Solution-phase microwave assisted parallel synthesis, biological evaluation and in silico docking studies of N,N′-disubstituted thioureas derived from 3-chlorobenzoic acid

Article abstract of DOI:10.1016/j.bmc.2016.07.042

A facile and robust microwave-assisted solution phase parallel synthesis protocol was exercised for the development of a 38-member library of N,N′-disubstituted thiourea analogues (1–38) by using an identical set of conditions. The reaction time for synthesis of N,N′-disubstituted thiourea analogues was drastically reduced from a reported duration of 8–12 h for conventional methods to only 1.5–2.0 min. All the derivatives (1–38) were characterized by physico-analytical techniques such as elemental analysis in combination with FT-IR,¹H,¹³C NMR and by single crystal XRD analysis have also been performed. These compounds were screened for their in vitro urease inhibition activities. Majority of compounds exhibited potent urease inhibition activities, however, the most significant activity was found for 16, with an IC₅₀value of 1.23 ± 0.1 μM. Furthermore, the synthesized compounds were screened for their cytotoxic potential against lungs cancer cell lines. Cell culture studies demonstrated significant toxicity of the compounds on the cell lines, and the levels of toxicity were altered in the presence of various side groups. The molecular docking studies of the most potent inhibitors were performed to identify the probable binding modes in the active site of the urease enzymes. These compounds have a great potential and significance for further investigations.

Full text of DOI:10.1016/j.bmc.2016.07.042

Bioorganic & Medicinal Chemistry xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Solution-phase microwave assisted parallel synthesis, biological
evaluation and in silico docking studies of N,N0-disubstituted
thioureas derived from 3-chlorobenzoic acid
Muhammad Khawar Rauf a, Sumera Zaib b, Ammara Talib a, Masahiro Ebihara c, Amin Badshah a,
,
⇑
Michael Bolte d, Jamshed Iqbal b,
⇑
a Department of Chemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan
b Centre for Advanced Drug Research, COMSATS Institute of Information Technology, Abbottabad 22060, Pakistan
c Department of Chemistry, Faculty of Engineering, Gifu University, Yanagido, Gifu 501-1193, Japan
d Institut für Anorganische Chemie, J.W. Goethe-Universität Frankfurt, Max-von-Laue-Str. 7, 60438 Frankfurt/Main, Germany
a r t i c l e i n f o
a b s t r a c t
Article history:
A facile and robust microwave-assisted solution phase parallel synthesis protocol was exercised for the
development of a 38-member library of N,N0-disubstituted thiourea analogues (1–38) by using an iden-
tical set of conditions. The reaction time for synthesis of N,N0-disubstituted thiourea analogues was dras-
tically reduced from a reported duration of 8–12 h for conventional methods to only 1.5–2.0 min. All the
derivatives (1–38) were characterized by physico-analytical techniques such as elemental analysis in
combination with FT-IR, 1H, 13C NMR and by single crystal XRD analysis have also been performed.
These compounds were screened for their in vitro urease inhibition activities. Majority of compounds
exhibited potent urease inhibition activities, however, the most significant activity was found for 16, with
Received 24 May 2016
Revised 19 July 2016
Accepted 20 July 2016
Available online xxxx
Keywords:
Cytotoxicity
Microwave assisted parallel synthesis
N,N0-Disubstituted thioureas
Single crystal analysis
Urease inhibition
an IC50 value of 1.23 0.1 lM. Furthermore, the synthesized compounds were screened for their cyto-
toxic potential against lungs cancer cell lines. Cell culture studies demonstrated significant toxicity of
the compounds on the cell lines, and the levels of toxicity were altered in the presence of various side
groups. The molecular docking studies of the most potent inhibitors were performed to identify the prob-
able binding modes in the active site of the urease enzymes. These compounds have a great potential and
significance for further investigations.
Ó 2016 Elsevier Ltd. All rights reserved.
1. Introduction
agriculture.5–8 To counteract these negative effects, inhibition of
urease is an attractive strategy and a diverse variety of urease inhi-
Urease (urea amidohydrolase; E.C.3.5.1.5), a nickel-containing
metalloenzyme with an ability to catalyze the hydrolysis of urea
to ammonia and carbamates, is considered as an important viru-
lence factor in the pathogenesis of several diseases.1,2 It is predom-
inantly found in numerous living organisms, such as bacteria,
fungi, higher plants, and some invertebrates.3,4 Urease is important
for human and animal health due to its involvement in the devel-
opment of several infections associated with stones and plays a
vibrant role in pathogenesis of urolithiasis, pyelonephritis, and
hepatic encephalopathy.4 In addition, high concentration of ammo-
nia from the hydrolysis reaction as well as elevated pH level also
imparts several negative effects in the fields of medicine and
bitors have been reported1,9–12, however, due to their toxicity or
instability, the development of potent urease inhibitors with fewer
toxic effects is highly needed.
Similarly, cancer still remains a potentially life-threatening dis-
ease and is the second cause of death in the US, with an expected
1,665,540 cancer cases in 2014. The high volume of data related to
cancer deaths indicates that 90% of the patients die due to chronic
tumor metastases.13 Cancer is chronic disease, initiated by com-
plex processes, which are directly associated to a fundamental
cycle of life, i.e., cell division and may be induced by exogenous
agents and is characterized by uncontrolled cell proliferation lead-
ing to tumor formation.13,14 Several external (chemicals, radiation,
infectious microorganisms, and tobacco) and internal (hormones,
mutations and immune conditions) factors contribute enormously
to the eruption of cancers.14,15 Despite the development of several
anticancer drugs, the continued efforts to identify new anticancer
⇑
Corresponding authors.
E-mail addresses: aminbadshah@yahoo.com (A. Badshah), drjamshed@ciit.net.pk
(J. Iqbal).
http://dx.doi.org/10.1016/j.bmc.2016.07.042
0968-0896/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

2
M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
agents with less side effects and toxicity is the subject of current
medicinal chemistry research.16,17
power settings. The experiments were carried out in standard
Pyrex capable glass vials (20 mL).
Aryl/aroyl substituted thioureas have found potential applica-
tions for the structural developments in heterocyclic chemistry
and modifications with remarkable pharmacological and biological
implications. In the past few decades, in vitro studies have shown
that polyfunctional thioureas are potential herbicides,18 insecti-
cides,19 plant-growth regulators20 and antihypertensive agents,21
antitubercular, antimalarial, anticancer,22 antithyroid, anti-
helmintic and TRPV1 receptor antagonists.23 Thioureas may be
used as precursors to yield active compounds of pharmacological
importance by treating with other intermediates bearing different
NMR spectra were recorded on a Bruker ARX, 300 MHz spec-
trometer. 1H NMR (300.13 MHz): internal standard solvent CDCl3
(7.28 ppm from TMS): internal standard TMS; 13C NMR
(75.47 MHz): internal standard solvent CDCl3 (77.0 ppm from
TMS): internal standard TMS; the splitting of proton resonances
in the reported 1H NMR spectra are defined as s = singlet, d = dou-
blet, t = triplet, q = quartet and m = complex pattern; coupling con-
stants are reported in Hz. FT-IR spectra were recorded as KBr
pellets on a Bio-Rad Excalibur FT-IR Model FTS 3000 MX (400–
4000 cmꢀ1) and the elemental analyses were performed using a
LECO-932 CHNS analyzer. The melting points were determined
on a Bio Cote SMP10-UK and are uncorrected. The chemicals like
3-chlorobenzoic acid, ammonium thiocyanate, thionyl chloride
and primary/secondary amines, were purchased from Sigma–
Aldrich and used as received.
functionalities. For example, thioureas condense with
a-halocar-
bonyl compounds to produce 2-amino-1,3-thiazoles24 and arylth-
ioureas can be converted into benzothiazoles in the presence of
bromine as an oxidizing agent.25 The synthesis of iminothiazoli-
nes,26 thiohydantoins,27 1,3,5-triazines28 and 2-amino-oxazolidi-
nes29 from thioureas has been well explored.
Mostly, N,N0-disubstituted thioureas are synthesized by treating
primary and secondary amines with Na+/K+/NH4+ thiocyanates,21
aroyl isothiocyanates with primary and secondary amines followed
by acid hydrolysis30 and isothiocyanates with ammonia or
amines.31 The de-benzoylation of N-substituted and N,N0-disubsti-
tuted thioureas with hydrazine hydrate under solvent-free condi-
2.2. Microwave assisted solution-phase parallel synthesis of N,
N0-disubstituted thioureas (1–38)
Freshly prepared 3-chlorobenzoyl isothiocyanate was mixed in
a 1:1 molar ratio with the desired primary amines in dry THF, in
half capped cylindrical glass vessels. The reaction mixtures were
subjected to microwave heating using the low absorption mode
at 60–65 °C for about 1.5–2.0 min. On cooling, the reaction mix-
tures were slowly poured into HCl acidified (pH 4–5) chilled water
in to ditto labeled beakers (100 mL) and stirred well with a glass
rod. The solids were separated by filtration and dried at room tem-
perature (Scheme 1). The yields were above 85% for each of the
substituted aroylthioureas (1–38).
tions yields mono- and N,N0-disubstituted thioureas32 and
a
synthesis of unsubstituted thioureas with primary alkyl amines
at high temperature have also been reported.33 In the recent dec-
ades, numerous new methodologies have been exercised for the
design and synthesis of N,N0-substituted thioureas.34 However,
many of these methods are not very popular due to some disad-
vantages, specifically, long reaction time under high thermal con-
ditions and the use of large quantity of solvents. The
development of efficient, mild and environment friendly protocols
are still need of the day. The microwave supported organic synthe-
ses have received great attention in the recent years that greatly
facilitated the organic chemists to increase the number and purity
of the products. It has also inherent advantages of high chemical
yield and shorter reaction time. Recently, solvent free microwave
assisted synthesis of substituted thioureas have been reported.35–
2.3. Spectroscopic data (1–38)
The spectroscopic data for the compounds (1–38) is given
below.
2.3.1. 1-(3-Chlorobenzoyl)-3-phenylthiourea (1)
Yield 88%; white solid; mp 135–136 °C; FT-IR (KBr, cmꢀ1) 3217,
3134, 3014, 1972, 1590, 1530, 1329, 1250, 1157, 1016, 869, 716,
692, 542, 490; 1H NMR (300 MHz, CDCl3) d 7.31 (t, 1H, J = 7.2 Hz,
37
These solvent free syntheses have many objections regarding
purity of the products due to the presence of inorganic byproducts,
non-uniform heating and restricted molecular activities in the vis-
cous phase. Keeping in view these issues and in perpetuation to our
previous work on different types of thioureas, their coordination
and medicinal chemistry,38,39 we report here the microwave
assisted solution-phase parallel synthesis of N,N0-disubstituted
thioureas and their biological activities such as urease inhibition
both in vitro & in virtual and anticancer activities because of their
inherent drug like significance. We speculated that our designed
thiourea compounds could effectively inhibit the urease due to
structural similarity with the natural substrate of urease, i.e., urea
and the diverse structural features introduced on both diversity
points of thiourea would probably exert positive effects on activity
through the binding of molecules to active site of the enzyme. The
results of this study are presented in the subsequent discussion.
0
ArHa), 7.45 (t, 2H, J = 8.1 Hz, ArHb@b ), 7.51 (t, 1H, J = 7.8 Hz, ArHe),
0
7.64 (d, 2H, J = 8.1 Hz, ArHf), 7.74 (d, 2H, J = 8.1 Hz, ArHc@c ), 7.77
0
(d, 1H, J = 7.5 Hz, ArHd), 7.91 (s, 1H, ArHd ), 9.06 (s, CONH), 12.47
(s, CSNH); 13C NMR (75 MHz, CDCl3) d 124.16 (2C), 125.33,
127.03, 127.98, 128.99 (2C), 130.53, 133.42, 133.80,135.66,
137.47 (Ar-C), 165.61 (C@O), 178.03 (C@S); Anal. Calcd for C14H11
-
N2OSCl; C, 57.83; H, 3.81; N, 9.63; S, 11.03 Found: C, 57.81; H, 3.80;
N, 9.65; S, 11.00.
