Synthesis and antimicrobial activity of 5-((3-aryl-1-phenyl-1Hpyrazol-4-yl) methylene)thiazolidine-2,4-diones

Article abstract of DOI:10.1007/s00044-011-9829-4

A series of 5-((3-aryl-1-phenyl-1H-pyrazol-4- yl)methylene)thiazolidine-2, 4-diones was synthesized by Knoevenagel condensation of various 3-aryl-1-phenyl- 1H-pyrazole-4-carbaldehydes (1a-h) with thiazolidine-2,4- dione (2) in ethanol in the presence of piperidine. All compounds were screened for their in vitro antibacterial (Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa and Escherichia coli) activity and compared with the commercially available antibiotic, ciprofloxacin. All compounds showed good activity against gram-positive bacteria, however, none of the compounds were found to be effective against gram-negative bacteria. Compound 3g was found to be most potent member among all the compounds showing MIC of 16 μg/ml against S. aureus and 32 μg/ml against B. subtilis. All the synthesized compounds were also tested for their in vitro antifungal (Aspergillus niger and A. flavus) activity and compared with commercially available fluconazole. They showed excellent antifungal activity. Compounds 3b, 3e, 3f and 3g were active against both fungi showing more than 60% inhibition. Compound 3e was found to be superior to the reference drug.

Full text of DOI:10.1007/s00044-011-9829-4

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res (2012) 21:2961–2968
DOI 10.1007/s00044-011-9829-4
ORIGINAL RESEARCH
Synthesis and antimicrobial activity of 5-((3-aryl-1-phenyl-1H-
pyrazol-4-yl)methylene)thiazolidine-2,4-diones
•
Om Prakash Deepak K. Aneja Poonam Lohan
•
•
•
•
•
Khalid Hussain Sanjiv Arora Chetan Sharma
Kamal R. Aneja
Received: 25 May 2011 / Accepted: 20 October 2011 / Published online: 3 November 2011
Ó Springer Science+Business Media, LLC 2011
Abstract A series of 5-((3-aryl-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-diones was synthesized by
Knoevenagel condensation of various 3-aryl-1-phenyl-
1H-pyrazole-4-carbaldehydes (1a–h) with thiazolidine-2,4-
dione (2) in ethanol in the presence of piperidine. All
compounds were screened for their in vitro antibacterial
(Staphylococcus aureus, Bacillus subtilis, Pseudomonas
aeruginosa and Escherichia coli) activity and compared
with the commercially available antibiotic, ciprofloxacin.
All compounds showed good activity against gram-positive
bacteria, however, none of the compounds were found to
be effective against gram-negative bacteria. Compound 3g
was found to be most potent member among all the com-
pounds showing MIC of 16 lg/ml against S. aureus and
32 lg/ml against B. subtilis. All the synthesized com-
pounds were also tested for their in vitro antifungal
(Aspergillus niger and A. flavus) activity and compared
with commercially available fluconazole. They showed
excellent antifungal activity. Compounds 3b, 3e, 3f and 3g
were active against both fungi showing more than 60%
inhibition. Compound 3e was found to be superior to the
reference drug.
Keywords Pyrazole ꢀ Thiazolidine-2,4-dione ꢀ
Antibacterial activity ꢀ Antifungal activity
Introduction
Thiazolidines have been the subject of extensive research
because of their biological activities associated with this
class of compounds (Bhargava and Chaurasia, 1969). Sev-
eral studies have been reported that 2,4-thiazolidinedione
derivatives have acquired much importance because of their
diverse pharmaceutical applications such as antihypergly-
cemic (Lee et al., 2005), bactericidal (Mahalle et al., 2008),
pesticidal (Hyo et al., 2007), fungicidal (Deohate et al.,
2004), insecticidal (Sahu et al., 2007), anticonvulsant
(Altintas et al., 2006), tuberculostatic (Thaker et al., 2003),
anti-inflammatory (Szeto and K-T 2008, Pastromas et al.,
2008; Murugan et al., 2009), antithyroidal and potentiation
of pentobarbital induced sleeping time, etc. (Metwally et al.,
2006).
