Synthesis of benzamide derivatives of anacardic acid and their cytotoxic activity

Article abstract of DOI:10.1016/j.ejmech.2009.01.033

Several benzamide derivatives were synthesized from anacardic acid (1a) which was the product of hydrogenation of the naturally occurring anacardic acid mixture (1a-d), a major constituent of cashew nut shell liquid. Anacardic acid (1a) was first alkylated followed by hydrolysis of the ester to obtain synthones namely, 2-ethoxy-6-pentadecylbenzoic acid (5) and 2-isopropoxy-6-pentadecylbenzoic acid (6). These salicylic acid derivatives were then coupled with a variety of anilines to obtain novel benzamide compounds (7-39). Cytotoxic effect of these synthesized compounds was tested on HeLa cell line of wild type with relatively high expression of p300 and on HCT-15, which is p300 negative. Of all the compounds, 2-isopropoxy-6-pentadecyl-N-pyridin-4-ylbenzamide (27), 2-ethoxy-N-(3-nitrophenyl)-6-pentadecylbenzamide (22) and 2-ethoxy-6-pentadecyl-N-pyridin-4-ylbenzamide (10) were found to be more potent with the respective IC₅₀ values 11.02 μM, 13.55 μM, 15.29 μM on HeLa cell line. Their activities are comparable with garcinol which is a cell permeable histone acetyltransferase (HAT) inhibitor and 10 fold more active than p300 HAT activators so far reported.

Full text of DOI:10.1016/j.ejmech.2009.01.033

European Journal of Medicinal Chemistry 44 (2009) 2711–2719
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Synthesis of benzamide derivatives of anacardic acid and their cytotoxic activity
*
Venkateshappa Chandregowda, Anil Kush, Goukanapalli Chandrasekara Reddy
Vittal Mallya Scientific Research Foundation, P.O. Box 406, K.R. Road, Bangalore 560004, Karnataka, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Several benzamide derivatives were synthesized from anacardic acid (1a) which was the product of
hydrogenation of the naturally occurring anacardic acid mixture (1a–d), a major constituent of cashew
nut shell liquid. Anacardic acid (1a) was first alkylated followed by hydrolysis of the ester to obtain
synthones namely, 2-ethoxy-6-pentadecylbenzoic acid (5) and 2-isopropoxy-6-pentadecylbenzoic acid
(6). These salicylic acid derivatives were then coupled with a variety of anilines to obtain novel benza-
mide compounds (7–39). Cytotoxic effect of these synthesized compounds was tested on HeLa cell line of
wild type with relatively high expression of p300 and on HCT-15, which is p300 negative. Of all the
compounds, 2-isopropoxy-6-pentadecyl-N-pyridin-4-ylbenzamide (27), 2-ethoxy-N-(3-nitrophenyl)-6-
pentadecylbenzamide (22) and 2-ethoxy-6-pentadecyl-N-pyridin-4-ylbenzamide (10) were found to be
Received 22 May 2008
Received in revised form
20 November 2008
Accepted 29 January 2009
Available online 5 February 2009
Keywords:
Cashew nut shell liquid
Anacardic acid
more potent with the respective IC50 values 11.02 mM, 13.55 mM, 15.29 mM on HeLa cell line. Their
Benzamides
HeLa and HCT-15 cell lines
Cytotoxicy
activities are comparable with garcinol which is a cell permeable histone acetyltransferase (HAT)
inhibitor and 10 fold more active than p300 HAT activators so far reported.
Ó 2009 Elsevier Masson SAS. All rights reserved.
1. Introduction
Cl
O
O
Cashew nut shell liquid (CNSL) is a by-product of cashew nut
industry and anacardic acid mixture (1a–d) isolated from CNSL
which are salicylic acid derivatives with a nonisoprenoid alk(en)yl
side chain [1].
NH
CF3
2
Anacardic acid and its derivatives have been tested for various
biological activities viz., antimicrobial activity [2,3] and soybean
lipoxygenase-1 inhibitory activity [4,5]. Earlier from our lab we
have reported the synthesis of sildenafil analogues [6], dihy-
dropyridine analogues [7] as calcium channel blockers, iso-
nicotinoylhydrazones for antimycobacterial activity [8] starting
from anacardic acid. Kubo et al., reported the separation of ana-
cardic acid into monoene (15:1), diene (15:2) and triene (15:3) by
preparative HPLC and tested against cancer cells, and found to show
moderate cytotoxic activity on BT-20 breast and HeLa epithelioid
cervix carcinoma cells [9]. It has been proved that Anacardic acid
has a unique function of mediating changes in membrane potential
and pH gradient across liposomal membranes [10]. Anacardic acid
was found to form highly lipophilic metal derivatives with selec-
tivity towards the first row transition metals such as Feþ2, Cuþ2 and
Znþ2 having selective ionophoric properties [11]. An amide deriv-
ative of anacardic acid (2) has been reported to be the first specific
activator of histone acetyltransferase (HAT) activity of p300 [12]
while anacardic acid (1a) showed to be HAT inhibitor.
Recently, a few anacardic acid derivatives such as 2 were
reported and elucidated their mechanism of HAT activation and
suggested that, 2 and its derivatives binds to p300 predominantly
to the amide group of a-helix and b-sheets and affect the structure
of the enzyme [13]. But all the reported compounds were not cell
permeable and showed inhibition at higher concentrations of the
compound. More recently, it has been shown that anacardic acid is
a specific activator of kinase activity of Aurora Kinase A [14],
suppresses expression of nuclear factor-kB regulated gene products
leading to potentiation of apoptosis [15] and inhibitor of the HAT
activity of recombinant Plasmodium falciparum GCN5 [16]. Sbar-
della et al. recently showed that long chain alkylidenemalonates
which are structurally related to anacardic acid as modulators of
histone acetyltransferases [17].
In view of potential of anacardic acid and its derivatives as
anticancer agents, it was proposed to make and identify potent
benzamide derivatives from a natural product anacardic acid iso-
lated from Anacardium occideltale L., test them for anticancer
activity against HeLa cell line [18].
The aim of the present work is to make use of abundantly and
cheaply available anacardic acid and to make synthons for gener-
ating novel cell permeable benzamide compounds and check for
* Corresponding author. Tel.: þ91 80 2661 1664; fax: þ91 80 2661 2806.
E-mail address: gcreddy@vmsrf.org (G.C. Reddy).
0223-5234/$ – see front matter Ó 2009 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2009.01.033

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V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
their antiproliferative activity. This was achieved by, (a) Saturation
of naturally occurring anacardic acid mixture followed by alkyl-
ation of hydroxyl and carboxylic acid groups (b) Hydrolysis of ester
to acid (c) Converting acid to acid chloride and then coupling with
substituted anilines and (d) testing these compounds for their
cytotoxic effect on HeLa and HCT-15 cell lines.
cytotoxic effect, we found that compound 27 was the most potent
(IC50 ¼ 11.02 M), which was comparable to a cell permeable HAT
M) and 10 fold more active than
m
inhibitor garcinol (IC50 ¼ 9.13
m
p300 HAT activators reported earlier [13].
3. Conclusion
2. Results and discussion
We have synthesized novel benzamide derivatives from ana-
cardic acid synthones. Compounds 27, 22 and 10 were found to be
2.1. Chemistry
more potent with IC50 values 11.02 mM, 13.55 mM, 15.29 mM
respectively on HeLa cell line and their activities are comparable
with garcinol, which is a cell permeable HAT inhibitor and 10 fold
more active than p300 HAT activators so far reported.
Thus, these novel compounds would be useful as biological
switching molecules for probing the role of p300 HAT in cellular
physiology.
The anacardic acid mixture (Fig. 1, 1a–d) was isolated from CNSL
by a reported method [19]. Accordingly CNSL was treated with
calcium hydroxide, during which anacardic acid present in CNSL
becomes calcium anacardate, which was isolated and hydrolyzed
with dil. hydrochloric acid to generate anacardic acid ene mixture,
which was a mixture of monoene, diene and triene located at (80),
(80, 110) and (80, 110, 140) of the C15 alkyl chain respectively. O-ethyl
anacardic acid (5) and O-isopropyl anacardic acid (6) (Scheme 1)
were prepared by hydrogenation of anacardic acid mixture (1a–d)
using hydrogen and Pd/C to get saturated anacardic acid (1a), which
was then reacted with diethyl sulfate and isopropyl bromide
respectively in presence of base to obtain diethyl anacardate (3) and
diisopropyl anacardate (4).
The title compounds (7–39, Table 1) were synthesized from
compounds 5 and 6 (Scheme 2) by treating corresponding 2-
alkoxy-6-pentadecylbenzoic acid (5/6) with thionyl chloride to get
acid chloride and then coupled with substituted anilines to get
benzamides. Pure compounds were obtained by recrystallization
using hexane. All the compounds were fully characterized by using
IR, NMR and Mass spectroscopy.
