Studies on novel 7-acyl-5-chloro-2-oxo-3H-benzoxazole derivatives as potential analgesic and anti-inflammatory agents

Article abstract of DOI:10.1002/ardp.200300748

In this study, a series of (7-acyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)alkanoic acid derivatives were synthesized and evaluated for their analgesic and anti-inflammatory activities by using the p-benzoquinone-induced writhing test and the carrageenan hind paw edema model, respectively. Acetic acid-induced peritoneal capillary permeability test and serotonin-induced hind paw edema test were also employed for the most active compounds. The test results indicated that (7-acyl-2-oxo-3H-benzoxazol-3-yl)alkanoic acids (Compounds 6 a-c, 8 a-c, 10 a-c) were equally or more potent analgesic and anti-inflammatory agents than aspirin and indomethacin respectively. The compounds 8 a and 8 c, but not 8 b which have the longest carbon chain on alkanoic acid moiety did not induce gastric lesion in mice.

Full text of DOI:10.1002/ardp.200300748

310 Ünlü et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Serdar Ünlüa,
Studies on Novel
Sultan Nacak Baytasa,
Esra Kupelib,
7-Acyl-5-chloro-2-oxo-3H-benzoxazole Derivatives
as Potential Analgesic and Anti-Inflammatory
Agents
Erdem Yesiladab
a
Department of
Pharmaceutical Chemistry
and
Department of
Pharmacognosy, Division of
Pharmaceutical Sciences,
Faculty of Pharmacy, Gazi
University, Ankara, Turkey
In this study, a series of (7-acyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)alkanoic acid
derivatives were synthesized and evaluated for their analgesic and anti-inflammato-
ry activities by using the p-benzoquinone-induced writhing test and the carrageenan
hind paw edema model, respectively. Acetic acid-induced peritoneal capillary per-
meability test and serotonin-induced hind paw edema test were also employed for
the most active compounds. The test results indicated that (7-acyl-2-oxo-3H-ben-
zoxazol-3-yl)alkanoic acids (Compounds 6 a–c,8 a–c,10 a–c) were equally or more
potent analgesic and anti-inflammatory agents than aspirin and indomethacin re-
spectively. The compounds 8 a and 8 c, but not 8 b which have the longest carbon
chain on alkanoic acid moiety did not induce gastric lesion in mice.
b
Keywords: 7-Acyl-2-oxo-3H-benzoxazoles; Alkanoic acids; Propionitriles; QSAR;
Analgesic and Anti-inflammatory Activity
Received: November 20, 2003; Accepted January 23, 2003 [FP748]
DOI 101002/ardp.200300748 *
As already demonstrated in 1973, 6-acyl-2-oxo-3H-
benzoxazole derivatives (II, III) prepared by Bonte et al.
[9] showed higher analgesic activity than that of aspirin.
Introduction
Although there are many nonsteroidal anti-inflammatory
drugs (NSAIDs) on the market, a considerable amount
of research focusses on these drugs due to the serious
side effects of NSAIDs, i.e. gastrointestinal irritation and
kidney damage.Therefore, investigations on new anti-in-
flammatory agents are still a challenge. Recently, in ef-
forts to synthesize anti-inflammatory drugs without or
with minimal side effect, 2-oxo-3H-benzoxazoles have
emerged as a promising group of substances.
Renard et al. [10] showed that the analgesic and anti-in-
flammatory activity of (6-acyl-2-oxo-3H-benzoxazol-3-
yl)alkanoic acid was higher than those of aspirin and ke-
toprofen. Following this study, a numbers of (6-acyl-2-
oxo-3H-benzoxazol-3-yl)alkanoic acid and their ester
and amide derivatives were synthesized and their high
analgesic and anti-inflammatory activities were demon-
strated [11–16].
Since the pioneering discovery of the hypnotic proper-
ties of 2-oxo-3H-benzoxazole over the last twenty-five
years, the 2-oxo-3H-benzoxazole ring has become an
important building block in medicinal chemistry and led
to the discovery of a number of derivatives endowed with
antispasmodic, antitubercular, antibacterial, antimicro-
bial, antifungal, and normolipemic effects [1-7]. Anti-in-
flammatory, analgesic, and antipyretic activity have been
described in 2-oxo-3H-benzoxazole and its 6-acyl deriv-
atives.Studies on analgesic activity of 2-oxo-3H-benzox-
azole was first reported by Close et al. [8] showing that
substitution of alkyl, allyl, and acetyl groups at position 3
and alkyl and halogen groups at position 6 of 2-oxo-3H-
benzoxazole (I) derivatives possess equal or higher an-
algesic activity than aspirin.
Although many reports on 2-oxo-3H-benzoxazoles in-
volved the synthesis and biological activity of 3-, 5-, and
6-acyl-2-oxo-3H-benzoxazole derivatives, there is no
published report on the synthesis and biological activity
of 7-acyl-5-chloro-2-oxo-3H-benzoxazoles and the other
7-acyl derivatives of 2-oxo-3H-benzoxazole in the litera-
ture.Therefore, we aimed to synthesize 7-benzoyl deriv-
atives of 5-chloro-2-oxo-3H-benzoxazole and their alka-
noic acid derivatives which possess a better structural
analogy with well known anti-inflammatory drugs, such
as indomethacin, than 6-acyl-2-oxo-3H-benzoxazole
derivatives.
