Synthesis and bio-evaluation of alkylaminoaryl phenyl cyclopropyl methanones as antitubercular and antimalarial agents

Article abstract of DOI:10.1016/j.bmc.2010.09.071

A series of 4-alkylaminoaryl phenyl cyclopropyl methanones (6a-6u and 8a-8c) were synthesized from 4-fluorochalcones (3a and 3b) by cyclopropanation of double bond followed by nucleophilic substitution of F with different amines. The compounds were screened for their antitubercular and antimalarial activities against Mycobacterium tuberculosis H37Rv and Plasmodium falciparum 3D7 strains in vitro respectively. Several compounds (6a, 6d-6h, 6p, 6q and 8a-8c) exhibited good in vitro antitubercular activities with MIC values 3.12-12.5 μg/mL and preferentially inhibited the growth of P. falciparum in vitro (4a, 4c, 6a-6d, 6f, 6s, 8a and 8c) with IC₅₀ as low as 0.080 and 0.035 μg/mL and SI values 4975 and 6948, respectively. Molecular docking studies and in vitro evaluation against FAS-II enzymes using reporter gene assays were carried out to elucidate the mode of action of these molecules. Two compounds 4a and 6g showed significant inhibition at 25 μM concentration of the compound.

Full text of DOI:10.1016/j.bmc.2010.09.071

Bioorganic & Medicinal Chemistry 18 (2010) 8289–8301
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and bio-evaluation of alkylaminoaryl phenyl cyclopropyl
methanones as antitubercular and antimalarial agents
Arya Ajay a, Vandana Singh b, Shubhra Singh c, Swaroop Pandey d, Sarika Gunjan d, Divya Dubey e,
Sudhir Kumar Sinha c, Bhupendra N. Singh b, Vinita Chaturvedi c, Renu Tripathi d,
Ravishankar Ramchandran e, Rama P. Tripathi a,
⇑
a Medicinal and Process Chemistry Division, Central Drug Research Institute Chattar Manzil, PO Box 173, Mahatma Gandhi Marg, Lucknow 226001, India
b Microbiology Division, Central Drug Research Institute Chattar Manzil, PO Box 173, Mahatma Gandhi Marg, Lucknow 226001, India
c Drug Target Discovery and Development Divison, Central Drug Research Institute Chattar Manzil, PO Box 173, Mahatma Gandhi Marg, Lucknow 226001, India
d Parasitology Division, Central Drug Research Institute Chattar Manzil, PO Box 173, Mahatma Gandhi Marg, Lucknow 226001, India
e Molecular and Structural Biology Division, Central Drug Research Institute Chattar Manzil, PO Box 173, Mahatma Gandhi Marg, Lucknow 226001, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of 4-alkylaminoaryl phenyl cyclopropyl methanones (6a–6u and 8a–8c) were synthesized from
4-fluorochalcones (3a and 3b) by cyclopropanation of double bond followed by nucleophilic substitution
of F with different amines. The compounds were screened for their antitubercular and antimalarial activ-
ities against Mycobacterium tuberculosis H37Rv and Plasmodium falciparum 3D7 strains in vitro respec-
tively. Several compounds (6a, 6d–6h, 6p, 6q and 8a–8c) exhibited good in vitro antitubercular
Received 8 September 2010
Revised 28 September 2010
Accepted 30 September 2010
Available online 14 October 2010
activities with MIC values 3.12–12.5
in vitro (4a, 4c, 6a–6d, 6f, 6s, 8a and 8c) with IC50 as low as 0.080 and 0.035
4975 and 6948, respectively. Molecular docking studies and in vitro evaluation against FAS-II enzymes
using reporter gene assays were carried out to elucidate the mode of action of these molecules. Two com-
l
g/mL and preferentially inhibited the growth of P. falciparum
g/mL and SI values
Keywords:
Cyclopropylphenyl methanones
Antitubercular
Antimalarial
FAS-II
l
pounds 4a and 6g showed significant inhibition at 25
lM concentration of the compound.
Ó 2010 Elsevier Ltd. All rights reserved.
Mycobacterium tuberculosis H37Rv
Plasmodium falciparum 3D7
1. Introduction
Nearly all the fatal cases are caused by the malaria parasite Plasmo-
dium falciparum.16 The widespread resistance of malaria parasite to
most common antimalarials17,18 (chloroquine, pyrimethamine),
and cross resistance to structurally unrelated drugs makes the cur-
rent situation more worsening.19–21 Artemisinine or their struc-
tural analogs are potent antimalarial22,23 but these are associated
with the problem of high cost24 and recently reported resistance
to artesunate in Pailin and Western Combodia.25 Moreover, the ad-
verse effect of co-infection with HIV and malaria is become
increasingly apparent.26 Therefore, new agents that will (i) shorten
the current TB treatment, (ii) compatible with HIV drugs and (iii)
have novel target with high selectivity for both of diseases are
needed.
Mycobacterium tuberculosis and P. falciparum allocate enzymatic
components of the type II fatty acid biosynthetic pathway
(FAS-II)27,28 for their fatty acid biosynthesis. In Mycobacterium,
FAS-II pathway comprises a group of enzymes as MabA (FabG),
InhA (FabI), FabH as well as Kas A or Kas B which sequentially
catalyses the reduction and condensation steps of fatty acid
synthesis.29,30 Among these enzymes InhA and Kas A or Kas B is
the most important target for tuberculosis chemotherapy. For
Plasmodium, FAS-II is the only fatty acid pathway whose enzy-
matic components reside at a unique plastid organelle known as
The current arsenal of antiinfective agents in our hand for the
treatment of tuberculosis and malaria is insufficient to protect us
over the long term. With more than 1.6 million deaths and 9.2 mil-
lion new cases being reported each year, tuberculosis is a leading
infectious disease claiming millions of death today, mostly in
developing countries harboring latent TB infections.1,2 HIV epi-
demics in many countries have led to the emergence of new waves
of TB infection as co-infection with HIV and mycobacteria render
latent TB to an active condition, often with fatal consequences.3,4
The current cocktail of antitubercular drug regimens can achieve
more than 99% efficacy but this is often reduced due to alarming
growth of MDR and XDR-TB.5–9 Although a number of lead mole-
cules exist today10–12 to develop new drugs but no new chemical
entity has emerged for clinical use over the last 40 years. In con-
junction with tuberculosis, malaria is also a devastating infectious
disease having high morbidity and mortality rate with 300–500
million clinical cases and 1.5–2.7 million deaths every year.13–15
⇑
Corresponding author. Tel.: +91 0522 2612411; fax +91 522 2623405/2623938/
2629504.
E-mail address: rpt.cdri@gmail.com (R.P. Tripathi).
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.09.071

8290
A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
‘apicoplast’ and essential for development of the parasite.28 An-
other most important metabolic pathway, the isoprenoid pathway
DOXP is also known in apicoplast.31 Both the DOXP and FAS-II
pathways are valuable drug targets in antiinfective drug develop-
ment since these two pathways are absent in the human host.32
Out of several inhibitors of the FAS-II and DOXP, triclosan (I)33
and fosmidomycin34 are very important (Fig. 1). Simple acetophe-
various amines led to the formation of targeted compounds
6b–6f, 6i, 6k–6m and 6o–6u in quantitative yields (Scheme 1).
In order to see the effect of substitutents in the alkylamine
chain we have prepared another series of compounds starting from
the above compound 6s having 2-hydroxyethylamino chain at the
4th position of one of the aromatic ring A. Thus 6s was reacted with
MeSO2Cl in presence of TEA in CH2Cl2 at 0 °C to give respective 2-
methanesulphonyloxyethyl amino derivative (7). The nucleophilic
substitution of –SO2Me group in compound 7 with several azoles
in presence of NaH/DMF at 0–120 °C gave respective 2-(azolyl)-
ethylaminoaryl phenyl cyclopropyl methanones (8a–8c) in good
yields (Scheme 2).
nones, benzylideneacetophenones and p-nitro-a-acetylamino-
b-hydroxypropiophenones are reported to have good antitubercular
activties.35–38 The cyclopropane derivative of fosmidomycin (II)
(Fig. 1) with limited conformational flexibility in the backbone is
as active as fosmidomycin.39 Moreover, cyclopropyl ring is a com-
mon structural element in the mycobacterial cell wall40 and its
chemotherapeutic importance is also well known.41–43 Aryl cyclo-
propyl ketones play a prominent role as intermediates in the syn-
thesis of many other biologically active compounds.44 As a part of
our ongoing research devoted for the synthesis of antiinfective
agents45–50 we have already identified the aryloxyphenyl cyclopro-
pyl methanones (III)51 (Fig. 1) as potent antitubercular agent.
Keeping in view the above we were prompted to synthesize
4-alkylaminoaryl phenyl cyclopropyl methanones and evaluate
them for anti-mycobacterial and antimalarial activities.
The structures of all the compounds as given in Table 1 were
determined on the basis of their spectroscopic data and microanal-
yses. The IR spectra of the compounds, in general, exhibited the
absorption band at around 3432 cmÀ1 indicating the presence of
aromatic amino group, absorption band of carbonyl group at
around 1638–1640 cmÀ1. The ESMS (mass spectra) of the com-
pounds showed [M+H]+ peaks corresponding to their molecular
formulae. The 1H and 13C NMR spectra are consistent with the pro-
posed structures. The 1H NMR spectrum of a prototype molecule
6g, displayed exchangeable N–H proton signal as a singlet at d
5.39. The four protons of cyclopropyl ring were visible at four dif-
ferent chemical shifts as multiplets at d 2.76–2.71, 2.60–2.53, 1.85–
1.76 and 1.40–1.34 ppm. The aromatic protons appeared as dou-
blet at d 7.83, 7.23, 7.07 and 6.51 ppm with coupling constant in
range of 8.28–8.58 Hz. The protons of alkylamino chain were visi-
ble at d 3.25, 2.42 as triplet and at d 1.85–1.76 as multiplet along
with one proton of cyclopropyl ring. The protons of the N,N-
dimethyl group were observed as a singlet at d 2.25 ppm. The
geometry of the parent chalcone was found to be trans from the
coupling constant between the olefinic protons of the propenone
moiety (compound 4a, J = 15.8 Hz) and the TMSOI mediated cyclo-
propanation (Corey–Chaykovsky reaction) leads the trans product
with retained stereochemistry as evident from the litrature.54–56
In the 13C NMR spectrum, the only carbonyl carbon appeared at d
195.6, while the quaternary aromatic carbon attached to the amine
group was observed at d 153.0. Three carbons of cyclopropyl ring
were visible at d 28.7, 28.2 and 18.7 ppm. The other aromatic qua-
ternary carbons (ArC) were observed at their usual chemical shifts
of d 140.1, 132.3 and 126.8 ppm, while sp2 aromatic carbons
(ArCH) appeared at d 131.0, 128.9, 127.9 and 111.7 ppm. Almost
similar patterns were observed in 1H NMR and 13C NMR spectra
of other compounds (8a–8c) of the series.
2. Results and discussion
2.1. Chemistry
The protocol for the synthesis of 4-alkylamino substituted aryl
phenyl cyclopropyl methanones begins with the synthesis of chal-
cones (3a and 3b).52,53 The aldol reaction between 4-fluoro aceto-
phenone (1) and substituted aromatic aldehydes (2a and 2b)
followed by in situ dehydration, leads the desired chalcones (3a
and 3b) in quantitative yields. There are three possible pathways
to synthesize the title compounds as shown in Figure 2. The first
one we discarded as amines may react with ketones and a complex
mixture would be obtained (Path 1). The second approach (Path 2)
involves the ipso nucleophilic substitution at 4-fluoro group in one
of the phenyl ring of chalcone 3a by various amines followed by
cyclopropanation. The third approach (Path 3) involves the cylo-
propanation of double bond in chalcones followed by ipso nucleo-
philic substitution at 4-fluoro group in one of the phenyl ring of
chalcone 3a to give the title molecules.
The ipso nucleophilic substitution at 4-fluoro group in one of the
phenyl ring of chalcone 3a by various amines in presence of K2CO3/
DMF at 120 °C in an inert atmosphere of nitrogen gave the desired
alkylamino substituted chalcones (4a–4e) in moderate yields only
(Scheme 1). The unsatisfactory yield of the products may possibly
be due to Michael addition product as we could not isolate the
other products of the reaction observed during reaction (TLC).
The reaction of the alkylamino substituted chalcones with trim-
ethylsulphoxonium iodide (TMSOI) and 50% aq NaOH at 80 °C in
presence of tetrabutylammonium bromide as phase transfer cata-
lyst and CH2Cl2 as solvent gave the title compounds (6a, 6g, 6h,
6j and 6n) (Scheme 1). Therefore, the third approach was thought
wherein the fluorochalcones 3a and 3b were first subjected to
cyclopropanation using TMSOI as described above to give the
respective cycloproyl derivatives 5a and 5b in good yields. The
latter on ipso nucleophilic substitution of 4-fluoro group using
2.2. Biological activities
All of the above synthesized compounds including few of the
intermediate chalcones were screened for their antitubercular and
antimalarial activities against M. tuberculosis H37Rv and
P. falciparum 3D7 strains in vitro. The antitubercular screening was
carried out in vitro using agar microdilution method57 while the
antimalarial activity was assayed as per earlier reported
protocols.58,61
2.2.1. Antitubercular activity
Antitubercular screening results of all the synthesized com-
pounds and few of the intermediate cahlcones are listed in Table
O
OH
Cl
O
P
OH
OH
N
O
Na
O
O
(III)
O
Cl
Cl
(II)
(I)
O
Figure 1. Chemical structure of some FAS-II inhibitor and biologically active cyclopropane ring containing molecules.

