New N-arylamino biquinoline derivatives: Synthesis, antimicrobial, antituberculosis, and antimalarial evaluation

Article abstract of DOI:10.1016/j.ejmech.2012.05.004

A new series of N-arylamino biquinoline derivatives 5a-x were synthesized by reaction of 2-chloro-3-formyl quinolines 2a-d with malononitrile and various enhydrazinoketones 4a-f in absolute ethanol. The newly synthesized compounds were evaluated for their in vitro antimicrobial activity against a representative panel of pathogenic strains and antituberculosis activity against Mycobacterium tuberculosis H37Rv. Compounds 5h and 5s exhibited excellent antibacterial activity and some of the compounds demonstrated moderate antituberculosis activities compared with the first line drugs. The compounds were evaluated in vitro for their activity against the growth of Plasmodium falciparum, the malaria causing parasite. Some of them showed antimalarial activity with IC₅₀ values as low as 0.005-0.009 μg/mL.

Full text of DOI:10.1016/j.ejmech.2012.05.004

European Journal of Medicinal Chemistry 54 (2012) 239e247
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
New N-arylamino biquinoline derivatives: Synthesis, antimicrobial,
antituberculosis, and antimalarial evaluation
*
Nimesh M. Shah, Manish P. Patel, Ranjan G. Patel
Department of Chemistry, Sardar Patel University, Vallabh Vidyanagar 388120, Gujarat, India
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
< A series of N-arylamino biquinoline
derivatives have been synthesized.
< All the synthesized compounds
were screened for antimicrobial
activities as well as antituberculosis
activity.
< They were also compared pharma-
cologically concerning antimalarial
activity
against
Plasmodium
falciparum.
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 N-arylamino biquinoline derivatives 5a-x were synthesized by reaction of 2-chloro-3-
formyl quinolines 2aed with malononitrile and various enhydrazinoketones 4aef in absolute ethanol.
The newly synthesized compounds were evaluated for their in vitro antimicrobial activity against
a representative panel of pathogenic strains and antituberculosis activity against Mycobacterium
tuberculosis H37Rv. Compounds 5h and 5s exhibited excellent antibacterial activity and some of the
compounds demonstrated moderate antituberculosis activities compared with the first line drugs. The
compounds were evaluated in vitro for their activity against the growth of Plasmodium falciparum,
the malaria causing parasite. Some of them showed antimalarial activity with IC50 values as low as
Received 28 September 2011
Received in revised form
30 April 2012
Accepted 2 May 2012
Available online 12 May 2012
Keywords:
Quinoline
Antimicrobial activity
Multicomponent reaction
IC50
0.005e0.009 mg/mL.
Ó 2012 Elsevier Masson SAS. All rights reserved.
1. Introduction
underscores the urgency and continuous need for the discovery of
new therapeutics.
Malaria is a devastating disease, which is endemic in about 100
developing countries putting 3.3 billion people at risk. It is esti-
mated that malaria causes 300e500 million clinical cases and 1-2
million deaths annually [1]. Plasmodium falciparum is the parasite
responsible for most malaria cases (80%), which often prove fatal.
The burden of malaria has not declined, partly because the para-
sites have become resistant to the available drugs [2e4]. The
alarming spread of drug resistant strains of P. falciparum
Emerging infectious diseases and the increasing number of
multi-drug resistant microbial pathogens still make the treatment
of infectious diseases an important and pressing global health
problem. Therefore, there is a vital need to discover new antimi-
crobial agents to avert the emergence of resistance and ideally
shorten the duration of therapy [5,6]. Furthermore, the ever-
increasing drug resistance, toxicity and side effects of currently
used antituberculosis drugs, and the absence of their bactericidal
activity highlight the need for new, safer and more effective anti-
mycobacterial compounds. So this spot of research is accorded an
immense meaning and keeps on attracting much attention of
growing number of medicinal chemists.
* Corresponding author. Tel.: þ91 02692 226856.
E-mail address: patelranjanben@yahoo.com (R.G. Patel).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.05.004

240
N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
Among the important pharmacophores responsible for antimi-
heterylidenenitrile, containing the electron-poor C]C double
bond, from the Knoevenagel condensation between 2-chloro-3-
formyl quinolines 2aed and malononitrile by loss of water mole-
cules. Michael addition of enhydrazinoketones 4aef to the ylidenic
bond in forming an acyclic intermediate which cyclizes by nucle-
ophilic attack of the NH group on the cyano carbon, followed by
tautomerisation to the final products 5aex.
crobial & antimalarial activity, the quinoline scaffold is still
considered a viable lead structure for the synthesis of more effi-
cacious and broad spectrum antimicrobial & antimalarial agents. In
the recent time, quinoline nucleus has gathered an immense
attention among chemists as well as biologists as it is one of the key
building elements for many naturally occurring compounds. The
quinoline ring is endowed with various activities, such as antimi-
crobial [7], antituberculosis [8], antimalarial [9], anti- inflammatory
[10], anticancer [11], antibiotic [12], antihypertensive [13], tyroki-
nase PDGF-RTK inhibiting agents [14], and antiHIV [15,16].
Amongst the various activities of their derivatives, antimicrobial &
antimalarial activity is noteworthy.
3. Pharmacology
3.1. Antimicrobial activity
The antimicrobial activity of synthesized compounds were
carried out by broth microdilution method according to National
Committee for Clinical Laboratory Standards (NCCLS) [27]. Anti-
bacterial activity was screened against three Gram positive (Bacillus
subtilis MTCC 441, Clostridium tetani MTCC 449, Streptococcus
pneumoniae MTCC 1936) and three Gram negative (Escherichia coli
MTCC 443, Salmonella typhi MTCC 98, Vibrio cholerae MTCC 3906)
bacteria by using ampicillin, ciprofloxacin, norfloxacin and chlor-
amphenicol as a standard antibacterial drug. Antifungal activity
was screened against two fungal species (Aspergillus. fumigats
MTCC 3008 and Candida albicans MTCC 227) where griseofulvin
and nystatin used as standard antifungal drugs. The antimicrobial
screening data are shown in Table 2. All MTCC cultures were
collected from Institute of Microbial Technology, Chandigarh and
tested against above mentioned known drugs. Mueller Hinton
broth was used as nutrient medium to grow and dilute the drug
suspension for the test. DMSO was used as diluents to get desired
concentration of compounds to test upon standard bacterial strains.
Further, there are few examples of syntheses of N-substituted
quinoline derivatives in the literature [17,18]. However, after
extensive literature search, it was observed that till date enough
efforts have not been made to synthesize N-arylamino biquinoline
scaffold and to study its antimalarial activity. In the radiance of the
aforementioned facts and as a prolongation of our efforts in
developing heterocycles of biological interest [19e25] also consid-
ering the significant role of quinolines in biological applications, we
were provoked to synthesize new series of N-arylamino biquinoline
derivatives and evaluate them as antimicrobial, antitubercular and
antimalarial agents. The variation in the nature and size of
substituents at such structures was thought to be of interest rep-
resenting variable electronic, lipophilic, and steric environment
that would influence the anticipated biological activities.
2. Chemistry
The synthetic approach adopted to obtain the target compounds
is depicted in Scheme 1. The starting material 2-chloro-3-formyl
quinolines 2aed were prepared according to literature procedure
[26] by VilsmeiereHaack reaction of acetanilide 1aed. The required
3.2. Antituberculosis activity
enhydrazinoketones 4aef was prepared by the reaction of
b
-dike-
A primary in vitro antituberculosis activity of all the newly
synthesized compounds against Mycobacterium tuberculosis H37Rv
strain was determined by using LowensteineJensen medium
(conventional method) as described by Rattan [28] and the
observed results are presented in Table 3 in the form of % inhibition,
relative to that of standard antitubercular drug isoniazid and
rifampicin. Compounds effecting <90% inhibition in the primary
screen were not evaluated further.
tone with phenyl hydrazine hydrochloride in aqueous condition.