Results and discussion
Chemistry
Correspondence: Serdar Ünlü, Department of Pharmaceu-
tical Chemistry, Faculty of Pharmacy, Gazi University,
06330 Etiler, Ankara, Turkey. Phone: +90 312 212-6645, Fax:
+90 312 223-5018, e-mail: sunlu@gazi.edu.tr.
The acylation of the 2-oxo-3H-benzoxazole ring was in-
vestigated by various aromatic electrophilic substitution
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
0365-6233/06–07/0310 $ 17.50+.50/0
* In der gedruckten Version war der DOI falsch. Richtig muss es heißen: DOI 10.1002/ardp.200300748

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Novel analgesic and anti-inflammatory benzoxazole derivatives 311
Scheme 1. Synthetic route leading to the compounds 10 a–c and 11 a–c.
conditions. Acyl derivatives of 2-oxo-3H-benzoxazoles
were obtained in good yields with aliphatic, aromatic,
and arylaliphatic carboxylic acids in the presence of
polyphosphoric acid (PPA) [9]. Additionally, C-acylation
of 2-oxo-3H-benzoxazole under various Friedel-Crafts
reaction conditions using carboxylic acids in PPA or acid
halides or anhydrides in the presence of AlCl3-DMF or
Figure 1. Derivative I.
AlCl3-CS2 was published in the literature [17].The regio-
selectivity of the C-acylation of 2-oxo-3H-benzoxazole
was also reported;the electrophilic aromatic substitution
always led to 6-acyl derivatives and the structures of the
2-oxo-3H-benzoxazole derivatives were synthesized by
the C-acylation of 2-(N-acetylamino)phenol at position 4
without any isomer and by-product [20] and the subse-
quent cyclization of this acyl derivative to 2-oxo-3H-
1
acyl derivatives were elucidated by H-NMR, IR, X-ray
diffraction, and theoretical calculations [17–19]. 5-Acyl-

312 Ünlü et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
benzoxazole ring [17]. Based on these reports, the im-
possibility of preparing 7-acyl-2-oxo-3H-benzoxazole
derivatives using 2-oxo-3H-benzoxazole as a starting
material prompted us to synthesize 7-benzoyl-2-oxo-
3H-benzoxazole by using 2-oxo-3H-benzoxazole pre-
cursor, such as 2-acetylamino-4-chlorophenol in which
the amino group was protected by acetyl group (Scheme
1). Since the phenolic hydroxy group has a better elec-
tron donating effect than the acetyl amino group and the
position 4 is closed by the chlorine substituent,
2-acetylamino-5-chloro-6-benzoylphenol
derivatives
(compounds 2 a–c) were synthesized in good yields and
Figure 2. Derivatives II and III.
Table 1. Yields, melting points, and crystallization solvents of the synthesized compounds.
Compound
R1
R2
Yield
%
MP.
°C
Crystallization Solvent
4 a
4 b
4 c
5 a
5 b
5 c
6 a
6 b
6 c
7 a
7 b
7 c
8 a
8 b
8 c
9 a
9 b
9 c
10 a
10 b
10 c
11 a
11 b
11 c
H
H
H
H
74.00
64.00
71.00
78.20
57.33
77.30
62.80
61.32
55.30
82.50
78.10
76.38
67.30
48.00
48.00
65.70
72.00
62.80
63.90
66.00
43.60
51.20
51.20
55.80
183-84
169–70
233–34
128–29
119–20
137–39
209–12
190–91
224–26
106–07
104–05
185–86
213–15
142–43
214–19
182–83
134–35
213–14
168–70
167–69
214–15
84–85
Benzene
Toluene
Toluene
2-Cl
4-Cl
H
2-Cl
4-Cl
H
2-Cl
4-Cl
H
2-Cl
4-Cl
H
2-Cl
4-Cl
H
2-Cl
4-Cl
H
2-Cl
4-Cl
H
2-Cl
4-Cl
CH2COOC2H
Ethanol
5
CH2COOC2H5
CH2COOC2H5
CH2COOH
CH2COOH
CH2COOH
(CH2)3COOC2H5
(CH2)3COOC2H5
(CH2)3COOC2H5
(CH2)3COOH
(CH2)3COOH
(CH2)3COOH
(CH2)2CN
Ethanol/Water
Ethanol/Water
Benzene
Toluene
Toluene
Ethanol
1-Propanol
1-Propanol
Acetonitrile
Toluene
Toluene
Ethanol
Ethanol
Ethanol
Benzene
Toluene
Toluene
n-Hexane
n-Hexane
n-Hexane
(CH2)2CN
(CH2)2CN
(CH2)2COOH
(CH2)2COOH
(CH2)2COOH
(CH2)2COOC2H5
(CH2)2COOC2H5
(CH2)2COOC2H5
112–13
132–33

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Novel analgesic and anti-inflammatory benzoxazole derivatives 313
without any isomer by the reaction of 2-acetylamino-4-
chlorophenol (1) with benzoylchloride derivatives in the
presence of AlCl3-DMF prepared by a method published
previously [21].Subsequently, these intermediates were
hydrolyzed to aromatic amine derivatives (compounds
3 a–c) in concentrated HCl or sodium hydroxide solution.
