Synthesis and antimicrobial activity of novel quinoline derivatives bearing pyrazoline and pyridine analogues

Article abstract of DOI:10.1007/s00044-016-1732-6

The present investigation is in the interest of some synthesized novel derivatives containing (5-(2-chloroquinolin-3-yl)-3-(aryl)-4,5-dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanones (4a–o) moieties incorporated with different biological active heterocycles such as quinoline, pyrazoline and pyridine derivatives. For the determination of the compounds reported in this paper was based on IR, ¹H NMR, ¹³C NMR and mass spectral data and same compounds were screened for their antibacterial and antifungal activity on four bacteria (Staphylococcus aureus, Streptococcus pyogenes, Escherichia coli, Pseudomonas aeruginosa) and three fungi (Candida albicans, Aspergillus niger, Aspergillus clavatus) using ampicillin and griseofulvin as the standard drugs. Cytotoxicity study was carried out using MTT colorimetric assay (HeLa cell line). Among the screened compounds, 4e, 4f and 4n showed most potent antibacterial activity, while compounds 4d and 4g emerged as the most active against fungal strains. The results demonstrated that compound 4o was remarkably active against all microbial strains. From the viewpoint of SAR studies, it was observed that the presence of electron withdrawing groups remarkably enhanced the antimicrobial activity of synthesized compounds. Additionally, preliminary MTT cytotoxicity studies on HeLa cells suggested that effective antimicrobial activity of 4e–g, 4n and 4o was accompanied by low cytotoxicity.

Full text of DOI:10.1007/s00044-016-1732-6

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-016-1732-6
ORIGINAL RESEARCH
Synthesis and antimicrobial activity of novel quinoline derivatives
bearing pyrazoline and pyridine analogues
Nisheeth C. Desai1 Bonny Y. Patel1 Bharti P. Dave2
●
●
Received: 4 May 2016 / Accepted: 11 October 2016
© Springer Science+Business Media New York 2016
Abstract The present investigation is in the interest of
some synthesized novel derivatives containing (5-(2-chlor-
oquinolin-3-yl)-3-(aryl)-4,5-dihydro-1H-pyrazol-1-yl)(pyr-
idin-4-yl)methanones (4a–o) moieties incorporated with
different biological active heterocycles such as quinoline,
pyrazoline and pyridine derivatives. For the determination
of the compounds reported in this paper was based on IR,
1H NMR, 13C NMR and mass spectral data and same
compounds were screened for their antibacterial and anti-
fungal activity on four bacteria (Staphylococcus aureus,
Streptococcus pyogenes, Escherichia coli, Pseudomonas
aeruginosa) and three fungi (Candida albicans, Aspergillus
niger, Aspergillus clavatus) using ampicillin and griseo-
fulvin as the standard drugs. Cytotoxicity study was carried
out using MTT colorimetric assay (HeLa cell line). Among
the screened compounds, 4e, 4f and 4n showed most potent
antibacterial activity, while compounds 4d and 4g emerged
as the most active against fungal strains. The results
demonstrated that compound 4o was remarkably active
against all microbial strains. From the viewpoint of SAR
studies, it was observed that the presence of electron
withdrawing groups remarkably enhanced the antimicrobial
activity of synthesized compounds. Additionally, pre-
liminary MTT cytotoxicity studies on HeLa cells suggested
that effective antimicrobial activity of 4e–g, 4n and 4o was
accompanied by low cytotoxicity.
●
●
Keywords Pyridine Pyrazoline 2-Chloroquinoline-3-
●
●
carbaldehyde Antimicrobial evaluation Cytotoxicity
Abbreviations
Anti-HIV Anti-human immunodeficiency virus
NCE’S
PDGF-
RTK
New chemical entity’s
Platelet-derived growth factor receptors tyrosine
kinases
IR
Infrared
NMR
DMF
DMSO
Nuclear magnetic resonance
Dimethylformamide
Dimethyl sulfoxide
Introduction
A wide-ranging array of microbes including bacteria, viru-
ses and fungi are becoming resistant to the standard mar-
keted drugs used to treat infections. This resistance is a
major hurdle for the treatment of highly infectious diseases
globally. The World Health Organization (WHO) has con-
sidered this antimicrobial drug resistance and the dwindling
number of active antimicrobial drugs to be one of the
greatest threats to human health. Moreover, problems of
multi-drug resistance of bacteria and fungi such as
Methicillin-resistant Staphylococcus aureus (MRSA),
Vancomycin-resistant Staphylococcus aureus (VRSA),
Vancomycin resistant enterococcus (VRE) and Fluconazole
Electronic supplementary material The online version of this article
(doi:10.1007/s00044-016-1732-6) contains supplementary material,
which is available to authorized users.
* Nisheeth C. Desai
dnisheeth@rediffmail.com
1
Division of Medicinal Chemistry, Department of Chemistry,
(DST-FIST Sponsored & UGC NON-SAP), Mahatma Gandhi
Campus, Maharaja Krishnakumarsinhji Bhavnagar University,
Bhavnagar 364002, India
2
Department of Life Sciences, Maharaja Krishnakumarsinhji
Bhavnagar University, Bhavnagar 364002, India

Med Chem Res
Fig. 1 Chemical remodeling of
existing antibacterial drugs from
potent molecules either in phase
II or phase III of clinical trials
resistant Candida have reached an alarming level becoming
a serious medical problem worldwide (Mishra et al. 2013;
Cetinkaya et al. 2000; Gulshan and Moye-Rowley 2007).
Therefore, it is necessary to develop antimicrobial agents
with improved potency and the combat is still on to discover
novel and potent antimicrobial agents for medicinal che-
mists (Moellering 2011). The structural modifications in
existing drugs have shown astounding results in the field of
drug discovery. Newly investigational drugs or trial drugs
which utilize this strategy are safer with a broader spectrum
of activity. Figure 1 represents potent molecules either in
phase II or phase III of clinical trials from the existing
NCE’S (Butler et al. 2013).
Searching for structure with significant bioactivity, we
focused onto the development of molecules through combi-
nation of different active pharmacophores like quinolines,
pyrazoline and pyridine into one core structure. That may
lead to compounds with improved antimicrobial activity.
Among the important heterocyclic moieties of biological and
pharmacological attention, the quinoline ring is endowed
with various activities such as antimicrobial (Desai et al.
2014), antiplasmodial (Vandekerckhove et al. 2015), anti-
tuberculosis (Keri and Patil 2014), anti-inflammatory
(El-Feky et al. 2015), antimalarial (Vandekerckhove and
D’hooghe 2015), anticancer (Spanò et al. 2015), antioxidant
(Vivekanand et al. 2015), and anti-HIV (Ahmed et al. 2010).
