1,3,4-Oxadiazole-3(2H)-carboxamide derivatives as potential novel class of monoamine oxidase (MAO) inhibitors: Synthesis, evaluation, and role of urea moiety

Article abstract of DOI:10.1016/j.bmc.2008.07.026

A new series of 1,3,4-oxadiazole-3(2H)-carboxamide derivatives have been synthesized by direct heterocyclization reaction of substituted benzoylisocyanate with various aroylhydrazones as novel monoamine oxidase inhibitors (MAOIs). The target molecules have been identified on the basis of satisfactory analytical and spectra (IR, ¹H NMR, ¹³C NMR, and HR-MS) data. The newly synthesized compounds were evaluated for their MAO inhibitory activity by kynuramine fluorimetric assay method. The preliminary results showed that most of the compounds have moderate inhibitory activities toward MAO at the concentration of 10^-5-10^-3 M. This work may provide a novel class of lead compounds with potential MAO inhibitions for further optimization.

Full text of DOI:10.1016/j.bmc.2008.07.026

Bioorganic & Medicinal Chemistry 16 (2008) 7565–7572
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
1,3,4-Oxadiazole-3(2H)-carboxamide derivatives as potential novel class
of monoamine oxidase (MAO) inhibitors: Synthesis, evaluation, and role
of urea moiety
*
Shaoyong Ke, Zhong Li, Xuhong Qian
Shanghai Key Laboratory of Chemical Biology, Institute of Pesticides and Pharmaceuticals, School of Pharmacy, East China University of Science and Technology,
Shanghai 200237, China
a r t i c l e i n f o
a b s t r a c t
Article history:
A new series of 1,3,4-oxadiazole-3(2H)-carboxamide derivatives have been synthesized by direct hetero-
cyclization reaction of substituted benzoylisocyanate with various aroylhydrazones as novel monoamine
oxidase inhibitors (MAOIs). The target molecules have been identified on the basis of satisfactory analyt-
ical and spectra (IR, 1H NMR, 13C NMR, and HR-MS) data. The newly synthesized compounds were eval-
uated for their MAO inhibitory activity by kynuramine fluorimetric assay method. The preliminary results
showed that most of the compounds have moderate inhibitory activities toward MAO at the concentra-
tion of 10ꢀ5–10ꢀ3 M. This work may provide a novel class of lead compounds with potential MAO inhi-
bitions for further optimization.
Received 22 May 2008
Revised 13 July 2008
Accepted 15 July 2008
Available online 18 July 2008
Keywords:
1,3,4-Oxadiazole-3(2H)-carboxamide
Heterocycle
MAO inhibitor
Urea
Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction
it has been linked to a variety of adverse health conditions includ-
ing alcoholism, smoking, aggressive behavior, certain neurodegen-
Monoamine oxidase (MAO) is a ubiquitous membrane-bound,
flavin-containing enzyme, which is particularly abundant in the li-
ver and brain.1 MAO is located intracellularly in the mitochondrial
outer membranes of neuronal, glial, and other cells and catalyzes
the oxidative deamination of monoamine neurotransmitters both
from endogenous and exogenous sources to the corresponding
aldehydes with consumption of oxygen and production of hydro-
gen peroxide, thereby affecting the concentrations of neurotrans-
mitter amines as well as many xenobiotic ones.2–4
In mammals, two distinct isoforms of MAO are present in most
tissues, namely, MAO-A and MAO-B, which were defined in 1968,
based on their differential substrate and inhibitor specificity,5–7 tis-
sue and cell distribution,8 and gene expression9,10 characteristics.
MAO-A preferentially deaminates aromatic monoamines such as
the neurotransmitters serotonin, noradrenaline, and adrenaline,
while MAO-B mainly oxidizes b-phenylethylamines and benzylam-
ines. Both isoforms act on dopamine and tryptamine.1
erative disorders, and even to the growth inhibition and
progression of cancer.13,14 Due to the important function played
by MAO in the metabolism of neurotransmitters, MAO inhibitors
represent a useful tool for the treatment of several neurological
diseases.15,16 Recognition of the importance of monoamine oxi-
dases as targets for drug intervention for treatment of a variety
of conditions has produced an enormous interest in development
of inhibitors of these enzymes during these years.
The first generation of MAO inhibitors, originally synthesized in
the 1950s, were used as antidepressants due to their inhibitory ef-
fect on the metabolism of monoamine neurotransmitters. The next
discovery was made by Johnston,17 who showed that the MAO en-
zyme exists in two forms: A and B. This led to the synthesis of a
new generation of MAO inhibitors exhibiting greater selectivity to-
ward the individual isoforms. Following an initial experience with
non-selective, irreversible MAO inhibitors in the treatment of
depression associated with severe side effects,18 it can be stated
that today the selective MAO-A inhibitors are currently used for
treating neurological disorders such as anxiety and depression,
while selective inhibitors of the B isoform are administered alone
or together with L-DOPA for the treatment of Parkinson’s syn-
drome and Alzheimer’s disease.19–23
In the central nervous system and peripheral tissues, MAO cat-
alyzes the metabolic transformation of amine neurotransmitters;
whereas in the liver, MAO functions as a detoxicating enzyme
against foreign amine compounds that enter the body.11,12 It is well
recognized that MAO plays an important role in metabolism in that
During the past decades, extensive studies focused on the
search for MAO inhibitors have led to the discovery of various
structural classes of potent MAO inhibitors, including hydrazide,
* Corresponding author. Fax: +86 21 64252603.
E-mail address: xhqian@ecust.edu.cn (X. Qian).
0968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.07.026

7566
S. Ke et al. / Bioorg. Med. Chem. 16 (2008) 7565–7572
amide, thiazole, imidazole, oxazolidinone, oxadiazolone, diacylu-
rea derivatives, etc.24–37 In spite of considerable progress in under-
standing the interactions of the two enzyme forms with their
corresponding substrates, no general rules are yet available for
the rational design of potent and selective inhibitors of MAO. There
are many different structures of MAO inhibitors partly due to the
fact that the active sites of the MAO are ambiguous, which limits
the design of potent selective MAO inhibitors. Figure 1 shows the
chemical structures of some representative MAO inhibitors used
in research or clinical practice.24–37
The aim of the present study was the identification of novel po-
tent MAO inhibitors that could serve as potential lead molecules
for drug discovery. Among above depicted representative struc-
tures of MAO inhibitors, many heterocycle nitrogen-containing
derivatives, including thiadiazole, oxadiazole, thiazole, etc., behave
as potential MAO inhibitors, and a common structural feature of
substrates and inhibitors is an amino or imino group that is as-
sumed to play an essential role in orientation and complex forma-
tion at the active site of the enzyme,38 however, little attention has
been paid to the synthesis of 1,3,4-oxadiazoline derivatives bearing
urea moiety. Considering that aroylurea derivatives as dipeptide
mimics tend to contain multi-structure in a molecule, the biologi-
cal activity of these compounds may be improved by the promo-
tion of these combinations with the cell’s microstructure and the
accumulation of various biological activities resulting from the
incorporation of different heterocyclic and non-heterocyclic
changes on biological activity of the compounds. Furthermore,
the introduction of urea moiety, known to be an important partial
structure of many bioactive compounds,39–48 may be stabilized
within the active site of the enzyme through several hydrogen
bonds.49
and discover a new lead compound by combining the different
possible active units in an oxadiazoline model as shown in Figure
2, in order to find novel class of lead compounds with potential
MAO inhibitions. We utilized 1,3,4-oxadiazoline scaffold as key
prototype structural unit and planed for the attachment of an urea
functionality to the heterocycle, and speculated that we might be
able to improve on the properties of compounds by extending
the active system. Thus to test this hypothesis, we prepared com-
pounds
(Scheme 1) and evaluated their MAO inhibitory properties.
The synthetic route is shown in Scheme 1, and the structures of
all newly synthesized compounds were confirmed by 1H NMR, 13
1,3,4-oxadiazole-3(2H)-carboxamide
derivatives
3
C
NMR, IR, EIMS, and high-resolution mass spectra (HR-MS) data.