A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
8291
O
complex mixture may formed
Different aldehyde
Z
1
-
O
h
t
a
O
P
)
(Z
Alkyl amines
F
Different aldehyde
O
Z
F
R
R
Path-2
Path-3
O
O
F
R
Z
R
Figure 2. Possible pathways for the formation of alkylaminoaryl phenyl cyclopropyl methanones.
O
(iii)
Path-2
(ii)
O
O
Z
R
CHO
4a-4e: R= 4-Cl
(i)
O
O
F
F
R
R
(iii)
(ii)
2a, R= 4-Cl
2b, R= 3,4 diOMe
1
3a, R= 4-Cl
Path-3
F
Z
R
R
3b, R= 3,4diOMe
6a-6s: R= 4-Cl
6t,6u: R= 3,4 DiOMe
5a: R= 4-Cl
5b: R= 3,4 DiOMe
Scheme 1. Synthesis of 4-alkylaminoaryl phenyl cyclopropyl methanones (6a–6u). Reagents and conditions: (i) KOH, EtOH, rt; (ii) different amines (Z), K2CO3, DMF, 120 °C,
inert atmosphere; (iii) TMSOI, TBAB (20 mol %), CH2Cl2:50% aq NaOH (1:1), 80 °C.
O
O
O
(ii)
(i)
X'
X''
X
N
HO
MsO
N
H
Cl
N
H
Cl
N
H
Cl
7
6s
8a-8c
X=X''= N and X'= CH2 ;1,2,4 Triazole
X=X'= CH2 and X''= N; Imidazole/
Benzimidazole
Scheme 2. Synthesis of azolylalkylaminoaryl phenyl cyclopropyl methanones (8a–8c). Reagents and conditions: (i) CH3SO2Cl, Et3N, CH2Cl2, 0 °C–rt; (ii) different azoles, NaH,
DMF, 0–120 °C.
2. As evident from the results out of all the alkylaminoaryl phenyl
cyclopropyl methanones screened, compounds 6a, 6d–6h, 6p, 6q
and 8a–8c showed minimum inhibitory concentration in the range
6.25
the imidazole unit reduces the activity as compounds 8b and 8c
have MIC values 12.5 g/mL each. Similar observation was made
with the above compounds having piperazinyl substitutents.
Replacement of N-methyl group (compound 6a, MIC 6.25 g/mL)
in piprazine by aryl group (compound 6e) resulted in loss of
activity with MIC 12.5 g/mL.
Encouraged by recent report of antimalrial and antitubercular
activities in acetylenic chalcones by Kelly’s group59 we were
prompted to see the effect of 4-alkylamino chalcones (4a–4e)
against malaria parasite and M. tuberculosis with a view to find
out the correlation between the unsaturated system and cyclo-
propanated system. The antitubercular screening results did not
offer any definite correlation between activities with the unsatu-
rated or cyclopropanated compounds. It is evident by the fact that,
in compounds 4a and 4c (chalcones having double bond, MIC value
lg/mL. However, introduction of any lipophilic moiety to
l
of 3.12–12.5 lg/mL. Among compounds having 4-chloro and
3,4-dimethoxy substituent in aromatic ring B, compounds with
4-chloro substituent (6a–6s and 8a–8c) were more active than
the compounds with 3,4-dimethoxy substituents (6t and 6u). The
most active compound of the series was found to be compound
l
l
6h having a furfuryl amine substituent with MIC value 3.12 lg/
mL. The compounds 6a and 6g having dialkylamino chain as 4-sub-
stitutent in one of the phenyl rings (ring A), whether it is cyclic or
acyclic have almost similar antitubercular activity with MIC
6.25 lg/mL. On the other hand compounds 6d, 6f and 6p having
monoalkylamine chain, dialkylamine chain and cyclic amine with
C-chain length 6four carbon atoms with comparable lipophilicity
displayed only moderate activity with the same MIC value
12.5
substitutent also displayed only moderate antitubercular activity
(MIC 12.5 g/mL). In the second series of compounds having
2-(azolyl)ethylamino substitutent at 4-position in benzene ring A
showed promising to moderate antitubercular activities. Com-
pound 8a with imidazolyl ethylamino substituent has MIC value
l
g/mL. However, in contrast compounds with the oleylamine
12.5
propanated analogues 6g and 6h (MICs 6.25 and 3.12
respectively. On the contrary, the chalcones 4b and 4d with MIC
value 3.12 and 6.25 g/mL are better antitubercular as compared
to their cyclopropanated analogues 6a and 6j with MICs values
6.25 and >12.5 g/mL, respectively.
l
g/mL each) the activity is less as compared to their cyclo-
lg/mL),
l
l
l

8292
A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
Table 1
Compound 6c was also found to be a potent antimalarial agent
with IC50 0.08 g/mL and very low MIC (0.3 g/mL). Majority of
Synthesis of alkylamino substituted chalcones (4a–4e) and alkylaminoaryl phenyl
cyclopropyl methanones (6a–6u and 8a–8c)
l
l
the active compounds have shown good correlation between their
MIC and IC50 values (Table 3). A closure look into structure–activity
relationship of the compounds revealed that compounds with lipo-
philic alkylamino substituent (6b, 6c, 6d and 6f) showed good anti-
malarial activity indicating that the lipophilicity has significant
impact on antimalrial activity. Similarly compounds with basic
alkylamino chain (compounds 4a and 6a) or azole moiety (com-
pounds 8a and 8c) also exhibited potent antimalarial activities. It
seems that basic alkylamino cahin and lipophilicity are important
factors governing the biological activities.
Compound R
No.
Z (amine)
Mp (°C) Yield
(%)
4a
4-Cl
N,N-Dimethyl
63–65 35
aminopropylamine
N-Me piprazine
Furfuryl amine
Morpholine
4b
4c
4d
4e
6a
6b
6c
6d
6e
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
100–102 54
72–74 40
98–100 45
111–113 60
112–114 80
98–100 84
97–99 82
Pipridine
N-Me piprazine
Cyclohexyl amine
Heptyl amine
Butyl amine
4-(2,3-Dichlorophenyl)
piperazin
Dibutyl amine
N,N-Dimethyl
aminopropylamine
Furfuryl amine
1,5-DiMe hexyl amine
Morpholine
Dodecyl amine
Hexadecyl amine
Octyl amine
128–130 81
134–136 77
2.2.3. Cytotoxicity
The cytotoxic evaluation of these compounds was carried out
on VERO cell line as per earlier reported protocol62 and the results
are shown in Table 3. On the basis of antimalarial/antitubercular
and cytotoxic activities, the calculated selectivity indices (SI val-
ues) (CC50/IC50) have shown that all the compounds active against
malaria and tuberculosis have moderate to high degree of safety.
The compound 8a has shown the best selectivity index (6948.57)
as well as potent antimalarial activity. Out of ten antimalarial com-
pounds, nine have shown median cytotoxic concentration to be
6f
6g
4-Cl
4-Cl
152–154 78
94–96 77
6h
6i
4-Cl
4-Cl
96–98 85
128–130 80
115–117 83
92–94 80
6j
4-Cl
6k
6l
4-Cl
4-Cl
122–124 85
154–156 87
118–120 79
132–134 86
138–140 86
153–155 80
140–142 76
166–168 73
156–158 81
147–149 79
110–112 67
6m
6n
6o
6p
6q
6r
6s
6t
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
4-Cl
3,4 Di-OMe
3,4 Di-OMe
4-Cl
Pipridine
N,N-Dimethylamine
Pyrolidine
82.00–404.03 lg/mL (Table 3).
Oleyl amine
2.2.4. FAS-II inhibitory activities
Veratryl amine
Ethanolamine
N-Me piprazine
Heptyl amine
(1H-Imidazol-1-
yl)ethylamine
(1H-1,2,4-Triazol-1-
yl)ethylamine
(1H-Benzo[d]imidazol-1yl)
ethylamine
2.2.4.1. In silico docking studies. To identify the drug target of
the synthesized compounds first the intermediate chalcone 4a
was selected for in silico docking studies.63,64 Furthermore to char-
acterize the proteinaceous drug target which is most likely to
interact with compound 4a, the ligand file was uploaded into Inhib-
itor Identification Tool (Is-it?): (unpublished and implemented in
the host laboratory) server and docking jobs were queued by
selecting all the 85 drug targets from M. tuberculosis enlisted in
the server. The top candidates identified by Is-it?: server, ranked
by interaction energies, included 10 out of the 12 targets from FasII
pathway (Fig. 3). It is thus likely that aryl amino derivatives may
interact with multiple proteins including fabG or 1UZL (star
marked in the Fig. 3), Enoyl-acp reductase (1P44, 2NSD, inhA)
and fabD (2QC3) from FAS-II pathway. The compound also showed
high affinity with another protein Gyrase or gyrB (another red star
marked in the Fig. 3). The target proteins so identified are enlisted
in Supplementary Table. The top potential binding partner identi-
fied is MabA, which is also known as FabG1, a part of FAS-II en-
zyme, catalyzing the NADPH-specific reduction of long chain
beta-ketoacyl derivatives. The compound 4a is predicted to be
interacting with most of the reported important residues of the
NADP binding site with an estimated inhibition constant, Ki value
of 261.41 nM. The chlorine atom of the compound 4a is present
in the vicinity of TYR 153 which is reported to be the part of cata-
lytic triad residues. The molecular interactions of the compound 4a
with MabA are shown in Figure 4.
6u
8a
8b
8c
4-Cl
4-Cl
117–119 68
134–136 70
Table 2
Antitubercular activity of the synthesized molecules 4a–4e, 6a–6u and 8a–8c
S.
No.
Compound
No.
Antitubercular S.
activity No.
Compound
No.
Antitubercular
activity
(MIC lg/mL)
(MIC
l
g/mL)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
4a
4b
4c
4d
4e
6a
6b
6c
6d
6e
6f
12.5
3.12
12.5
6.25
>12.5
6.25
>12.5
>12.5
12.5
12.5
12.5
6.25
3.12
>12.5
>12.5
16
17
18
19
20
21
22
23
24
25
26
27
28
29
6k
6l
In*
In
>12.5
>12.5
In
12.5
12.5
>12.5
>12.5
>12.5
>12.5
6.25
12.5
12.5
0.025
6m
6n
6o
6p
6q
6r
6s
6t
6u
8a
8b
8c
6g
6h
6i
As no ligand variation was done in the final molecule compared
to their parent chalcones, the synthesized final molecules directly
undergoes to biological assay against FAS-II pathway.
6j
Inh*
In* = Insoluble, Inh* = Isoniazid.
2.2.4.2. FAS-II inhibition assay. FAS-II inhibitory activity was as-
sessed using a recombinant non-pathogenic mycobacterial strain,
Mycobacterium aurum, which contains M. tuberculosis kas operon
promoter in fusion with lacZ reporter gene.65 The strain shows con-
tinued expression of reporter gene under the influence of kas oper-
on promoter during basal conditions, while an increased
expression of the reporter gene is noticed only after treatment with
FAS-II pathway inhibitors. The preliminary screening of the com-
pounds shows FAS-II inhibitory activity at two different concentra-
2.2.2. Antimalarial activity
The above alkylamino chalcones and their cyclopropyl deriva-
tives were also screened against malaria parasite P. falciparum
using the earlier reported protocols.58,60,61 Most of the compounds
showed moderate to high antimalarial activity. Compounds 4a, 4c,
6a–6d, 6f, 6s, 8a and 8c were active against the parasite with IC50
in the range of 0.035–0.76
pounds, compound 8a was the most potent antimalarial agent with
MIC and IC50 values of 0.062 and 0.035 g/mL, respectively.
lg/mL. Among all the screened com-
l
tion, 25 and 50 lM and results are listed in Table 4. In present