The N-arylamino biquinoline derivatives were synthesized by
reaction of 2-chloro-3-formyl quinolines 2aed, malononitrile with
appropriate enhydrazinoketones 4aef in absolute ethanol in
presence of base piperidine.
A plausible mechanism for the reaction is outlined in Scheme 2.
The reaction occurs via an in situ initial formation of the
Scheme 1. General synthetic route for the title compounds 5aex.

N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
241
Scheme 2. Plausible mechanistic pathway of the synthesis of N-arylamino biquinoline derivatives.
3.3. Antimalarial activity
compounds 5g (R1 ¼ OCH3, R2 ¼ Cl), 5r (R1 ¼ CH3, R2 ¼ OCH3) and 5u
(R1 ¼ H, R2 ¼ Br) MIC 100 g/mL were found equipotent to ampicillin
MIC 100 g/mL towards S. pneumoniae. In case of inhibiting Gram
m
All new N-arylamino biquinoline derivatives 5aex were evalu-
ated for their antimalarial activity in vitro upon the P. falciparum
strain. In vitro activity against P. falciparum was determined using
chloroquine and quinine as reference compounds. The results of the
biological evaluation are expressed as the drug concentration
resulting in 50% inhibition (IC50) of parasite growth and are listed in
Table 4.
m
possitive bacteria B. subtilis, compound 5a (R1 ¼ H, R2 ¼ H), 5g
(R1 ¼ OCH3, R2 ¼ Cl), 5h (R1 ¼ Cl, R2 ¼ Cl), 5j (R1 ¼ CH3, R2 ¼ CH3), 5k
(R1 ¼ OCH3, R2 ¼ CH3), 5p (R1 ¼ Cl, R2 ¼ F), 5r (R1 ¼ CH3, R2 ¼ OCH3), 5s
(R1 ¼ OCH3, R2 ¼ OCH3), 5u (R1 ¼ H, R2 ¼ Br) and 5v (R1 ¼ CH3, R2 ¼ Br)
MIC < 250
mg/mL displayed significant activity compared to the
standard ampicillin MIC 250
m
g/mL. Compound 5c (R1 ¼ OCH3,
R2 ¼ H), 5i (R1 ¼ H, R2 ¼ CH3), 5n (R1 ¼ CH3, R2 ¼ F), 5o (R1 ¼ OCH3,
R2 ¼ F), 5t (R1 ¼ Cl, R2 ¼ OCH3), 5w (R1 ¼ OCH3, R2 ¼ Br) and 5x
(R1 ¼ Cl, R2 ¼ Br) MIC 250 mg/mL exhibited comparable activityas the
4. Results and discussion
4.1. Analytical results
standard ampicillin MIC 250 mg/mL against B. subtilis. Compounds 5g
(R1 ¼ OCH3, R2 ¼ Cl), 5h (R1 ¼ Cl, R2 ¼ Cl), 5k (R1 ¼ OCH3, R2 ¼ CH3),
A series of new N-arylamino biquinoline derivatives 5aex have
been synthesized substantially through the synthetic route as illus-
trated in Scheme 1. The structures of all the newly synthesized
compounds were confirmed by FTIR, 1H NMR, 13C NMR, mass and
elemental analysis. The absorption bands for all the compounds in
IR-spectra were observed in the range of 2180e2210 cmꢀ1 corre-
sponding to C^N. The IR spectrum of compound 5aex exhibited
characteristic absorption band in the range 3260e3425 cmꢀ1 is
mainly attributed to the presence of NH2. Carbonyl group stretching
frequency was observed at 1640e1665 cmꢀ1. The 1H NMR spectrum
5m (R1 ¼ H, R2 ¼ F), 5n (R1 ¼ CH3, R2 ¼ F), 5s (R1 ¼ OCH3, R2 ¼ OCH3),
5t (R1 ¼ Cl, R2 ¼ OCH3) and 5w (R1 ¼ OCH3, R2 ¼ Br) MIC < 250
mg/mL
revealed good inhibitoryeffects while compounds 5a(R1 ¼ H, R2 ¼ H),
5e (R1 ¼ H, R2 ¼ Cl), 5f (R1 ¼ CH3, R2 ¼ H), 5i (R1 ¼ H, R2 ¼ CH3), 5j
(R1 ¼ CH3, R2 ¼ CH3), 5l (R1 ¼ Cl, R2 ¼ CH3), 5q (R1 ¼ H, R2 ¼ OCH3),
5v (R1 ¼ CH3, R2 ¼ Br) and 5x (R1 ¼ Cl, R2 ¼ Br) MIC 250
mg/mL
have the inhibitory effects as that of the standard ampicillin MIC
250
g/mL against C. tetani. Against E. coli compound 5h (R1 ¼ Cl,
R2 ¼ Cl) MIC 62.5 g/mL possessed pronounced activity compared
to ampicillin MIC 100
m
m
m
g/mL where as compounds 5b (R1 ¼ CH3,
ofcompound5aexshowedasharp singletat
proton, and a broad singet at 8.80e8.99 ppm arising due to NH. In
the 13C NMR spectral data of the title compounds 5aex, most char-
acteristic signal around 34.22e34.60 ppm indicated the formation
of quinoline ring and the carbonyl carbon was observed at around
194.90e195.05 ppm. The signal at around 56.22e57.55 ppm is
assigned to carbon attached with carbonitrile.
d
5.02e5.13ppmofeCH
R2 ¼ H), 5f (R1 ¼ CH3, R2 ¼ H), 5s (R1 ¼ OCH3, R2 ¼ OCH3) and 5x
d
(R1 ¼ Cl, R2 ¼ Br) showed inhibitory effects as that of the standard
ampicillin MIC 100
MIC 50
compared to ampicillin MIC 100
(R1 ¼ OCH3, R2 ¼ Cl) and 5h (R1 ¼ Cl, R2 ¼ Cl) MIC 100
found equally active as ampicillin MIC 100 g/mL. Compound 5s
(R1 ¼ OCH3, R2 ¼ OCH3) MIC 62.5 g/mL showed excellent activity
while compounds 5g (R1 ¼ OCH3, R2 ¼ Cl), 5h (R1 ¼ Cl, R2 ¼ Cl), 5o
(R1 ¼ OCH3, R2 ¼ F) and 5u (R1 ¼ H, R2 ¼ Br) MIC 100 g/mL were
found equipotent against Gram negative bacteria V. cholerae rela-
tive to ampicillin MIC 100 g/mL.
m
g/mL. Compound 5s (R1 ¼ OCH3, R2 ¼ OCH3)
d
m
g/mL displayed excellent activity towards S. typhi
g/mL while compounds 5g
g/mL were
m
d
d
m
m
The obtained elemental analysis values are in good agreement
with theoretical data. Mass spectra of all the title compounds gave
expected Mþ peak corresponding with proposed molecular mass.
Physicochemical properties of all the derivatives are shown in
Table 1.
m
m
m
The data indicate that a change in the substituent might also
affect the antibacterial activity of title compounds 5aex. Compar-
ison of biological activities among 5aex shows functional groups as
R1 ¼ OCH3 or R2 ¼ Br to be potentially more active against B. subtilis.
Also antibacterial potency of compounds among 5aex shows
functional groups as R2 ¼ Cl/CH3 found more active against C. tetani.
A close examination of the structures of the active compounds in
4.2. Biological results
4.2.1. Antimicrobial activity
Reviewing of the antibacterial activities of biquinoline derivatives
(Table 1) indicate that compound 5s (R1 ¼ OCH3, R2 ¼ OCH3) showed
highest activity i.e. 25
mg/mL against S. pneumoniae. Moreover,

242
N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
Table 1
Physicochemical characteristics of N-arylamino biquinoline derivatives 5aex.
Compd.