2.3.2. 1-(3-Chlorobenzoyl)-3-(2-chlorophenyl)thiourea (2)
Yield 92%; white solid; mp 131–132 °C; FT-IR (KBr, cmꢀ1) 3285,
3236, 3022, 1676, 1658, 1591, 1531, 1439, 1347, 1256, 1158, 1079,
876, 769, 746, 692, 563, 493; 1H NMR (300 MHz, CDCl3) d 7.27 (d,
1H, J = 7.5 Hz, ArHc), 7.37 (t, 1H, J = 7.8 Hz, ArHa), 7.48–7.54 (m, 2H,
0
ArHe, b), 7.64 (d, 1H, J = 8.1 Hz, ArHb ), 7.79 (d, 1H, J = 7.8 Hz,
0
ArHf),7.94 (s, 1H, ArHd ), 8.41 (d, 1H, J = 7.8 Hz, ArHd) 9.15 (s,
CONH), 12.64 (s, CSNH); 13C NMR (75 MHz, CDCl3) d 125.43,
126.13, 126.98, 127.86, 127.98, 128.10, 129.73, 130.52, 133.25,
133.86, 135.89, 135.67 (Ar-C), 165.42 (C@O), 178.32 (C@S); Anal.
Calcd for C14H10N2OSCl2; C, 57.70; H, 3.10; N, 8.61; S, 9.86 Found:
C, 57.70; H, 3.10; N, 8.63; S, 9.85.
2. Experimental
2.1. Materials and methods
All experiments were carried out under the specified conditions
of temperature and normal pressure. Solvents were purified and
distilled from the drying agents, stored over molecular sieves 4 Å
and degassed before use. Microwave syntheses were carried out
using a CEM MARS 6 microwave synthesizer with low absorption
2.3.3. 1-(3-Chlorobenzoyl)-3-(3-chlorophenyl)thiourea (3)
Yield 95%; white solid; mp 132–133 °C; FT-IR (KBr, cmꢀ1) 3310,
3232, 3048, 1657, 1593, 1530, 1423, 1252, 1159, 1078, 893, 779,
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
3
No.
R
R' No.
R
R' No.
R
No.
R
R'
c′
b′
c′
Cl
Cl
O2N
b′
b′
b′
a′
a
1
H
11
Cl
Cl
H
21
H
31
H
NO2
g
a
c
c
b
c
c
b
Cl
c
b
b
Cl
H3CO
b′
b′
b′
N
S
a
a
a
H
H
H
H
H
H
H
2
3
12
13
22
23
32
33
a′
b
c
b
b
Cl
Cl
b
′
c′
Cl
OCH3
b′
Cl
c′
c′
a′
a
a
a
CH3
Cl
c
Cl
b
c
c
a
b
c′
c′
b′
F3C
b′
b′
c′
b′
H2
a
CH3
Cl
4
H
14
H
24
a
H
34
H
H
C
c
b
c
c
b
b
c
b
CF3
Cl
Cl
Cl
H3C
CH3
c′
b′
H2
C
a
a
a
H
H
H
H
H
H
H
H
H
5
6
7
15
16
17
25
26
27
a
35
36
37
c
c
b
b
c
b
c
b
b′
c′
Cl
c′
b′
b′
b′
a
CF3
H
c′
c
Cl
c
b
b
c
c
b
b
Cl
c′
Br
b′
c′
b′
b′
b′
c′
a
a
a
a
c
b
c
c
b
b
b
c
Cl
Cl
Br
b′
b′
b′
b′
N
N
a
a
H
H
H
H
8
18
28
38
Br
b
b
b
c
b
Cl
O2N
c′
b′
b′
Cl
c′
a
F
9
H
H
19
20
H
H
29
30
H
H
Cl
c
c
b
b
c
b
Cl
c′
c′
F
c′
b′
a
a
NO2
10
c
c
c
b
b
Cl
Scheme 1. Synthesis of N,N0-disubstituted thioureas (1–38).
757, 676565, 484; 1H NMR (300 MHz, CDCl3) d 7.29 (d, 1H,
J = 6.9 Hz, ArHc), 7.36 (t, 1H, J = 8.1 Hz, ArHb), 7.51 (t, 1H,
J = 8.1 Hz, ArHe), 7.59 (d, 1H, J = 8.1 Hz, ArHa), 7.65 (d, 1H,
J = 8.1 Hz, ArHf), 7.76 (d, 1H, J = 8.1 Hz, ArHd), 7.87 (s, 1H,
ArHf, b ), 7.77 (d, 1H, J = 7.8 Hz, ArHd), 7.91 (s, 1H, ArHd ), 9.18
0
0
b,
(s, CONH), 12.50 (s, CSNH); 13C NMR (75 MHz, CDCl3) d 125.41
(2C), 128.02, 129.09 (2C), 130.53, 132.35, 133.24, 133.28, 135.65,
135.97 (Ar-C), 165.78 (C@O), 178.21 (C@S); MS (EI) m/z 323.6
(M+), 138.8 (base peak); Anal. Calcd for C14H10N2OSCl2; C, 57.70;
H, 3.10; N, 8.61; S, 9.86 Found: C, 57.69; H, 3.09; N, 8.65; S, 9.87.
J = 8.1 Hz, ArHd ), 9.09 (s, CONH), 12.53 (s, CSNH); 13C NMR
0
(75 MHz, CDCl3) d 122.19, 124.10, 125.38, 127.09, 128, 129.93,
130.56, 133.21, 133.92, 134.50, 135.69, 138.55 (Ar-C), 165.71
(C@O), 178.10 (C@S); Anal. Calcd for C14H10N2OSCl2; C, 57.70; H,
3.10; N, 8.61; S, 9.86 Found: C, 57.69; H, 3.09; N, 8.62; S, 9.86.
2.3.5. 1-(3-Chlorobenzoyl)-3-(2,3-dichlorophenyl)thiourea (5)
Yield 85%; white solid; mp 183–184 °C; FT-IR (KBr, cmꢀ1) 3229,
3152, 3022, 1677, 1586, 1518, 1344 1251, 1160, 1160, 875, 787,
696, 459; 1H NMR (500 MHz, DMSO-d6) d 7.43 (t, 1H, J = 8.0 Hz,
ArHb), 7.55–7.60 (m, 2H, ArHe, c), 7.73 (d, 1H, J = 8.0 Hz, ArHa),
7.89 (d, 1H, J = 8.1 Hz, ArHf), 8.0 (d, 1H, J = 8.0 Hz, ArHd), 8.04 (s,
2.3.4. 1-(3-Chlorobenzoyl)-3-(4-chlorophenyl)thiourea (4)
Yield 96%; white solid; mp 166–167 °C; FT-IR (KBr, cmꢀ1) 3214,
3046, 1667, 1605, 1532, 1487, 1358, 1260, 1161, 1090, 1005, 874,
826, 697, 602, 503; 1H NMR (300 MHz, CDCl3) d 7.40 (d, 2H,
1H, ArHd ), 12.01 (s, CONH), 12.51 (s, CSNH); 13C NMR (125 MHz,
0
0
J = 9.0 Hz, ArHc@c ), 7.50 (t, 1H, J = 7.8 Hz, ArHe), 7.63–7.69 (m, 3H,
DMSO-d6) d 127.7, 128.0, 128.2, 129.0, 129.2, 130.8, 132.4, 133.4,
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

4
M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
133.6, 134.3, 138.0 (Ar-C), 167.6 (C@O), 180.9 (C@S); Anal. Calcd
for C14H9N2OSCl3; C, 46.75; H, 2.52; N, 7.79; S, 8.92 Found: C,
46.75; H, 2.51; N, 7.80; S, 8.92.
2.3.11. 1-(3-Chlorobenzoyl)-3-(2,4,5-trichlorophenyl)thiourea
(11)
Yield 88%; white solid; mp 213–214 °C; FT-IR (KBr, cmꢀ1) 3295,
3086, 3068, 2990, 1982, 1564, 1522, 1470, 1368, 1308,1252, 1155,
1130, 1079, 873, 737, 683, 606, 559, 478; 1H NMR (500 MHz,
DMSO-d6) d 7.56 (t, 1H, J = 8.0 Hz, ArHe), 7.71 (d, 1H, J = 7.5 Hz,
ArHf), 7.92 (d, 1H, J = 7.5 Hz, ArHd), 7.98 (s, 1H, ArHc), 8.03 (s, 1H,
2.3.6. 1-(3-Chlorobenzoyl)-3-(2,4-dichlorophenyl)thiourea (6)
Yield 95%; white solid; mp 189–190 °C; FT-IR (KBr, cmꢀ1) 3258,
3092, 3001, 1686, 1578, 1527, 1469, 1323, 1248, 1161, 1094, 866,
812, 723, 693, 601553, 443; 1H NMR (500 MHz, DMSO-d6) d 7.49
(d, 1H, J = 8.6 Hz, ArHc), 7.57 (t, 1H, J = 8.0 Hz, ArHe), 7.73 (d, 1H,
ArHb ), 8.41 (s, 1H, ArHd ), 12.00(s, CONH), 12.52 (s, CSNH); 13C
NMR (125 MHz, DMSO-d6) d 127.9, 128.0, 128.6, 128.8, 129.0,
130.0. 130.8, 131.0, 133.5, 133.6, 134.2, 136.1 (Ar-C), 167.7
(C@O), 180.7 (C@S); MS (EI) m/z 356.6 (M+ = M-Cl), 138.8 (base
peak); Anal. Calcd for C14H8N2OSCl4; C, 42.67; H, 2.05; N, 7.11; S,
8.14 Found: C, 42.66; H, 2.00; N, 7.13; S, 8.13.
0
0
0
J = 8.0 Hz, ArHb), 7.75 (s, 1H, ArHb ), 7.92 (d, 1H, J = 8.0 Hz, ArHf),
0
8.02 (d, 1H, J = 8.6 Hz, ArHd), 8.04 (s, 1H, ArHd ), 12.01 (s, CONH),
12.49 (s, CSNH); 13C NMR (125 MHz, DMSO-d6) d 127.9, 128.0,
129.0, 129.5, 129.8, 130.2, 130.8, 132.0, 133.4, 133.6, 134.4, 135.2
(Ar-C), 167.6 (C@O), 180.7 (C@S); MS (EI) m/z 359.5 (M+), 138.8
(base peak); Anal. Calcd for C14H9N2OSCl3; C, 46.75; H, 2.52; N,
7.79; S, 8.92 Found: C, 46.74; H, 2.50; N, 7.81; S, 8.91.
2.3.12. 1-(3-Chlorobenzoyl)-3-(2,4,6-trichlorophenyl)thiourea
(12)
Yield 95%; white solid; mp 179–180 °C; FT-IR (KBr, cmꢀ1
)
2.3.7. 1-(3-Chlorobenzoyl)-3-(2,5-dichlorophenyl)thiourea (7)
Yield 86%; white solid; mp 201–202 °C; FT-IR (KBr, cmꢀ1) 3244,
3095, 2988, 1680, 1590, 1545, 1528, 1408, 1344, 1256, 1165, 1088,
898, 812, 704, 545; 1H NMR (500 MHz, DMSO-d6) d 7.42 (d, 1H,
J = 8.6 Hz, ArHa), 7.57 (t, 1H, J = 8.0 Hz, ArHe), 7.63 (d, 1H,
3412, 3147, 3040, 1677, 1571, 1482, 1341, 1234, 1159, 1079,
880, 818, 728, 652, 565, 490; 1H NMR (300 MHz, CDCl3) d 7.46
0
(s, 2H, ArHb@b ), 7.51 (t, 1H, J = 8.1 Hz, ArHe), 7.66 (d, 1H,
0
J = 7.8 Hz, ArHf), 7.77 (d, 1H, J = 7.8 Hz, ArHd), 7.93 (s, 1H, ArHd ),
9.29 (s, CONH), 11.86 (s, CSNH); 13C NMR (75 MHz, CDCl3) d
125.49 (2C), 128.19, 128.71 (2C), 130.53, 132.16, 133.02, 134.01,
134.89, 135.01, 135.70 (Ar-C), 165.76 (C@O), 180.42 (C@S); Anal.
Calcd for C14H8N2OSCl4; C, 42.67; H, 2.05; N, 7.11; S, 8.14 Found:
C, 42.64; H, 2.01; N, 7.12; S, 8.14.
0
J = 8.5 Hz, ArHb ), 7.73 (d, 1H, J = 8.6 Hz, ArHf), 7.93 (d, 1H,
0
J = 8.0 Hz, ArHd), 8.05 (s, 1H, ArHc), 8.22 (s, 1H, ArHd ), 12.05 (s,
CONH), 12.50 (s, CSNH); 13C NMR (125 MHz, DMSO-d6) d 127.6,
127.7, 128.0, 128.4, 129.0, 130.9, 131.3, 131.7, 133.4, 133.6,
134.4, 137.1 (Ar-C), 167.7 (C@O), 180.6 (C@S); Anal. Calcd for C14
-
H9N2OSCl3; C, 46.75; H, 2.52; N, 7.79; S, 8.92 Found: C, 46.75; H,
2.51; N, 7.83; S, 8.93.
2.3.13. 1-(3-Chlorobenzoyl)-3-(2,3,5,6-tetrachlorophenyl)
thiourea (13)
Yield 89%; white solid; mp 201–202 °C; FT-IR (KBr, cmꢀ1) 3487,
3416, 3119, 3046, 1677, 1568, 1512, 1470, 1341, 1242, 1157, 883,
740, 715, 589, 415; 1H NMR (300 MHz, CDCl3) d 7.55 (t, 1H,
J = 7.8 Hz, ArHe), 7.66–7.68 (m, 2H, ArHa, f), 7.8 (d, 1H, J = 7.8 Hz,
2.3.8. 1-(3-Chlorobenzoyl)-3-(2,6-dichlorophenyl)thiourea (8)
Yield 87%; white solid; mp 200–201 °C; FT-IR (KBr, cmꢀ1) 3152,
3068, 1677,1564, 1514, 1250, 1159, 875, 789, 720, 664, 462; 1H
NMR (300 MHz, CDCl3) d 7.31 (t, 1H, J = 7.5 Hz, ArHa), 7.46 (d, 2H,
ArHd), 7.95 (s, 1H, ArHd ), 9.26 (s, CONH), 12.02 (s, CSNH); 13C
0
0
J = 8.1 Hz, ArHb@b ), 7.51 (t, 1H, J = 8.1 Hz, ArHe), 7.65 (d, 1H,
NMR (75 MHz, CDCl3) d 125.5 (2C), 128.19 (2C), 130.58, 130.68,
132.06, 132.27, 134.10, 135.76 (Ar-C), 165.82 (C@O), 180.01
(C@S); Anal. Calcd for C14H7N2OSCl5; C, 39.24; H, 1.65; N, 6.54; S,
7.48 Found: C, 39.28; H, 1.65; N, 6.55; S, 7.48.
J = 7.8 Hz, ArHf), 7.80 (d, 1H, J = 7.8 Hz, ArHd), 9.28 (s, CONH),
11.90 (s, CSNH); 13C NMR (75 MHz, CDCl3) d 125.49 (2C), 128.19,
128.64 (2C), 129.80, 130.51, 133.18, 133.26, 133.91, 134.40,
135.67 (Ar-C), 165.69 (C@O), 180.39 (C@S); Anal. Calcd for C14H9-
N2OSCl3; C, 46.75; H, 2.52; N, 7.79; S, 8.92 Found: C, 46.74; H,
2.50; N, 7.81; S, 8.91.
2.3.14. 1-(3-Chlorobenzoyl)-3-(4-methylphenyl)thiourea (14)
Yield 95%; white solid; mp 153–154 °C; FT-IR (KBr, cmꢀ1) 3307,
3046, 1672, 1557, 1467, 1427, 1358, 1276, 1250, 1148, 1078, 815,
722, 680, 604, 508, 418; 1H NMR (300 MHz, CDCl3) d 2.39 (s, 3H, p-
2.3.9. 1-(3-Chlorobenzoyl)-3-(3,4-dichlorophenyl)thiourea (9)
Yield 88%; white solid; mp 176–177 °C; FT-IR (KBr, cmꢀ1) 3242,
3096, 3028, 1666, 1607, 1531, 1474, 1356, 1300, 1260, 1162, 1132,
897, 874, 810, 683, 436; 1H NMR (300 MHz, CDCl3) d 7.48–7.51 (m,
2H, ArHc, b), 7.57 (t, 1H, J = 8.7 Hz, ArHe), 7.67 (d, 1H, J = 7.8 Hz,
0
CH3), 7.25 (d, 2H, J = 8.4 Hz, ArHc@c ), 7.50 (t, 1H, J = 8.1 Hz, ArHe),
0
7.57 (d, 2H, J = 8.4 Hz, ArHb@b ), 7.63 (d, 1H, J = 7.8 Hz, ArHf), 7.77
0
(d, 1H, J = 7.8 Hz, ArHd), 7.91 (s, 1H, ArHd ), 9.13 (s, CONH), 12.37
(s, CSNH); 13C NMR (75 MHz, CDCl3) d 20.51 (o-CH3), 124.23 (2C),
125.36, 128.00, 129.58 (2C), 130.50, 133.46, 133.74, 134.89,
135.61, 137.1 (Ar-C), 165.62(C@O), 178.18 (C@S); Anal. Calcd for
0
ArHf), 7.77 (d, 1H, J = 7.8 Hz, ArHd), 7.90 (s, 1H, ArHc ), 7.99 (s, 1H,
ArHd ) 9.07 (s, CONH), 12.55 (s, CSNH); 13C NMR (75 MHz, CDCl3)
0
d 123.30, 125.37, 125.65, 127.99, 130.50, 130.59, 132.79, 133.08,
134.01, 135.74, 136.79 (Ar-C), 165.76 (C@O), 178.13 (C@S); Anal.
Calcd for C14H9N2OSCl3; C, 46.75; H, 2.52; N, 7.79; S, 8.92 Found:
C, 46.75; H, 2.51; N, 7.82; S, 8.92.
C15H13N2OSCl; C, 59.11; H, 4.30; N, 9.19; S, 10.52 Found: C,
59.10; H, 4.29; N, 9.21; S, 10.51.
2.3.15. 1-(3-Chlorobenzoyl)-3-(2,3-dimethylphenyl)thiourea
(15)
2.3.10. 1-(3-Chlorobenzoyl)-3-(3,5-dichlorophenyl)thiourea
(10)
Yield 96%; white solid; mp 161–162 °C; FT-IR (KBr, cmꢀ1) 3217,
3065, 2935, 1674, 1566, 1514, 1381, 1330, 1247, 1169, 1064, 832,
723, 602, 550, 444; 1H NMR (300 MHz, CDCl3) d 2.25 (s, 3H, m-CH3),
2.36 (s, 3H, o-CH3) 7.17–7.23 (m, 2H, ArHa, c), 7.42 (t, 1H, J = 7.1 Hz,
ArHb), 7.51 (t, 1H, J = 8.1 Hz, ArHe), 7.65 (d, 1H, J = 8.1 Hz, ArHf),
7.80 (d, 1H, J = 8.1 Hz, ArHd), 9.21 (s, CONH), 12.10 (s, CSNH); 13C
NMR (75 MHz, CDCl3) d 14.24 (m-CH3), 20.51 (o-CH3), 124.44,
125.41,125.91, 128.09, 129.59, 130.49, 132.58, 133.44, 133.75,
135.65, 138.10 (Ar-C), 165.73 (C@O), 179.63 (C@S); MS (EI) m/z
317.8 (M+), 110.9 (base peak); Anal. Calcd for C16H15N2OSCl; C,
60.28; H, 4.74; N, 8.79; S, 10.06 Found: C, 60.27; H, 4.74; N, 8.80;
S, 10.01.
Yield 86%; white solid; mp 185–186 °C; FT-IR (KBr, cmꢀ1) 3241,
3089, 3050, 1673, 1588, 1565, 1531, 1445, 1351, 1267, 1160, 1119,
852, 797, 732, 689, 559, 431; 1H NMR (500 MHz, DMSO-d6) d 7.50
(s, 1H, ArHa), 7.57 (t, 1H, J = 8.0 Hz, ArHe), 7.73 (d, 1H, J = 8.0 Hz,
0
ArHf), 7.81 (s, 2H, ArHc, c ), 7.91 (d, 1H, J = 8.0 Hz, ArHd), 8.02 (s,
1H, ArHd ), 11.87 (s, CONH), 12.44 (s, CSNH); 13C NMR (125 MHz,
0
DMSO-d6) d 123.7 (2C), 126.2, 127.9, 128.9, 130.9, 133.4, 133.6,
134.5, 140.9 (Ar-C), 167.0 (C@O), 180.0 (C@S); Anal. Calcd for C14
H9N2OSCl3; C, 46.75; H, 2.52; N, 7.79; S, 8.92 Found: C, 46.73; H,
2.49; N, 7.81; S, 8.92.
-
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
5
2.3.16. 1-(3-Chlorobenzoyl)-3-(2,4,6-trimethylphenyl)thiourea
(16)
Yield 90%; white solid; mp 160–161 °C; FT-IR (KBr, cmꢀ1) 3165,
2.3.21. 1-(3-Chlorobenzoyl)-3-(2, 4-dinitrophenyl)thiourea (21)
Yield 83%; yellow solid; mp 161–162 °C; FT-IR (KBr, cmꢀ1
3450, 3335, 3203, 3107, 1691, 1594, 1557, 1510, 1324, 1258,
1161, 1107, 881, 853, 752, 732, 676, 608, 496, 462; 1H NMR
(300 MHz, CDCl3) d 6.95 (d, 1H, J = 7.8 Hz, ArHc), 7.45 (t, 1H,
J = 8.1 Hz, ArHe), 7.760 (d, 1H, J = 8.1 Hz, ArHf), 7.75 (d, 1H,
)
3026, 2917, 1675, 1531, 1514, 1341, 1250, 1171, 1147, 1070, 860,
748, 721, 701, 655, 615, 436; 1H NMR (500 MHz, CDCl3) d 2.27 (s,
0
6H, 2[o-CH3]), 2.32 (s, 3H, p-CH3), 6.96 (s, 2H, ArHb@b ), 7.48 (t,
0
1H, J = 8.1 Hz, ArHe), 7.62 (d, 1H, J = 6.9 Hz, ArHf), 7.78 (d, 1H,
J = 7.8 Hz, ArHd), 7.87 (s, 1H, ArHd ), 8.21 (d, 2H, J = 9.1 Hz, ArHb),
9.11 (s, 1H, ArHb ), 9.25 (s, CONH), 13.3 (s, CSNH); 13C NMR
0
0
J = 6.9 Hz, ArHd), 7.93 (s, 1H, ArHd ), 9.35 (s, CONH), 11.69 (s, CSNH);
13C NMR (125 MHz, CDCl3) d 18.0 (2C, o-CH3), 21.01 (p-CH3), 125.4,
128.0 (2C), 129.0 (2C), 130.3, 132.6, 133.3, 133.6, 134.8, 135.4,
138.0 (Ar-C), 165.7 (C@O), 179.8 (C@S); Anal. Calcd for C17H17N2-
OSCl; C, 61.34; H, 5.15; N, 8.42; S, 9.63 Found: C, 61.34; H, 5.14;
N, 8.44; S, 9.64.
(75 MHz, CDCl3) d 122.3, 125.5, 128.3, 128.9, 130.8, 131.6, 133.2,
134.3, 135.6, 143.2, 145.3, 147.2 (Ar-C), 166.3 (C@O), 179.3
(C@S); Anal. Calcd for C14H9N4O5SCl; C, 44.16; H, 2.38; N, 14.71;
S, 8.42 Found: C, 44.14; H, 2.35; N, 14.73; S, 8.40.
2.3.22. 1-(3-Chlorobenzoyl)-3-(2-methoxyphenyl)thiourea (22)
Yield 89%; white solid; mp 131–132 °C; FT-IR (KBr, cmꢀ1) 3256,
3013, 2941, 1678, 1904, 1558, 1535, 1487, 1464, 1364, 1316, 1246,
1180, 1153, 1111, 1022, 739, 708, 683, 488; 1H NMR (300 MHz,
CDCl3) d 3.97 (s, 3H, o-OCH3), 6.98 (d, 1H, J = 8.1 Hz, ArHc), 7.04
(t, 1H, J = 8.1 Hz, ArHa), 7.24 (t, 1H, J = 8.1 Hz, ArHb), 7.49 (t, 1H,
J = 8.1 Hz, ArHe), 7.63 (d, 1H, J = 7.5 Hz, ArHf), 7.78 (d, 1H,
2.3.17. 1-(3-Chlorobenzoyl)-3-(3-bromophenyl)thiourea (17)
Yield 89%; white solid; mp 165–166 °C; FT-IR (KBr, cmꢀ1) 3250,
3161, 1679, 1578, 1530, 1419, 1322, 1252, 1158, 1078, 800, 712,
684, 600, 478, 448; 1H NMR (500 MHz, DMSO-d6) d 7.36 (t, 1H,
J = 8.2 Hz, ArHb), 7.45 (d, 1H, J = 7.5 Hz, ArHc), 7.54 (t, 1H,
J = 8.1 Hz, ArHe), 7.61 (d, 1H, J = 8.1 Hz, ArHa), 7.70 (d, 1H,
0
0
0
J = 7.5 Hz, ArHf), 7.91 (d, 1H, J = 7.5ArHd), 8.01 (s, 1H, ArHc ), 8.05
J = 7.5 Hz, ArHd), 7.94 (s, 1H, ArHd ), 8.77 (d, 1H, J = 8.1 Hz, ArHb ),
(s, 1H, ArHd ), 11.80 (s, CONH), 12.49 (s, CSNH); 13C NMR
9.05 (s, CONH), 12.78 (s, CSNH); 13C NMR (75 MHz, CDCl3) d 56.0
0
(125 MHz, DMSO-d6) d 121.5, 123.9, 127.3, 127.9, 128.7, 129.5,
130.8, 131.0, 133.3, 133.6, 134.5, 139.9 (Ar-C), 167.2 (C@O), 179.7
(C@S); MS (EI) m/z 369.6 (M+), 138.8 (base peak); Anal. Calcd for
(o-OCH3),
110.62,
120.26,
122.89,
125.34,
126.95,
127.01,128.04,130.44, 132.59, 133.63, 135.57, 150.69 (Ar-C),
165.16 (C@O), 176.27 (C@S); Anal. Calcd for C15H13N2O2SCl; C,
56.16; H, 4.08; N, 8.73; S, 10.00 Found: C, 56.15; H, 4.00; N, 8.75;
S, 9.98.
C14H10N2OSBrCl; C, 45.49; H, 2.73; N, 7.58; S, 8.67 Found: C,
45.47; H, 2.72; N, 7.60; S, 8.65.
2.3.18. 1-(3-Chlorobenzoyl)-3-(2,4-dibromophenyl)thiourea
(18)
2.3.23. 1-(3-Chlorobenzoyl)-3-(3-methoxyphenyl)thiourea (23)
Yield 85%; white solid; mp 110–111 °C; FT-IR (KBr, cmꢀ1) 3275,
3045, 2961, 1677, 1597, 1561, 1524, 1466, 1352, 1279, 1246, 1155,
1034, 908, 827, 802, 676, 554, 450; 1H NMR (300 MHz, CDCl3) d
3.85 (s, 3H, p-OCH3), 6.85 (d, 1H, J = 8.1 Hz, ArHc), 7.22 (d, 1H,
J = 8.1 Hz, ArHa), 7.33 (t, 1H, J = 8.1 Hz, ArHb), 7.48–7.53 (m, 2H,
Yield 90%; white solid; mp 160–161 °C; FT-IR (KBr, cmꢀ1) 3250,
3040, 3020, 3013, 1682, 1568, 1527, 1466, 1323, 1252, 1160, 1075,
1046, 868, 764, 698, 590, 453; 1H NMR (500 MHz, DMSO-d6) d 7.57
(t, 1H, J = 8.2 Hz, ArHe), 7.66 (d, 1H, J = 8.1 Hz, ArHc), 7.72 (d, 1H,
J = 8.1 Hz, ArHb), 7.82 (d, 1H, J = 8.1 Hz, ArHf), 7.92 (d, 1H,
0
ArHe, c ), 7.64 (d, 1H, J = 7.8 Hz, ArHf), 7.75 (d, 1H, J = 7.8 Hz, ArHd),
J = 8.1 Hz, ArHd),7.99 (s, 1H, J = 8.1ArHb ), 8.04 (s, 1H, J = 8.1 Hz,
7.91 (s, 1H, ArHd ), 9.10 (s, CONH), 12.54 (s, CSNH); 13C NMR
0
0
ArHd ) 12.07 (s, CONH), 12.41 (s, CSNH); 13C NMR (125 MHz,
(75 MHz, CDCl3)
127.98, 129.36, 129.68, 130.52, 133.39, 133.79, 135.64, 138.53,
159.92 (Ar-C), 165.62 (C@O), 177.73 (C@S); Anal. Calcd for C15H13
d 55.47 (p-OCH3), 109.47, 112.71, 116.16,
0
DMSO-d6) d 120.5, 121.2, 128.0, 129.0, 130.7, 130.9, 131.4, 133.4,
133.6, 134.4, 135.0, 137.0 (Ar-C), 167.6 (C@O), 180.8 (C@S); Anal.
Calcd for C14H9N2OSBr2Cl; C, 37.49; H, 2.02; N, 6.25; S, 7.15 Found:
C, 37.45; H, 2.00; N, 6.27; S, 7.10.
-
N2O2SCl; C, 56.16; H, 4.08; N, 8.73; S, 10.00 Found: C, 56.14; H,
4.02; N, 8.76; S, 9.99.
2.3.19. 1-(3-Chlorobenzoyl)-3-(2-nitrophenyl)thiourea (19)
Yield 84%; white solid; mp 155–156 °C; FT-IR (KBr, cmꢀ1) 3283,
3232, 3020, 1666, 1660, 1590, 1530, 1438, 1345, 1252, 1155, 1076,
873, 766, 746, 690, 562, 483; 1H NMR (300 MHz, CDCl3) d 7.26 (d,
1H, J = 7.5 Hz, ArHc), 7.36 (t, 1H, J = 7.8 Hz, ArHa), 7.47–7.53 (m, 2H,
2.3.24. 1-(3-Chlorobenzoyl)-3-(2-trifluoromethylphenyl)
thiourea (24)
Yield 86%; white solid; mp 150–151 °C; FT-IR (KBr, cmꢀ1) 3251,
3119, 3057, 1675, 1568, 1523, 1320, 1245, 1159, 1115, 1032, 874,
760, 744, 710, 603, 548; 1H NMR (500 MHz, CDCl3) d 7.42–7.49 (m,
2H, ArH b, e), 7.60–7.65 (m, 2H, ArHa, c), 7.73 (d, 1H, J = 7.5 Hz, ArHf),
0
ArHe, b), 7.63 (d, 1H, J = 8.1 Hz, ArHb ), 7.76 (d, 1H, J = 7.8 Hz, ArHf),
0
0
7.92 (s, 1H, ArHd ), 8.43 (d, 1H, J = 7.8 Hz, ArHd) 9.18 (s, CONH),
7.77 (d, 1H, J = 7.5 Hz ArHd), 7.92 (d, 1H, J = 7.6 Hz, ArHb ), 7.95 (s,
12.62 (s, CSNH); 13C NMR (75 MHz, CDCl3) d 124.3, 125.1, 126.2,
126.8, 127.5, 128.3, 129.6, 130.8, 133.1, 133.7, 135.6, 135.8 (Ar-
C), 165.3 (C@O), 178.5 (C@S).
1H, ArHd ), 9.38 (s, CONH), 12.44 (s, CSNH); 13C NMR (125 MHz,
0
CDCl3) d 123.93, 124.60 & 124.81 (13C-19F, 1J = 26.5 Hz, CF3),
125.5, 126.30 & 126.33 (13C-19F, 2J = 3.75 Hz), 127.5, 128.0, 129.6,
130.3, 132.1, 133.0, 133.7, 135.3, 135.6 (Ar-C), 165.7 (C@S), 180.3
(C@O); Anal. Calcd For C15H10N2OSClF3; C, 50.22; H, 2.81; N,
7.81; S, 8.90 Found: C, 50.19; H, 2.79; N, 7.83; S, 8.89.
2.3.20. 1-(3-Chlorobenzoyl)-3-(4-nitrophenyl)thiourea (20)
Yield 83%; yellow solid; mp 169–170 °C; FT-IR (KBr, cmꢀ1
)
3287, 3045, 3032, 1691, 1594, 1557, 1510, 1324, 1258, 1161,
1107, 881, 853, 752, 732, 676, 608, 496, 462; 1H NMR
2.3.25. 1-(3-Chlorobenzoyl)-3-(3- trifluoromethylphenyl)
thiourea (25)
0
(300 MHz, CDCl3) d 7.53 (t, 1H, J = 7.8 Hz, ArHe ), 7.67 (d, 1H,
J = 8.1 Hz, ArHf), 7.79 (d, 1H, J = 8.1 Hz, ArHd),7.92 (s, 1H, ArHd ),
Yield 85%; white solid; mp 121–122 °C; FT-IR (KBr, cmꢀ1) 3334,
3219, 3033, 1676, 1601, 1574, 1527, 1454, 1329, 1252, 1158, 1111,
890, 799, 742, 718, 691, 608, 486; 1H NMR (300 MHz, CDCl3) d
7.49–7.57 (m, 3H, ArHe, c, b), 7.66 (d, 1H, J = 7.8 Hz, ArHf), 7.78 (d,
0
0
0
8.06 (d, 2H, J = 9.3 Hz, ArHc@c ), 8.31 (d, 2H, J = 9.3 Hz, ArHb@b ),
9.19 (s, CONH), 12.96 (s, CSNH); 13C NMR (75 MHz, CDCl3) d
123.23(2C), 124.65 (2C), 125.45, 128.04, 130.64, 132.94, 134.15,
135.77, 143.09, 145.25 (Ar-C), 165.92 (C@O), 177.89 (C@S); MS
(EI) m/z 334.7 (M+), 138.8 (base peak); Anal. Calcd for C14H10N3-
O3SCl; C, 50.08; H, 3.00; N, 12.51; S, 9.55 Found: C, 50.02; H,
3.01; N, 12.53; S, 9.54.
0
1H, J = 7.8 Hz, ArHd), 7.91 (s, 1H, ArHd ), 7.96 (d, 1H, J = 7.8 Hz,
ArHa), 8.05 (s, 1H, ArHc ) 9.14 (s, CONH), 12.64 (s, CSNH); 13C
0
NMR (75 MHz, CDCl3) d 120.93, 123.6, 125.42& 126.62 (13C-19F,
1J = 25.1 Hz), 127.30,128.01, 129.51, 130.58, 131.20, 131.63,
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

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M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
0
133.15, 133.98, 135.71, 137.99 (Ar-C), 165.81 (C@O), 178.37 (C@S);
MS (EI) m/z 357.7 (M+), 138.9 (base peak); Anal. Calcd For C15H10
(500 MHz, CDCl3) d 7.11 (m, 2H, ArHc@c ), 7.49 (t, 1H, J = 7.6 Hz,
b0,
-
ArHe), 7.62–7.66 (m, 3H, ArHb,
f), 7.75 (d, 1H, J = 8.9 Hz, ArHd),
7.88 (s, 1H, ArHd ) 9.16 (s, CONH), 12.38 (s, CSNH); 13C NMR
(125 MHz, CDCl3) d 115.8 (d, 2C, 2J = 25 Hz, 13C-19F), 125.3, 126.2
(d, 2C, 3J = 6.0 Hz, 13C-19F), 127.9, 120.4, 133.2, 133.4, 133.7,
135.6, 159.0 (d, 1J = 250 Hz, 13C-19F) (Ar-C), 165.7 (C@O), 178.6
(C@S); Anal. Calcd for C14H10N2OSClF; C, 54.46; H, 3.26; N, 9.07;
S, 10.39 Found: C, 54.45; H, 3.25; N, 9.10; S, 10.38.
0
N2OSClF3; C, 50.22; H, 2.81; N, 7.81; S, 8.90 Found: C, 50.17; H,
2.78; N, 7.81; S, 8.90.
2.3.26. 1-(3-Chlorobenzoyl)-3-(4-trifluoromethylphenyl)
thiourea (26)
Yield 88%; white solid; mp 112–113 °C; FT-IR (KBr, cmꢀ1) 3225,
3116, 3042, 1669, 1605, 1568, 1531, 1414, 1356, 1327, 1261, 1165,
1113, 1067, 876, 837, 789, 730, 700, 608, 589, 459, 422; 1H NMR
(500 MHz, CDCl3) d 7.48 (t, 1H, J = 6.2 Hz, ArH e), 7.63 (d, 1H,
2.3.31. 1-(3-Chlorobenzoyl)-3-(1-naphthyl)thiourea (31)
Yield 90%; intense violet; mp 164–165 °C; FT-IR (KBr, cmꢀ1
)
0
J = 6.2 Hz, ArHf), 7.67 (d, 2H, J = 6.9 Hz, ArHc@c ), 7.76 (d, 1H,
3231, 3165, 2919, 1670, 1568, 1509, 1328, 1246, 1173, 1156,
J = 6.5 Hz, ArHd), 7.90 (m, 3H, ArHb, b , d ), 9.24 (s, CONH), 12.71 (s,
1066, 883, 773, 746, 713, 475, 444; 1H NMR (500 MHz, CDCl3) d
0
0
CSNH); 13C NMR (125 MHz, CDCl3) d 123.7 (2C), 125.71 & 125.91
7.38 (t, 1H, J = 8.2 Hz, ArHb ), 7.47 (t, 1H, J = 8.2 Hz, ArHb), 7.53–
7.65 (m, 3H, ArHe, a, a), 7.70 (d, 1H, J = 7.5 Hz, ArHc ), 7.86 (d, 1H,
J = 8.1 Hz, ArHf), 7.92 (d, 1H, J = 7.5 Hz, ArHd), 7.98 (s, 1H, ArHd ),
0
13C-19F, 1J = 25 Hz, CF3), 125.4 (2C), 126.0& 126.09 (2C,13C-19F,
0
(
2J = 3.75), 130.0, 130.5, 133.1, 133.8, 135.6, 140.5 (Ar-C), 165.8
(C@O), 178.0 (C@S); Anal. Calcd For C15H10N2OSClF3; C, 50.22; H,
2.81; N, 7.81; S, 8.90 Found: C, 50.20; H, 2.80; N, 7.82; S, 8.90.
0
8.02 (d, 2H, J = 7.5 Hz, ArHg,), 9.5 (s, CONH), 12.66 (s, CSNH); 13C
NMR (125 MHz, CDCl3) d 121.5, 123.8, 125.2, 125.5, 126.1, 126.4,
126.9, 128.0, 128.2, 128.4, 128.6, 129.9, 130.4, 133.0, 133.7,134.1
(Ar-C), 165.0 (C@O), 180.2 (C@S); Anal. Calcd for C18H13N2OSCl;
C, 63.43; H, 3.84; N, 8.22; S, 9.41 Found: C, 63.42; H, 3.83; N,
8.23; S, 9.40.
2.3.27. 1-(3-Chlorobenzoyl)-3-(n-butylphenyl)thiourea (27)
Yield 85%;, white solid; mp 142.5–142.7 °C; FT-IR (KBr, cmꢀ1
)
3316, 3226, 3025, 2954, 2928, 2854, 1668, 1597, 1553, 1510,
1446, 1423, 1350, 1278, 1248, 1146, 1078, 832, 810, 719, 679,
526, 493; 1H NMR (500 MHz, CDCl3) d 0.94 (t, 3H, J = 7.5 Hz, -
C H2-CbH2-C H2-CdH3), 1.37 (sex, 2H, J = 6.9 Hz, -C H2-), 1.61(p,
2.3.32. 1-(3-Chlorobenzoyl)-3-(2-benzothiazol)thiourea (32)
Yield 86%; white solid; mp 160–161 °C; FT-IR (KBr, cmꢀ1) 3285,
3042, 3033, 1698, 1592, 1552, 1511, 1329, 1253, 1164, 1109, 885,
853, 752, 735, 674, 608, 492, 463; 1H NMR (300 MHz, CDCl3) d 7.52
a
c
c
2H, J = 7.6 Hz, -CbH2-), 2.62(t, 2H, J = 7.5 Hz, -C H2-), 7.22 (d, 2H,
a
0
J = 8.1 Hz, ArHc@c ), 7.45 (t, 1H, J = 8.1 Hz, ArHe), 7.59(d, 2H,
0
0
0
0
J = 8.2 Hz, ArHb@b ), 7.71–7.83 (m, 2H, ArHd, ), 7.89 (s, 1H, ArHd ),
(t, 1H, J = 7.8 Hz, ArHe ), 7.60 (d, 2H, J = 9.1 Hz, ArHa@a ), 7.66 (d, 1H,
f
9.28 (s, CONH), 12.42 (s, CSNH); 13C NMR (125 MHz, CDCl3) d
13.8, 22.23, 33.33, 35.11(n-butyl), 123.7(2C), 125.4, 127.9, 128.72
(2C), 130.3, 133.4, 133.5, 134.9, 135.3, 141.8(Ar-C), 165.7(C-7),
177.9 (C-8); MS (EI) m/z (M+); Anal. Calcd for C18H19ClN2OS; C,
62.33, H 5.52, N 8.08, S 9.24 Found: C 62.28, H 5.51, N 8.01, S 9.22.
J = 8.1 Hz, ArHf), 7.75 (d, 1H, J = 8.1 Hz, ArHd), 7.95 (s, 1H, ArHd ),
0
0
8.31 (d, 2H, J = 9.3 Hz, ArHb@b ), 9.18 (s, CONH), 12.94 (s, CSNH);
13C NMR (75 MHz, CDCl3) d 123.1, 124.65, 125.4, 128.0, 130.6,
132.9, 134.1, 135.7, 136.6, 143.0, 145.2, 148.9, 165.8 (C@O),
174.5 (CNNS), 177.9 (C@S); MS (EI) m/z 334.7 (M+), 138.8 (base
peak); Anal. Calcd for C14H10N3O3SCl; C, 50.08; H, 3.00; N, 12.51;
S, 9.55 Found: C, 50.02; H, 3.01; N, 12.53; S, 9.54.
2.3.28. 1-(3-Chlorobenzoyl)-3-(2, 6-diethylphenyl)thiourea (28)
Yield 89%; white solid; mp 156–157 °C; FT-IR (KBr, cmꢀ1) 3192,
3158, 3066, 2968, 2934, 2873, 1672, 1530, 1509, 1420, 1337,
12471202, 1159, 1111, 1065, 875, 812, 714, 617, 478, 444; 1H
NMR (500 MHz, CDCl3) d 1.26 (t, 6H, J = 4.5 Hz, 2 [o-CH2–CH3]),
2.67 (q, 4H, J = 4.5 Hz, 2 [o-CH2–CH3]), 7.21 (d, 2H, J = 7.6 Hz, ArHb-
2.3.33. 1-(3-Chlorobenzoyl-3-(N-methylphenyl)thiourea (33)
Yield 88%; white solid; mp 130–131 °C; FT-IR (KBr, cmꢀ1) 3240,
3200, 3027, 2933, 1697, 1570, 1515, 1437, 1387, 1277, 1248, 1177,
1118, 1059, 916, 763, 735, 638, 550, 465; 1H NMR (300 MHz,
CDCl3) d 3.78 (s, 3H, N-CH3), 7.26–8.40 (m, 9H, Ar-H), 8.5 (s, CONH);
13C NMR (75 MHz, CDCl3) d 45.7 (N-CH3), 125.4, 125.5, 127.8 (2C),
127.9 (2C),128.0 (2C), 129.5 (2C), 130.0, 132.7, 133.4, 134.5, 134.9
(Ar-C), 161.59 (C@O), 175.76 (C@S); Anal. Calcd for C15H13N2OSCl;
C, 59.11; H, 4.30; N, 9.19; S, 10.52 Found: C, 59.11; H, 4.29; N, 9.22;
S, 10.52.
0
@b ), 7.33 (t, 1H, J = 7.5 Hz, ArHa), 7.50 (t, 1H, J = 7.5 Hz, ArHe), 7.64
(d, 1H, J = 8.2 Hz, ArHf), 7.80 (d, 1H, J = 8.2 Hz, ArHd), 7.95 (s, 1H,
ArHd ), 9.28 (s, CONH), 11.80 (s, CSNH); 13C NMR (125 MHz, CDCl3)
0
d 20.5 (2C,-CH3), 24.7 (2C, –CH2–), 125.4, 126.5 (2C), 128.0, 128.8,
130.4 (2C), 133.3, 133.7, 134.0, 135.5, 140.9 (Ar-C), 165.7 (C@O),
180.18 (C@S); MS (EI) m/z 346.1 (M+), 139.0 (base peak); Anal.
Calcd for C18H19N2OSCl; C, 62.33; H, 5.52; N, 8.08; S, 9.24 Found:
C, 62.31; H, 5.51; N, 8.11; S, 9.23.
2.3.34. 1-(3-Chlorobenzoyl)-3-benzylthiourea (34)
Yield 88%; white solid; mp 113–114 °C; FT-IR (KBr, cmꢀ1) 3310,
3236, 3209, 3020, 2960, 1674, 1641, 1541, 1322, 1252, 1170, 1070,
804, 748, 697, 617, 478; 1H NMR (300 MHz, CDCl3) d 4.93 (d, 2H,
J = 5.4 Hz, –CH2–), 7.34–7.85 (m, 9H, ArH), 9.12 (s, CONH), 10.94
(s, CSNH); 13C NMR (75 MHz, CDCl3) d 49.89 (–CH2–), 125.33,
127.94 (2C), 128.94 (2C), 129.96, 130.44, 131.63, 133.62, 135.53,
135.99, 137.81 (Ar-C), 165.54 (C@O), 179.76 (C@S); MS (EI) m/z
269.9 (M+ = M-Cl), 104.9 (base peak); Anal. Calcd for C15H13N2-
OSCl; C, 59.11; H, 4.30; N, 9.19; S, 10.52 Found: C, 59.12; H, 4.28;
N, 9.20; S, 10.50.
2.3.29. 1-(3-Chlorobenzoyl)-3-(3-fluorophenyl)thiourea (29)
Yield 89%; white solid; mp 141–142 °C; FT-IR (KBr, cmꢀ1) 3298,
3228, 3061, 1668, 1541, 1528, 1470, 1246, 1163, 1051, 897, 801,
746, 683, 609, 538, 455; 1H NMR (500 MHz, CDCl3) d 7.0 (d, 1H,
0
J = 7.5 Hz, ArHc), 7.36–7.39 (m, 2H, ArHa, c ), 7.48 (t, 1H, J = 8.1 Hz,
ArHe), 7.62 (d, 1H, J = 7.5 Hz, ArHf), 7.71–7.75 (m, 2H, ArHd, b),
7.89 (s, 1H, ArHd ) 9.14 (s, CONH), 12.58 (s, CSNH); 13C NMR
0
(125 MHz, CDCl3) d 111.3 (d, 13C-19F, 2J = 25 Hz), 113.9 (d, 13C-19F,
2J = 25 Hz), 119.5, 125.5, 128.0, 130.1, 130.3, 130.6, 133.3, 133.5,
135.7, 163.0 (d, 13C-19F, 1J = 250 Hz) (Ar-C), 165.9 (C = O), 177.9
(C@S); Anal. Calcd for C14H10N2OSClF; C, 54.46; H, 3.26; N, 9.07;
S, 10.39 Found: C, 54.47; H, 3.25; N, 9.11; S, 10.39.
2.3.35. 1-(3-Chlorobenzoyl)-3-(2-chlorobenzyl)thiourea (35)
Yield 87%; white solid; mp 124–125 °C; FT-IR (KBr, cmꢀ1
)
3294, 3040, 2981, 1680, 1605, 1561, 1522, 1346, 1244, 1149,
1110, 914, 829, 735, 673, 604, 453; 1H NMR (500 MHz, CDCl3)
d 4.98 (d, 2H, J = 5.4 Hz, –CH2–), 7.23–7.56 (m, 6H, ArH), 7.67
2.3.30. 1-(3-Chlorobenzoyl)-3-(4-fluorophenyl)thiourea (30)
Yield 90%; white solid; mp 151–152 °C; FT-IR (KBr, cmꢀ1) 3216,
3044, 1667, 1615, 1569, 1537, 1504, 1360, 1261, 1227, 1159, 1115,
1082, 878, 820, 730, 697, 606, 539, 511, 475, 434; 1H NMR
0
(d, 1H, J = 7.5 Hz, ArHd), 7.82 (s, 1H, ArHd ), 9.21 (s, CONH),
10.06 (s, CSNH); 13C NMR (125 MHz, CDCl3) d 47.3 (–CH2–),
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
7
125.4, 126.2, 126.9, 127.9, 128.4, 129.4,129.6, 130.3, 130.5, 133.0,
133.4, 135.3 (Ar-C), 165.6 (C@O), 179.8 (C@S); Anal. Calcd for
15H12N2OSCl2; C, 53.11; H, 3.57; N, 8.26; S, 9.45 Found: C,
2.5. Biological assays
C
2.5.1. Measurement of urease activity
53.12; H, 3.57; N, 8.28; S, 9.45.