1H NMR, 13C NMR, mass spectra of compounds 7–39 are avail-
able free of charge via Internet at http:/www.sciencedirect.org.
4. Experimental
4.1. General methods/instrument
CNSL was obtained from commercial source and all other
chemicals and reagents of laboratory grade were obtained from
local dealers and were used without further purification. IR spectra
were recorded on a Nicolet Avatar 320 FT-IR spectrometer. 1H and
13C NMR spectra were recorded in CDCl3/DMSO-d6 at 200 MHz on
a Bruker A G Spectrometer. Chemical shifts are reported in
d units
downfield from TMS as internal standard. Mass spectra were
recorded using GC–MS-QP2010S (Direct probe) and on Q-TOF
microÔ AMPS MAX 10/6A system. CHN analysis was recorded using
ThermoFinnigan FLASH EA 1112 CHNS analyzer. Melting points
were recorded on Acro Steel Pvt. Ltd. melting point apparatus and
are uncorrected.
2.2. Antiproliferative activity
Benzamide compounds (7–39) were tested on HeLa cell line of
wild type with relatively more p300 expressed [18] and with p300
negative HCT-15. Inhibition of cell proliferation was measured by
MTT assay. Several benzamide molecules exhibited very good
cytotoxic activity against HeLa cell line (Table 1). Mantelingu et al.
[13] have reported that benzamide molecules derived from ana-
cardic acid, bearing chloro and trifluoromethyl groups on the
aniline moiety are poor cell permeable and are specific p300 HAT
activators. We have also observed the same effect with our
compounds 8 and 12. In order to find out the influence of other
substituents on phenyl ring we have made several compounds
having substitution other than halogen and trifluoromethyl groups.
We found that, compounds 10, 22 and 27 showed very good
inhibitory activities followed by compounds 16 and 31.
4.2. Solvent extracted CNSL
Solvent extracted CNSL containing anacardic acid mixture 63%,
cardanol 10.5% and cardols 22% was obtained from cashew pro-
cessing industry, Mangalore (Karnataka), India.
4.3. Isolation of anacardic acid mixture from natural CNSL
Commercially available solvent extracted CNSL (100 g) was
dissolved in 5% aqueous methanol (600 mL). Activated charcoal
(20 g) was added and stirred for 15 min. This solution was filtered
over celite to remove insoluble plant materials. The clear filtrate
was transferred into a round bottomed flask fitted with a double
But all these compounds exhibited poor cytotoxic activity on
p300 negative cell line (HCT-15) with inhibitory concentrations of
surface reflux condenser and
a mechanical stirrer. Calcium
(IC50) >300
m
M. Thus, these preliminary results give an indication
hydroxide (50 g) was added in portions under stirring. After
complete addition of calcium hydroxide, the temperature of the
reaction mixture was raised to 50 ꢀC and stirred for 3 h. Superna-
tant solution was monitored by TLC for the absence of anacardic
acid. After the completion of the reaction, the precipitated calcium
anacardate was filtered and washed thoroughly with methanol
(200 mL) and the cake was dried under vacuum at 45–50 ꢀC for 2 h
(dry weight 110 g). The calcium anacardate was suspended in
distilled water (440 mL) and 11 M HCl (60 mL) was added and
stirred for 1 h. The resultant solution was extracted with ethyl
acetate (2 ꢁ150 mL). The combined organic layers were washed
with distilled water (2 ꢁ 100 mL), dried over anhydrous sodium
sulfate and concentrated under reduced pressure to yield anacardic
acid mixture (60 g). The identity of the compound was confirmed
by HPLC and in comparison with standard samples [19,21].
that, some of benzamide compounds derived from anacardic acid
are cell permeable and likely to act as modulators of HAT activity of
p300. Hence, of the several compounds synthesized and tested for
C15H31-n
a
OH
n=0
COOH
b
n=2
C15H31-n
c
n=4
1
d
n=6
Fig. 1. Anacardic acid mixture.

V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
2713
OEt
OEt
OH
COOEt
C15H31
COOH
C15H31
COOH
C15H31
Anacardic acid
ene mixture
iii
v
i
ii
1a-d
5
3
1a
iv
O-iPr
O-iPr
COO-iPr
C15H31
COOH
C15H31
4
6
Scheme 1. Synthetic pathway of 5 and 6; Reagents and conditions: (i) MeOH/Pd/C/H2 2.5 kg/cm2 at RT for 2 h; (ii) Diethyl sulfate/K2CO3/Acetone/Reflux for 3 h; (iii) DMSO/t-BuOK/
40 ꢀC for 2 h; (iv) i-PrBr/MIBK/K2CO3/Bu4NHSO4/Reflux for 24 h; (v) DMSO/t-BuOK/40 ꢀC for 2 h.
4.4. Preparation of intermediates
compound 5 as a brown solid; (6.5 g, Yield ¼ 70%); m.p. 60–62 ꢀC;
IR (KBr): 2916, 2846, 1705, 1585, 1461, 1396, 1265, 1118, 1076 cmꢂ1
;
4.4.1. Preparation of anacardic acid (1a)
1H NMR (CDCl3, 200 MHz):
d
0.88 (t, J ¼ 6.0 Hz, 3H), 1.25 (brs, 24H),
Anacardic acid mixture (30 g) was dissolved in methanol
(120 mL). 5% Pd/C (0.75 g) was added slowly and this solution was
transferred into hydrogenation flask. Hydrogenation was carried
out at 2.5 kg/cm2 pressure for 2 h, and the solution was filtered
through a celite bed to obtain catalyst-free solution. Organic
solvent was evaporated under vacuum to obtain anacardic acid
(1a), which was recrystallized from petroleum ether (40–60 ꢀC);
Yield ¼ 27 g; m.p. 90–91 ꢀC; IR (KBr): 3140, 2917, 1850, 1655, 1604,
1.45 (t, J ¼ 7.0 Hz, 3H), 1.59 (m,2H), 2.78 (t, J ¼ 8.0 Hz, 2H), 4.15 (q,
J ¼ 7.0 Hz, 2H), 6.80 (d, J ¼ 8.0 Hz,1H), 6.88 (d, J ¼ 7.8 Hz,1H), 7.30 (t,
J ¼ 8.0 Hz, 1H); GC–MS (DI, m/z): 376 (Mþ).
4.4.5. Preparation of 2-isopropoxy-6-pentadecylbenzoic acid (6)
Using the procedure described for compound 5, compound 6
was prepared from compound 4 in 70% yield; m.p. 52–54 ꢀC; IR
(KBr): 2916, 2850, 1701, 1581, 1465, 1384, 1265, 1118, 1026 cmꢂ1; 1H
1466, 1308, 1248, 1206, 894, 815, 757 cmꢂ1
;
1H NMR (200 MHz,
NMR (CDCl3, 200 MHz):
d
0.88 (t, J ¼ 6.2 Hz, 3H), 1.25 (brs, 24H),
CDCl3):
d
0.88 (t, 3H, J ¼ 6.0 Hz), 1.25(brs, 24H), 1.6(m, 2H), 2.98(t,
1.40 (d, J ¼ 6.0 Hz, 6H), 1.56 (m, 2H), 2.85 (t, J ¼ 8.0 Hz, 2H), 4.58–
4.76 (m, 1H), 6.84 (d, J ¼ 8.0 Hz, 1H), 6.89 (d, J ¼ 8.0 Hz, 1H), 7.35 (t,
J ¼ 8.0 Hz, 1H); GC–MS (m/z): 390 (Mþ).
2H, J ¼ 8.0 Hz), 6.7–6.8 (dd, 2H, J ¼ 8.0 Hz), 7.36(t, 1H, J ¼ 8.0 Hz).
4.4.2. Preparation of ethyl-2-ethoxy-6-pentadecylbenzoate (3)
To a stirred solution of 1a (12 g, 34.8 mmol) in acetone (60 mL)
was added anhydrous powdered potassium carbonate (14.4 g,
104 mmol). Diethyl sulfate (10.7 g, 69.6 mmol) was added in
portions for about 10 min at RT. After the addition was complete,
the reaction mixture was heated to reflux and stirred for 3 h. The
reaction mixture was cooled to RT and organic solvent was
concentrated under reduced pressure. Water (100 mL) was added
to the reaction mixture and extracted with ethyl acetate (80 mL).
The organic layer was separated, washed with water and dried over
anhydrous sodium sulfate. The organic layer was evaporated to
obtain product 3 as a viscous liquid; Yield: Quantitative; IR (KBr):
2920, 1730, 1260 cmꢂ1; GC–MS (DI, m/z): 404 (Mþ).