Furthermore, for a few comparative activity measurements of com-
pounds without urea moiety, some 1,3,4-oxadiazoline derivatives
4 as shown in Scheme 1 were also synthesized. The newly synthe-
sized compounds were evaluated for their MAO inhibitory activity
by kynuramine fluorimetric assay method. The preliminary results
showed that all the urea-containing compounds have moderate
inhibitory activities toward MAO at the concentration of 10ꢀ4
–
10ꢀ3 M, however, the activities of compounds without aroylurea
moiety were almost lost. The present work demonstrated that
assembling the biological active unit of oxadiazoles and ureas
might be able to result in a novel class of lead compounds with po-
tential MAO inhibitions for further optimization of MAO inhibitors
with good activity.
2. Results and discussion
2.1. Synthesis of substituted 1,3,4-oxadiazole derivatives
As part of our medicinal chemistry and agrochemistry program
aimed at the search for novel heterocycle oxa(thia)diazole-based
bioactive molecules,50–55 we report herein our effort to design
Although various methods have been reported for the synthesis
of oxadiazoline derivatives,56–60 these methods require high reac-
tion temperature, long reaction time, and expensive reagent. Reddy
S
O
O
O
H
N
F
N
N
N
NH2
N
H
N
H
N
H
N
O
Cl
Iproniazide (A/B)
Pargiline (A/B)
Moclobemide (A)
Ro 41-1049 (A)
O
O
N
O
R2
N
Cl
N
N
O
S
N
O
O
N
N
R1
n
OH
N
O
N
O
R3
CSC
Toloxatone (A)
LU 53439 (B)
Ar1
O
O
(CH2)n-Z
X
N
X
S
N
N
H
N
O
N
H
Y
R
N
NH
R
O
O
O
O
R
Ar
R
Figure 1. Some representative structures of MAO inhibitors used in research or clinical practice.
Hydrophobic
chain
R4
N
N
Incorporate with
Acylurea
moiety
R3
R2
O
benzoylurea scaffold
O
N
N
H
N
R
R1
1,3,4-Oxadiazolines
O
R'
1,3,4-Oxadiazole
heterocycle
O
Figure 2. Design strategy of 1,3,4-oxadiazoline derivatives containing urea moiety.

S. Ke et al. / Bioorg. Med. Chem. 16 (2008) 7565–7572
7567
O
(2)
Ar
H
N
R
N
N
O
N
3
4
O
O
(1)
Ar
N
H
Ar COOH
1a-f
1, 2
2a-f
O
(3)
Ar
N
N
a-f
a. 4-Cl-C6H4 b. 3,4,5-(OMe)3-C6H2 c. 2,4-Cl2-C6H3
O
Ar
d. 3,4-(OCH2O)-C6H3 e. 3,5-Me2-C6H3
f. 2-Furan
Scheme 1. Reagents and conditions: (1) a—EtOH, concd H2SO4; b—NH2NH2ꢁH2O, reflux for 8 h; c—nonan-5-one, EtOH, reflux for 6–8 h; (2) aroylisocyanate, toluene, reflux for
0.5–1.5 h; (3) Ac2O, reflux for 1–2 h.
et al.58 have synthesized some novel 1,3,4-oxadiazol-2-yl-4(3H)-
quinazolinones by polyphosphoric acid-catalyzed cyclization
method (reflux at 120–130 °C for 3–4 h and the yields are 51–
67%). Hall et al.59 reported that the hydrazides were converted to
1,3,4-oxadiazoles derivatives under drastic reaction conditions
(in neat phosphorus oxychloride under microwave conditions at
100 °C). Ali et al.60 obtained some novel oxadiazolines containing
sugar moiety under reflux temperature with acetic anhydride,
and the yields are 68–71%. However, in the present-described
experimental conditions the heterocyclization reaction reached
completion with a relative simple operation giving, high yield
(64–80%) at shorter time (0.5–1.5 h) compared to the reported
cyclization methods for 1,3,4-oxadiazole ring. The synthetic route
of the designed 1,3,4-oxadiazoline derivatives 2,2-dibutyl-5-
(substituted-phenyl)-N-(substituted-benzoyl)-1,3,4-oxadiazole-3
(2H)-carboxamides 3a–k is outlined in Scheme 1.
The various synthesized benzoates were treated with hydrazine
hydrate in EtOH to afford the hydrazides. The following condensa-
tion reaction between hydrazides and nonan-5-one led to the
important substrates, substituted benzoylhydrazone 2. Another
key intermediate, substituted benzoylisocyanates, was easily ob-
tained by three steps under the conditions of experimental. The
subsequent reaction of substituted benzoylhydrazone 2 with vari-
ous substituted benzoylisocyanates were heating at about 75–
80 °C in toluene for about 0.5–1.5 h afforded the target compounds
characterized as 2,2-dibutyl-5-(substituted-phenyl)-N-(substi-
tuted-benzoyl)-1,3,4-oxadiazole-3(2H)-carboxamides 3a–k in
excellent yields of 64–80%. The heterocyclization reaction is most
probably initiated by a nucleophilic attack of the imino hydrazone
nitrogen to the carbonyl carbon of the isocyanate leading to an
intermediate, with subsequent proton transfer and cyclization of
the dipolar intermediate leading to the formation of target com-
pounds. The possible mechanism of cyclization reaction to afford
target compounds is shown in Scheme 2. In a conventional meth-
reaction in the presence of acetic anhydride to afford the 3-acet-
yl-5-aryl-2,3(2H)-1,3,4-oxadiazoles 4a–c.
In the above-described experimental conditions the heterocyc-
lization reaction reached completion giving a very high yield at
shorter time compared to the reported cyclization methods for
1,3,4-oxadiazole ring derivatives.
2.2. Spectroscopy
The structures of all the newly synthesized compounds were
characterized as 2,2-dibutyl-5-(substituted-phenyl)-N-(substi-
tuted-benzoyl)-1,3,4-oxadiazole-3(2H)-carboxamide
3 and 2,2-
dibutyl-3-acetyl-5-aryl-2,3(2H)-1,3,4-oxadiazole 4 on the basis of
satisfactory analytical and spectral data including 1H NMR, 13C
NMR, IR, EIMS, and HR-EIMS.
The IR spectra of the 1,3,4-oxadiazole-3(2H)-carboxamide
derivatives 3a–k show characteristic absorption bands at 3380–
3230, 1750–1660, and 1635–1610 cmꢀ1, which have been assigned
to N–H bond, amido carbonyl, and imino groups, respectively. The
1H NMR spectra of 1,3,4-oxadiazole-3(2H)-carboxamides 3a–k
show signals in the range 10.50–11.25 ppm, assigned to the NH
group proton, 6.80–8.50 ppm for the protons of the aromatic ring,
and at 0.95–3.20 ppm for the aliphatic chain protons. Due to the
instability of the oxadiazolines-benzoylurea derivatives, the
molecular ion peak in their mass spectra was of low intensity.
The appearance of peaks for [M–ArCONHCO]+ shows that the
amide C–N bond undergoes fission readily.
2.3. MAO inhibition activity
The in vitro inhibition activities against MAO of all synthesized
compounds were investigated by kynuramine fluorimetric assay
method.61,62 Enzymatic assays revealed that all test compounds
3a–k were weak to moderate MAO inhibitors at low micromolar
concentrations. On the contrary, compounds 4a–c almost lost
od, aroylhydrazone
2
undergoes convenient heterocyclization
O
C
N
O
R2
O
O
O
HN
O
O
N
N
O
N
O
N
H
N
R1
R1
O
N
N
O
HN
O
R2
R1
N
N
R1
H
R2
R2
Scheme 2. The possible mechanism of cyclcondensation reaction with substituted benzoylisocyanate to afford 1,3,4-oxadiazolines.

7568
S. Ke et al. / Bioorg. Med. Chem. 16 (2008) 7565–7572
Table 2
inhibitory activities at the same concentration level, except 4c with
weak activity. The inhibition activity results of compounds 3a–k
and 4a–c against MAO are shown in Tables 1 and 2, respectively.
