Synthesis and biological evaluation of novel triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene thiazolidinone derivatives as antibacterial and cytotoxic agents

Article abstract of DOI:10.1007/s11164-017-2978-4

Novel triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene thiazolidinone derivatives 6a–n and 7a–l were synthesized and characterized by Fourier transform infrared, ¹H and ¹³C nuclear magnetic resonance, mass spectrometry and elemental (CHN) analysis. The in vitro antibacterial (6a–n and 7a–l) and cytotoxic (6a–n) activities were evaluated for these compounds. The results revealed that the compounds 6b, 6i, 6k, 7b, 7h displayed good antibacterial activity. The compounds 6c (IC₅₀?=?5.4?μM), 6l (IC₅₀?=?6.3?μM) and 6f (IC₅₀?=?9.85?μM) were effective for inhibition of human breast cancer cell line MCF-7. Similarly, the compounds 6b (IC₅₀?=?8.7?μM) and 6c (IC₅₀?=?9.06?μM) were shown to have effective inhibition on human cervical cancer cell line Hela.

Full text of DOI:10.1007/s11164-017-2978-4

Res Chem Intermed
DOI 10.1007/s11164-017-2978-4
Synthesis and biological evaluation of novel triazole
substituted pyrazolyl-methylenehydrazinyl-5-arylidene
thiazolidinone derivatives as antibacterial and cytotoxic
agents
Vinay Pogaku1 Ravi Kumar Eslavath2 G. Dayakar3
•
•
•
Surya S. Singh2 Srinivas Basavoju1
•
Received: 31 December 2016 / Accepted: 3 May 2017
Ó Springer Science+Business Media Dordrecht 2017
Abstract Novel triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene
thiazolidinone derivatives 6a–n and 7a–l were synthesized and characterized by
1
Fourier transform infrared, H and 13C nuclear magnetic resonance, mass spec-
trometry and elemental (CHN) analysis. The in vitro antibacterial (6a–n and 7a–l)
and cytotoxic (6a–n) activities were evaluated for these compounds. The results
revealed that the compounds 6b, 6i, 6k, 7b, 7h displayed good antibacterial activity.
The compounds 6c (IC50 = 5.4 lM), 6l (IC50 = 6.3 lM) and 6f (IC50 = 9.85 lM)
were effective for inhibition of human breast cancer cell line MCF-7. Similarly, the
compounds 6b (IC50 = 8.7 lM) and 6c (IC50 = 9.06 lM) were shown to have
effective inhibition on human cervical cancer cell line Hela.
Keywords 3-Methyl-1-phenyl-1H-pyrazol-5(4H)-one Á 1H-1,2,4-triazole Á
1H-benzo[d][1,2,3]triazole Á Pyrazolyl-methylenehydrazinyl-5-arylidene
thiazolidinones Á Antibacterial Á Cytotoxic activity
Electronic supplementary material The online version of this article (doi:10.1007/s11164-017-2978-4)
contains supplementary material, which is available to authorized users.
&
Srinivas Basavoju
basavoju_s@yahoo.com
1
Department of Chemistry, National Institute of Technology, Warangal, Telangana 506 004,
India
2
3
Department of Biochemistry, Osmania University, Hyderabad, Telangana 500 007, India
Department of Biotechnology, Kakatiya University, Warangal, Telangana 506 009, India
123

V. Pogaku et al.
Introduction
A number of life-threatening infectious diseases are caused by Gram-positive and
Gram-negative bacteria [1]. These infections are the most prominent death-causing
diseases in the world, because of their ability to spread rapidly and develop their
own resistance towards multiple drugs. According to recent studies, an increase of
antibiotic resistance by the Gram-positive bacteria and Gram-negative bacteria [2],
raises the global alarm and explains the importance and urgent need for the
development of more potent, novel broad spectrum antibacterial drugs with fewer
side effects and improved efficacy to cure microbial infections. On the other hand,
based on the World Health Organization reports, cancer is a second leading
prominent death cause disease of humanity [3]. The literature shows that anticancer
drugs have many limitations, such as drug resistance, exhibiting many side effects
and more systematic toxicity and no 100% efficacy and isolation procedures [4, 5].
Therefore, it is essential to develop novel and inexpensive anticancer agents with
higher activity and lower side effects.
Pyrazole scaffolds are important heterocyclic moieties, and they occupy a major
position in medicinal and pesticide chemistry due to their various biological
activities such as antibacterial [6, 7], antiparasitic [8], antiviral [9], antineoplastic
[10], antitumor, anti-inflammatory [11, 12], antimalarial [13], analgesic [14],
anticonvulsant [15], anti-depressant [16] and antituberculosis [17] activity. Pyra-
zolylthiazoles were reported as cardiovascular drugs [18] and these derivatives also
showed antinociceptive activity [19] and phototoxicity [20]. Lee et al., introduced
various N-containing heterocycles by replacing the chlorine atom at 5-position on
3-methyl-5-chloropyrazole-3-carboxaldehyde by nucleophilic aromatic substitution
[21]. A significant work by Kalaria et al. [22], on pyrazole-based thiazolopyran
derivatives showed antimicrobial and antituberculosis activity. However, the
pyrazole derivatives have also been used as insecticides [23], fungicides [24, 25],
and herbicides [26–29]. Some of the fungicides such as pyrazophos, furametpyr,
penthiopyrad and pyraclostrobin are available on the market. Prajapati and
coworkers synthesized the fused pyrazolinoquinolizine and 1,4-oxazinopyrazoline
derivatives by substituting several cyclic sec-amines (viz pyrrolidine, piperidine and
morpholine) at the 5-position of pyrazole ring [30]. Not only heterocyclic rings but
also phenylthio groups have been attached to the pyrazole ring at the 5-position
showing antiviral (Tobacco mosaic virus) activity [31].
Furthermore, recent literature embraces the 4-thiazolidinone and its arylidene
derivatives due to their wide range of biological and pharmacological activities such
as antidiabetic [32]. antibacterial [33, 34], antifungal [35], anti-inflammation [36],
antihypertension [37], anti-HIV [38], treatment of pain [39], antihistaminic [40],
anticancer [41, 42] and inhibition of BRD-4 [43], with inhibition of bacterial
enzyme MurB [44, 45]. The reported 2-thiazolylimino-5-arylidene-4-thiazolidi-
nones showed antimicrobial activity [46]. Lima and co-workers have synthesized
5-benzylidene-2-[(pyridine-4-ylmethylene)hydrazono]-thiazolidin-4-one derivatives
and evaluated them for their antibiotic and cytotoxic activities [47]. The above-
mentioned results from the literature encouraged us to design triazole substituted
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Synthesis and biological evaluation of novel triazole…
pyrazolyl moieties linking with methylenehydrazinyl-5-arylidene thiazolidinones
expecting antibacterial and cytotoxic activities. Therefore, in this paper we report
the synthesis of triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene
thiazolidinone derivatives 6a–n and 7a–l with their antibacterial and cytotoxic
activities.
Results and discussion
Synthesis
The starting material 5-chloro-3-methyl-1-phenyl-1H-pyrazole-4-carbaldehyde 1
was prepared from 3-methyl-1-phenyl-1H-pyrazol-5(4H)-one, via Vilsmeier-Haack
formylation reaction. The compounds 3a, 3b were prepared by the nucleophilic
substitution of the compound 1 with 1H-1,2,4-triazole 2a and 1H-benzo[d][1,2,3]-
triazole 2b under reflux condition in DMF and anhydrous K2CO3 for 4 h.
Furthermore, compounds 3a, 3b on condensation with thiosemicarbazide a catalytic
amount of glacial acetic acid in ethanol under reflux for 4 h afforded 2-((3-methyl-
1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4-yl)methylene)hydrazinecarboth-
ioamide 4a and 2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyra-
zol-4-yl)methylene)hydrazinecarbothioamide 4b. The synthesized thiosemicar-
bazones were confirmed by NMR and mass spectral data. In the 1H NMR spectrum
of compound 4a, a singlet at d 7.8 for H–C=N proton, and a singlet at d 11.26 ppm
for the –NH proton indicate the formation of thiosemicarbazones. The two singlets
at d 6.43 and 6.57 ppm correspond to –NH and –SH group protons that indicate en-
thiol tautomerism [48]. The mass spectrum showed a M - 1 peak at m/z 325 that
confirms the molecular weight of the compound.
The compound 2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-
4-yl)methylene)hydrazinyl)thiazol-4(5H)-one 5a and 2-(2-((5-(1H-benzo[d][1–3]-
triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-
one 5b were prepared by refluxing thiosemicarbazones 4a, 4b with ethyl
chloroacetate and anhydrous sodium acetate in ethanol for 6 h. The formation of
the thiazolidinone ring in compounds 5a and 5b was confirmed by the IR, NMR and
1
mass spectrometry. In the H NMR spectrum of the compound 5a, the presence of
singlet peak at d 3.75 equivalent to two protons indicates the C-5 protons (-CH2) of
the thiazolidinone ring.
The formation of thiazolidinone ring further confirmed by a strong band in the IR
spectrum at 1727 cm-1 corresponds to a carbonyl group. However, the mass
spectrum showed a molecular ion peak at m/z 367 (M ? 1) confirming the
molecular weight of the compound.
Finally, after confirmation of the compounds 5a, 5b, the target compounds
triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene thiazolidinones 6a–
n and 7a–l were achieved by the reaction of compounds 5a, 5b with various
substituted benzaldehydes in the presence of a catalytic amount of piperdine in
methanol at reflux conditions for 3–5 h via Knoevenagel condensation. The
formation of the arylidene derivatives 6a–n and 7a–l confirmed by the following
123