A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
8293
Table 3
Antimalarial activity and selectivity index of the synthesized molecules 4a–4e, 6a–6u
and 8a–8c
S. No.
Compound No.
MIC
g/mL)
IC50
g/mL)
CC50
(lg/mL)
SI
(
l
(
l
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
4a
4b
4c
4d
4e
6a
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
6m
6n
6o
6p
6q
6r
1.56
50
10
50
50
10
10
0.3
0.6
50
10
50
10
0.64
11.78
0.76
4.86
8.92
0.54
0.75
0.08
0.33
8.82
0.73
2.86
1.71
ND
3.02
10.4
4.57
2.13
6.05
1.43
9.75
75.13
8
5.92
9.25
8.69
531.62
7.33
26.3
151.96
261.57
4975
958.69
43.38
590.04
5.53
102.46
404.03
35.64
234.59
82.06
196.18
398
316.37
382.61
430.73
15.83
255.89
ND
452.76
385.7
409.83
214.14
284.28
343.78
431.49
393.35
421.86
305
149.64
ND
*
ND
10
>50
50
10
10
149.92
37.08
89.68
100.53
46.99
240.4
44.26
5.24
52.73
508.33
37.23
121.67
6948.57
11.7
10
>50
>50
10
2
50
10
0.062
10
2
0.05
6s
6t
6u
8a
8b
8c
0.6
2.09
2.24
0.035
3.41
0.74
0.008
77.81
272.54
243.2
39.9
Figure 4. The molecular interactions of compound 4a within the MabA binding
pocket. The interacting residues are labeled and hydrogen bonds are marked by the
dashed lines.
285.23
385.45
Chloroquine
*
ND = Not determined.
increasing concentration of compound and declined at higher con-
centration owing to killing of cells at exceeding dose of drugs. It
may be noted that we repeated the same experiment using another
M. aurum recombinant strain carrying hsp60 promoter wherein no
inducibility was observed under similar conditions, rather a de-
cline in reporter gene activity was noticed in line with diminishing
viability of bacterial cells (Fig. 5B) as the treatment dose increased.
Isoniazid (INH), a known FAS-II pathway inhibitor was used as a
positive control in both conditions. All the ongoing results of
screening suggest that FAS-II is a viable target of the intermediate
studies molecule 4a was selected for the assessment, the critical
concentration of compound at which it inhibited the growth of
M. aurum culture by P85% was determined by cfu analysis and
then b-gal enzyme assay was performed to assess the inducibility
under treated condition. We observed an induced level of b-gal en-
zyme activity after treatment with compound 4a with respect to
untreated control (Fig. 5A). The activity gradually increased with
Figure 3. The plot of docking energy calculated by AUTODOCK for aryl amino derivative and the protein targets. Only acceptable scores are used for plot. The most potential
binders are 1UZL (a fasII pathway enzyme) and gyrB (Gyrase B) and are denoted by asterisk.

8294
A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
Table 4
reduced pressure. Thin layer chromatography was performed using
Silica Gel 60 F254 plates with detecting agent iodine vapors or by
spraying with dragendorf reagent. Silica gel (60–120 mesh) was
used for column chromatography. Tetramethylsilane (0.0 ppm)
was used as an internal standard in 1H NMR and CDCl3
(77.0 ppm) was used in 13C NMR. The abbreviations used to indi-
cate the peak multiplicity were; s, singlet; br s, broad singlet; d,
doublet; dd, double doublet; t, triplet; q, quartet; m, multiplet;
Hz, Hertz. FAB MS was recorded on Jeol (Japan)/SX-102. Infrared
spectrum was taken with KBr on Perkin-Elmer RX-1. Melting
points were determined on a Buchi 535 digital melting point appa-
ratus and were uncorrected. Elemental analysis was performed on
a Perkin-Elmer 2400 C, H, N analyzer and values were within 0.4%
of the calculated values.
FAS-II inhibitory activitiy of selected compounds at two different concentrations
S. No.
Compound No.
% Inhibition (25
l
M)
% Inhibition (50 lM)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
4a
4b
4d
6a
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
83.40 3.0
13.61 0.64
47.14 3.19
0
88.1 0.077
14.65 0.86
42.85 7.99
0.17 0.24
0
0
0
0
39.49 2.37
15.66 8.52
41.52 5.16
46.32 7.75
16.48 9.34
17.5 24.74
29.82 4.2
26.99 6.7
36.0 10.17
30.25 7.9
48.63 2.94
17.87 2.02
1.03 1.46
60.86 1.15
31.82 2.12
37.55 10.79
57.37 3.48
27.45 6.04
27.45 6.04
3.81 4.2
32.71 6.9
36.72 3.2
23.9 4.6
4.1.1. General procedure for the synthesis of substituted (E)-1-
phenyl-3-phenylprop-2-en-1-one (3a, 3b)
6m
8a
8b
8c
50.22 2.31
30.1 2.57
16.93 0.54
To
a stirring mixture of 4-fluoroacetophenone (8.6 ml,
54.5 mmol) and 4-chlorobenzaldehyde (10 g, 71.13 mmol) or 3,4-
dimethoxy benzaldehyde (11.7 g, 70.40 mmol) in ethanol was
added KOH (1.98 g, 3.5 mmol) and stirred continued for 10–
30 min at room temperature till the disappearance of starting
materials. The light green/yellow solid was precipitated, filtered
washed with water and dried to get the substituted chalcones
(3a and 3b) in good yields.
chalcone 4a. The assessment of the FAS-II activity of the cyclopro-
pylated analogue of 4a, that is, 6g was determined by the same
protocol and compound shows 57.37% inhibition against M. aurum
at 50 lM concentration of the compound. Moreover b-gal enzyme
assay shows induced level of b-gal enzyme activity after treatment
with compound 6g with respect to untreated control (Fig. 5C). To
determine the ligand efficacy against FAS-II pathway some active
chalcones and cyclopropyl methanones were screened against
the culture of M. aurum. Among the screened molecules some com-
pound (4d and 6f) shows moderate inhibition (41–47%) of FAS-II
pathway and few of the compounds (4a, 6d, 6g and 8a) shows good
inhibition in the range of 50–88%. While profiling the activity of
synthesized compounds several ligands were identified having
good affinity towards the enzymes of FAS-II pathway. In general,
the aminoalkylamine chain has prominent role in FAS-II inhibitory
activity as evidenced by comparing the effect of compounds 4a, 4b
and 6g (Fig. 5C) wherein 4a and 6g having N,N-dimethylaminopro-
pyl chain have prominent inhibitory effect while 4b has no signif-
icant inhibition. Thus, the double bond or the cyclopropyl moiety
has no significant role in eliciting the FAS-inhibitory response.
Some of the active compounds which did not respond towards
the FAS-II pathway might have some other target and needs fur-
ther investigation. Moreover, the FAS-II pathway being absent in
human may be targeted with such small synthetic molecules to
get new chemotherapeutics for tuberculosis and malaria.
4.1.2. General procedure for the synthesis of substituted (E)-3-
(4-chlorophenyl)-1-(4-alkylamino)prop-2-en-1-one (4a–4e)
A mixture of (E)-3-(4-chlorophenyl)-1-(4-fluorophenyl)prop-
2-en-1-one (3a) (1 g, 3.84 mmol), K2CO3 (0.637 g, 4.60 mmol) and
desired amine (1.2 equiv) in DMF (5 ml) was stirred magnetically
at 100–120 °C for 16–18 h under inert atmosphere of nitrogen.
After the completion of the reaction (TLC), the reaction mixture
was cooled to room temperature and poured in water and ex-
tracted with ethyl acetate. The organic layer was dried over Na2SO4
and concentrated under reduced pressure to get the crude product.
The latter was purified by column chromatography (SiO2, 100–200
mesh) using gradient of hexane/ethyl acetate (9:1?6:4)/
0.2:9.8?0.6:9.4% methanol/chloroform to give the desired com-
pounds (4a–4e, yield 40–56%).
4.1.2.1. (E)-3-(4-Chlorophenyl)-1-(4-(3-(dimethylamino) pro-
pylamino)phenyl)prop-2-en-1-one (4a). Yellow solid, mp 60–
62 °C; yield 46%; Rf 0.43 (0.2:9.8, MeOH/CHCl3); IR (KBr) mmax
cmÀ1: 3426 (N–H), 2928 (C–H), 1605 (C@O). 1H NMR (200 MHz,
CDCl3): d 7.92 (2H, d, J = 8.84 Hz, ArH), 7.71 (1H, d, J = 15.8 Hz,
CH@CH), 7.64–7.45 (3H, m, ArH and CH@CH), 7.36 (2H, d,
J = 8.90 Hz, ArH), 6.56 (2H, d, J = 8.86 Hz, ArH), 5.53 (1H, br s,
NH), 3.27 (2H, q, J = 3.12 Hz NHCH2), 2.44 (2H, t, J = 6.24 Hz, CH2),
2.24 (6H, s, N(CH3)2), 1.81 (2H, m, CH2). 13C NMR (50 MHz, CDCl3):
d 187.5 (ArCO), 153.1, 141.3 (ArC), 136.1 (CH@CH), 134.4, 131.0,
129.7, (ArCH), 131.0, 128.9, 127.9 (ArCH), 128.7 (ArC), 127.0
(CH@CH), 111.8 (ArCH), 58.4 (NHCH2), 45.0 ((CH3)2NCH2), 42.9
(NCH2), 26.3 (CH2). ESMS (m/z): 343 (M+H)+. Anal. Calcd for
3. Conclusion
In conclusion, we have synthesized and evaluated a series of
alkylaminoaryl phenyl cyclopropyl cahlcones and methanones in
good yields. The compounds were evaluated against M. tuberculosis
and P. falciparum. Few of the compounds showed moderate to sig-
nificant antimalarial and antitubercular activities. We have also
shown one of the possible mode of actions of these compounds
may be FAS-II inhibition by in silico screen and in vitro FAS-II en-
zyme inhibitory activities. Further exploration of this study is cur-
rently underway.
C20H23ClN2O: C, 70.06; H, 6.76; N, 8.17. Found: C, 70.13; H, 6.89;
N, 8.26.
4.1.2.2. (E)-3-(4-Chlorophenyl)-1-(4-(4-methylpiperazin-1-yl)-
phenyl)prop-2-en-1-one (4b). Yellow solid, mp 84–86 °C; yield
56%; Rf 0.40 (0.1:9.9, MeOH/CHCl3); IR (KBr) mmax cmÀ1: 2938
(C–H), 1610 (C@O); 1H NMR (200 MHz, CDCl3): d 7.92 (2H, d,
J = 8.86 Hz, ArH), 7.73 (1H, d, J = 15.54 Hz, CH@CH), 7.54 (2H, d,
J = 8.76 Hz, ArH), 7.30–7.16 (3H, m, ArH and CH@CH), 6.85 (2H,
d, J = 8.84 Hz, ArH), 3.36–3.30 (4H, m, 2 Â CH2), 2.58–2.53 (4H, m,
2 Â CH2), 2.30 (3H, s, NCH3). 13C NMR (50 MHz, CDCl3): d 187.8
(ArCO), 154.6 (ArC), 142.1 (CH@CH), 136.4, 134.2 (ArC), 131.5,
4. Experimental
4.1. Chemistry
All glasswares were dried in an open flame before use in con-
nection with an inert atmosphere. Solvents were evaporated under