4.5. General procedure for the preparation of final compounds
(7–39)
To a stirred solution of 2-alkoxy-6-pentadecylbenzoic acid (5 or
6) (5 mmol) in hexane (10 vol) was added thionyl chloride (1/2 vol)
and catalytic amount of DMF (2 drops). The reaction mixture was
refluxed for about 2 h. Distilled off excess thionyl chloride along
with hexane under reduced pressure to obtain 2-alkoxy-6-pentad
ecyl benzoyl chloride. The acid chloride obtained was dissolved in
dichloromethane (5 vol) and added slowly to a pre-mixed stirred
solution of substituted aniline (6 mmol) and triethylamine
(5 mmol) in dichloromethane1 (5 vol), by keeping the temperature
of the reaction mixture at 15–20 ꢀC. After the complete addition of
acid chloride, the reaction mixture was allowed to stir at 35–40 ꢀC
for about 1 h. Water was added to the reaction mixture. The organic
phase was separated and washed with dil. HCl, and water. Finally
dried over anhydrous sodium sulfate, filtered and concentrated
organic solvent to obtain crude material of title compounds. Pure
compounds were obtained by recrystallization in hexane with
yields more than 80%.
4.4.3. Preparation of isopropyl-2-isopropoxy-6-pentadecylbenzoate
(4)
To a stirred solution of 1a (100 g, 0.29 mol) in methyl isobutyl
ketone (700 mL), potassium carbonate (130 g, 0.94 mol) and tet-
rabutylammonium hydrogen sulfate (2 g), was added isopropyl
bromide (90 mL, 0.96 mol). The reaction mass was refluxed for
about 24 h, cooled to RT and added water (500 mL) and ethyl
acetate (500 mL). Organic layer was separated, washed with water
(2 ꢁ 100 mL) and brine solution (1 ꢁ100 mL). Organic layer was
evaporated to obtain product 4 as a brown colored liquid in
quantitative yield; IR (KBr): 2925, 1732, 1264 cmꢂ1; GC–MS (DI, m/
z): 432 (Mþ).
4.5.1. N-(3-chloro-4-fluorophenyl)-2-ethoxy-6-
pentadecylbenzamide (7)
Using 5 and 3-chloro-4-fluoroaniline as starting materials, the
title compound
7 was obtained as a white solid (2.27 g,
Yield ¼ 85%); m.p. 85–89 ꢀC; IR (KBr): 3259, 2920, 1654, 1596, 1527,
1500, 1465, 1392, 1265, 1114, 1080, 813, 763, 698 cmꢂ1 1H NMR
;
4.4.4. Preparation of 2-ethoxy-6-pentadecylbenzoic acid (5)
To a stirred solution of 3 (10 g, 24.7 mmol) in DMSO (50 mL) was
added potassium tert-butoxide (3 g, 24.7 mmol). The reaction
mixture was stirred at RT for about 2 h and then heated to 40 ꢀC,
stirred for another 2 h. The reaction mass was cooled to RT and
quenched in ice-water, adjusted pH to 2 with hydrochloric acid. The
resulting precipitate was filtered and dried at RT to obtain
(CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.23 (brs, 24H), 1.37 (t,
J ¼ 6.9 Hz, 3H), 1.61 (m, 2H), 2.71 (t, J ¼ 7.7 Hz, 2H), 4.07 (q,
J ¼ 6.9 Hz, 2H), 6.78 (d, J ¼ 8.4 Hz,1H), 6.87 (d, J ¼ 7.6 Hz, 1H), 7.11 (t,
1
DMF was used as solvent for the preparation of compounds 10 and 28.

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V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
Table 1
Cytotoxic effects of compounds 7–39 on HeLa cell linea (Cervical epithelial adenocarcinoma cell line) and HCT-15 (Colon carcinoma cell line).b
Compound
R
R1
[IC50
(m
M) ꢃ SE]c HeLa
Compound
R
R1
[IC50
(
m
M) ꢃ SE]c HeLa
Cl
G*
9.13 ꢃ 0.61
12
Et
351.01 ꢃ 11.95
CF3
CH3
F
7
8
Et
Et
112.38 ꢃ 6.82
335.12 ꢃ 9.59
13
14
Et
Et
128.45 ꢃ 6.17
140.51 ꢃ 11.83
Cl
Br
CF3
OCH3
CH3
NH2
N
H3C
Ph
N
9
Et
Et
44.25 ꢃ 3.50
15
16
Et
Et
65.41 ꢃ 2.71
O
N
10
15.29 ꢃ 0.67
19.39 ꢃ 0.53
NH2
Br
11
18
Et
342.02 ꢃ 10.10
17
Et
100.51 ꢃ 8.68
CF3
CH3
OCH3
Et
35.60 ꢃ 3.14
29
i-Pr
60.44 ꢃ 5.84
CH
F
CH3
NH2
19
20
Et
Et
36.40 ꢃ 2.30
50.51 ꢃ 4.73
30
31
i-Pr
i-Pr
60.48 ꢃ 6.38
32.12 ꢃ 1.81
Br
21
Et
155.34 ꢃ 8.21
32
i-Pr
57.97 ꢃ 3.87
I
NH2

V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
2715
Table 1 (continued )
Compound
R
R1
[IC50
(m
M) ꢃ SE]c HeLa
Compound
R
R1
[IC50
(
m
M) ꢃ SE]c HeLa
22
Et
Et
13.55 ꢃ 0.89
33
i-Pr
43.19 ꢃ 2.77
NO2
CH
23
79.03 ꢃ 1.70
34
i-Pr
275.31 ꢃ 14.61
CF3
CH3
24
25
Et
135.25 ꢃ 8.60
90.51 ꢃ 6.09
35
36
i-Pr
i-Pr
200.20 ꢃ 10.98
170.03 ꢃ 7.52
Br
F
F
i-Pr
I
Cl
CH3
N
H3C
Ph
N
26
i-Pr
220.28 ꢃ 10.77
37
i-Pr
92.59 ꢃ 4.54
NO2
O
N
27
28
i-Pr
i-Pr
11.02 ꢃ 0.68
38
39
i-Pr
i-Pr
135.81 ꢃ 5.87
361.41 ꢃ 9.66
CH3
Br
200.21 ꢃ 17.37
F
CH3
*G ¼ Garcinol, a cell permeable HAT inhibitor was used as control (lit IC50 ¼ 7
mM) [20].
a
Determined by MTT assay.
b
All compounds showed IC50 values >300
m
M on HCT-15 which has p300 negative.
c
Values are average of at least three independent experiments ꢃ standard error, conducted in triplicate samples for each concentration.
J ¼ 8.6 Hz, 1H), 7.23–7.27 (m, 1H), 7.31–7.36 (m, 1H), 7.52 (s, 1H),
688 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.23
7.80 (dd, J ¼ 6.5 and 2.5 Hz, 1H); 13C NMR (CDCl3, 50 MHz):
d
14.55,
(brs, 24H), 1.37 (t, J ¼ 6.9 and 7.0 Hz, 3H), 1.61 (m, 2H), 2.71 (t,
J ¼ 7.7 Hz, 2H), 4.07 (q, J ¼ 6.9 and 7.0 Hz, 2H), 6.78 (d, J ¼ 8.0 Hz,
1H), 6.88 (d, J ¼ 7.6 Hz, 1H), 7.28 (t, J ¼ 8.0 Hz, 1H), 7.65–7.84 (m,
15.28, 23.12, 29.88, 30.10, 31.92, 32.35, 33.72, 64.79, 110.03, 116.84,
117.28, 119.84, 119.97, 122.52, 122.66, 130.93, 135.00, 143.42, 155.88,
157.20, 166.51; GC–MS (DI, m/z): 503 (Mþ).
4H); 13C NMR (CDCl3, 50 MHz):
d 14.52, 15.23, 23.10, 29.84, 30.08,
31.89, 32.34, 33.72, 64.85, 110.10, 114.00, 119.23, 119.28, 120.20,
122.79, 124.25, 125.79, 131.11, 135.96, 138.04, 143.62, 155.91, 166.63;
GC–MS (DI, m/z): 597 (Mþ).
4.5.2. N-[4-bromo-3-(trifluoromethyl)phenyl]-2-ethoxy-6-
pentadecylbenzamide (8)
Using 5 and 4-bromo-3-(trifluoromethyl)aniline as starting
materials, the title compound 8 was obtained as a white solid
(3.67 g, Yield ¼ 85%); m.p. 60–63 ꢀC; IR (KBr): 3348, 3271, 3105,
2919, 2850, 1683, 1652, 1582, 1511, 1464, 1407, 1253, 1122, 821,
4.5.3. N-(4-antipyrinyl)-2-ethoxy-6-pentadecylbenzamide (9)
Using 5 and antipyrine as starting materials, the title compound
9 was obtained as a dark brown solid. (2.53 g, Yield ¼ 85%); m.p.