V. Pogaku et al.
1
spectral data, from the H NMR spectrum of compound 6d, the appearance of
singlet peaks in the region of d 7.72 indicating the olefinic proton attached to the
1
thiazolidinone ring. On the other hand, the disappearance of peaks at d 3.75 in H
NMR and at d 33.5 in 13C NMR confirms the involvement of -CH2 protons of
compound 5a in the reaction.
1
The H NMR spectra of compounds 6a–n and 7a–l show an arylidene proton
(olefinic proton), that was deshielded by an adjacent C=O group, showing the values
in the range of d 7.5–8.02 ppm. However, a literature survey was carried out to
determine the geometry of the C=C double bond in compounds 6 and 7. The
literature showed that the arylidene-4-thiazolidinones have Z-configuration at the
C=C double bond and the 1H NMR for the arylidene protons observed the chemical
shift values in the range of d 7.63–7.97 [36, 46, 49–52]. This supports
Z-configuration at the arylidene double bond in the compounds 6a–n and 7a–l.
Furthermore, we made attempts to develop the crystals for the compounds 6a–n
and 7a–l in various solvents and also solvent mixtures. As the solubility of the
compounds was very low, the compounds were not crystallized, instead they were
precipitated or agglomerated. But, a literature study was carried out on the crystal
structures of 5-arylidene-4-thiazolidinone moieties to establish the geometry of the
olefinic double bond of the compounds 6a–n and 7a–l. The analysis of the reported
[53–58] crystal structures of arylidene-4-thiazolidinones shows that the olefinic
double bond has Z-configuration, and it seems thermodynamically more stable than
the E-configuration.
During the synthesis of target compounds (6a–n and 7a–l), it was observed that
the yields of 1H-benzo[d][1–3]triazole substituted derivatives 7a–l were higher than
1H-1,2,4-triazole substituted derivatives 6a–n. The synthetic route for the
compounds 6a–n and 7a–l is illustrated in Scheme 1. The structures of the
derivatives and physical data of the compounds 6a–n and 7a–l are presented in
Fig. 1. These structures of all the synthesized compounds 6a–n and 7a–l were
characterized by spectral data (IR, NMR, and Mass). A generalized spectral
description is given below by taking the compound 6a, the appearance of a broad
peak in IR spectra at 3403 cm-1 corresponding to –NH, and sharp peaks at 1705,
1639 and 1614 cm-1 corresponding to C=O (carbonyl in thiazolidinone ring), C=N
and C=C (attached to thiazolidinone ring) stretching frequencies, respectively. The
1H NMR spectrum shows a signal at d 12.59 and d 9.10 corresponding to -NH
proton and carbazone (H-C=N) protons, respectively, the two singlets of the triazole
ring protons appeared at d 9.10 and 8.38, respectively. In the 13C NMR the presence
of peaks at d 153.74 (C=O) and d 149.65 (C=N) and molecular ion peak at m/z 455
(M ? 1) in the mass spectrum as well as elemental analyses data confirmed the
formation of the products. Spectral data for all the synthesized novel compounds
were in full agreement with the proposed structures.
Antibacterial activity
The triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene thiazolidinone
derivatives 6a–n and 7a–l were screened for their in vitro antibacterial activity
against both Gram positive and Gram negative bacteria like Bacillus subtilis,
123

Synthesis and biological evaluation of novel triazole…
Scheme 1 Synthetic route to compounds 6a–n and 7a–l. Reaction conditions: (a) 1H-1,2,4-triazole (2a)
or 1H-benzo[d][1,2,3]triazole (2b), DMF, K2CO3, reflux, 4 h. (b) Thiosemicarbazide, ethanol, a catalytic
amount of acetic acid, reflux, 4 h. (c) Ethyl chloroacetate, anhydrous NaOAc, ethanol, reflux, 6 h.
(d) Substituted benzaldehydes, a catalytic amount of piperdine, ethanol, reflux, 3–5 h
Bacillus megaterium and Escherichia coli, and Pseudomonas aeruginosa by using
the agar well plate method [59–61]. The experiments were carried out in triplicate;
the results were taken as a mean standard deviation (SD) and are summarized in
Table 1. The antibacterial screening data revealed that most of the tested
compounds exhibited good to moderate antibacterial activity against all the tested
bacterial stains. The compound 6b exhibited good activity against B. subtilis
(ZOI = 18 mm and MIC = 50 lg mL-1), B. meg (ZOI = 18 mm and
MIC = 50 lg mL-1), E. coli (ZOI = 18 mm and MIC = 50 lg mL-1), and
moderate
activity
against
P.
aeruginosa
(ZOI = 12 mm
and
MIC = 100 lg mL-1).
The compound 6i has shown good activity against B. subtilis (ZOI = 17 mm and
MIC = 50 lg mL-1), B. meg (ZOI = 16 mm and MIC = 50 lg mL-1), P.
aeruginosa (ZOI = 16 mm and MIC = 50 lg mL-1) and moderate activity against
E. coli (ZOI = 13 mm and MIC = 100 lg mL-1). The compound 6k has also
exhibited
good
activity
against
B.
subtilis
(ZOI = 18 mm
and
MIC = 50 lg mL-1), B. meg (ZOI = 16 mm and MIC = 50 lg mL-1); and
moderate activity against E. coli (ZOI = 13 mm and MIC = 100 lg mL-1) and
P. aeruginosa (ZOI = 15 mm and MIC = 50 lg mL-1) with respect to the
standard antibacterial drug streptomycin.
123

