz, CDCl3): δ 1.79–1.88 (m,
4H, 2CH2), 2.78 (t, J = 6.0 Hz, 2H, CH2), 2.95 (t,
J = 6.1 Hz, 2H, CH2), 4.71 (s, 2H, CH2), 5.47 (s, 2H,
CH2-triazole), 7.75 (t, J = 8.15 Hz, 1H, Ar–H), 7.89 (s,
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2017
Synthesis, Antibacterial Activity, and Docking Studies of 1,2,3-triazole-tagged
Thieno[2,3-d]pyrimidinone Derivatives
13C NMR (100 MHz, CDCl3): δ 21.5, 22.5, 23.7, 37.5,
42.3, 61.5, 109.0, 117.6, 119.1, 125.4, 127.3, 129.7,
133.3, 135.6, 138.6, 139.4, 144.4, 147.6, 155.6, 161.6,
169.5; LCMS (m/z): 501.1 [M + H]+, 523.1 [M + Na]+
for C21H17N6O5SCl. Elem. Anal. for C21H17N6O5SCl:
(calcd) C, 50.35; H, 3.42; N, 16.78; S, 6.40. Found: C,
50.40; H, 3.69; N, 16.64; S, 6.52.
Acknowledgments. One of the authors (MAK) is gratefully
acknowledge to University Grant Commission, New Delhi, for
providing financial assistance in the form of UGC-BSR (RFSMS
NO. F. 25-1/2013-14(BSR)/7-187/2007(BSR)) fellowship.
REFERENCES AND NOTES
In vitro antibacterial activity.
Evaluation of
[1] Dewal, B. M.; Wani, S. A.; Vidaillac, C.; Oupicky, D.; Rybak,
J. M.; Firestine, M. S. Eur J Med Chem 2012, 51, 145.
[2] Mitscher, L. A.; Pilla, S. P.; Gentry, E. J.; Shankel, D. M. Med
Res Rev 1999, 19, 477.
[3] Hubschwerlen, C.; Specklin, J. L.; Sigwalt, C.; Schroeder, S.;
Locher, H. H. Bioorg Med Chem 2003, 11, 2313.
[4] Veerachamy, A.; Durairaj, S.; Viswas, R. S. ARKIVOC 2006,
16, 149.
[5] Srujana, M.; Shirisha, T.; Jyothi, B.; Kasula, J. M. J Pharm Res
2014, 3, 66.
[6] Ashalatha, B. V.; Narayana, B.; Raj, K. K. V.; Kumari, N. S.
Eur J Med Chem 2007, 42, 719.
[7] Petrie, C. R.; Cottam, H. B.; Mckernan, P. A.; Robins, R. K.;
Revankar, G. R. J Med Chem 1985, 28, 1010.
[8] Chambhare, R. V.; Khadse, B. G.; Bobde, A. S.; Bahekar, R. H.
Eur J Med Chem 2003, 38, 89.
[9] Abbas, S. E.; Gawad, N. M. A.; George, R. F.; Akar, Y. A. Eur
J Med Chem 2013, 65, 195.
[10] El-Sherbeny, M. A.; El-Ashmawy, M. B.; El-Subbagh, H. I.;
El-Emam, A. A.; Badria, F. A. Eur J Med Chem 1995, 30, 445.
[11] Mulla, J. A. S.; Khazi, M. I. A.; Panchamukhi, S. I.; Gong, Y. D.;
Khazi, I. A. M. Med Chem Res 2014, 23, 3235.
[12] Hou, J.; Liu, X.; Shen, J.; Zhao, G.; Wang, P. G. Expert Opin
Drug Discovery 2012, 7, 489.
[13] Vatmurge, N. S.; Hazra, B. G.; Pore, V. S.; Shirazi, F.; Chavan,
P. S.; Deshpande, M. V. Bioorg Med Chem Lett 2008, 18, 2043.
[14] (a) Zhang, J.; Zhang, H.; Cai, W.-X.; Yu, L.-P.; Zhen, X.-C.;
Zhang, A. Bioorg Med Chem 2009, 17, 4873; (b) Jagasia, R.; Holub, J. M.;
Bollinger, M.; Krishenbaum, K.; Finn, M. G. J Org Chem 2009, 74,
2964; (c) Huber, D.; Hubner, H.; Gmeiner, P. J Med Chem 2009, 52, 6860.
[15] Chen, M. D.; Lu, S. J.; Yuag, G. P.; Yang, S. Y.; Du, X. L.
Heterocyclic Commun 2000, 6, 421.
[16] Buckle, D. R.; Outred, D. J.; Rockell, C. J. M.; Smith, H.;
Spicer, B. A. J Med Chem 1983, 26, 251.
[17] Menendez, C.; Chollet, A.; Rodriguez, F.; Inard, C.; Pasca,
M. R.; Lherbet, C. Eur J Med Chem 2012, 52, 275.
[18] Passannanti, P.; Diana, P.; Barraja, F.; Mingoia, A.; Lauria, G.
Cirrincione Heterocycles 1998, 48, 1229.