R1
R2
Formula (Mw)
Mp (ꢁC)
Yielda%
Analysis (%) calc./found
C
H
N
5a
5b
5c
5d
5e
5f
H
H
C27H24ClN5O (469.97)
C28H26ClN5O (483.99)
C28H26ClN5O2 (499.99)
C27H23Cl2N5O (504.41)
226ꢀ228
240e242
210e212
264ꢀ266
252e254
224e226
249e251
212e214
198e200
231e233
225e227
260e262
228e230
280e282
247e249
212e214
269e271
255e257
282e284
221e223
256e258
237e239
240e242
208e210
84
81
78
85
80
73
78
82
70
74
77
80
77
72
80
74
71
79
70
81
83
79
83
80
69.00
68.79
69.48
69.33
67.26
67.50
64.29
64.06
64.29
64.35
64.87
65.03
62.93
63.02
60.18
60.35
69.48
69.21
69.94
70.11
67.76
67.55
64.87
64.80
66.46
66.64
66.99
67.15
64.92
64.85
62.08
62.31
67.26
67.42
67.76
68.03
65.72
65.55
62.93
63.10
59.08
58.89
59.75
59.90
58.09
57.88
55.59
55.47
5.15
4.92
5.41
5.24
5.24
5.12
4.60
4.77
4.60
4.74
4.86
4.70
4.71
4.54
4.12
4.26
5.41
5.50
5.67
5.75
5.49
5.58
4.86
5.04
4.75
4.47
5.02
5.16
4.86
4.77
4.24
4.36
5.24
5.40
5.49
5.26
5.32
5.20
4.71
4.92
4.22
4.38
4.48
4.55
4.35
4.53
3.80
3.69
14.90
14.81
14.47
14.64
14.01
14.19
13.88
13.74
13.88
13.99
13.51
13.62
13.10
13.28
13.00
12.94
14.47
14.39
14.06
13.97
13.62
13.44
13.51
13.78
14.35
14.22
13.95
13.73
13.52
13.70
13.41
13.18
14.01
14.23
13.62
13.48
13.21
13.41
13.10
12.99
12.76
12.59
12.44
12.27
12.10
12.01
12.01
12.28
CH3
OCH3
Cl
H
H
H
H
Cl
C27H23Cl2N5O (504.41)
CH3
OCH3
Cl
Cl
C28H25Cl2N5O (518.44)
C28H25Cl2N5O2 (534.44)
C27H22Cl3N5O (538.86)
C28H26ClN5O (483.99)
C29H28ClN5O (498.02)
C29H28ClN5O2 (514.02)
C28H25Cl2N5O (518.44)
5g
5h
5i
Cl
Cl
H
CH3
CH3
CH3
CH3
F
5j
CH3
OCH3
Cl
5k
5l
5m
5n
5o
5p
5q
5r
H
C27H23ClFN5O (487.96)
CH3
OCH3
Cl
F
C28H25ClFN5O (501.98)
C28H25ClFN5O2 (517.98)
C27H22Cl2FN5O (522.40)
C28H26ClN5O2 (499.99)
C29H28ClN5O2 (514.02)
C29H28ClN5O3 (530.02)
C28H25Cl2N5O2 (534.44)
F
F
H
OCH3
OCH3
OCH3
OCH3
Br
CH3
OCH3
Cl
5s
5t
5u
5v
5w
5x
H
C27H23BrClN5O (548.86)
CH3
OCH3
Cl
Br
C28H25BrClN5O (562.89)
C28H25BrClN5O2 (578.89)
Br
Br
C27H22BrCl2N5O (583.31)
a
All the yields are on isolated basis.
Table 1 revealed that, their antimicrobial activity is strongly bound
to the nature of the substituent at the quinoline-C6, together with
the substituent linked to the arylamino part of the structure. In
general, it could be clearly recognized that potential antibacterial
activity was encountered with 5s quinoline containing a methoxy
substituent and aryl ring also containing a methoxy substituent.
However, replacement of methoxy substituent on arylamino part
with H i.e 5c resulted in remarkable reduction in antibacterial
spectrum while replacement with chloro substituent resulted in 5g
which also showed comparably good activity. The dihalo derivative
5h which contains a chloro substituent both on arylamino part and
quinoline-C6 also exhibited promising antibacterial activity.
Compound 5d without a substituent on arylamino part also showed
a marked decrease in antimicrobial activity. Thus substituent in the
aryamino ring seems to have greater influence on the antibacterial
activity.
fumigatus. Compounds which exhibited excellent activity towards
C. albicans compared to griseofulvin MIC 500 g/mL include 5c
(R1 ¼ OCH3, R2 ¼ H), 5e (R1 ¼ H, R2 ¼ Cl), 5o (R1 ¼ OCH3, R2 ¼ F) and
g/mL. The data indicate that
m
5v (R1 ¼ CH3, R2 ¼ Br) MIC 250
m
functional groups as R2 ¼ CH3 involve in the antifungal potency of
respective compounds.
4.2.2. Antituberculosis activity
The encouraging results from the antimicrobial studies promp-
ted us to go for the preliminary screening of the title compounds for
their in vitro antituberculosis activity against M. tuberculosis H37Rv.
Of the compounds screened for antituberculosis activity, compound
5h (R1 ¼ Cl, R2 ¼ Cl) found to possess better activity against
M. tuberculosis H37Rv with 87% inhibition. Also compounds 5c
(R1 ¼ OCH3, R2 ¼ H) and 5x (R1 ¼ Cl, R2 ¼ Br) displayed moderate
inhibition of 80 and 85% respectively. Even though, the observed
activities exhibited by the tested compounds were lower than that
of the standard drug isoniazid, the most potent compound of the
It also appears from antifungal activity data compounds were
more active towards C. albicans and less active aganist Aspergillus

N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
243
Table 2
In vitro antimicrobial activity of N-arylaminobiquinolines 5aex MICs,
m
g/mL.
Gram negative bacteria
Compd.
Gram positive bacteria
Fungi
B.S. MTCC 441
C.T. MTCC 449
S.P. MTCC 1936
E.C. MTCC 443
S.T. MTCC 98
V.C. MTCC 3906
A.F. MTCC 3008
C.A. MTCC 227
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
5q
5r
5s
5t
5u
5v
5w
5x
Ampi.
Chlor.
Cipro.
Norflo.
Grise.
Nyst.
200
500
250
1000
500
500
100
200
250
200
200
500
500
250
250
100
500
200
200
250
200
200
250
250
250
50
250
500
500
1000
250
250
200
200
250
250
100
250
200
200
500
500
250
500
200
200
500
250
200
250
250
50
200
250
250
1000
500
250
100
200
500
200
200
500
500
250
200
200
200
100
25
250
100
250
250
200
100
50
50
10
n. t.
n. t.
200
100
250
200
200
100
250
62.5
200
500
150
250
200
250
125
250
250
250
100
250
250
250
200
100
100
50
250
200
250
250
500
200
100
100
200
500
200
250
200
250
125
500
200
500
50
200
250
250
125
250
100
50
25
10
n. t.
n. t.
250
250
500
1000
500
200
100
100
250
500
200
250
200
200
100
125
250
500
62.5
250
100
250
200
250
100
50
>1000
>1000
500
1000
500
1000
>1000
>1000
500
500
500
250
1000
250
500
>1000
>1000
500
500
500
500
500
500
500
1000
1000
500
>1000
>1000
>1000
500
>1000
>1000
>1000
500
1000
1000
250
500
>1000
>1000
500
100
>1000
250
1000
1000
n. t.
n. t.
n. t.
500
n. t.a
n. t.
50
100
50
n. t.
n. t.
25
10
n. t.
n. t.
25
10
n. t.
n. t.
n. t.
n. t.
100
100
100
n. t.
n. t.
n. t.
500
100
B.S.: Bacillus subtilis, C.T.: Clostridium tetani, S.P.: Streptococcus pneumoniae.
E.C.: Escherichia coli, S.T.: Salmonella typhi, V.C.: Vibrio cholerae.
A.F.: Aspergillus fumigatus, C.A.: Candida albicans.
MTCC: Microbial Type Culture Collection.
Ampi.: Ampicillin, Chlor.: Chloramphenicol, Cipro.: Ciprofloxacin, Norflo.: Norfloxacin.