V. Pogaku et al.
Fig. 1 Structures, time and yields of the compounds 6a–n and 7a–l
123

Synthesis and biological evaluation of novel triazole…
Table 1 In vitro antibacterial activity of 6a–n and 7a–l
Zone of inhibition (in mm) and MIC in lg/mLa
Entry
Gram-positive
B. subtilis
Gram-negative
E. coli
B. megaterium
P. aeruginosa
ZOI MIC SD
ZOI MIC SD
ZOI MIC SD
ZOI MIC SD
6a
6b
6c
6d
6e
6f
10
18
16
14
0
200 0.36
50 0.21
100 0.43
100 0.54
NA
10
18
16
14
0
200 0.52
50 0.15
100 0.55
100 0.22
NA
14
18
14
12
0
100 0.26
50 0.30
100 0.41
200 0.53
NA
14
12
16
12
0
100 0.33
100 0.46
100 0.46
200 0.43
NA
10
6
200 0.50
200 0.70
200 0.42
50 0.22
100 0.45
50 0.15
200 0.54
NA
12
5
100 0.65
200 0.50
200 0.61
50 0.18
100 0.56
50 0.52
200 0.40
NA
10
13
12
13
10
13
9
200 0.56
150 0.97
200 0.55
100 0.32
200 0.41
100 0.70
200 0.37
NA
10
6
200 0.79
200 0.66
150 0.65
50 0.15
200 0.60
50 0.65
200 0.46
NA
6g
6h
6i
8
7
13
16
7
17
12
18
8
16
10
16
8
6j
6k
6l
15
7
6m
6n
7a
7b
7c
7d
7e
7f
0
0
0
0
8
200 0.41
200
6
200 0.36
200
8
200 0.15
100
5
200 0.25
200
10
18
14
10
08
10
10
18
12
0
10
16
12
08
08
10
08
16
12
0
13
16
14
08
11
08
08
18
10
0
10
16
10
08
10
08
07
15
09
0
50 0.30
100 0.55
200 0.95
200 0.51
200 0.80
200 0.30
50 0.21
100 0.51
NA
50 0.21
100 0.28
200 0.25
200 0.70
200 0.45
200 0.38
50 0.37
100 0.50
NA
50 0.41
200 0.25
200 0.56
200 0.61
200 0.35
200 0.46
50 0.30
200 0.76
NA
50 0.15
200 0.62
200 0.94
200 0.21
200 0.60
200 0.60
100 0.45
200 0.36
NA
7g
7h
7i
7j
7k
7l
0
NA
0
NA
0
NA
0
NA
12
100 0.65
08
200 0.45
10
200 0.40
10
200 0.51
12.5 0.40
Streptomycin 20
12.5 0.25 20
12.5 0.15 21
12.5 0.15 20
a
The experiments were carried out in triplicate
Values are mean SD of three replicates
NA not active
The MIC values were expressed in lg mL-1. The zone of inhibition (ZOI) values given in mm. Bacterial
stains: B. subtilis—Bacillus subtilis, B. meg—Bacillus megaterium, E. coli—Escherichia coli and P.
aeruginosa—Pseudomonas aeruginosa, The zone of inhibition (ZOI) values (in mm) for analogs (6a–n
and 7a–l) at 100 lg mL-1
The compound 7b has shown good antibacterial activity against B. subtilis
(ZOI = 18 mm and MIC = 50 lg mL-1), and moderate antibacterial activity
against B. meg, E. coli, P. aeruginosa with (ZOI = 16 mm and
123

V. Pogaku et al.
MIC = 50 lg mL-1). The compound 7h exhibited good activity against B. subtilis
and E. coli, with (ZOI = 18 mm and MIC = 50 lg mL-1) and moderate activity
on B. meg.
Cytotoxic activity
The in vitro cytotoxicity tests were screened for 6a–n derivatives against two human
cancer cell lines MCF-7 and Hela by using the MTT assay [62–64]. The graphs of
percentage of cell viability versus concentration of the compounds 6a–n were
plotted and the IC50 values were determined (Figs. 2, 3). The IC50 values obtained
for these derivatives are illustrated in Table 2.
All the synthesized compounds showed good to moderate anticancer activity with
IC50 values ranging from 5.4 to 120 lM. Among all the synthesized compounds, the
compound 6c showed excellent cytotoxic activity against the MCF-7 cell line,
having IC50 value 5.4 lM. Similarly, compounds 6l and 6f showed the good
cytotoxic activity with IC50 value 6.3 lM and 9.85 lM, respectively, against the
MCF-7 cell line. The remaining derivatives exhibited moderate activity against the
MCF-7 cell line.
Similarly, for the results obtained from MTT assay against the HeLa cell line, the
compounds 6b and 6c showed good cytotoxic activity with IC50 value 8.7 and
9.06 lM, respectively. 6f and 6l also exhibited good cytotoxic activity with IC50
value 10.7 and 10.1 lM, respectively. However, the compounds 6c and 6l showed
good activity against both cell lines MCF-7 and HeLa.
Structure–activity relationship (SAR) could be observed by examining the effect
of substituents like chlorine, bromine, methyl, nitro, hydroxyl, methoxy and N,N-
dimethylamine groups on the 5-benzylidene ring of these derivatives on the zone of
inhibition and cell growth inhibition. The derivatives with electron withdrawing
groups 3-NO2, 4-Br (6i, 6k) displayed good antibacterial activity against Bacillus
subtilis, Bacillus megaterium, Pseudomonas aeruginosa, and similarly 4-chloro (6c)
also shows moderate activity against both Gram positive and Gram negative
bacteria than other electron withdrawing groups derivatives. The derivatives with an
Fig. 2 Percentage of viability of compounds on MCF-7 lines
123

Synthesis and biological evaluation of novel triazole…
Fig. 3 Percentage of viability of compounds on HeLa lines
Table 2 Inhibition values
Entry
IC50 values in lM SD
(IC50) of the compounds (6a–n)
on two human cancer cell lines
MCF-7
HeLa
6a
30.40 0.39
15.86 0.32
5.40 0.15
19.50 0.35
49.22 0.45
9.85 0.25
20.40 0.35
22.80 0.40
24.42 0.38
44.27 0.43
120.00 0.51
6.30 0.20
14.48 0.41
49.88 0.38
4.5 0.10
24.00 0.34
8.70 0.26
9.06 0.21
17.20 0.41
63.85 0.46
10.76 0.30
21.10 0.32
38.16 0.46
34.42 0.41
20.89 0.36
85.00 0.50
10.10 0.25
19.47 .036
42.40 0.39
4.50 0.10
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
6m
6n
Doxorubicin
electron releasing group, i.e., 4-Me group (6b, 7b) displayed good antibacterial
activity compared to the derivatives with –OCH3, –N(CH3)2, –OH and the
unsubstituted parent compound 6a and 7a [65]. Further, 2-Cl (6e), 2-OH (6m, 7k)
and 4-N(CH3)3 (6l, 7j) groups on the phenyl ring of thiazolidinone are not
favourable for the antibacterial activity. In the case of bromo- and chloro-substituted
derivatives, the para-position is more favourable for the antibacterial activity than
the meta- and ortho- positions of the 5-benzylidine ring. In general, we conclude
from the results that the electron withdrawing groups like NO2, Br, Cl (6i, 6k, 6c) on
the 5-benzylidine ring favour the inhibition of the Gram-negative and Gram-positive
123