Furthermore, quinolines are used as an inhibitor in tyrokinase
PDGF-RTK inhibiting agents (Maguire et al. 1994), SH2-
containing inositol 5′-Phosphatase (SHIP) (Russo et al. 2015)
and Mycobacterium tuberculosis DNA gyraseB inhibitors
(Medapi et al. 2015).
2-Pyrazoline derivatives have been reported to exhibit
various pharmacological activities such as antimicrobial
(Karthikeyan et al. 2007), anti-inflammatory (Barsoum et al.
2006), antihypertensive, anti-tumor (Lin et al. 2007),
anticancer (Gowramma et al. 2009) and anticonvulsant
(Ozdemir et al. 2007). In addition, pyrazolines are also
reported to possess cytotoxic properties against human lung
tumor cell line (A549) (Greenlee et al. 2001). Nowadays
some newly steroidal pyrazoline are also synthesized for
finding a novel drug molecule (Shamsuzzaman and Mohd
2012). Pyridine is a versatile bioactive heterocycle having
its wide presence in many synthetic drugs such as rosigli-
tazone (antidiabetic), pioglitazone (hypoglycemic),
lansoprazole (proton-pump inhibitor), and etoricoxib (COX-
2 selective inhibitor). The compounds containing pyridine
scaffold exhibit anticancer (Kadayat et al. 2015), anti-
inflammatory, anti-ulcer (Mohareb et al. 2015), anti-
mycobacterial (Jose et al. 2015b) and antimicrobial (Jose
et al. 2015a) activities.
Prompted by the above-mentioned observations, we have
focused onto construct some new antimicrobial derivatives
bearing quinoline, pyrazoline and pyridine scaffolds and
study the structure activity relationship due to substituent
variations. Earlier, our research group had synthesized
quinoline and other bioactive derivatives as potential anti-
microbial agents (Desai et al. 2013a, b, 2015a, b). In con-
tinuation to this, (5-(2-chloroquinolin-3-yl)-3-(aryl)-4,5-
dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanones (4a–o)
have been synthesized and screened for their antimicrobial
activity. In addition, cytotoxicity studies were also con-
ducted on HeLa cell lines to assess the ability of these
compounds to inhibit the cell growth. The newly synthe-
sized compounds (4a–o) were elucidated by IR, NMR and
mass spectroscopy for their structural characterization.
Results and discussion
Chemistry
Synthetic strategies adopted to achieve the target com-
pounds are depicted in Scheme 1. Here, scaffold 4 is a part
of synthesis of new chemical entities in the form of anti-
microbial agents. According to Scheme 1, the key chalcone
derivatives 3-(2-chloroquinolin-3-yl)-1-(aryl)prop-2-en-1-
ones (3a–o) are used as precursors for the synthesis of title
compounds (4a–o). Compounds (3a–o) were synthesized
through the Claisen–Schmidt condensation of equimolar
amounts of different acetophenone derivatives (2a–o) and
compound 2-chloroquinoline-3-carbaldehyde 1 by stirring
the reactants in aqueous alcoholic solution containing
sodium hydroxide at room temperature. Compound 1 is
Vilsmeier–Haack reactions adduct; it provides a vital and
efficient intermediate for the synthesis of several new sub-
stituted heterocyclic analogs. Chalcone compounds (3a–o)
on cyclocondensation with isoniazid yielded final com-
pounds (5-(2-chloroquinolin-3-yl)-3-aryl-4,5-dihydro-1H-

Med Chem Res
Scheme 1 Synthetic pathway of
novel compounds 4a–o
pyrazol-1-yl)(pyridin-4-yl)methanones (4a–o). Further it
has been observed that compounds (4a–o) have pyrazoline
core nucleus and were generated through cyclization,
migration of α, β unsaturated double bonds, and diminish-
ing of carbonyl group.
carbon of methylene group present in pyrazoline nucleus
and methine carbon showed a chemical shift at δ =
60.0–66.6 ppm. Furthermore, the mass spectrum of (4a–o)
showed a molecular ion peak corresponding to molecular
formula (4a–o) along with of other fragment peaks, which
supported the proposed structure of compounds. The detail
discussion of characterization data are given in experi-
mental section. The structure and carbon numbering of
compound (4a–o) as an example are described in Fig. 2.
Designed series of molecules (4a–o) were characterized
1
by IR, H NMR, 13C NMR, and mass spectrometry tech-
niques. IR spectrums of title compounds (4a–o) gave
stretching vibrations at 3035–3059 cm−1 due to aromatic
C–H stretching vibrations corresponds to methylene group
that appears over the range at 2872–2926 cm−1. The strong
intensity absorption bands at 1645–1696 cm−1 is due to
stretching vibrations of C=O group. The characteristic
Discussion of antibacterial screening
Amongst synthesized compounds (4a–o), several com-
pounds exhibited antimicrobial potency that diverged from
good to excellent. On the basis of antibacterial screening
results from Table 1, compound 4o (-2,4-(Cl)2) displayed
good activity against E. coli, while compounds 4f (-4-F) and
4n (-4-NO2) possessed very good activity against E. coli at
50 µg ml−1 MIC. It is noteworthy that compound 4e (-4-Cl)
showed highest inhibition at MIC = 12.5 µg ml−1 against the
same Gram-negative bacterium, i.e. E. coli. In the case of P.
aeruginosa, compound 4o (-2,4-(Cl)2) showed same inhi-
bition as to the standard drug. While compounds 4e (-4-Cl)
and 4n (-4-NO2) inhibited bacteria at MIC = 50 µg ml−1
(very good activity) and maximum inhibition (MIC = 12.5
µg ml−1) was shown by compound 4f (-4-F) against P.
aeruginosa. Moreover, compound 4e (-4-Cl) displayed
good activity and compounds 4f (-4-F) and 4n (-4-NO2)
showed very good activity at 250 and 100 µg ml−1 MIC
1
signals in H NMR of compounds (4a–o) were of three
pyrazoline protons which displayed doublet of doublet. Out
of three, two pyrazoline protons (C-13) Ha and Hb give AB
system. Part one of AB system displayed a signal at δ =
3.29–3.40 ppm as doublet of doublet with coupling con-
stants 17.42–17.55 and 3.04–3.10 Hz and part two of AB
system appeared as doublet of doublet at δ = 3.74–3.82 ppm
with coupling constants 17.40–17.52 and 11.01–11.16 Hz.