The varied biological activities of rhodanines (2-thioxo-
thiazolidin-4-one) and their analogues have been known
from the beginning of 20th century. Rhodanines and
2,4-thiazolidinediones have become a pharmacologically
important class of heterocyclic compounds since the intro-
duction of various glitazone and epalrestat into clinical use
for the treatment of type II diabetes and diabetic complica-
tions, respectively (Terashima et al., 1984). Different pos-
sibilities of heterocyclic modifications with a wide spectrum
of pharmacological properties are the most important
grounds for investigation of this class of compounds
(Bozdag-Dundar et al., 2008; Chandrappa et al., 2008; Jeon
et al., 2006; Tuncbilek and Altanlar 2006; Ramsh et al. 2004;
Clark et al., 1991). There have been many reports in litera-
ture depicting that the presence of heterocyclic moieties like
O. Prakash (&)
Institute of Pharmaceutical Sciences, Kurukshetra University,
Kurukshetra 136119, India
e-mail: dromprakash50@rediffmail.com
D. K. Aneja ꢀ P. Lohan ꢀ K. Hussain ꢀ S. Arora
Department of Chemistry, Kurukshetra University,
Kurukshetra 136119, India
e-mail: dk_aneja@rediffmail.com
C. Sharma ꢀ K. R. Aneja
Department of Microbiology, Kurukshetra University,
Kurukshetra 136119, India
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Med Chem Res (2012) 21:2961–2968
Scheme 1 Synthesis of
pyrazolyl-2,4-
thiazolidinediones (3a–h) using
Knoevenagel condensation
O
N
EtOH/ Piperidine
reflux, 5-6 h
N
N
R
N
S
R
+
NH
H
S
H
O
O
O
N
O
H
3
1
2
thienyl, furyl, pyridyl and pyrimidinyl at fifth position
proves to be more potent and efficacious than a simple aryl
group (Rawal et al., 2007; El-Gaby et al., 2009; Kaneria
et al., 2003). Although there are not many thiazolidinediones
fused to pyrazoles, a number of them are incorporated into a
wide variety of therapeutically important compounds pos-
sessing a broad spectrum of biological activities. In a recent
report, pyrazolyl-2,4-thiazolidinediones have been reported
as anti-inflammatory and neuroprotective agents (Youssef
et al., 2010). These observations encouraged us to undertake
the synthesis of heterocyclic compounds containing pyra-
zole moiety (Prakash et al., 2008, 2009a, b, 2010a, b). We
report herein the synthesis of 5-((3-aryl-1-phenyl-1H-pyra-
zol-4-yl)methylene)thiazolidine-2,4-diones (3a–h) with an
aim to find new and more potent antibacterial and antifungal
agents.
Table 1 Physical data of 5-((3-aryl-1-phenyl-1H-pyrazol-4-yl)
methylene)thiazolidine-2,4-diones (3a–h)
Compounds
R
H
Mp (°C) Lit. Mp (°C)
Yields %
85
3a
292–294 263–265
(Youssef et al., 2010)
3b
3c
3d
CH3
288–290
–
–
94
90
88
OCH3 293–295
Cl
325–327 280–282
(Youssef et al., 2010)
3e
3f
F
298–300
301–303
299–301
323–325
–
–
–
–
92
93
83
88
Br
3g
3h
NO2
OH
formed between the hydrogen bond of =CH and oxygen
atom in TZD (El-Deeb et al., 2010; Prashantha Kumar
et al., 2011). Some of the characteristics values of 1H
NMR, 13C NMR and IR of compound 3c are shown in
Fig. 1 for illustration.
Results and discussion
Chemistry
The starting material thiazolidine-2,4-dione (2) was
prepared according to literature procedure in one step from
thiourea and 2-chloroacetic acid in concentrated hydro-
chloric acid (Brown, 1961). 4-Formylpyrazoles 1a–h were
prepared according to literature procedure involving Vils-
meier-Haack reaction of various substituted acetophenone
hydrazones using POCl3/DMF at 50–60°C for 4–5 h (Kira
et al., 1969).