From Table 1, which shows the structures and the MAO inhibi-
tion data, it can be seen that most of the 1,3,4-oxadiazole-3(2H)-
carboxamide derivatives 3a–k show obvious activity against
MAO with values in the range of 6.03–41.02% at the low concentra-
In vitro inhibition activity of 1,3,4-oxadiazoline derivatives 4a–c on rat brain
mitochondria by kynuramine fluorimetric assay
O
N
N
tion of 100
100 M and almost lost activity at the lower concentration of
M. Compound 3d was the most potent of the series, with values
which reached 41.02% and 8.37% at 100 M and 10 M, respec-
lM. Compounds 3a and 3b have weaker activities at
R
O
l
8
l
Entry
Compound
Substituents (R)
Inhibitions against MAO (%)
l
l
tively. Moreover, compounds 1,3,4-oxadiazole-3(2H)-carboxamide
3a–k and 3-acetyl-5-aryl-2,3(2H)-1,3,4-oxadiazoles 4a–c show a
striking contrast, compounds 4a–c without urea moiety almost lost
inhibitory activities at the same concentration level, except 4c with
weak activity (Table 2), which indicates that the acylurea moiety
may be stabilized within the active site of the enzyme through
hydrogen bonds. Additionally, we can find from Table 1 that the
inhibition activities of the target compounds 3a–k were influenced
by the substituents on the phenyl ring gave the moderate activity.
When the substituent R is 2,6-difluoro substituent, the aromatic
ring (Ar) bearing piperonyl moiety (3d) is more favorable for inhi-
bition activities. Furthermore, when Ar has the same substituents
such as compounds 3d and 3h, 3e and 3g, the compounds (3d
and 3e) with strong electron-withdrawing group (2,6-difluor) in-
crease the potency. Among all the active compounds, the higher
inhibition activity in this series was obtained with the piperonyl
derivatives 3d and 3h, which is similar to the conclusion given in
the literature.11
0.008 (mM)
0.08 (mM)
0.8 (mM)
1
2
3
4a
4b
4c
3,4,5-(MeO)3
3,4-OCH2O
3,5-Me2
0.00
1.21
2.89
0.00
0.00
17.69
0.00
0.00
42.81
The comparative semi-logarithmic plot of the inhibitory activity
of target compounds 3a–k and 4a–c against rat brain mitochon-
drial MAO is given in Figure 3, which further testified compound
3d was the most potential compound of the series of 1,3,4-oxadi-
azole-3(2H)-carboxamide derivatives. On the basis of these results,
we can state that most of the assayed compounds 3a–k possessed
potential monoamine oxidase inhibitory activity. Futhermore,
1,3,4-oxadiazole-3(2H)-carboxamide derivatives, particularly with
piperonyl derivatives, may be novel lead compounds with MAO
inhibitory activity.
Figure 3. Semi-logarithmic plot of the inhibitory activity of target compounds 3c–k
against rat brain mitochondrial MAO at concentration of 1 ꢂ 10ꢀ5–10ꢀ3 M and of
compounds 3a–b, 4a–c at 0.8 ꢂ 10ꢀ5–10ꢀ3 M, respectively.
Table 1
In vitro inhibition activity of 1,3,4-oxadiazole-3(2H)-carboxamide derivatives 3a–k on rat brain mitochondria by kynuramine fluorimetric assay
O
H
N
R
Ar
N
N
O
O
Entry
Compound
Substituents
Inhibitions against MAO (%)
0.1 (mM)
Ar
R
0.01 (mM)
1 (mM)
1
2
3
4
5
6
7
8
9
3a
3b
3c
3d
3e
3f
3g
3h
3i
p-Cl-C6H4
3,4,5-(MeO)3-C6H2
2,4-Cl2-C6H3
3,4-OCH2O-C6H3
3,5-Me2-C6H3
2-Furanc
3,5-Me2-C6H3
3,4-OCH2O-C6H3
2,4-Cl2-C6H3
2,6-2F
2,6-2F
2,6-2F
2,6-2F
2,6-2F
2,6-2F
o-Cl
o-Cl
o-Cl
2,3,5-F3-4-MeO
2,3,5-F3-4-MeO
0.00a
0.00b
5.43
8.37
5.85
1.84
4.12
6.73
5.86
2.92
NDd
6.03
21.72
25.14
41.02
26.24
23.68
24.59
27.49
24.90
25.85
ND
58.79
69.18
72.48
78.39
69.17
75.07
72.78
75.37
77.96
81.84
ND
10
11
3j
3k
2,4-Cl2-C6H3
3,4,5-(MeO)3-C6H2
a,b
These two compounds were tested at the concentrations of 0.008, 0.08, and 0.8 mM, respectively.
This aromatic ring is furan.
ND, not determined.
c
d

S. Ke et al. / Bioorg. Med. Chem. 16 (2008) 7565–7572
7569
4.2.1. 4-Chloro-N’-(nonan-5-ylidene)benzohydrazide (2a)
3. Conclusion
This compound was obtained as white powder following the
above mentioned, yield 81%, mp 158.4–159.6 °C; 1H NMR
(400 MHz, CDCl3): d = 8.70 (s, 1H, N–H), 7.70 (d, J = 8.8 Hz, 2H,
Ph-H), 7.44 (d, J = 8.4 Hz, 2H, Ph-H), 2.27–2.41 (m, 4H, N@C(CH2)2),
1.32–1.56 (m, 8H, CH2), 0.93–0.99 (m, 6H, CH3); MS: m/z = 294
(M+), 237, 210, 168, 139, 111; EI-HRMS: calcd for C16H23ClN2O
(M+), 294.1499; found, 294.1500.
In summary, we have described the molecular design, synthe-
sis, and MAO inhibitory activities of a series of novel 1,3,4-oxa-
diazole-3(2H)-carboxamide derivatives and developed a practical
and efficient procedure for the synthesis of 1,3,4-oxadiazole-
3(2H)-carboxamide derivatives through the direct heterocycliza-
tion reaction of substituted benzoylhydrazone with various ben-
zoylisocyanates in toluene for 0.5–1.5 h, according to the above-
described experimental conditions, the cyclization reaction
reached completion giving a very high yield at shorter time com-
pared to the reported cyclization methods for 1,3,4-oxadiazole
ring derivatives. The preliminary bioassay results indicated that
some of 1,3,4-oxadiazole-3(2H)-carboxamide derivatives exhib-
ited moderate MAO inhibition activity at the concentration of
10ꢀ4–10ꢀ3 M. The present work demonstrated that assembling
the biological active unit of oxadiazole and urea could result in
new lead compounds with high lipophilicity and potent MAO
inhibition activity. To our best knowledge, this is the first report
about the syntheses and MAO inhibition activity of 1,3,4-oxadi-
azoles derivatives bearing urea moiety. Further structural opti-
mization and structure–inhibition activity relationships about
the designed 1,3,4-oxadiazole-3(2H)-carboxamide derivatives
are well under way.
4.2.2. 3,4,5-Trimethoxy-N0-(nonan-5-ylidene)benzohydrazide
(2b)
This compound was obtained as white powder following the
above mentioned, yield 84%, mp 123.4–124.8 °C; 1H NMR
(400 MHz, CDCl3): d = 8.67 (s, 1H, N–H), 6.99 (s, 2H, Ph-H), 3.91
(s, 6H, OCH3), 3.90 (s, 3H, OCH3), 2.24–2.41 (m, 4H, N@C(CH2)2),
1.38–1.58 (m, 8H, CH2), 0.98 (t, J = 7.2 Hz, 3H, CH3), 0.93 (t,
J = 7.2 Hz, 3H, CH3); MS: m/z = 350 (M+), 293, 209, 195, 167; EI-
HRMS: calcd for C19H30N2O4 (M+), 350.2206; found, 350.2207.
4.2.3. 2,4-Dichloro-N0-(nonan-5-ylidene)benzohydrazide (2c)
This compound was obtained as white powder following the
above mentioned, yield 78%, mp 105.2–106.4 °C; 1H NMR
(400 MHz, CDCl3): d = 9.04 (s, 1H, N–H), 7.46 (d, 4J = 2 Hz, 1H, Ph-
H), 7.37–7.42 (m, 2H, Ph-H), 2.42 (t, J = 7.8 Hz, 2H, N@C–CH2),
2.27 (t, J = 8 Hz, 2H, N@C–CH2), 1.36–1.62 (m, 8H, CH2), 0.93–
0.98 (m, 6H, CH3); MS: m/z = 328 (M+), 271, 244, 229, 202, 173,
145, 109, 85, 73, 55; EI-HRMS: calcd for C16H22Cl2N2O (M+),
328.1109; found, 328.1100.
4. Experimental
4.1. Instrumentation and chemicals
4.2.4. N0-(Nonan-5-ylidene)benzo[d][1,3]dioxole-5-
carbohydrazide (2d)
All melting points (mp) were obtained using a Büchi Melting
Point B540, and are uncorrected. 1H and 13C NMR spectra were
recorded on a Brucker AM-400 (400 MHz) spectrometer with
CDCl3 as the solvent and TMS as the internal standard. Chemical
shifts are reported in d (parts per million) values. Coupling con-
stants nJ are reported in Hz. High-resolution electron impact
mass spectra (HR-EIMS) were recorded under electron impact
(70 eV) condition using a MicroMass GCT CA 055 instrument.
Infrared (IR) spectra were measured on a Nicolet FT-IR-20SX
instrument using a potassium bromide (KBr) disk, scanning from
400 to 4000 cmꢀ1. Analytical thin-layer chromatography (TLC)
was carried out on precoated plates, and spots were visualized
with ultraviolet (UV) light. All chemicals or reagents were pur-
chased from standard commercial suppliers and treated with
standard methods before use. Solvents were dried in a routine
way and redistilled.
This compound was obtained as white solid following the above
mentioned, yield 80%, mp 143.4–144.2 °C; 1H NMR (400 MHz,
CDCl3): d = 8.64 (s, 1H, N–H), 7.28–7.33 (m, 2H, Ph-H), 6.86 (d,
J = 8.4 Hz, 1H, Ph-H), 6.04 (s, 2H, OCH2O), 2.26–2.38 (m, 4H,
N@C–(CH2)2), 1.51–1.59 (m, 4H, CH2), 1.35–1.45 (m, 4H, CH2),
0.98 (t, J = 7.2 Hz, 3H, CH3), 0.94 (t, J = 7.4 Hz, 3H, CH3); MS: m/
z = 304 (M+), 247, 149, 121, 96; EI-HRMS: calcd for C17H24N2O3
(M+), 304.1787; found, 304.1788.
4.2.5. 3,5-Dimethyl-N0-(nonan-5-ylidene)benzohydrazide (2e)
This compound was obtained as white solid following the above
mentioned, yield 75%, mp 87.3–88.1 °C; 1H NMR (400 MHz, CDCl3):
d = 8.73 (s, 1H, N–H), 7.39 (s, 2H, Ph-H), 7.16 (s, 1H, Ph-H), 2.21–
2.41 (m, 10H, N@C–(CH2)2 and Ph-CH3), 1.36–1.58 (m, 8H, CH2),
0.98 (t, J = 7.2 Hz, 3H, CH3), 0.94 (t, J = 7.4 Hz, 3H, CH3); MS: m/
z = 288 (M+), 259, 246, 231, 204, 189, 162, 133, 105, 91, 77; EI-
HRMS: calcd for C18H28N2O (M+), 288.2202; found, 288.2203.
4.2. General synthetic procedure for substituted
aroylhydrazones 2a–f
4.2.6. N0-(Nonan-5-ylidene)furan-2-carbohydrazide (2f)
Substituted aroylhydrazones 2 were prepared via esterifica-
tion, hydrazination and condensation three step reactions. Fol-
lowing are the brief descriptions: (1) Synthesis of
aroylhydrazides. These were prepared by the usual esterification
of appropriate benzoic acid (0.1 mol) with alcohol (200 mL) and
concentrated sulfuric acid (10 drops) followed by condensation
of the resulting ethyl benzoates (0.1 mol) with hydrazine hydrate
(0.5 mol). The product was collected by filtration, washed with
water and was recrystallined from ethanol. (2) Synthesis of aro-
ylhydrazones 2. Equal amounts of appropriate aroylhydrazide
(0.01 mol) and nonan-5-one (1.42 g, 0.01 mol) were refluxed in
absolute alcohol (15 mL) for several hours, which was detected
by TLC. Then the solution was concentrated, and the condensa-
tion product aroylhydrazone separated out on cooling and was
recrystallized from ethanol. Their physico-chemical properties
and the spectra data are as follows:
This compound was obtained as white crystal following the
above mentioned, yield 78%, mp 46.3–48.5 °C; 1H NMR
(400 MHz, CDCl3): d = 9.09 (s, 1H, N–H), 7.48 (s, 1H, Furan-H),
7.29 (s, 1H, Furan-H), 6.55 (t, J = 1.6 Hz, 1H, Furan-H), 2.30–2.42
(m, 4H, N@C(CH2)2), 1.35–1.57 (m, 8H, CH2), 0.99 (t, J = 7 Hz, 3H,
CH3), 0.93 (t, J = 7.2 Hz, 3H, CH3); MS: m/z = 250 (M+), 193, 166,
151, 137, 95, 81, 55; EI-HRMS: calcd for
250.1681; found, 250.1682.
C
14H22N2O2 (M+),
4.3. General synthetic procedure for substituted
benzoylisocyanate
These key intermediates were prepared by the usual method
used for isocyanate. Substituted benzamides (0.01 mol), which
were obtained by the usual chlorination of appropriate benzoic

7570
S. Ke et al. / Bioorg. Med. Chem. 16 (2008) 7565–7572
acid (0.01 mol) with SOCl2 (8–10 mL) and followed by ammonoly-
sis of the resulting acyl chloride (0.01 mol) with concentrated
ammonia liquor (15 mL, 28%), and 15 mL of toluene were placed
in a dried round-bottomed flask containing a magnetic stirrer bar
under nitrogen, and to this 10 mL of oxalyl chloride was added
dropwise for 20 min at ice-bath. After addition, the resulting clear
solution was heated at about 75 °C for 6–8 h, and then the exces-
sive oxalyl chloride was removed under reduced pressure to give
a clear solution of substituted benzoyl isocyanate, which was used
for the next step reaction without further purification.
145.95, 137.90, 134.23, 132.12, 131.22, 127.34, 121.52, 113.85,
111.67, 105.16, 36.25, 24.53, 22.26, 13.90; IR(KBr):
m = 3259 (N–
H), 1703, 1677 (C@O), 1618 (C@N) cmꢀ1; MS: m/z = 511, 328,
299, 286, 271, 244, 201, 172, 141, 113; EI-HRMS: calcd for
C
24H25Cl2F2N3O3 (M+), 511.1241; found, 511.1241.
4.4.4. 5-(Benzo[d][1,3]dioxol-5-yl)-2,2-dibutyl-N-(2,6-
difluorobenzoyl)-1,3,4-oxadiazole-3(2H)-carboxamide (3d)
This compound was obtained as white solid following the above
mentioned, yield 76%, mp 159.5–160.7 °C; 1H NMR (400 MHz,
CDCl3): d = 9.11 (s, 1H, N–H), 7.37–7.45 (m, 2H, Ph-H), 7.27 (s,
1H, Ph-H), 6.97 (t, J = 8.2 Hz, 2H, Ph-H), 6.88 (d, J = 8 Hz, 1H, Ph-
H), 6.06 (s, 2H, OCH2O), 2.28–2.35 (m, 2H, CH2), 1.90–1.98 (m,
2H, CH2), 1.27–1.36 (m, 8H, CH2), 0.88 (t, J = 7 Hz, 6H, CH3); 13C
NMR (100 MHz, CDCl3): d = 160.83, 158.34, 154.74, 150.82,
148.06, 145.75, 131.90, 122.15, 117.68, 114.04, 111.61, 108.52,
106,92, 105.08, 101.83, 36.27, 24.48, 22.28, 13.91; IR(KBr):
4.4. General synthetic procedure for 1,3,4-oxadiazole-3(2H)-
carboxamide derivatives 3a–k
A solution of the appropriately substituted benzoyl isocyanate
(2 mmol) in dry toluene or 1,2-dichloroethane (5 mL) was added
dropwise to a stirred solution of the aroylhydrazone 2 (1.5 mmol)
in dry toluene (20 mL) under reflux for over a period of 20 min.
Then the reaction mixture was stirred at 75–85 °C for 0.5–1.5 h,
which was monitored by TLC until the disappearance of aroylhyd-
razone 2. The mixture was cooled to ambient temperature and
washed with a brine solution. The organic layer was separated,
dried over anhydrous sodium sulfate, and evaporated under re-
duced pressure. The residue was purified by column chromatogra-
phy on silica gel using petroleum ether (60–90 °C)/ethyl acetate (v/
v) as eluent, slowly increasing the polarity to give the target com-
pounds 3a–k. Their physico-chemical properties and the spectra
data are as follows:
m
= 3237 (N–H), 1699, 1680 (C@O), 1624 (C@N) cmꢀ1; MS: m/
z = 487, 304, 262, 247, 220, 205, 149, 121; EI-HRMS: calcd for
25H27F2N3O5 (M+), 487.1919; found, 487.1919.
C
4.4.5. 2,2-Dibutyl-N-(2,6-difluorobenzoyl)-5-(3,5-
dimethylphenyl)-1,3,4-oxadiazole-3(2H)-carboxamide (3e)
This compound was obtained as white solid following the above
mentioned, yield 68%, mp 141.2–142.5 °C; 1H NMR (400 MHz,
CDCl3): d = 9.15 (s, 1H, N–H), 7.46 (s, 2H, Ph-H), 7.37–7.43 (m,
1H, Ph-H), 7.17 (s, 1H, Ph-H), 6.97 (t, J = 8 Hz, 2H, Ph-H), 2.39 (s,
6H, Ph-CH3), 2.39 (s, 6H, Ph-CH3), 2.29–2.36 (m, 2H, CH2), 1.91–
1.98 (m, 2H, CH2), 1.27–1.36 (m, 8H, CH2), 0.88 (t, J = 7 Hz, 6H,
CH3); 13C NMR (100 MHz, CDCl3): d = 160.84, 155.26, 145.82,
138.55, 133.62, 131.92, 131.82, 124.64, 123.73, 111.69, 111.45,
4.4.1. 2,2-Dibutyl-5-(4-chlorophenyl)-N-(2,6-difluorobenzoyl)-
1,3,4-oxadiazole-3(2H)-carboxamide (3a)
This compound was obtained as white solid following the above
mentioned, yield 66%, mp 126.5–128.0 °C; 1H NMR (400 MHz,
CDCl3): d = 9.11 (s, 1H, N–H), 7.77 (d, J = 8.4 Hz, 2H, Ph-H), 7.45
(d, J = 8.4 Hz, 2H, Ph-H), 7.40 (q, 1H, Ph-H), 6.97 (t, J = 8 Hz, 2H,
Ph-H), 2.29–2.40 (m, 2H, CH2), 1.92–1.99 (m, 2H, CH2), 1.30–1.34
(m, 8H, CH2), 0.88 (t, J = 7.2 Hz, 6H, CH3); 13C NMR (100 MHz,
CDCl3): d = 160.80, 158.36, 154.15, 145.82, 138.13, 132.04,
129.14, 128.19, 122.45, 111.75, 111.50, 105.62, 36.29, 24.48,
104.97, 36.33, 24.45, 22.29, 21.17, 13.91; IR(KBr): m = 3237 (N–H),
1699, 1677 (C@O), 1625 (C@N) cmꢀ1; MS: m/z = 471, 288, 246,
231, 204, 189, 141, 133, 105, 79; EI-HRMS: calcd for C26H31F2N3O3
(M+), 471.2333; found, 471.2333.
4.4.6. 2,2-Dibutyl-N-(2,6-difluorobenzoyl)-5-(furan-2-yl)-1,3,4-
oxadiazole-3(2H)-carboxamide (3f)
This compound was obtained as pale yellow solid following the
above mentioned, yield 70%, mp 124.5–126.1 °C; 1H NMR
(400 MHz, CDCl3): d = 9.08 (s, 1H, N–H), 7.61–7.62 (m, 1H, Ar-H),
7.37–7.44 (m, 1H, Ar-H), 6.93–6.99 (m, 3H, Ar-H), 6.57 (q, 1H, Ar-
H), 2.28–2.36 (m, 2H, CH2), 1.91–1.98 (m, 2H, CH2), 1.29–1.38
(m, 8H, CH2), 0.89 (t, J = 6.8 Hz, 6H, CH3); 13C NMR (100 MHz,
CDCl3): d = 160.86, 160.80, 158.35, 158.29, 147.97, 145.88,
139.42, 132.11, 132.01, 114.80, 111.95, 111.74, 111.49, 105.56,
22.26, 13.89; IR(KBr):
m = 3355 (N–H), 1714, 1695 (C@O), 1621
(C@N) cmꢀ1 ESI-HRMS: calcd for C24H26ClF2N3O3 ([M+Na]+),
;
500.1528; found, 500.1517.
4.4.2. 2,2-Dibutyl-N-(2,6-difluorobenzoyl)-5-(3,4,5-
trimethoxyphenyl)-1,3,4-oxadiazole-3(2H)-carboxamide (3b)
This compound was obtained as white solid following the above
mentioned, yield 78%, mp 141.2–142.4 °C; 1H NMR (400 MHz,
CDCl3): d = 9.15 (s, 1H, N–H), 7.38–7.45 (m, 1H, Ph-H), 7.06 (s,
2H, Ph-H), 6.97 (t, J = 8 Hz, 2H, Ph-H), 3.96 (s, 6H, OCH3), 3.92 (s,
3H, OCH3), 2.29–2.37 (m, 2H, CH2), 1.90–1.98 (m, 2H, CH2), 1.27–
1.35 (m, 8H, CH2), 0.89 (t, J = 7 Hz, 6H, CH3); 13C NMR (100 MHz,
CDCl3): d = 161.09, 160.72, 154.89, 153.46, 145.77, 141.40,
131.94, 118.97, 111.66, 111.42, 105.32, 104.18, 60.98, 56.42,
36.27, 24.43, 22.26, 13.89; IR(KBr):
m = 3370 (N–H), 1710, 1688
(C@O), 1621 (C@N) cmꢀ1; MS: m/z = 433, 324, 297, 250, 221, 208,
193, 166, 141, 113, 95; EI-HRMS: calcd for C22H25F2N3O4 (M+),
433.1813; found, 433.1813.
4.4.7. 2,2-Dibutyl-N-(2-chlorobenzoyl)-5-(3,5-dimethylphenyl)-
1,3,4-oxadiazole-3(2H)-carboxamide (3g)
36.29, 24.44, 22.26, 13.90; IR(KBr):
m
= 3370 (N–H), 1710, 1688
This compound was obtained as white solid following the above
mentioned, yield 75%, mp 108.5–110.2 °C; 1H NMR (400 MHz,
CDCl3): d = 9.33 (s, 1H, N–H), 7.62 (d, J = 6.8 Hz, 1H, Ph-H), 7.45
(s, 2H, Ph-H), 7.43 (dd, 3J = 6 Hz, 4J = 8 Hz, 2H, Ph-H), 7.36–7.40
(m, 1H, Ph-H), 7.16 (s, 1H, Ph-H), 2.39 (s, 6H, Ph-CH3), 2.31–2.37
(m, 2H, CH2), 1.91–1.98 (m, 2H, CH2), 1.30–1.36 (m, 8H, CH2),
0.88 (t, J = 7 Hz, 6H, CH3); 13C NMR (100 MHz, CDCl3): d = 155.06,
145.93, 138.52, 135.15, 133.52, 131.50, 130.53, 129.78, 129.57,
126.97, 124.59, 123.85, 104.95, 36.40, 24.54, 22.33, 21.16, 13.97;
(C@O), 1621 (C@N) cmꢀ1; ESI-HRMS: calcd for C27H33F2N3O6
([M+Na]+), 556.2235; found, 556.2263.
4.4.3. 2,2-Dibutyl-5-(2,4-dichlorophenyl)-N-(2,6-
difluorobenzoyl)-1,3,4-oxadiazole-3(2H)-carboxamide (3c)
This compound was obtained as white solid following the above
mentioned, yield 74%, mp 97.2–98.4 °C; 1H NMR (400 MHz, CDCl3):
d = 9.10 (s, 1H, N–H), 7.74 (d, J = 8.4 Hz, 1H, Ph-H), 7.55 (s, 1H, Ph-
H), 7.40–7.45 (m, 1H, Ph-H), 7.35–7.37 (m, 1H, Ph-H), 6.97 (t,
J = 8.2 Hz, 2H, Ph-H), 2.29–2.37 (m, 2H, CH2), 1.93–2.01 (m, 2H,
CH2), 1.27–1.40 (m, 8H, CH2), 0.89 (t, J = 6.8 Hz, 6H, CH3); 13C
NMR (100 MHz, CDCl3): d = 160.92, 160.38, 158.37, 152.34,
IR(KBr):
MS: m/z = 469, 288, 246, 231, 204, 189, 139, 133, 105, 79, 57;
m ;
= 3244 (N–H), 1699, 1680 (C@O), 1630 (C@N) cmꢀ1
EI-HRMS: calcd for
469.2130.