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V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
OR
O
4.5.7. 2-Ethoxy-N-(4-methylphenyl)-6-pentadecylbenzamide (13)
Using 5 and 4-methylaniline as starting materials, the title
OR
R1
COOH
C15H31
N
H
i, ii
compound 13 was obtained as
a off-white solid (2.27 g,
Yield ¼ 85%); m.p. 78–80 ꢀC; IR (KBr): 3282, 2915, 2846, 1651, 1596,
C15H31
1523, 1465, 1400, 1257, 1114, 810, 759, 690 cmꢂ1 1H NMR (CDCl3,
;
7-39
200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 24H), 1.36 (t, J ¼ 6.9
5/6
and 7.0 Hz, 3H), 1.62 (m, 2H), 2.34 (s, 3H), 2.72 (t, J ¼ 7.8 Hz, 2H),
4.06 (q, J ¼ 6.9 and 7.0 Hz, 2H), 6.77 (d, J ¼ 8.2 Hz, 1H), 6.86 (d,
J ¼ 7.6 Hz, 1H), 7.16 (d, J ¼ 8.2 Hz, 2H), 7.21–7.30 (m, 1H), 7.48 (d,
Scheme 2. Synthetic pathway of 7–39, where R and R1 are as described in Table 1.
Reagents and conditions: (i) SOCl2/Hexane/Reflux for 2 h/Distill off excess SOCl2; (ii)
CH2Cl2/TEA/Aniline/40 ꢀC for 1 h.
J ¼ 8.2 Hz, 2H); 13C NMR (CDCl3, 50 MHz):
d 14.56, 15.28, 21.33,
23.13, 29.90, 30.11, 31.91, 32.36, 33.71, 64.73, 100.04, 120.52, 122.49,
126.98, 129.96, 130.52, 134.35, 136.01, 143.20, 155.89, 166.36; GC–
MS (DI, m/z): 465 (Mþ); Anal. C31H47NO2: C, 79.95; H, 10.17; N, 3.01.
Found: C, 79.62; H, 9.92; N, 3.22.
93–96 ꢀC; IR (KBr): 3200, 2920, 2850, 1670, 1625, 1610, 1550, 1465,
1390, 1300, 1261, 1080, 705, 585 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.24 (brs, 24H), 1.42 (t, J ¼ 6.9 Hz, 3H), 1.62
(m, 2H), 2.40 (s, 3H), 2.71 (t, J ¼ 7.7 Hz, 2H), 3.11 (s, 3H), 4.07 (q,
J ¼ 6.9 Hz, 2H), 6.75 (d, J ¼ 8.2 Hz, 1H), 6.84 (d, J ¼ 7.6 Hz, 1H), 7.16–
4.5.8. 2-Ethoxy-N-(4-methoxyphenyl)-6-pentadecylbenzamide
(14)
7.32 (m, 3H), 7.39–7.50 (m, 4H); 13C NMR (CDCl3, 50 MHz):
d 13.00,
14.55, 15.30, 23.12, 29.75, 30.11, 31.98, 32.35, 33.83, 36.70, 64.60,
109.07, 109.60, 122.15, 124.46, 126.37, 127.08, 129.62, 130.48, 135.28,
142.73, 150.10, 156.04, 162.01, 167.12; GC–MS (DI, m/z): 561 (Mþ).
Using 5 and 4-methoxyaniline as starting materials, the title
compound 14 was obtained as a grey solid. (2 g, Yield ¼ 80%); m.p.
84–87 ꢀC; IR (KBr): 3286, 2923, 2850, 1647, 1596, 1515, 1465, 1407,
1249, 1114, 1033, 825, 690 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d 0.87
4.5.4. 2-Ethoxy-6-pentadecyl-N-pyridin-4-ylbenzamide (10)
Using 5 and 4-aminopyridine as starting materials, the title
compound 10 was obtained as a cream colored solid. (1.92 g,
Yield ¼ 80%); m.p. 87–90 ꢀC; IR (KBr): 3232, 3155, 2920, 2850, 1689,
1589, 1527, 1461, 1396, 1307, 1272, 1207, 1114, 995, 829, 763, 594,
(t, J ¼ 6.1 Hz, 3H), 1.29 (brs, 24H), 1.42 (t, J ¼ 6.9 Hz, 3H), 1.67 (m,
2H), 2.77 (t, J ¼ 7.7 Hz, 2H), 3.85 (s, 3H), 4.10 (q, J ¼ 6.9 Hz, 2H), 6.81
(d, J ¼ 8.2 Hz, 1H), 6.89 (d, J ¼ 7.6 Hz, 1H), 6.94 (d, J ¼ 8.8 Hz, 2H),
7.26–7.34 (m, 1H), 7.44 (s, 1H), 7.54 (d, J ¼ 8.8 Hz, 2H); 13C NMR
(CDCl3, 50 MHz):
d 14.56, 15.30, 23.12, 29.90, 30.11, 31.92, 32.36,
540 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.23
33.73, 55.94, 64.75, 110.03, 114.65, 122.39, 125.50, 127.20, 130.49,
(brs, 24H), 1.35 (t, J ¼ 6.9 Hz, 3H), 1.59 (m, 2H), 2.70 (t, J ¼ 7.7 Hz,
2H), 4.05 (q, J ¼ 6.9 Hz, 2H), 6.77 (d, J ¼ 8.2 Hz, 1H), 6.89 (d,
J ¼ 7.6 Hz,1H), 7.26 (t, J ¼ 7.6 Hz,1H), 7.53 (brs, 2H), 7.95 (s, 1H), 8.48
131.66, 143.17, 155.91, 156.94, 166.37; GC–MS (DI, m/z): 481 (Mþ).
4.5.9. N-(4-aminophenyl)-2-ethoxy-6-pentadecylbenzamide (15)
Using 5 and p-phenylenediamine as starting materials, the title
compound 15 was obtained as a light brown solid. (2.18 g,
Yield ¼ 88%); m.p. 90–93 ꢀC; IR (KBr): 3282, 2920, 2850, 1651, 1593,
(brs, 2H); 13C NMR (CDCl3, 50 MHz):
d 14.54, 15.20, 23.10, 29.83,
30.08, 31.85, 32.33, 33.69, 64.69, 109.95, 113.96, 122.61, 125.96,
131.04, 143.30, 145.82, 150.83, 155.87, 167.37; HRMS (m/z): Calcu-
lated for C29H44N2O2 (M þ H)þ ¼ 453.2481, Found ¼ 453.3460.
1523, 1461, 1423, 1323, 1265, 1114, 1076, 825, 759 cmꢂ1 1H NMR
;
(CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 24H), 1.37 (t,
4.5.5. N-(4-bromo-2-ethylphenyl)-2-ethoxy-6-
pentadecylbenzamide (11)
J ¼ 7.0 Hz, 3H), 1.59 (m, 2H), 2.72 (t, J ¼ 7.8 Hz, 2H), 3.59 (brs, 2H),
4.05 (q, J ¼ 7.0 Hz, 2H), 6.70 (d, J ¼ 8.6 Hz, 2H), 6.76 (d, J ¼ 8.6 Hz,
1H), 6.85 (d, J ¼ 7.6 Hz, 1H), 7.13–7.32 (m, 2H), 7.36 (d, J ¼ 8.6 Hz,
Using 5 and 4-bromo-2-ethylaniline as starting materials, the
title compound 11 was obtained as a light brown solid (2.61 g,
Yield ¼ 88%); m.p. 99–101 ꢀC; IR (KBr): 3232, 2920, 1654, 1593,
2H); 13C NMR (CDCl3, 50 MHz):
d 14.56, 15.30, 23.12, 29.80, 30.11,
31.92, 32.36, 33.71, 64.73, 110.01, 115.91, 122.40, 122.71, 127.00,
129.50, 130.39, 143.08, 143.82, 155.90, 167.00; HRMS (m/z): Calcu-
lated for C30H46N2O2 (M þ H)þ ¼ 467.3637, Found ¼ 467.3641.
1512, 1461, 1392, 1269, 1114, 1076, 810, 721 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 27H), 1.38 (t, J ¼ 6.9
and 7.0 Hz, 3H), 1.64 (m, 2H), 2.67 (m, 4H), 4.08 (q, J ¼ 6.9 and
7.0 Hz, 2H), 6.79 (d, J ¼ 8.2 Hz, 1H), 6.87 (d, J ¼ 7.6 Hz, 1H), 7.23–7.38
4.5.10. N-(2-aminophenyl)-2-ethoxy-6-pentadecylbenzamide (16)
Using 5 and o-phenylenediamine as starting materials, the title
compound 16 was obtained as a white solid. (2.1 g, Yield ¼ 85%);
m.p. 88–90 ꢀC; IR (KBr): 3267, 2920, 2850, 1647, 1589, 1523, 1461,
(m, 4H), 7.61 (d, J ¼ 11.2 Hz, 1H); 13C NMR (CDCl3, 50 MHz):
d 14.29,
14.56, 15.27, 23.10, 24.48, 29.80, 30.12, 32.10, 32.36, 33.82, 64.72,
109.91, 119.18, 122.49, 126.10, 127.00, 130.09, 130.67, 131.83, 134.65,
138.35, 143.15, 155.84, 166.95; GC–MS (DI, m/z): 558 (Mþ); Anal.
