3-(1,3,4-Thiadiazole-2-yl)quinoline derivatives: Synthesis, characterization and anti-microbial activity

Article abstract of DOI:10.1016/j.ejmech.2011.04.013

A new series of thiadiazoles and intermediate thiosemicarbazones were synthesized from the chloroquinone molecule, with an aim to explore their effect on in vitro growth of microorganisms causing microbial infection. The chemical structures of the compound were elucidated by elemental analysis, FTIR, 1H and 13C NMR and ESI-MS spectral data. In vitro anti-microbial activity was performed against Staphylococcusaureus, Streptococcuspyogenes, Salmonellatyphimurium, and Escherichiacoli. The MIC was detected using the double dilution method. The results were compared by calculating percent inhibit area/μg of the compounds and the standard "amoxicillin". The selected compounds were tested for cytotoxic results using MTT assay H9c2 cardiac myoblasts cell line and the results showed that all the compounds offered remarkable >80% viability to a concentration of 200 μg/mL.

Full text of DOI:10.1016/j.ejmech.2011.04.013

European Journal of Medicinal Chemistry 46 (2011) 3158e3166
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Preliminary communication
3-(1,3,4-Thiadiazole-2-yl)quinoline derivatives: Synthesis, characterization
and anti-microbial activity
,
b,
Abdul R. Bhat a, Tazeem b, Amir Azam c, Inho Choi a , Fareeda Athar
**
*
a School of Biotechnology, Yeungnum University, Gyeongsan 712-749, South Korea
b Center for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, Jamia Nagar, New Delhi 110025, India
c Department of Chemistry, Jamia Millia Islamia, New Delhi 110025, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A new series of thiadiazoles and intermediate thiosemicarbazones were synthesized from the chlor-
oquinone molecule, with an aim to explore their effect on in vitro growth of microorganisms causing
microbial infection. The chemical structures of the compound were elucidated by elemental analysis,
FTIR, 1H and 13C NMR and ESI-MS spectral data. In vitro anti-microbial activity was performed against
Staphylococcus aureus, Streptococcus pyogenes, Salmonella typhimurium, and Escherichia coli. The MIC was
detected using the double dilution method. The results were compared by calculating percent inhibit
Received 26 January 2011
Received in revised form
31 March 2011
Accepted 4 April 2011
Available online 13 April 2011
area/
cytotoxic results using MTT assay H9c2 cardiac myoblasts cell line and the results showed that all the
compounds offered remarkable >80% viability to a concentration of 200 g/mL.
mg of the compounds and the standard “amoxicillin”. The selected compounds were tested for
Keywords:
Chloroquinoline-3-carboxaldehyde
Thiosemicarbazone
Thiadiazole
m
Ó 2011 Elsevier Masson SAS. All rights reserved.
Anti-microbial activity
1. Introduction
Various successful attempts have been made to produce the potent
therapeutic agents such as torvafloxin [17], moxifloxacin [18],
gemifloxacin [19] and gatifloxacin [20] etc. from quinoline. The
incidence of gram-positive bacterial resistance to these antibacte-
rial agents is growing and will be of major concern in near future
[21,22]. Therefore, there is an urgent need for novel chemical
entities that are particularly effective against gram-positive path-
ogens. In continuation of our efforts in developing heterocycles of
biological interest [23e25] and considering the significant role of
thiadiazoles [26e28] and quinolines in biological applications, we
wish to report here the synthesis of a new series of quinoline-
thiadiazole derivatives and their anti-microbial activities.
The innovative research for antibiotics has improved mankind’s
health status by confining life threatening infections. However, the
emergence and spread of bacterial resistance represent a severe
global problem [1]. Multidrug-resistant gram-positive pathogens,
such as methicillin-resistant Staphylococci aureus (MRSA), penicillin
resistant Streptococcus pneumoniae (PRSP), and vancomycin-
resistant enterrococci (VRE), compounded problems in the thera-
peutics [2e4]. Heterocyclic thiadiazoles are widely exposed to
therapeutic world, because of their known anti-HIV [5], anti-
inflammatory [6e8], anti-cancer [9], anti-tuberculosis [10,11],
anti-convulsant [12] and anti-hypertensive [13,14] activities.
Nitroheteroaryl-1,3,4-thiadiazole derivatives have shown impres-
sive anti-microbial and antiparasitic activity particularly against
Trypanosomatid protozoa [15]. Thiadiazole derivative, 2-(4-chlor-
opheny1amino)-5-(4-aminophenyl)-1,3,4-thiadiazole showed 57%
inhibition against Mycobacterium tuberculosis [16].
2. Result and discussion
2.1. Chemistry
The synthetic route of compounds (1Be24B) is shown in Fig. 1.
Quinolines have been of great interest as antibacterial and
antiviral compounds for several decades, starting from the intro-
duction of nalidixic acid in 1962 for the urinary tract infections.
Compounds were obtained from
a
starting material of
2,8-substituted-quinoline-3-carbaldehyde. The synthesized thio-
semicarbazones (Fig. 1, Table 1) of substituted-quinoline were
cyclized in presence of acetic anhydride to get the final thiadiazole
compounds. All the reactions were monitored by TLC (aluminum
sheet, silica gel 60 F254, Merck). The precursor, thiosemicarbazone
(1Ae24A) and the final compounds (1Be24B) were purified by
column chromatography using silica gel (pore size 60 Å, 200e400
* Corresponding author. Tel.: þ91 11 26981717x4492; fax: þ91 11 26980164.
** Corresponding author. Tel.: þ82 53 810 3024; fax: þ82 53 810 4769.
E-mail addresses: inhochoi@ynu.ac.kr (I. Choi), fareeda.cirbs@jmi.ac.in, fareeda_
athar@yahoo.co.in (F. Athar).
0223-5234/$ e see front matter Ó 2011 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2011.04.013

A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
3159
S
S
ClCH2COONa
HCl
KOH + S=C=S + H2NR
HOOC
O
S
NHR
NaOOC
S
NHR
NHR
NH2NH2.H20
O
N
S
N
Cl
S
N
O
N
X
H2N
N
S
N
H
NHR
N
R
H
N
Cl
1A-24A
N
Cl
X
X
1B-24B
X= H, Cl, CH3, R= Different substituents given in Table 1
Fig. 1. Schematic Representation of the Synthetic route adopted for the synthesis N-4-acetyl-5-(2,8-substitutedquinolin-3-yl)-4,5-dihydro-1,3,4-thiadiazole-2-yl)acetamide
derivatives.
mesh) and eluted with proper solvent (CHCl3:CH3OH) system. The
elutants were concentrated in vacuo and crystallized at 4 ꢀC using
chloroform hexane mixture to obtain a pure compound. The purity
of the compounds was decided on the basis of melting points and
elemental data analysis.
ion signal at M þ 1, however in some cases [M þ Na] signal was
found. In 1H NMR, the characterize signal around 7.28e6.72 ppm
(CH]N) shows the condensation of substituted-quinoline-3-car-
baldehyde and the different thiosemicarbazides. Other signals were
found in accordance to the established structures. When the thio-
semicarbazides were condensed in excess of acetic anhydride, the
corresponding singlet shows an up-field shift from 7.28e6.72 ppm
to 5.69e4.93 ppm range. The proton shift showed the change in
status of carbon from unsaturated to saturated one. The cyclization
was confirmed by the disappearance of NH signals and the
appearance of a new singlet signal due to the six proton (eCH3)
around 2.13e3.10 ppm. To confirm the results, some molecules
were analyzed by 13C NMR. The 13C NMR spectra of thiadiazoles
showed some prominent signals, such as signals in 50e60 ppm,
and around 22e26 ppm range, which were absent in corresponding
thiosemicarbazones. These signals represent the change in carbon
environment and were attributed to CH(NS) carbon and alkyl
carbon, as the inclusion of two methyl groups was expected in the
process of cyclization.
All the compounds were characterized using the 1H NMR, IR and
mass spectrophotometry. Some of the compounds were analyzed
by using 13C NMR. Almost all the compounds showed the molecular
Table 1
Different substituent’s “X” and “R” as indicated in Fig. 1.
Compound
X
R
1A, 1B
Cl
H
9A, 9B
H
17A, 17B
CH3
3A, 3B
Cl
11A, 11B
19A, 19B
H
CH3
The structures were further confirmed by the appearance and
disappearance of
a
band around 3400 cmꢁ1 in the thio-
5A, 5B
Cl
semicarbazones and thiadiazoles, respectively. This suggests that
the NH group had reacted in the process of cyclization. Besides the
other characteristic signals for aromatic rings and C]N, a strong
signal around 2930e2965 cmꢁ1 indicates the presence of aliphatic
methyl groups. The signal appeared as a new signal in the
compound (1Be24B). The existence of a strong band in the region
1102e1133 cmꢁ1 due to C]S and the absence of any band in the
region 2500e2600 cmꢁ1 due to CeSH suggest that all thio-
semicarbazones retains the thione nature. In the compounds
(1Be24B), two sharp carbonyl peaks were observed, backing
inclusion of two acetyl groups to the newly cyclized thiadiazoles at
the two different positions. Carbonyl group stretching frequency
corresponded to esteric carbonyl stretching and amidic carbonyl
13A, 13B
21A, 21B
H
CH3
7A, 7B
15A, 15B
23A, 23B
Cl
H
CH3
2A, 2B
10A, 10B
18A, 18B
Cl
H
CH3
stretching were observed at 1640e1678 cmꢁ1
.