C
26H32ClN3O3 (M+), 469.2132; found,

S. Ke et al. / Bioorg. Med. Chem. 16 (2008) 7565–7572
7571
4.4.8. 5-(Benzo[d][1,3]dioxol-5-yl)-2,2-dibutyl-N-(2-
4.5. General synthetic procedure for 3-acetyl-5-aryl-2,3(2H)-
chlorobenzoyl)-1,3,4-oxadiazole-3(2H)-carboxamide (3h)
This compound was obtained as white solid following the
above mentioned, yield 76%, mp 119.4–121.3 °C; 1H NMR
(400 MHz, CDCl3): d = 9.32 (s, 1H, N–H), 7.64 (d, J = 6.8 Hz, 1H,
Ph-H), 7.42–7.44 (m, 2H, Ph-H), 7.36–7.40 (m, 2H, Ph-H), 7.26
(d, 4J = 1.6 Hz, 1H, Ph-H), 6.88 (d, J = 8 Hz, 1H, Ph-H), 6.05 (s,
2H, OCH2O), 2.32–2.40 (m, 2H, CH2), 1.92–1.99 (m, 2H, CH2),
1.27–1.38 (m, 8H, CH2), 0.88 (t, J = 7 Hz, 6H, CH3); 13C NMR
(100 MHz, CDCl3): d = 154.54, 150.74, 148.04, 145.86, 135.05,
131.59, 130.49, 129.84, 129.75, 127.03, 122.08, 117.79, 108.52,
106.88, 105.08, 101.81, 36.35, 24.57, 22.33, 13.99; IR(KBr):
1,3,4-oxadiazoles derivatives 4a–c
Appropriate aroylhydrazone 2 (1 mmol) was refluxed in acetic
anhydride (5 mL) for 1–2 h, which was detected by TLC. The mix-
ture was cooled, poured into crushed ice, and allowed to stand at
room temperature overnight. The mixture was extracted with
ethyl acetate (3ꢂ 15 mL). The organic layer was separated, dried
over anhydrous sodium sulfate, and evaporated under reduced
pressure. The residue was purified by column chromatography
on silica gel using petroleum ether (60–90 °C)/ethyl acetate (v/v)
as eluent, slowly increasing the polarity to give the target com-
pounds 4a–c. Their physico-chemical properties and the spectra
data are as follows:
m
= 3259 (N–H), 1703, 1665 (C@O), 1628 (C@N) cmꢀ1; MS: m/
z = 485, 304, 262, 247, 220, 205, 149, 138, 121, 111, 91, 65;
EI-HRMS: calcd for
485.1717.
C
25H28ClN3O5 (M+), 485.1717; found,
4.5.1. 2,2-Dibutyl-3-acetyl-5-(3,4,5-trimethoxyphenyl)-1,3,4-
oxadiazoline (4a)
4.4.9. 2,2-Dibutyl-N-(2-chlorobenzoyl)-5-(2,4-dichlorophenyl)-
1,3,4-oxadiazole-3(2H)-carboxamide (3i)
This compound was obtained as white powder following the
above mentioned, yield 86%, mp 128.6–129.3 °C; 1H NMR
(400 MHz, CDCl3): d = 7.06 (s, 2H, Ph-H), 3.94 (s, 6H, OCH3), 3.90
(s, 3H, OCH3), 2.41–2.49 (m, 2H, CH2), 2.35 (s, 3H, COCH3), 1.92–
1.99 (m, 2H, CH2), 1.26–1.33 (m, 8H, CH2), 0.89 (t, J = 6.8 Hz, 6H,
CH3); 13C NMR (100 MHz, CDCl3): d = 154.49, 153.35, 140.91,
120.01, 105.20, 103.97, 60.94, 56.31, 36.14, 24.63, 22.37, 22.26,
This compound was obtained as white solid following the
above mentioned, yield 64%, mp 104.3–105.7 °C; 1H NMR
(400 MHz, CDCl3): d = 9.36 (s, 1H, N–H), 7.74 (d, J = 8.8 Hz,
1H, Ph-H), 7.66 (d, J = 7.2 Hz, 1H, Ph-H), 7.54 (d, J = 6 Hz,
1H, Ph-H), 7.41–7.44 (m, 2H, Ph-H), 7.35–7.40 (m, 2H, Ph-
H), 2.34–2.42 (m, 2H, CH2), 1.96–2.05 (m, 2H, CH2), 1.25–
1.41 (m, 8H, CH2), 0.89 (t, J = 7 Hz, 6H, CH3); 13C NMR
(100 MHz, CDCl3): d = 165.79, 152.10, 146.04, 137.79, 134.76,
134.14, 131.78, 131.30, 131.08, 130.47, 129.99, 129.91,
127.34, 127.08, 121.62, 105.08, 36.33, 24.62, 22.31, 13.97;
13.99; IR(KBr):
(M+), 335, 293, 251, 195, 167, 152, 137, 122; EI-HRMS: calcd for
21H32N2O5 (M+), 392.2311; found, 392.2311.
m
= 1651 (C@O), 1587 (C@N) cmꢀ1; MS: m/z = 392
C
4.5.2. 2,2-Dibutyl-3-acetyl-5-(benzo[d][1,3]dioxol-5-yl)-1,3,4-
oxadiazoline (4b)
IR(KBr):
m = 3362 (N–H), 1710, 1691 (C@O), 1625 (C@N)
cmꢀ1; MS: m/z = 509, 328, 286, 271, 244, 180, 139, 111, 75;
This compound was obtained as yellow liquid following the
above mentioned, yield 74%, 1H NMR (400 MHz, CDCl3): d = 7.38
(dd, 3J = 8.4 Hz, 4J = 1.4 Hz, 1H, Ph-H), 7.29 (d, 4J = 1.4 Hz, 1H, Ph-
H), 6.86 (d, J = 8.4 Hz, 1H, Ph-H), 6.04 (s, 2H, OCH2O), 2.38–2.45
(m, 2H, CH2), 2.32 (s, 3H, COCH3), 1.90–2.02 (m, 2H, CH2), 1.23–
1.33 (m, 8H, CH2), 0.88 (t, J = 6.8 Hz, 6H, CH3); 13C NMR
(100 MHz, CDCl3): d = 166.70, 154.40, 150.27, 147.91, 121.68,
118.69, 108.37, 106.84, 104.94, 101.65, 36.09, 24.65, 22.39, 22.22,
EI-HRMS: calcd for
509.1025.
C
24H26Cl3N3O3 (M+), 509.1040; found,
4.4.10. 2,2-Dibutyl-5-(2,4-dichlorophenyl)-N-(2,3,5-trifluoro-4-
methoxybenzoyl)-1,3,4-oxadiazole-3(2H)-carboxamide (3j)
This compound was obtained as white solid following the above
mentioned, yield 75%, mp 88.1–89.0 °C; 1H NMR (400 MHz, CDCl3):
d = 9.89 (s, 1H, N–H), 7.77 (d, J = 8.4 Hz, 1H, Ph-H), 7.62–7.68 (m,
1H, Ph-H), 7.56 (d, 4J = 2 Hz, 3H, Ar-H), 7.38 (dd, 4J = 2 Hz,
3J = 8.4 Hz, 1H, Ph-H), 4.08 (s, 3H, OCH3), 2.38–2.47 (m, 2H, CH2),
1.99–2.05 (m, 2H, CH2), 1.31–1.41 (m, 8H, CH2), 0.89 (t, J = 7 Hz,
6H, CH3); 13C NMR (100 MHz, CDCl3): d = 171.11, 159.34, 158.34,
151.88, 146.48, 145.89, 137.81, 134.10, 131.37, 130.99, 127.39,
121.45, 112.22, 112.03, 104.89, 60.37, 36.39, 24.62, 22.33, 13.95;
13.99; IR(KBr):
m
= 1658 (C@O), 1595 (C@N) cmꢀ1; MS: m/z = 346
(M+), 289, 247, 149, 141, 114, 85, 57; EI-HRMS: calcd for
C19H26N2O4 (M+), 346.1893; found, 346.1893.