A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
8295
Figure 5. Reporter gene expression and viability assay after treatment with 4a. (A) Note the induced level of b-gal activity in treated sample in recombinant strain M. aurum-
kaspr:lacZ with respect to untreated control. INH as a positive control shows maximum inducibility. Recombinant M. aurum strain with hsp60 promoter, M. aurum-
hsp60pr:lacZ, shows decline in b-gal activity in all treated samples including INH with respect to untreated control. (B) Both recombinant strains exhibit similar decline in the
viability of bacterial cells; greater inhibition is observed with increasing concentration of compounds. (C) Induced level of b-gal activity in treated cyclopropylated analogue of
4a, that is, 6g in recombinant strain M. aurum-kaspr:lacZ with respect to untreated control. Recombinant M. aurum strain with hsp60 promoter, M. aurum-hsp60pr:lacZ, shows
decreased in b-gal activity.
129.8, 129.5 (ArCH), 128.8 (ArC), 122.7 (CH@CH), 113.8 (ArCH),
51.4, 48.8 (2 Â CH2), 45.1 (NCH3). ESMS (m/z): 341 (M+H)+. Anal.
Calcd for C20H21ClN2O: C, 70.48; H, 6.21; N, 8.22. Found: C,
70.38; H, 6.12; N, 8.14.
7.57–7.25 (6H, m, ArH, CH@CH and ArF-H), 7.06 (2H, d,
J = 8.86 Hz, ArH), 6.46–6.37 (2H, m, ArF-H), 5.06 (2H, s, CH2). 13C
NMR (50 MHz, CDCl3): d 188.2 (ArCO), 162.5 (ArC), 149.8 (CH@CH),
143.6, 142.8 (ArCH), 136.6, 134.0 (ArC), 131.7 (ArC), 131.1, 129.8,
129.6, 122.6, 115.0, 111.0, 110.8 (ArCH), 62.7 (CH2). ESMS (m/z):
351 (M+H)+. Anal. Calcd for C21H18ClNO2: C, 71.69; H, 5.16; N,
3.98. Found: C, 71.58; H, 5.10; N, 3.86.
4.1.2.3. (E)-3-(4-Chlorophenyl)-1-(4-(furan-2-ylmethylamino)-
phenyl)prop-2-en-1-one (4c). White solid, mp 82–84 °C; yield
40%; Rf 0.37 (0.1:9.9, MeOH/CHCl3); IR (KBr) mmax cmÀ1: 3445
(N–H), 2932 (C–H), 1600 (C@O). 1H NMR (200 MHz, CDCl3): d
8.03 (2H, d, J = 8.80 Hz, ArH), 7.76 (1H, d, J = 15.60 Hz, CH@CH),
4.1.2.4. (E)-3-(4-Chlorophenyl)-1-(4-morpholinophenyl)prop-2-
en-1-one (4d). Light green, mp 82–84 °C; yield 42%; Rf 0.45 (4:6,

8296
A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
EtOAc/Hexane); IR (KBr) mmax cmÀ1: 2932 (C–H), 1608 (C@O); 1H
NMR (200 MHz, CDCl3): d 7.99 (2H, d, J = 9.26 Hz, ArH), 7.76 (1H,
d, J = 15.62 Hz, CH@CH), 7.58 (2H, d, J = 8.72 Hz, ArH), 7.39–7.25
(3H, m, ArH and CH@CH), 6.91 (2H, d, J = 9.00 Hz), 3.88 (4H, t,
J = 4.72 Hz, 2 Â CH2), 3.35 (4H, t, J = 5.06 Hz, 2 Â CH2). 13C NMR
(50 MHz, CDCl3): d 187.8 (ArCO), 154.6 (ArC), 142.1 (CH@CH),
136.4, 134.2 (ArC), 131.5, 129.8, 129.5 (ArCH), 128.8 (ArC), 122.7
(CH@CH), 113.8 (ArCH), 66.9, 47.9 (2 Â CH2). ESMS (m/z): 328
(M+H)+. Anal. Calcd for C19H18ClNO2: C, 69.62; H, 5.53; N, 4.27.
Found: C, 69.56; H, 5.48; N, 4.14.
28.1, 27.3 (C-2 and C-3), 18.2 (C-4). ESMS (m/z): 275 (M+H)+. Anal.
Calcd for C16H12ClFO: C, 69.95; H, 4.40. Found: C, 69.97; H, 4.42.
4.1.4.2. (2-(3,4-Dimethoxyphenyl)cyclopropyl)(4-fluorophenyl)-
methanone (5b). White solid, mp 78–80 °C; yield 90%; Rf 0.55
(1.5:8.5, EtOAc/Hexanae); IR (KBr) mmax cmÀ1: 2934 (C–H), 1652
(C@O); 1H NMR (200 MHz, CDCl3): d 8.04–7.97 (2H, m, ArH),
7.15–7.07 (2H, m, ArH), 6.80 (1H, d, J = 8.8 Hz, ArH), 6.69–6.65
(2H, m, ArH), 3.86 (3H, s, OCH3), 3.85 (3H, s, OCH3), 2.76–2.70
(1H, m, H-2), 2.64–2.60 (1H, m, H-3), 1.91–1.84 (1H, m, H-4a),
1.55–1.48 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d 188.9 (ArCO),
165.5, 148.5, 146.7, 136.8, 133.5 (ArC), 131.7, 129.1, 127.2, 126.4,
113.3 (ArCH), 28.3, 27.2 (C-2 and C-3), 18.4 (C-4). ESMS (m/z):
301 (M+H)+. Anal. Calcd for C18H17FO3: C, 71.99; H, 5.71. Found:
C, 72.06; H, 5.74.
4.1.2.5. (E)-3-(4-Chlorophenyl)-1-(4-(piperidin-1-yl)phenyl)prop-
2-en-1-one (4e). White solid, mp 116–118 °C; yield 54%; Rf 0.55
(10%, EtOAc/Hexane); IR (KBr) m
max cmÀ1: 2921 (C–H), 1610 (C@O);
1H NMR (300 MHz, CDCl3): d 7.82 (2H, d, J = 8.76 Hz, ArH), 7.74
(1H, d, J = 15.34 Hz, CH@CH), 7.24 (2H, d, J = 8.72 Hz, ArH),
7.21–7.15 (3H, m, ArH and CH@CH), 6.88 (2H, d, J = 8.78 Hz), 3.67
(4H, br s, 2 Â NCH2), 1.78 (6H, br s, 3 Â CH2). 13C NMR (50 MHz,
CDCl3): d 185.5 (ArCO), 151.6 (ArC), 143.4 (CH@CH), 136.2, 133.2
(ArC), 131.8, 128.8, 128.3 (ArCH), 127.9 (ArC), 122.3 (CH@CH),
113.1 (ArCH), 54.3 (2 Â CH2), 23.7 (2 Â CH2), 22.8 (CH2). ESMS
(m/z): 326 (M+H)+. Anal. Calcd for C20H20ClNO: C, 73.72; H, 6.19;
N, 4.30. Found: C, 73.76; H, 6.09; N, 4.26.
4.1.4.3. (2-(4-Chlorophenyl)cyclopropyl)(4-(4-methylpiperazin-
1-yl)phenyl)methanone (6a). Pale white solid, mp 98–100 °C;
yield 84%; Rf 0.38 (0.1:9.9, MeOH/CHCl3); IR (KBr) mmax cmÀ1
:
2936 (C–H), 1647 (C@O); 1H NMR (300 MHz, CDCl3): d 7.90 (2H,
d, J = 8.67 Hz, ArH), 7.27 (2H, d, J = 8.28 Hz, ArH), 7.11 (2H, d,
J = 8.19 Hz, ArH), 6.86 (2H, d, J = 8.73 Hz, ArH), 3.38–3.35 (4H, m,
2 Â CH2), 2.78–2.72 (1H, m, H-2), 2.62–2.53 (5H, m, 2 Â CH2 and
H-3), 2.34 (3H, s, NCH3), 1.89–1.83 (1H, m, H-4a), 1.44–1.38 (1H,
m, H-4b). 13C NMR (50 MHz, CDCl3): d 195.7 (ArCO), 152.5, 139.8,
131.9 (ArC), 130.7, 128.4, 127.2 (ArCH), 126.5 (ArC), 111.3 (ArCH),
51.1, 48.6 (2 Â CH2), 45.3 (NCH3), 28.3, 27.5 (C-2 and C-3), 18.6
(C-4). ESMS (m/z): 355 (M+H)+. Anal. Calcd for C21H23ClN2O: C,
71.07; H, 6.53; N, 7.89. Found: C, 71.05; H, 6.58; N, 7.83.
4.1.3. General procedure for the synthesis of substituted aryl
phenyl cyclopropyl methanone (5a, 5b) and alkylamino
substituted aryl phenyl cyclopropyl methanone (6a, 6g, 6h, 6j,
6n) from 4a–4e
A mixture of compound 3a or 3b (1 equiv), TMSOI (trimethyl
sulphoxonium iodide, 2 equiv) and TBAB (tetra butylammonium
bromide, 20 mol%) in CH2Cl2 (10 ml) was stirred magnetically at
ambient temperature for 15 min. 50% aq NaOH (10 ml) solution
was subsequently added dropwise and the reaction mixture was
refluxed at 80 °C, till the disappearance of chalcone (3a or 3b).
After the completion of the reaction (TLC), the reaction mixture
was cooled to room temperature, excess of dichloromethane was
added and the organic layer was extracted. The organic layer was
dried (anhyd Na2SO4) and concentrated under reduced pressure
to get a crude product. The latter was purified by column chroma-
tography (SiO2, 100–200 mesh) using gradient of hexane/ethylace-
tate/chloroform/methanol to give the desired compounds (5a, 5b
and 6a, 6g, 6h, 6j and 6n) in good yields.
4.1.4.4. (2-(4-Chlorophenyl)cyclopropyl)(4-(cyclohexylamino)-
phenyl)methanone (6b). White solid, mp 128–130 °C; yield
81%; Rf 0.50 (15%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3348
(N–H), 2927 (C–H), 1651 (C@O); 1H NMR (300 MHz, CDCl3): d
7.83 (2H, d, J = 8.70 Hz, ArH), 7.24 (2H, d, J = 8.31 Hz, ArH), 7.08
(2H, d, J = 8.13 Hz, ArH), 6.52 (2H, d, J = 8.52 Hz, ArH), 4.11 (1H,
br s, NH), 3.33 (1H, br s, NHCH), 2.74–2.70 (1H, m, H-2),
2.59–2.56 (1H, m, H-3), 2.06–2.02 (2H, m, CH2), 1.84–1.64 (4H,
m, CH2, H-4a and H-4b), 1.40–1.16 (6H, m, CH2). 13C NMR
(50 MHz, CDCl3): d 195.6 (ArCO), 151.6, 140.1, 132.4 (ArC), 131.0,
128.9, 127.9 (ArCH), 126.8 (ArC), 112.0 (ArCH), 51.6 (NHCH), 33.5
(CH2), 28.8, 28.3 (C-2 and C-3), 26.1 (CH2), 25.2 (CH2), 18.2 (C-4).
ESMS (m/z): 354 (M+H)+. Anal. Calcd for C22H24ClNO: C, 74.67; H,
6.84; N, 3.96. Found: C, 74.63; H, 6.81 N, 3.94.
4.1.4. General procedure for the synthesis of alkylamino substi-
tuted aryl phenyl cyclopropyl methanone (6b–6f, 6i, 6k–6l and
6o–6u) from 5a and 5b
4.1.4.5. (2-(4-Chlorophenyl)cyclopropyl)(4-(heptylamino)phe-
nyl)methanone (6c). White solid, mp 134–136 °C; yield 77%; Rf
0.50 (1:4, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3353 (N–H), 2924
(C–H), 1569 (C@O); 1H NMR (300 MHz, CDCl3): d 7.88 (2H, d,
J = 8.61 Hz, ArH), 7.27 (2H, d, J = 8.40 Hz, ArH), 7.11 (2H, d,
J = 8.28 Hz, ArH), 6.56 (2H, d, J = 8.61 Hz, ArH), 4.20 (1H, br s,
NH), 3.18 (2H, t, J = 6.96 Hz, NHCH2), 2.79–2.74 (1H, m, H-2),
2.63–2.56 (1H, m, H-3), 1.89–1.83 (1H, m, H-4a), 1.67–1.63 (2H,
m, CH2), 1.44–1.32 (9H, m, 4 Â CH2 and H-4b), 0.91 (3H, t, CH3).
13C NMR (50 MHz, CDCl3): d 195.7 (ArCO), 152.6, 140.1, 132.4
(ArC), 131.0, 128.9, 127.9 (ArCH), 127.2 (ArC), 111.7 (ArCH), 43.7,
32.2, 29.7, 29.5 (NHCH2 and 3 Â CH2), 28.8, 28.4 (C-2 and C-3),
27.4, 23.0 (2 Â CH2), 18.7 (C-4), 14.5 (CH3). ESMS (m/z): 370
(M+H)+. Anal. Calcd for C23H28ClNO: C, 74.68; H, 7.63; N, 3.79.
Found: C, 74.71; H, 7.65; N, 3.76.
A mixture of 5a or 5b (1 equiv), K2CO3 (1.2 equiv) and desired
amine (1.2 equiv) in DMF (5 ml) was stirred magnetically at 100–
120 °C for 16–18 h under inert atmosphere of nitrogen. After the
completion of the reaction (TLC), the reaction mixture was cooled
to room temperature and poured in water and extracted with ethyl
acetate. The organic layer was dried (anhyd Na2SO4) and concen-
trated under reduced pressure to give a crude product. The latter
was purified by column chromatography (SiO2, 100–200 mesh)
using gradient of hexane/ethyl acetate (9:1?6:4)/0.2:9.8?0.6:
9.4% methanol/chloroform to give the desired compounds (6b–6f,
6i, 6k–6l and 6o–6u) yield 78–86%.
4.1.4.1. (2-(4-Chlorophenyl)cyclopropyl)(4-fluorophenyl)meth-
anone (5a). White solid, mp 118–120 °C; yield 84%; Rf 0.40
(1.0:9.0, EtOAc/Hexanae); IR (KBr) mmax cmÀ1: 2938 (C–H), 1657
(C@O); 1H NMR (200 MHz, CDCl3): d 8.03–7.95 (2H, m, ArH),
7.28–7.23 (2H, m, ArH), 7.18–7.06 (4H, m, ArH), 2.82–2.73 (1H,
m, H-2), 2.69–2.60 (1H, m, H-3), 1.95–1.86 (1H, m, H-4a),
1.55–1.45 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d 188.7 (ArCO),
165.8, 136.8, 134.5 133.3, (ArC), 131.1, 129.2, 128.4, 115.3 (ArCH),
4.1.4.6. (4-(Butylamino)phenyl)(2-(4-chlorophenyl)cyclopropyl)-
methanone (6d). White solid, mp 97–99 °C; yield 82%; Rf 0.48
(15%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3436 (N–H), 3020
(C–H), 1597 (C@O); 1H NMR (300 MHz, CDCl3): d 7.85 (2H, d,
J = 8.22 Hz, ArH), 7.26 (2H, d, J = 7.74 Hz, ArH), 7.11 (2H, d,