The synthetic route followed for the preparation of the
5-((3-aryl-1-phenyl-1H-pyrazol-4-yl)methylene)thiazolidine-
2,4-diones (3a–h) illustrated in Scheme 1 involves
condensation of the 3-aryl-1-phenyl-1H-pyrazole-4-carbalde-
hydes (1a–h) and thiazolidine-2,4-dione (2). Piperidine in
ethanol was used as a catalyst in this condensation. The
desired products 3a–h were obtained in excellent yields as
shown in Table 1. Although, geometrical isomerism (E/Z-
isomers) was possible because of restricted rotation about
the exocyclic C=C bond of the pyrazolyl-2,4-thiazolididi-
ones, all the derivative prepared in this study were obtained
exclusively in Z-form as confirmed by the analytical data.
The 1HNMR spectra of the pyrazolyl-2,4-thiazolidindiones
showed the most characteristics olefinic proton =CH was
deshielded more (d = 7.3–7.6 ppm) as expected in Z-form,
relative to the slightly shielded protons of the E-form
(d = 6.2–6.3 ppm). This deshielding of the olefinic proton
is caused by the anisotropic effect exerted by the nearby
carbonyl group of the 2,4-thiazolidinediones in Z-isomer.
Furthermore, the Z-isomers are thermodynamic more stable
because of intramolecular hydrogen bond that can be
It is to be mentioned that the compounds 3a and 3d have
been reported recently by Youssef et al., 2010. They syn-
thesized 3a and 3d by the condensation in anhydrous tol-
uene in the presence of glacial acetic acid and piperidine.
The spectral data (IR, 1H NMR and 13C NMR of the
compounds 3a and 3d reported in literature match with the
spectral data of our products. However, melting points of
3a (263–265°C) and 3d (280–282°C) (Youssef et al., 2010)
were found to be different from melting points of products
synthesized by us (3a, 292–294°C and 3d, 325–327°C).
Therefore, it became essential to synthesize authentic
sample of 3a and 3d by using reported conditions. Since,
TLC and spectral data of the compounds prepared by the
known procedures (Youssef et al., 2010) were identical to
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N
N
O
CH3
H
3.834 (s, 3H)
8.623 (s, 1H)
H
In 1H NMR (ppm)
S
7.971-7.991 (Mixed with aromatic protons)
O
N
O
H
12.4-12.6 (1H, bs)
N
N
O
55.42
In 13C NMR (ppm)
139.20
S
O
O
N
H
168.56
167.86
N
N
O
CH3
In IR (cm-1)
S
1690
O
N
O
H
1744
1
Fig. 1 Characteristics value of H NMR, 13C NMR and IR of compound 3c
those obtained by our conditions, the reported melting
points might be in error.
Table 2 In vitro antibacterial activity of compounds (3a–h) through
agar well diffusion method
Compounds
Diameter of growth of inhibition zone (mm)a
S. aureus
B. subtilis
E. coli
P. aeruginosa
Biological evaluations
3a
20.3
16.3
13.6
14.3
20.6
18.6
23.6
15.0
26.0
16.0
15.6
14.3
15.0
21.6
21.6
21.3
14.6
24.0
–
–
In vitro antibacterial activity
3b
–
–
3c
–
–
All the compounds 3a–h were screened for their in vitro
antibacterial activity against two gram-positive bacte-
ria namely, Staphylococcus aureus (MTCC 96), Bacillus
subtilis (MTCC 121) and two gram-negative bacteria
namely, Escherichia coli (MTCC 1652) and Pseudomonas
aeruginosa (MTCC 741) (Table 2). Minimum inhibitory
concentration (MIC) of those compounds was determined
which are showing activity in primary screening (Table 3).
Standard antibiotic namely, ciprofloxacin was used for
comparison with antibacterial activity shown by compounds
3a–h. The results were recorded for each tested compounds
as the average diameter of the zones of inhibition of bacterial
growth surrounding the wells in millimetre.
3d
–
–
3e
–
–
3f
–
–
3g
–
–
3h
–
–
Ciprofloxacin
25.0
22.0
– No activity
a
Values including diameter of the well (8 mm) are means of three
replicates
From the results, it has been revealed that all the tested
compounds possess moderate to good antimicrobial activity
against both gram-positive bacteria (S. aureus and B. subtilis)
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Table 3 MIC (lg/ml) of compounds (3a–h) by using macro dilution
method
S. aureus
B. subtilis
140
120
100
80
Compounds
S. aureus
B. subtilis
3a
32
128
128
128
32
128
128
128
128
32
3b
60
3c
40
20
3d
0
3e
3f
64
32
3g
16
32
3h
128
5
128
5
Compounds/ Antibiotic
Ciprofloxacin
Fig. 2 Comparison of MIC of tested compounds (3a–h) with
antibiotics up to MIC 128 (lg/ml)
(Table 2). However, none of them was found to be effective
against gram-negative bacteria (E. coli and P. aeruginosa).