V. Pogaku et al.
bacteria more than electron releasing groups with the exception of compounds
having the methyl group [49].
In the case of cytotoxic activity, the derivatives with electron releasing –CH3, –
OCH3 and –N(CH3)2 groups at para-position exhibited potent activity rather than
the other electron releasing groups like hydroxy, dimethoxy and trimethoxy.
However, among all the electron withdrawing groups, 4-Cl (6c) derivative shows
excellent activity. From this, it can be concluded that the derivatives with electron
releasing or withdrawing groups present at para-position of 5-benzylidine ring are
more favorable for cytotoxic activity than the ortho- and meta-positions except 6n.
Experimental section
General considerations
All the chemicals and solvents were purchased from Aldrich/Merck and used
without further purifications. All melting points were determined by using Stuart
SMP30 melting point apparatus and were uncorrected. The progress of the reactions
was checked with TLC plates (E. Merck, Mumbai, India). The products were
purified by column chromatography on silica gel (100–200 mesh) eluted with a
solvent system of ethyl acetate–hexane (4:6). IR spectra were recorded on KBr disc
by a using Perkin-Elmer 100S spectrophotometer. 1H, 13C NMR spectra were
recorded on an Avance-III Bruker-400 MHz spectrometer using DMSO-d6 as
solvent and TMS as an internal standard, and chemical shifts are expressed as ppm.
Mass spectra were determined on a Jeol JMSD-300 spectrometer. The 5-chloro-3-
methyl-1-phenyl-1H-pyrazole-4-carbaldehyde 1 [66] and 3-methyl-1-phenyl-5-(1H-
1,2,4-triazol-1-yl)-1H-pyrazole-4-carbaldehyde 2 [22] were prepared by reported
procedures.
Experimental procedures
2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-yl)methylene)hydrazinecarbo
thioamide (4a, 4b) The compound 3a, 3b (0.01 mol) and thiosemicarbazide
(0.01 mol) were taken in 5 mL of ethanol. This reaction mixture was refluxed for
3 h. The progress of the reaction was monitored by TLC. After the completion of
the reaction, the reaction mixture was concentrated to half of the volume and
cooled. The filtered separated solid product was dried, and recrystallized from
methanol to give compound 4a, 4b.
2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4yl)methylene)hy-
drazinyl) thiazol-4(5H)-one (5a, 5b) A mixture of compound 4a, 4b (1.63 g,
0.005 mol), ethyl chloroacetate (0.53 g, 0.005 mol), anhydrous sodium acetate
(1.23 g, 0.015 mol), taken in absolute ethanol (10 mL) was refluxed for 6 h. The
progress of the reaction was observed by TLC. After the completion of the reaction,
it cooled to room temperature and the separated solid product was filtered, washed
with water, dried, and recrystallized from methanol to give compound 5a, 5b.
123

Synthesis and biological evaluation of novel triazole…
General procedure for synthesis of 5-arylidene-2-((E)-2-((3-methyl-1-phenyl-5-(1H-
1,2,4-triazol-1-yl)-1H-pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6a–
n and 7a–l) The compound 5a, 5b (0.18 g, 0.5 mmol), and substituted benzalde-
hydes (0.5 mmol) were taken in ethanol, and a catalytic amount of piperdine was
added to the mixture, then reflux of the reaction mixture for 3–5 h. The progress of
the reaction was monitored by TLC. After the completion of the reaction, it was
cooled to room temperature with filtering of the separated solid product, and washed
with ethanol. The products were purified by the column chromatography on silica
gel (100–200 mesh).
General procedures for biological evaluation
Antibacterial evaluation
The triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene thiazolidinone
derivatives 6a–n and 7a–l were screened for their in vitro antibacterial activity
against both gram positive and gram negative bacteria like Bacillus subtilis, Bacillus
megaterium and Escherichia coli, and Pseudomonas aeruginosa by using the well-
plate method in Mueller–Hinton Agar in triplicates; streptomycin was used as a
positive control [59–61]. The test solution was prepared by using dimethyl
sulphoxide (DMSO) as solvent. DMSO was used as a negative control whereas
streptomycin was used as a positive control. The standard pathogenic microbial
cultures were procured from the Microbial Type Culture Collection (MTCC),
Chandigarh, India. The sterile nutrient agar media was used for the incubation of
bacteria cultures and inoculated into the fresh nutrient broth of 10 mL, in order to
yield a bacterial suspension of about 10–100 colony forming units (CFU) per mL.
The inoculums were spread over the surface of the agar media by diluting the initial
microbial suspension ten times with distilled water. Then 30 mL of bacterial
suspension of 100 lg mL-1 concentrations was transferred into the 6 mm diameter
well made by the sterile cork borer and incubated for about 24 h at 37 °C.
Antibacterial evolutions were conducted in nutrient agar at 100 lg mL-1 concen-
trations of the synthesized compounds 6a–n and 7a–l with respect to the standard
drug streptomycin. The minimum inhibitory concentration (MIC) was measured in
lg mL-1 and zone of inhibition (ZOI) values were measured in mm [59, 61].
In vitro cytotoxicity assay
To study the in vitro cytotoxicity activity of the triazole derivatives 6a–n on cancer
cells MCF-7 and HeLa cells the MTT assay was adopted [62–64]. The cell cultures
were collected and maintained by the norms of the National Centre for Cell
Sciences Pune, India instructions. The cancer cells were maintained at 5% CO2 and
37 °C in the incubator. The triazole derivatives 6a–n were dissolved in DMSO in
the range of 10–100 lg. The cell was cultured in a 96-well tissue plate at the density
of 8 9 103 per well in the plate and kept in an incubator for growing at 37 °C in a
5% CO2 for 48 h. After attachment, the cells were treated with a different
123

V. Pogaku et al.
concentration of triazole derivatives and incubated for 24 h. MTT reagent was
added to each well, and the plate was incubated for 4 h. Then the medium was
discarded and DMSO was added to dissolve the purple product. The absorbance of
each well was measured at 570 nm using an Elisa reader. The IC50 values were
calculated from the plot of cell viability with different concentrations of triazole
derivatives.
Conclusions
A new series of triazole substituted pyrazolyl-methylenehydrazinyl-5-arylidene
thiazolidinone derivatives 6a–n and 7a–l were synthesized and evaluated for their
antibacterial activity against Gram-positive, Gram-negative bacteria, and cytotox-
icity activity. The compounds 6b, 6i, 6k and 7a, 7h exhibited good antibacterial
activity. The compounds 6c showed excellent cytotoxicity against MCF-7, whereas
6l and 6f exhibited good cytotoxic activity against MCF-7, and the compounds 6b
and 6c exhibited good cytotoxicity against HeLa cells. This synthetic protocol and
biological evaluation may help in the development of synthetic organic and
medicinal chemistry areas.
Spectral data for all the synthesized compounds
2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4-yl)methylene)hydrazine
1
carbothioamide (4a) White solid; mp: 242–244 °C; H NMR (DMSO-d6) d:2.53
(s, 3H, CH3), 6.43 (s, 1H), 6.57 (s, 1H), 7.25 (broad s, 2H, ArH), 7.36 (broad s, 3H,
ArH), 7.84 (s, 1H, HC=N), 8.18 (s, 1H, triazole), 8.20 (s, 1H, triazole), 11.26 (s, 1H,
NH-exchangeable); 13C NMR (400 MHz, DMSO-d6) d:178.03, 153.91, 148.41,
137.44, 134.63, 133.54, 129.88, 128.89, 123.58, 113.07, 14.43; ESI-MS: m/z 325
(M - 1); yield: 86%.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)hy-
1
drazinecarbothioamide (4b) White solid; mp: 238–240 °C; H NMR (DMSO-d6)
d:2.58 (s, 3H, CH3), 6.46 (s, 1H), 6.62 (s, 1H), 7.45 (broad s, 2H, ArH), 7.46 (broad
s, 3H, ArH), 7.49–7.66 (m, 4H, ArH), 7.90 (s, 1H, HC=N), 11.16 (s, 1H, NH-
exchangeable); ESI-MS: m/z 375 (M - 1); yield: 90%.
2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4-yl)methylene)hy-
drazinyl) thiazol-4(5H)-one (5a) White solid; mp: 250–252 °C; IR (KBr, cm-1
)
1
t
max: 3423 (NH), 1727 (C=O), 1630 (C=N); H NMR (DMSO-d6) d:2.60 (s, 3H,
CH3), 3.75 (s, 2H, CH2 protons), 7.21–7.36 (m, 5H, ArH), 8.15 (s, 1H, CH=N), 8.36
(s, 1H, triazole proton), 8.50 (s, 1H, triazole proton), 11.26 (s, 1H, NH-
exchangeable); 13C NMR (400 MHz, DMSO-d6) d:174.71, 153.80, 149.03,
148.18, 147.18, 137.50, 133.98, 129.84, 128.95, 123.74, 112.99, 33.49, 14.39;
ESI-MS: m/z 367 (M ? 1); yield: 72%.
2-(2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4 yl)methy-
lene) hydrazinyl)thiazol-4(5H)-one (5b) White solid; mp: 256–258 C; IR(KBr
°
123