Newly synthesized series of N,N0-disubstituted thioureas (1–38)
were screened for inhibitory activity against urease. The activity
was measured by determining the amount of ammonia liberated
during the indophenol by using standard protocol.39,43 The com-
pounds were initially screened at 1 mM concentration. Thioureas
exhibiting P50% inhibition were selected for further determina-
tion of dose–response curves against all the compounds. For this
purpose 7–9 serial dilutions of each compound were prepared in
assay buffer and their dose response curves were obtained by
adopting the same methods used for initial screening. All experi-
ments were performed in triplicate. Results reported are mean of
three independent experiments ( SEM) and expressed as percent
inhibitions calculated by the formula.
2.3.36. 1-(3-Chlorobenzoyl)-3-cyclohexylthiourea (36)
Yield 89%; white solid; mp 90–91 °C; FT-IR (KBr, cmꢀ1) 3323,
3227, 3170, 3048, 2928, 2854, 1672, 1627, 1542, 1341, 1262,
1169, 1022, 778, 707, 664, 623; 1H NMR (300 MHz, CDCl3) d
1.23–2.17 (m, 10H, Cyclohexane –CH2–), 4.26–4.35 (m, 1H, Cyclo-
0
hexane –CH–), 7.51 (t, 2H, J = 7.6 Hz, ArHe@e ), 7.63 (t, 1H,
0
J = 7.6 Hz, ArHf), 7.84 (d, 2H, J = 7.6 Hz, ArHd@d ), 8.94 (s, CONH),
10.72 (s, CSNH); 13C NMR (75 MHz, CDCl3) d 24.3 (2C), 25.4,
33.26 (2C), 57.62 (Cyclohexane-C), 127.3 (2C), 129.1 (2C), 131.8,
133.5 (Ar-C), 166.8 (C@O), 178.22 (C@S); Anal. Calcd for C14H17N2-
OSCl; C, 56.65; H, 5.77; N, 9.44; S, 10.80 Found: C, 56.64; H, 5.77; N,
9.46; S, 10.79.
Inhibitionð%Þ ¼ ½100 ꢀ ðAbs of test compound=Abs of controlÞ
ꢁ 100ꢂ
2.3.37. 1-(3-Chlorobenzoyl)-3-dicyclohexylthiourea (37)
Yield 80%; white solid; mp 120–121 °C; FT-IR (KBr, cmꢀ1) 3328,
3177, 3055, 2946, 2864, 1662, 1617, 1526, 1335, 1264, 1168, 1020,
774, 709, 668, 626; 1H NMR (300 MHz, CDCl3) d 1.24–2.20 (m, 20H,
Cyclohexane –CH2-), 4.22–4.34 (m, 2H, Cyclohexane –CH-), 7.53 (t,
IC50 values of potential inhibitors (P50%) were obtained with
the help of non-linear regression analysis program of Graph Pad
prism 5.0 Software Inc., San Diego, California, USA.
0
2H, J = 7.6 Hz, ArHe@e ), 7.64 (t, 1H, J = 7.6 Hz, ArHf), 7.82 (d, 2H,
2.5.2. Cytotoxicity assay by sulforhodamine B (SRB)
J = 7.6 Hz, ArHd@d ), 8.98 (s, CONH); 13C NMR (75 MHz, CDCl3) d
0
Lung carcinoma (H-157), (ATCC CRL-5802) cell lines were kept
in RPMI-1640 having heat-inactivated fetal bovine serum (10%)
glutamine (2 mM), pyruvate (1 mM), 100 U/mL penicillin and
24.3 (4C), 25.4, 33.26 (4C), 57.62 (Cyclohexane-C), 127.3 (4C),
129.1 (4C), 131.8, 133.5 (Ar-C), 167.0 (C@O), 178.2 (C@S); Anal.
Calcd for C20H27N2OSCl; C, 63.39; H, 7.18; N, 7.39; S, 8.46 Found:
C, 63.37; H, 7.17; N, 7.40; S, 8.78.
100
in a 5% CO2 incubator at 37 °C.44 96 well plates were used for grow-
ing H-157 cells by inoculating 104 cells per 100
L per well and
l
g/mL streptomycin in T-75 cm2 sterile tissue culture flasks
l
2.3.38. 1-(3-Chlorobenzoyl)-3-(pyrimidin-2-yl)thiourea (38)
plates were incubated in CO2 incubator at 37 °C. Within 24 h, a uni-
form monolayer was formed which was used for experiments. The
Yield 88%; yellow solid; mp 180–181 °C; FT-IR (KBr, cmꢀ1
)
3158, 3048, 2991, 2939, 1722, 1656, 1587, 1561, 1515, 1439,
1412, 1331, 1244, 1206, 1175, 1126, 916, 808, 729, 677, 567,
503, 456; 1H NMR (500 MHz, DMSO-d6) d 7.30 (t, 1H, J = 6.3 Hz,
pyrimidin–Ha), 7.61 (t, 1H, J = 6.3 Hz, ArHe), 7.73 (d, 1H,
compounds (100 lM) were inoculated in test wells while control
and blank wells were also prepared containing standard drug (vin-
cristine) and culture media with cells, respectively.44,45 The plates
were then incubated for 48 h. After that cells were fixed with 50 lL
0
J = 6.8 Hz, ArHf), 7.91 (d, 1H, J = 6.8 Hz, ArHd), 7.97 (s, 1H, ArHd ),
of 50% ice cold trichloroacetic acid solution (TCA) at 4 °C for 1 h.
The plates were washed 5 times with phosphate-buffered saline
(PBS) and air dried. Fixed cells were further treated with 0.4% w/
v sulforhodamine B dye prepared in 1% acetic acid solution and left
at room temperature for 30 min. After that the plates were rinsed
with 1% acetic acid solution and allowed to dry. In order to solubi-
lize the dye, the dried plates were treated with 10 mM Tris base
solution for 10 min at room temperature. Absorbance was mea-
sured at 490 nm subtracting the background measurement at
630 nm. All experiments were performed in triplicate. Results
reported are mean of three independent experiments ( SEM) and
expressed as percent inhibitions calculated by the formula.
0
8.76 (d, 2H, J = 4.2 Hz, pyrimidin-Hb@b ), 12.2 (s, 1H, CONH),
13.5 (s(broad)
, d
1H, CSNH); 13C NMR (125 MHz, DMSO-d6)
117.6,157.5,158.9 (2C) (pyrimidin), 127.4, 128.5, 131.4, 133.2,
134.0, 135.7 (Ar-C), 166.3 (C@O), 178.4 (C@S); MS (EI) m/z
291.7 (M+), 138.8 (base peak); Anal. Calcd for C12H9N4OSCl; C,
49.23; H, 3.10; N, 19.14; S, 10.95 Found: C, 49.23; H, 3.08; N,
19.19; S, 10.94.
2.4. Data collection and structural refinement of 13, 16, 24, 28
and 30
Crystals of the compounds 13, 16, 24, 28 and 30 were grown by
the slow evaporation in ethanol/dichloromethane (1:1). The color-
less crystals were mounted in a random orientation on a glass fiber
and the diffraction data collections were performed with a Stoe
IPDS-II two circle (13) and Rigaku AFC7R Mercury CCD diffrac-
Inhibition ð%Þ ¼ ½100 ꢀ ðAbs of test compound=Abs of controlÞ
ꢁ 100ꢂ
tometer (16, 24, 28 and 30) with graphite monochromatic MoK
a
2.5.3. Docking protocols
radiation (k = 0.71073 Å).40 The structures were solved by direct
methods using SHELXS-97 and refined with full-matrix least-
squares on F2 with SHELXL-97 and absorption correction with
MULABS option in PLATON and REQABS.41 Structures were solved
by direct and Fourier methods and refined by full-matrix least-
squares based on F2.42 All non-hydrogen atoms were refined
anisotropically, hydrogen atoms were determined from difference
Fourier maps and refined at idealized positions riding on their
attached C or N atoms. For 16 there are three crystallographically
independent but chemically equal molecules along with a solvate
per asymmetric unit.
2.5.3.1. Structure selection and preparation.
Molecular
docking studies were conducted to investigate putative interac-
tions of the compounds in complex with the urease enzyme. In
order to perform efficient docking studies, the crystallographic
structure of Jack bean urease (PDB ID: 3LA4) was obtained from
the RCSB PDB database46 and prepared. Prior to docking experi-
ments, the structures of the enzyme and the compounds were pre-
pared as follows. The enzyme structure was protonated with the
Protonate3D47 algorithm implemented in the molecular modeling
tool MOE.48 The structure was energy minimized using Amber99
force field including all crystallographic solvent molecules. The
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

8
M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
Figure 1. ORTEP structures of 13, 16, 24, 28 and 30 showing anisotropic displacement ellipsoids drawn at the 35% probability level. Structure of 16 consists of three
independent conformers with chloroform as a solvate.
backbone atoms were restrained with a small force in order to
avoid collapse of the binding pockets during energy minimization
calculations. After minimization, the water molecules and co-crys-
tallized bound compounds were removed.
lowest DG values were considered as the most stable pose with
highest affinity for interaction with the receptor.
2.6. Supplementary data
2.5.3.2. Compounds preparation.
The 3D structural coordi-
Crystallographic data (excluding structure factors) for the struc-
tures reported in this paper have been deposited to the Cambridge
Crystallographic Data Centre as supplementary publication CCDC
no. 605433, 1453877–1453880 for 13, 16, 24, 28 and 30, respec-
tively. Copies of available materials can be obtained, free of charge,
on application to the director, CCDC, 12 Union Road, Cambridge
CB21EZ, UK, (Fax:+44-(0)-1223-336033 or e-mail: deposit@
ccdc.cam.ac.uk).