On the basis of zone of inhibition produced against the
test bacterium, compounds 5-((3-(4-fluorophenyl)-1-phenyl-
1H-pyrazol-4-yl)methylene)thiazolidine-2,4-dione (3e), 5-((3-
(4-bromophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)
thiazolidine-2,4-dione (3f) and 5-((3-(4-nitrophenyl)-1-phenyl-
1H-pyrazol-4-yl)methylene)thiazolidine-2,4-dione (3g) were
found to be most effective. The compound 3g showed the
maximum zone of inhibition of 24 mm against S. aureus and
21.3 mm against B. subtilis in comparison of ciprofloxacin
which produced the zone of inhibition 26 mm against
S. aureus and 24 mm against B. subtilis while the compound
3e which shows the maximum zone of inhibition of 20.6 mm
against S. aureus and 21.6 mm against B. subtilis. The
compound 3f which was found to be less potent than 3e and
3g showed the maximum zone of inhibition of 18.6 mm
against S. aureus and 21.6 mm against B. subtilis. In addition
to these, compounds 3a and 3b showed moderate antibac-
terial activity against S. aureus and B. subtilis with zone of
inhibition[16 mm.
In the whole series, the MIC of various tested compounds
ranged between 16 and 128 lg/ml against gram-positive
bacteria. The compound 3g was found to be best, showed the
lowest MIC of 16 lg/ml against S. aureus and compounds 3e,
3f and 3g showed the lowest MIC of 32 lg/ml against
B. subtilis (Table 3, Fig. 2). The reason for different sensi-
tivity between gram-positive and gram-negative bacteria
could be ascribed to the morphological difference between
these micro-organisms. Gram-negative bacteria have an outer
phospholipidic membrane carrying the structural lipopoly-
saccharide components. This makes the cell wall imperme-
able to lipophilic solutes, while porins constitute a selective
barrier to the hydrophilic solutes with an exclusion limit of
about 600 Da (Nikaido and Vaara, 1985). The Gram-positive
bacteria had been found to be more susceptible since they
have only an outer peptidoglycan layer which is not an
effective permeability barrier (Scherrer and Gerhardt 1971).
Table 4 In vitro antifungal
Compounds Mycelial growth
activity of compounds (3a–h)
inhibition (%)
through poisoned food method
A. flavus A. niger
3a
3b
3c
3d
3e
3f
60.0
65.5
57.7
60.0
78.8
62.5
63.3
54.4
62.5
68.8
60.0
65.5
82.5
61.1
61.1
55.5
81.1
3g
3h
Fluconazole 77.7
In vitro antifungal activity
All the compounds were also tested for their in vitro anti-
fungal activity against two fungi namely, Aspergillus flavus
and A. niger (Table 4, Fig. 3). Fluconazole was used as ref-
erence drug for comparison of the activity of the tested
compounds and the results were recorded as the percentage of
the mycelial growth inhibitions. From the results, it has been
revealed that most of the compounds possessed remarkable
antifungal activity against both fungal strains. When com-
pared with standard drug which showed 77.7 and 81.1%
mycelial growth inhibition against A. flavus and A. niger,
respectively, it was found that compound 3e showed excellent
activity with 78.8 and 82.5% inhibition of mycelial growth
against A. flavus and A. niger, respectively. In addition to this,
three compounds 3b, 3f and 3g showed comparable activity
with[60% inhibition of mycelial growth against both fungi
while remaining compounds showed moderate activity.
So, in whole series tested, compound 3e was found to the
most potent antifungal agent among all members having
greater activity than commercially available drug (Table 4,
Fig. 3).
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(300 MHz, DMSO-d6): 12.6–12.4 (bs, 1H, NH, exch.