4.5.3. 2,2-Dibutyl-3-acetyl-5-(3,5-dimethylphenyl)-1,3,4-
oxadiazoline (4c)
This compound was obtained as white solid following the above
mentioned, yield 72%, mp 50.2–51.3 °C; 1H NMR (400 MHz, CDCl3):
d = 7.46 (s, 2H, Ph-H), 7.13 (s, 1H, Ph-H), 2.41–2.47 (m, 2H, CH2),
2.38 (s, 6H, Ph-CH3), 2.35 (s, 3H, COCH3), 1.92–1.98 (m, 2H, CH2),
1.29–1.34 (m, 8H, CH2), 0.88 (t, J = 6.8 Hz, 6H, CH3); 13C NMR
(100 MHz, CDCl3): d = 166.82, 154.94, 138.34, 133.02, 124.65,
124.39, 104.78, 36.16, 24.63, 22.40, 22.25, 21.18, 13.99; IR(KBr):
IR(KBr):
MS: m/z = 559, 362, 286, 271, 244, 231, 189, 172, 144, 113;
m ;
= 3377 (N–H), 1743, 1684 (C@O), 1620 (C@N) cmꢀ1
EI-HRMS: calcd for
559.1252.
C
25H26Cl2F3N3O4 (M+), 559.1252; found,
4.4.11. 2,2-Dibutyl-N-(2,3,5-trifluoro-4-methoxybenzoyl)-5-
(3,4,5-trimethoxyphenyl)-1,3,4-oxadiazole-3(2H)-carboxamide
(3k)
m
= 1654 (C@O), 1595 (C@N) cmꢀ1; MS: m/z = 330 (M+), 273, 231,
189, 133, 105, 85, 57; EI-HRMS: calcd for C20H30N2O2 (M+),
330.2307; found, 330.2307.
This compound was obtained as white solid following the above
mentioned, yield 63%, mp 113.6–116.2 °C; 1H NMR (400 MHz,
CDCl3): d = 9.83 (s, 1H, N–H), 7.58 (q, 3J = 16 Hz, 4J = 8.8 Hz, 1H,
Ph-H), 7.08 (s, 2H, Ph-H), 4.06 (s, 3H, OCH3), 3.96 (s, 6H, OCH3),
3.92 (s, 3H, OCH3), 2.33–2.46 (m, 2H, CH2), 1.94–2.07 (m, 2H,
CH2), 1.30–1.39 (m, 8H, CH2), 0.89 (t, J = 6.4 Hz, 6H, CH3); 13C
NMR (100 MHz, CDCl3): d = 159.65, 154.72, 153.44, 145.78,
141.37, 111.99, 111.77, 105.60, 104.10, 60.99, 56.21, 36.48, 29.68,
4.6. MAO inhibitory activity determination
The prepared compounds were submitted to the Chinese Na-
tional Center for Drug Screening for in vitro MAO inhibitory activ-
ity assays. MAO inhibitory activity of rat brain mitochondrial was
determined by a fluorimetric procedure using kynuramine as a
substrate according to the reported method61,62 with some modifi-
cations. Fluorescence was detected using a Perkin-Elmer Victor II
spectrofluorimeter. Test compounds were dissolved in DMSO/
24.54, 22.33, 13.96; IR(KBr):
m = 3332 (N–H), 1747, 1691 (C@O),
1632 (C@N) cmꢀ1; MS: m/z = 581, 393, 350, 293, 251, 231, 195,
189, 167, 152; EI-HRMS: calcd for C28H34F3N3O7 (M+), 581.2349;
found, 581.2350.
H2O and different inhibitor concentrations (10ꢀ5–10ꢀ3 mol Lꢀ1
)

7572
S. Ke et al. / Bioorg. Med. Chem. 16 (2008) 7565–7572
28. Löscher, W.; Lehmann, H.; Teschendorf, H. J.; Traut, M.; Gross, G. J. Pharmacol.
Exp. Ther. 1999, 288, 984.
29. Chen, J. F.; Steyn, S.; Staal, R.; Petzer, J. P.; Xu, K.; Van der Schyf, C. J.; Castagnoli,
K.; Sonsalla, P. K.; Castagnoli, N. Jr.; Schwarzschild, M. A. J. Biol. Chem. 2002,
277, 36040.
30. Khan, K. M.; Rahat, S.; Choudhary, M. I.; Atta-ur-Rahman; Ghani, U.; Perveen,
S.; Khatoon, S.; Dar, A.; Malik, A. Helv. Chem. Acta 2002, 85, 559.
31. Gökhan, N.; Yesßilada, A.; Ucar, G.; Erol, K.; Bilgin, A. A. Arch. Pharm. Pharm. Med.
Chem. 2003, 336, 362.
32. Chimenti, F.; Maccioni, E.; Secci, D.; Bolasco, A.; Chimenti, P.; Granese, O. B.;
Turini, P.; Alcaro, S.; Ortuso, F.; Cirilli, R.; Torre, F. L.; Cardia, M. C.; Distinto, S. J.
Med. Chem. 2005, 48, 7113.
were used. A control was run for each test and MAO inhibiting
activity of samples was expressed as % of the control.
Acknowledgments
This work was financially supported by National Key Project for
Basic Research (2003CB114405), National Natural Science Founda-
tion of China. The authors also thank for the partial support from
Shanghai Education Committee and Shanghai Foundation of Sci-
ence and Technology.
33. Dar, A.; Khan, K. M.; Ateeq, H. S.; Khan, S.; Rahat, S.; Perveen, S.; Supuran, C. T. J.
Enzym. Inhib. Med. Chem. 2005, 20, 269.
34. Chimenti, F.; Secci, D.; Bolasco, A.; Chimenti, P.; Granese, A.; Carradori, S.;
Befani, O.; Turini, P.; Alcaro, S.; Ortuso, F. Bioorg. Med. Chem. Lett. 2006, 16,
4135.
Supplementary data
35. Volk, N.; Malan, S. F.; Castagnoli, N. Jr.; Bergh, J. J.; Petzer, J. P. Bioorg. Med.
Chem. 2006, 14, 3512.
36. Chimenti, F.; Maccioni, E.; Secci, D.; Bolasco, A.; Chimenti, P.; Granese, A.;
Befani, O.; Turini, P.; Alcaro, S.; Ortuso, F.; Cardia, M. C.; Distinto, S. J. Med.
Chem. 2007, 50, 707.
37. Van den Berg, D.; Zoellner, K. R.; Ogunrombi, M. O.; Malan, S. F.; Terre’Blanche,
G.; Castagnoli, N. Jr.; Bergh, J. J.; Petzer, J. P. Bioorg. Med. Chem. 2007, 15, 3692.
38. Belleau, B.; Moran, J. J. Med. Pharm. Chem. 1962, 5, 215.
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmc.2008.07.026.
References and Notes
1. Kalgutkar, A. S.; Dalvie, D. K.; Castagnoli, N., Jr.; Taylor, T. J. Chem. Res. Toxicol.
2001, 14, 1139.
2. Haung, R. H.; Faulkner, R. J. Biol. Chem. 1981, 256, 9211.
3. Cohen, G.; Farooqui, R.; Kesler, N. Proc. Natl. Acad. Sci. U.S.A. 1997, 94, 4890–4894.
4. Edmondson, D. E.; Mattevi, A.; Binda, C.; Li, M.; Hubálek, F. Curr. Med. Chem.
2004, 11, 1983.
5. Youdim, M. B. H.; Finberg, J. P. M. Biochem. Pharmacol. 1991, 41, 155.
6. Gottowik, J.; Cesura, A. M.; Malherbe, P.; Lang, G.; Prada, M. D. FEBS Lett. 1993,
317, 152.
7. Geha, R. M.; Rebrin, I.; Chen, K.; Shih, J. C. J. Biol. Chem. 2001, 276, 9877.
8. Westlund, K. N.; Denney, R. M.; Kochersperger, L. M.; Rose, R. M.; Abell, C. W.
Science 1985, 230, 181.