Proton Hc of pyrazoline (C-11, methine proton) also
appeared as doublet of doublet at δ = 6.02–6.09 ppm
with coupling constants in range of 11.08–11.22 and
3.08–3.17 Hz. Moreover, 13C NMR confirmed the final
compounds structure by the appearance of signals at
δ = 171.2–172.9 ppm due to carbonyl carbon of isoniazid
directly attached to pyrazoline nitrogen. Compounds (4a–o)
showed signal at δ = 39.1–39.9 ppm corresponding to

Med Chem Res
values against S. aureus. Compound 4o (-2,4-(Cl)2)
exhibited inhibition at MIC = 12.5 µg ml−1 against Gram-
positive bacterium, i.e. S. aureus. In case of Gram-negative
bacterium (S. pyogenes), compounds 4e (-4-Cl) and 4o
(-2,4-(Cl)2) exhibited good activity at MIC = 50 µg ml−1
and compound 4f (-4-F) displayed very good activity
against S. pyogenes. Compound 4n (-4-NO2) had shown
MIC 12.5 µg ml−1 which observed as the highest inhibition
against S. pyogenes. The remaining compounds of the series
possessed feeble antibacterial activity as shown in Table 1.
Tabular data revealed that the presence of functional group
at para position improved antibacterial activity as compared
to ortho and meta substituted compounds.
Discussion of antifungal screening
Minimum inhibitory concentration (MIC) values of anti-
fungal activity spotted against Candida albicans, Aspergillus
niger and Aspergillus clavatus were by conventional broth
micro dilution method. We have discussed and compared
antifungal activities based on standard drug Griseofulvin
shown in Table 1. Compounds 4m (-3-NO2), 4n (-4-NO2)
and 4o (-2,4-(Cl)2) exhibited same MIC value as standard
drug against C. albicans. Excellent and very good activity
possessed against C. albicans by compounds 4d (-2-Cl) and
4g (-4-Br) at MIC = 50 and 100 µg ml−1, respectively. Same
compounds 4d and 4g also displayed very good activity
against A. niger, while highest inhibition (12.5 µg ml−1)
exists in di-chloro substituted compounds, i.e. 4o (-2,4-
(Cl)2). Furthermore, compounds 4g and 4o having
Fig. 2 Carbon numbering of the final compounds 4a–o
Table 1 Antimicrobial
Sr. No.
-R
Minimum inhibitory concentrations
screening of the compounds
4a–o
For bacteria (MIC) in µg ml−1
Gram-negative Gram-positive
For fungi (MIC) in µg ml−1
Fungi
E.c.
P.a.
S.a.
S.p.
C.a.
A.n.
A.c.
4a
–H
250
500
250
100
12.5
50
500
250
250
100
50
500
500
250
>1000
250
100
50
1000
1000
1000
50
500
500
250
50
1000
250
500
12.5
250
250
50
4b
-2-CH3
-4-CH3
-2-Cl
>1000
500
4c
4d
250
4e
-4-Cl
250
>1000
1000
100
250
250
50
4f
-4-F
12.5
250
1000
500
250
250
250
250
50
100
4g
-4-Br
250
500
>1000
250
250
500
250
50
250
100
250
500
500
>1000
250
100
12.5
100
100
–
4h
-2-OCH3
-4-OCH3
-2-OH
-3-OH
-4-OH
-3-NO2
-4-NO2
-2,4-(Cl)2
500
1000
>1000
>1000
1000
1000
500
500
250
250
>1000
500
250
250
12.5
–
250
500
1000
500
250
250
1000
50
4i
1000
1000
500
4j
4k
4l
>1000
500
4m
4n
100
500
4o
100
100
–
100
100
–
12.5
250
500
Ampicillin
Griseofulvin
–
–
–
500
100
100
E.c. Escherichia coli MTCC 443; P.a. Pseudomonas aeruginosa MTCC 1688; S.a. Staphylococcus aureus
MTCC 96; S.p. Staphylococcus pyogenes MTCC 442; C.a. Candida albicans MTCC 227; A.n. Aspergillus
niger MTCC 282; A.c. Aspergillus clavatus MTCC 1323
Bold values indicate the most active compounds

Med Chem Res
Table 2 Levels of cytotoxicity prompted by compounds 4a–o on
HeLa cells
purity of all compounds was checked on aluminum-coated
TLC plates 60, F245 (E. Merck) using various solvent sys-
tems as mobile phase and visualized under ultraviolet (UV)
light, or iodine vapor. Perkin-Elmer 2400 CHN analyzer
was used for elemental analysis (% C, H, N). IR spectra
were recorded on Perkin Elmer FT-IR spectrophotometer.
1H NMR spectra were recorded on a Bruker Avance II 400
MHz while 13C NMR spectra on Varian Mercury-400, 100
MHz in CDCl3 as a solvent and tetramethylsilane (TMS) as
an internal standard using 5 mm tube. Chemical shifts were
reported in ppm units with use of δ scale. Mass spectra were
also scanned on a Shimadzu LCMS 2010 spectrometer. The
essential chemicals were purchased from E. Merck. Buchi
Rota vapor instrument was used for distillation purpose.
Sr. No.
Cytotoxicity
(IC50 µM)a
HeLab
4a
60.74
81.14
78.16
87.53
>100
>100
>100
69.09
58.74
88.84
66.34
75.98
58.65
>100
>100
3.24
4b
4c
4d
4e
4f
4g
4h
4i
4j
Preparation of 2-chloroquinoline-3-carbaldehyde (1)
4k
4l
To a solution of acetanilide (5 mmol) in dry DMF (15
mmol) at 0–5 °C with stirring phosphoryl chloride (POCl3)
(60 mmol) was added drop-wise and the mixture stirred at
100 °C for 16–20 h. The mixture was poured into crushed
ice, stirred for 5 min and the resulting solid filtered, washed
well with water and dried. The yellow crystals formed were
filtered, washed with water and crystallized using ethyl
acetate (Meth-Cohn 1993). 1H NMR (CDCl3, 400 MHz): δ
= 7.59 (t, 1H, Ar–H), 7.69 (t, 1H, Ar–H), 8.01 (d, 1H,
Ar–H), 8.17 (d, 1H, Ar–H), 8.55 (s, 1H, Ar–H), 9.61 (s, 1H,
Ar–H); anal. calcd. for C10H6ClNO: C, 62.68; H, 3.16; N,
7.31; found: C, 62.73; H, 3.24; N, 7.36.