nates of the compounds were generated for all the compounds
using MOE followed by assignment of protonation and ionization
states in physiological pH range by using the ‘wash’ module. After-
wards, the compounds’ structures were energy minimized with the
MMFF94x force field for docking studies.48
2.5.3.3. Docking studies.
Docking studies of all the com-
pounds were performed using LeadIT from BioSolveIT, GmbH Ger-
many.49 Receptor was loaded by Load or Prepare Receptor utility of
the LeadIT software, while selecting the metal ions as part of the
receptor. The binding site for Jack bean urease was defined in
7.5 Å spacing of the amino acid residues surrounding the bound
phosphate. By FlexX utility of LeadIT, docking of compounds was
performed. Default docking parameters were not changed and
top 30 highest scoring docked positions were kept for further anal-
ysis. The Discovery studio visualizer v4, was used for visualizing
the results.50 Using HYDE visual affinity51 program of LeadIT, the
binding mode analysis of the docked poses were evaluated. Bind-
3. Results and discussion
3.1. Chemistry
Solution-phase microwave assisted parallel synthesis of N,N0-
disubstituted thioureas was performed using CEM MARS 6 micro-
wave reactor, specifically for organic synthesis. A small library of
thirty eight members (1–38) was successfully developed and fully
characterized by modern analytical techniques. The purity level of
the compounds was established by TLC with Merck Kieselgel GF
254 plates and elemental analysis. Their structures were
ing free energy, i.e.,
DG was determined for each pose. Poses with
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
9
Table 1
Crystallographic data and structure refinement for compounds 13, 16, 24, 28 and 30
Compound
13
16
24
28
30
Formula
FW
C
14H7Cl5N2OS
C17H17ClN2OS, C0.33 H0.33 Cl
372.62
C15H10ClF3N2OS
358.76
C18H19ClN2OS
346.86
C14H10ClFN2OS
308.75
428.53
Crystal system
Space group
a (Å)
b (Å)
c (Å)
Triclinic
P-1
Triclinic
P-1
Triclinic
P-1
7.636(3)
8.275(3)
12.148(4)
88.564(11),
79.230(10),
83.530(9)
749.3(5)
Triclinic
P-1
8.1763(17)
9.071(2)
12.155(3)
85.069(7),
77.163(7),
78.649(7)
861.0(3)
Triclinic
P-1
7.8062(11)
8.4597(12)
12.7760(19)
93.374(11),
99.840(11),
97.379(11)
821.583
10.860(2)
15.783(4)
16.253(4)
101.677(3),
101.111(3),
90.700(3)
2673.22
8.6742(15)
8.7810(19)
9.1296(18)
86.151(6),
73.428(7),
79.084(6)
654.4(2)
a, b, c (°)
V (Å3)
Z
2
6
2
2
2
T (K)
173(2)
1.732
1.012
123(2)
1.389
0.487
123(2)
1.590
0.431
123(2)
1.338
0.349
123(2)
1.567
0.459
DC Mg/m3
l
(mmꢀ1
)
F(000)
428
1160
364
364
316
Crystal size (mm3)
h range (°)
0.37 ꢁ 0.32 ꢁ 0.22
3.8 to 25.7
ꢀ9<=h 6 9,
ꢀ10 6 k 6 9,
ꢀ15 6 l 6 15
5914
0.40 ꢁ 0.32 ꢁ 0.25
3.01 to 27.48°
ꢀ14 6 h 6 12,
ꢀ15 6 k 6 20,
ꢀ21 6 l 6 13
21729
0.30 ꢁ 0.24 ꢁ 0.15
3.00 to 27.47°
ꢀ9 6 h 6 9,
ꢀ7 6 k 6 10,
ꢀ15 6 l 6 15
5870
0.38 ꢁ 0.28 ꢁ 0.12
3.13 to 27.46°
ꢀ8 6 h 6 10,
ꢀ11 6 k 6 10,
ꢀ15 6 l 6 14
6831
0.50 ꢁ 0.45 ꢁ 0.35
3.11 to 27.46°
ꢀ8 6 h 6 10,
ꢀ11 6 k 6 10,
ꢀ15 6 l 6 14
5034
Index ranges
Reflections collected
Indept. Reflect. [R(int)]
3025
12097
3355
3865
2897
[R(int) = 0.0397]
98.5%
0.7057, 0.8080
3025/0/217
1.021
[R(int) = 0.0249]
98.8%
0.8879, 0.8290
12097/0/664
1.088
[R(int) = 0.0216]
97.5%
0.8816, 0.9382
3355/0/208
1.038
[R(int) = 0.0171]
97.9%
0.9594, 0.8789
3865/0/210
1.088
[R(int) = 0.0196]
% 96.5
0.8560, 0.8031
2897/0/189
1.051
Completeness to h = 28°
Max. & min. transmission
Data/restraints/parameters
Goodness-of-fit on F2
R1 [I > 2
r
(I)]
0.0336
0.0430
0.0373
0.0350
0.0360
wR2 [all data]
0.0908
0.294 and ꢀ0.434
0.1059
0.423 and ꢀ0.515
0.0960
0.368 and ꢀ0.319
0.0831
0.319 and ꢀ0.231
0.0981
0.963 and ꢀ0.271
Largest peak and hole (e Åꢀ3
)
investigated by FT-IR, 1H- and 13C-NMR (Bruker ARX, 300 MHz
spectrometer) and single crystal XRD. The reaction intermediate,
3-cholorobenzoyl isothiocyanate was synthesized by conventional
method and purified by steam distillation technique. Stoichiomet-
ric quantities of 3-cholorobenzoyl and respective primary/sec-
ondary amines were subjected to cylindrical glass vials with THF
as solvent. Irradiated the glass vials under identical set of condi-
tions of power and exposure time. The nature of substrates pri-
around 1710 cmꢀ1. The band around 22530–2590 cmꢀ1 associated
to SH was not appeared which indicated the absence of the N@CSH
tautomerism in all the compounds (1–38).52–54
The 1H NMR data for the compounds (1–38) showed that the
NH proton resonates considerably downfield and chemical shifts
were found around 9.00–9.55 and 10.80–12.78 ppm for free and
hydrogen bonded NH, respectively, and aromatic protons appeared
downfield 7.25 to 8.55 ppm. The coordinating or highly polar sol-
vents like DMSO-d6 had prominent effect on the free NH protons
chemical shift and appeared more downfield as compared with
the solvents like C6D6, CDCl3 and CD2Cl2. Such shifts could be
ascribed to the probable hydrogen bonding between the NH and
sulfoxide (S@O) moiety.52
´
mary/secondary amines (H2NR/ HNRR), strongly influence the
yield of the final products (Scheme 1). The substituents like F, Cl,
Br, CF3, OR, R and –NO3 attached at the ortho and para positions
on the aromatic ring in H2NR/HNRR0 moiety show strong meso-
meric effect (+R). This has been reflected in the increase of product
yield in the following order; Br/Cl > R > OR > F > CF3 > cyclo-
hexyl > –NO3. The cyclohexyl and –NO3 containing substrate has
shown lowest yield due to conformational isomerism and high
electron with drawing effect, respectively.39 The complete descrip-
tion of the substituents for derivatives (1–38) is illustrated in
Scheme 1.
The 13C NMR data depicted all the signals due to magnetically
distinctive carbons present in the compounds (1–38). The carbon
resonances associated with the aromatic moieties of thiourea
ligands were consigned on the basis of signal intensities and were
compared with the literature values.55 The chemical shifts related
to the carbons in CONH and CSNH groups of the thioureas resonate
around 165–170 and 175–185 ppm, respectively.
3.2. Spectroscopic studies
3.3. Structural studies
The major absorption bands recorded in the FT-IR spectra of the
synthesized N,N0-disubstituted thioureas (1–38) are delineated in
spectroscopic data separately. The assignments are in good agree-
The five molecular structures 13, 16, 24, 28 and 30 (Fig. 1) con-
sist of similar thiourea cores (–CONHCSNH–) with different substi-
tution patterns. The cores are mostly planar with O–C–N–C and C–
N–C–S torsion angles of ꢀ4.5(4)° and 174.7(2)° for 13, ꢀ6.0(3)°
(molecule I), 3.1(3)° (molecule II), 7.5(3)° (molecule III) and
175.87(16)° (I), ꢀ178.19(15)° (II), 178.26(14)° (III) for 16, 12.2(2)°
and 172.65(12)° for 24, ꢀ9.4(2)° and ꢀ179.38(11)° for 28, 2.5(2)°
and 175.07(12)° for 30 as well. The aromatic ring planes make
dihedral angles of 59.74(1)°, 54.82(5)°, 88.27(3)°, 71.32(1)° and
21.31(5)° for (13), (16I), (24), (28) and (30), respectively. In general,
there are no unexpected variations in geometric parameters. All
ment with the literature reported values.52 The absence of
m (S-H)
absorption band at 2530–2590 cmꢀ1 confirmed the change of
isothiocyanate moiety into thiourea (–NHCSNH–) group. The char-
acteristic IR bands of N,N0-disubstituted thioureas were between/
near to; 3120–3382 (NH), 2950–3100 Ph(CH), 1650–1730 (C@O),
1550–1630 (CN), 1250–1275 (C@S) and 1150–1190 (CAS). A med-
ium strong band at 1670–1680 cmꢀ1 suggested a possible hydro-
gen bond interactions between the NH-group and the carbonyl
O-atom, instead of the expected normal carbonyl (C@O) absorption
Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042

10
M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
Table 2
Selected bond lengths (Å) and angles (°) for compounds 13, 16, 24, 28 and 30
13
C(2)–S(1)
C(1)–O(1)
1.650(2)
1.229(3)
O(1)–C(1)–N(1)
C(1)–N(1)–C(2)
121.9(2)
128.7(2)
C(1)–N(1)
C(2)–N(1)
1.386(3)
1.402(3)
N(2)–C(2)–N(1)
N(2)–C(2)–S(1)
114.9(2)
125.73(17)
16
28
C(1)–S(1)
C(2)–O(1)
C(1)–N(1)
C(1)–N(2)
C(2)–N(2)
1.6690(16)
1.222(2)
1.325(2)
1.399(2)
1.385(2)
C(1)–N(1)–C(3)
N(1)–C(1)–N(2)
N(1)–C(1)–S(1)
O(1)–C(2)–N(2)
N(2)–C(1)–S(1)
124.61(14)
116.56(14)
124.15(13)
122.43(17)
119.29(13)
C(1)–S(1)
C(2)–O(1)
C(1)–N(1)
C(1)–N(2)
C(2)–N(2)
1.6734(14)
1.2239(17)
1.3286(17)
1.3929(16)
1.3799(17)
C(1)–N(1)–C(3)
N(1)–C(1)–N(2)
N(1)–C(1)–S(1)
O(1)–C(2)–N(2)
N(2)–C(1)–S(1)
123.99(11)
116.85(11)
124.14(10)
122.78(12)
119.01(10)
24
30
C(1)–S(1)
C(2)–O(1)
C(1)–N(1)
C(1)–N(2)
C(2)–N(2)
1.6669(17)
1.225(2)
1.337(2)
1.3920(19)
1.383(2)
C(1)–N(1)–C(3)
N(1)–C(1)–N(2)
N(1)–C(1)–S(1)
O(1)–C(2)–N(2)
N(2)–C(1)–S(1)
123.41(13)
116.13(14)
124.24(12)
122.39(14)
119.59(12)
C(1)–S(1)
C(2)–O(1)
C(1)–N(1)
C(1)–N(2)
C(2)–N(2)
1.6713(15)
1.2244(19)
1.333(2)
1.4032(19)
1.3780(19)
C(1)–N(1)–C(3)
N(1)–C(1)–N(2)
N(1)–C(1)–S(1)
O(1)–C(2)–N(2)
N(2)–C(1)–S(1)
131.83(13)
114.66(13)
128.38(12)
122.65(14)
116.95(11)
Table 3
The intermolecular and intramolecular hydrogen bonds for compounds 13, 16, 24, 28 and 30. Hydrogen-bond geometry: Distance, Å; angle,°
Compound
D–Hꢃ ꢃ ꢃA
D–H(Å)
Hꢃ ꢃ ꢃA(Å)
Dꢃ ꢃ ꢃA(Å)
D–Hꢃ ꢃ ꢃA(°)
13
16
N2–H2ꢃ ꢃ ꢃO1
N1–H1ꢃ ꢃ ꢃO1
N2–H2ꢃ ꢃ ꢃS2
N3–H3ꢃ ꢃ ꢃO2
N3–H3ꢃ ꢃ ꢃO3i
N4–H4ꢃ ꢃ ꢃS1
N5–H5ꢃ ꢃ ꢃO3
N5–H5ꢃ ꢃ ꢃO2ii
N1–H1ꢃ ꢃ ꢃO1
N2–H2ꢃ ꢃ ꢃS1i
N1–H1ꢃ ꢃ ꢃO1
N2–H2ꢃ ꢃ ꢃS1i
N1–H1ꢃ ꢃ ꢃO1
N2–H2ꢃ ꢃ ꢃS1i
0.85(3)
0.83(2)
0.85(2)
0.818(18)
0.818(18)
0.85(2)
0.84(2)
0.84(2)
0.88
0.88
0.88
0.88
0.86(2)
0.79(2)
1.96(3)
1.97(2)
2.53(2)
2.015(19)
2.39(2)
2.55(2)
2.00(2)
2.58(2)
1.92
2.57
1.95
2.56
1.88(2)
2.77(2)
2.644(2)
136(2)
139(2)
167.9(18)
135.4(18)
135.7(17)
170.9(18)
136(2)
142(2)
135.4
163.5
134.9
159.0
144(2)
151(2)
2.6509(19)
3.3730(18)
2.6598(19)
3.0280(19)
3.3878(18)
2.674(2)
3.285(2)
24
28
30
2.6227(18)
3.4251(17)
2.6425(15)
3.3951(13)
2.6227(18)
3.4787(15)
Symmetry codes: (i) x, y, z ꢀ 1; (ii) x, y, z + 1(16); (i) ꢀx + 1, ꢀy + 1, ꢀz + 1(24); (i) ꢀx + 1, ꢀy + 1, ꢀz + 1(28); (i) ꢀx + 1, ꢀy + 1, ꢀz + 1(30).
the molecular structures show an intramolecular NAHꢃ ꢃ ꢃO hydro-
gen bond stabilizing the (–CONHCSNH–) geometry, forming a
pseudo six membered ring (Table 3). Crystallographic and refine-
ment details are shown in Table 1. Selected bond lengths and
angles are given in Table 2.
chlorophenyl derivatives. Similarly, all the trifluoromethylphenyl
derivatives were potent inhibitors of urease in the series.