2-oxo-3H-benzoxazoles can be prepared by cyclization
of 2-aminophenol derivatives as a source of carbonyl
group such as urea [22], N,NЈ-carbonyldiimidazole [23],
or ethyl chloroformate [24] in different reaction condi-
tions.
an edema produced by carrageenan is a biphasic event
and it was reported that the inhibitory effects of agents
which act on the first stage of the carrageenan-induced
hind paw inflammation are attributable to the inhibition of
the chemical mediators such as histamine, serotonin,
and bradykinin [31, 32]. On the other hand, the second
stage of the edema might be related to the arachidonic
acid metabolites, since inhibition occurs by aspirin, in-
domethacin, and other cyclooxygenase inhibitors [31,
32].Therefore, in addition to edema measurements eve-
ry 90 min during 360 min by the carrageenan-induced
hind paw edema test, serotonin-induced edema test [30]
was also applied to explore the stage of inflammation on
which the compounds exhibited their activity. Moreover,
acute toxicity and gastric ulcerogenic effects of the title
compounds were investigated in test animals.
In this study, 7-benzoyl-5-chloro-2-oxo-3H-benzoxazole
derivatives (4 a–c) were obtained in good yields by the
cyclization of 3 a–c using ethyl chloroformate in pyridine
under mild conditions. The sodium salts of compound
4 a–c were reacted with ethyl 2-bromoacetate and ethyl
4-chlorobutanoate to prepare compound 5 a–c and
7 a–c, respectively. These ester derivatives were hydro-
lyzed to corresponding free acids (compound 6 a–c and
8 a–c) in conc. HCl.
Among the synthesized compounds, anti-inflammatory
and analgesic activity of nonsubtituted derivatives on po-
sition 3 (4 a–c) were found weaker than those bearing
cyanoalkyl, ester, or acid residues on this position at
100 mg/kg dose.
Instead of nucleophilic substitution reaction, Michael-
type addition of compound 4 a–c to acrylonitrile was ap-
plied to prepare propionitrile derivatives (9 a–c). Finally,
these propionitrile derivatives were converted to corre-
sponding acids (10 a–c) and ethyl ester derivatives
(11 a–c) by treatment with aqueous acid and absolute
ethanol in the presence of acid catalyst, respectively.
A comparable analgesic activity to aspirin was observed
for 7-acyl-2-oxo-3H-benzoxazole derivatives (9 a–c)
bearing a cyanoethyl group on position 3.It is noteworthy
that chlorine (2-Cl or 4-Cl) substitution on the acyl group
did not induce any remarkable change in the activity of
these derivatives.
The structure and the other properties of the title com-
pounds are given in Table 1.
At 100 mg/kg dose level, the highest anti-inflammatory
and analgesic activity was observed, as expected, by the
derivatives bearing alkanoic acid residue at position 3
(6 a–c, 8 a–c, 10 a–c) (Table 2). Therefore, these com-
pounds were further tested in higher (200 mg/kg) and
lower (50 mg/kg) dose levels (Table 3). Moreover, inhibi-
tory effects on serotonin-induced hind paw edema and
acetic acid-induced capillary permeability (modified
Whittle test) are shown in Table 4.
Pharmacology
Analgesic activities of the resulting compounds were in-
vestigated by p-benzoquinone-induced writhing test [25]
which is a well established method of testing the analge-
sic activity of compounds and sufficiently sensitive to de-
tect the effect of analgesics which are less active than
aspirin. However, the test can not distinguish between
narcotic and nonnarcotic drugs and may even give a
positive result for compounds which are not regarded as
clinically useful analgesics [26–28]. Therefore, we also
applied the acetic acid-induced peritoneal capillary per-
meability test in mice which is a relatively reliable way of
testing nonnarcotic analgesics [29] for those carboxylic
acids that are more active compounds than the other de-
rivatives in the p-benzoquinone-induced writhing test.
Among the nonsubstituted benzoyl derivatives (6 a, 8 a,
10 a) higher analgesic and anti-inflammatory activities
were observed in connection with a longer chain length
in alkanoic acid residue which also coincided with the
Whittle test results.Interestingly, in spite of the gastric le-
sion inducing effect of 6 a, propanoic acid (10 a) and bu-
tanoic acid (8 a) derivatives were found free of such a
side effect. However, aspirin which is known to induce
gastric ulceration was also found free of gastric lesion ef-
fect at a dose of 100 mg/kg under our test conditions.
2-Chlorobenzoyl derivatives (6 b, 8 b, 10 b) showed the
highest inhibitory activity in all analgesic and anti-inflam-
matory tests employed in the present study.It is notewor-
thy that the incidence of gastric damage increased sig-
nificantly for these derivatives. Although the gastric le-
sion incidence was established for these derivatives,
compound 10 b showed a smaller gastric lesion-induc-
Anti-inflammatory activities of the compounds were as-
sessed by utilizing carrageenan-induced hind paw ede-
ma model [30]. Since the carrageenan edema has been
used in the development of indomethacin, many re-
searchers adapted this procedure for screening poten-
tial anti-inflammatory compounds. It is well known that

314 Ünlü et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Table 2. Effects of compounds on carrageenan-induced hind paw edema model and PBQ-induced writhings.