4m
4n
4o
Doxorubicin
a
IC50 is the concentration required to inhibit 50 % of cell growth
HeLa human cervical cancer cell line
b
-4-Br, -2,4-(Cl)2 functional groups respectively, possessed
very good activity against A. clavatus, while excellent
activity against A. clavatus was demonstrated by -2-Cl (4d)
derivative. Compound 4o having electron withdrawing
groups in structure proves its significance to almost all
bacterial and fungal strains that makes a great imaginary
impact in antimicrobial activity.
General procedure of 3-(2-chloroquinolin-3-yl)-1-(aryl)
prop-2-en-1-ones (3a–o)
Discussion of in vitro cytotoxicity studies
Compounds 3-(2-chloroquinolin-3-yl)-1-(aryl)prop-2-en-1-
ones (3a–o) were prepared according to the literature
method (Elgazwy 2008). All other derivatives were pre-
pared by the same method and monitored by thin layer
In vitro cytotoxic activity of newly synthesized compounds
(4a–o) was evaluated against human cervical cancer cell line
(HeLa) by the MTT colorimetric assay (Mosmann 1983),
which measures the reduction of tetrazolium bromide salt
into a formazan dye by mitochondrial dehydrogenases in
treated vs. untreated cells. The IC50 values achieved for
these compounds were shown in Table 2. Cytotoxicity
results displayed that the derivatives 4e (4-Cl), 4f (4-F), 4g
(4-Br), 4n (4-NO2) and 4o (2,4-(Cl)2) accounted no toxicity
at concentration of 100 µg/ml (IC50 > 100 µg/ml), while
other derivatives exhibited moderate toxicity against HeLa
cell lines.
1
chromatography (TLC). H NMR (CDCl3, 400 MHz): δ =
7.31–7.74 (m, 7H, Ar–H), 7.78 (d, 1H), 7.97 (d, 3H), 8.21
(s, 1H, quinoline-H4); anal. calcd. for C18H12ClNO: C,
73.60; H, 4.12; N, 4.77; found: C, 73.69; H, 4.19; N, 4.70.
General procedure of (5-(2-chloroquinolin-3-yl)-3-(aryl)-
4,5-dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanones
(4a–o)
A mixture of differently substituted quinolinyl chalcones
3a–o (0.01 mol) and isoniazid (0.02 mol) were taken in 20
ml glacial acetic acid and refluxed at 120 °C over a period
of 8 h. The mixture was concentrated under vacuum and
diluted with ice cold water. The separated solid was filtered,
dried, and crystallized from ethanol (95 %).
Experimental
Melting points were noted on Gallenkamp apparatus and
were left uncorrected. The completion of reaction and the

Med Chem Res
(5-(2-Chloroquinolin-3-yl)-3-phenyl-4,5-dihydro-1H-
(3-(2-Chlorophenyl)-5-(2-chloroquinolin-3-yl)-4,5-dihydro-
pyrazol-1-yl)(pyridin-4-yl)methanone (4a)
1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4d)
Yellow crystals; yield: 73 %; mp 118–120 °C; IR (KBr)
νmax 3064 (C–H, aromatic), 2871 (C–H stretching, –CH2-
group), 1692 (C=O stretching), 1579 (C=N), 1513 (C=C),
Light yellow crystals; yield: 69 %; mp 164–166 °C; IR
(KBr) νmax 3066 (C–H, aromatic), 2900 (C–H stretching,
–CH2-group), 1690 (C=O stretching), 1585 (C=N), 1520
(C=C), 745 (C–Cl stretching) cm−1; 1H NMR (CDCl3, 400
MHz): δ = 3.40 (dd, Jab = 17.45 Hz, Jac = 3.06 Hz, 1H, Ha),
3.78 (dd, Jab = 17.43 Hz, Jbc = 11.06 Hz, 1H, Hb), 6.08 (dd,
Jac = 3.11 Hz, Jbc = 11.12 Hz, 1H, Hc), 7.18–8.43 (m, 13H,
Ar–H); 13C NMR (CDCl3, 100 MHz): δ = 43.1 (C-12, CH2
of pyrazoline), 62.6 (C-11, CH–N of pyrazoline),
122.2–148.9 (Ar–C), 128.6 (C-15, Ar–C–Cl), 152.5 (C-8,
C–Cl of quinoline), 156.3 (C-13, C=N of pyrazoline), 167.4
(C-23, C=O); LCMS: m/z 446.07 [M+]; anal. calcd. for
C24H16Cl2N4O: C, 64.44; H, 3.61; N, 12.53; found: C,
64.49; H, 3.62; N, 12.58.
1
735 (C–Cl stretching) cm−1; H NMR (CDCl3, 400 MHz):
δ = 3.29 (dd, Jab = 17.45 Hz, Jac = 3.05 Hz, 1H, Ha), 3.78
(dd, Jab = 17.41 Hz, Jbc = 11.02 Hz, 1H, Hb), 6.06 (dd, Jac
= 3.09 Hz, Jbc = 11.08 Hz, 1H, Hc), 7.06–8.32 (m, 14H,
Ar–H); 13C NMR (CDCl3, 100 MHz): δ = 43.6 (C-12, CH2
of pyrazoline), 62.7 (C-11, CH–N of pyrazoline),
122.1–148.2 (Ar–C), 152.1 (C-8, C–Cl of quinoline), 156.6
(C-13, C=N of pyrazoline), 167.3 (C-23, C=O); LCMS: m/
z 412.11 [M+]; anal. calcd. for C24H17ClN4O: C, 69.82; H,
4.15; N, 13.57; Found: C, 69.86; H, 4.14; N, 13.66.