Synthesis and biological evaluation of novel triazole…
cm-1) tmax: 3429 (NH), 1721 (C=O), 1586 (C=N); 1H NMR (DMSO-d6) d: 2.60 (s,
3H, CH3), 3.73 (s, 2H, CH2 protons), 7.14 (d, J = 8 Hz, 2H, ArH), 7.30–7.32 (m,
2H, ArH) 7.47–7.61 (m, 4H, ArH), 8.06 (s, 1H, CH=N), 8.19 (d, J = 8 Hz, 1H, CH,
ArH), 11.84 (s, 1H, NH-exchangeable); 13C NMR (100 MHz, DMSO) d: 174.58,
149.71, 147.01, 145.23, 137.42, 134.46, 132.18, 130.16, 129.88, 129.10, 125.56,
123.76, 120.40, 113.82, 110.71, 33.33, 14.27; ESI-MS: m/z 415 (M-); yield: 74%.
5-benzylidene-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4-
yl)methylene)hydrazinyl)thiazol-4(5H)-one (6a) Pale yellow solid; mp:
290–292 °C; IR (KBr, cm-1) tmax: 3403 (NH), 1705 (C=O), 1639 (C=C), 1614
(C=N);1H NMR (DMSO-d6) d: 2.55 (s, 3H, CH3), 7.24 (d, J = 6.8 Hz, 2H, ArH),
7.40–7.51 (m, 4H, ArH), 7.57–7.62 (m, 3H, ArH), 7.67 (d, J = 7.2 Hz, 2H, ArH),
8.28 (s, 1H, CH=N), 8.38 (s, 1H, triazoleproton), 9.10 (s, 1H, triazoleproton), 12.59
(s, 1H, NH exchangeable); 13C NMR (400 MHz, DMSO-d6) d: 153.74, 149.65,
148.53, 148.34, 137.49, 134.00, 133.60, 130.48, 130.31, 129.85, 129.72, 129.32,
129.06, 123.82, 112.76, 13.64; ESI-MS: m/z 455 (M ? 1); Anal. Calcd. For
C23H18N8OS: C, 60.78; H, 3.99; N, 24.65; Found: C, 60.56; H, 3.82; N, 24.37.
2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4-yl)methylene)hy-
drazinyl)-5-(4-methylbenzylidene)thiazol-4(5H)-one (6b) Pale yellow solid; mp:
310–312 °C; IR (KBr, cm-1) tmax: 3428 (NH), 1707 (C=O), 1640 (C=C), 1598
1
(C=N); H NMR (DMSO-d6) d: 2.55 (s, 3H, pyrazole CH3), 2.40 (s, 3H, Ar-CH3),
7.25 (Broad singlet, 2H, ArH), 7.42 (Broad singlet, 5H, ArH) 7.55 (Broad singlet,
3H, ArH),8.28 (s, 1H, CH=N), 8.38 (s,1H, triazole proton), 9.10 (s, 1H, triazole
proton), 12.53 (s, 1H, NH-exchangeable); ESI-MS: m/z 496 (M - 1); Anal. Calcd.
For C24H20N8OS: C, 61.52; H, 4.30; N, 23.92; Found: C, 61.24; H, 4.17; N, 24.19.
5-(4-chlorobenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyra-
zol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6c) Pale yellow solid; mp:
258–259 °C; IR (KBr, cm-1) tmax: 3403 (NH), 1708 (C=O), 1643 (C=C), 1607
1
(C=N); H NMR (DMSO-d6) d: 2.55 (s, 3H, CH3), 7.23–7.65 (m, 10H, ArH), 8.29
(s, 1H, CH=N), 8.42 (s, 1H, triazole proton), 9.10 (s, 1H, triazole proton), 12.64 (s,
1H, NH-exchangeable); ESI-MS: m/z 487 (M - 1); Anal. Calcd. For C23H17ClN8
OS: C, 56.50; H, 3.50; N, 22.92; Found: C, 56.72; H, 3.68; N, 22.68.
5-(3-chlorobenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyra-
zol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6d) Pale yellow solid; mp:
263–265 °C; IR (KBr, cm-1) tmax: 3415 (NH), 1705 (C=O), 1640 (C=C), 1599
(C=N);1H NMR (DMSO-d6) d: 2.55 (s, 3H, CH3), 7.23–7.77 (m, 9H, ArH), 7.72 (s,
1H, olefinic), 8.29 (s, 1H, CH=N), 8.37 (s, 1H, triazole proton), 9.08 (s, 1H, triazole
proton), 12.67 (s, 1H, NH-exchangeable); 13C NMR (400 MHz, DMSO-d6) d:
153.70, 149.68, 148.31, 136.21, 134.42, 131.50, 130.18, 129.86, 129.07, 128.37,
123.83, 112.88, 13.58; ESI-MS: m/z 487 (M - 1); Anal. Calcd. For C23H17ClN8OS:
C, 56.50; H, 3.50; N, 22.92; Found: C, 56.25; H, 3.62; N, 23.21.
5-(2-chlorobenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyra-
zol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one
(6e) Yellow
solid;
mp:
123

V. Pogaku et al.
272–274 °C; IR (KBr, cm-1) tmax: 3399 (NH), 1708 (C=O), 1640 (C=C), 1598
(C=N); 1H NMR (DMSO-d6) d: 2.53 (s, 3H, CH3), 7.18–7.53 (m, 9H, ArH), 8.02 (s,
1H, olefinic), 8.28 (s, 1H, CH=N), 8.35 (s, 1H, triazole proton), 9.07 (s, 1H, triazole
proton), 12.74 (s, 1H, NH-exchangeable); ESI-MS: m/z 487 (M - 1); Anal. Calcd.
For C23H17ClN8OS: C, 56.50; H, 3.50; N, 22.92; Found: C, 56.79; H, 3.64; N,
22.66.
5-(4-methoxybenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-
pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6f) Pale yellow solid; mp:
215–217 °C; IR (KBr, cm-1) tmax: 3396 (NH), 1706 (C=O), 1637 (C=C), 1589
1
(C=N); H NMR (DMSO-d6) d: 2.55 (s, 1H, CH3), 3.86 (s, 3H, OCH3), 7.16 (d,
J = 8.8 Hz, 2H, ArH), 7.24 (d, J = 9.6 Hz, 2H, ArH), 7.43 (m, 3H, ArH), 7.53 (S,
1H, olefinic), 7.63 (d, J = 8.8 Hz, 2H, ArH), 8.27 (s, 1H, CH=N), 8.40 (s, 1H,
triazole proton), 9.10 (s, 1H, triazole proton) 12.47 (s, 1H, NH-exchangeable); 13C
NMR (400 MHz, DMSO-d6) d: 153.78, 149.59, 148.34, 137.50, 132.45, 129.84,
129.38, 129.03, 126.51, 123.80, 115.27, 112.82, 55.95, 13.67; ESI-MS: m/z 483
(M - 1); Anal. Calcd. For C24H20N8O2S: C, 59.49; H, 4.16; N, 23.13; Found: C,
59.21; H, 4.05; N, 22.88.
5-(3,4-dimethoxybenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-
1H-pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6g) Pale yellow solid;
mp: 268–270 °C; IR (KBr, cm-1) tmax: 3416 (NH), 1703 (C=O), 1643 (C=C),
1593(C=N); 1H NMR (DMSO-d6) d: 2.56 (s, 3H, CH3), 3.85 (s, 3H, OCH3), 3.87 (s,
3H, OCH3), 7.17–7.27 (m, 5H, ArH), 7.38–7.45 (m, 3H, ArH), 7.54 (s, 1H, CH=C
proton), 8.22 (s, 1H, CH=N), 8.37 (s, 1H, triazole proton), 9.09 (s, 1H, triazole
proton), 12.50 (s, 1H, NH-exchangeable); 13C NMR (400 MHz, DMSO-d6) d:
153.81, 150.78, 149.40, 149.35, 148.32, 148.13, 137.48, 133.84, 129.85, 129.01,
126.70, 124.14, 123.76, 120.51, 113.51, 112.85, 112.45, 56.13, 55.95, 14.00; ESI-
MS: m/z 513 (M - 1); Anal. Calcd. For C25H22N8O3S: C, 58.35; H, 4.31 N, 21.78;
Found: C, 58.08; H, 4.19; N, 21.52.
2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4-yl)methylene)hydrazinyl)-5-
(3,4,5-trimethoxybenzylidene)thiazol-4(5H)-one (6h) Pale yellow solid; mp:
294–296 °C; IR (KBr, cm-1) tmax: 3423 (NH), 1709 (C=O), 1647 (C=C), 1603
(C=N); 1H NMR (DMSO-d6) d: 2.57 (s, 3H, CH3), 3.74 (s, 3H, OCH3), 3.90 (s, 6H,
two methoxy), 6.99 (s, 2H, Ar–H), 7.22 (d, J = 6.8 Hz, 2H, Ar–H), 7.36–7.48 (m,
3H, Ar–H), 7.56 (S, 1H, olefinic), 8.17 (s, 1H, CH=N), 8.32 (s, 1H, triazole proton),
9.08 (s, 1H, triazole proton) 12.60 (s, 1H, NH-exchangeable); ESI-MS: m/z 543
(M - 1); Anal. Calcd. For C26H24N8O4S: C, 57.34; H, 4.44; N, 20.58; Found: C,
57.66; H, 4.59; N, 20.84.
5-(4-bromobenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-
pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6i) Pale yellow solid;
mp: 272–274 °C; IR (KBr, cm-1) tmax: 3428 (NH), 1708 (C=O), 1643 (C=C),
1597 (C=N); 1H NMR (DMSO-d6) d: 2.55 (s, 3H, CH3), 7.23–7.81 (m, 10H, ArH),
8.29 (s, 1H, CH=N), 8.42 (s,1H, triazole proton), 9.10 (s, 1H, triazole proton), 12.63
(s, 1H, NH exchangeable); 13C NMR (400 MHz, DMSO-d6) d: 153.80, 148.33,
123