A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
8297
J = 7.83 Hz, ArH), 6.54 (2H, d, J = 8.04 Hz, ArH), 4.11 (1H, br s, NH),
3.21 (2H, d, J = 8.04 Hz, NHCH2), 2.73 (1H, m, H-2), 2.58 (1H, m, H-
3), 1.86–1.84 (1H, m, H-4a), 1.68–1.63 (2H, m, CH2), 1.51–1.39 (3H,
m, CH2 and H-4b), 1.00 (3H, t, J = 7.11 Hz, CH3). 13C NMR (50 MHz,
CDCl3): d 194.7 (ArCO), 152.1, 139.6, 132.0 (ArC), 130.5, 128.5,
127.5 (ArCH), 126.8 (ArC), 111.2 (ArCH), 42.9 (NHCH2), 31.4
(CH2) 28.2, 27.7 (C-2 and C-3), 20.2 (CH2), 18.2 (C-4), 13.9 (CH3).
ESMS (m/z): 328 (M+H)+. Anal. Calcd for C20H22ClNO: C, 73.27; H,
6.76; N, 4.27. Found: C, 73.21; H, 6.73; N, 4.24.
128.9, 128.0, 127.9, 112.2, 110.8, 107.8 (ArCH), 41.1 (CH2), 28.8,
28.4 (C-2 and C-3), 18.7 (C-4). ESMS (m/z): 352 (M+H)+. Anal. Calcd
for C21H18ClNO2: C, 71.69; H, 5.16; N, 3.98. Found: C, 71.63; H,
5.13; N, 3.94.
4.1.4.11. (2-(4-Chlorophenyl)cyclopropyl)(4-(6-methylheptan-
2-ylamino)phenyl)methano-ne (6i). White solid, mp 128–
130 °C; yield 80%; Rf 0.30 (5%, EtOAc/Hexane); IR (KBr) mmax
cmÀ1: 3422 (N–H), 2926 (C–H), 1596 (C@O); 1H NMR (300 MHz,
CDCl3): d 7.82 (2H, d, J = 8.70 Hz, ArH), 7.24 (2H, d, J = 8.37 Hz,
ArH), 7.08 (2H, d, J = 8.37 Hz, ArH), 6.49 (2H, d, J = 8.67 Hz, ArH),
3.95 (1H, br s, NH), 3.53–3.51 (1H, m, NHCH), 2.73–2.67 (1H, m,
H-2), 2.57–2.51 (1H, m, H-3), 1.85–1.79 (1H, m, H-4a), 1.56–1.21
(12H, m, 3 Â CH2, CH3, 2 Â CH and and H-4b), 0.87 (6H, d,
J = 6.6 Hz (CH3)2). 13C NMR (50 MHz, CDCl3): d 195.0 (ArCO),
151.6, 140.1, 132.4 (ArC), 131.0, 128.9, 127.9 (ArCH), 127.1 (ArC),
112.1 (ArCH), 48.7 (CH), 39.3, 37.7 (2 Â CH2), 28.7 (CH), 28.3,
28.2 (C-2 and C-3), 24.3 (CH2), 23.1, 21.1 (CH3), 18.5 (CH2). ESMS
(m/z): 384 (M+H)+. Anal. Calcd for C24H30ClNO: C, 75.08; H, 7.88;
N, 3.65. Found: C, 74.98; H, 7.84; N, 3.61.
4.1.4.7. (2-(4-Chlorophenyl)cyclopropyl)(4-(4-(2,3-dichlorophe-
nyl)piperazin-1-yl)phenyl) methanone (6e). Light green solid,
mp 152–154 °C; yield 78%; Rf 0.50 (2%, MeOH/CHCl3); IR (KBr) mmax
cmÀ1: 2936 (C–H), 1593 (C@O); 1H NMR (300 MHz, CDCl3): d 7.93
(2H, d, J = 8.70 Hz, ArH), 7.25 (2H, d, J = 8.34 Hz, ArH), 7.17 (2H, d,
J = 7.44, ArH), 7.09 (2H, d, J = 8.37 Hz, ArH), 6.96–6.89 (3H, m, ArH),
3.52–3.49 (4H, m, 2 Â CH2), 3.19–3.16 (4H, m, 2 Â CH2), 2.80–2.74
(1H, m, H-2), 2.63–2.57 (1H, m, H-3), 1.88–1.82 (1H, m, H-4a),
1.45–1.25 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d 196.0 (ArCO),
154.5, 151.1, 139.9, 134.7, 132.5 (ArC), 130.5, 129.0 (ArCH), 128.7,
128.2 (ArC), 128.0, 127.9, 125.5, 118.9, 114.2 (ArCH), 51.4, 48.2
(2 Â CH2), 28.9, 28.6 (C-2 and C-3), 19.0 (C-4). ESMS (m/z): 485
(M+H)+. Anal. Calcd for C26H23Cl3N2O: C, 64.28; H, 4.77; N, 5.77.
Found: C, 64.25; H, 4.79; N,5.73.
4.1.4.12. (2-(4-Chlorophenyl)cyclopropyl)(4-morpholinophenyl)-
methanone (6j). Light green solid, mp 115–117 °C; yield 83%; Rf
0.50 (2%, MeOH/CHCl3); IR (KBr) mmax cmÀ1: 3019 (C–H), 1649
(C@OC@O). 1H NMR (300 MHz, CDCl3): d 7.89 (2H, d, J = 9.09 Hz,
ArH), 7.24 (2H, d, J = 8.46 Hz, ArH), 7.08 (2H, d, J = 8.40 Hz, ArH),
6.84 (2H, d, J = 8.91 Hz, ArH), 3.83–3.80 (4H, m, 2 Â CH2), 3.28–
3.25 (m, 4H, 2 Â CH2), 2.76–2.70 (1H, m, H-2), 2.60–2.54 (1H, m,
H-3), 1.88–1.73 (1H, m, H-4a), 1.42–1.36 (1H, m, H-4b). 13C NMR
(50 MHz, CDCl3): d 195.7 (ArCO), 153.3, 140.1, 132.4 (ArC), 130.4,
129.0, 127.9 (ArCH), 126.9 (ArC), 113.8 (ArCH), 66.7 (2 Â CH2),
48.1 (2 Â CH2), 28.8, 28.5 (C-2 and C-3), 18.8 (C-4). ESMS (m/z):
342 (M+H)+. Anal. Calcd for C20H20ClNO2: C, 70.27; H, 5.90; N,
4.10. Found: C, 70.25; H, 5.89; N, 4.07.
4.1.4.8. (2-(4-Chlorophenyl)cyclopropyl)(4-(dibutylamino)phe-
nyl)methanone (6f). Light green solid, mp 94–96 °C; yield 77%;
Rf 0.55 (10%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 2959 (C–H),
1594 (C@O); 1H NMR (300 MHz, CDCl3): d 7.87 (2H, d, J = 9.00 Hz,
ArH), 7.24 (2H, d, J = 8.46 Hz, ArH), 7.08 (2H, d, J = 8.43 Hz, ArH),
6.59 (2H, d, J = 9.03 Hz, ArH), 3.31 (4H, m, N(CH2)2), 2.77–2.71
(1H, m, H-2), 2.58–2.51 (1H, m, H-3), 1.87–1.81 (1H, m, H-4a),
1.63–1.53 (4H, m, 2 Â CH2), 1.41–1.29 (5H, m, 2 Â CH2 and H-4b),
0.93 (6H, t, 2 Â CH3).13C NMR (50 MHz, CDCl3): d 195.3 (ArCO),
151.6, 140.2, 132.3 (ArC), 130.9, 128.9, 127.9 (ArCH), 125.6 (ArC),
111.0 (ArCH), 51.3 (N(CH2)2), 30.0 (2 Â CH2), 28.7, 28.2 (C-2 and
C-3), 20.7 (CH2), 18.4 (C-4), 14.3 (CH3). ESMS (m/z): 384 (M+H)+.
Anal. Calcd for C24H30ClNO: C, 75.08; H, 7.88; N, 3.65. Found: C,
75.01; H, 7.90; N, 3.62.
4.1.4.13. (2-(4-Chlorophenyl)cyclopropyl)(4-(dodecylamino)phe-
nyl)methanone (6k). White solid, mp 122–124 °C; yield 85%; Rf
0.50 (3%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3355 (N–H), 2921
(C–H), 1639 (C@O); 1H NMR (300 MHz, CDCl3): d 7.82 (2H, d,
J = 8.61 Hz, ArH), 7.23 (2H, d, J = 8.34 Hz, ArH), 7.07 (2H, d,
J = 8.31 Hz, ArH), 6.50 (2H, d, J = 8.61 Hz, ArH), 4.13 (1H, br s,
NH), 3.15 (2H, t, J = 6.87 Hz, NHCH2), 2.77–2.68 (1H, m, H-2),
2.61–2.52 (1H, m, H-3), 1.84–1.78 (1H, m, H-4a), 1.67–1.57 (2H,
m, CH2), 1.38–1.25 (19H, m, 9 Â CH2 and H-4b), 0.88 (3H, t,
J = 6.85 Hz, CH3). 13C NMR (50 MHz, CDCl3): d 195.2 (ArCO),
152.4, 140.1, 132.4 (ArC), 130.9, 128.9, 127.9 (ArCH), 127.3 (ArC),
111.8 (ArCH), 43.8, 32.3, 32.2, 30.0, 30.0, 29.8, 29.7 (CH2), 28.7,
28.2 (C-2 and C-3), 27.5, 27.4, 23.1, 18.6 (CH2), 18.5 (C-4), 14.6
(CH3). ESMS (m/z): 440 (M+H)+. Anal. Calcd for C28H38ClNO: C,
76.42; H, 8.70; N, 3.18. Found: C, 76.38; H, 8.68; N, 3.14.
4.1.4.9. (2-(4-Chlorophenyl)cyclopropyl)(4-(3-(dimethylamino)-
propylamino)phenyl)met-hanone (6g). White solid, mp 96–
98 °C; yield 83%; Rf 0.33 (0.1:9.9, MeOH/CHCl3); IR (KBr) mmax
cmÀ1: 3432 (N–H), 2928 (C–H), 1690 (C@O). 1H NMR (300 MHz,
CDCl3): d 7.83 (2H, d, J = 8.58 Hz, ArH), 7.23 (2H, d, J = 8.34 Hz,
ArH), 7.07 (2H, d, J = 8.28 Hz, ArH), 6.51 (2H, d, J = 8.58 Hz, ArH),
5.39 (1H, s, NH), 3.25 (2H, br s, NHCH2), 2.76–2.71 (1H, m, H-2),
2.60–2.53 (1H, m, H-3), 2.42 (2H, t, J = 6.24 Hz, NCH2), 2.25 (6H,
s, N(CH3)2), 1.85–1.76 (3H, m, CH2 and H-4a), 1.40–1.34 (1H, m,
H-4b). 13C NMR (50 MHz, CDCl3): d 195.6 (ArCO), 153.0, 140.1,
132.3 (ArC), 131.0, 128.9, 127.9 (ArCH), 126.8 (ArC), 111.7 (ArCH),
58.8 (NHCH2), 45.7 ((CH3)2NCH2), 43.0 (NCH2), 28.7, 28.2 (C-2 and
C-3), 26.4 (CH2), 18.7 (C-4). ESMS (m/z): 357 (M+H)+. Anal. Calcd
for C21H25ClN2O: C, 70.67; H, 7.06; N, 7.85. Found: C, 70.59; H,
7.02; N, 7.81.
4.1.4.14. (2-(4-Chlorophenyl)cyclopropyl)(4-(hexadecylamino)-
phenyl)methanone (6l). White solid, mp 154–156 °C; yield
87%; Rf 0.55 (2%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3459 (N–
H), 2890 (C–H), 1693 (C@O); 1H NMR (300 MHz, CDCl3): d 7.82
(2H, d, J = 8.64 Hz, ArH), 7.23 (2H, d, J = 8.34 Hz, ArH), 7.08 (2H,
d, J = 8.37 Hz, ArH), 6.50 (2H, d, J = 8.67 Hz, ArH), 4.13 (1H, br s,
NH), 3.15 (2H, t, J = 6.96 Hz, NHCH2), 2.71–2.7 (1H, m, H-2),
2.56–2.53 (1H, m, H-3), 1.83–1.80 (1H, m, H-4a), 1.64–1.60 (2H,
m, CH2), 1.38–1.25 (27H, m, 13 Â CH2 and H-4b), 0.90–0.86 (3H,
m, CH3). 13C NMR (50 MHz, CDCl3): d 195.0 (ArCO), 152.3, 140.1,
132.4 (ArC), 130.9, 128.9, 127.9 (ArCH), 127.4 (ArC), 111.7 (ArCH),
43.8, 32.3, 30.0, 30.0, 29.7 (CH2), 28.6, 28.1 (C-2 and C-3), 27.5,
23.1, (CH2), 18.5 (C-4), 14.5 (CH3). ESMS (m/z): 497 (M+H)+. Anal.
Calcd for C32H46ClNO: C, 77.46; H, 9.34; N, 2.82. Found: C, 77.40;
H, 9.29; N, 2.79.
4.1.4.10. (2-(4-Chlorophenyl)cyclopropyl)(4-(furan-2-ylmethyl-
amino)phenyl)methanone (6h). White solid, mp 96–98 °C; yield
85%; Rf 0.55 (20%, EtOAc/Hexane); IR (KBr) m
max cmÀ1: 3445 (N–H),
2928 (C–H), 1600 (C@O); 1H NMR (300 MHz, CDCl3): d 7.96 (2H, d,
J = 8.88 Hz, ArH), 7.43 (1H, m, Furyl-H), 7.25 (2H, d, J = 8.46 Hz,
ArH), 7.07 (2H, d, J = 8.46 Hz, ArH), 6.99 (2H, d, J = 8.91 Hz, ArH),
6.42 (1H, m, Furyl-H), 6.36 (1H, m, Furyl-H), 5.04 (2H, s, NHCH2),
2.80–2.74 (1H, m, H-2), 2.64–2.57 (1H, m, H-3), 1.89–1.83 (1H,
m, H-4a), 1.47–1.41 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d
195.7 (ArCO), 151.8, 151.7, 142.5, 140.0, 132.4 (ArC), 130.9,

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A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
4.1.4.15. (2-(4-Chlorophenyl)cyclopropyl)(4-(octylamino)phe-
nyl)methanone (6m). White solid, mp 134–136 °C; yield 83%;
Rf 0.50 (5%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3354 (N–H),
2922 (C–H),1588 (C@O); 1H NMR (300 MHz, CDCl3): d 7.82 (2H,
d, J = 8.67 Hz, ArH), 7.24 (2H, d, J = 8.40 Hz, ArH), 7.07 (2H, d,
J = 8.40 Hz, ArH), 6.50 (2H, d, J = 8.67 Hz, ArH), 4.16 (1H, br s,
NH), 3.15 (2H, t, J = 7.02 Hz, NHCH2), 2.73–2.67 (1H, m, H-2),
2.57–2.51 (1H, m, H-3), 1.84–1.78 (1H, m, H-4a), 1.64–1.57 (2H,
m, CH2), 1.38–1.28 (11H, m, 5 Â CH2 and H-4b), 0.95–0.86 (3H, t,
CH3). 13C NMR (50 MHz, CDCl3): d 195.0 (ArCO), 152.4, 140.1,
132.4 (ArC), 130.9, 128.9, 127.9 (ArCH), 127.3 (ArC), 111.7 (ArCH),
43.7, 32.2, 29.8, 29.6 (NHCH2 and 3 Â CH2), 28.6, 28.1 (C-2 and
C-3), 27.5, 23.0 (2 Â CH2), 18.5 (C-4), 14.5 (CH3). ESMS (m/z): 384
(M+H)+. Anal. Calcd for C24H30ClNO: C, 75.08; H, 7.88; N, 3.65.
Found: C, 75.06; H, 7.86; N, 3.67.
CH2), 1.47–1.21 (24H, m, 12 Â CH2), 0.92 (3H, t, J = 6.64 Hz, CH3).
13C NMR (50 MHz, CDCl3): d 195.6 (ArCO), 152.6, 140.1, 132.4
(ArC), 131.0 (ArCH), 130.4, 130.0 (CH@CH) 128.9, 127.9 (ArCH),
127.1 (ArC), 111.7 (ArCH), 43.7, 33.04, 32.3, 30.1, 30.0, 29.7, 29.6
(NHCH2 and 6 Â CH2), 28.7, 28.3 (C-2 and C-3), 27.6, 27.6, 27.5,
23.1 (4 Â CH2), 18.7 (C-4), 14.5 (CH3). ESMS (m/z): 522 (M+H)+.
Anal. Calcd for C34H48ClNO: C, 78.20; H, 9.26; N, 2.68. Found: C,
78.11; H, 9.21; N, 2.65.
4.1.4.20.
(2-(4-Chlorophenyl)cyclopropyl)(4-(3,4-dimethoxy-
benzylamino)phenyl)methan-one (6r). White solid, mp 135–
137 °C; yield 73%; Rf 0.50 (20%, EtOAc/Hexane); IR (KBr) mmax
cmÀ1: 3350 (N–H), 2938 (C–H), 1587 (C@O); 1H NMR (200 MHz,
CDCl3): d 7.74 (2H, d, J = 8.74 Hz, ArH), 7.26–7.19 (3H, m, ArH),
7.10 (2H, d, J = 8.48 Hz, ArH), 6.89–6.79 (2H, m, ArH), 6.61 (2H, d,
J = 8.72 Hz, ArH), 4.46 (1H, br s, NH), 4.32 (2H, s, NHCH2), 3.87
(3H, s, OCH3), 3.86 (3H, s, OCH3), 2.76–2.70 (1H, m, H-2), 2.63–
2.55 (1H, m, H-3), 1.89–1.80 (1H, m, H-4a), 1.42–1.34 (1H, m,
H-4b). 13C NMR (50 MHz, CDCl3): d 195.7 (ArCO), 152.6, 148.6,
148.1, 140.1, 133.1, 132.4 (ArC), 130.9, 128.9, 127.9 (ArCH), 127.2
(ArC), 120.0, 112.1, 111.7, 111.0 (ArCH), 56.2 (2 Â OCH3), 48.0
(NHCH2), 28.8, 28.4 (C-2 and C-3), 18.7 (C-4). ESMS (m/z): 422
(M+H)+. Anal. Calcd for C25H24ClNO3: C, 71.17; H, 5.73; N, 3.32.
Found: C, 71.11; H, 5.70; N, 3.28.
4.1.4.16. (2-(4-Chlorophenyl)cyclopropyl)(4-(piperidin-1-yl)phe-
nyl)methanone (6n). Pale white solid, mp 134–136 °C; yield
85%; Rf 0.50 (5%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 2921 (C–H),
1596 (C@O); 1H NMR (300 MHz, CDCl3): d 7.78 (2H, d, J = 8.76 Hz,
ArH), 716 (2H, d, J = 8.28 Hz, ArH), 7.01 (2H, d, J = 8.28 Hz, ArH),
6.74 (2H, d, J = 8.79 Hz, ArH), 3.26 (4H, br s, 2 Â NCH2), 2.68–2.62
(1H, m, H-2), 2.51–2.45 (1H, m, H-3), 1.78–1.72 (1H, m, H-4a),
1.58 (6H, br s, 3 Â CH2), 1.32–1.26 (1H, m, H-4b). 13C NMR
(50 MHz, CDCl3): d 195.5 (ArCO), 152.4, 140.0, 132.1 (ArC), 130.6,
128.8, 127.3 (ArCH), 127.1 (ArC), 111.5 (ArCH), 51.2 (2 Â NCH2),
28.6, 28.4 (C-2 and C-3), 23.8 (2 Â CH2), 23.3 (CH2), 18.7 (C-4).
ESMS (m/z): 341 (M+H)+. Anal. Calcd for C21H22ClNO: C, 74.21; H,
6.52; N, 4.12. Found: C, 74.17; H, 6.49; N, 4.14.
4.1.4.21.
(2-(4-Chlorophenyl)cyclopropyl)(4-(2-hydroxyethyl-
amino)phenyl)methanone (6s). White solid, mp 144–146 °C;
yield 75%; Rf 0.50 (5%, MeOH/CHCl3); IR (KBr) mmax cmÀ1: 3419
(O–H), 3330 (N–H), 2930 (C–H), 1588 (C@O); 1H NMR (300 MHz,
CDCl3): d 7.83 (2H, d, J = 8.58 Hz, ArH), 7.25 (2H, d, J = 8.46 Hz,
ArH), 7.09 (2H, d, J = 8.28 Hz, ArH), 6.59 (2H, d, J = 8.43 Hz, ArH),
3.87–3.71 (2H, m, CH2), 3.46 (1H, br s, NH), 3.33–3.32 (2H, m,
CH2), 2.79–2.74 (1H, m, H-2), 2.63–2.56 (1H, m, H-3), 1.89–1.83
(1H, m, H-4a), 1.46–1.36 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3):
d 195.9 (ArCO), 152.6, 140.1, 132.4 (ArC), 131.0, 128.9, 127.9
(ArCH), 127.2 (ArC), 111.7 (ArCH), 58.7 (CH2OH), 43.3 (NHCH2),
28.8, 28.4 (C-2 and C-3), 18.7 (C-4). ESMS (m/z): 316 (M+H)+. Anal.
Calcd for C18H18ClNO2: C, 68.46; H, 5.75; N, 4.44. Found: C, 68.39;
H, 5.73; N, 4.41.
4.1.4.17. (2-(4-Chlorophenyl)cyclopropyl)(4-(dimethylamino)-
phenyl)methanone (6o). White solid, mp 92–94 °C; yield 80%;
Rf 0.45 (15%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3020 (C–H),
1596 (C@O). 1H NMR (300 MHz, CDCl3): d 7.87 (2H, d, J = 8.97 Hz,
ArH), 7.24 (2H, d, J = 8.43 Hz, ArH), 7.08 (2H, d, J = 8.37 Hz, ArH),
6.63 (2H, d, J = 8.97 Hz, ArH), 3.05 (6H, s, NHCH2), 2.76–2.70 (1H,
m, H-2), 2.58–2.52 (1H, m, H-3), 1.85–1.79 (1H, m, H-4a), 1.39–
1.32 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d 195.6 (ArCO),
150.8, 139.5, 132.6 (ArC), 130.6 (ArCH), 129.8 (ArC), 129.0, 127.8,
115.5 (ArCH), 43.0 (N(CH3)2), 29.5, 28.9 (C-2 and C-3), 19.5 (C-4).
ESMS (m/z): 300 (M+H)+. Anal. Calcd for C18H18ClNO: C, 72.11; H,
6.05; N, 4.67. Found: C, 72.08; H, 6.01; N, 4.65.
4.1.4.22. (2-(3,4-Dimethoxyphenyl)cyclopropyl)(4-(4-methylpi-
perazin-1-yl)phenyl)meth-anone (6t). White solid, mp 112–
114 °C; yield 81%; Rf 0.30 (0.1:9.9, MeOH/CHCl3); IR (KBr) mmax
cmÀ1: 2939 (C–H), 1651 (C@O); 1H NMR (200 MHz, CDCl3): d
7.89 (2H, d, J = 8.67, 8.70 Hz, ArH), 6.87–6.69 (5H, m, ArH), 3.83
(6H, s, DiOMe), 3.36 (4H, br s, 2 Â CH2), 2.57–2.32 (9H, m, NCH3,
2 Â CH2 and H-2, H-3), 1.80–1.76 (1H, m, H-4a), 1.46–1.40 (1H,
m, H-4b). 13C NMR (50 MHz, CDCl3): d 195.5 (ArCO), 152.3, 144.7,
139.9, 131.7 (ArC), 128.1, 127.5 (ArCH), 126.7 (ArC), 111.8 (ArCH),
56.3, 56.9 (2 Â OMe), 50.5, 48.1 (2 Â CH2), 45.7 (NCH3), 28.3, 27.6
(C-2 and C-3), 18.3 (C-4). ESMS (m/z): 381 (M+H)+. Anal. Calcd
for C23H28N2O3: C, 72.60; H, 7.42; N, 7.36. Found: C, 72.57; H,
7.39; N, 7.33.
4.1.4.18. (2-(4-Chlorophenyl)cyclopropyl)(4-(pyrrolidin-1-yl)phe-
nyl)methanone (6p). White solid, mp 134–136 °C; yield 81%; Rf
0.40 (5%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 2960 (C–H), 1607
(C@O); 1H NMR (200 MHz, CDCl3): d 7.91 (2H, d, J = 8.90 Hz,
ArH), 7.27 (2H, d, J = 8.48 Hz, ArH), 7.11 (2H, d, J = 8.48 Hz, ArH),
6.52 (2H, d, J = 8.94 Hz, ArH), 3.39–3.33 (4H, m, 2 Â NCH2), 2.79–
2.73 (1H, m, H-2), 2.63–2.50 (1H, m, H-3), 2.10–1.98 (4H, m,
2 Â CH2), 1.89–1.80 (1H, m, H-4a), 1.43–1.39 (1H, m, H-4b). 13C
NMR (50 MHz, CDCl3): d 195.7 (ArCO), 151.1, 140.2, 132.3 (ArC),
130.8, 128.9, 127.9 (ArCH), 125.9 (ArC), 111.5 (ArCH), 48.2
(2 Â NCH2), 28.8, 28.4 (C-2 and C-3), 25.8 (2 Â CH2), 18.7 (C-4).
ESMS (m/z): 326 (M+H)+. Anal. Calcd for C20H20ClNO: C, 73.72; H,
6.19; N, 4.30. Found: C, 73.68; H, 6.17; N, 4.32.
4.1.4.23. (2-(3,4-Dimethoxyphenyl)cyclopropyl)(4-(heptylami-
no)phenyl)methanone (6u). White solid, mp 144–146 °C; yield
79%; Rf 0.40 (1:4, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3357 (N–
H), 2921 (C–H), 1556 (C@O); 1H NMR (300 MHz, CDCl3): d 7.88
(2H, d, J = 8.70 Hz, ArH), 6.79–6.66 (3H, m, ArH), 6.54 (2H, d,
J = 8.73 Hz, ArH), 4.27 (1H, br s, NH), 3.86 (6H, s, DiOMe), 3.17
(2H, m, NHCH2), 2.74–2.71 (1H, m, H-2), 2.57–2.52 (1H, m, H-3),
1.82–1.79 (1H, m, H-4a), 1.65–1.60 (2H, m, CH2), 1.43–1.30 (9H,
m, 4 Â CH2 and H-4b), 0.92 (3H, t, J = 6.27 Hz, CH3). 13C NMR
(50 MHz, CDCl3): d 195.5 (ArCO), 152.3, 146.3, 140.3, 132.7 (ArC),
128.3, 127.5 (ArCH), 127.1 (ArC), 111.9 (ArCH), 56.4, 56.5
(2 Â OMe), 43.5, 32.0, 29.6, 29.3 (NHCH2 and 3 Â CH2), 28.7, 28.3
4.1.4.19. (Z)-(2-(4-Chlorophenyl)cyclopropyl)(4-(octadec-9-enyl-
amino)phenyl)methanone (6q). White solid, mp 137–139 °C;
yield 80%; Rf 0.50 (10%, EtOAc/Hexane); IR (KBr) mmax cmÀ1: 3353
(N–H), 2923 (C–H), 1590 (C@O); 1H NMR (200 MHz, CDCl3): d
7.86 (2H, d, J = 8.72 Hz, ArH), 7.26 (2H, d, J = 8.32 Hz, ArH), 7.10
(2H, d, J = 8.48 Hz, ArH), 6.54 (2H, d, J = 8.74 Hz, ArH), 5.35–5.32
(2H, m, CH@CH), 4.20 (1H, br s, NH), 3.18–3.15 (2H, m, NHCH2),
2.81–2.69 (1H, m, H-2), 2.63–2.51 (1H, m, H-3), 2.10–1.89 (3H,
m, 1H, H-4a + CH2), 1.91–1.78 (1H, m, H-4b), 1.73–1.56 (2H, m,

A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
8299
(C-2 and C-3), 27.1, 23.2 (2 Â CH2), 18.6 (C-4), 14.4 (CH3). ESMS (m/
z): 396 (M+H)+. Anal. Calcd for C25H33NO3: C, 75.91; H, 8.41; N,
3.54. Found: C, 75.36; H, 8.38; N, 3.57.
4.1.5.3. (4-(2-(1H-Benzo[d]imidazol-1-yl)ethylamino)phenyl)(2-
(4-chlorophenyl)cyclop-ropyl)methanone (8c). White solid, mp
134–136 °C; yield 70%; Rf 0.45 (3%, MeOH/CHCl3); IR (KBr) mmax
cmÀ1: 3337 (N–H), 2932 (C–H), 1585 (C@O); 1H NMR (300 MHz,
CDCl3):
d 7.89–7.86 (3H, m, ArH), 7.80–7.65 (2H, m, ArH),
4.1.5. General procedure for the synthesis of 2-(azolyl)-
7.56–7.50 (1H, m, ArH), 7.36–7.25 (4H, m, ArH), 7.12 (2H, d,
J = 8.43 Hz, ArH), 6.58 (2H, d, J = 8.79 Hz, ArH), 4.43 (2H, t,
J = 5.31 Hz, CH2), 3.71 (2H, t, J = 5.70 Hz, CH2), 3.39 (1H, m, NH),
2.86–2.80 (1H, m, H-2), 2.59–2.53 (1H, m, H-3), 1.85–1.79 (1H,
m, H-4a), 1.48–1.42 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d
195.6 (ArCO), 152.3 (ArC), 144.4 (ArBzC), 143.6 (benzimidazole
CH), 140.5 (ArC), 134.1 (ArBzC), 132.2 (ArC), 131.2, 128.7, 127.5
(ArCH), 127.1 (ArC), 123.6, 123,2, 118.7, 109.9 (ArBzCH), 111.4
(ArCH), 48.7 (NBenzimidCH2), 43.3 (NHCH2), 28.8, 28.4 (C-2 and
C-3), 18.7 (C-4). ESMS (m/z): 416 (M+H)+. Anal. Calcd for
ethylaminoaryl phenyl cyclopropyl methanones (8a–8c)
A mixture of 2-(4-chlorophenyl)cyclopropyl)-4-(2-hydroxyeth-
ylamino)phenyl methanone (6s) (5.0 g, 15.87 mmol), TEA (triethyl-
amine, 2.65 ml, 19.0 mmol) in CH2Cl2 (20 ml) was stirred
magnetically at 0 °C for 15–20 min. A solution of methanesulphonyl
chloride (3.17 ml, 31.7 mmol in 5 ml CH2Cl2) was subsequently
added dropwise and the reaction mixture was stirred at 0 °C, for
4 h. After completion of the reaction (TLC), the reaction mixture
was diluted with excess of dichloromethane and extracted with
water. The organic layer was separated and dried (anhyd Na2SO4)
and concentrated under reduced pressure to get the intermediate
2-(4-(2-(4-chlorophenyl)cyclopropanecarbonyl)phenylami-
no)ethyl methanesulfonate (7) (4.80 g) and it was used as such with-
out further purification in the subsequent reaction with azoles.
To stirring slurry of NaH (0.11 g, 4.58 mmol) in DMF (4 mL) at
0 °C, the desired azole (1.2 equiv) was added and the reaction
mixture was stirred at 0 °C for 15–20 min. The above obtained 2-
(4-(2-(4-chlorophenyl)cyclopropanecarbonyl) phenylamino) ethyl
methanesulfonate (7) was added and stirring continued at
100–120 °C till the disappearance of starting material (TLC). The
reaction mixture was cooled to room temperature, quenched with
ethyl acetate and extracted with water. The organic layer was dried
(anhyd Na2SO4) and concentrated under reduced pressure to give a
crude mass which was purified by column chromatography (SiO2,
100–200 mesh) using gradient of hexane/ethyl acetate
(9:1?6:4)/0.2:9.8?0.6:9.4% methanol/chloroform to give the
desired compound (8a–8c) in varying yields.
C25H22ClN3O: C, 72.19; H, 5.33; N, 10.10. Found: C, 72.23; H,
5.36; N, 10.13.
4.2. Biology (assay methods)
4.2.1. Antimycobacterial assay
Determination of antitubercular activity against M. tuberculosis
H37Rv strain (Agar microdilution method): Drug susceptibility and
determination of MIC of the test compounds/drugs against M.
tuberculosis H37Rv was done by agar microdilution method.57 The
MIC of the test compounds was determined by incorporating
twofold dilution of this suspension were added to (in tubes)
7H10 middle brook’s medium (containing 1.7 mL medium and
0.2 mL OADC supplement) at different concentration of the test
compounds keeping the volume constant, that is, 0.1 mL. Medium
was allowed to cool keeping the tubes in slanting position. A
culture of M. tuberculosis H37Rv growing on L-J medium was
harvested in 0.85% saline with 0.05% Tween-80. A suspension of
1 lg/mL concentration of extracts/compounds was prepared in di-
4.1.5.1. (4-(2-(1H-Imidazol-1-yl)ethylamino)phenyl)(2-(4-chlo-
rophenyl)cyclopropyl)me-thanone (8a). White solid, mp 110–
methyl sulphoxide (DMSO). These tubes were then incubated at
37 °C for 24 h followed by streaking of M. tuberculosis H37Rv
(5 Â 105 bacilli per tube). The tubes were then incubated at
37 °C. Growth of bacilli was seen after 30 days of incubation. Tubes
having the compounds were compared with control tubes where
medium alone was incubated with H37Rv. The lowest concentra-
tion of the compound at which complete inhibition of colonies
occurred was taken as minimum inhibitory concentration (MIC)
of test compound.
112 °C; yield 67%; Rf 0.40 (5%, MeOH/CHCl3); IR (KBr) mmax cmÀ1
:
3334 (N–H), 2930 (C–H), 1588 (C@O); 1H NMR (300 MHz, CDCl3):
d 7.84 (2H, d, J = 8.82 Hz, ArH), 7.50 (1H, s, imidazole-H), 7.27
(2H, d, J = 8.46 Hz, ArH), 7.11 (2H, d, J = 8.46 Hz, ArH), 6.97 (2H, s,
imidazole-H), 6.57 (2H, d, J = 8.79 Hz, ArH), 4.19 (2H, t,
J = 5.61 Hz, CH2), 3.58 (2H, t, J = 5.67 Hz, CH2), 3.35 (1H, m, NH),
2.81–2.75 (1H, m, H-2), 2.59–2.53 (1H, m, H-3), 1.85–1.79 (1H,
m, H-4a), 1.48–1.42 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d
195.7 (ArCO), 152.6, 140.1 (ArC), 138.1 (imidazole-CH), 132.4
(ArC), 131.0 (ArCH), 128.5 (imidazole-CH), 128.9, 127.9 (ArCH),
127.2 (ArC), 121.2 (imidazole-CH), 111.7 (ArCH), 46.3 (NimidCH2),
43.3 (NHCH2), 28.8, 28.4 (C-2 and C-3), 18.7 (C-4). ESMS (m/z):
366 (M+H)+. Anal. Calcd for C21H20ClN3O: C, 68.94; H, 5.51; N,
11.49. Found: C, 68.87; H, 5.49; N, 11.53.
4.2.1.1. Antimalarial assay. The antimalarial assay was per-
formed in vitro against chloroquine sensitive Plasmodium falcipa-
rum 3D7 strains reported previously.58 Culture was maintained
in vitro as described by Trager and Jansen60 with some modifica-
tions
lL asynchronous culture of P. falciparum 3D7 was added.
The parasitaemia was maintained to ꢀ0.5% and hematocrit was
adjusted to 1.5%. Plates were incubated in CO2 incubator
maintained at 37 °C for 24 h. Radiolabelled hypoxanthine solution
4.1.5.2.
(4-(2-(1H-1,2,4-Triazol-1-yl)ethylamino)phenyl)(2-(4-
chlorophenyl)cyclopropyl) methanone (8b). White solid, mp
117–119 °C; yield 68%; Rf 0.45 (2%, MeOH/CHCl3); IR (KBr) mmax
cmÀ1: 3331 (N–H), 2934 (C–H), 1593 (C@O); 1H NMR (300 MHz,
CDCl3): d 8.17 (1H, s, triazole-H), 7.92 (1H, s, triazole-H), 7.81
(2H, d, J = 8.79 Hz, ArH), 7.25 (2H, d, J = 8.43 Hz, ArH), 7.10 (2H,
d, J = 8.43 Hz, ArH), 6.56 (2H, d, J = 8.79 Hz, ArH), 4.40 (2H, t,
J = 5.64 Hz, CH2), 3.58 (2H, t, J = 5.88 Hz, CH2), 3.34 (1H, m, NH),
2.79–2.74 (1H, m, H-2), 2.57–2.51 (1H, m, H-3), 1.83–1.78 (1H,
m, H-4a), 1.46–1.39 (1H, m, H-4b). 13C NMR (50 MHz, CDCl3): d
195.3 (ArCO), 152.6 (ArC), 150.9, 144.2 (triazole CH), 140.1, 132.4
(ArC), 131.0, 128.9, 127.9 (ArCH), 127.2 (ArC), 111.7 (ArCH), 48.7
(NTriazoleCH2), 43.3 (NHCH2), 28.8, 28.4 (C-2 and C-3), 18.7 (C-4).
ESMS (m/z): 367 (M+H)+. Anal. Calcd for C20H19ClN4O: C, 65.48;
H, 5.22; N, 15.27. Found: C, 65.54; H, 5.20; N, 15.30.
containing 0.5
l Ci/well was added. These plates were further
incubated for another 48 h. Cells were harvested on Whattmann
filter papers, transferred in scintillation vials, dried overnight.
Scintillation cocktail was added in these vials after 24 h, radioactiv-
ity was counted under scintillation b-counter. IC50 values of tested
compounds were calculated on the basis of radiolabelled hypoxan-
thine uptake by the parasites.
4.2.1.2. Antimalarial assay using minimum inhibitory concen-
tration (MIC). MICs of compounds were determined as described
by Rieckmann et al. (1978)61 with some modifications. Briefly,
stock solution of the compounds were prepared at 10 mg/ml in
DMSO and stored at 0 °C until use. The test was performed in 96-
well microtiter plates, twofold serial drug dilutions were prepared