(5-(2-Chloroquinolin-3-yl)-3-(o-tolyl)-4,5-dihydro-1H-
pyrazol-1-yl)(pyridin-4-yl)methanone (4b)
(3-(4-Chlorophenyl)-5-(2-chloroquinolin-3-yl)-4,5-dihydro-
1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4e)
Cream crystals; yield: 67 %; mp 184–185 °C; IR (KBr) νmax
3065 (C–H, aromatic), 2922, 2875 (C–H stretching, –CH3
group, –CH2-group), 1690 (C=O stretching), 1584 (C=N),
1516 (C=C), 736 (C–Cl stretching) cm−1; 1H NMR (CDCl3,
400 MHz): δ = 2.30 (s, 3H, Ar–CH3), 3.38 (dd, Jab = 17.44
Yellow crystals; yield: 67 %; mp 122–124 °C; IR (KBr)
νmax 3059 (C–H, aromatic), 2926 (C–H stretching, –CH2-
group), 1645 (C=O stretching), 1587 (C=N), 1568 (C=C),
1
750 (C–Cl stretching) cm−1; H NMR (CDCl3, 400 MHz):
Hz, Jac = 3.07 Hz, 1H, Ha), 3.74 (dd, Jab = 17.43 Hz, Jbc
=
δ = 3.33 (dd, Jab = 17.46 Hz, Jac = 3.05 Hz, 1H, Ha), 3.77
(dd, Jab = 17.45 Hz, Jbc = 11.07 Hz, 1H, Hb), 6.04 (dd, Jac
= 3.10 Hz, Jbc = 11.13 Hz, 1H, Hc), 7.22–8.42 (m, 13H,
Ar–H); 13C NMR (CDCl3, 100 MHz): δ = 43.3 (C-12, CH2
of pyrazoline), 62.7 (C-11, CH–N of pyrazoline),
122.1–148.8 (Ar–C), 136.2 (C-17, Ar–C–Cl), 152.2 (C-8,
C–Cl of quinoline), 156.4 (C-13, C=N of pyrazoline), 167.2
(C-23, C=O); LCMS: m/z 446.07 [M+]; anal. calcd. for
C24H16Cl2N4O: C, 64.44; H, 3.61; N, 12.53; Found: C,
64.53; H, 3.69; N, 12.60.
11.04 Hz, 1H, Hb), 6.05 (dd, Jac = 3.10 Hz, Jbc = 11.09 Hz,
1H, Hc), 7.10–8.38 (m, 13H, Ar–H); 13C NMR (CDCl3,
100 MHz): δ = 20.1 (C-15, Ar–CH3), 43.3 (C-12, CH2 of
pyrazoline), 62.5 (C-11, CH–N of pyrazoline), 122.2–148.4
(Ar–C), 152.2 (C-8, C–Cl of quinoline), 156.7 (C-13, C=N
of pyrazoline), 167.4 (C-23, C=O); LCMS: m/z 426.12
[M+]; anal. calcd. for C25H19ClN4O: C, 70.34; H, 4.49; N,
13.12; found: C, 70.42; H, 4.46; N, 13.20.
(5-(2-Chloroquinolin-3-yl)-3-(p-tolyl)-4,5-dihydro-1H-
pyrazol-1-yl)(pyridin-4-yl)methanone (4c)
(5-(2-Chloroquinolin-3-yl)-3-(4-fluorophenyl)-4,5-dihydro-
1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4f)
Light cream crystals; yield: 65 %; mp 172–174 °C; IR
(KBr) νmax 3063 (C–H, aromatic), 2920, 2878 (C–H
stretching, –CH2-group, –CH3 group), 1670 (C=O stretch-
ing), 1582 (C=N), 1518 (C=C), 734 (C–Cl stretching)
cm−1; 1H NMR (CDCl3, 400 MHz): δ = 2.25 (s, 3H,
Ar–CH3), 3.30 (dd, Jab = 17.43 Hz, Jac = 3.06 Hz, 1H, Ha),
3.78 (dd, Jab = 17.43 Hz, Jbc = 11.03 Hz, 1H, Hb), 6.04 (dd,
Jac = 3.09 Hz, Jbc = 11.10 Hz, 1H, Hc), 7.10–8.32 (m, 13H,
Ar–H); 13C NMR (CDCl3, 100 MHz): δ = 21.5 (C-17,
Ar–CH3), 43.4 (C-12, CH2 of pyrazoline), 62.7 (C-11,
CH–N of pyrazoline), 122.0–148.6 (Ar–C), 152.1 (C-8,
C–Cl of quinoline), 156.5 (C-13, C=N of pyrazoline), 167.2
(C-23, C=O); LCMS: m/z 426.12 [M+]; anal. calcd. for
C25H19ClN4O: C, 70.34; H, 4.49; N, 13.12; found: C,
70.40; H, 4.42; N, 13.16.
Dark yellow crystals; yield: 71 %; mp 150–152 °C; IR
(KBr) νmax 3060 (C–H, aromatic), 2874 (C–H stretching,
–CH2-group), 1695 (C=O stretching), 1584 (C=N), 1528
(C=C), 740 (C–Cl stretching), 1152 (C–F stretching) cm−1;
1H NMR (CDCl3, 400 MHz): δ = 3.40 (dd, Jab = 17.48 Hz,
Jac = 3.08 Hz, 1H, Ha), 3.81 (dd, Jab = 17.42 Hz, Jbc
=
11.10 Hz, 1H, Hb), 6.02 (dd, Jac = 3.12 Hz, Jbc = 11.20 Hz,
1H, Hc), 7.13–8.30 (m, 13H, Ar–H); 13C NMR (CDCl3,
100 MHz): δ = 43.6 (C-12, CH2 of pyrazoline), 62.6 (C-11,
CH–N of pyrazoline), 121.9–147.5 (Ar–C), 152.3 (C-8,
C–Cl of quinoline), 156.8 (C-13, C=N of pyrazoline), 164.5
(C-17, Ar–C–F), 167.8 (C-23, C=O); LCMS: m/z 430.10
[M+]; anal. calcd. for C24H16ClFN4O: C, 66.90; H, 3.74; N,
13.00; found: C, 64.86; H, 3.65; N, 13.10.

Med Chem Res
(3-(4-Bromophenyl)-5-(2-chloroquinolin-3-yl)-4,5-dihydro-
z 442.12 [M+]; anal. calcd. for C25H19ClN4O2: C, 67.80; H,
1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4g)
4.32; N, 12.65; found: C, 67.83; H, 4.39; N, 12.62.
Gray crystals; yield: 65 %; mp 141–143 °C; IR (KBr) νmax
3064 (C–H, aromatic), 2879 (C–H stretching, –CH2-group),
1693 (C=O stretching), 1585 (C=N), 1530 (C=C), 734
(5-(2-Chloroquinolin-3-yl)-3-(2-hydroxyphenyl)-4,5-
dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4j)
1
(C–Cl stretching), 663 (C–Br stretching) cm−1; H NMR
Yellow crystals; yield: 62 %; mp 155–157 °C; IR (KBr)
νmax 3407 (–OH), 3062 (C–H, aromatic), 2875 (C–H
stretching, –CH2-group), 1695 (C=O stretching), 1586
(C=N), 1527 (C=C), 745 (C–Cl stretching) cm−1; 1H NMR
(CDCl3, 400 MHz): δ = 3.36 (dd, Jab = 17.48 Hz, Jac = 3.06
Hz, 1H, Ha), 3.77 (dd, Jab = 17.44 Hz, Jbc = 11.07 Hz, 1H,
Hb), 6.07 (dd, Jac = 3.10 Hz, Jbc = 11.11 Hz, 1H, Hc),
7.12–8.48 (m, 13H, Ar–H), 9.17 (s, 1H, OH); 13C NMR
(CDCl3, 100 MHz): δ = 43.8 (C-12, CH2 of pyrazoline),
62.3 (C-11, CH–N of pyrazoline), 121.7–148.5 (Ar–C),
152.5 (C-8, C–Cl of quinoline), 156.4 (C-13, C=N of
pyrazoline), 162.5 (C-15, Ar–C–OH), 167.2 (C-23, C=O);
LCMS: m/z 428.10 [M+]; anal. calcd. for C24H17ClN4O: C,
67.21; H, 4.00; N, 13.06; found: C, 67.29; H, 4.08; N,
13.10.
(CDCl3, 400 MHz): δ = 3.39 (dd, Jab = 17.50 Hz, Jac = 3.05
Hz, 1H, Ha), 3.77 (dd, Jab = 17.48 Hz, Jbc = 11.08 Hz, 1H,
Hb), 6.08 (dd, Jac = 3.14 Hz, Jbc = 11.22 Hz, 1H, Hc),
7.16–8.44 (m, 13H, Ar–H); 13C NMR (CDCl3, 100 MHz):
δ = 43.8 (C-12, CH2 of pyrazoline), 62.7 (C-11, CH–N of
pyrazoline), 122.1–147.5 (Ar–C), 126.2 (C-17, Ar–C–Br),
152.6 (C-8, C–Cl of quinoline), 157.0 (C-13, C=N of
pyrazoline), 167.5 (C-23, C=O); LCMS: m/z 490.02 [M+];
anal. calcd. for C24H16ClBrN4O: C, 58.62; H, 3.28; N,
11.39; found: C, 58.70; H, 3.19; N, 11.46.
(5-(2-Chloroquinolin-3-yl)-3-(2-methoxyphenyl)-4,5-
dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4h)
Light brown crystals; yield: 62 %; mp 195–197 °C; IR
(KBr) νmax 3066 (C–H, aromatic), 2937, 2879 (C–H
stretching, –OCH3 group, –CH2-group), 1692 (C=O
stretching), 1584 (C=N), 1531 (C=C), 736 (C–Cl stretch-
(5-(2-Chloroquinolin-3-yl)-3-(3-hydroxyphenyl)-4,5-
dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4k)
Dark yellow crystals; yield: 60 %; mp 219–221 °C; IR
(KBr) νmax 3410 (–OH), 3060 (C–H, aromatic), 2878 (C–H
stretching, –CH2-group), 1693 (C=O stretching), 1588
(C=N), 1524 (C=C), 740 (C–Cl stretching) cm−1; 1H NMR
(CDCl3, 400 MHz): δ = 3.37 (dd, Jab = 17.44 Hz, Jac = 3.05
Hz, 1H, Ha), 3.75 (dd, Jab = 17.46 Hz, Jbc = 11.09 Hz, 1H,
Hb), 6.08 (dd, Jac = 3.11 Hz, Jbc = 11.13 Hz, 1H, Hc),
7.14–8.49 (m, 13H, Ar–H), 9.07 (s, 1H, OH); 13C NMR
(CDCl3, 100 MHz): δ = 43.6 (C-12, CH2 of pyrazoline),
62.2 (C-11, CH–N of pyrazoline), 121.5–148.8 (Ar–C),
152.6 (C-8, C–Cl of quinoline), 156.6 (C-13, C=N of
pyrazoline), 160.2 (C-16, Ar–C–OH), 167.4 (C-23, C=O);
LCMS: m/z 428.10 [M+]; anal. calcd. for C24H17ClN4O: C,
67.21; H, 4.00; N, 13.06; found: C, 67.18; H, 4.08; N,
13.03.
ing) cm−1; 1H NMR (CDCl3, 400 MHz): δ = 3.35 (dd, Jab
=
17.54 Hz, Jac = 3.10 Hz, 1H, Ha), 3.67 (s, 3H, –OCH3), 3.80
(dd, Jab = 17.52 Hz, Jbc = 11.15 Hz, 1H, Hb), 6.03 (dd, Jac
= 3.17 Hz, Jbc = 11.20 Hz, 1H, Hc), 7.17–8.48 (m, 13H,
Ar–H); 13C NMR (CDCl3, 100 MHz): δ = 43.9 (C-12, CH2
of pyrazoline), 56.2 (Ar–OCH3), 62.5 (C-11, CH–N of
pyrazoline), 122.0–147.7 (Ar–C), 152.8 (C-8, C–Cl of
quinoline), 156.7 (C-13, C=N of pyrazoline), 167.7 (C-23,
C=O); LCMS: m/z 442.12 [M+]; anal. calcd. for
C25H19ClN4O2: C, 67.80; H, 4.32; N, 12.65; found: C,
67.91; H, 4.41; N, 12.58.
(5-(2-Chloroquinolin-3-yl)-3-(4-methoxyphenyl)-4,5-
dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4i)
Brownish crystals; yield: 63 %; mp 166–168 °C; IR (KBr)
νmax 3064 (C–H, aromatic), 2942, 2877 (C–H stretching,
–OCH3 group, –CH2-group), 1694 (C=O stretching), 1585
(C=N), 1528 (C=C), 732 (C–Cl stretching) cm−1; 1H NMR
(CDCl3, 400 MHz): δ = 3.34 (dd, Jab = 17.50 Hz, Jac = 3.08
Hz, 1H, Ha), 3.61 (s, 3H, –OCH3), 3.82 (dd, Jab = 17.48 Hz,
(5-(2-Chloroquinolin-3-yl)-3-(4-hydroxyphenyl)-4,5-
dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4l)
Light yellow crystals; yield: 70 %; mp 227–229 °C; IR
(KBr) νmax 3415 (–OH), 3063 (C–H, aromatic), 2879 (C–H
stretching, –CH2-group), 1696 (C=O stretching), 1585
(C=N), 1520 (C=C), 736 (C–Cl stretching) cm−1; 1H NMR
(CDCl3, 400 MHz): δ = 3.40 (dd, Jab = 17.46 Hz, Jac = 3.06
Hz, 1H, Ha), 3.77 (dd, Jab = 17.47 Hz, Jbc = 11.07 Hz, 1H,
Hb), 6.07 (dd, Jac = 3.13 Hz, Jbc = 11.12 Hz, 1H, Hc),
7.20–8.47 (m, 13H, Ar–H), 9.14 (s, 1H, OH); 13C NMR
(CDCl3, 100 MHz): δ = 43.8 (C-12, CH2 of pyrazoline),
Jbc = 11.12 Hz, 1H, Hb), 6.08 (dd, Jac = 3.14 Hz, Jbc
=
11.18 Hz, 1H, Hc), 7.10–8.44 (m, 13H, Ar–H); 13C NMR
(CDCl3, 100 MHz): δ = 43.6 (C-12, CH2 of pyrazoline),
55.5 (Ar–OCH3), 62.4 (C-11, CH–N of pyrazoline),
121.8–148.1 (Ar–C), 152.6 (C-8, C–Cl of quinoline), 156.6
(C-13, C=N of pyrazoline), 167.4 (C-23, C=O); LCMS: m/

Med Chem Res
62.3 (C-11, CH–N of pyrazoline), 121.4–148.7 (Ar–C),
152.5 (C-8, C–Cl of quinoline), 156.7 (C-13, C=N of
pyrazoline), 161.1 (C-17, Ar–C–OH), 167.5 (C-23, C=O);
LCMS: m/z 428.10 [M+]; Anal. Calcd. for C24H17ClN4O:
C, 67.21; H, 4.00; N, 13.06; found: C, 67.29; H, 4.05; N,
13.12.