Grise.: Griseofulvin, Nyst.: Nystatin.
a
n. t. not tested.
series, compound 5h (R1 ¼ Cl, R2 ¼ Cl) opens up new door to opti-
(R1 ¼ H, R2 ¼ CH3), 5l (R1 ¼ Cl, R2 ¼ CH3), 5s (R1 ¼ OCH3, R2 ¼ OCH3),
5v (R1 ¼ CH3, R2 ¼ Br) and 5w (R1 ¼ OCH3, R2 ¼ Br) exhibited
superior activity against P. falciparum strain as compared to chlo-
roquine IC50 0.020. Interestingly, it appears that electronic factors
have minimal effect, with both electron-donating and electron-
withdrawing groups giving the same range of activities.
mize this series for new class of antituberculars.
4.2.3. Antimalarial activity
All new compounds 5aex were evaluated for their antimalarial
activity against chloroquine-sensitive (3D7) strain of P. falciparum.
The IC50 values were calculated from experiments carried out in
duplicate and the results are presented in Table 4. As shown in
Table 4, they were found to have IC50 between 0.005 and 0.54 upon
P. falciparum strain. It is important to note that compounds 5a
(R1 ¼ H, R2 ¼ H), 5f (R1 ¼ CH3, R2 ¼ H), 5g (R1 ¼ OCH3, R2 ¼ Cl), 5i
5. Conclusion
In conclusion, novel series of N-arylamino biquinoline 5aex
were synthesized and tested for their in vitro antimicrobial,
Table 3
Table 4
In vitro antituberculosis activity (% inhibition) of N-arylamino biquinolines 5aex
In vitro antimalarial activity of N-arylamino biquinolines 5aex.
M. tuberculosis H37Rv (at concentration 6.25 mg/mL).
Compd.
IC50
(m
g/mL)
Compd.
IC50 (mg/mL)
Compd.
% inhibition
Compd.
% inhibition
5a
5b
5c
5d
5e
5f
5g
5h
5i
0.015
0.15
0.22
0.21
0.42
0.008
0.009
0.028
0.005
0.22
0.19
0.014
0.027
5n
5o
5p
5q
5r
5s
5t
5u
0.047
0.25
0.154
0.025
0.54
0.019
0.026
0.54
0.012
0.014
0.08
5a
5b
5c
5d
5e
5f
5g
5h
5i
8
12
80
5
22
15
21
87
8
32
50
72
62
5n
5o
5p
5q
5r
5s
5t
5u
33
22
65
44
3
7
22
33
47
17
85
98
99
5v
5w
5x
5v
5w
5x
Rifampicin
Isoniazid
5j
5k
5l
5j
5k
5l
Chloroquine
Quinine
0.020
0.268
5m
5m

244
N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
antituberculosis and antimalarial activity. Promising pharmaco-
logical results were obtained. The noteworthy compound 5s
exhibited good antimicrobial potency against all the microor-
ganism tested. Most of the compounds showed better antimalarial
activity, further optimization and development of this series is
required for drug development. Furthermore, this work contributes
to validate the choice of the arylamino biquinoline moiety as
a template useful for designing new antimicrobial/antimalarial
compounds.
139.39, 146.27, 146.99, 147.23, 150.05, 153.46, 155.11 (17C, Ar-C),
194.92 (C]O); MS (m/z): 470 (Mþ).
6.1.1.2. 2-Amino-4-(2-chloro-6-methyl(3-quinolyl))-7,7-dimethyl-5-
oxo-1-phenylamino-1,4,6,7,8-pentahydro
quinoline-3-carbonitrile
(5b). IR (KBr, nmax, cmꢀ1): 3410 & 3275 (asym. & sym. stretching
of eNH2), 2200 (C^N stretching), 1645 (C]O stretching), 755
(CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.91 (s, 3H, CH3),
0.98 (s, 3H, CH3), 1.97e2.81 (m, 4H, 2 ꢃ CH2), 2.52 (s, 3H, AreCH3),
5.08 (s,1H, CH), 6.33e8.25 (m,11H, AreH and NH2), 8.82 (s,1H, NH).
13C NMR (100 MHz, DMSO-d6)
d: 21.59 (Ar-CH3), 27.76, 28.97 (2C,
6. Experimental
CH3), 32.11 (C(CH3)2), 34.29 (C4), 38.03 (CH2), 49.45 (CH2eCO),
57.11 (CeCN), 111.19, 112.45, 120.91, 121.27, 126.36, 126.61, 127.66,
129.98, 133.68, 136.68, 137.62, 139.27, 144.84, 146.97, 149.14, 153.42,
155.06 (17C, AreC), 194.89 (C]O).
6.1. Chemistry
All reactions were performed with commercially available
reagents and they were used without further purification. Organic
solvents were purified by standard methods [29] and stored over
molecular sieves. All reactions were monitored by thin-layer
chromatography (TLC, on aluminium plates coated with silica gel
60 F254, 0.25 mm thickness, Merck) carried on fluorescent coated
plates and detection of the components was made by exposure to
iodine vapours or UV light. Melting points were determined in open
capillaries and the declared values are not corrected. Infrared
spectra were recorded on Shimadzu FTIR 8401 spectrophotometer
using potassium bromide pellets in the range 4000e400 cmꢀ1 and
frequencies of only characteristic peaks are expressed in cmꢀ1. 1H &
13C Nuclear Magnetic Resonance spectra were recorded in DMSO-d6
on a Bruker Avance 400F (MHz) spectrometer (Bruker Scientific
Corporation Ltd., Switzerland) using residual solvent signal as an
internal standard at 400 MHz and 100 MHz respectively. Chemical
shifts are reported in parts per million (ppm). Mass spectra were
scanned on a Shimadzu LCMS 2010 spectrometer (Shimadzu,
Tokyo, Japan) at Oxygen Healthcare Research Pvt. Ltd., Ahmedabad.
Elemental analyses were performed by PerkineElmer 2400 series-
II elemental analyzer (PerkineElmer, USA) at Sophisticated
Instrumentation Centre for Applied Research & Training (SICART),
Vallabh Vidyanagar and all compounds are within ꢂ0.4% of the
theoretical compositions. Yields are not optimized. Ampicillin,
griseofulvin, nystatin and isoniazid were commercial.
6.1.1.3. 2-Amino-4-(2-chloro-6-methoxy(3-quinolyl))-7,7-dimethyl-
5-oxo-1-phenylamino-1,4,6,7,8,-pentahydro quinoline-3-carbnitrile
(5c). IR (KBr, nmax, cmꢀ1): 3425 & 3270 (asym. & sym. stretching
of eNH2), 2195 (C^N stretching),1645 (C]O stretching), 750 (CeCl
stretching). 1H NMR (400 MHz, DMSO-d6):
d
0.90 (s, 3H, CH3), 0.98
(s, 3H, CH3), 1.95e2.80 (m, 4H, 2 ꢃ CH2), 3.87 (s, 3H, AreOCH3), 5.09
(s, 1H, CH), 6.40e8.18 (m, 11H, AreH and NH2), 8.84 (s, 1H, NH). 13
NMR (100 MHz, DMSO-d6) : 27.63, 29.06 (2C, CH3), 32.12 (C(CH3)2),
C
d
34.36 (C4), 38.02 (CH2), 49.47 (CH2-CO), 56.04 (AreOCH3), 57.17
(CeCN), 105.52, 111.24, 112.18, 120.32, 121.90, 123.05, 129.21, 129.65,
131.09, 134.30, 136.42, 139.98, 142.71, 146.87, 148.55, 154.13, 158.59
(17C, AreC), 194.92 (C]O).