Compound
Anti-inflammatory Activity
Thickness of the edema ± SEM
(Inhibition, %)
Analgesic
Activity
Number of
stretching
(Inhibition, %)
Gastric-
ulcerogenic
Effect
90 min
180 min
270 min
360 min
Control
4 a
37.5 2.74
33 4.1
(12.0)
51.3 2.88
46.3 3.02
(9.7)
60.7 2.64
55.2 2.4
(9.1)
60.3 2.55
54 4.25
(10.4)
37 4.89
24.2 1.54
(34.6)*
0/6
0/6
4 b
4 c
5 a
5 b
5 c
6 a
6 b
6 c
7 a
7 b
7 c
8 a
8 b
8 c
9 a
9 b
9 c
10 a
10 b
31.8 2.15
(15.2)
44.7 2.99
(12.9)
51 3.62
(15.9)
50.5 2.54
(16.3)*
20.7 2.74
(44.2)*
1/6
0/6
0/6
0/6
0/6
2/6
1/6
0/6
0/6
1/6
0/6
0/6
2/6
0/6
2/6
0/6
0/6
0/6
1/6
31.2 3.89
(16.8)
43.3 2.64
(15.6)
50.3 3.58
(17.1)*
48.3 2.25
(19.9)**
20.2 1.58
(45.4)**
32 3.52
(14.7)
41.3 3.20
(19.6)
47.0 2.88
(22.5)
44.6 1.94
(25.9)*
19.2 1.96
(48.1)**
30.6 2.67
(18.4)
40.2 2.4
(21.6)
42.9 1.8
(29.3)*
39.4 1.65
(34.6)**
14.7 1.28
(60.3)*
30.6 2.9
(18.4)
38.7 2.81
(24.5)
42 1.85
(30.8)*
40.1 1.97
(33.5)**
24.2 1.54
(34.6)*
30.8 3.7
(17.9)
42.8 3.18
(16.6)
47 2.75
(22.6)**
42.5 2.5
(29.5)***
15.58 1.21
(58.1)**
26 1.88
(30.7)**
35.3 2.26
(31.2)**
26.5 3.29
(56.3)***
26.2 3.29
(56.6)***
9.14 1.22
(75.3)***
32.2 2.27
(14.1)
41.5 3.18
(19.1)*
50.2 3.18
(17.3)*
49.5 3.52
(17.9)*
23.4 1.94
(36.6)*
34 2.06
(9.3)
45.6 1.95
(11.2)
49.7 1.63
(18.1)
48.3 1.74
(19.9)
18.7 2.12
(49.5)**
33.6 2.5
(10.5)
45.8 1.94
(10.8)
46.1 2.46
(24.1)
45.3 2.12
(24.9)*
16.3 1.78
(55.9)**
34 2.54
(9.3)
44 2.1
(14.3)
43.5 1.25
(28.3)*
43.5 1.5
(27.8)*
19.8 1.01
(46.5)**
28.3 3.37
(24.5)
38.3 3.65
(25.3)*
35.8 3.54
(41.0)***
34.2 3.34
(43.3)***
11.83 0.76
(68.0)***
26.8 2.92
(28.5)*
36.8 3.24
(28.3)**
31.7 4.04
(47.8)***
30.7 2.04
(49.1)***
8.42 1.51
(77.2)***
29 3.44
(22.7)
37.5 3.24
(26.9)**
40 3.2
(34.1)***
35.3 3.31
(41.5)***
12.6 1.24
(65.9)***
33.7 2.41
(10.0)
43.9 1.65
(14.3)
44.9 1.25
(25.9)*
42.1 1.57
(30.2)**
14 1.63
(62.2)**
32.6 4.18
(12.9)
42.1 2.82
(18.0)
45.8 2.68
(24.6)
41.7 1.6
(30.8)**
13.2 0.7
(64.3)***
30.3 2.14
(19.1)
44.6 2.46
(13.1)
47 2.52
(22.5)
43.1 1.85
(28.5)*
13.3 1.17
(64.1)***
30.3 1.99
(19.2)
40.8 3.04
(20.5)*
40.7 3.77
(32.9)**
41 2.56
(32.0)***
13.8 2.14
(62.8)**
25 2.38
(33.3)**
29 2.89
(43.5)***
20.8 2.02
(65.7)***
25.5 1.98
(57.7)***
7.17 1.06
(80.6)***

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Novel analgesic and anti-inflammatory benzoxazole derivatives 315
Table 2. (continued).
Compound
Anti-inflammatory Activity
Thickness of the edema ± SEM
(Inhibition, %)
Analgesic
Activity
Number of
stretching
(Inhibition, %)
Gastric-
ulcerogenic
Effect
90 min
180 min
270 min
360 min
10 c
11 a
11 b
11 c
30.7 3.89
(18.1)
41.3 3.43
(19.5)*
45.5 2.83
(25.0)**
47.3 4.09
(21.6)*
18.6 2.06
(49.7)**
0/6
0/6
0/6
1/6
33.4 2.55
(11.0)
44.8 2.41
(12.7)
51.2 2.45
(15.6)
51 2.02
(15.4)
20.2 2.09
(45.4)
36.3 3.66
(3.2)
43.5 2.81
(15.3)
47.6 2.11
(21.5)
50.6 1.6
(16.2)
26.2 1.99
(29.2)
30.1 4.21
(19.8)
41 3.64
(20.2)
43.8 2.85
(25.8)
44 2.3
(27.0)*
13.3 1.26
(64.1)***
Indomethacin
Aspirin
26.3 2.33
(29.8)*
26.3 2.78
(48.7)***
25.3 3.28
(58.3)***
29.5 3.25
(51.1)***
–
2/6
0/6
13 1.79
(64.9)***
*: p < 0.05; **: p < 0.01; ***: p < 0.001.