C32H49NO2: C, 80.12; H, 10.30; N, 2.92. Found: C, 79.94; H, 10.25; N,
3.11.
1299, 1265, 1114, 1076, 752 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d 0.87
(t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 24H), 1.43 (t, J ¼ 7.0 Hz, 3H), 1.60–1.71
(m, 2H), 2.75 (t, J ¼ 7.8 Hz, 2H), 4.10 (q, J ¼ 7.0 Hz, 2H), 6.76–6.89 (m,
4H), 7.08–7.31 (m, 4H); 13C NMR (CDCl3, 50 MHz):
d 14.54, 15.29,
4.5.6. N-[4-chloro-2-(trifluoromethyl)phenyl]-2-ethoxy-6-
pentadecylbenzamide (12)
Using 5 and 4-chloro-2-(trifluoromethyl)aniline as starting
materials, the title compound 12 was obtained as a white solid
(2.5 g, Yield ¼ 85%); m.p. 84–86 ꢀC; IR (KBr): 3225, 3082, 2965,
2921, 2850, 1657, 1585, 1506, 1463, 1413, 1264, 1132, 1056, 818,
23.11, 29.78, 30.10, 32.07, 32.34, 33.86, 64.72, 109.68, 117.51, 119.32,
122.40, 123.37, 126.80, 127.08,128.33, 130.53, 142.60, 142.75, 155.80,
167.21; HRMS (m/z): Calculated for C30H46N2O2 (M þ Na)þ
¼ 489.3457, Found ¼ 489.3463.
4.5.11. 2-Ethoxy-6-pentadecyl-N-[3-
(trifluoromethyl)phenyl]benzamide (17)
663 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.25
(brs, 24H), 1.35 (t, J ¼ 6.9 Hz, 3H), 1.58 (m, 2H), 2.70 (t, J ¼ 7.8 Hz,
2H), 4.08 (q, J ¼ 6.9 Hz, 2H), 6.78 (d, J ¼ 8.3 Hz, 1H), 6.87 (d,
J ¼ 7.6 Hz, 1H), 7.28 (t, J ¼ 8.0 Hz, 1H), 7.54–7.60 (m, 2H), 7.78 (s, 1H),
Using 5 and 3-(trifluoromethyl)aniline as starting materials, the
title compound 17 was obtained as a cream colored solid. (2.34 g,
Yield ¼ 85%); m.p. 59–61 ꢀC; IR (KBr): 3286, 3066, 2920, 2850,
8.43 (d, J ¼ 8.5 Hz, 1H); 13C NMR (CDCl3, 50 MHz):
d
14.52, 14.92,
1658, 1581, 1535, 1469, 1392, 1334, 1172, 1118, 879, 794, 698 cmꢂ1
;
23.10, 29.83, 30.08, 31.87, 32.34, 33.69, 64.63, 109.83, 121.00, 122.49,
126.06, 126.20, 126.57, 126.65, 130.00,131.05,133.29, 134.50, 143.34,
155.90, 167.00; GC–MS (DI, m/z): 553 (Mþ).
1H NMR (CDCl3, 200 MHz):
d
0.86 (t, J ¼ 6.1 Hz, 3H), 1.23 (brs, 24H),
1.37 (t, J ¼ 6.9 Hz, 3H), 1.61 (m, 2H), 2.73 (t, J ¼ 7.7 Hz, 2H), 4.08 (q,
J ¼ 6.9 Hz, 2H), 6.78 (d, J ¼ 8.2 Hz, 1H), 6.88 (d, J ¼ 7.6 Hz, 1H), 7.28–

V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
2717
7.49 (m, 3H), 7.65 (s, 1H), 7.85 (brs, 2H); GC–MS (DI, m/z): 519
(t, J ¼ 8.1 Hz, 1H), 7.79 (s, 1H), 7.99 (d, J ¼ 8.1 Hz, 1H), 8.06 (d,
(Mþ).
J ¼ 7.8 Hz,1H), 8.40 (s,1H); 13C NMR (CDCl3, 50 MHz):
d 14.53,15.26,
23.11, 29.87, 30.09, 31.90, 32.34, 33.74, 64.82, 110.06, 114.89, 119.20,
122.75, 125.80, 125.94, 130.29, 131.13, 139.72, 143.59, 149.08, 155.93,
166.83; Anal. C30H44N2O4: C, 72.55; H, 8.93; N, 5.64. Found: C,
72.65; H, 9.05; N, 5.54.
4.5.12. 2-Ethoxy-N-(3-ethynylphenyl)-6-pentadecylbenzamide (18)
Using 5 and 3-ethynylaniline as starting materials, the title
compound 18 was obtained as a light brown solid (2.27 g,
Yield ¼ 90%); m.p. 88–90 ꢀC; IR (KBr): 3281, 3060, 2955, 2920,
2848, 2106, 1654, 1532, 1461, 1401, 1271, 1114, 875, 687 cmꢂ1
;
1H
4.5.17. 2-Ethoxy-N-(2-ethylphenyl)-6-pentadecylbenzamide (23)
Using 5 and 2-ethylaniline as starting materials, the title
NMR (CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.23 (brs, 24H),
1.36 (t, J ¼ 6.9 Hz, 3H), 1.60 (m, 2H), 2.72 (t, J ¼ 7.7 Hz, 2H), 3.06 (s,
1H), 4.06 (q, J ¼ 6.9 Hz, 2H), 6.77 (d, J ¼ 8.3 Hz, 1H), 6.86 (d,
J ¼ 7.6 Hz, 1H), 7.22–7.35 (m, 3H), 7.52 (s, 1H), 7.64 (brd, J ¼ 7.3 Hz,
compound 23 was obtained as a off-white solid (2.42 g,
Yield ¼ 95%); m.p. 77–79 ꢀC; IR (KBr): 3244, 2923, 2846, 1647, 1523,
1461, 1392, 1265, 1114, 1076, 798, 744 cmꢂ1 1H NMR (CDCl3,
;
1H), 7.71 (s, 1H); 13C NMR (CDCl3, 50 MHz):
d
14.51, 15.25, 23.10,
200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 27H), 1.39 (t, J ¼ 6.9
29.88, 30.09, 31.87, 32.34, 33.70, 64.83, 77.81, 83.69, 110.13, 120.88,
120.61, 123.31, 123.66, 126.57, 128.40, 129.48, 130.74, 138.64, 143.36,
155.90, 166.49; GC–MS (DI, m/z): 475 (Mþ).
and 7.0 Hz, 3H), 1.62 (m, 2H), 2.70 (m, 4H), 4.09 (q, J ¼ 6.9 and
7.0 Hz, 2H), 6.79 (d, J ¼ 8.2 Hz, 1H), 6.87 (d, J ¼ 7.6 Hz, 1H), 7.13–7.30
(m, 5H), 7.88 (d, J ¼ 7.5 Hz, 1H); 13C NMR (CDCl3, 50 MHz):
d 14.10,
14.20, 14.86, 22.70, 24.24, 29.37, 29.63, 31.69, 31.94, 33.40, 64.33,
109.54, 121.99, 124.41, 125.80, 126.70, 128.60, 130.03, 135.12, 136.09,
140.00, 142.67, 155.49, 166.54; GC–MS (DI, m/z): 479 (Mþ).
4.5.13. 2-Ethoxy-N-(3-fluorophenyl)-6-pentadecylbenzamide (19)
Using 5 and 3-fluoroaniline as starting materials, the title
compound 19 was obtained as
a light brown solid (3 g,
Yield ¼ 90%); m.p. 65–67 ꢀC; IR (KBr): 3284, 3066, 2919, 2850, 1663,
4.5.18. 2-Ethoxy-N-(2-fluorophenyl)-6-pentadecylbenzamide (24)
Using 5 and 2-fluoroaniline as starting materials, the title
compound 24 was obtained as a cream colored solid (2.37 g,
Yield ¼ 95%); m.p. 65–68 ꢀC; IR (KBr): 3244, 2920, 2846, 1658, 1593,
1599, 1528, 1435, 1257, 1114, 1061, 858, 768, 683 cmꢂ1 1H NMR
;
(CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.23 (brs, 24H), 1.36 (t,
J ¼ 6.9 and 7.0 Hz, 3H), 1.60 (m, 2H), 2.71 (t, J ¼ 7.7 Hz, 2H), 4.06 (q,
J ¼ 6.9 and 7.0 Hz, 2H), 6.75–6.88 (m, 3H), 7.17–7.34 (m, 3H), 7.58 (d,
1523, 1461, 1392, 1265, 1110, 1072, 914, 748, 690 cmꢂ1 1H NMR
;
J ¼ 10.6 Hz, 1H), 7.61 (s, 1H); 13C NMR (CDCl3, 50 MHz):
d
14.51,
(CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 24H), 1.36 (t,
15.24, 23.10, 29.85, 30.08, 31.87, 32.34, 33.71, 64.83, 110.13, 111.10,
111.53, 122.65, 126.44, 130.37, 130.55, 130.81, 140.02, 140.24, 143.42,
155.90, 166.50; GC–MS (DI, m/z): 469 (Mþ).