D2O), 8.664 (s, 1H, Pyrazolyl-H), 8.015–7.989 (m, 2H),
7.681–7.623 (m, 2H), 7.584–7.491 (m, 6H), 7.430–7.380
(m, 1H); 13C NMR d (75 MHz, DMSO-d6): 168.95,
168.61, 153.93, 139.27, 131.90, 130.09, 129.40, 129.13,
128.33, 127.89, 124.41, 121.57, 119.80 and 116.25; Anal.
Found: C, 65.5; H, 3.66; N, 11.9. C19H13N3O2S (347.39)
requires C, 65.69; H, 3.77; N, 12.10%.
A. flavus A. niger
90
80
70
60
50
40
30
20
10
0
(Z)-5-((1-phenyl-3-p-tolyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3b)
Compounds/ Antifungal Drug
IR cm-1 (KBr): 1736 (C=O, thiazolidinedione ring), 1682
(C=O, thiazolidinedione ring), 1597, 1528, 1450, 1412,
Fig. 3 Comparison of mycelial growth inhibition of tested com-
pounds (3a–h) with antifungal drugs
1
1327, 1234, 1149, 1065 and 1034; H NMR d (300 MHz,
DMSO-d6): 12.6–12.4 (bs, 1H, NH, exch. D2O), 8.646 (s,
1H, Pyrazolyl-H), 7.976–8.002 (d, 2H, J = 7.8 Hz), 7.571–
7.344 (m, 8H), 2.387 (s, 3H, CH3); 13C NMR d (75 MHz,
DMSO-d6): 154.07, 139.15, 130.13, 129.98, 128.97,
128.23, 127.93, 119.75 and 21.27; Anal. Found: C, 66.30;
H, 4.30; N, 11.45. C20H15N3O2S (361.42) requires C,
66.46; H, 4.18; N, 11.63%.
Experimental
General remarks
Melting points were taken on slides in an electrical appa-
ratus Labindia visual melting range apparatus and are
uncorrected. IR spectra were recorded on a Perkin-Elmer
1800 FT-IR spectrophotometer.1H NMR and 13C NMR
spectra were recorded on a Bruker 300 and 75 MHz
instrument, respectively, using TMS as an internal stan-
dard. All other chemicals used as such as were procured
from supplier.
(Z)-5-((3-(4-methoxyphenyl)-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3c)
IR cm-1 (KBr): 1744 (C=O, thiazolidinedione ring), 1690
(C=O, thiazolidinedione ring), 1589, 1528, 1450, 1319,
1
1242, 1180, 1142, 1068 and 1018; H NMR d (300 MHz,
DMSO-d6): 12.6–12.4 (bs, 1H, NH, exch. D2O), 8.623 (s,
1H, Pyrazolyl-H), 7.991-7.971 (m, 2H), 7.566–7.519
(m, 5H), 7.418–7.369 (m, 1H), 7.122–7.094 (d, 2H,
J = 8.4 Hz), 3.834 (s, 3H, OCH3). 13C NMR d (75 MHz,
DMSO-d6): 168.56, 167.86, 154.12, 139.20, 130.18,
129.88, 128.78, 128.32, 127.97, 119.87 and 55.42; Anal.
Found: C, 63.5; H, 4.2; N, 11.10. C20H15N3O3S (377.42)
requires C, 63.65; H, 4.01; N, 11.13%.
Preparation of (Z)-5-((3-aryl-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-diones (3a–h)
A mixture of 1,3-diphenyl-1H-pyrazol-4-carboxaldehyde
(1a, 1g, 4 mmol) and thiazolidine-2,4-dione (2, 0.51 g,
5 mmol) in ethanol (20 ml) and 2–3 drops of piperidine,
was refluxed. A solid separated out of the reaction mixture
within 15–20 min and the refluxing was continued for
5–6 h to complete the reaction. The reaction mixture was
cooled to room temperature, filtered and washed with
ethanol to give the pure product 3a (1.25 g, 90% yield).
The other derivatives 3b–h were synthesized by adopt-
ing the similar procedure.