3.79 (dd, Jab = 17.46 Hz, Jbc = 11.09 Hz, 1H, Hb), 6.09 (dd,
Jac = 3.15 Hz, Jbc = 11.12 Hz, 1H, Hc), 7.16–8.48 (m, 12H,
Ar–H); 13C NMR (CDCl3, 100 MHz): δ = 43.8 (C-12, CH2
of pyrazoline), 62.7 (C-11, CH–N of pyrazoline),
121.5–149.7 (Ar–C), 125.3, 127.8 (C-15, C-17, Ar–C–Cl),
152.7 (C-8, C–Cl of quinoline), 155.4 (C-13, C=N of
pyrazoline), 167.6 (C-23, C=O); LCMS: m/z 480.03 [M+];
anal. calcd. for C24H15Cl3N4O: C, 59.84; H, 3.14; N, 11.63;
found: C, 62.89; H, 3.17; N, 11.67.
(5-(2-Chloroquinolin-3-yl)-3-(3-nitrophenyl)-4,5-dihydro-
1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4m)
Brown crystals; yield: 61 %; mp 221–223 °C; IR (KBr) νmax
3065 (C–H, aromatic), 2877 (C–H stretching, –CH2-group),
1694 (C=O stretching), 1589 (C=N), 1518 (C=C), 1486,
1353 (N–O asymmetric, symmetric stretching, –NO2
group), 735 (C–Cl stretching) cm−1; 1H NMR (CDCl3, 400
MHz): δ = 3.38 (dd, Jab = 17.43 Hz, Jac = 3.05 Hz, 1H, Ha),
3.74 (dd, Jab = 17.49 Hz, Jbc = 11.06 Hz, 1H, Hb), 6.07 (dd,
Jac = 3.15 Hz, Jbc = 11.17 Hz, 1H, Hc), 7.25–8.56 (m, 13H,
Ar–H); 13C NMR (CDCl3, 100 MHz): δ = 43.5 (C-12, CH2
of pyrazoline), 62.7 (C-11, CH–N of pyrazoline),
121.5–149.1 (Ar–C), 140.2 (C-15, Ar–C–NO2), 152.8 (C-8,
C–Cl of quinoline), 155.8 (C-13, C=N of pyrazoline), 167.2
(C-23, C=O); LCMS: m/z 457.09 [M+]; anal. calcd. for
C24H16ClN5O3: C, 62.96; H, 3.52; N, 15.30; found: C,
62.93; H, 3.48; N, 15.26.
Biological assay
Antibacterial bioassay
The newly synthesized compounds (4a–o) were screened
for their antibacterial activity against Gram positive [Sta-
phylococcus aureus (MTCC-96), Streptococcus pyogenes
(MTCC-442)], and Gram-negative bacteria [Escherichia
coli (MTCC-443), Pseudomonas aeruginosa (MTCC-
1688)] at different concentrations of 1000, 500, 200, 100,
50, 25, 12.5 µg ml−1. Several of the newly synthesized
compounds (4a–o) were found to exhibit moderate to
excellent antimicrobial activity. Antibacterial activity was
carried out as per National Committee for Clinical
Laboratory Standards (NCCLS) protocol using serial broth
dilution method and all the standard strains used for the
antimicrobial activity were procured from Institute of
Microbial Technology (IMTECH), Chandigarh. “The drugs
which were found to be active in primary screening were
similarly diluted to obtain 100, 50, 25 and 12.5 µg ml−1
concentrations for secondary screening to test in a second
set of dilution against all microorganisms. Inoculum size for
test strain was adjusted to 106 CFU/ml (Colony Forming
Unit per milliliter) by likening the turbidity.
Mueller–Hinton Broth was used as a nutrient medium to
grow and dilute the synthesized compound suspension for
test organisms. 2 % DMSO was used as a diluent/vehicle to
acquire the desired concentration of synthesized compounds
and standard drugs to test upon standard microbial strains.
Diluted 1000 µg/ml concentrated solution of synthesized
compounds were used as stock solution. The control tube
containing not any antibiotic was instantly subcultured
[before inoculation] by spreading a loopful evenly over
quarter of a plate of medium suitable for the growth of test
organisms. The culture tubes were then incubated for 24 h at
37 °C, and the growth was observed visually and spectro-
photometrically. After that, 10 μg ml−1 suspensions were
further inoculated on appropriate media and development
was noted after 24 and 48 h. The lowest concentration
preventing appearance of turbidity was considered as
minimum inhibitory concentration (MIC, μg ml−1) i.e., the
amount of growth from the control tube before incubation
(5-(2-Chloroquinolin-3-yl)-3-(4-nitrophenyl)-4,5-dihydro-
1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4n)
Orange yellow crystals; yield: 65 %; mp 211–213 °C; IR
(KBr) νmax 3063 (C–H, aromatic), 2876 (C–H stretching,
–CH2-group), 1693 (C=O stretching), 1592 (C=N), 1519
(C=C), 1481, 1348 (N–O asymmetric, symmetric stretch-
1
ing, –NO2 group), 731 (C–Cl stretching) cm−1; H NMR
(CDCl3, 400 MHz): δ = 3.40 (dd, Jab = 17.44 Hz, Jac = 3.07
Hz, 1H, Ha), 3.76 (dd, Jab = 17.42 Hz, Jbc = 11.08 Hz, 1H,
Hb), 6.07 (dd, Jac = 3.12 Hz, Jbc = 11.09 Hz, 1H, Hc),
7.26–8.59 (m, 13H, Ar–H); 13C NMR (CDCl3, 100 MHz):
δ = 43.6 (C-12, CH2 of pyrazoline), 62.6 (C-11, CH–N of
pyrazoline),
121.4–149.3
(Ar–C),
150.1
(C-17,
Ar–C–NO2), 152.7 (C-8, C–Cl of quinoline), 155.6 (C-13,
C=N of pyrazoline), 167.4 (C-23, C=O); LCMS: m/z
457.09 [M+]; anal. calcd. for C24H16ClN5O3: C, 62.96; H,
3.52; N, 15.30; found: C, 62.91; H, 3.48; N, 15.31.