4A, 4B
12A, 12B
20A, 20B
Cl
H
CH3
2.2. Pharmacology
2.2.1. In vitro evaluation of antibacterial activity against gram-
positive and gram-negative bacteria
6A, 6B
14A, 14B
22A, 22B
Cl
H
CH3
F
F
All the synthesized thiosemicarbazones (1Ae24A) and their
thiadiazole derivatives (1Be24B) were screened for their anti-
microbial activity using the gram-positive and gram-negative
bacteria. Four different cultures, two each of gram-negative
(Escherichia coli and Salmonella typhimurium) and gram-positive
(Staphylococcus aureus and Staphylococcus pyogenes) were treated
8A, 8B
16A, 16B
24A, 24B
Cl
H
CH3
NO2

3160
A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
Table 2
Table 4
Antibacterial activity of 1-((2,8-dichloroquinolin-3-yl)methylene)thiosemicarb-
azide derivatives (1Ae24A), positive control (Amoxicillin), and negative control
(DMSO) measured by the Halo Zone Test (Unit, mm).
Minimum Inhibition Concentration
methylene)thiosemicarbazide derivatives (1Ae24A) and positive control.
(mg/mL) of 1-((2,8-dichloroquinolin-3-yl)
Compound
Effect of Compounds on Microorganism
Compounds
Effect of Compounds on Microorganism
S. aureus
S. pyogenes
S. typhimurium
E. coli
S. aureus
S. pyogenes
S. typhimurium
E. coli
1A
2A
3A
4A
5A
6A
7A
8A
64
128
64
256
32
64
64
32
128
64
256
256
256
32
32
64
64
256
128
256
64
32
32
64
64
128
256
128
64
64
32
128
128
256
256
32
64
32
64
128
128
128
128
64
128
256
256
128
64
1A
2A
3A
4A
5A
6A
7A
8A
21.13 ꢂ 0.56
17.68 ꢂ 0.73
17.23 ꢂ 0.33
e
19.58 ꢂ 0.79
13.15 ꢂ 0.76
13.73 ꢂ 0.51
17.54 ꢂ 0.36
10.26 ꢂ 0.78
15.92 ꢂ 0.73
15.76 ꢂ 0.47
15.26 ꢂ 0.62
16.97 ꢂ 0.45
21.07 ꢂ 0.64
12.38 ꢂ 0.50
13.67 ꢂ 0.74
16.48 ꢂ 0.75
14.07 ꢂ 0.93
17.43 ꢂ 0.59
12.25 ꢂ 1.11
17.10 ꢂ 0.86
18.71 ꢂ 0.73
11.08 ꢂ 0.50
14.45 ꢂ 0.61
18.81 ꢂ 0.41
10.13 ꢂ 0.36
17.65 ꢂ 1.34
13.27 ꢂ 0.89
19.57 ꢂ 0.58
e
16.66 ꢂ 0.53
15.17 ꢂ 0.87
15.72 ꢂ 0.53
17.54 ꢂ 0.34
10.53 ꢂ 1.04
14.92 ꢂ 0.71
17.55 ꢂ 0.59
15.99 ꢂ 0.68
12.42 ꢂ 0.70
22.58 ꢂ 0.54
15.93 ꢂ 0.60
19.73 ꢂ 0.96
14.73 ꢂ 0.49
11.56 ꢂ 0.57
18.02 ꢂ 0.50
e
15.80 ꢂ 0.58
17.18 ꢂ 0.71
15.23 ꢂ 0.94
17.62 ꢂ 0.39
13.47 ꢂ 0.89
13.75 ꢂ 0.15
15.88 ꢂ 0.18
e
18.90 ꢂ 0.40
12.15 ꢂ 0.18
15.01 ꢂ 0.18
19.70 ꢂ 0.60
11.67 ꢂ 0.59
19.52 ꢂ 0.81
12.37 ꢂ 0.50
09.90 ꢂ 0.65
e
64
64
256
256
128
128
256
32
64
64
32
256
256
128
256
64
64
64
32
32
9A
9A
17.76 ꢂ 0.38
21.12 ꢂ 0.57
17.68 ꢂ 0.78
13.23 ꢂ 0.34
18.84 ꢂ 0.41
10.13 ꢂ 0.26
16.65 ꢂ 1.19
13.27 ꢂ 0.79
e
10A
11A
12A
13A
14A
15A
16A
17A
18A
19A
20A
21A
22A
23A
24A
Amoxicillin
10A
11A
12A
13A
14A
15A
16A
17A
18A
19A
20A
21A
22A
23A
24A
þive C
ꢁive C
32
64
16.75 ꢂ 0.37
17.93 ꢂ 0.38
13.46 ꢂ 0.42
12.76 ꢂ 0.51
20.04 ꢂ 0.42
12.75 ꢂ 0.14
15.78 ꢂ 0.16
14.80 ꢂ 0.41
e
64
256
256
256
256
128
32
64
32
64
32
64
128
256
256
128
64
32
64
32
32
18.02 ꢂ 0.88
17.44 ꢂ 0.73
15.66 ꢂ 0.56
22.51 ꢂ 0.53
13.94 ꢂ 0.62
15.88 ꢂ 0.58
17.18 ꢂ 0.91
14.23 ꢂ 0.94
22.37 ꢂ 0.27
e
256
128
256
32
256
32
16.97 ꢂ 0.65
21.27 ꢂ 0.64
16.61 ꢂ 0.53
15.17 ꢂ 0.47
14.72 ꢂ 0.53
18.67 ꢂ 0.74
17.48 ꢂ 0.75
22.05 ꢂ 0.14
e
19.24 ꢂ 0.95
17.31 ꢂ 0.97
20.85 ꢂ 0.64
e
64
32
control, the DMSO poured disk. The MIC was evaluated by the
macro-dilution test using standard inoculums of 10ꢁ5 CFL mLꢁ1
Serial dilutions of the test compounds, previously dissolved in
.
with synthesized compounds using disk diffusion method [29]. The
minimum inhibitory concentration (MIC), concentration at which
the halo zone formation starts, was determined by using different
dilutions of the treatment compounds. The results were compared
with positive control, the standard drug amoxicillin and negative
dimethyl sulfoxide (DMSO), wereprepared to final concentrations of
512, 256, 128, 64, 32, 16, 8, 4, 2 and 1 m
g mLꢁ1. The dilutions were
added to 24 h old inoculums. The susceptibility of the bacteria to the
test compounds was determined by the formation of an inhibitory
zone after 18 h of incubation at 37 ꢀC. Tables 2 and 3 report the
Table 3
Antibacterial activity of 1,3,4-thiadiazole derivatives (1Be24B), positive control
(Amoxicillin), and negative control (DMSO) measured by the Halo Zone Test (Unit,
mm).
Table 5
Minimum Inhibition Concentration
(mg/mL) of 1,3,4-thiadiazole derivatives
(1Be24B) and positive control.
Compounds
Effect of Compounds on Microorganism
Compound
Minimum Inhibition Concentration (mg/mL)
S. aureus
S. pyogenes
S. typhimurium
E. coli
S. aureus
S. pyogenes
S. typhimurium
E. coli
1B
2B
3B
4B
5B
6B
7B
8B
18.13 ꢂ 0.57
18.03 ꢂ 0.50
17.39 ꢂ 0.62
17.62 ꢂ 0.87
19.76 ꢂ 0.64
12.99 ꢂ 0.85
15.67 ꢂ 0.72
19.81 ꢂ 0.52
15.67 ꢂ 0.60
20.63 ꢂ 0.67
14.03 ꢂ 0.82
14.57 ꢂ 0.88
15.66 ꢂ 0.33
17.41 ꢂ 0.95
18.27 ꢂ 0.90
14.29 ꢂ 0.66
13.66 ꢂ 0.68
19.00 ꢂ 0.69
15.42 ꢂ 0.43
18.77 ꢂ 0.91
15.80 ꢂ 0.73
21.99 ꢂ 0.57
18.92 ꢂ 0.53
16.54 ꢂ 0.72
21.52 ꢂ 0.53
e
19.91 ꢂ 0.72
14.15 ꢂ 0.27
17.73 ꢂ 0.52
17.94 ꢂ 0.41
13.84 ꢂ 0.74
13.92 ꢂ 0.38
17.09 ꢂ 0.92
16.32 ꢂ 0.32
16.97 ꢂ 0.59
19.93 ꢂ 0.62
17.38 ꢂ 0.50
14.33 ꢂ 0.36
17.14 ꢂ 0.93
14.74 ꢂ 0.51
17.00 ꢂ 0.54
14.92 ꢂ 0.49
16.77 ꢂ 1.00
17.71 ꢂ 0.44
13.09 ꢂ 0.72
15.45 ꢂ 0.72
19.14 ꢂ 0.82
16.13 ꢂ 0.71
18.32 ꢂ 1.10
15.27 ꢂ 1.12
20.90 ꢂ 0.60
e
17.27 ꢂ 0.42
15.17 ꢂ 0.87
16.72 ꢂ 0.61
18.54 ꢂ 0.73
16.86 ꢂ 0.51
17.59 ꢂ 0.67
16.88 ꢂ 0.49
17.25 ꢂ 0.89
13.42 ꢂ 0.63
18.60 ꢂ 0.52
14.59 ꢂ 0.55
18.82 ꢂ 0.64
16.06 ꢂ 0.72
16.67 ꢂ 0.50
16.89 ꢂ 0.50
17.03 ꢂ 0.62
18.08 ꢂ 0.82
15.13 ꢂ 0.98
12.01 ꢂ 0.78
17.84 ꢂ 0.66
16.11 ꢂ 0.84
16.60 ꢂ 0.51
17.27 ꢂ 0.16
19.16 ꢂ 0.23
21.82 ꢂ 0.42
e
16.72 ꢂ 0.52
18.18 ꢂ 0.95
15.23 ꢂ 1.13
16.36 ꢂ 0.90
15.59 ꢂ 0.64
16.45 ꢂ 0.62
18.11 ꢂ 0.58
12.56 ꢂ 0.61
17.76 ꢂ 0.38
19.79 ꢂ 0.62
15.63 ꢂ 0.39
17.86 ꢂ 0.72
15.84 ꢂ 0.65
20.13 ꢂ 0.36
16.40 ꢂ 1.07
14.34 ꢂ 0.91
18.34 ꢂ 0.56
16.08 ꢂ 0.55
16.75 ꢂ 0.69
19.00 ꢂ 0.85
17.90 ꢂ 0.67
15.36 ꢂ 0.52
18.16 ꢂ 0.42
13.03 ꢂ 0.30
23.39 ꢂ 0.68
e
1B
2B
3B
4B
5B
6B
7B
8B
64
64
64
64
16
64
64
32
64
32
256
128
32
16
32
64
64
128
128
256
64
64
64
64
32
64
64
128
128
64
64
64
64
64
64
64
128
256
64
32
64
64
32
64
32
64
64
32
32
64
64
256
128
256
128
32
32
64
64
32
128
128
64
32
64
64
32
64
64
128
256
32
32
32
9B
10B
11B
12B
13B
14B
15B
16B
17B
18B
19B
20B
21B
22B
23B
24B
þive C
ꢁive C
9B
10B
11B
12B
13B
14B
15B
16B
17B
18B
19B
20B
21B
22B
23B
24B
Amoxicillin
32
256
128
16
64
16
64
64
64
128
256
32
16
64
64
256
128
256
256
64
64
64
64
32
64
32

A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
3161
Fig. 2. The graph showing comparative percent area inhibition per
m
g of the compounds and the Amoxicillin, in case of gram-positive bacteria, S. aureus and S. pyogenes.