Table 3. Carrageenan-induced hind paw edema model test results.
Compound
Dose.
mg/kg.
Per os
Inhibition of swelling (%)
Analgesic
Activity
% inhibition
of writhings
Gastric-
ulcerogenic
Effect
90 min
180 min
270 min
360 min
6 a
6 b
6 c
8 a
8 b
8 c
50
100
200
12.4
17.9
32.8
11.8
16.6
35.9
11.8
22.6**
34.3
17.6
29.5***
33.5**
27.5**
58.1**
62.3***
1/6
2/6
3/6
50
100
200
16.2
30.7**
33.1*
17.6
31.2**
38.2***
18.4
56.3***
42.7***
22.1
56.6***
50.2***
38.7***
75.3***
76.9***
0/6
1/6
3/6
50
100
200
12.9
14.1
16.8
11.2
19.1*
24.9*
13.7
17.3*
20.8***
14.9
17.9*
22.1**
15.0
36.6*
46.3***
0/6
0/6
0/6
50
100
200
12.9
24.5
41.5**
12.5
25.3*
38.5***
12.2
41.0***
40.9***
18.3
43.3***
44.7***
35.6***
68.0***
70.4***
0/6
0/6
0/6
50
100
200
11.2
28.5*
44.2**
14.4
28.3**
44.1***
16.9
47.8***
43.6***
19.6
49.1***
46.3***
40.5***
77.2***
75.0***
0/6
2/6
3/6
50
100
200
12.2
22.7
37.5**
11.2
26.9**
38.2***
15.2
34.1***
41.4***
16.8
41.4***
40.2***
32.2**
65.9***
70.6***
0/6
0/6
0/6

316 Ünlü et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Table 3. (continued).
Compound
Dose.
mg/kg.
Per os
Inhibition of swelling (%)
Analgesic
Activity
% inhibition
of writhings
Gastric-
ulcerogenic
Effect
90 min
180 min
270 min
360 min
10 a
10 b
10 c
50
100
200
50
100
200
50
100
200
10
100
200
50
12.2
19.2
27.4*
12.9
20.5*
25.1**
17.8
43.5***
42.8***
12.9
19.5*
22.2
48.7***
13.7
32.9**
30.8***
20.3
65.7***
50.6***
15.6
25.0**
27.2
58.3***
17.9
32.0***
35.8***
26.6
57.7***
48.7***
15.9
21.6*
34.2**
51.1***
26.4**
62.8**
64.8***
45.1***
80.6***
78.8***
22.9*
49.7**
58.3***
–
0/6
0/6
0/6
0/6
1/6
2/6
0/6
0/6
0/6
2/6
0/6
2/6
0/6
10.5
33.3**
35.2*
11.7
18.1
24.1
Indomethacin
ASA
29.8*
64.9***
69.4***
31.3**
ASA
ASA
*: p < 0.05; **: p < 0.01; ***: p < 0.001.
Table 4. Serotonin-induced paw edema model and acetic acid-induced capillary permeability test results.
Com-
pound mg/kg.
Per os
Dose.
Swelling in thickness (× 10–2 mm) ± SEM
(% inhibition)
Dye
leakage ± SEM
(% inhibition)
0 min
6 min
12 min
18 min
24 min
30 min
Control
4.0 0.4
4.3 0.3
11.2 1.7 19.7 1.2 30.2 2.6 35.7 3.1 40.2 2.7
12.7 1.3 17.7 1.1 26.2 2.1 31.2 2.1 33.3 2.6
8.37 0.43
6.60 0.44
(21.9)*
4.73 0.31
(43.4)***
7.06 0.72
(15.7)
5.70 0.41
(31.9)***
5.19 0.38
(38.0)***
5.53 0.34
(33.9)***
6.10 0.29
(27.1)***
4.45 0.32
(46.8)***
6 a
100
100
100
100
100
100
100
100
100
10
(10.2)
11.7 1.2 17.0 1.9 24.3 1.5 28.3 0.9 30.5 2.5
(13.7) (19.5) (20.7)* (24.1)*
17.7 1.9 25.2 2.3 25.3 2.8 29.2 2.0 35.0 2.1
(16.2) (18.2) (12.9)
10.7 2.0 17.2 1.9 25.3 2.7 29.3 2.1 33.8 2.6
(13.2)
(12.6)
(17.2)
6 b
3.7 0.3
(8.3)
3.5 0.4
(12.5)
3.5 0.4
(12.5)
4.0 0.51
6 c
8 a
(4.5)
1.8 1.2
(12.7)
17.5 1.2 25.7 1.9 31.7 1.3 34.5 1.3
(11.2) (14.9) (11.2) (14.2)
26.0 1.6 31.2 2.7 33.2 2.6
(13.9) (12.6) (17.4)
(16.2)
(17.9)
(15.9)
8 b
8 c
4.2 0.5 12.7 1.11 8.3 1.9
(7.1)
3.8 0.5
(4.3)
3.7 0.5
(8.3)
4.5 0.6
10 a
10 b
10 c
12.5 1.9 16.5 1.7 25.5 2.2 29.3 2.5 35.3 2.5
(16.2) (15.6) (17.9) (12.2)
11.8 1.2 15.3 1.5 22.8 3.7 27.5 1.6 31.5 1.8
(22.3)* (24.5) (22.9)* (21.6)*
12.8 1.2 17.7 2.4 27.8 1.9 30.5 2.3 33.5 2.4
(10.2) (7.9) (14.6) (16.7)
11.7 1.2 15.2 1.3 22.2 0.9 24.8 1.2 26.2 1.8
(22.8)* (26.5)* (30.5)** (34.8)**
6.02 0.23
(28.0)***
4.34 0.27
(48.1)***
Indo-
methacin
4.0 0.4
*:p < 0.05;**:p < 0.01;***:p < 0.001.