Synthesis and biological evaluation of novel triazole…
136.92, 132.66, 132.21, 129.85, 126.39, 123.79, 118.80, 108.70, 13.34; ESI-MS: m/
z 533 (M - 1); Anal. Calcd. For C23H17BrN8OS: C, 51.79; H, 3.21; N, 21.01;
Found: C, 51.52; H, 3.36; N, 21.29.
5-(3-bromobenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyra-
zol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one
(6j) Yellow
solid;
mp:
285–287 °C; IR (KBr, cm-1) tmax: 3399 (NH), 1708 (C=O), 1640 (C=C), 1598
1
(C=N); H NMR (DMSO-d6) d: 2.54 (s, 3H, CH3), 7.24–7.86 (m, 10H, ArH), 8.26
(s, 1H, CH=N), 8.38 (s,1H, triazole), 9.08 (s, 1H, triazole) 12.67 (s, 1H, NH-
exchangeable); 13C NMR (400 MHz, DMSO-d6) d:153.73, 149.64, 148.75, 148.27,
137.47, 136.45, 133.62, 133.04, 132.79, 131.69, 129.85, 129.05, 128.67, 127.63,
125.35, 123.82, 122.95, 112.69, 13.63; ESI-MS: m/z 533 (M - 1); Anal. Calcd. For
C23H17BrN8OS: C, 51.79; H, 3.21; N, 21.01; Found: C, 51.56; H, 3.38; N, 21.28.
3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-pyrazol-4-yl)methylene)hydrazinyl)-5-
(3-nitrobenzylidene)thiazol-4(5H)-one (6k) Red solid; mp: 280–282 °C; IR (KBr,
cm-1) tmax: 3421 (NH), 1594 (C=O), 1509 (C=C); 1H NMR (DMSO-d6) d: 2.56 (s,
3H, CH3), 7.21 (d, J = 7.2 Hz, 2H, ArH), 7.25 (s, 1H, olefinic), 7.35–7.44 (m, 3H,
ArH), 7.86 (d, J = 8.82 Hz, 2H, ArH), 8.06 (s, 1H, CH=N), 8.32 (d, J = 8.82 Hz,
2H, ArH), 8.39 (s, 1H, triazole proton), 9.09 (s, 1H, triazole proton); ESI-MS: m/z
498 (M - 1); Anal. Calcd. For C23H17N9O3S: C, 55.30; H, 3.43; N, 25.24; Found:
C, 55.06; H, 3.28; N, 25.02.
5-(4-(dimethylamino)benzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-
yl)-1H-pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6l) Yellow solid;
mp: 297–299 °C; IR (KBr, cm-1) tmax: 3422 (NH), 1638 (C=O), 1579 (C=C),
1522 (C=N); 1H NMR (DMSO-d6) d: 2.56 (s, 3H, CH3), 3.04 (s, 6H, 2CH3) 6.87 (d,
J = 8.4 Hz, 2H, ArH) 7.24 (d, J = 6.8 Hz, 2H, ArH), 7.42–7.51 (m, 6H, ArH), 8.23
(s, 1H, CH=N), 8.40 (s, 1H, triazole proton), 9.10 (s, 1H, triazole proton), 12.29 (s,
1H, NH-exchangeable); 13C NMR (400 MHz, DMSO-d6) d: 153.78, 151.53, 148.34,
137.51, 137.30, 132.46, 129.84, 129.32, 123.78, 123.55, 112.94, 112.50, 13.75; ESI-
MS: m/z 496 (M - 1); Anal. Calcd. For C25H23N9OS: C, 60.35; H, 4.66; N, 25.33;
Found: C, 60.57; H, 4.52; N, 25.12.
5-(2-hydroxybenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-
pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6m) Pale yellow solid;
mp: 306–308 °C; IR (KBr, cm-1) tmax: 3138 (NH), 1709 (C=O), 1645 (C=C),
1
1597 (C=N); H NMR (DMSO-d6) d: 2.55 (s, 3H, CH3), 6.98 (d, J = 8.0 Hz, 1H,
ArH), 7.07 (t, J = 7.2 Hz, 1H, ArH), 7.23 (d, J = 6.8 Hz, 2H, ArH), 7.32 (t,
J = 7.2 Hz,1H, ArH), 7.42 (d, J = 7.6 Hz, 3H, ArH), 7.49 (d, J = 7.6 Hz, 1H,
ArH), 7.8 (s, 1H, CH=C proton), 8.25 (s, 1H, CH=N) 8.35 (s,1H, triazole proton),
9.08 (s, 1H, triazole proton) 10.41 (s, 1H, OH proton) 12.50 (s, 1H, NH-
exchangeable); ESI-MS: m/z 469 (M - 1); Anal. Calcd. For C23H18N8O2S: C,
58.71; H, 3.86; N, 23.82; Found: C, 58.92; H, 3.98; N, 23.58.
5-(4-hydroxybenzylidene)-2-(2-((3-methyl-1-phenyl-5-(1H-1,2,4-triazol-1-yl)-1H-
pyrazol-4-yl)methylene)hydrazinyl)thiazol-4(5H)-one (6n) Pale yellow solid;
123