6.1.1.4. 2-Amino-4-(2,6-dichloro(3-quinolyl))-7,7-dimethyl-5-oxo-1-
phenylamino-1,4,6,7,8,-pentahydro quinoline-3-carbonitrile (5d). IR
(KBr, nmax, cmꢀ1): 3415 & 3260 (asym. & sym. stretching of eNH2),
2185 (C^N stretching), 1640 (C]O stretching), 755 (CeCl stretch-
ing). 1H NMR (400 MHz, DMSO-d6):
d
0.90 (s, 3H, CH3), 1.00 (s, 3H,
CH3), 1.96-2.78 (m, 4H, 2 ꢃ CH2), 5.10 (s, 1H, CH), 6.43e8.28 (m,11H,
AreH and NH2), 8.88 (s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d
:
27.68, 28.96 (2C, CH3), 32.15 (C(CH3)2), 34.40 (C4), 37.93 (CH2), 49.45
(CH2eCO), 56.72 (CeCN), 111.43, 112.02, 120.26, 124.41, 126.35,
127.58, 128.71, 130.05, 131.25, 132.22, 134.62, 136.76, 144.12, 146.36,
148.53, 153.71, 155.64 (17C, AreC), 195.02 (C]O).
6.1.1. General procedure for the synthesis of N-arylamino
6.1.1.5. 2-Amino-4-(2-chloro-(3-quinolyl))-7,7-dimethyl-5-oxo-1-(4-
biquinoline derivatives 5aex
chlorophenylamino)-1,4,6,7,8-pentahydro
quinoline-3-carbonitrile
A 100 mL round bottomed flask, fitted with a reflux condenser,
was charged with a mixture of 2-chloro-3-formyl quinolines 2aed
(1 mmol), malononitrile (1 mmol), enhydrazinoketones 4aef
(1 mmol), and a catalytic amount of piperidine in ethanol (10 mL).
The mixture was heated under reflux for 2e3 h and the progress of
the reaction was monitored by TLC. After the completion of reaction
(as evidenced by TLC), the reaction mixture was cooled to room
temperature and stirred magnetically for further 20 min, the solid
mass separated was collected by filtration, washed well with
ethanol (10 mL) and purified by leaching in equal volume ratio of
chloroform and methanol (10 mL) to obtain pure solid sample.
(5e). IR (KBr, nmax, cmꢀ1): 3420 & 3265 (asym. & sym. stretching
of eNH2), 2190 (C^N stretching),1650 (C]O stretching), 760 (CeCl
stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.90 (s, 3H, CH3), 0.99
(s, 3H, CH3), 1.91e2.82 (m, 4H, 2 ꢃ CH2), 5.13 (s, 1H, CH), 6.40e8.25
(m, 11H, AreH and NH2), 8.80 (s, 1H, NH). 13C NMR (100 MHz,
DMSO-d6) d: 27.80, 29.08 (2C, CH3), 32.17 (C(CH3)2), 34.28 (C4),
37.98 (CH2), 49.55 (CH2eCO), 56.88 (CeCN), 111.54, 113.10, 120.25,
125.32, 126.71, 127.48, 128.55, 129.79, 130.05, 131.11, 134.25, 136.74,
144.41, 146.30, 147.26, 150.50, 154.33 (17C, AreC), 194.94 (C]O).
6.1.1.6. 2-Amino-4-(2-chloro-6-methyl(3-quinolyl))-7,7-dimethyl-5-
oxo-1-(4-chlorophenylamino)-1,4,6,7,8-pentahydro
quinoline-3-
6.1.1.1. 2-Amino-4-(2-chloro(3-quinolyl))-7,7-dimethyl-5-oxo-1-
phenylamino-1,4,6,7,8-pentahydro quinoline-3-carbonitrile (5a). IR
(KBr, nmax, cmꢀ1): 3420 & 3280 (asym. & sym. stretching of eNH2),
2180 (C^N stretching), 1640 (C]O stretching), 750 (CeCl
carbonitrile (5f). IR (KBr, nmax, cmꢀ1): 3420 & 3260 (asym. & sym.
stretching of eNH2), 2205 (C^N stretching), 1665 (C]O stretch-
ing), 755 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.92 (s,
3H, CH3), 0.99 (s, 3H, CH3), 1.95e2.84 (m, 4H, 2 ꢃ CH2), 2.54 (s, 3H,
stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.91 (s, 3H, CH3),
AreCH3), 5.09 (s, 1H, CH), 6.38e8.21 (m, 10H, AreH and NH2), 8.90
0.98 (s, 3H, CH3), 1.93e2.81 (m, 4H, 2 ꢃ CH2), 5.11 (s, 1H, CH),
(s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d: 21.57 (AreCH3), 27.72,
6.35e8.23 (m, 12H, AreH and NH2), 8.83 (s, 1H, NH). 13C NMR
29.05 (2C, CH3), 32.14 (C(CH3)2), 34.39 (C4), 38.07 (CH2), 49.53
(CH2eCO), 57.55 (CeCN), 111.25, 113.64, 120.72, 124.29, 126.37,
127.82, 129.30, 130.20, 133.05, 137.62, 138.13, 139.36, 144.38, 147.25,
148.90, 152.21, 155.42 (17C, AreC), 195.02 (C]O).
(100 MHz, DMSO-d6) d: 27.76, 29.23 (2C, CH3), 32.26 (C(CH3)2),
34.37 (C4), 38.24 (CH2), 49.55 (CH2eCO), 57.08 (CeCN), 111.85,
112.95, 120.92, 121.91, 126.26, 127.98, 129.98, 131.01, 137.44, 138.88,

N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
245
6.1.1.7. 2-Amino-4-(2-chloro-6-methoxy(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-chlorophenylamino)-1,4,6,7,8-pentahydro quinoline-3-
carbonitrile (5g). IR (KBr, nmax, cmꢀ1): 3425 & 3265 (asym. & sym.
stretching of eNH2), 2200 (C^N stretching), 1660 (C]O stretch-
6.1.1.12. 2-Amino-4-(2,6-dichloro(3-quinolyl))-7,7-dimethyl-5-oxo-1-
(4-methylphenylamino)-1,4,6,7,8-pentahydro quinoline-3-carbonitrile
(5l). IR (KBr,
max, cmꢀ1): 3420 & 3280 (asym. & sym. stretching
of -NH2), 2200 (C^N stretching), 1660 (C]O stretching), 750 (CeCl
stretching). 1H NMR (400 MHz, DMSO-d6):
0.92 (s, 3H, CH3), 0.98
n
ing), 755 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d
0.87 (s,
d
3H, CH3), 0.99 (s, 3H, CH3), 1.99e2.78 (m, 4H, 2 ꢃ CH2), 3.90 (s, 3H,
(s, 3H, CH3), 1.92e2.85 (m, 4H, 2 ꢃ CH2), 2.54 (s, 3H, AreCH3), 5.08
AreOCH3), 5.08 (s, 1H, CH), 6.42e8.12 (m, 10H, AreH and NH2), 8.99
(s, 1H, CH), 6.26e8.26 (m, 10H, AreH and NH2), 8.96 (s, 1H, NH). 13C
(s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d: 27.60, 28.90 (2C, CH3),
NMR (100 MHz, DMSO-d6) d: 21.54 (Ar-CH3), 27.75, 29.04 (2C, CH3),
32.14 (C(CH3)2), 34.54 (C4), 38.00 (CH2), 49.49 (CH2eCO), 56.04
(AreOCH3), 57.21 (CeCN), 105.56, 111.19, 112.55, 120.14, 121.17,
123.37, 129.35, 131.73, 134.36, 136.33, 139.23, 142.20, 147.32, 148.70,
153.10, 154.72, 158.19 (17C, AreC), 194.99 (C]O).
32.19 (C(CH3)2), 34.22 (C4), 37.92 (CH2), 49.50 (CH2-CO), 56.66
(CeCN), 111.40, 113.25, 120.34, 122.36, 124.27, 127.52, 128.02, 130.25,
131.41, 134.66, 138.59, 142.22, 144.46, 148.72, 150.13, 154.25, 155.95
(17C, AreC), 194.98 (C]O).