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Novel analgesic and anti-inflammatory benzoxazole derivatives 317
ing effect than compounds 6 b and 8 b.In addition, espe-
cially among the synthesized compounds, the highest
and significant activity against serotonin-induced edema
was shown by 6 b and 10 b.
Experimental
Chemistry
All chemicals were obtained from Aldrich (Germany) or Merck
Chemical Co. (Darmstadt, Germany); they were stored over a
4Å-molecular sieve and used as received. 2-Acetylamino-4-
chlorophenol was prepared according to the method reported
earlier [21]. Melting points were determined with Thomas Hoo-
ver Capillary Melting Points Apparatus (Arthur H.Thomas Co.,
Philadelphia, PA, USA) and the values given are uncorrected.
IR spectra were recorded on a BrukerVector 22 IR (Opus Spec-
troscopic Software Version 2.0) spectrometer (KBr, υ, cm–1)
(Bruker Analytic GmbH, Ettlingen, Germany).Nuclear magnet-
ic resonance (1H-NMR) spectra were recorded on a Bruker
400 FT-NMR spectrometer (Bruker) Spectra were calibrated
using tetramethylsilane (TMS) (δ = 0.00). All chemical shifts
were reported as δ (ppm) values.Elemental analyses were per-
formed with Leco-932 (C,H,N,S-O-Elemental Analyzer) at Sci-
entific and Technical Research Council of Turkey, Instrumental
Analysis Center (Ankara, Turkey) and were within the range of
0.4 % of theoretical value.
A less remarkable analgesic and anti-inflammatory ac-
tivity was observed for 4-chlorobenzoyl derivatives (6 c,
8 c and 10 c) which also correlated with the results of
peritoneal capillary permeability test, although, even the
highest doses of these derivatives did not induce any ap-
parent gastric toxicity. The activity of these compounds
increased considerably upon increasing alkanoic acid
chain length. In fact, this kind of conclusion may be
drawn for the afore-mentioned other derivatives as well.
If a comparison would be made between the butanoic
acid derivatives, 8 b caused gastric damage while the
other two compounds 8 a and 8 c did not.
In general, the following conclusions may be drawn:
2-Acetylamino-6-benzoyl-4-chlorophenol derivatives (2 a–c)
1.All of the compounds exhibited their analgesic and an-
ti-inflammatory activity at the second stage of the inflam-
mation according to the test results. Higher analgesic
and anti-inflammatory activity of 2-chlorobenzoyl deriva-
tives (6 b, 8 b, and 10 b) may be attributed to the effect of
chlorine substituent that forces the phenyl ring to take a
position in a different plane than the 2-oxo-3H-benzox-
azole ring causing a structure that resembles indometh-
acin derivatives. This effect of the chlorine substituent
might explain the activity observed with these com-
pounds.
N,N-dimethylformamide (DMF) (0.22 mol) was added dropwise
to anhydrous aluminium chloride (0.8 mol) at 0–10 °C, stirring
constantly and protecting the reaction mixture from atmospher-
ic moisture. After addition was completed, the mixture was
stirred for further 10 minutes.The appropriate benzoyl chloride
derivative (0.14 mol) was added portionwise to this mixture
over 15 minutes. After addition of 2-acetylamino-4-chlorophe-
nol (0.1 mol), the mixture was heated for six hours at 120 °C,
cooled and poured into 1 L ice-water containing 10 mL concen-
trated hydrochloric acid.The product precipitated, it was filtered
and dissolved in 1 L aqueous sodium hydroxide solution
(0.2 N).The solution was filtered and acidified with concentrat-
ed hydrochloric acid.The precipitate was filtered, washed with
water to neutral pH, dried, and crystallized from the appropriate
solvent.
2.It should be noticed that the analgesic and anti-inflam-
matory activities of alkanoic acid derivatives bearing
benzoyl and 4-chlorobenzoyl substituents (6 a, 6 c, 8 a,
8 c, 10 a, and 10 c) were enhanced, apparently by elon-
gating the alkanoic acid chain while especially propanoic
acid (10 a and 10 c) and butanoic acid derivatives (8 a
and 8 c) were devoid of any gastric lesion.
2-Amino-6-benzoyl-4-chlorophenol derivatives (3 a, 3 c)
2-Acetylamino-6-benzoyl-4-chlorophenol derivative (0.1 mol)
was added to 300 mL of 40 % sodium hydroxide solution and
200 mL ethanol.This solution was refluxed for 7 h, evaporated
under vacuum to remove ethanol, and cooled.After adding 500
mL water, the solution was filtered to remove insoluble material
and acidified with acetic acid to pH = 5.5. The product precipi-
tated was filtered, washed with water to neutral pH, dried, and
crystallized from appropriate solvent.
According to the results of in vivo studies conducted in
the present study, the analgesic and anti-inflammatory
activities of 8 a and 8 c were comparable to that of known
drugs, i.e. aspirin and indomethacin, without inducing
any visible gastric damage. Further detailed studies are
under way in order to investigate the effect of the synthe-
sized alkanoic acid derivatives on chronic inflammatory
test models, as well as antipyretic effect, and COX-2 se-
lective inhibitory effects.
2-Amino-6-benzoyl-4-chlorophenol hydrochloride (3 b)
The mixture of 2-acetylamino-4-chloro-6-(2-chlorobenzoyl)-
phenol (0.1 mol) and 250 mL concentrated hydrochloric acid
was refluxed for 3 h, and then diluted with 500 mL water.The in-
soluble material was removed by hot filtration.The solution was
allowed to cool and crystals separated from the solution were
collected by suction filtration.
7-Benzoyl-5-chloro-2-oxo-3H-benzoxazole derivatives (4 a–c)
Ethyl chloroformate (0.2 mol) was added dropwise to the solu-
tion of 2-amino-6-benzoyl-4-chlorophenol derivative (0.1 mol)
in pyridine while stirring and cooling in an ice bath. After stirring
for further 30 minutes, the solution was heated under a con-
denser at 100 °C in an oil bath for 2.5–5 h.After cooling to room
temperature, the solution was poured into 500 mL ice-water
and acidified with 150 mL concentrated hydrochloric acid.The
Acknowledgement
This investigation was supported by the Scientific and
Technical Research Council of Turkey, TÜBITAK
(Research Grant # SBAG-2497).
·

318 Ünlü et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
precipitate was collected by suction filtration, washed with wa-
ter, dried, and crystallized from the appropriate solvent.
was added to this solution until the precipitation was complete.
The precipitate was filtered, washed with water, dried, and
crystallized from appropriate solvents.
Ethyl 2-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)ace-
tate Derivatives (5 a–c)
2-Acetylamino-6-benzoyl-4-chlorophenol (2 a)
7-Benzoyl-5-chloro-2-oxo-3H-benzoxazole derivative (10 mmol)
was dissolved in an ethanolic solution of sodium ethoxide pre-
pared by the reaction of Na° (10 mmol) with 10 mL ethanol.
After addition of ethyl bromoacetate (11 mmol), the solution
was refluxed for 3–8 h and cooled to room temperature.To pre-
cipitate the product, 50 mL water was added to this solution.
The precipitate was filtered, dried, and crystallized from the ap-
propriate solvent.
1H-NMR (CDCl3):δ 12.7 (s, 1 H, -OH), 8.76 (d, 1 H, J = 2.5, H5),
8.02 (s, 1 H, -NH-CO), 7.79–7.61 (m, 5 H, H2Ј-H6Ј), 7.36 (d, 1 H,
J = 2.5, H3) and 2.35 (s, 3 H, -CO-CH3). IR νmax cm–1 (KBr):
3308 (N-H), 3145 (O-H), 1676 (C=O, amide) 1625 (C=O, ke-
tone), 1608, 1598 (C=C, aromatic). Anal.: C15H12ClNO3.
2-Acetylamino-4-chloro-6-(2-chlorobenzoyl)phenol (2 b)
1H-NMR (DMSO-d6): δ 11.44 (s, 1 H, -OH), 9.57 (s, 1 H, -NH-
CO), 8.13 (d, 1 H, J = 2.43, H5), 7.54–7.41 (m, 4 H, H3Ј-H6Ј),
6.84 (d, 1 H, J = 2.59, H3), 2.06 (s, 3 H, CO-CH3). IR νmax cm–1
(KBr): 3347 (N-H), 3196 (O-H), 1691 (C=O, amide), 1642
(C=O, ketone), 1593 (C=C, aromatic). Anal.: C15H11Cl2NO3.
Ethyl
4-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)bu-
tanoate Derivatives (7 a–c)
7-Benzoyl-5-chloro-2-oxo-3H-benzoxazole derivative (10 mmol)
was treated with sodium ethoxide (10 mmol) in 20 mL ethanol.
After evaporation to dryness under vacuum, the residue was
dissolved in 30 mL N,N-dimethylformamide and then ethyl 4-
chlorobutanoate (11 mmol) was added, heated for 12–20 h. at
80–100 °C in an oil-bath, cooled to room temperature, poured
into 50 mL ice-water. The precipitate was collected by suction
filtration, dried and crystallized from the appropriate solvent.
2-Acetylamino-4-chloro-6-(4-chlorobenzoyl)phenol (2 c)
1H-NMR (DMSO-d6): δ 12.43 (s, 1 H, OH), 8.73 (d, 1 H, J =
2.35, H5), 7.92 (s, 1 H, -NH-CO), 7.67–7.53 (m, 4 H, H2Ј-H6Ј),
7.26 (d, 1 H, J = 2.49, H3), 2.28 (s, 3 H, CO-CH3). IR νmax cm–1
(KBr): 3269 (N-H), 3125 (O-H), 1672 (C=O, amide), 1625
(C=O, ketone), 1607 (C=C, aromatic). Anal.: C15H11Cl2NO3.