J ¼ 6.9 and 7.0 Hz, 3H), 1.61 (m, 2H), 2.75 (t, J ¼ 7.7 Hz, 2H), 4.08 (q,
J ¼ 6.9 and 7.0 Hz, 2H), 6.78 (d, J ¼ 8.2 Hz, 1H), 6.87 (d, J ¼ 7.6 Hz,
1H), 7.07–7.31 (m, 4H), 7.86 (s, 1H), 8.48 (t, J ¼ 7.4 and 7.9 Hz, 1H);
HRMS (m/z): Calculated for C30H44NO2F (M þ H)þ ¼ 470.3434,
Found ¼ 470.3419.
4.5.14. N-(3-bromophenyl)-2-ethoxy-6-pentadecylbenzamide (20)
Using 5 and 3-bromoaniline as starting materials, the title
compound 20 was obtained as a cream color solid (2.59 g,
Yield ¼ 92%); m.p. 75–77 ꢀC; IR (KBr): 3271, 2920, 2846, 1654, 1585,
4.5.19. N-(3-chloro-4-fluorophenyl)-2-isopropoxy-6-
pentadecylbenzamide (25)
1523, 1396, 1269, 1114, 1076, 860, 759, 682 cmꢂ1
;
1H NMR (CDCl3,
Using 6 and 3-chloro-4-fluoroaniline as starting materials, the
200 MHz):
d
0.87 (t, J ¼ 6 Hz, 3H), 1.23 (brs, 24H), 1.37 (t, J ¼ 6.9 and
title compound 25 was obtained as a brown solid (2.63 g,
7.0 Hz, 3H), 1.61 (m, 2H), 2.71 (t, J ¼ 7.7 Hz, 2H), 4.07 (q, J ¼ 6.9 and
7.0 Hz, 2H), 6.77 (d, J ¼ 8.2 Hz, 1H), 6.87 (d, J ¼ 7.6 Hz, 1H), 7.17–7.31
(m, 3H), 7.47–7.53 (m, 2H), 7.87 (s, 1H); 13C NMR (CDCl3, 50 MHz):
Yield ¼ 90%); m.p. 77–80 ꢀC; IR (KBr): 3271, 2920, 2846, 1654, 1593,
1500, 1461, 1384, 1269, 1114, 810, 760 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.23 (brs, 24H), 1.31 (d,
d
14.17, 14.88, 22.74, 29.50, 29.72, 31.50, 31.97, 33.31, 64.42, 109.69,
J ¼ 6.0 Hz, 6H), 1.56 (brs, 2H), 2.72 (t, J ¼ 7.7 Hz, 2H), 4.54 (m, 1H),
6.80 (d, J ¼ 8.4 Hz,1H), 6.87 (d, J ¼ 7.6 Hz, 1H), 7.12 (t, J ¼ 8.6 Hz, 1H),
7.22–7.38 (m, 2H), 7.53 (s, 1H), 7.80 (dd, J ¼ 6.5 and 2.5 Hz, 1H); 13C
118.30, 120.90, 122.24, 122.76, 125.96, 127.24, 130.35, 130.45, 139.50,
143.00, 155.49, 166.11; GC–MS (DI, m/z): 529 (Mþ).
NMR (CDCl3, 50 MHz):
d 14.10, 22.18, 22.68, 29.45, 29.67, 31.46,
4.5.15. 2-Ethoxy-N-(3-iodophenyl)-6-pentadecylbenzamide (21)
Using 5 and 3-iodoaniline as starting materials, the title
compound 21 was obtained as a cream colored solid (2.76 g,
Yield ¼ 90%); m.p. 83–85 ꢀC; IR (KBr): 3263, 2920, 2846, 1654, 1577,
31.92, 33.34, 71.67, 111.83, 116.43, 116.87, 119.35, 119.48, 122.06,
122.46, 130.41, 134.84, 143.34, 154.53, 157.20, 166.13; GC–MS (DI, m/
z): 517 (Mþ).
1519, 1461, 1400, 1269, 1114, 1076, 860, 759 cmꢂ1
;
1H NMR (CDCl3,
4.5.20. N-(4-antipyrinyl)-2-isopropoxy-6-pentadecylbenzamide
(26)
200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.23 (brs, 24H), 1.37 (t, J ¼ 6.9
and 7.0 Hz, 3H), 1.61 (m, 2H), 2.71 (t, J ¼ 7.7 Hz, 2H), 4.08 (q, J ¼ 6.9
and 7.0 Hz, 2H), 6.77 (d, J ¼ 8.2 Hz, 1H), 6.86 (d, J ¼ 7.6 Hz, 1H), 7.07
(t, J ¼ 8.0 Hz, 1H), 7.27 (t, J ¼ 8.2 and 7.6 Hz, 1H), 7.45–7.58(m, 3H),
Using 6 and antipyrine as starting materials, the title compound
26 was obtained as a brown solid (2.36 g, Yield ¼ 80%); m.p. 105–
107 ꢀC; IR (KBr): 3210, 2923, 2850, 1655, 1630, 1593, 1465, 1390,
8.01 (s, 1H); 13C NMR (CDCl3, 50 MHz):
d
14.12, 14.85, 22.69, 29.46,
1295, 1261, 1114, 1022, 702, 590 cmꢂ1 1H NMR (CDCl3, 200 MHz):
;
29.68, 31.45, 31.93, 33.27, 64.42, 94.24, 109.70, 119.02, 122.21,
126.00, 128.46, 130.41, 130.50, 133.27, 139.36, 142.98, 155.47, 166.01;
GC–MS (DI, m/z): 577 (Mþ).
0.87 (t, J ¼ 6.2 Hz, 3H), 1.24 (brs, 24H), 1.36 (d, J ¼ 6 Hz, 6H), 1.60 (m,
2H), 2.40 (s, 3H), 2.71 (t, J ¼ 7.7 Hz, 2H), 3.11 (s, 3H), 4.58 (s,1H), 6.77
(d, J ¼ 8.4 Hz, 1H), 6.82 (d, J ¼ 7.7 Hz, 1H), 7.13 (s, 1H), 7.19–7.32 (m,
2H), 7.43–7.50 (m, 4H); GC–MS (DI, m/z): 575 (Mþ); Anal.
C36H53N3O3: C, 75.09; H, 9.28; N, 7.30. Found: C, 74.99; H, 9.43; N,
7.69.
4.5.16. 2-Ethoxy-N-(3-nitrophenyl)-6-pentadecylbenzamide (22)
Using 5 and 3-nitroaniline as starting materials, the title
compound 22 was obtained as a light yellow solid (2.32 g,
Yield ¼ 88%); m.p. 94–96 ꢀC; IR (KBr): 3282, 2920, 2846, 1654, 1535,
4.5.21. 2-Isopropoxy-6-pentadecyl-N-pyridin-4-ylbenzamide (27)
Using 6 and 4-aminopyridine as starting materials, the title
1465, 1400, 1346, 1257, 1110, 871, 732 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
(t, J ¼ 6.2 Hz, 3H), 1.23 (brs, 24H), 1.38 (t, J ¼ 6.9 Hz,
compound 27 was obtained as a light brown solid (1.91 g,
3H),1.60 (m, 2H), 2.73 (t, J ¼ 7.7 Hz, 2H), 4.09 (q, J ¼ 6.9 Hz, 2H), 6.80
Yield ¼ 80%); m.p. 70–73 ꢀC; IR (KBr): 3280, 3140, 2916, 2850, 1666,
(d, J ¼ 8.2 Hz,1H), 6.89 (d, J ¼ 7.5 Hz, 1H), 7.30 (t, J ¼ 8.2 Hz, 1H), 7.53
1589, 1525, 1465, 1400, 1269, 1110, 850 cmꢂ1 1H NMR (CDCl3,
;

2718
V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
200 MHz):
d
0.86 (t, J ¼ 6.6 Hz, 3H), 1.24 (brs, 24H), 1.31 (d,
129.49, 129.85, 142.98, 143.38, 154.49, 165.96; GC–MS (DI, m/z): 480
(Mþ); Anal. C31H48N2O2: C, 77.45; H, 10.06; N, 5.83, Found: C, 77.40;
H, 10.35; N, 5.99.
J ¼ 6.0 Hz, 6H), 1.58 (m, 2H), 2.72 (t, J ¼ 7.7 Hz, 2H), 4.56 (m, 1H),
6.81 (d, J ¼ 8.0 Hz,1H), 6.88 (d, J ¼ 7.6 Hz,1H), 7.29 (t, J ¼ 8.0 Hz,1H),
7.63 (brs, 2H), 8.12 (s, 1H), 8.51 (brs, 2H); 13C NMR (CDCl3, 50 MHz):
d
14.12, 22.15, 22.69, 29.44, 29.68, 31.48, 31.93, 33.43, 71.72, 111.69,
4.5.26. N-(2-aminophenyl)-2-isopropoxy-6-pentadecylbenzamide
(32)
114.00, 122.69, 126.15, 130.84, 143.54, 146.56, 148.89, 154.66,167.03;
HRMS (m/z): Calculated for C30H46N2O2 (M þ H)þ ¼ 467.3637,
Found ¼ 467.3647.