V. Pogaku et al.
mp, 276–278 °C; IR (KBr, cm-1) tmax: 3416 (NH), 1703 (C=O), 1643 (C=C), 1593
1
(C=N); H NMR (DMSO-d6) d: 2.55 (s, 3H, CH3), 6.97 (d, J = 8 Hz, 2H, ArH),
7.24 (d, J = 8 Hz, 2H, ArH), 7.42–7.53 (m, 6H, ArH), 8.26 (s, 1H, CH=N) 8.37 (s,
1H, triazole proton), 9.11 (s, 1H, CH, triazole proton), 10.25 (s, 1H, OH proton),
12.42 (s, 1H, NH-exchangeable); 13C NMR (400 MHz, DMSO-d6) d: 159.76,
153.72, 149.58, 148.37, 137.52, 133.51, 132.69, 129.85, 129.02, 125.01, 123.80,
116.68, 112.88, 13.66; ESI-MS: m/z 469 (M - 1); Anal. Calcd. For C23H18N8O2S:
C, 58.71; H, 3.86; N, 23.82; Found: C, 58.47; H, 3.94; N, 23.63.
2-(2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methy-
lene) hydrazinyl)-5-benzylidenethiazol-4(5H)-one (7a) Pale yellow solid; mp:
°
299–301 C; IR (KBr cm-1) tmax: 3419 (NH), 1718 (C=O), 1641 (C=C), 1508
(C=N); 1H NMR (DMSO-d6) d: 2.61 (s, 3H, CH3), 7.13–7.15 (m, 2H, ArH) 7.33 (d,
J = 6.8 Hz, 3H, ArH), 7.46 (d, J = 7.2 Hz, 3H, ArH), 7.53–7.62 (m, 4H, ArH),
7.69 (t, J = 7.6 Hz, 2H), 8.19 (d, J = 8.4 Hz, 1H, ArH), 8.35 (s, 1H, CH=N), 12.64
(s, 1H, NH exchangeable); 13C NMR (100 MHz, DMSO-d6) d: 167.89, 159.57,
150.47, 148.73, 145.36, 137.36, 134.31, 133.80, 131.54, 130.39, 130.08, 129.89,
129.74, 129.24, 125.53, 123.87, 123.25, 120.16, 113.45, 111.00, 13.37; ESI-MS: m/
z 503 (M-); Anal. Calcd. For C27H20N8OS: C, 64.27; H, 4.00; N, 22.21; Found: C,
64.04; H, 3.84; N, 22.02.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)
hydrazinyl)-5-(4-methylbenzylidene)thiazol-4(5H)-one (7b) Pale yellow solid; mp:
317–319 °C; IR (KBr, cm-1) tmax: 3407 (NH), 1709 (C=O), 1644 (C=C), 1599
(C=N); 1H NMR (DMSO-d6) d: 2.46 (s, 3H, tolyl CH3), 2.61 (s, 3H, pyrazole CH3),
7.13–7.16 (m, 2H, ArH) 7.33–7.37 (m, 5H, ArH), 7.43 (s, 1H, H-C=C), 7.46–7.51
(m, 3H, ArH), 7.57–7.64 (m, 2H, ArH), 8.21 (d, J = 8.0 Hz, 1H, ArH), 8.34 (s, 1H,
CH=N), 12.41 (s, 1H, NH exchangeable); 13C NMR (100 MHz, DMSO-d6) d:
153.21, 145.27, 131.02, 130.46, 130.36, 129.90, 129.24, 123.87, 111.01, 21.60,
13.36; ESI-MS: m/z 517 (M-); Anal. Calcd. For C28H22N8OS: C, 64.85; H, 4.28; N,
21.61; Found: C, 65.08; H, 4.45; N, 21.39.
2-(2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methy-
lene) hydrazinyl) 5-(4-chlorobenzylidene)thiazol-4(5H)-one (7c) Pale yellow solid;
°
mp: 266–268 C; IR (KBr cm-1) tmax: 3407 (NH), 1708 (C=O), 1643 (C=C), 1597
1
(C=N); H NMR (DMSO-d6) d: 2.62 (s, 3H, CH3), 7.15–7.75 (m, 13H, ArH) 8.21
(d, J = 8.0 Hz, 1H, ArH), 8.33 (s, 1H, CH=N), 12.52 (s, 1H, NH exchangeable);
ESI-MS: m/z 537 (M-); Anal. Calcd. For C27H19ClN8OS: C, 60.16; H, 3.55; N,
20.79; Found: C, 60.41; H, 3.72; N, 20.98.
2-(2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methy-
lene) hydrazinyl) 5-(3-chlorobenzylidene)thiazol-4(5H)-one (7d) Yellow solid; mp:
°
279–280 C; IR (KBr cm-1) tmax: 3404 (NH), 1708 (C=O), 1644 (C=C), 1502
(C=N);1H NMR (DMSO-d6) d: 2.62 (s, 3H, CH3), 7.13–7.16 (m, 2H, ArH)
7.31–7.36 (m, 4H, ArH), 7.43–7.47 (m, 2H, ArH), 7.56–7.62 (m, 3H, ArH), 7.67 (s,
1H, CH=C), 7.76 (t, J = 8.0 Hz, 1H), 8.17 (d, J = 8.0 Hz, 1H, ArH), 8.33 (s, 1H,
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Synthesis and biological evaluation of novel triazole…
CH=N), 12.56 (s, 1H, NH exchangeable); ESI-MS: m/z 537 (M-); Anal. Calcd. For
C27H19ClN8OS: C, 60.16; H, 3.55; N, 20.79; Found: C, 60.43; H, 3.74; N, 21.02.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)
hydrazinyl)-5-(4-methoxybenzylidene)thiazol-4(5H)-one (7e) Pale yellow solid;
mp: 260–262 °C; IR (KBr, cm-1) tmax: 3394 (NH), 1702 (C=O), 1631 (C=C),
1594 (C=N); 1H NMR (DMSO-d6) d: 2.62 (s, 3H, CH3), 3.92 (S, 3H, CH3),
7.14–7.16 (m, 2H, ArH), 7.23 (d, J = 8.8 Hz, 2H, ArH), 7.32–7.37 (m, 3H, ArH),
7.44–7.51 (m, 3H, ArH), 7.57–7.64 (m, 3H, ArH), 8.22 (d, J = 8.8 Hz, 1H, ArH),
8.33 (s, 1H, CH=N), 12.35 (s, 1H, NH exchangeable); 13C NMR (100 MHz,
DMSO-d6) d: 161.09, 132.40, 129.90, 129.22, 126.31, 125.56, 123.86, 120.22,
115.28, 113.50, 110.98, 56.02, 13.44; ESI-MS: m/z 533 (M-); Anal. Calcd. For
C28H22N8O2S: C, 62.91; H, 4.15; N, 20.96; Found: C, 62.65; H, 4.01; N, 20.73.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)
hydrazinyl)-5-(3,4-dimethoxybenzylidene)thiazol-4(5H)-one (7f) Pale yellow solid;
mp: 288–290 °C; IR (KBr, cm-1) tmax: 3240 (NH), 1713 (C=O), 1637 (C=C), 1592
(C=N); 1H NMR (DMSO-d6) d: 2.64 (s, 3H, CH3), 3.87 (S, 3H, CH3), 3.93 (s, 3H,
CH3), 7.05 (d, J = 8.0 Hz, 1H, ArH), 7.13–7.16 (m, 2H, ArH), 7.23–7.33 (m, 5H,
ArH), 7.46–7.51 (m, 2H, ArH), 7.55–7.62 (m, 2H, ArH), 8.20 (d, J = 8.0 Hz, 1H,
ArH), 8.23 (s, 1H, CH=N), 12.39 (s, 1H, NH exchangeable); ESI-MS: m/z 563
(M-) Anal. Calcd. For C29H24N8O3S: C, 61.69; H, 4.28; N, 19.85; Found: C, 61.45;
H, 4.11; N, 19.64.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)
hydrazinyl)-5-(3,4,5-trimethoxybenzylidene)thiazol-4(5H)-one (7g) Pale yellow
solid; mp: 306–308 °C; IR (KBr, cm-1) tmax: 3426 (NH), 1713 (C=O), 1650
(C=C), 1612 (C=N); 1H NMR (DMSO-d6) d: 2.66 (s, 3H, CH3), 3.77 (S, 3H, CH3),
3.92 (S, 6H, CH3), 6.95 (s, 2H), 7.15 (broad s, 2H), 7.32 (broad s, 3H), 7.44 (s, 1H),
7.52–7.61 (m, 3H), 8.12 (s, 2H), 12.52 (s, 1H, NH exchangeable); 13C NMR
(100 MHz, DMSO-d6) d 153.65, 145.23, 129.93, 125.67, 123.77, 120.26, 110.67,
107.87, 60.71, 56.51, 14.82;ESI-MS: m/z 595 (M ?)Anal. Calcd. For C30H26N8
O4S: C, 60.59; H, 4.41; N, 18.84; Found: C, 60.85; H, 4.57; N, 18.62.
2-(2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methy-
lene) hydrazinyl)-5-(4-bromobenzylidene)thiazol-4(5H)-one (7h) Pale yellow
solid; mp: 283–285 °C; IR (KBr, cm-1) tmax: 3404 (NH), 1709 (C=O), 1646
(C=C);1H NMR (DMSO-d6) d: 2.62 (s, 3H, CH3), 7.15 (broad d, 2H, ArH) 7.32
(broad s, 3H, ArH), 7.41–7.50 (m, 4H, ArH), 7.57–7.64 (m, 2H, ArH), 7.87 (d,
J = 8.0 Hz, 2H, ArH), 8.21 (d, J = 8.0 Hz, 1H, ArH), 8.33 (s, 1H, CH=N), 12.51
(s, 1H, NH exchangeable); ESI-MS: m/z 581 (M-); Anal. Calcd. For C27H19BrN8
OS: C, 55.58; H, 3.28; N, 19.20; Found: C, 55.31; H, 3.11; N, 19.43.
2-(2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methy-
lene) hydrazinyl)-5-(3-bromobenzylidene)thiazol-4(5H)-one (7i) Yellow solid; mp:
°
323–325 C; IR (KBr cm-1) tmax: 3398 (NH), 1705 (C=O), 1639 (C=C), 1503
1
(C=N); H NMR (DMSO-d6) d: 2.62 (s, 3H, CH3), 7.15 (broad d, 2H, ArH) 7.32
123