[19] Guantai, E. M.; Ncokazi, K.; Egan, T. J.; Gut, J.; Rosenthal, P. J.;
Smith, P. J.; Chibale, K. Design, Bioorganic Med Chem 2010, 18, 8243.
[20] Piotrowska, D. G.; Balzarini, J.; Glowacka, I. E. Eur J Med
Chem 2012, 47, 501.
[21] Manfredini, S.; Vicentini, C. B.; Manfrini, M.; Bianchi, N.;
Rutigliano, C.; Mischiati, C.; Gambari, R. Bioorg Med Chem 2000, 8,
2343.
antibacterial activities of the synthesized compounds was
carried out in 96-well micro titer plates using the broth
dilution method [27]. The bacterial cultures were grown
overnight at 37°C and serially diluted to a concentration
of 106 colony forming units/mL with sterile MHB
(Mueller-Hinton broth) medium. A different synthesized
compounds (1, 2.5, 5, 10, 25, 50, 75, 100 μg/mL) were
diluted with MHB medium was performed in triplicate
for each compound in a sterile 96-well plate to a final
volume of 50 μL in each well. The final volume was
adjusted to 250 μL by adding an aliquot of 100 μL
bacterial suspensions and 100 μL MHB broths to each
well, and the plates were incubated at 37°C for 18–20 h.
Controls
were
performed
without
synthesized
compounds, and the MIC of chemical compounds was
defined as the lowest concentration of the synthesized
compounds required for complete inhibition or killing the
bacterial inoculums. The growth inhibition was
monitored by measuring the absorbance at 492 nm. The
MIC values reported were reproducible between three
independent triplicates.
Docking assay.
The synthesized molecules and
standard Gentamicin were selected for performing
molecular docking studies. In this present study, GLIDE
5.6 [28] was used for molecular docking. Crystal
structures of B. subtilis Purine riboswitch (pdb id: 2XNW
[29]) and E. coli TPP riboswitch (pdb id: 4NYG [30])
were downloaded from protein data bank (http://www.
rcsb.org). The reason behind choosing pdb id’s 2XNW,
4NYG, is that these were crystallized with RNA, mRNAs
(termed riboswitches), which are targets for antibiotics and
RNA-ligand docking is a poorly explored area. Molecules
were built using Maestro build panel and prepared by Lig
Prep 2.0 application, which uses MMFF 94s [31] force
field and gave the corresponding low-energy 3D
conformers of the ligands. The protein was prepared using
protein preparation wizard of Schrödinger’s molecular
docking software. In this target preparation, all water
molecules were removed, and hydrogen atoms were added
to the target. Grid was generated around the active site of
the protein by selecting the active site amino acids.
Receptor Vander Waals scaling for the nonpolar atoms was
kept 0.9 [32]. Low-energy conformation of the molecules
were selected and docked into the grid using standard
precision (SP) docking protocol. Dock pose of molecules
were analyzed for interactions with the receptor.
[22] Danoun, S.; Baziard-Mouysset, G.; Stigliani, J.; Payard, M.;
Selkti, M.; Viossat, B.; Tomas, A. Heterocyclic Commun 1998, 4, 45.
[23] Rostovsev, V. V.; Green, L. G.; Fokin, V. V.; Sharpless, K. B.
Angew Chem Int Ed 2002, 41, 2596.
[24] Tornoe, C. W.; Christensen, C.; Meldal, M. J Org Chem 2002,
67, 3057.
[25] Gewald, K.; Neumann, G. Chem Ber 1968, 101, 1933.
[26] Sirisha, B.; Narasaiah, B.; Yakaiah, T.; Gayatri, G.; Narahari
Sastry, G.; Raghu Prasad, M.; Raghuram Rao, A. Eur J Med Chem
2010, 45, 1739.
[27] Shiva Shankar, S.; Benke, N. S.; Nagendra, N.; Srivastava, P. L.;
Thulasiram, V. H.; Gopi, N. H. J Med Chem 2013, 56, 8468.
[28] Schrödinger, L. L. C. Glide, Version 4.0: New York, NY, 2005.
[29] Peter, D.; Francis, E. R.; David, A. R.; Colin, M. H.; David,
M. L.; Robert, T. B.; Ruth, B. Chem Biol 2011, 18, 324.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

M. Aruna Kumari, S. Triloknadh, N. Harikrishna, M. Vijjulatha, and C. Venkata Rao
Vol 000
[30] Katherine, D. W.; Philip, H.; Kevin, M. W.; Alison, G. S.;
Chris, A.; Adrian, R. F. -D. A. Chem Biol 2014, 21, 591.
SUPPORTING INFORMATION
[31] Thomas, A. H. J Comput Chem 1999, 20, 720.
Additional Supporting Information may be found online
in the supporting information tab for this article.
[32] Richard, A. F.; Jay, L. B.; Robert, B. M.; Thomas, A. H.; Jasna,
J. K.; Daniel, T. M.; Matthew, P. R.; Eric, H. K.; Mee, S.; Jason, K. P.;
David, E. S.; Perry, F.; Peter, S. S. J Med Chem 2004, 47, 1739.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet