5-Aryl-imidazolin-2-ones as a scaffold for potent antioxidant and memory-improving activity

Article abstract of DOI:10.1016/j.bmcl.2007.12.064

A series of 5-phenyl-substituted-N-alkyl-imidazolin-2-ones with potent radical-scavenging activity and lipid peroxidation inhibitory activity was synthesized. Many of the compounds showed memory-improving effect in animal models independent of the inhibitory activity on lipid peroxidation.

Full text of DOI:10.1016/j.bmcl.2007.12.064

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 18 (2008) 1478–1483
5-Aryl-imidazolin-2-ones as a scaffold for potent antioxidant
and memory-improving activity
Kazutoshi Watanabe,a,* Yasuhiro Morinaka, Yoshio Hayashi,b Masaki Shinoda,c
Hiroyoshi Nishi,d Nobuko Fukushima,e Toshiaki Watanabe,f Akira Ishibashi,g
Satoshi Yukia and Masahiko Tanakah
aPharmaceuticals Research Division, Mitsubishi Pharma Corporation, 1000, Kamoshida-cho, Aoba-ku, Yokohama 227-0033, Japan
bTsukuba Industrial Liaison and Cooperative Research Center, University of Tsukuba, 1-1-1 Tennodai, Tsukuba,
Ibaraki 305-8577, Japan
cAPI Corporation, 2-3-4, Nihonbashi, Chuo-ku, Tokyo 103-0027, Japan
dSales & Marketing Division, Mitsubishi Pharma Corporation, 2-2-6, Nihonbashi-Honcho, Chuo-ku, Tokyo 103-8405, Japan
eDIA Research Martech Inc., 1000, Kamoshida-cho, Aoba-ku, Yokohama 227-0033, Japan
fFaculty of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan
gPharmacovigilance & Quality Assurance Division, Mitsubishi Pharma Corporation, 2-6-9,
Hiranomachi, Chuo-ku, Osaka 541-0046, Japan
hPharmaceuticals Development Division, Mitsubishi Pharma Corporation, 2-2-6,
Nihonbashi-Honcho, Chuo-ku, Tokyo 103-8405, Japan
Received 27 August 2007; revised 19 December 2007; accepted 22 December 2007
Available online 28 December 2007
Abstract—A series of 5-phenyl-substituted-N-alkyl-imidazolin-2-ones with potent radical-scavenging activity and lipid peroxidation
inhibitory activity was synthesized. Many of the compounds showed memory-improving effect in animal models independent of the
inhibitory activity on lipid peroxidation.
Ó 2008 Elsevier Ltd. All rights reserved.
Free radical species are known to play important roles
in maintaining biological systems such as mitochondria,
signal transductions, and the immune system. On the
other hand chronic overproduction of free radical spe-
cies and the resulting oxidative/nitrative stress is gener-
ally considered to be one of the major contributors to
the pathophysiological processes that lead to irreversible
cell damage.1 Free radical production induces the for-
mation of highly reactive oxygen species which readily
react with many biological substrates, particularly with
the amines and polyunsaturated fatty acids which are
the major constituents of the lipid bilayer of the highly
vulnerable brain cell membrane. Chronic oxidative
stress consequently induces progressive irreversible dam-
age leading to impairment of the normal physiological
function of the central nervous system.2–4 Moreover,
oxidative/nitrative stress seems to play a pivotal role in
a variety of central nervous system diseases including
Alzheimer’s disease,5–12 Parkinson’s disease,9,13–18
amyotrophic lateral sclerosis,19,20 and stroke.18,21–23
Free radical-scavenging compounds are therefore
looked to as the source of new tools for the treatment
of these illnesses, whose prevalence is increasing
worldwide.
For many years, our group has been researching free
radical-scavenging compounds, one of which, edaravone
(1, MCI-186) (Fig. 1), shows potent effects against the
deterioration caused by edema and infarction in a rat
model of cerebral ischemia.24–26 Edaravone has also
proven very effective in the treatment of cerebral ische-
mia in humans27 and has been marketed in Japan since
2001 as RadicutTM.
Keywords: Imidazolin-2-one; Antioxidant; Radical scavenger.
The major chemical feature of edaravone is the presence
of an aromatic phenol-like hydroxyl group, which in
turn is due to the presence of a keto-enol tautomerism
*
Corresponding author. Tel.: +81 45 963 3463; fax: +81 45 963 4556;
e-mail: Watanabe.Kazutoshi@mb.mt-pharma.co.jp
Deceased, September 30,2004.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.12.064

K. Watanabe et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1478–1483
1479
ment with trifluoroacetic acid afforded the desired imi-
dazolin-2-ones 7. Compound 37 (Table 2) was
obtained by reaction of the corresponding aminoketone
with methyl chloroformate followed by reaction with
methylamine and successive cyclization.
O
N
1
N
H3C
keto-enol
tautomerization
OH
X
O
X
OH
2
The arylketones 10 with alkyl, aralkyl, and aryl groups
were prepared by Friedel-Crafts acylation of the corre-
sponding alkyl, aralkyl, and aryl benzene 9. The 20-
and 40-alkoxypropiophenones 12 were obtained by
alkylation of the corresponding hydroxypropiophenon-
es 11. The 30-isopropoxypropiophenone 15 was obtained
by alkylation of the 30-hydroxybenzonitrile 13 followed
by reaction with Grignard reagent (Scheme 2).
X=CH
H
R
R
R
N, etc
enol-3
keto-3
1 and design of phenol-like
Figure 1. Structure of edaravone
compound.
(Fig. 1).28 We predict that this phenol-like function will
show good scavenging and antioxidant properties
accompanied by a lower intrinsic toxicity than com-
pounds containing pure phenol.
Radical-scavenging activity against 2,2-diphenyl-1-pic-
rylhydrazyl (DPPH) was measured by absorbance of
solution (OD) at a wavelength of 516.8 nm. An ethanol
solution of the test compound was added to an ethanol
solution of 100 lM of DPPH (final concentration of test
compound 100 lM), and the optical density was mea-
sured. The radical-scavenging activity of the compounds
was expressed as a percentage of the optical density of a
control using an excess of vitamin C.
With this aim in mind, we reinvestigated several hetero-
aromatic scaffolds in which keto-enol tautomerism gave
a hydroxy function and synthesized 5-aryl-imidazolin-2-
one compounds which showed potent lipid peroxidation
inhibitory activity and improvement of memory impair-
ment combined with low toxicity in an animal model.29
The lipid peroxidation inhibitory activity was measured
by inhibition of the autooxidation of rat brain homoge-
nate in the presence and absence of the test compound
according to a modified version of the thiobarbituric
acid test (TBARS, thiobarbituric acid reactive sub-
stance) of Masugi and Nakamura.30 The compound
was added to brain homogenate prepared from male
Wistar rats and the mixture was incubated at 37 °C for
30 min. The IC50 values were calculated by non-linear
regression.
1,4-Disubstituted-5-aryl-imidazolin-2-ones 7 were pre-
pared from the 2-amino-1-arylketone 6 (Scheme 1).
The reaction of bromine with the aryl ketones 4 gave
the corresponding 2-bromoketones which were con-
verted to the N-protected aminoketones 5 with trifluoro-
acetamide under basic conditions. The hydrochloride
salt of the aminoketones 6 was obtained by removal of
the trifluoroacetyl group by hydrolysis with concen-
trated hydrochloric acid. The aminoketones 6 were trea-
ted with the corresponding isocyanates and
intramolecular cyclization of the resulting urea by treat-
The effect of the test compound on cycloheximide-in-
duced amnesia was investigated in a step-down passive
avoidance task in male mice (ddy, 5–6 weeks old,
O
a)
R2
R2
10
12
9
O
O
b)
d)
RO
HO
11
O
RO
CN
O
13 R=H
14 R= -Pr
15
c)
Scheme 1. Reagents and conditions: (a) Br2, CHCl3 or CH2Cl2 or
AcOH, 0 °C to room temperature; (b) CF3CONH2, K2CO3, acetone,
reflux; (c) conc. HCl, EtOH, reflux; (d) R1NCO, Et3N, acetone, 0 °C;
(e) CF3COOH; (f) ClCOOMe, Et3N, acetone, 0 °C; (g) MeNH2 HCl,
DMF, 170 °C.
i
Scheme 2. Reagents and conditions: (a) CH3CH2COCl, AlCl3,
CH2Cl2, 0 °C; (b) RBr or RI, K2CO3, acetone or 2-butanone, reflux;
(c) i-PrI, K2CO3, 2-butanone, reflux; (d) EtMgBr, THF then aq HCl.

1480
K. Watanabe et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1478–1483
n = 15 in each group).31 In brief, 30 min after cyclohex-
imide (120 mg/kg, s.c.) or saline treatment, mice kept in
a light room were allowed to move freely into a dark
room, in which they were given a foot shock (0.5 mA,
3 s) on a grid floor (acquisition trial). One day later,
the mice were given a test compound (0.1–30 mg/kg)
or vehicle orally 30 min before a retention trial, in which
the latent tendency of each subject to move from the
light room to the dark room was measured.
fixed substituent. Both meta and para positions showed
similar activity (57 and 58), while ortho-substitution
gave 56 with decreased inhibitory activity (Table 2).
Further optimization of the imidazolinone structure
was therefore performed with the para position fixed be-
cause of easier synthetic access to several diverse
substituents.
Alkyl groups with various chain lengths were introduced
directly and with an oxygen linker. It was clear that, in
both the methyl and cyclohexyl series, there were certain
alkyl chain lengths suitable for eliciting higher inhibitory
activity; shorter alkyl groups such as methyl 28 and
methoxy 29, and longer alkyl groups such as n-docecyl
72, n-dodecyloxy 73, n-tetradecyl 48 and 74, n-octadecyl
49 and 75, n-octadecyloxy 50 and 76 had poor activity,
while alkyl chains with 4–10 carbon atoms generally
showed good inhibitory activity.
Several 1,4-disubstituted 5-phenylimidazolines were pre-
pared and their lipid peroxidation inhibitory activity
was measured (Table 1). The unsubstituted phenyl
group (R2 = H) showed no or only low activity regard-
less of the substituent at the 1-position (16, 21, and
23). The introduction of a para substituent, such as a
4-n-butyl group on the phenyl group, gave compound
17, which showed good lipid peroxidation inhibitory
activity (Table 1), while a 4-n-butoxy group afforded a
greater increase in activity than the 4-n-butyl group
(17 vs 19, 24 vs 25). The methyl group in the R1 position
of the imidazolinone structure could be efficiently re-
placed by other more lipophilic groups such as n-butyl,
n-hexyl, isopropyl, phenyl, and benzyl (22, 24, 20, 26,
and 27, respectively). No significant difference in the
inhibitory activity was observed when R3 was methyl
or ethyl (17 and 18, respectively). Some of the com-
pounds were tested for radical-quenching ability and
17, 20, 25, and 27 showed good DPPH scavenging activ-
ity. These results seemed to indicate that lipophilicity
could play a role in increasing inhibitory activity against
lipid peroxidation and that methyl and cyclohexyl
groups were preferable as the R1 substituent for good
free radical-quenching activity and crystallinity.
The activity did not seem to depend on chain isomerism
since no significant difference was observed between lin-
ear and branched alkyl groups or linear and ramified
alkoxy groups in either the methyl or cyclohexyl series
(17 vs 32 and 33, 35 vs 36; 24 vs 59, and 61 vs 62).
Nor did the change from linear to cyclic structure mod-
ify inhibitory activity (37 vs 39 and 63 vs 65). The com-
pound group with the highest activity was composed of
various aryl, aryloxy, and arylalkoxy substituents (51,
52, 53, 78, and 79) together with substituents with a 4-
to 10-carbon alkyl chain moiety.
Furthermore, compounds with a cyclohexyl group as R1
substituent showed only slightly lower activity than
those with a methyl group, which led us to suppose that
steric effects did not play an important role in inhibitory
activity.
To confirm the effects of lipophilicity, we also investi-
gated the change in lipid peroxidation inhibitory activity
relative to the position of the substituents in the phenyl
moiety.
We plotted the IC50 of lipid peroxidation inhibitory
activity against the calculated clogP values. The result
showed a distribution of the highest inhibitory activity
around a clogP value of 3–9 (Fig. 2). Compounds with
clogP value of below 2.5 or above 9 showed lower inhib-
The effect of the position of the substituents on the phe-
nyl group was verified using the isopropoxy group as the
Table 1. SAR of lipid peroxidation inhibitory activity and radical scavenging activity of 1,4-disubstituted-5-aryl-imidazolin-2-one compounds
Compound
R1
R2
R3
Lipid peroxidation inhibitory
activity IC50 (lM)
DPPH scavenging
activity % of control
16
17
18
19
20
21
22
23
24
25
26
27
CH3
CH3
H
CH3
CH3
C2H5
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
>100
10.7
13.9
5.1
NT
98%
NT
NT
78%
NT
NT
NT
NT
88%
NT
59%
n-C4H9
n-C4H9
n-C4H9O
n-C4H9
H
CH3
CH3
i-C3H7
n-C4H9
n-C4Hg
c-C6H11
c-C6H11
c-C6H11
C6H5
10.7
>100
12.0
47.6
27.1
13.9
16.6
12.4
n-C4H9
H
n-C4H9
n-C4H9O
n-C4H9
n-C4H9
C6H5CH2
NT, not tested.

K. Watanabe et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1478–1483
1481
Table 2. Effect of phenyl moiety substituents on lipid peroxidation
inhibitory activity
Compound R1
R2
Lipid peroxidation
inhibitory activity
IC50 (lM)
16
28
29
30
31
17
19
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
23
54
55
56
57
58
24
25
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
CH3
H
4-CH3
>100
85.3
147
21.5
20.4
10.7
5.1
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
c-C6H11
4-CH3O
4-n-C3H7
4-i-C3H7O
4-n-C4H9
4-n-C4H9O
4-i-C4H9
4-t-C4H9
4-n-C5H11
Figure 2. Relationship between clogP and IC50 of lipid peroxidation
inhibitory activity. clogP was calculated by ChemDraw Ultra 7.0.1.
IC50 values larger than 100 lM were rated 100 lM and plotted.
itory activity. These results seem to indicate that the
activity of the compounds was greatly affected by their
lipophilicity, which influences the optimal distribution
of the compounds in the lipid bilayer of the membrane.
7.8
8.4
5.9
4.5
4-n-C5H11
O
4-(CH3)2CH(CH2)2O 6.6
4-n-C6H13
4-n-C6H13
4-c-C6H11
4-n-C7H15
4-n-C7H15
4-n-C8H17
4-n-C8H17
9.0
7.3
Although scavenging of free radical species is thought to
be an effective mechanism for the protection of the cen-
tral nervous system, it needs to be proven in a relevant
animal model that compounds displaying this action
are able to cross the blood–brain barrier and show some
efficacy in improving the function of an injured central
nervous system.
O
10.0
8.4
O
O
5.7
19.4
4.0
4-n-C10H21
4-n-C10H21
4-n-C12H25
4-n-C12H25
4-n-C14H29
4-n-C18H37
4-n-C18H37
4-Ph
7.9
6.9
O
O
13.0
9.7
We tested the compounds for pharmacological efficacy
in memory improvement by oral administration in a
passive avoidance model in rats. Compounds with li-
pid peroxidation inhibitory activity (IC50) of up to
11 lM were selected for the study and the results
listed in Table 3. Many of the substituted imidazoli-
none compounds had memory-improving activity,
some with effect starting from 0.1 mg/kg. These find-
ings showed that there was no obvious correlation
with lipid peroxidation inhibitory activity or with lipo-
philicity. The lack of correlation was not surprising as
other physico-chemical properties of the compounds
such as solubility, pharmacokinetics, and distribution
in the brain need to be known before the results can
be explained.
43.8
>100
>100
6.3
O
4-PhO
4-PhCH2O
H
8.1
6.7
47.6
20.4
13.9
21.5
10.4
13.0
27.1
13.9
10.9
10.6
9.1
c-C6H11 4-CH3O
c-C6H11 4-n-C3H7
c-C6H11 2-i-C3H7O
c-C6H11 3-i-C3H7O
c-C6H11 4-i-C3H7O
c-C6H11 4-n-C4H9
c-C6H11 4-n-C4H9O
c-C6H11 4-i-C4H9
c-C6H11 4-n-C5H11
Of the compounds in Table 3, used in acute toxicity
studies via oral administration in rats, 17, 39, and 51 dis-
played a lethal dose (LD50) larger than 1000 mg/kg,
which indicated a much lower intrinsic toxicity of the
phenol-like imidazolinone structure than in the classic
phenol containing compounds (LD50 530 mg/kg).32
c-C6H11 4-n-C5H11
O
c-C6H11 4-(CH3)2CH(CH2)2O 7.2
c-C6H11 4-n-C6H13
c-C6H11 4-n-C6H13
c-C6H11 4-c-C6H11
c-C6H11 4-n-C7H15
c-C6H11 4-n-C7H15
c-C6H11 4-n-C8H17
c-C6H11 4-n-C8H17
13.7
8.5
O
28.9
8.7
O
O
11.6
10.3
8.5
In conclusion, we synthesized free radical scavengers
based on the hypothesis that compounds with an aro-
matic hydroxy group generated by keto-enol tauto-
merization would show radical-scavenging and
antioxidant activities similar to those of phenols but
less acute toxicity. Our study resulted in the synthesis
of a series of novel 5-aryl-imidazolin-2-ones with
good free radical-scavenging activities and potent li-
pid peroxidation inhibitory activity, many of which
combined memory-improving activity with low acute
toxicity.
c-C6H11 4-n-C10H21
c-C6H11 4-n-C10H21
c-C6H11 4-n-C12H25
c-C6H11 4-n-C12H25
c-C6H11 4-n-C14H29
c-C6H11 4-n-C18H37
c-C6H11 4-n-C18H37
c-C6H11 4-Ph
27.0
10.5
>100
52.4
>100
>100
>100
10.9
7.9
O
O
O
c-C6H11 4-PhO
c-C6H11 4-PhCH2O
8.3

1482
K. Watanabe et al. / Bioorg. Med. Chem. Lett. 18 (2008) 1478–1483
Table 3. Memory-improving activity
Compound
R1
R2
Lipid peroxidation inhibitory
activity IC50 (lM)
Memory-improving activity
MED (mg/kg, po)
17
19
32
35
36
37
38
39
40
41
43
44
45
47
51
52
53
61
62
64
69
71
77
78
79
CH3
CH3
n-C4H9
n-C4H9O
i-C4H9
10.7
5.1
7.8
4.5
6.6
9.0
7.3
10.0
8.4
5.7
4.0
7.9
6.9
9.7
6.3
8.1
6.7
91
0.1
0.3
CH3
CH3
30
a
n-C5H11O
(CH3)2CH(CH2)2O
n-C6H13
a
CH3
CH3
0.3
a
CH3
CH3
n-C6H13
c-C6H11
n-C7H15
n-C7H15
n-C8H17
O
0.1
30
CH3
CH3
a
O
O
CH3
CH3
30
n-C10H21
n-C10H21
n-C12H25
Ph
0.3
0.1
0.1
CH3
CH3
O
O
CH3
CH3
0.3
a
PhO
PhCH2O
a
a
a
a
CH3
c-C6H11
c-C6H11
c-C6H11
c-C6H11
c-C6H11
c-C6H11
c-C6H11
c-C6H11
n-C5H11
(CH3)2CH(CH2)2O
O
7.2
8.5
8.5
10.5
10.9
7.9
8.3
n-C6H13
n-C8H17
O
O
30
n-C10H21
O
0.1
0.1
0.1
0.1
Ph
PhO
PhCH2O
MED, minimal effective dose.
a Not active at 0.1 mg/kg, po.
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Acknowledgments
The authors thank Dr. Dino Nisato and Dr. Hiroshi
Tanaka for critical reading of the manuscript.
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