8300
A. Ajay et al. / Bioorg. Med. Chem. 18 (2010) 8289–8301
in complete RPMI 1640 and 50
well. Parasitized RBCs (50 lL) were added to each well. The final
l
L of each dilution was used in each
4.2.4.3. Reporter gene expression analysis. Recombinant M. aur-
um strains were grown in Sauton’s medium with Kanamycin at
37 °C to 0.5 OD600 after which culture was diluted to 0.04–0.05
OD with fresh medium. Ten millilitre of diluted culture were dis-
tributed to separate tubes, equilibrated for 2 h at 37 °C and then
culture suspension had a hematocrit of 3–4% with 1.0–2.0%
infection (>95% rings). Micro culture plates were incubated for
24–32 h at 37 °C in an incubator supplied with 5% CO2 to allow
the development of malaria parasites, culture plates were taken
out and maximum supernatant medium was removed, thin blood
smear of each well content were made and stained with 15% Giem-
sa stain. These smears were checked for the maturation of schizo-
nts relative to their controls.
varying concentrations (10, 25, 50 and 75 lM) of compounds were
added to different tubes. Following 12 h incubation at 37 °C, 5 ml
cultures from each tube were pelleted, washed and resuspended
in PBS (Phosphate Buffer Saline, pH 7.2), sonicated at 4 °C and
supernatant was collected by centrifugation at 13000 rpm for
10 min at 4 °C. Protein contents were quantified using Bradford As-
say reagent (Sigma B6916) as per manufacturer protocol. b-Gal as-
say was performed from total cellular protein as described
earlier.65 Briefly, same amount of protein were mixed with
4.2.3. Cell cytotoxicity assay
The cytotoxic assay was carried out against monkey kidney cell
line C1008 (Vero cells). The cells were cultivated in 25 cm2 tissue
culture flask supplemented with MEM-medium (9.7 g MEM,
2.2 g/l NaHCO3, 6 g HEPES, Gentamycine sulphate 50 mg, ampho-
tericin B 2.5 mg, TDW-1000 ml) + 15%FBS provided with 5% CO2
at 37 °C. The culture medium was changed on alternate days. The
growth rate was determined as described.62 For the cytotoxicity as-
say the cells were washed with PBS, trypsinized with 0.25% trypsin
and a cell suspension was made in culture medium. The cells were
counted in Neubaur chamber and appropriate dilution was made
200
lL of ONPG (4 mg/ml) and incubated for 30 min at 37 °C. Reac-
tion was stopped by adding 500
l
L of 1 M Na2CO3 and optical den-
sity was measured at 410 nm. Experiments were carried out in
triplicates for each treatment and b-galactosidase units were calcu-
lated for each set individually. The culture at each point was also
plated to confirm the decline in viability of cells after drug treat-
ment. The whole experiment was repeated twice and similar
trends in results were obtained. Mean value and standard
deviation were calculated and plotted for each set of data.
(1 Â 105 cells/ml). Vero cell suspension (100
lL) was added to
the microtiter plates and allowed to adhere overnight. Serial dilu-
tions of test compounds were prepared in these plates and Vero
cells were incubated with these compounds for 72 h. Resazurin
was added in wells and after 4 h these plates were read under
florescence reader (Biotek). Cytotoxic concentration (CC50) was
determined using MS-EXEL.
4.2.5. Data analysis
All data were expressed as mean SD with at least three sepa-
rate experiments. IC50 were determined with linear regression
analysis using Microsoft Excel. Statistically significant comparison
was calculated using student’s t-test for unpaired variants. Values
of p <0.05 were regarded as statistically significant.
4.2.4. FAS-II inhibitory studies
4.2.4.1. In silico studies. The three-dimensional structure of the
compound 4a was built and optimized using the Builder module
of Insight II (M/s Accelrys Inc.). The compound was then taken as
probe and submitted to an in house developed web-based tool
called Inhibitor Identification Tool (IS-IT) (unpublished). The tool of-
fers docking against 85 potential drug targets specifically chosen
from Mycobacterium tuberculosis. Missing residues and atoms of
each protein structure were repaired using the biopolymer module
of Sybyl 6.8 (Tripos Associates), and Kollman63,64 charges were
assigned to the protein. The grid for docking calculations was cen-
tered on the binding site of each protein. Docking simulations were
carried out by using AUTODOCK 4.0, and interaction energies between
compounds and the proteins were calculated using the scoring
function of AUTODOCK 4.0. The protein target with interaction
energies greater than the control docking energies was selected
for further enzymatic assays.
Acknowledgments
Author thanks, CSIR and DRDO New Delhi for financial assis-
tance. Arya Ajay is thankful to UGC New Delhi for SRF. It is a CDRI
communication 7967.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmc.2010.09.071.
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