(Z)-5-((3-(4-chlorophenyl)-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3d)
IR cm-1 (KBr): 1751 (C=O, thiazolidinedione ring), 1690
(C=O, thiazolidinedione ring), 1605, 1504, 1450, 1412,
1335, 1227, 1142, 1095, 1057 and 1018; 1H NMR d
(300 MHz, DMSO-d6): 12.7–12.5 (bs, 1H, NH, exch. D2O),
8.694 (s, 1H, Pyrazolyl-H), 8.019–7.993 (m, 2H), 7.710–
7.665 (m, 2H), 7.586–7.529 (m, 3H), 7.440–7.384 (m, 3H);
13C NMR d (75 MHz, DMSO-d6): 168.12, 167.91, 153.00,
139.21, 131.34, 131.23, 130.09, 128.51, 128.31, 127.97,
122.06, 119.84, 116.58, 116.29 and 116.05; Anal. Found: C,
59.65; H, 3.36; N, 10.8. C19H12ClN3O2S (381.84) requires
C, 59.76; H, 3.17; N, 11.00%.
Characterization data of synthesized compounds (3a–h)
(Z)-5-((1,3-diphenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3a)
IR cm-1 (KBr): 1736 (C=O, thiazolidinedione ring), 1690
(C=O, thiazolidinedione ring), 1605, 1528, 1504, 1443,
1
1412, 1366, 1327, 1234, 1149, 1065 and 1026; H NMR d
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(Z)-5-((3-(4-fluorophenyl)-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3e)
7.459–7.366 (m, 3H), 6.951–6.923 (d, 2H, J = 8.4 Hz),
13C NMR d (75 MHz, DMSO-d6): 154.01, 139.05, 130.03,
129.88, 128.77, 128.13, and 127.83; Anal. Found: C, 62.5;
H, 3.75; N, 11.45. C19H13N3O3S (363.39) requires C,
62.80; H, 3.61; N, 11.56%.
IR cm-1 (KBr): 1759 (C=O, thiazolidinedione ring), 1697
(C=O, thiazolidinedione ring), 1589, 1520, 1443, 1404,
1
1327, 1234, 1149, 1065 and 1003; H NMR d (300 MHz,
DMSO-d6): 12.8–12.7 (bs, 1H, NH, exch. D2O), 8.653
(s, 1H, Pyrazolyl-H), 7.999–7.972 (d, 2H, J = 8.1 Hz), 7.770–
7.744 (d, 2H, J = 7.8 Hz), 7.598–7.496 (m, 5H), 7.431–7.382
(m, 1H). 13C NMR d (75 MHz, DMSO-d6): 168.22, 168.11,
154.00, 139.31, 131.44, 131.33, 130.19, 128.62, 128.42,
127.88, 122.02, 119.78, 116.78, 116.55 and 116.36; Anal.
Found: C, 62.34; H, 3.52; N, 11.6. C19H12FN3O2S (365.38)
requires C, 62.46; H, 3.31; N, 11.50%.
Biological assay
Test microorganisms
Four bacteria, S. aureus (MTCC 96), B. subtilis (MTCC
121) (gram-positive), E. coli (MTCC 1652) and P. aeruginosa
(MTCC 741) (gram-negative) procured from Microbial
Type Culture Collection (MTCC), Chandigarh and two
fungi, A. niger and A. flavus, the ear pathogens isolated
from the Kurukshetra patients, were used in the present
study (Aneja et al., 2010).
(Z)-5-((3-(4-bromophenyl)-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3f)
IR cm-1 (KBr): 1751 (C=O, thiazolidinedione ring), 1690
(C=O, thiazolidinedione ring), 1597, 1528, 1497, 1450,
In vitro antibacterial activity
1
1412, 1335, 1227, 1142, 1088, 1057 and 1011; H NMR d
(300 MHz, DMSO-d6): 12.5–12.7 (bs, 1H, NH, exch.
D2O), 8.703 (s, 1H, Pyrazolyl-H), 8.019–7.993 (d, 2H,
J = 7.8 Hz), 7.656–7.647 (m, 4H), 7.588–7.543 (m, 3H),
7.443–7.396 (m, 1H). 13C NMR d (75 MHz, DMSO-d6):
167.97, 167.56, 152.69, 139.16, 134.37, 130.81, 130.10,
129.52, 128.69, 128.04, 123.62, 122.14, 119.88 and 116.04.
Anal. Found: C, 53.45; H, 2.96; N, 9.77. C19H12
BrN3O2S (426.29) requires C, 53.53; H, 2.84; N, 9.86%.
The antibacterial activity of synthesized compounds was
evaluated by the agar well diffusion method. All the cul-
tures were adjusted to 0.5 McFarland standard, which is
visually comparable to a microbial suspension of approx-
imately 1.5 9 108 CFU/ml. 20 ml of Mueller–Hinton agar
medium was poured into each Petri plate and the agar
plates were swabbed with 100 ll inocula of each test
bacterium and kept for 15 min for adsorption. Using sterile
cork borer of 8 mm diameter, wells were bored into the
seeded agar plates and these were loaded with a 100 ll
volume with concentration of 4.0 mg/ml of each compound
reconstituted in the dimethylsulphoxide (DMSO). All the
plates were incubated at 37°C for 24 h. Antibacterial
activity of each synthetic compound was evaluated by
measuring the zone of growth inhibition against the test
organisms with zone reader (Hi Antibiotic zone scale).
DMSO was used as a negative control whereas ciproflox-
acin was used as a positive control. This procedure was
performed in three replicate plates for each organism
(Ahmad and Beg, 2001; Andrews 2001).
(Z)-5-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3g)
IR cm-1 (KBr): 1748 (C=O, thiazolidinedione ring), 1690
(C=O, thiazolidinedione ring), 1574, 1512, 1450, 1342 and
1
1227; H NMR d (300 MHz, DMSO-d6): 12.8–12.7 (bs,
1H, NH, exch. D2O), 8.537 (s, 1H, Pyrazolyl-H), 8.412–
8.386 (m, 2H), 7.977–7.929 (m, 4H), 7.671–7.284 (m, 4H);
13C NMR d (75 MHz, DMSO-d6): 167.89, 167.66, 152.68,
139.25, 134.41, 130.91, 130.22, 129.62, 128.58, 128.12,
123.25, 122.12, 119.98 and 116.25; Anal. Found: C, 58.0;
H, 2.98; N, 14.20. C19H12N4O4S (392.39) requires C,
58.16; H, 2.84; N, 14.28%.
Determination of minimum inhibitory concentration
(MIC)
(Z)-5-((3-(4-hydroxyphenyl)-1-phenyl-1H-pyrazol-4-
yl)methylene)thiazolidine-2,4-dione (3h)
Minimum inhibitory concentration is the lowest concen-
tration of an antimicrobial compound that will inhibit the
visible growth of a microorganism after overnight incu-
bation. MIC of the various compounds against bacterial
strains was tested through a macro dilution tube method as
recommended by NCCLS (2000). In this method, various
test concentrations of synthesized compounds were made
from 128 to 0.25 lg/ml in sterile tubes 1–10. 100 ll sterile
IR cm-1 (KBr): 1751 (C=O, thiazolidinedione ring), 1690
(C=O, thiazolidinedione ring), 1589, 1528, 1450, 1319,
1242, 1142, 1065 and 1026; 1H NMR d (300 MHz,
DMSO-d6): 12.6–12.4 (bs, 1H, –NH, exch. D2O), 9.864
(bs, 1H, –OH, exch. D2O), 8.609 (s, 1H, Pyrazolyl-H),
7.992–7.966 (d, 2H, J = 7.8 Hz), 7.570–7.541 (m, 3H),
123

Med Chem Res (2012) 21:2961–2968
2967
India for providing financial assistance to carry out this study. One of
the authors Deepak K. Aneja is grateful to the UGC, New Delhi, India
for providing JRF. Thanks are due to CDRI, Lucknow for carrying out
elemental analysis.
Mueller–Hinton Broth was poured in each sterile tube
followed by addition of 200 ll test compound in tube 1.
Two fold serial dilutions were carried out from the tube no.
1 to the tube no. 10 and excess broth (100 ll) was dis-
carded from the last tube no. 10. To each tube, 100 ll
of standard inoculums (1.5 9 108 CFU/ml) was added.
Ciprofloxacin was used as control. Turbidity was observed
after incubating the inoculated tubes at 37°C for 24 h.
Conflict of interest The authors have declared no conflict of
interest.
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