inhibition zones (mm) of each compound and the minimum
concentration at which the inhibition zones appeared are presented
in Tables 4 and 5.
a thorough investigation like enzyme assay and other biological
assays would be helpful in designing more potent antibacterial
agents for therapeutic use.
The zone of inhibition was measured at the minimum inhibitory
concentration. To compare the anti-microbial effect of the
compounds with that of the ampicillin, the inhibition zone and MIC
were considered. We calculated percent inhibition area per micro-
gram of the compound and compared it with the positive control. All
the thiosemicarbazones (1Ae24A) were found less effective than
amoxicillinwhen % inhibition area was compared. Although some of
them showed 80% resemblance with the standard drug. From the
initial screening of thiadiazoles (1Be24B) and comparing the %
2.2.2. Toxicity profile
To ensure the toxicity of the compounds 5B, 8B, 10B, 13B, 14B,
15B, 21B and 22B with better or nearer to, ratio of percent area
inhibited per mg compared with positive control, were selected for
cytotoxic studies. The compounds were tested against H9c2 cardiac
myoblasts. The results were compared with a negative control
(DMSO) and a positive control (amoxicillin). A subconfluent pop-
ulation of H9c2 cells was treated with increasing concentrations of
test compounds and the positive control and the number of viable
cells was measured after 48 h by MTT cell viability assay. Fig. 3
depicted that all these compounds exhibited >80% viability at the
concentration range of 1.57e200 mg mLꢁ1. An ELISA plate reader
(Labsystems Multiskan RC, Helsinki, Finland) at 570 nm with
a reference wavelength of 655 nm was used.
inhibition area per mg, it was found that compound 10B and 21B
shows better inhibition in S. pyogens than amoxicillin, while 13B,15B
and 22B were approximately similar to the amoxicillin inhibition.
Similarly comparing the percent inhibition area in S. aureus,
compound 5B and 14B showed better results than the control
positive and compound 10B and 8B were found approximately
similar to the control positive (Fig. 2). When the percent area
inhibited ratio of gram-negative bacteria was compared, none of the
compounds showed better result than the positive control.
The distinct difference in the antibacterial property of the thi-
osemicarbazones and the thiadiazoles justifies the purpose of this
study. The importance of such work lies in the possibility that the
new compound might be more effective against bacteria for which
2.2.3. Structure activity relationship
The new series of thiadiazoles were designed in a way to find-out
the effect of substituted aliphatic and aromatic moieties on the
biological activity. The thiadiazole molecule was substituted at C5
with three different 8-substitutedchloroquinoline (Cl, H, and CH3)
derivatives and at C2 with N-(substituted)acetamide (aromatic and
Fig. 3. The graph showing effect of different concentrations of thiadiazoles on percent viability of H9c2 cardiac myoblast cell line.

3162
A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
cyclic) derivatives. From the results it was found that thiadiazole
derivatives substituted with N-(aromatic)acetamide group at C2
shows better inhibitory effects. Among the aromatic compounds the
N-(o-tolyl)acetamide and N-(p-tolyl)acetamide showed better
activity. However N-(o-tolyl)acetamide in combinationwith the 2,8-
dichloroquinoline substituted at C5 of the thiadiazole ring showed
much better inhibitory effect in S. aureus, compared on the basis of
percent area of inhibitionper microgram. This was followed by N-(o-
tolyl)acetamide in combination with 2-chloroquinoline and N-(p-
tolyl)acetamide in combination with chloroquinoline. When o-tolyl
group was used in combination with 2-chloro-8-methylquinoline,
the effect was less compared to 2-chloro and 2,8-dichloroquinoline.
The trend was found opposite in case of N-(2,5-difluorophenyl)
acetamide substituted at C2 of thiadiazole molecule. When
difluorophenyl molecule was used in combination with 2-chloro,8-
methyl quinolone attached at C5 of the thiadiazole ring, the effect
was higher compared to 2-chloroquinoline. N-(p-nitropheyl)acet-
amide when used with 2-chloroquinoline also showed effect on the
inhibition of growth of S. pyogenes. But the effect was not as prom-
inent with other aromatic substituted groups. This shows that the
substituent group play a definite role on the biological activity of
these molecules.
4.2. Preparation of 4-Substituted1-((2,8-dichloroquinolin-3-yl)
methylene)thiosemicarbazide
Synthesizedthiosemicarbazides(1 equiv.)weredissolved in10mL
of absolute EtOH, followed by addition of a solution of 2,8-disubsti-
tutedquinoline-3-carboxaldehyde(1equiv.)inEtOH. Afterstirringfor
aninitialperiodof15e30min, AcOHwasadded incatalyticamount to
increase the yield of the reaction. The mixture was heated to reflux at
60 ꢀC for 12 h. The resultant mixture was evaporated in vacuo to give
crude solid products, which were further purified by column chro-
matography (CH2Cl2/MeOH) to yield compounds 1Ae24A.
4.2.1. 1-((2,8-Dichloroquinolin-3-yl) methylene)thiosemicarbazide
(1A)
Light Yellowish; yield 79%; mp 165 ꢀC; Anal. Calc. for
C11H8Cl2N4S: C 44.16, H 2.70, N 18.73, found C 44.13, H 2.71, N 18.71;
IR
NMR (CDCl3)
n
(cmꢁ1): 3466 (NH2),3163 (NH), 1465 (C]N), 1109 (C]S); 1H
d(ppm): 7.39e7.69 (m, 4H, Ar-H), 7.1 (s, 1H, CH]N),
5.96 (s, 1H, NH2), 2.4 (s, 2H, NH2); ESI-MS m/z: [Mþ þ Na] 322.1
4.2.2. 1-((2,8-Dichloroquinolin-3-yl)methylene)-4-cyclopentyl-
thiosemicarbazide (2A)
Dark Brown; yield 76%; mp 158 ꢀC; Anal. Calc. for C16H16Cl2N4S:
C 52.32, H 4.39, N 15.25, found C 52.29, H 4.41, N 15.24; IR
3197 (NH), 2829 (CH2), 1447 (C]N), 1110 (C]S); 1H NMR (CDCl3)
/ppm: 11.74 (s, 1H, NNH), 8.6 (s, 1H, Ar-H), 7.43e7.56 (m, 3H, Ar-H),
n
(cmꢁ1):
3. Conclusion
d
A new library of thiosemicarbazones and thidiazoles were
synthesized with an aim to target the microorganisms that cause
microbial infection. The compounds were synthesized keeping in
mind the molecular weight, antibiotics literally mean low molec-
ular weight chemical compounds. Considering the individual drug
activity of quinolines, thiosemicarbazones and thiadiazoles, we
designed the synthesis in such a way that products will possess
a quinoline moiety and the corresponding thiosemicarbazones and
thiadizoles and will posses low molecular weight. In all the thio-
semicarbazones (1Ae24A) and thiadiazoles (1Be24B) synthesized
and screened for anti-microbial activity compounds 5B and 14B
showed better inhibitory effects for S. aureus and 13B and 15B
showed better results for S. pyogenes.
6.91(s, 1H, N]CH), 5.99 (s, 1H, CNH), 2.82e2.93 (m, 1H, NCH),
1.93e2.3 (m, 8H, CH2); ESI-MS m/z: [Mþ þ H] 368.3
4.2.3. 1-((2,8-Dichloroquinolin-3-yl)methylene)-4-cyclohexyl-
thiosemicarbazide (3A)
Light Yellowish; yield 81%; mp 160 ꢀC; Anal. Calc. for
C17H18Cl2N4S: C 60.92, H 4.26, N 15.79, found C 62.94, H 4.24,
N 15.77; IR
n
(cmꢁ1): 3259 (NH), 2863 (CH2), 1475 (C]N), 1117 (C]
S); 1H NMR (DMSO)
d(ppm): 11.17 (s, 1H, NNH), 8.19 (s, 1H, Ar-H),
7.39e7.53 (m, 3H, Ar-H), 7.28 (s, 1H, CH]N), 6.13 (s, 1H, CNH),
2.93 (m, 1H, NCH), 2.29e2.50 (m, 10H, CH2); ESI-MS m/z: [Mþ þ H]
382.2
4.2.4. 1-((2,8-Dichloroquinolin-3-yl)methylene)-4-cyclooctyl-
thiosemicarbazide (4A)
4. Experimental
Blackish; yield 62%; mp 161 ꢀC; Anal. Calc. for C19H22Cl2N4S:
n
(cmꢁ1):
Reactions were conducted in oven dried glassware. Solvents were
purified prior to use. Anhydrous sodium sulfate was used as the
drying agent. All the melting points were recorded on a KSW appa-
ratus and were uncorrected. Elemental analyses were performed by
Regional Sophisticated Instrumentation Centre, Central drug
Research Institute, Lucknow, India. IR spectra were recorded on
PerkineElmer model 1600 FI-IR (KBr Pellet in the 4000e400 cmꢁ1
range). The 1H NMR spectra were obtained at 300 MHz in CDCl3,
DMSO-d6 by using Bruker spectrospin and Varian instrument using
TMS as internal standard and chemical shift values are given in ppm
downfield to TMS (tetramethylsilane). ESI-MS was recorded on
a Micromass Quattro II triple quadrupole mass spectrometer. Split-
ting patternsaredesignatedasfollows:s,singlet;d,doublet;t, triplet;
m, multiplet. Chemical shift values are given in parts per million. The
reactions were monitored by precoated aluminum-sheet TLC using
chloroform:methanol (5:1) as eluant and column chromatography
was accomplished using Silica gel, 60 Ǻ (200e400 mesh).
C 55.74, H 5.42, N 13.69, found C 55.76, H 5.44, N 13.71; IR
3250 (NH), 2822 (CH2), 1777 (C]C), 1115 (C]S); 1H NMR (CDCl3)
(ppm): 11.53 (s,1H, NNH), 8.68 (s,1H, Ar-H), 7.45e7.69 (m, 3H, Ar-H),
d
7.13 (s, 1H, CH]N), 5.67 (s, 1H, CNH), 2.42e2.53 (m, 1H, NCH),
2.07e2.31 (m, 4H, CH2), 1.73e1.92 (m, 10H, CH2); ESI-MS m/z:
[Mþ þ Na] 432.3
4.2.5. 1-((2,8-Dichloroquinolin-3-yl)methylene)-4-o-tolylthiosemi-
carbazide 5A
White; yield 69%; mp 163 ꢀC; Anal. Calc. for C18H14Cl2N4S :
C 55.53, H 3.62, N 14.39, found C 55.55, H 3.59, N 14.37; IR
3260 (NH), 3020 (Ar,CeH), 2928 (CH3), 1446 (C]N), 1113 (C]S); 1H
NMR (CDCl3) (ppm): 11.93 (s, 1H, NNH), 9.06 (s, 1H, CNH),
n
(cmꢁ1):
d
7.34e8.69 (m, 8H, Ar-H), 7.12 (s, 1H, CH]N), 2.31 (s, 3H, CH3); ESI-
MS m/z: [Mþ þ H] 390.4
4.2.6. 1-((2,8-Dichloroquinolin-3-yl)methylene)-4-(2,5-difluoro-
phenyl)thiosemicarbazide (6A)
4.1. Preparation of thiosemicarbazides
White; yield 73%; mp 165 ꢀC; Anal. Calc. for C17H10Cl2F2N4S:
C 49.65, H 2.45, N 13.62, found C 49.69, H 2.50, N 13.58; IR
n
(cmꢁ1):
Substituted thiosemicarbazides were prepared by a two-step
synthetic route as reported in our previous paper [30]. The data
was compared with our previous work.
3199 (NH), 3092 (Ar, CeH),1476 (C]N), 1125 (C]S); 1H NMR
(DMSO) d(ppm): 11.23 (s, 1H, NNH), 7.9e8.65 (m, 7H, Ar-H), 7.21 (s,
1H, CH]N); ESI-MS m/z: [Mþ þ H] 412.26.

A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
3163
4.2.7. 1-((2,8-Dichloroquinolin-3-yl)methylene)-4-p-tolylthiosemi-
4.2.15. 1-((2-Chloroquinolin-3-yl)methylene)-4-p-
tolylthiosemicarbazide (15A)
Pinkish; yield 77%; mp 163 ꢀC; Anal. Calc. for C18H15ClN4S: C 60.92,
H 4.26, N 15.79, found C 60.89, H 4.30, N 15.76; IR
(cmꢁ1): 3155 (NH),
3064 (Ar,CeH),1506 (C]N), 1132 (C]S); 1H NMR (CDCl3)
(ppm):
carbazide (7A)
Brownish; yield 76%; mp 164 ꢀC; Anal. Calc. for C18H14Cl2N4S:
C 55.53, H 3.62, N 14.39, found C 55.51, H 3.60, N 16.43; IR
3255 (NH), 3062 (Ar,CeH), 2914 (CH3),1486 (C]N), 1127 (C]S); 1H
NMR (CDCl3) (ppm): 12.13 (s, 1H, NNH), 9.01(s, 1H, Ar-H), 8.31 (s,
n
(cmꢁ1):
n
d
d
11.57 (s,1H, NNH), 8.71 (s,1H, NH), 8.21 (s,1H, Ar-H), 7.29e7.64 (m, 8H,
1H, NH), 7.38e7.86 (m, 7H, Ar-H.), 7.13 (s, 1H, CH]N), 2.12 ( s, 3H,
CH3); ESI-MS m/z: [Mþ] 389.1.
Ar-H), 6.97 (s,1H, CH]N), 2.37 (s, 3H, CH3); ESI-MS m/z:[Mþ þ 1]355.9
4.2.16. 1-((2-Chloroquinolin-3-yl)methylene)-4-p-
nitrophenylthiosemicarbazide (16A)
4.2.8. 1-((2,8-Dichloroquinolin-3-yl)methylene)-4-p-nitrophenyl-
thiosemicarbazide (8A)
Light Black; yield 87%; mp 162 ꢀC; Anal. Calc. for C17H12ClN5O2S:
LightPinkish;yield67%;mp171 ꢀC; Anal. Calc. forC17H11Cl2N5O2S
C 52.92, H 3.13, N 18.15, found C 52.96, H 3.17, N 18.12; IR
n
(cmꢁ1):
: C 48.58, H 2.64, N 16.66, found C 48.62, H 2.61, N 16.68; IR
n
(cmꢁ1):
3187 (NH), 3057 (Ar,CeH), 1457 (C]N), 1131 (C]S); 1H NMR
3187 (NH), 3065 (Ar,CeH),1463 (C]C),1131 (C]S); 1H NMR (DMSO)
(DMSO) d(ppm): 11.13 (s, 1H, NNH), 9.54 (s, 1H, NH), 7.37e8.10 (m,
d
(ppm): 11.69 (s,1H, NNH), 9.69 (s,1H, NH), 7.51e8.09 (m, 8H, Ar-H),
9H, Ar-H), 6.97 (s, 1H, CH]N); ESI-MS m/z: [Mþ þ 1] 386.6
6.72 (s,1H, CH]N); ESI-MS m/z: [Mþ þ H] 421.3.
4.2.17. 1-((2-Chloro-8-methylchloroquinolin-3-yl)methylene)
thiosemicarbazide (17A)
4.2.9. 1-((2chloroquinolin-3-yl)methylene)thiosemicarbazide (9A)
Brown; yield 73%; mp 160 ꢀC; Anal. Calc. for C11H9ClN4S: C 49.91,
Yellowish; yield 74%; mp 162 ꢀC; Anal. Calc. for C12H11ClN4S :
H 3.43, N 21.16, found C 49.89, H 3.47, N 21.14; IR
n
(cmꢁ1): 3477
(ppm):
C 51.70, H 3.98, N 20.10, found C 51.66, H 4.02, N 20.13; IR
3465 (NH2), 3199 (NH), 2956 (CH3), 1421 (C]N), 1121 (C]S); 1H
NMR (CDCl3) (ppm): 11.74 (s, IH, NNH), 9.3 (s, 1H, Ar-H), 7.19e7.39
n
(cmꢁ1):
(NH2), 3160 (NH),1469 (C]N),1108 (C]S); 1H NMR (CDCl3)
d
10.96 (s, 1H, NNH), 9.3 (s, 1H, Ar-H), 7.13e7.26 (m, 4H, Ar-H), 7.11 (s,
1H, CH]N), 2.38 (s, 2H, NH2); ESI-MS m/z: [Mþ þ H] 266.6.