9. Bach, A. W. J.; Lan, N. C.; Johnson, D. L.; Abell, C. W.; Bembenek, M. E.; Kwan, S.
W.; Seeburg, P. H.; Shih, J. C. Proc. Natl. Acad. Sci. U.S.A. 1988, 85, 4934.
10. Grimsby, J.; Chen, K.; Wang, L. J.; Lan, N. C.; Shin, J. C. Proc. Natl. Acad. Sci. U.S.A.
1991, 88, 3637.
11. Raciti, G.; Mazzone, P.; Raudino, A.; Mazzone, G.; Cambria, A. Bioorg. Med.
Chem. 1995, 3, 1485.
12. Shih, J. C.; Chen, K.; Ridd, M. J. Ann. Rev. Neurosci. 1999, 22, 197.
13. Valley, M. P.; Zhou, W.; Hawkins, E. M.; Shultz, J.; Cali, J. J.; Worzella, T.; Bernad,
L.; Good, T.; Good, D.; Riss, T. L.; Klaubert, D. H.; Wood, K. V. Anal. Biochem.
2006, 359, 238.
14. Toninello, A.; Pietrangeli, P.; Marchi, U. D.; Salvi, M.; Mondovi, B. Biochim.
Biophys. Acta 2006, 1765, 1.
39. Kaymakçiog˘lu, B. K.; Rollas, S.; Körceg˘ez, E.; Ariciog˘lu, F. Eur. J. Pharma. Sci.
2005, 26, 97.
40. Takahashi, T.; Sakuraba, A.; Hirohashi, T.; Shibata, T.; Hirose, M.; Haga, Y.;
Nonoshita, K.; Kanno, T.; Ito, J.; Iwaasa, H.; Kanatani, A.; Fukami, T.; Sato, N.
Bioorg. Med. Chem. 2006, 14, 7501.
41. Ban, H.; Muraoka, M.; Ioriya, K.; Ohashi, N. Bioorg. Med. Chem. Lett. 2006, 16,
44.
42. Dinges, J.; Ashworth, K. L.; Akritopolou-Zanze, I.; Arnold, L. D.; Baumeister, S.
A.; Bousquet, P. F.; Cunha, G. A.; Davidsen, S. K.; Djuric, S. W.; Gracias, V. J.;
Michaelides, M. R.; Rafferty, P.; Sowin, T. J.; Stewart, K. D.; Xia, Z.; Zhang, H. Q.
Bioorg. Med. Chem. Lett. 2006, 16, 4266.
43. Galiano, S.; Ceras, J.; Cirauqui, N.; Pérez, S.; Juanenea, L.; Rivera, G.; Aldana, I.;
Monge, A. Bioorg. Med. Chem. 2007, 15, 3896.
44. Fortin, S.; Moreau, E.; Lacroix, J.; Teulade, J.-C.; Patenaude, A.; C-Gaudreault, R.
Bioorg. Med. Chem. Lett. 2007, 17, 2000.
45. Kim, Y. J.; Ryu, J.-H.; Cheon, Y. J.; Lim, H. J.; Jeon, R. Bioorg. Med. Chem. Lett.
2007, 17, 3317.
46. Cao, P.; Huang, X.; Ding, H.; Ge, H.; Li, H.; Ruan, B.; Zhu, H. Chem. Biodivers.
2007, 4, 881.
47. Berglund, M.; Dalence-Guzemán, M. F.; Skogvall, S.; Sterner, O. Bioorg. Med.
Chem. 2008, 16, 2529.
48. Zarghi, A.; Kakhgi, S.; Hadipoor, A.; Daraee, B.; Dadrass, O. G.; Hedayati, M.
Bioorg. Med. Chem. Lett. 2008, 18, 1336.
49. Wouters, J.; Ooms, F.; Jegham, S.; Koenig, J. J.; George, P.; Durant, F. Eur. J. Med.
Chem. 1997, 32, 721.
50. Li, Z.; Zhu, Z. X.; Song, G. H.; Qian, X. H.; Sun, J. J. Chem. Res. 1999, 66.
51. Shi, W.; Qian, X. H.; Zhang, R.; Song, G. H. J. Agric. Food Chem. 2001, 49, 124.
52. Cao, S.; Qian, X. H.; Song, G. H.; Huang, Q. C. J. Fluorine Chem. 2002, 117, 63.
53. Cao, S.; Qian, X. H.; Song, G. H.; Chai, B.; Jiang, Z. S. J. Agric. Food Chem. 2003, 51,
152.
54. Li, Z. G.; Yang, Q.; Qian, X. H. Bioorg. Med. Chem. 2005, 13, 3149.
55. Yang, Q.; Qian, X. H.; Xu, J. Q.; Sun, Y. S.; Li, Y. G. Bioorg. Med. Chem. 2005, 13,
1615.
56. Kaspentakis, G. C.; Tsoleridis, C. A.; Stephanidou-Stephanaton, J. J. Heterocyclic
Chem. 2007, 44, 425.
57. Feng, D. X.; Huang, Y.; Chen, R. Y.; Yu, Y.; Song, H. B. Synthesis 2007, 12,
1779.
58. Komaraiah, A.; Sailu, B.; Reddy, P. S. N. Synth. Commun. 2008, 38, 114.
59. Hall, A.; Brown, S. H.; Chowdhury, A.; Giblin, G. M. P.; Gibson, M.; Healy, M. P.;
Livermore, D. G.; McArthur Wilson, R. J.; Naylor, A.; Anthony Rawlings, A.;
Roman, S.; Ward, E.; Willay, C. Bioorg. Med. Chem. Lett. 2007, 17, 4450.
60. Ali, O. M.; Amer, H. H.; Abdel-Rahman, A. A.-H. Synthesis 2007, 18, 2823.
61. Weissbach, H.; Smith, T. E.; Daly, J. W.; Witkop, B.; Udenfriend, S. J. Biol. Chem.
1960, 235, 1160.
15. Ye, S.; Yooshida, S.; Frohlich, R.; Haufe, G.; Kirk, K. L. Bioorg. Med. Chem. 2005,
13, 2489.
16. Hassan, S. H.; Khattab, S. N.; Bekhit, A. A.; Amer, A. Bioorg. Med. Chem. Lett.
2006, 16, 1753.
17. Johnston, J. P. Biochem. Pharmacol. 1968, 17, 1285.
18. Anderson, M. C.; Hasan, F.; McCrodden, J. M.; Tipton, K. F. Neurochem. Res. 1993,
18, 1145.
19. Pacher, P.; Kohegyi, E.; Kecskemeti, V.; Furst, S. Curr. Med. Chem. 2001, 8, 89.
20. Riederer, P.; Lachenmayer, L.; Laux, G. Curr. Med. Chem. 2004, 11, 2033.
21. James, W. T.; Koller, W. C. Neurology 2004, 63, S2.
22. Riederer, P.; Danielczyk, W.; Grünblatt, E. Neurotoxicology 2004, 25, 271.
23. Santo, R. D.; Costi, R.; Roux, A.; Artico, M.; Befani, O.; Meninno, T.; Agostinelli,
E.; Palmegiani, P.; Turini, P.; Cirilli, R.; Ferretti, R.; Gallinella, B.; Torre, F. L. J.
Med. Chem. 2005, 48, 4220.
24. Prada, M. D.; Kettler, R.; Keller, H. H.; Burkard, W. P.; Muggli-Maniglio, D.;
Haefely, W. E. J. Pharmacol. Exp. Ther. 1989, 248, 400.
25. Prada, M. D.; Kettler, R.; Keller, H. H.; Cesura, A. M.; Richards, J. G.; Saura, M. J.;
Muggli-Maniglio, D.; Wyss, P. C.; Kyburz, E.; Imhof, R. J. Neural Transm., Suppl.
1990, 29, 279.
26. Moureau, F.; Wouters, J.; Vercauteren, D. P.; Collin, S.; Evrard, G.; Durant, F.;
Ducrey, F.; Koenig, J. J.; Jarreau, F. X. Eur. J. Med. Chem. 1992, 27, 939.
27. Mazouz, F.; Gueddari, S.; Burstein, C.; Mansuy, D.; Milcent, R. J. Med. Chem.
1993, 36, 1157.
62. Schmidt, K.; Li, Z.; Schubert, B.; Huang, B.; Stoyanova, S.; Hamburger, M. J.
Ethnopharmacol. 2003, 89, 251.