6.1.1.8. 2-Amino-4-(2,6-dichloro(3-quinolyl))-7,7-dimethyl-5-oxo-1-
(4-chlorophenylamino)-1,4,6,7,8-pentahydro quinoline-3-carbonitrile
(5h). IR (KBr, nmax, cmꢀ1): 3400 & 3280 (asym. & sym. stretching of
eNH2), 2190 (C^N stretching), 1660 (C]O stretching), 755 (CeCl
6.1.1.13. 2-Amino-4-(2-chloro-(3-quinolyl))-7,7-dimethyl-5-oxo-1-
(4-fluorophenylamino)-1,4,6,7,8-pentahydro quinoline-3-carbonitrile
(5m). IR (KBr, nmax, cmꢀ1): 3415 & 3280 (asym. & sym. stretching
of eNH2), 2190 (C^N stretching),1665 (C]O stretching), 750 (CeCl
stretching). 1H NMR (400 MHz, DMSO-d6):
d
0.87 (s, 3H, CH3), 1.00
(s, 3H, CH3),1.99e2.79 (m, 4H, 2 ꢃ CH2), 5.08 (s,1H, CH), 6.42e8.12 (m,
10H, AreH and NH2), 8.99 (s,1H, NH).13C NMR (100 MHz, DMSO-d6)
stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.89 (s, 3H, CH3), 0.96
(s, 3H, CH3), 1.90e2.86 (m, 4H, 2 ꢃ CH2), 5.09 (s, 1H, CH), 6.28e8.22
d:
(m, 11H, AreH and NH2), 8.98 (s, 1H, NH). 13C NMR (100 MHz,
27.62, 28.99 (2C, CH3), 32.33 (C(CH3)2), 34.45 (C4), 37.93 (CH2), 49.42
(CH2eCO), 56.72 (CeCN), 110.43, 111.49,120.21, 126.81, 130.09,131.30,
131.72, 132.11, 134.35, 136.71, 140.03, 144.66, 148.53, 148.59, 150.55,
153.35, 155.09 (17C, AreC), 195.03 (C]O); MS (m/z): 539 (Mþ).
DMSO-d6) d: 27.72, 28.94 (2C, CH3), 32.27 (C(CH3)2), 34.30 (C4),
37.95 (CH2), 49.54 (CH2-CO), 56.42 (CeCN), 111.21, 113.35,
117.05, 120.21, 122.82, 124.60, 126.77, 128.52, 128.96, 130.06, 131.68,
138.46, 144.37, 147.21, 150.61, 155.63, 156.34 (17C, AreC), 195.05
(C]O).
6.1.1.9. 2-Amino-4-(2-chloro-(3-quinolyl))-7,7-dimethyl-5-oxo-1-(4-
methylphenylamino)-1,4,6,7,8-pentahydro quinoline-3-carbonitrile
(5i). IR (KBr, nmax, cmꢀ1): 3405 & 3275 (asym. & sym. stretching
of eNH2), 2200 (C^N stretching), 1655 (C]O stretching), 750
6.1.1.14. 2-Amino-4-(2-chloro–6-methyl(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-fluorophenylamino)-1,4,6,7,8-pentahydro
quinoline-3-
carbonitrile (5n). IR (KBr, nmax, cmꢀ1): 3410 & 3290 (asym. & sym.
stretching of eNH2), 2200 (C^N stretching), 1660 (C]O stretch-
(CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.93 (s, 3H, CH3),
1.00 (s, 3H, CH3), 1.98e2.79 (m, 4H, 2 ꢃ CH2), 2.54 (s, 3H, AreCH3),
ing), 755 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.89 (s,
5.05 (s, 1H, CH), 6.38e8.20 (m, 11H, AreH and NH2), 8.92 (s, 1H,
3H, CH3), 0.98 (s, 3H, CH3), 1.88e2.85 (m, 4H, 2 ꢃ CH2), 2.55 (s, 3H,
NH). 13C NMR (100 MHz, DMSO-d6)
d: 21.57 (AreCH3), 27.72, 29.07
Ar-CH3), 5.05 (s, 1H, CH), 6.25e8.28 (m, 10H, Ar-H and NH2), 8.93 (s,
(2C, CH3), 32.14 (C(CH3)2), 34.60 (C4), 37.90 (CH2), 49.47 (CH2eCO),
57.41 (CeCN), 111.22, 112.87, 120.41, 126.32, 127.06, 128.26, 129.41,
130.01, 131.25, 132.53, 134.63, 136.74, 144.06, 146.32, 148.02, 150.85,
155.22 (17C, AreC), 195.00 (C]O).
1H, NH). 13C NMR (100 MHz, DMSO-d6)
d: 21.61 (AreCH3), 27.78,
28.95 (2C, CH3), 32.21 (C(CH3)2), 34.39 (C4), 38.00 (CH2), 49.45
(CH2-CO), 56.56 (CeCN), 111.63, 113.20, 117.81, 120.25, 124.35,
126.85, 128.70, 130.22, 132.48, 134.42, 136.92, 141.59, 144.61, 150.34,
152.73, 155.55, 157.16 (17C, AreC), 194.92 (C]O).
6.1.1.10. 2-Amino-4-(2-chloro-6-methyl(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-methylphenylamino)-1,4,6,7,8-pentahydro quinoline-3-
carbonitrile (5j). IR (KBr, nmax, cmꢀ1): 3400 & 3285 (asym. & sym.
stretching of eNH2), 2210 (C^N stretching), 1660 (C]O stretch-
6.1.1.15. 2-Amino-4-(2-chloro–6-methoxy(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-fluorophenylamino)-1,4,6,7,8-pentahydro
quinoline-3-
carbonitrile (5o). IR (KBr, nmax, cmꢀ1): 3405 & 3275 (asym. & sym.
stretching of eNH2), 2210 (C^N stretching), 1665 (C]O stretching),
ing), 760 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.89 (s,
3H, CH3), 0.98 (s, 3H, CH3), 1.96e2.82 (m, 4H, 2 ꢃ CH2), 2.52 (s, 3H,
745 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.91 (s, 3H,
AreCH3), 2.56 (s, 3H, AreCH3), 5.09 (s, 1H, CH), 6.35e8.21 (m, 10H,
CH3), 0.98 (s, 3H, CH3), 1.89e2.85 (m, 4H, 2 ꢃ CH2), 3.91 (s, 3H,
AreH and NH2), 8.94 (s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d:
AreOCH3), 5.08 (s, 1H, CH), 6.32e8.37 (m, 10H, AreH and NH2), 8.91
21.57 (AreCH3), 21.59 (AreCH3), 27.74, 28.96 (2C, CH3), 32.18
(C(CH3)2), 34.41 (C4), 38.00 (CH2), 49.52 (CH2eCO), 56.41 (CeCN),
111.06, 112.72, 120.66, 124.21, 126.30, 128.85, 130.06, 130.95,
132.46, 134.52, 136.34, 142.08, 144.18, 146.23, 148.52, 150.02, 155.71
(17C, AreC), 194.95 (C]O).
(s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d: 27.74, 29.05 (2C, CH3),
32.29 (C(CH3)2), 34.39 (C4), 37.98 (CH2), 49.50 (CH2eCO), 55.92
(AreOCH3), 56.50 (CeCN), 105.80, 112.01, 113.23, 117.58, 120.56,
124.44, 128.12, 132.48, 135.23, 138.10, 140.83, 143.15, 146.33, 150.22,
152.25, 155.46, 156.76 (17C, AreC), 194.99 (C]O); MS (m/z): 518
(Mþ).
6.1.1.11. 2-Amino-4-(2-chloro-6-methoxy(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-methylphenylamino)-1,4,6,7,8-pentahydro quinoline-3-
carbonitrile (5k). IR (KBr, nmax, cmꢀ1): 3405 & 3275 (asym. & sym.
stretching of eNH2), 2205 (C^N stretching), 1665 (C]O stretch-
6.1.1.16. 2-Amino-4-(2,6-dichloro(3-quinolyl))-7,7-dimethyl-5-oxo-1-
(4-fluorophenylamino)-1,4,6,7,8-pentahydro quinoline-3-carbonitrile
(5p). IR (KBr, nmax, cmꢀ1): 3415 & 3280 (asym. & sym. stretching of
eNH2), 2195 (C^N stretching), 1655 (C]O stretching), 755 (CeCl
ing), 745 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.90 (s,
3H, CH3), 1.00 (s, 3H, CH3), 1.95e2.84 (m, 4H, 2 ꢃ CH2), 2.54 (s, 3H,
stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.89 (s, 3H, CH3), 1.00
AreCH3), 3.89 (s, 3H, AreOCH3), 5.06 (s,1H, CH), 6.30e8.24 (m,10H,
(s, 3H, CH3), 1.87e2.84 (m, 4H, 2 ꢃ CH2), 5.08 (s, 1H, CH), 6.30e8.33
AreH and NH2), 8.92 (s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d:
(m, 10H, AreH and NH2), 8.97 (s, 1H, NH). 13C NMR
21.55 (AreCH3), 27.70, 29.01 (2C, CH3), 32.11 (C(CH3)2), 34.37 (C4),
38.01 (CH2), 49.47 (CH2eCO), 55.14 (AreOCH3), 56.22 (CeCN),
105.21, 111.32, 113.56, 120.24, 121.69, 128.11, 130.02, 130.91, 132.42,
134.53, 137.22, 142.55, 145.21, 148.28, 150.32, 155.36, 156.03 (17C,
AreC), 194.90 (C]O); MS (m/z): 514 (Mþ).
(100 MHz, DMSO-d6) d: 27.79, 29.08 (2C, CH3), 32.25 (C(CH3)2), 34.42
(C4), 38.09 (CH2), 49.52 (CH2eCO), 56.38 (CeCN), 111.27, 113.42,
117.56, 120.21, 124.32, 126.18, 128.10, 130.26, 132.84, 134.45,
138.30, 141.05, 144.59, 146.72, 151.24, 155.76, 156.19 (17C, AreC),
194.96 (C]O).

246
N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
6.1.1.17. 2-Amino-4-(2-chloro-(3-quinolyl))-7,7-dimethyl-5-oxo-1-
6.1.1.22. 2-Amino-4-(2-chloro-6-methyl(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-bromophenylamino)-1,4,6,7,8-pentahydro quinoline-3-
(4-methoxyphenylamino)-1,4,6,7,8-pentahydro
quinoline-3-
carbonitrile (5q). IR (KBr, nmax, cmꢀ1): 3410 & 3280 (asym. & sym.
stretching of eNH2), 2205 (C^N stretching), 1660 (C]O stretch-
carbonitrile (5v). IR (KBr, n
max, cmꢀ1): 3405 & 3275 (asym. &
sym. stretching of -NH2), 2210 (C^N stretching), 1665 (C]O
ing), 740 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.92 (s,
stretching), 750 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
3H, CH3), 0.99 (s, 3H, CH3), 1.89e2.82 (m, 4H, 2 ꢃ CH2), 3.86 (s, 3H,
d
0.90 (s, 3H, CH3), 0.98 (s, 3H, CH3), 1.83e2.82 (m, 4H, 2 ꢃ CH2),
2.55 (s, 3H, AreCH3), 5.08 (s, 1H, CH), 6.31e8.36 (m, 10H, AreH and
NH2), 8.95 (s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
: 21.65
AreOCH3), 5.04 (s, 1H, CH), 6.28e8.31 (m, 11H, AreH and NH2), 8.89
(s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d: 27.75, 29.00 (2C, CH3),
d
32.36 (C(CH3)2), 34.29 (C4), 37.95 (CH2), 49.52 (CH2eCO), 55.93
(AreOCH3), 56.41 (CeCN), 111.35. 113.21, 115.45, 117.66, 125.11,
126.34, 126.82, 127.44, 128.37, 130.12, 136.63, 140.23, 144.56, 146.77,
150.46, 154.85, 157.03 (17C, AreC), 194.92 (C]O).
(AreCH3), 27.72, 29.01 (2C, CH3), 32.39 (C(CH3)2), 34.22 (C4), 37.99
(CH2), 49.48 (CH2eCO), 56.51 (CeCN), 111.10, 113.26, 118.14, 120.41,
124.55, 126.32, 128.74, 130.24, 132.67, 134.79, 136.90, 142.41, 144.12,
146.69. 150.63, 154.28, 156.88 (17C, AreC), 194.99 (C]O).
6.1.1.18. 2-Amino-4-(2-chloro-6-methyl(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-methoxyphenylamino)-1,4,6,7,8-pentahydro quinoline-3-
6.1.1.23. 2-Amino-4-(2-chloro-6-methoxy(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-bromophenylamino)-1,4,6,7,8-pentahydro quinoline-3-
carbonitrile (5r). IR (KBr,
n
max, cmꢀ1): 3410 & 3270 (asym. &
carbonitrile (5w). IR (KBr, n
max, cmꢀ1): 3410 & 3285 (asym. &
sym. stretching of -NH2), 2200 (C^N stretching), 1650 (C]O
sym. stretching of -NH2), 2200 (C^N stretching), 1665 (C]O
stretching), 740 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
stretching), 745 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d
0.91 (s, 3H, CH3), 0.98 (s, 3H, CH3), 1.87e2.86 (m, 4H, 2 ꢃ CH2),
d
0.91 (s, 3H, CH3),1.00 (s, 3H, CH3),1.82e2.85 (m, 4H, 2 ꢃ CH2), 3.91
(s, 3H, AreOCH3), 5.05 (s, 1H, CH), 6.34e8.38 (m, 10H, AreH and
NH2), 8.90 (s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
: 27.71, 28.95
2.53 (s, 3H, AreCH3), 3.86 (s, 3H, AreOCH3), 5.08 (s, 1H, CH),
6.26e8.34 (m, 10H, AreH and NH2), 8.91 (s, 1H, NH). 13C NMR
d
(100 MHz, DMSO-d6)
d
: 21.63 (AreCH3), 27.76, 28.98 (2C, CH3),
(2C, CH3), 32.35 (C(CH3)2), 34.28 (C4), 37.97 (CH2), 49.50 (CH2eCO),
55.86 (AreOCH3), 56.49 (CeCN), 105.55, 111.14, 113.56, 117.85,
120.64, 125.63, 128.51, 130.21, 132.45, 134.57, 136.34, 140.50, 144.26,
147.89, 150.55, 156.39, 157.80 (17C, AreC), 195.00 (C]O).
32.30 (C(CH3)2), 34.24 (C4), 37.93 (CH2), 49.55 (CH2eCO), 55.91
(AreOCH3), 56.60 (CeCN), 111.26, 113.64, 117.61, 120.29, 124.52,
126.33, 128.72, 130.66, 132.47, 134.82, 138.61, 140.25, 144.79, 146.13,
150.28, 155.61, 156.89 (17C, AreC), 194.95 (C]O).
6.1.1.24. 2-Amino-4-(2,6-dichloro(3-quinolyl))-7,7-dimethyl-5-oxo-1-
(4-bromophenylamino)-1,4,6,7,8-pentahydro quinoline-3-carbonitrile
(5x). IR (KBr, nmax, cmꢀ1): 3405 & 3280 (asym. & sym. stretching of
eNH2), 2195 (C^N stretching), 1660(C]O stretching), 740 (CeCl
6.1.1.19. 2-Amino-4-(2-chloro-6-methoxy(3-quinolyl))-7,7-dimethyl-
5-oxo-1-(4-methoxyphenylamino)-1,4,6,7,8-pentahydro quinoline-3-
carbonitrile (5s). IR (KBr, n
max, cmꢀ1): 3405 & 3280 (asym. &
sym. stretching of -NH2), 2200 (C^N stretching), 1650 (C]O
stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.92 (s, 3H, CH3), 1.01
stretching), 755 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
(s, 3H, CH3), 1.84e2.86 (m, 4H, 2 ꢃ CH2), 5.10 (s, 1H, CH), 6.32e8.39
d
0.86 (s, 3H, CH3), 1.01 (s, 3H, CH3), 1.86e2.84 (m, 4H, 2 ꢃ CH2),
(m,10H, AreH and NH2), 8.95 (s,1H, NH).13C NMR (100 MHz, DMSO-
3.86 (s, 3H, AreOCH3), 3.91 (s, 3H, AreOCH3), 5.07 (s, 1H, CH),
d6) d: 27.78, 29.06 (2C, CH3), 32.30 (C(CH3)2), 34.32 (C4), 38.07
6.34e8.39 (m, 10H, AreH and NH2), 8.96 (s, 1H, NH). 13C NMR
(CH2), 49.52 (CH2eCO), 56.57 (CeCN), 111.22, 113.52, 117.63, 120.21,
124.52, 126.38, 128.95, 130.25, 132.65, 134.84, 136.80, 142.50, 144.78,
146.31, 151.25. 154.58. 156.95 (17C, AreC), 195.05 (C]O); MS (m/z):
583 (Mþ).