Using 6 and o-phenylenediamine as starting materials, the title
compound 32 was obtained as a white solid (2.0 g, Yield ¼ 85%);
m.p. 76–78 ꢀC; IR (KBr): 3247, 2920, 2846, 1647, 1585, 1523, 1461,
4.5.22. N-(4-bromo-2-ethylphenyl)-2-isopropoxy-6-
pentadecylbenzamide (28)
1384, 1269, 1114, 1026, 744 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d 0.88
(t, J ¼ 6.2 Hz, 3H), 1.24 (brs, 24H), 1.36 (d, J ¼ 6.0 Hz, 6H), 1.63 (m,
2H), 2.75 (t, J ¼ 7.9 Hz, 2H), 4.08 (brs, 2H), 4.62 (m, 1H), 6.78–6.88
(m, 4H), 7.07–7.30 (m, 4H); HRMS (m/z): Calculated for C31H48N2O2
(M þ Na)þ ¼ 503.3613, Found ¼ 503.3630.
Using 6 and 4-bromo-2-ethylaniline as starting materials, the
title compound 28 was obtained as a light grey solid (2.49 g,
Yield ¼ 85%); m.p. 77–80 ꢀC; IR (KBr): 3236, 2920, 2846, 1651, 1600,
1510, 1461, 1400, 1265, 1114, 820, 725 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 27H), 1.31 (d,
4.5.27. N-(3-ethynylphenyl)-2-isopropoxy-6-pentadecylbenzamide
(33)
Using 6 and 3-ethynylaniline as starting materials, the title
compound 33 was obtained as a cream colored solid (2.25 g,
Yield ¼ 90%); m.p. 70–72 ꢀC; IR (KBr): 3283, 3061, 2920, 2849, 2107,
1655, 1531, 1464, 1402, 1264, 1115, 873, 688 cmꢂ1; 1H NMR (CDCl3,
J ¼ 6.0 Hz, 6H), 1.60 (m, 2H), 2.67 (m, 4H), 4.56 (m, 1H), 6.80 (d,
J ¼ 8.3 Hz, 1H), 6.86 (d, J ¼ 7.6 Hz, 1H), 7.22–7.38 (m, 4H), 7.84 (d,
J ¼ 9.18 Hz, 1H); 13C NMR (CDCl3, 50 MHz):
d 13.84, 14.10, 22.13,
22.67, 23.97, 29.34, 29.66, 31.61, 31.90, 33.37, 71.25, 111.31, 118.59,
122.01, 125.41, 127.34, 129.63, 130.09, 131.34, 134.25, 137.68, 142.82,
154.40, 166.56; GC–MS (DI, m/z): 572 (Mþ).
200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.23 (brs, 24H), 1.31 (d,
J ¼ 6.0 Hz, 6H), 1.58 (m, 2H), 2.73 (t, J ¼ 7.7 Hz, 2H), 3.07 (s, 1H), 4.54
(m,1H), 6.80 (d, J ¼ 8.3 Hz,1H), 6.86 (d, J ¼ 7.5 Hz,1H), 7.22–7.36 (m,
3H), 7.53 (s, 1H), 7.62 (brd, J ¼ 7.3 Hz, 1H), 7.71 (s, 1H); 13C NMR
4.5.23. 2-Isopropoxy-N-(4-methoxyphenyl)-6-
pentadecylbenzamide (29)
Using 6 and 4-methoxyaniline as starting materials, the title
(CDCl3, 50 MHz): d 14.17, 22.21, 22.74, 29.51, 29.72, 31.50, 31.97,
compound 29 was obtained as
a
light grey solid (2.28 g,
33.37, 71.65, 77.45, 83.32, 111.82, 120.43, 122.39, 123.00, 123.21,
127.20, 127.99, 129.12, 130.29, 138.26, 143.27, 154.53, 166.19; GC–MS
(DI, m/z): 489 (Mþ).
Yield ¼ 90%); m.p. 76–78 ꢀC; IR (KBr): 3300, 3100, 2920, 2850, 1643,
1508, 1461, 1400, 1242, 1118, 1037, 840, 770 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
0.88 (t, J ¼ 6.1 Hz, 3H), 1.23 (brs, 24H), 1.31 (d,
J ¼ 6.0 Hz, 6H), 1.61 (m, 2H), 2.74 (t, J ¼ 7.7 Hz, 2H), 3.81 (s, 3H), 4.54
(m, 1H), 6.79 (d, J ¼ 8.0 Hz, 1H), 6.86 (d, J ¼ 7.6 Hz, 1H), 6.90 (d,
J ¼ 8.0 Hz, 2H), 7.24 (t, J ¼ 8.0 Hz, 1H), 7.40 (s, 1H), 7.49 (d, J ¼ 8.0 Hz,
4.5.28. 2-Isopropoxy-6-pentadecyl-N-[3-
(trifluoromethyl)phenyl]benzamide (34)
Using 6 and 3-trifluoromethyl aniline as starting materials, the
title compound 34 was obtained as a white solid (2.18 g,
2H); 13C NMR (CDCl3, 50 MHz):
d
14.09, 22.15, 22.66, 29.45, 29.65,
31.43, 31.89, 33.30, 55.47, 71.51, 111.79, 114.20, 121.89, 122.20, 127.61,
129.93, 131.23, 143.01, 154.47, 156.45, 165.97; GC–MS (DI, m/z): 495
(Mþ).
Yield ¼ 88%); m.p. 54–56 ꢀC; IR (KBr): 3282, 2920, 2846, 1658, 1610,
1535, 1460, 1400, 1334, 1250, 1118, 800, 700 cmꢂ1; 1H NMR (CDCl3,
200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.23 (brs, 24H), 1.31 (d,
J ¼ 6.0 Hz, 6H), 1.60 (m, 2H), 2.74 (t, J ¼ 7.7 Hz, 2H), 4.55 (m, 1H),
6.81 (d, J ¼ 8.3 Hz, 1H), 6.87 (d, J ¼ 7.5 Hz, 1H), 7.25–7.52 (m, 3H),
7.67 (s, 1H), 7.82 (d, J ¼ 9.8 Hz, 1H), 7.84 (s, 1H); 13C NMR (CDCl3,
4.5.24. 2-Isopropoxy-N-(4-methylphenyl)-6-pentadecylbenzamide
(30)
Using 6 and 4-methylaniline as starting materials, the title
compound 30 was obtained as a cream colored solid (2.21 g,
Yield ¼ 90%); m.p. 88–90 ꢀC; IR (KBr): 3320, 3100, 2920, 2850, 1651,
50 MHz): d 14.16, 22.17, 22.73, 29.49, 29.71, 31.51, 31.97, 33.37, 71.68,
111.79, 116.49, 120.81, 122.46, 122.90, 123.00, 126.86, 129.64, 130.46,
131.50, 138.76, 143.39, 154.56, 166.34; GC–MS (DI, m/z): 533 (Mþ).
1596, 1527, 1461, 1400, 1261, 1114, 850, 750 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
0.92 (t, J ¼ 6.2 Hz, 3H), 1.28 (brs, 24H), 1.34 (d,
4.5.29. N-(3-bromophenyl)-2-isopropoxy-6-pentadecylbenzamide
(35)
J ¼ 6.0 Hz, 6H), 1.67 (m, 2H), 2.38 (s, 3H), 2.78 (t, J ¼ 7.7 Hz, 2H), 4.57
(m, 1H), 6.83 (d, J ¼ 8.4 Hz, 1H), 6.90 (d, J ¼ 7.6 Hz, 1H), 7.24 (t,
J ¼ 8.4 Hz,1H), 7.27 (d, J ¼ 8.4 Hz, 2H), 7.49 (s,1H), 7.51 (d, J ¼ 8.4 Hz,
Using 6 and 3-bromoaniline as starting materials, the title
compound 35 was obtained as a cream colored solid. (2.65 g,
Yield ¼ 95%); m.p. 61–64 ꢀC; IR (KBr): 3274, 2920, 2846, 1654, 1585,
2H); 13C NMR (CDCl3, 50 MHz):
d 14.09, 20.86, 22.15, 22.67, 29.45,
29.65, 31.42, 31.90, 33.31, 71.60, 111.90, 120.06, 122.27, 127.71,
129.51, 129.96, 133.84, 135.61, 143.08, 154.49, 165.96; GC–MS (DI, m/
z): 479 (Mþ).