d
(m, 3H, Ar-H), 6.85 (s, 1H, N]CH), 2.51 (s, 3H, CH3), 2.19 (s, 2H,
NH2); ESI-MS m/z: [Mþ þ Na] 301.9
4.2.10. 1-((2-Chloroquinolin-3-yl)methylene)-4-cyclopentyl-
thiosemicarbazide (10A)
4.2.18. 1-((2-Chloro-8-methylchloroquinolin-3-yl)methylene)-4-
cyclopentylthiosemicarbazide (18A)
Reddish brown; yield 82%; mp 162 ꢀC; Anal. Calc. for C16H17ClN4S : C
57.73, H 5.15, N 16.83, found C 57.77, H 5.13, N 16.85; IR
(NH), 2886 (CH2), 1431 (C]N), 1113 (C]S); 1H NMR (CDCl3)
n
(cmꢁ1): 3179
(ppm):
Brownish; yield 78%; mp 154 ꢀC; Anal. Calc. for C17H19ClN4S :
d
C 58.86, H 5.52, N 16.15, found C 58.82, H 5.55, N 16.19; IR
3497 (NH), 2946 (CH3), 2857 (CH2),1477 (C]N), 1110 (C]S); 1H
NMR (CDCl3) (ppm): 11.11 (s, 1H, NNH), 8.69 (s, 1H, Ar-H),
n
(cmꢁ1):
11.14 (s, 1H, NNH), 7.21e7.37(m5H,Ar-H),7.03(s,1H,N]CH), 2.76e2.81
(m, 1H, NeCH), 1.53e1.69 (m, 8H, CH2); ESI-MS m/z: [Mþ] 332.5
d
7.29e7.46 (m, 3H, Ar-H), 7.06 (s, 1H, N]CH), 2.81e2.94 (m, 1H,
NeCH), 1.73e1.96 (m, 8H, CH2); ESI-MS m/z: [Mþ þ 1] 347.7
4.2.11. 1-((2-Chloroquinolin-3-yl)methylene)-4-cyclohexyl-
thiosemicarbazide (11A)
Creamy White; yield 73%; mp 162 ꢀC; Anal. Calc. for C17H19ClN4S:
4.2.19. 1-((2-Chloro-8-methylchloroquinolin-3-yl)methylene)-4-
cyclohexylthiosemicarbazide (19A)
C 58.86, H 5.52, N 16.15, found C 58.89, H 5.54, N 16.14; IR
3189 (NH), 2867 (CH2),1461 (C]N), 1107 (C]S); 1H NMR (DMSO)
(ppm): 11.07 (s,1H, NNH), 7.49e7.61 (m. 4H, Ar-H), 6.88 (s,1H, CH]
n
(cmꢁ1):
Blackish; yield 75%; mp 155 ꢀC; Anal. Calc. for C18H21ClN4S:
d
C 59.90, H 5.86, N 15.52, found C 59.93, H 5.89 N 15.449; IR
3189 (NH), 2852 (CH2),1411 (C]N), 1107 (C]S); 1H NMR (DMSO)
(ppm): 12.07 (s, 1H, NNH), 8.19 (s, 1H, Ar-H), 7.56e7.72 (m, 3H, Ar-
n
(cmꢁ1):
N), 4.93 (s, 1H, CNH), 2.93e3.04 (m, 1H, NCH), 1.89e2.10 (m, 10H,
CH2); ESI-MS m/z: [Mþ þ Na] 370.1
d
H), 7.14 (s, 1H, CH]N), 6.23 (s, 1H, CNH), 2.89e3.01 (m, 1H, NCH),
2.45 (s, 3H, CH3), 1.91e2.05 (m, 10H, CH2); ESI-MS m/z: [Mþ þ 1]
361.9
4.2.12. 1-((-Hloroquinolin-3-yl)methylene)-4-cyclooctyl-
thiosemicarbazide (12A)
Light Brownish; yield 82%; mp 165 ꢀC; Anal. Calc. for C19H23ClN4S
: C 60.87, H 6.18, N 14.94, found C 60.90, H 6.16, N 14.90; IR
3250 (NH),2861 (CH2), 1465 (C]N), 1130 (C]S); 1H NMR (CDCl3)
(ppm): 10.89 (s,1H, NNH), 7.63e7.71 (m, 5H, Ar-H), 7.18 (s,1H, CH]
n
(cmꢁ1):
4.2.20. 1-((2-Chloro-8-methylchloroquinolin-3-yl)methylene)-4-
cyclooctylthiosemicarbazide (20A)
d
Yellowish; yield 80%; mp 158 ꢀC; Anal. Calc. for C19H23ClN4S: C 60.87,
N), 5.78 (s,1H, CNH), 2.34e2.52 (m,1H, NCH),1.97e2.13 (m, 4H, CH2),
1.61e1.74 (m, 10H, CH2); ESI-MS m/z: [Mþ þ H] 375.8
H6.18,14.94, found C 60.91, H6.21, N 14.91; IR
n
(cmꢁ1): 3245(NH), 2859
(CH2), 1465 (C]N), 1125 (C]S); 1H NMR (CDCl3)
d(ppm): 11.12 (s, 1H,
NNH), 8.68 (s, 1H, Ar-H), 7.48e7.65 (m, 3H, Ar-H), 7.09 (s, 1H, CH]N),
5.32 (s, 1H, CNH), 2.54e2.61 (m, 1H, NCH), 2.39 (s, 3H, CH3), 2.05e2.25
(m, 4H, CH2), 1.76e1.91 (m, 10H, CH2); ESI-MS m/z: [Mþ þ 1] 375.8
4.2.13. 1-((2-Chloroquinolin-3-yl)methylene)-4-o-tolylthiosemi-
carbazide (13A)
Whitish; yield 79%; mp 171 ꢀC; Anal. Calc. for C18H15ClN4S :
C 60.92, H 4.26, N 15.79, found C 60.92, H 4.25, N 18.81; IR
3160 (NH), 3064 (Ar,CeH), 1472 (C]N), 1123 (C]S); 1H NMR
(CDCl3) (ppm): 11.93 (s, 1H, NNH), 9.06 (s, 1H, CNH) 7.12e7.68 (m,
n
(cmꢁ1):
4.2.21. 1-((2-Chloro-8-methylchloroquinolin-3-yl)methylene)-4-o-
tolylthiosemicarbazide (21A)
d
Creamy; yield76%;mp 155ꢀC; Anal. Calc. forC19H17ClN4S :C 61.86
9H, Ar-H), 7.05 (s, 1H, CH]N), 2.56 (s, 3H, CH3); ESI-MS m/z:
[Mþ þ H] 355.8
H 4.65 N 15.19 found C 61.85, H 4.68, N 15.23; IR
n
(cmꢁ1): 3160 (NH),
3095(Ar,CeH), 2964 (CH3),1433(C]N),1113(C]S);1H NMR(CDCl3)
d
(ppm):11.53(s,1H, NNH), 9.72 (s,1H, CNH), 7.85e8.10 (m, 8H, Ar-H),
4.2.14. 1-((2-Chloroquinolin-3-yl)methylene)-4-(2,5-difluoro-
phenyl)thiosemicarbazide (14A)
6.85 (s, 1H CH]N), 2.46 (s, 6H, CH3); ESI-MS m/z: [Mþ þ 1] 369.6
Bluish; yield 74%; mp 179 ꢀC; Anal. Calc. for C17H11ClF2N4S: C 54.19,
H2.94, N14.87, foundC54.15,H2.89, N14.90, IR:3458(NH),1504(C]
4.2.22. 1-((2-Chloro-8-methylchloroquinolin-3-yl)-4-(2,5-difluoro-
phenyl)thiosemicarbazide (22A)
C),1035(C]S).1HNMR(DMSO)
d
(ppm):10.89(s,1H, NNH),7.63e7.85
Light black; yield 83%; mp 158 ꢀC; Anal. Calc. for C18H13ClF2N4S :
(m, 8H, Ar-H), 6.93 (s, 1H, CH]N); ESI-MS m/z: [Mþ þ Na] 399.6
C 55.32, H 3.35, N 14.34, found C 55.35, H 3.51, N 14.36; IR n
(cmꢁ1):

3164
A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
3158 (NH), 3035 (Ar,CeH), 2934 (CH3), 1474 (C]N), 1115 (C]S); 1H
4.3.4. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-cyclooctylacetamide (4B)
NMR (DMSO) d(ppm): 11.71(s,1H, NNH), 7.76e7.98 (m, 7H, Ar-H),
7.17 (s, 1H, CH]N), 2.64 (s, 3H, CH3); ESI-MS m/z: [Mþ þ H] 391.9
Light black; yield 57%; mp 210 ꢀC; Anal. Calc. for C23H26Cl2N4O2S:
C 55.98, H 5.31, N 11.35, found: C 55.94, H 5.33, N 11.332; IR
2933 (CH3), 2872 (CH2), 1674, 1641 (CO), 1492 (C]N); 1H NMR
(CDCl3) (ppm): 8.61 (s, 1H, Ar-H), 7.48e7.69 (m, 3H, Ar-H), 5.36 (s,
n
(cmꢁ1):
4.2.23. 1-((2-Chloro-8-methylchloroquinolin-3-yl)-4-p-tolyl-
thiosemicarbazide (23A)
d
White; yield 76%; mp 160 ꢀC; Anal. Calc. for C19H17ClN4S: C
1H, SCHN), 2.69 (s, 6H, CH3), 2.32e2.41 (m, 1H, NCH), 1.99e2.11 (m,
4H, CH2), 1.58e1.85 (m, 10H, CH2); ESI-MS m/z: [Mþ þ 1] 494.2
61.86, H 4.65, N 15.19, found C 61.82, H 4.69, N 15.16; IR
3155 (NH), 3062 (Ar,CeH), 2912 (CH3), 1486 (C]N), 1102 (C]S); 1H
NMR (CDCl3) (ppm): 11.96 (s, 1H, NNH), 9.69 (s, 1H, NH), 7.28e7.56
n
(cmꢁ1):
d
4.3.5. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-o-tolylacetamide (5B)
(m, 8H, Ar-H), 6.94 (s, 1H, CH]N), 2.69 (s, 6H, CH3); ESI-MS m/z:
[Mþ þ H] 369.7
White; yield 58%; mp 196 ꢀC; Anal. Calc. for: C22H18Cl2N4O2S:
C 55.82, H 3.83, N 11.84, found: C 55.84, H 3.80, N 11.80; IR
n
(cmꢁ1):
4.2.24. 1-((2-Chloro-8-methylchloroquinolin-3-yl)-4-p-nitrophenyl-
thiosemicarbazide (24A)
3135 (Ar,CeH), 2963 (CH3), 1655&1643 (CO), 1435 (C]N); 1H NMR
(CDCl3)d(ppm):8.53(s,1H, Ar-H), 6.95e7.31(m, 7H,Ar-H), 5.69(s,1H,
White; yield 80%; mp 165 ꢀC; Anal. Calc. for C18H14ClN5O2S: C
SCHN), 2.35 (s, 3H, CH3), 2.13 (s, 6H, CH3); ESI-MS m/z: [Mþ þ H] 474.0
54.07, H 3.53, N 17.51, found C 54.04, H 3.55, N 17.49; IR
3187 (NH), 3021 (Ar,CeH), 2953 (CH3), 1477 (C]N), 1131 (C]S); 1H
NMR (DMSO) (ppm): 11.43 (s, 1H, NNH), 9.34 (s, 1H, NH),
n
(cmꢁ1):
4.3.6. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-(2-5-difluorophenyl)acetamide (6B)
d
7.68e7.96 (m, 8H, Ar-H), 6.79 (s, 1H, CH]N); ESI-MS m/z:
White; yield 67%; mp 206 ꢀC; Anal. Calc. for C21H14Cl2F2N4O2S:
[Mþ þ Na] 422.7.
C 50.92, H 2.85, N 11.31, found C 50.88, H 2.82, N 11.35; IR
n
(cmꢁ1):
3092 (Ar,CeH), 2952 (CH3), 1664, 1642 (CO), 1435 (C]N); 1H NMR
4.3. Synthesis of N-4-acetyl-5-(2,8-substitutedquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadi-azole-2-yl)N-substituted-acetamide
derivatives (1Be24B)
(DMSO) d(ppm): 7.78e8.62 (m, 7H, Ar-H), 5.69 (s, 1H, SCHN), 2.61
(s, 6H, CH3); ESI-MS m/z: [Mþ þ H] 496.1
4.3.7. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-p-tolylacetamide (7B)
Compounds (1Ae24A) were refluxed with excess of acetic aicd
anhydride, with a constant stirring for 15e24 h. The reaction was
monitored by TLC. The solution was poured into ice-water and
stirred for 3 h. The solution was filtered and the solid obtained was
washed with diethyl ether. The precipitated solid was recrystalized
from appropriate solvents to get the desired products. Some
compounds were subjected to column chromatography to get the
desired compounds.
Brown; yield 53%; mp 201 ꢀC; Anal. Calc. for C22H18Cl2N4O2S: C
55.82, H 3.83, N 11.84, found C 55.84, H 3.79, N11.81; IR
3049 (Ar,CeH), 2942 (CH3), 1678, 1640 (CO), 1449 (C]N); 1H NMR
n
(cmꢁ1):
(CDCl3) d(ppm): 7.38e7.86 (m, 8H, Ar-H), 4.98 (s,1H, SCHN), 2.39 ( s,
6H, CH3), 2.13 ( s, 3H, CH3); ESI-MS m/z: [Mþ þ Na] 518.2
4.3.8. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-(4-nitrophenyl)acetamide (8B)
4.3.1. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)acetamide (1B)
Light pink; yield 56%; mp 194 ꢀC; Anal. Calc. for C21H15Cl2N5O4S:
C 50.01, H 3.00, N 13.89, found C 49.99, H 3.03, N 13.87; IR
3065 (Ar,Ctyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-(4-nitrophenyl)H), 2971 (CH3), 2551 (CH2)
1673, 1640 (CO), 1445 (C]N); 1H NMR (DMSO)
(ppm): 7.73e8.21
n
(cmꢁ1):
Brownish yellow; yield 66%; mp 193 ꢀC; Anal. Calc .for
C15H22Cl2N4O2S: C 47.01, H 3.16, N 14.62 found: C 46.98, H 3.13, N
14.68; IR
(C]N); 1H NMR (CDCl3)
1H, SCHN), 2.53 (s, 6H, CH3); 13C NMR (CDCl3)
n
(cmꢁ1): 3211 (NH), 2953 (CH3)1658 & 1641 (CO), 1479
d
d
(ppm): 7.13e7.26 (m, 4H, Ar-H), 5.31 (s,
/ppm: 169.24 (C]
(m, 8H, Ar-H), 5.41 (s, 1H, SCHN), 2.13 (s, 6H, CH3); ESI-MS m/z:
d
[Mþ þ 1] 505.7
O), 163.36 (C]O), 158.61 (C]N), 149.32 (C]N), 139.16, 134.95,
129.24, 128.84, 126.26, 125.21 (Ar-C), 65.98 (SCN), 24.32, 26.10
(Aliphatic). ESI-MS m/z: [Mþ þ H] 384.2
4.3.9. N-4-acetyl-5-(2-chloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)acetamide (9B)
Reddish brown; yield 61%; mp 198 ꢀC; Anal. Calc. for
C15H13ClN4O2S: C 51.65, H 3.76, N 16.06, found C 51.62, H 3.79,
4.3.2. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-cyclopentylacetamide (2B)
N 16.11; IR
n
(cmꢁ1): 3183 (NH), 2926 (CH3), 1658, 1641 (CO), 1479
Brownish; yield 61%; mp 189 ꢀC; Anal. Calc. for C20H20Cl2N4O2S:
(C]N); 1H NMR (CDCl3)
d(ppm): 8.76 (s, 1H, Ar-H), 6.98e7.39 (m,
C 53.22, H 4.47, N 12.41, found C 53.20, H 4.49, N 12.43; IR
2911 (CH3), 2851 (CH2), 1671&1661 (CO), 1440 (C]N); 1H NMR
(CDCl3) (ppm): 8.51 (s, 1H, Ar-H), 7.21e7.37 (m, 3H, Ar-H), 5.25(s,
1H, SCHN), 2.65e2.2.78 (m, 1H, NeCH), 2.35 (s, 6H, CH3), 1.99e2.28
(m, 8H, CH2); 13C NMR (CDCl3)
/ppm: 171.23 (C]O), 168.21 (C]O),
n
(cmꢁ1):
7H, Ar-H), 5.41(s, 1H, SCHN), 2.36 (s, 6H, CH3); ESI-MS m/z:
[Mþ þ H] 349.8
d
4.3.10. N-4-acetyl-5-(2-chloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-cyclopentylacetamide (10B)
d
158.86 (C]N), 149.96 (C]N), 143.01, 137.25. 135.56, 129.24, 126.21
(Ar-C), 65.49 (SCN), 34.27, 27.26, 24.37, 23.73 21.98 (aliphatic,
cyclic); ESI-MS m/z: [Mþ þ 1] 452.2
Brownish yellow; yield 66%; mp 205 ꢀC; Anal. Calc. for
C20H21ClN4O2S: C 57.62, H 5.08, N 13.44, found C 57.65, H 5.06, N
13.46, IR
n
(cmꢁ1): 2928 (CH3), 2896 (CH2) 1661, 1652 (CO), 1440
(C]N); 1H NMR (CDCl3)
d
(ppm): 7.29e7.46 (m 5H, Ar-H), 5.51(s,1H,
4.3.3. N-4-acetyl-5-(2,8-dichloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-cyclohexylacetamide (3B)
SCHN), 2.71e2.81 (m, 1H, NeCH), 2.35 (s, 6H, CH3), 1.73e1.86 (m,
8H, CH2); ESI-MS m/z: [Mþ þ H] 417.8
Yellowish; yield 61%; mp 208 ꢀC; Anal. Calc. for C21H22Cl2N4O2S:
C 54.20, H 4.76, N 12.04, found C 54.22, H 4.73, N 12.06; IR
n
(cmꢁ1):
4.3.11. N-4-acetyl-5-(2-chloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-cyclohexylacetamide (11B)
2952 (CH3), 2859 (CH2), 1651&1640 (CO), 1430 (C]N). 1H NMR
(DMSO)
d
(ppm): 7.56e7.72 (m. 4H, Ar-H), 5.56 (s, 1H, SCHN), 2.68
Creamy white; yield 53%; mp 221 ꢀC; Anal. Calc. for
C21H23ClN4O2S: C 58.53, H 5.38, N 13.00, found C58.49, H 5.34,
(s, 6H, CH3), 1.73e1.91 (m, 10H, CH2); ESI-MS m/z: [Mþ þ Na] 488.3

A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
3165
N 13.03; IR
(C]N); 1H NMR (DMSO)
1H, SCHN), 2.63 (s, 6H, CH3), 2.32e2.41 (m, 1H, NCH), 1.99e2.13 (m,
10H, CH2); 13C NMR (DMSO)
/ppm: 169.35 (C]O), 166.92 (C]O),
158.39 (C]N), 153.24 (C]N), 139.22, 136.95, 132.10,130.29, 129.13,
123.12 (Ar-C), 65.87 (SCN), 37.02, 29.95, 27.36, 26.95, 22.01, 21.23,
19.89 (aliphatic); ESI-MS m/z: [Mþ þ H] 431.6
n
(cmꢁ1): 2918 (CH3), 2876 (CH2), 1651, 1640 (CO), 1430
(ppm): 7.39e7.353 (m, 5H, Ar-H), 5.37 (s,
12.96; IR
n
(cmꢁ1): 2918 (CH3), 2851 (CH2), 1661, 1649 (CO), 1440
(ppm): 1.92e2.16 (m, 8H, CH2), 2.38 (s,
d
(C]N); 1H NMR (CDCl3)
d
6H, CH3), 2.64 (s, 3H, CH3), 2.81e2.94 (m, 1H, NeCH), 5.34 (s, 1H,
N]CH), 7.73e7.56 (m, 3H, Ar-H), 8.24 (s, 1H, Ar-H); ESI-MS m/z:
[Mþ þ H] 431.6
d
4.3.19. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadiazole-2-yl)-N-cyclohexylacetamide (19B)
Light black; yield 63%; mp 188 ꢀC; Anal. Calc. for C22H25Cl N4O2S:
4.3.12. N-4-acetyl-5-(2-chloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-cyclooctylacetamide (12B)
C 59.38, H 5.66, N 12.59 found C 59.42, H 5.63, N 12.56; IR
2933 (CH3), 2867 (CH2), 1660, 1651 (CO), 1430 (C]N); 1H NMR
(DMSO) (ppm): 8.36 (s, 1H, Ar-H), 7.49e7.61 (m. 3H, Ar-H), 6.23 (s,
n
(cmꢁ1):
Brownish; yield 69%; mp 208 ꢀC; Anal. Calc. for C23H27ClN4O2S:
C 60.20, H 4.36, N 12.76, found : C 60.24, H 4.33, N 12.79; IR
2937 (CH3), 2861 (CH2), 1672, 1651 (CO), 1492 (C]N); 1H NMR
(CDCl3) (ppm): 10.89 (s, 1H, NNH), 7.45e7.69 (m, 5H, A-H), 5.39 (s,
1H, SCHN), 2.56e2.61 (m, 1H, NCH), 2.43 (s, 6H, CH3), 1.72e1.81 (m,
4H, CH2), 1.49e1.65 (m, 10H, CH2); 13C NMR (CDCl3)
(ppm): 175.52
n
(cmꢁ1):
d
1H, CNH), 5.26 (s, 1H, SCHN), 2.92e3.01 (m, 1H, NCH), 2.59 (s, 6H,
CH3), 2.51 (s. 3H, CH3), 1.73e1.93 (m, 10H, CH2); ESI-MS m/z:
[Mþ þ H] 445.1
d
d
(C]O), 165.32 (C]O), 153.95 (C]N), 147.86 (C]N), 137.32, 135.20,
129.01, 128.24, 127.35 (Ar-C), 53.11 (SCN), 38.96, 33.25, 29.94, 27.27,
24.95, 21.43 (aliphatic); ESI-MS m/z: [Mþ þ H] 460.2
4.3.20. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadiazole-2-yl)-N-cyclooctylacetamide (20B)
Brownish; yield 54%; mp 219 ꢀC; Anal. Calc. for C24H29ClN4O2S:
C 60.94, H 6.18, N 11.84 found : C 60.92, H 6.21, N 11.80; IR
2937 (CH3), 2891 (CH2), 1670, 1640 (CO), 1492 (C]N); 1H NMR
(CDCl3) (ppm): 5.12 (s, 1H, SCHN), 7.63e7.82 (m, 4H, Ar-H),
n
(cmꢁ1):
4.3.13. N-4-acetyl-5-(2-chloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-o-tolylacetamide (13B)
d
Creamy white; yield 64%; mp 192 ꢀC; Anal. Calc. for:
C22H19ClN4O2S: C 60.20, H 4.36, N 12.78, found: C 60.23, H 4.34,
2.51e2.62 (m, 1H, NCH), 2.48 (s, 6H, CH3), 2.37 (s, 3H, CH3),
1.65e1.73 (m, 4H, CH2), 1.31e1.56 (m, 10H, CH2); ESI-MS m/z:
[Mþ þ H] 438.2
N 12.75; IR n
(cmꢁ1): 3064 (Ar,CeH), 2926 (CH3),1655,1642 (CO),1435
(C]N); 1H NMR (CDCl3)
d(ppm): 7.16e7.69 (m, 9H, Ar-H), 5.59 (s, 1H,
SCHN), 2.51(s,3H, CH3), 2.43(s, 6H, CH3); ESI-MS m/z:[Mþ þ Na] 461.7
4.3.21. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadiazole-2-yl)-N-o-tolylacetamide (21B)
Creamy white; yield 56%; mp 203 ꢀC; Anal. Calc. for:
C23H21ClN4O2S: C 60.99, H 4.67, N 12.37, found: C 60.95, H 4.71, N
4.3.14. N-4-acetyl-5-(2-chloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-(2,5-difluorophenyl)acetamide (14B)
Blue black; yield 71%; mp 204 ꢀC; Anal. Calc. for C21H15ClN4O2S:
12.34; IR n
(cmꢁ1): 3095 (Ar,CeH), 2964 (CH3),1655,1642 (CO),1435
C 54.73, H 3.28, N 12.61, found C 54.69, H 3.31, N 12.58; IR
n
(cmꢁ1):
(C]N); 1H NMR (CDCl3)
d(ppm): 2.68 (s, 6H, CH3), 2.73 (s, 6H, CH3),
3072 (Ar,CeH), 2909 (CH3), 1678, 1664 (CO), 1435 (C]N); 1H NMR
5.37 (s, 1H, SCHN), 7.12e7.44 (m, 8H, Ar-H); ESI-MS m/z: [Mþ þ Na]
(DMSO)
d
(ppm): 2.58 (s, 6H, CH3), 5.43 (s, 1H, SCHN), 7.89e8.61 (m,
475.7
8H, Ar-H); ESI-MS m/z: [Mþ þ H] 461.5
4.3.22. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadiazole-2-yl)-N-(2,5-difluorophenyl)acetamide
(22B)
4.3.15. N-4-acetyl-5-(2-chloroquidnolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-p-tolylacetamide (15B)
Light pink; yield 68%; mp 207 ꢀC; Anal. Calc. for C22H19ClN4O2S:
Blackish; yield 58%; mp 198 ꢀC; Anal. Calc. for C22H17ClF2N4O2S:
C 60.20, H 4.36 N 12.76, found C 60.23, H 4.33, N 12.79; IR
n
(cmꢁ1):
C 55.64, H 3.61, N 11.80, found C 55.69, H 3.59, N 11.82; IR
n
(cmꢁ1):
3056 (Ar,CeH), 2926 (CH3), 1678,1640 (CO), 1445 (C]N); 1H NMR
3075 (Ar,CeH), 2934 (CH3), 1664, 1640 (CO), 1435 (C]N); 1H NMR
(CDCl3)
d
(ppm): 7.29e7.64 (m, 9H, Ar-H), 5.19 (s, 1H, SCHN), 2.36 (s,
(DMSO) d(ppm): 11.71(s, 1H, NNH), 7.76e7.98 (m, 7H, Ar-H), 7.17 (s,
3H, CH3), 2.19 (s, 6H, CH3),; ESI-MS m/z: [Mþ þ H] 484.7
1H, CH]N), 2.64 (s, 3H, CH3); ESI-MS m/z: [Mþ þ H] 475.8
4.3.16. N-4-acetyl-5-(2-chloroquinolin-3-yl)-4,5-dihydro-1,3,4-
thiadiazole-2-yl)-N-(4-nitophenyl)aceamide (16B)
4.3.23. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadiazole-2-yl)-N-p-phenylacetamide (23B)
White; yield 64%; mp 201 ꢀC; Anal. Calc. for C23H21ClN4O2S:
C 60.99, H 4.67, N 12.37, found C 61.02, H 4.65 N 12.41; IR
Blackish; yield 65%; mp 204 ꢀC.; Anal. Calc. for C22H16ClN5O4S:
C 60.20, H 4.36, N 12.76, found C 60.16, H 4.39, N 12.71; IR
n
(cmꢁ1):
3027 (Ar,CeH), 2952 (CH3), 1671,1658 (CO), 1449 (C]N); 1H NMR
n
(cmꢁ1): 3062 (Ar,CeH), 2911 (CH3), 1678, 1610 (CO), 1445 (C]
(DMSO)
d
(ppm): 7.51e8.09 (m, 9H, Ar-H), 5.12 (s, 1H, SCHN), 2.49 (s,
N); 1H NMR (CDCl3)
d(ppm): 7.23e7.61 (m, 8H, Ar-H), 5.61 (s, 1H,
6H, CH3); ESI-MS m/z: [Mþ þ Na] 492.5
SCHN), 2.69 (s, 6H, CH3), 2.34 (s, 6H, CH3); 13C NMR (CDCl3)
d/
ppm: 174.36 (C]O), 168.95 (C]O), 153.48 (C]N), 149.95 (C]N),
141.49, 139.10,135.08, 130.65, 128.76, 125.91, 124.34, 122.11
(Ar-C), 49.02 (SCN), 24.03, 22.01, 21.24 (aliphatic); ESI-MS m/z:
[Mþ þ H] 453.8
4.3.17. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-dihydro-
1,3,4-thiadiazole-2-yl)acetamide (17B)
Yellowish; yield 63%; mp 213 ꢀC Anal. Calc. for C16H15ClN4O2S:
C 52.96, H 4.17, N 15.44, found C 53.01, H 4.13, N 15.48; IR
3194 (NH), 2961 (CH3), 1658, 1645 (CO), 1479 (C]N); 1H NMR
(CDCl3) (ppm): 8.13 (s, 1H, Ar-H), 7.39e7.69 (m, 3H, Ar-H), 5.28 (s,
n
(cmꢁ1):
4.3.24. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadiazole-2-yl)-N-(4-nitrophenyl)acetamide (24B)
Creamy white; yield 55%; mp 207 ꢀC; Anal. Calc. for
C22H18ClN5O2S: C 54.80, H 3.75, N 14.47, found C 54.74, H 3.78,
d
1H, N]CH), 2.65 (s, 6H, CH3), 2.48 (s, 3H, CH3), 2.19 (s, 1H, NH2);
ESI-MS m/z: [Mþ þ H] 363.5
N 14.51; IR
1449 (C]N); 1H NMR (DMSO)
4.93 (s, 1H, SCHN), 2.51 (s, 3H, CH3), 2.24 (s, 6H, CH3); 13C NMR
(DMSO) /ppm: 172.36 (C]O), 175.39 (C]O), 156.64 (C]N), 151.37
(C]N), 140.19, 139.74, 136.38, 133.27, 131.24, 129.84, 128.68, 124.84,
n
(cmꢁ1): 3054 (Ar,CeH), 2952 (CH3), 1640, 1604 (CO),
4.3.18. N-4-acetyl-5-(2-chloro-8-methylquinolin-3-yl)-4,5-
dihydro-1,3,4-thiadiazole-2-yl)-N-cyclpentylacetamide (18B)
Dark brown; yield 58%; mp 209 ꢀC; Anal. Calc. for
C21H23ClN4O2S: C 58.53, H 5.38, N 13.00, found C 58.56, H 5.34, N
d(ppm): 7.19e7.39 (m, 8H, Ar-H),
d

3166
A.R. Bhat et al. / European Journal of Medicinal Chemistry 46 (2011) 3158e3166
(Ar-C), 47.84 (SCN), 26.94, 25.72, 23.19, 21.96 (aliphatic carbon);
defined as the relative absorbance of treated versus untreated
control cells.
ESI-MS m/z: [Mþ þ H] 484.9
4.4. Biological activity
Acknowledgment
Organism culture and in vitro screening for antibacterial activity
was done by the disk diffusion method with minor modifications.
S. aureus, S. pyogenes, S. typhimurium, and E. coli were subcultured
in BHI medium and incubated for 18 h at 37 ꢀC. Following the
incubation the bacterial cells were suspended, according to the
McFarland protocol in saline solution to produce a suspension of
about 105 CFU/mL. 10 mL of this suspension was mixed with 10 mL
of sterile antibiotic agar at 40 ꢀC and poured on to an agar plate in
a laminar flow cabinet. Five paper disks (6.0 mm diameter) were
fixed onto nutrient agar plate. One milligram of each test
compound was dissolved in 100 mL DMSO to prepare stock solu-
tion. From the stock solution different dilutions of each test
compound were prepared and poured over disk plate. Amoxicillin
was used as a standard drug (positive control). DMSO poured disk
was used as negative control. The susceptibility of the bacteria to
the test compounds was determined by the formation of an
inhibitory zone after 18 h of incubation at 36 ꢀC. Tables 2 and 3
report the inhibition zones (mm) of thiosemicarbazones
(1Ae24A) and thiadiazoles (1Be24B) respectively. The results were
compared with the negative and positive controls and the zone of
inhibitions was measured at the minimum inhibitory
concentration.
This work was supported by the Department of Science and
Technology (Grant no. SR/FT/LS-069/2007). The authors are also
thankful to the Higher Education Department, Jammu and Kashmir,
for their cooperation.
References
[1] K. Ehlert, Curr. Pharm. Des 5 (1999) 45e55.
[2] K.R. Babu, B. Eeshwaraiah, D. Aravind, H.M. Meshram, R.M. Raju, A. Bhattacharya,
R. Bandichhor, Monatshefte für Chemie 139 (2008) 179e181.
[3] A. Dalhoff, Infection 22 (1994) S111eS121.
[4] C. Mohan, G. Bhargava, PM.S. Bedi, J. Life Sci. 1 (2009) 97e101.
[5] P. Zhan, X. Liu, Y. Cao, Y. Wang, C. Pannecouque, E.D. Clercq, Bioorg. Med.
Chem. Lett. 18 (2008) 5368e5371.
[6] M.D. Mullican, M.W. Wilson, D.T. Conner, C.R. Kosfan, D.J. Schrier, R.D. Dyer,
J. Med.Chem. 36 (1993) 1090e1099.
[7] Y. Song, D.T. Connor, R. Doubleday, R.J. Sorenson, A.D. Sercel, P.C. Unangst,
B.D. Roth, R.B. Gilbertsen, K. Chan, D.J. Schrier, A. Guglietta, D.A. Bornemeier,
R.D. Dyer, J. Med. Chem. 42 (1999) 1161e1169.
[8] L. Labanauskas, V. Kalcas, E. drenaite, P. Gaidelis, A. Brukstus, A. Dauksas,
Pharmazie 56 (2001) 617e619.
[9] J.Y. Chou, S.Y. Lai, S.L. Pan, G.M. Jow, J.W. Chern, J.H. Guh, Biochem. Pharmacol.
66 (2003) 115e124.
[10] H.K. Shucla, N.C. Desai, R.R. Astik, K.A. Thaker, J. Indian Chem. Soc. 61 (1984)
168e171.
[11] E.E. Orucu, S. Rollas, F. Kandermirli, N. Shvets, A.S. Dimoglo, J. Med. Chem. 47
(2004) 6760e6767.
The minimum inhibitory concentration (MIC) was evaluated by
the macro-dilution test using standard inoculums of 105 CFL/mL.
Serial dilutions of the test compounds, previously dissolved in
dimethyl sulfoxide (DMSO) were prepared to final concentrations of
[12] C.B. Chapleo, M. Myers, P.L. Myers, J.F. Saville, A.C.B. Smith, M.R. Stillings,
I.F. Tulloch, D.S. Walter, A.P. Welbourn, J. Med. Chem. 29 (1986)
2273e2280.
[13] S. Turner, M. Myers, B. Gadie, A.J. Nelson, R. Pape, J.F. Saville, J.C. Doxey,
T.L. Berridge, J. Med. Chem. 31 (1988) 902e906.
[14] S. Turner, M. Myers, B. Gadie, S.A. Hale, A. Horsley, A.J. Nelson, R. Pape,
J.F. Saville, J.C. Doxey, T.L. Berridge, J. Med. Chem. 31 (1988) 907e913.
[15] F. Poorrajab, S.K. Ardestani, A. Foroumadi, S. Emami, A. Kariminia,
M.B. Fardmoghadam, A. Shafiee, Exp. Parasitol. 121 (2009) 323e330.
[16] S. KaraKusu, S. Rollas, Il Farmaco 57 (2002) 577e581.
[17] Y.L. Chen, K.C. Fang, J.Y. Sheu, S.L. Hsu, C.C. Tzeng, J. Med. Chem. 44 (2001)
2374e2377.
512, 256, 128, 64, 32, 16, 8, 4, 2 and 1 mgm/L. To each tube was added
100 mL of a 24 h old inoculum. The MIC, defined as the lowest
concentration of the test compound, which inhibits the visible
growth after 18 h, was determined visually after incubation for 18 h,
at 37 ꢀC, and the results are presented in Tables 4 and 5. DMSO and
amoxicillin were used as negative and positive controls.
[18] B.A. Cunha, Crit. Care Clin. 14 (1998) 309e327.
[19] C.Y. Hong, Y.K. Kim, J.H. Chang, S.H. Kim, D.H. Choi, Y.Z. Kim, J.H. Kwak, J. Med.
Chem. 40 (1997) 3584e3593.
[20] K. Hirai, M. Hosaka, Y. Niwata, T. Yasue, H. Fukuda, T. Ishizaki, S. Suzue, K.
Nishino, Paper presented at 30th Meeting of the ICAAC, Atlanta, GA, October
1990, pp. 21e24.
[21] P.L. Ho, R. Yung, D.C. Tsang, T.L. Que, W.H. Ho, T.K. Seto, W.C. Ng, Yam, W.S. Ng,
J. Antimicrob. Chemother. 48 (2000) 659e665.
[22] K. Weiss, C. Restieri, R. Gauthier, M. Laverdière, A. McGeer, Clin. Infect. Dis. 33
(2001) 517e522.
[23] A.R. Bhat, F. Athar, A. Azam, Helv. Chem. Acta 92 (2009) 1644e1655.
[24] A.R. Bhat, F. Athar, R.L. Van Zyl, C.T. Chen, A. Azam, Chem. Biod 5 (2008) 764e776.
[25] I. Irfan, S. Nongyao, D. Bahl, A.R. Bhat, A. Azam, Eur. J. Med. Chem. 45 (2010)
1648e1653.
[26] V.S. Palekar, A.J. Damle, S.R. Shukla, Eur. J. Med. Chem. 44 (2009) 5112e5116.
[27] M. Amir, M.S.Y. Khan, M.S. Zaman, Indian J. Chem. 43B (2004) 2189.
[28] B. Tozcoparan, E. Kupeli, E. Yesilada, M. Ertan, Bioorg. Med. Chem. 15 (2007) 1808.
[29] A.W. Bauer, W.M.M. Kirby, J.C. Sherris, M. Turck, Am. J. Clin. Path 45 (1966)
493e496.
4.5. Cytotoxicity profile
MTT Assay: H9c2 rat cardiac myoblasts were cultured and
maintained as monolayer in Dulbecco’s modified Eagle’s medium
(DMEM), high glucose, supplemented with 10% fetal bovine serum,
100 units/mL penicillin, 100 mg/mL streptomycin, and 2.5 mg/mL
amphotericin B, at 37 ꢀC in humidified incubator with 5% CO2 [31].
Cells were incubated with different concentrations of compounds
5B, 8B, 10B, 13B,14B, 15B, 21B, 22B and amoxicillin for 48 h at 37 ꢀC
in 5% CO2 humidified incubator together with untreated control
sample. At appropriate time points, cells were washed in PBS,
treated with 600 mL MTT solution (5 mg/mL, tetrazolium salt) and
incubated for 45 min at 37 ꢀC. After 45 min of incubation at 37 ꢀC,
the cell supernatants were discarded, MTT crystals were dissolved
with acid isopropanol and the absorbance measured at 570 nm.
All assays were performed in triplicate. Percent viability was
[30] D.G. O’Sullivan, P.W. Sadler, C. Webley, Chem. Commun. 7 (1963) 17e26.
[31] M.K. Gupta, T.V. Neelakantan, M. Sanghamitra, R.K. Tyagi, A. Dinda, S. Mualik,
C.K. Mukhopadhyay, S.K. Goswami, Antioxid. Redox Signal.
8 (2006)
1081e1093.