V. Pogaku et al.
(broad s, 3H, ArH), 7.38 (d, J = 8.0 Hz, 1H, ArH) 7.43–7.50 (m, 2H, ArH),
7.56–7.62 (m, 2H, ArH), 7.68–7.72 (m, 2H, ArH) 7.82 (s, 1H, CH=C) 8.17 (d,
J = 8.0 Hz, 1H, ArH), 8.32 (s, 1H, CH=N), 12.55 (s, 1H, NH exchangeable); ESI-
MS: m/z 581 (M-); Anal. Calcd. For C27H19BrN8OS: C, 55.58; H, 3.28; N, 19.20;
Found: C, 55.34; H, 3.13; N, 19.39.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)
hydrazinyl)-5-(4-(dimethylamino)benzylidene)thiazol-4(5H)-one (7j) Yellow solid;
mp: 302–304 °C; IR (KBr, cm-1) tmax: 3377 (NH), 1698 (C=O), 1634 (C=C), 1597
1
(C=N); H NMR (DMSO-d6) d: 2.63 (s, 3H, CH3), 3.10 (S, 6H, 2CH3), 6.93 (d,
J = 8.0 Hz, 2H, ArH) 7.13–7.17 (m, 2H, ArH), 7.32–7.35 (m, 5H, ArH), 7.36 (s,
1H, CH=C), 7.49 (t, J = 8.0 Hz, 1H, ArH), 7.56–7.64 (m, 2H, ArH), 8.23 (d,
J = 8.4 Hz, 1H, ArH), 8.29 (s, 1H, CH=N), 12.16 (s, 1H, NH exchangeable);13C
NMR (100 MHz, DMSO-d6) d: 151.58, 150.23, 145.41, 137.41, 134.26, 132.44,
131.57, 130.48, 130.07, 129.88, 129.16, 125.50, 123.84, 120.78, 120.25, 115.81,
113.62, 112.44, 110.94, 13.55; ESI-MS: m/z 546 (M-); Anal. Calcd. For
C29H25N9OS: C, 63.60; H, 4.60; N, 23.02; Found: C, 63.88; H, 4.78; N, 23.25.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)
hydrazinyl)-5-(2-hydroxybenzylidene)thiazol-4(5H)-one (7k) Pale yellow solid; mp:
320–322 °C; IR (KBr, cm-1) tmax: 3141 (NH), 1708 (C=O), 1638 (C=C), 1579
(C=N);1H NMR (DMSO-d6) d: 2.61 (s, 3H, CH3), 6.99 (d, J = 8.0 Hz, 1H, ArH)
7.15 (broad d, 3H, ArH), 7.23 (t, J = 7.2 Hz, 1H, ArH), 7.32–7.40 (m, 4H, ArH),
7.46 (t, J = 7.2 Hz, 1H, ArH), 7.57–7.63 (m, 2H, ArH), 7.78 (s, 1H, CH=C), 8.16
(d, J = 8.0 Hz, 1H, ArH), 8.32 (s, 1H, CH=N), 10.38 (s, 1H, OH), 12.35 (s, 1H, NH
exchangeable); 13C NMR (100 MHz, DMSO-d6) d: 167.92, 157.53, 150.38, 148.55,
145.34, 137.39, 134.32, 132.16, 131.57, 130.05, 129.89, 129.19, 128.68, 125.48,
124.19, 123.85, 121.39, 120.79, 120.18, 116.46, 113.50, 110.99, 13.45; ESI-MS: m/
z 519 (M-); Anal. Calcd. For C27H20N8O2S: C, 62.30; H, 3.87; N, 21.53; Found: C,
62.04; H, 3.70; N, 21.36.
2-((5-(1H-benzo[d][1,2,3]triazol-1-yl)-3-methyl-1-phenyl-1H-pyrazol-4-yl)methylene)
hydrazinyl)-5-(4-hydroxybenzylidene)thiazol-4(5H)-one (7l) Pale yellow solid; mp:
313–315 °C; IR (KBr, cm-1) tmax: 3069 (NH), 1709 (C=O), 1643 (C=C), 1593
1
(C=N); H NMR (DMSO-d6) d: 2.62 (s, 3H, CH3), 7.05 (d, J = 8.8 Hz, 2H, ArH)
7.14–7.16 (m, 3H, ArH), 7.31–7.35 (m, 5H, ArH), 7.38 (s, 1H, CH=C), 7.49 (t,
J = 8.0 Hz, 1H, ArH), 7.57–7.64 (m, 2H, ArH), 8.17 (d, J = 8.4 Hz, 1H, ArH),
8.33 (s, 1H, CH=N), 10.31 (s, 1H, OH), 12.30 (s, 1H, NH exchangeable);13C NMR
(100 MHz, DMSO-d6) d: 168.11, 159.87, 150.39, 148.26, 145.36, 137.39, 134.31,
132.64, 131.46, 130.06, 129.90, 129.77, 129.21, 125.50, 124.81, 123.88, 120.04,
118.89, 116.66, 113.54, 111.04, 13.40; ESI-MS: m/z 519 (M-); Anal. Calcd. For
C27H20N8O2S: C, 62.30; H, 3.87; N, 21.53; Found: C, 62.08; H, 3.72; N, 21.75.
Acknowledgements S. B. thanks the MHRD, India for the Research Seed Grant (RSG). V. P. thanks the
MHRD, India for the fellowship. S.B. and V.P. thank Director, National Institute of Technology
Warangal, India for providing research facilities.
123

Synthesis and biological evaluation of novel triazole…
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