(100 MHz, DMSO-d6) d: 27.71, 28.93 (2C, CH3), 32.31 (C(CH3)2),
34.30 (C4), 38.03 (CH2), 49.54 (CH2eCO), 55.88 (AreOCH3), 55.91
(AreOCH3), 56.56 (CeCN), 105.21, 111.73, 113.65, 117.81, 120.67,
124.56, 128.95, 131.16, 136.52, 138.39, 140.44, 142.66. 146.37, 150.23,
151.38, 154.62, 156.77 (17C, AreC), 194.91 (C]O).
6.2. Biological assay
6.1.1.20. 2-Amino-4-(2,6-dichloro(3-quinolyl))-7,7-dimethyl-5-oxo-
6.2.1. In vitro evaluation of antimicrobial activity
1-(4-
methoxyphenylamino)-1,4,6,7,8-pentahydro
quinoline-3-
The MICs of synthesized compounds were carried out by broth
microdilution method. DMSO was used as diluents to get desired
concentration of compounds to test upon standard bacterial strains.
Serial dilutions were prepared in primary and secondary screening.
The control tube containing no antibiotic was immediately sub
cultured (before inoculation) by spreading a loopful evenly over
a quarter of plate of medium suitable for the growth of the test
organism and put for incubation at 37 ꢁC overnight. The tubes were
then incubated overnight. The MIC of the control organism was
read to check the accuracy of the compound concentrations. The
MIC was defined as the lowest concentration of the antibiotic or
test sample allowing no visible growth. All the tubes not showing
visible growth (in the same manner as control tube described
above) was sub cultured and incubated overnight at 37 ꢁC. The
amount of growth from the control tube before incubation (which
represents the original inoculum) was compared. Subcultures
might show: similar number of colonies indicating bacteriostatic;
a reduced number of colonies indicating a partial or slow bacteri-
cidal activity and no growth if the whole inoculum has been killed.
The test must include a second set of the same dilutions inoculated
with an organism of known sensitivity. Each synthesized
carbonitrile (5t). IR (KBr,
n
max, cmꢀ1): 3415 & 3275 (asym. & sym.
stretching of -NH2), 2195 (C^N stretching), 1665 (C]O stretching),
750 (CeCl stretching). 1H NMR (400 MHz, DMSO-d6):
d
0.88 (s, 3H,
CH3), 0.98 (s, 3H, CH3), 1.84e2.86 (m, 4H, 2 ꢃ CH2), 3.90 (s, 3H,
AreOCH3), 5.04 (s, 1H, CH), 6.31e8.35 (m, 10H, AreH and NH2), 8.88
(s, 1H, NH). 13C NMR (100 MHz, DMSO-d6)
d: 27.75, 28.99 (2C, CH3),
32.27 (C(CH3)2), 34.22 (C4), 38.01 (CH2), 49.55 (CH2eCO), 55.93
(AreOCH3), 56.41 (CeCN), 111.22, 113.56, 117.48, 120.54, 124.36,
128.82, 130.16, 131.65, 132.72, 134.28, 136.12, 144.21, 146.07, 150.34,
151.39, 154.46, 157.14 (17C, AreC), 194.96 (C]O).
6.1.1.21. 2-Amino-4-(2-chloro-(3-quinolyl))-7,7-dimethyl-5-oxo-1-
(4-bromophenylamino)-1,4,6,7,8-pentahydro quinoline-3-carbonitrile
(5u). IR (KBr, nmax, cmꢀ1): 3410 & 3290 (asym. & sym. stretching of
eNH2), 2200 (C^N stretching), 1660 (C]O stretching), 755 (C-Cl
stretching). 1H NMR (400 MHz, DMSO-d6):
d 0.87 (s, 3H, CH3), 0.97
(s, 3H, CH3), 1.82e2.84 (m, 4H, 2 ꢃ CH2), 5.02 (s, 1H, CH), 6.37e8.32
(m, 11H, AreH and NH2), 8.92 (s, 1H, NH). 13C NMR (100 MHz,
DMSO-d6) d: 27.78, 29.05 (2C, CH3), 32.31 (C(CH3)2), 34.37 (C4),
37.97 (CH2), 49.50 (CH2eCO), 56.44 (CeCN), 111.44, 113.25, 118.51,
120.36, 125.85, 126.71, 128.16, 130.17, 133.67, 135.46, 136.11, 143.21,
145.78, 147.88, 150.26, 154.37, 156.62 (17C, AreC), 195.01 (C]O).
compound was diluted obtaining 2000
mg/mL concentration as
a stock solution. In primary screening 500, 250 and 200
mg/mL

N.M. Shah et al. / European Journal of Medicinal Chemistry 54 (2012) 239e247
247
concentrations of the synthesized compounds were taken. The
Acknowledgement
active synthesized compounds found in this primary screening
were further tested in a second set of dilution against all microor-
ganisms. The compounds found active in primary screening were
similarly diluted to obtain 100, 62.5, 50 and 25 mg/mL concentra-
tions. The highest dilution showing at least 99% inhibition is taken
as MIC.
The authors are thankful to Head, Department of Chemistry,
Sardar Patel University for providing 1H NMR and 13C NMR spec-
troscopy and research facilities. We are also thankful to Oxygen
Healthcare Research Pvt. Ltd., Ahmedabad, for providing mass
spectroscopy facilities, Vaibhav Laboratories, Ahmedabad, Gujarat,
India for the FTeIR, SICART, Vallabh Vidyanagar, for elemental
analysis and Dhanji P. Rajani, Microcare Laboratory, Surat, Gujarat,
India for antimicrobial and antimycobacterial screening of the
compounds reported herein. One of the authors is grateful to UGC,
New Delhi for Research Fellowship in Sciences for Meritorious
Students.
6.2.2. In vitro evaluation of antituberculosis activity
A primary screen was conducted at 6.25
tuberculosis H37Rv by LowensteineJensen (LJ) MIC method [28]
where primary 6.25 g/ml dilution of each test compound were
mg/ml against M.
m
added to liquid LowensteineJensen Medium and then media were
sterilized by inspissation method. A culture of M. tuberculosis
H37Rv growing on LowensteineJensen Medium was harvested in
0.85% saline in bijou bottles. DMSO was used as vehicle to get
desired concentration. These tubes were then incubated at 37 ꢁC for
24 h followed by streaking of M. tuberculosis H37Rv (5 ꢃ 104 bacilli
per tube). These tubes were then incubated at 37 ꢁC. Growth of
bacilli was seen after 12, 22, and finally 28 days of incubation. Tubes
having the compounds were compared with control tubes where
medium alone was incubated with M. tuberculosis H37Rv. The
concentration at which complete inhibition of colonies occurred
was taken as active concentration of test compound. The standard
strain M. tuberculosis H37Rv was tested with known drug Iso-
niazide and Rifampicin. The screening results are summarized as %
inhibition relative to standard drug Isoniazide and Rifampicin.
Compounds effecting <90% inhibition in the primary screen were
not evaluated further. Compounds demonstrating at least 90%
inhibition in the primary screen were re-tested at lower concen-
tration (MIC) in a LowensteineJensen medium. The protocols were
summarized in Table 3.
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blood smears were read blind.