1523, 1465, 1410, 1265, 1114, 860, 763 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 24H), 1.31 (d,
J ¼ 6.0 Hz, 6H), 1.55 (brs, 2H), 2.72 (t, J ¼ 7.7 Hz, 2H), 4.54 (m, 1H),
6.80 (d, J ¼ 8.3 Hz, 1H), 6.86 (d, J ¼ 7.5 Hz, 1H), 7.17–7.30 (m, 3H),
4.5.25. N-(4-aminophenyl)-2-isopropoxy-6-pentadecylbenzamide
(31)
7.45–7.61 (m, 2H), 7.87 (s, 1H); 13C NMR (CDCl3, 50 MHz):
d 14.56,
22.59, 23.13, 29.88, 30.11, 31.88, 32.36, 33.75, 72.04, 112.18, 118.61,
121.00, 122.81, 123.10, 127.61, 128.90, 130.70, 130.76, 139.90, 143.72,
154.92, 166.55; GC–MS (DI, m/z): 544 (Mþ).
Using 6 and p-phenylenediamine as starting materials, the
title compound 31 was obtained as
a light brown solid
(2.1 g, Yield ¼ 88%); m.p. 69–71 ꢀC; IR (KBr): 3294, 2920, 2850,1651,
1593, 1519, 1465, 1423, 1323, 1265, 1114, 1026, 825 cmꢂ1
;
1H NMR
4.5.30. N-(3-iodophenyl)-2-isopropoxy-6-pentadecylbenzamide
(36)
(CDCl3, 200 MHz):
d
0.87 (t, J ¼ 6.1 Hz, 3H), 1.24 (brs, 24H), 1.31 (d,
J ¼ 6.0 Hz, 6H), 1.59–1.62 (m, 2H), 2.73 (t, J ¼ 7.8 Hz, 2H), 4.53 (m,
1H), 6.70 (d, J ¼ 8.6 Hz, 2H), 6.78 (d, J ¼ 8.1 Hz, 1H), 6.85 (d,
J ¼ 7.6 Hz, 1H), 7.22–7.26 (m, 1H), 7.33 (s, 1H), 7.35 (d, J ¼ 8.6 Hz,
Using 6 and 3-iodoaniline as starting materials, the title
compound 36 was obtained as a light brown solid (2.88 g,
Yield ¼ 95%); m.p. 66–68 ꢀC; IR (KBr): 3271, 2920, 2846, 1654, 1581,
2H); 13C NMR (CDCl3, 50 MHz):
d
14.13, 22.20, 22.70, 29.38, 29.69,
1523, 1469, 1400, 1268, 1140, 890, 760 cmꢂ1 1H NMR (CDCl3,
;
31.46, 31.93, 33.33, 71.53, 111.84, 115.49, 122.10, 122.24, 127.50,
200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.24 (brs, 24H), 1.31 (d,

V. Chandregowda et al. / European Journal of Medicinal Chemistry 44 (2009) 2711–2719
2719
J ¼ 6.0 Hz, 6H), 1.56 (brs, 2H), 2.72 (t, J ¼ 7.7 Hz, 2H), 4.53 (m, 1H),
6.80 (d, J ¼ 8.3 Hz,1H), 6.86 (d, J ¼ 7.6 Hz,1H), 7.08 (t, J ¼ 8.0 Hz,1H),
7.26 (t, J ¼ 7.6 and 8.3 Hz, 1H), 7.45–7.56 (m, 3H), 8.02 (s, 1H); 13C
cells were treated with different concentrations of the compound
(1 M to 300 M, final concentration of DMSO < 1%). The cells were
later incubated for 72 h. The cytotoxicity was measured by adding
5 mg/mL of MTT (Sigma–Aldrich Inc., USA) to each well and incu-
bated for another 3 h. The purple formazan crystals were dissolved
by adding 100 ml of DMSO to each well. The absorbance was read at
570 nm in a spectrophotometer [Spectra Max 340]. The cell death
was calculated as follows:
m
m
NMR (CDCl3, 50 MHz):
d 14.12, 22.15, 22.69, 29.45, 29.67, 31.44,
31.92, 33.29, 71.60, 94.50, 111.75, 118.93, 122.36, 127.00, 128.39,
130.31, 130.51, 133.22, 138.50, 143.25, 154.50, 166.50; GC–MS (DI, m/
z): 591 (Mþ).
4.5.31. 2-Isopropoxy-N-(3-nitrophenyl)-6-pentadecylbenzamide
(37)
Cell death ¼ 100 ꢂ ½ðtest absorbance=control absorbanceÞ
ꢁ 100ꢄ
Using 6 and 3-nitroaniline as starting materials, the title
compound 37 was obtained as
a light brown solid (2.19 g,
Yield ¼ 84%); m.p. 87–90 ꢀC; IR (KBr): 3286, 2920, 2846, 1654, 1593,
The test result is expressed as the concentration of a test
compound which inhibits the cell growth by 50% (IC50).
1535, 1465, 1410, 1360, 1253, 1118, 870, 732 cmꢂ1 1H NMR (CDCl3,
;
200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.23 (brs, 24H), 1.32 (d,
J ¼ 6.0 Hz, 6H), 1.62 (m, 2H), 2.74 (t, J ¼ 7.7 Hz, 2H), 4.55 (m, 1H),
6.82 (d, J ¼ 8.3 Hz, 1H), 6.84 (d, J ¼ 7.7 Hz, 1H), 7.25–7.57 (m, 1H),
7.53 (t, J ¼ 8.1 Hz, 1H), 7.81 (s, 1H), 7.96–8.05 (m, 2H), 8.41 (s, 1H);
Acknowledgement
We thank Mr. A. C. Karunakara, Vittal Mallya Scientific Research
Foundation, for his analytical support and Mrs. Neelima and Ms.
Shwetha for their help in doing biological assay.
13C NMR (CDCl3, 50 MHz):
d 14.11, 22.18, 22.69, 29.46, 29.67, 31.48,
31.93, 33.39, 71.72, 111.79, 114.41, 118.78, 122.56, 125.44, 126.39,
129.90,130.65,139.32,143.54,148.74,154.57,166.43; GC–MS (DI, m/
z): 510 (Mþ).
Appendix. Supplementary data
4.5.32. N-(2-ethylphenyl)-2-isopropoxy-6-pentadecylbenzamide
(38)
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.ejmech.2009.01.033.
Using 6 and 2-ethylaniline as starting materials, the title
compound 38 was obtained as
a off-white solid (2.27 g,
Yield ¼ 90%); m.p. 88–91 ꢀC; IR (KBr): 3283, 3061, 2920, 2849, 2107,
References
1655, 1531, 1464, 1402, 1264, 1115, 873, 688 cmꢂ1; 1H NMR (CDCl3,
200 MHz):
d
0.87 (t, J ¼ 6.2 Hz, 3H), 1.24 (brs, 27H), 1.32 (d,
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J ¼ 6.0 Hz, 6H), 1.64 (m, 2H), 2.70 (m, 4H), 4.58 (m, 1H), 6.80 (d,
J ¼ 8.3 Hz, 1H), 6.86 (d, J ¼ 7.6 Hz, 1H), 7.13–7.30 (m, 5H), 7.89 (d,
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4.5.33. N-(2-fluorophenyl)-2-isopropoxy-6-pentadecylbenzamide
(39)
Using 6 and 2-fluoroaniline as starting materials, the title
compound 39 was obtained as a white solid (2.35 g, Yield ¼ 95%);
m.p. 54–57 ꢀC; IR (KBr): 3244, 2920, 2846, 1658, 1610, 1520, 1461,
1400, 1265, 1118, 920, 748 cmꢂ1; 1H NMR (CDCl3, 200 MHz):
d 0.87
(t, J ¼ 6.0 Hz, 3H), 1.24 (brs, 24H), 1.32 (d, J ¼ 6.0 Hz, 6H), 1.59 (m,
2H), 2.75 (t, J ¼ 7.7 Hz, 2H), 4.58 (m,1H), 6.80 (d, J ¼ 8.2 Hz,1H), 6.86
(d, J ¼ 7.6 Hz, 1H), 7.05–7.30 (m, 4H), 7.88 (s, 1H), 8.47 (t, J ¼ 8.0 Hz,
1H); 13C NMR (CDCl3, 50 MHz):
d 14.09, 21.98, 22.68, 29.42, 29.65,
31.47, 31.92, 33.43, 111.28, 114.58, 114.96, 121.99, 122.24, 124.00,
124.14, 124.61, 126.77, 130.30, 143.49, 154.55, 166.08; GC–MS (DI, m/
z): 483 (Mþ).
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4.6.1. In vitro growth inhibition assay
The cells were maintained in Dulbecco’s Modified Eagle’s
Medium (Sigma–Aldrich Inc., USA) supplemented with 10% fetal
bovine serum (Sigma Chemical Co., USA) in a CO2 incubator. The
cytotoxicity of the compounds was measured by MTT assay [22].
The cells were plated in a 96-well plate at the density of 10,000 cells
per well (HeLa) and 10,000 cells per well (HCT-15). After 24 h, the