3.4.2. Anticancer activity of N,N0-disubstituted thioureas
The anticancer drugs used in chemotherapy are systemic
antiproliferative agents, which preferentially kill those cells that
are dividing. In the present study, antiproliferative activity of the
newly synthesized thiourea derivatives was measured by the cell
growth inhibition against lung carcinoma (H157) through SRB
assay and the % inhibition was presented in Table 5. Vincristine,
a standard anticancer drug, was used as reference for test com-
pounds. The results showed that all the compounds exhibit mild
3.4. Pharmacology
3.4.1. Urease inhibition assay
The synthesized N,N0-disubstituted thioureas (1–38) were
screened for inhibitory activity against Jack bean urease and the
result are given in Table 4. The results revealed that most of the
compounds in the series possess lower IC50 values than the stan-
to moderate anticancer activity against H157 cell lines at 100 lM.
dard thiourea (IC50 = 22.3 1.2
methylphenyl substitution was found to be most potent
compound among the series having an IC50 value of
l
M). Compound 16 with 2,4,6-tri-
Compound 12 having 2,4,6-trichlorophenyl substitution was
the potent compound against H157 with 60.9% inhibition. The nat-
ure and type of the side groups present in compounds 1–38 is one
of the most important factors in the appearance of anticancer
effect. The substitution at phenyl groups effects the inhibitory
potential of compounds. Halogenated substitution at phenyl ring
has greater influence on anticancer activity.
1.23 0.1
active than the standard inhibitor, thiourea having IC50 value of
22.3 1.2 M. Whereas, the compound 23 having 2,4-dinitro-
lM. The compound was found to be ꢄ18-fold more
l
phenyl substitution was the least potent with 21.7% inhibition.
Compound 5 having 2,3-dichlorophenyl and compound 38 with
2-pyrimidinyl substitution, presented IC50 value of 1.92 0.1 and
3.4.3. Molecular docking
2.11 0.1
lM, respectively. The substitution of different groups
3.4.3.1. Molecular docking studies.
The structures of di-sub-
including electron-donating and electron-withdrawing groups at
the phenyl ring resulted in varying inhibition patterns depending
upon type and position of substituent. Some of the substituents
on phenyl ring, improved inhibitory activities of compounds and
some resulted in reduction in inhibition capacity.
stituted thioureas were docked to the crystallographic structure of
Jack bean urease PDB ID: 3LA4. Active site of the receptor contain
bi-nickel center. The binding pattern of synthesized inhibitors
were established by molecular docking studies. As evident from
in vitro results compound 16 was the most suitable for docking
studies. After reproducing the co-crystallized reference ligand into
the active site of receptor, all the derivatives were docked in active
It was noted that dichlorophenyl and trichlorophenyl thiourea
derivatives were more active against urease as compared to tetra-
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M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
11
Table 4
Table 6
In vitro urease inhibitory activity of N,N0-disubstituted thioureas (1–38)
Docking and Hyde scores and their corresponding ranks by Hyde affinity assessment
No
IC50 SEM (
l
M)/%inhibition No IC50 SEM (
l
M)/%inhibition
Codes
FlexX score of the
top ranking pose
Poser rank
Binding free energy
G (kJ molꢀ1
D
)
1
2
3
4
5
6
7
8
3.24 0.3
25.7%
29.1%
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
—
28.1%
23.1%
1
2
3
4
5
6
7
8
ꢀ23.34
ꢀ26.51
ꢀ22.14
ꢀ22.06
ꢀ25.73
ꢀ25.10
ꢀ24.63
ꢀ21.85
ꢀ21.82
ꢀ24.47
ꢀ28.54
ꢀ19.90
ꢀ18.17
ꢀ36.54
ꢀ20.16
ꢀ20.98
ꢀ21.65
ꢀ18.79
ꢀ22.08
ꢀ24.14
ꢀ18.64
ꢀ20.58
ꢀ19.75
ꢀ24.84
ꢀ26.04
ꢀ16.69
ꢀ28.48
ꢀ20.40
ꢀ29.08
ꢀ28.82
ꢀ20.66
ꢀ16.58
ꢀ29.34
ꢀ22.33
ꢀ25.06
ꢀ26.38
ꢀ26.22
ꢀ22.13
2
5
1
2
1
5
2
6
3
1
3
1
3
3
17
18
4
ꢀ17
ꢀ14
ꢀ18
ꢀ17
ꢀ13
ꢀ7
3.79 0.8
21.7%
5.58 0.2
2.86 0.1
7.04 0.3
36.9%
12.6 0.5
2.91 0.4
29.9%
11.7 0.2
1.92 0.1
1.34 0.1
9.72 0.7
5.38 0.3
36.7%
6.53 0.4
7.41 0.5
3.64 0.2
32.7%
12.4 0.4
4.45 0.3
1.23 0.1
20.1 0.9
6.32 0.3
8.59 0.7
ꢀ13
ꢀ16
ꢀ11
ꢀ16
ꢀ11
ꢀ16
ꢀ10
ꢀ12
ꢀ10
ꢀ15
ꢀ18
ꢀ10
ꢀ14
ꢀ11
ꢀ16
ꢀ10
ꢀ1
9
10
11
12
13
14
15
16
17
18
19
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
37.1%
27.9%
17.5 0.4
16.4 0.6
27.4%
14.3 0.2
29.9%
2.11 0.1
—
2
6
Thiourea 22.3 1.2
22
8
10
10
10
8
Table 5
Anticancer activity of N,N0-disubstituted thioureas (1–38) against lung carcinoma (H-
157) at 100
ꢀ19
ꢀ25
ꢀ20
ꢀ7
lM
8
19
10
2
13
1
4
14
4
1
5
4
17
Codes
%Inhibition
Codes
%Inhibition
ꢀ16
ꢀ13
ꢀ1
1
2
3
4
5
6
7
8
45.2
45.4
52.9
52.6
50.7
38.9
46.6
42.2
39.4
48.5
43.3
60.9
45.5
39.4
41.7
43.7
32.4
38.2
36.4
74.6
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
—
32.7
36.1
42.8
42.2
36.9
48.9
44.6
47.7
41.2
42.7
36.3
42.2
48.8
45.1
40.5
27.6
48.5
47.8
57.2
—
ꢀ11
ꢀ8
ꢀ5
ꢀ7
ꢀ14
ꢀ13
ꢀ11
ꢀ2
9
10
11
12
13
14
15
16
17
18
19
site. There was a common inclination for most of residues in the
active site to which all the compounds were interacting. It was
noticed that all the compounds show interactions with amino acids
Ala440, Arg609, Asp494, His409, His492, His519, Ala636, Met637
and His593. The hydrogen bonding interactions were identified
between the oxygen and hydrogen of the inhibitors with residues
Arg609, Asp494, Ala440 and His593 as described previously.39,56
Additionally, hydrophobic interactions of the inhibitors were rec-
ognized within the active site residues. Figure 2 reports a priori-
tized binding mode model of the potent inhibitor 16.
Vincristine
3.4.3.2. Binding mode analysis.
The putative binding mode
of compound 16, the most potent urease inhibitor was shown in
Figure 2. Trimethylphenyl substituent was located towards the
bottom of the binding pocket, where aromatic interactions with
the surrounding residues and the nickel ions in the deep pocket
were seen. While the 3-chlorobenzoyl side of the compound was
oriented towards residue Arg609 and Thr522. All the compounds
do not interact with any Ni atom in the active site, however, weak
interactions were found by most of the inhibitors with ni842 in the
catalytic site of urease enzyme. Figure 2 represents the alignment
of potent inhibitor inside the active pocket of 3LA4 by forming
hydrogen bond interactions with His593 and Arg609. Moreover,
p–p
interactions were noticed by trimethyl phenyl ring of the
derivative with active site residues.
Figure 2. Putative binding mode of 16 bound to the active site of Jack bean Urease.
Carbon atoms of 16 are colored blue and that of protein are light green. The two
nickel cations in the active site are represented as small dark green spheres.
3.4.3.3. HYDE assessment.
performed for first 30 top ranking docked conformations. The
The HYDE affinity assessment was
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12
M. K. Rauf et al. / Bioorg. Med. Chem. xxx (2016) xxx–xxx
23. (a) Nakamura, H.; Noh, J. Y.; Itoh, K.; Fukata, S.; Miyauchi, A.; Hamada, N. J. Clin.
binding free energy
able poses are given in Table 6 for all the derivatives. Most of the
compounds binds to the receptor with a very high binding affinity.
DG, FlexX docking score and their most favor-
Endocrinol. MeTable 2007, 92, 2157; (b) Duan, L.-P.; Xue, J.; Xu, L.-L.; Zhang, H.-
B. Molecules 2010, 15, 6941; (c) Chung, J. U.; Kim, S. Y.; Lim, J. O.; Choi, H. K.;
Kang, S. U.; Yoon, H. S.; Ryu, H. C.; Kang, D. W.; Lee, J.; Kang, B.; Choi, S.; Toth,
A.; Pearce, L. V.; Pavlyukovets, V. A.; Lundberg, D. J.; Blumberg, P. M. Bioorg.
Med. Chem. 2007, 15, 6043.
4. Conclusion
24. (a) Kearney, P. C.; Frenandez, M.; Flygare, J. A. J. Org. Chem. 1998, 63, 196; (b)
Boga, C.; Forlani, L.; Silvestroni, C.; Corradi, A. B.; Sagarabotto, P. J. Chem. Soc.,
Perkin Trans. 1 1999, 1363.
A series of N,N0-disubsituted thiourea analogues were prepared
using facile microwave-assisted solution phase parallel synthesis.
The synthesized compounds were evaluated for in vitro urease
inhibitory activity. Most of the compounds exhibited excellent
urease inhibition, among which, compound 16 shows potent
25. Patil, D. G.; Chedekel, M. R. J. Org. Chem. 1984, 49, 997.
26. Kasmi, S.; Hamelin, J.; Benhaoua, H. Tetrahedron Lett. 1998, 39, 8093.
27. (a) Kidwai, M.; Venkataramanan, R.; Dave, B. Green Chem. 2001, 3, 278; (b) Paul,
S.; Gupta, M.; Gupta, R.; Loupy, A. Synthesis 2002, 75.
28. Anshu, D.; Kapil, A.; Meha, S. Synth. Commun. 2004, 34, 1141.
29. Heinelt, U.; Schultheis, D.; Jager, S.; Iindenmaire, M.; Pollex, A.; Beckmann, H. S.
G. Tetrahedron 2004, 60, 9883.
urease inhibitory activity with an IC50 value 1.23 0.1 lM. All
30. (a) Frank, R. L.; Smith, P. V. Org. Synth. Coll. III 1955, 735; (b) Rasmussen, C. R.;
Villani, F. J. J.; Weaner, L. E.; Reynolds, B. E.; Hood, A. R.; Hecker, L. R.; Nortey, S.
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the synthesized derivatives were screened for their anticancer
activities. Compound 12 was the potent compound against H157
with 60.91% inhibition. The docking studies further unlocked the
binding site interactions of the potent inhibitors. Furthermore,
the results suggest that these derivatives may be used as preven-
tive and chemotherapeutic agents for cancer. Hence it is recom-
mended to find the correlation of urease inhibitors and their role
in the therapy of cancer.
33. Erickson, J. G. J. Org. Chem. 1956, 21, 483.
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Acknowledgements
The M. Khawar Rauf is thankful to the Quaid-i-Azam University,
Islamabad, Pakistan for the grant of funds for Post-doctoral Fellow-
ship. Jamshed Iqbal is thankful to the Organization for the Prohibi-
tion of Chemical Weapons (OPCW), The Hague, The Netherlands
and the Higher Education Commission of Pakistan for the financial
support through Project No. 20-3733/NRPU/R&D/14/520.
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Please cite this article in press as: Rauf, M. K.; et al. Bioorg. Med. Chem. (2016), http://dx.doi.org/10.1016/j.bmc.2016.07.042