(5-(2-Chloroquinolin-3-yl)-3-(2,4-dichlorophenyl)-4,5-
dihydro-1H-pyrazol-1-yl)(pyridin-4-yl)methanone (4o)
White yellow crystals; yield: 68 %; mp 231–233 °C; IR
(KBr) νmax 3068 (C–H, aromatic), 2872 (C–H stretching,
–CH2-group), 1690 (C=O stretching), 1591 (C=N), 1523
(C=C), 734 (C–Cl stretching) cm−1; 1H NMR (CDCl3, 400
MHz): δ = 3.37 (dd, Jab = 17.49 Hz, Jac = 3.09 Hz, 1H, Ha),

Med Chem Res
(which represents the original inoculum) was compared.
Solvent had no influence on strain growth and the result of
this was greatly affected by the size of inoculum. DMSO
and sterilized distilled water were used as negative control
while ‘ampicillin’ antibiotic (1 U strength) was used as
positive control. Standard drug ‘ampicillin’ was used in this
study for evaluating antibacterial activity which showed
100, 100, 250 and 100 μg ml−1 MIC against E. coli, P.
aeruginosa, S. aureus, and S. pyogene, respectively as
shown in Table 1”.
MTT assay for cytotoxic activity
In vitro cytotoxicity activity of compounds (4a–o) was
measured by means of the IC50 using the MTT [3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide] assay
process. “The IC50 determination was achieved according to
the National Committee for Clinical Laboratory Standards
(NCCLS) recommendations. All synthesized compounds
were dissolved in 0.1 % DMSO with the stock concentration
of 10 g/l and diluted with medium freshly before drug
administration. Cell lines were sowed into 96-well plates at
density of 8000 cells/ well. After seeding it for 24 h, each
compound dilution was added in duplicate, and cultivation
continued at 37 °C in a moistened atmosphere containing 10
% FBS, 1 % glutamine, and 50 µM/ml gentamicin sulfate in a
5 CO2 and 95 % air. After 24 h, 20 µL MTT reagent at 5 mg/
ml in PBS (filter sterilized, light protected and stored at 4 °C)
per well was added, and after 4 h of incubation at 37 °C, MTT
is changed to a blue formazan product by mitochondrial
succinate dehydrogenase. This product was eluted from cells
by addition of 150 ml of DMSO. The absorbance at 570 nm
was determined by an ELISA using ELX800 micro plate
spectrophotometer”.
Antifungal bioassay
“The same newly synthesized compounds (4a–o) were
screened for their antifungal activity against in six sets
against C. albicans (MTCC-227), A. niger (MTCC-282)
and A. clavatus (MTCC-1323) at various primary con-
centrations of 1000, 500 and 250 μg ml−1. The primary
screen active compounds were similarly diluted to obtain
200, 125, 100, 62.5, 50, 25 and 12.5 μg ml−1 concentrations
for secondary screening to test in a second set of dilution
against all microorganisms. ‘Griseofulvin’ was used as a
standard drug for antifungal activity, which showed 500,
100 and 100 μg ml−1 MIC against C. albicans, A. niger and
A. clavatus, respectively (Table 1). For growth of fungi, in
the present procedure, we have used Sabourauds dextrose
broth at 28 °C in aerobic condition for 48 h. DMSO and
sterilized distilled water used as negative controls while
‘griseofulvin’ (1 U strength) was used as a positive control”.
SAR studies
Structure–activity relationship (SAR) studies revealed that
the antimicrobial activity in heterocyclic class of quinoline,
pyrazoline and pyridine molecules depend on the nature of
the peripheral substituents and their spatial relationship
Fig. 3 Antibacterial and
antifungal activity are expressed
in the form of bar chart (EW
electron withdrawing, ED
electron donating)

Med Chem Res
Compliance with ethical standards
within this skeleton. The pattern of substitution for the
derivatives is carefully selected to confer different electro-
nic environment of the molecules. The electronic nature of
the substituent groups lead to significant discrepancy in
antimicrobial activity. The presence of chloro, fluoro and
nitro substituents at para position on aromatic ring system
has amplified the antibacterial activity of compounds
compared to those of electron donating substituents. By
replacing substitution position to ortho and meta, com-
pound tends to lost its potency (Chart 1 and 3). Incor-
poration of electron donating groups such as methyl,
methoxy and hydroxy diminished the antibacterial property
(Chart 2 and 4). The presence of lipophilic substituent on
phenyl ring provides a positive effect on antifungal activity.
In rationality with the above results, electron withdrawing
groups like chloro and bromo on ortho and para substituted
position showed optimal activity. It is concluded from
Table 1 that, a compound without any substitution does not
display antimicrobial activity against a panel of all micro-
organisms. Figure 3 showed antimicrobial activity in the
form of bar chart.
Conflict of interest The authors declare that they have no com-
peting interests.
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bacterial and antifungal activity against various bacterial
and fungal strains. It is concluded from biological activity
table that structural and electronic diversity of these pro-
ducts influences their biological activities. Compounds 4d,
4e, 4f, 4g, 4n and 4o are the most characteristic derivatives
identified in present study because of their notable in vitro
antimicrobial potency. SAR studies revealed that when
electron withdrawing groups like fluoro, chloro, bromo and
nitro present in structure demonstrated effective anti-
microbial activity. Likewise di-substituted chloro group
(compound 4o) shows optimum inhibition over all strains of
bacteria and fungi. It may be considered as a promising lead
for further design and development of new lead molecules.
As discussed from toxicity studies, we were encouraged to
make modifications in electronic diversity on the basic
structure of the final compounds (4a–o) for generation of
non-toxic antimicrobial agents. It may be concluded that the
presence of three different pharmacophore scaffolds enri-
ched the biological activity.
Acknowledgments Authors are thankful to the University Grants
Commission, New Delhi and Department of Science & Technology,
New Delhi for financial support under the NON-SAP and DST-FIST
programs respectively.
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