2-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)acetic acid
(6 a–c) and 4-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-
yl)butanoic acid (8 a–c) Derivatives
2-Amino-6-benzoyl-4-chlorophenol (3 a)
1H-NMR (DMSO-d6) δ: 10.6 (s, 1 H, OH), 7.72–7.54 (m, 5 H,
benzoyl H2Ј-H6Ј), 6.88 (d, 1 H, J = 2.52, H5), 6.54 (d, 1 H, J =
2.52, H3), 5.4 (s, 2 H, NH2).IR νmax cm–1 (KBr):3441 (NH2 asym-
metric), 3332 (NH2 symmetric), 3079 (OH), 1621 (C=O,
ketone), 1596, 1574 (C=C, aromatic). Anal.: C13H10ClNO2.
The mixture of ethyl (7-benzoyl-5-chloro-2-oxo-3H-benzox-
azol-3-yl)acetate (3 mmol) or ethyl 4-(7-benzoyl-5-chloro-2-
oxo-3H-benzoxazol-3-yl)butanoate (3 mmol) and 30 mL hydro-
chloric acid (conc.) was refluxed for 3–5 h. and cooled to room
temperature. The product precipitated, it was filtered off and
dissolved in 50 mL aqueous sodium bicarbonate (5 %).The so-
lution was filtered and acidified with hydrochloric acid (2 N).The
precipitate was filtered, washed with water, dried, and crystal-
lized from the appropriate solvent.
2-Amino-4-chloro-6-(2-chlorobenzoyl)phenol
(3 b)
hydrochloride
1H-NMR (DMSO-d6) δ: 7.45–7.28 (m, 4 H, H3Ј-H6Ј), 7.27–6.95
(broad, 3 H, -+NH3), 6.92 (d, 1 H, J = 2.49, H5), 6.23 ( d, 1 H, J =
2.49, H3). IR νmax cm–1 (KBr): 3150–3090 (OH), 2793–2537
(+NH3), 1632 (C=O, ketone), 1613, 1588 (C=C, aromatic).
Anal.: C13H10Cl3NO2.
3-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)propionitrile
Derivatives (9 a–c)
2-Amino-4-chloro-6-(4-chlorobenzoyl)phenol (3 c)
The solution of 7-benzoyl-5-chloro-2-oxo-3H-benzoxazole
(10 mmol), acrylonitrile (15 mmol), and triethylamine
(50 mmol) in 40 mL ethyl acetate was refluxed for 10–15 h, and
cooled. The precipitated product was filtered, dried, and crys-
tallized from the appropriate solvents.
1H-NMR (CDCl3): δ 12.00 (s, 1 H, OH), 7.66–7.52 (m, 4 H, H2Ј-
H6Ј), 6.90 (s, 2 H, H5-H3), 4.12 (s, 2 H, -NH2). IR νmax cm–1 (KBr):
3423 (NH2 asymmetric), 3310 (NH2 symmetric), 3175 (OH),
1620 (C=O, ketone), 1597 (C=C, aromatic). Anal.:
C13H9Cl2NO2.
3-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)propanoic
acid Derivatives (10 a–c)
7-Benzoyl-5-chloro-2-oxo-3H-benzoxazole (4 a)
3-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)propionitrile
(3 mmol) was refluxed in the mixture of N,N-dimethylform-
amide (20 mL), sulphuric acid (10 mL), and water (10 mL) for
10 h, left overnight without refluxing and was then refluxed for
additional 8 h. After cooling to room temperature, the product
precipitated, it was filtered and dissolved in 50 mL aqueous so-
dium bicarbonate solution (5 %).The solution was filtered to re-
move insoluble material and acidified with hydrochloric acid
(conc.). The precipitate that formed was filtered, dried, and
crystallized from the appropriate solvent.
1H-NMR (CDCl3): δ 8.71 (s, 1 H, -NH-), 8.04–7.71 (m, 5 H, H2Ј-
H6Ј) 7.58 (d, 1 H, J = 2.05, H6), 7.44 (d, 1 H, J = 2.05, H4).IR νmax
cm–1 (KBr): 3318 (N-H, lactam), 1780 (C=O, lactam), 1663
(C=O, ketone), 1608 (C=C, aromatic). Anal.: C14H8ClNO3.
5-Chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazole (4 b)
1H-NMR (CDCl3): δ 8.98 (s, 1 H, -NH-), 7.43 (d, 1 H, J = 1.95,
H6), 7.42–7.30 (m, 4 H, H3Ј-H6Ј), 7.19 (d, 1 H, J = 2.07, H4). IR
ν
max cm–1 (KBr): 3176 (N-H, lactam), 1775 (C=O, lactam), 1658
(C=O, ketone), 1600, 1587 (C=C, aromatic). Anal.:
C14H7Cl2NO3.
Ethyl
3-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)pro-
5-Chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazole (4 c)
panoate Derivatives (11 a–c)
1H-NMR (CDCl3 + DMSO-d6): δ 11.78 (s, 1 H, -NH-), 7.70–7.42
(m, 4 H, H2Ј-H6Ј), 7.18 (d, 1 H, J = 2.09, H6), 7.15 (d, 1 H, J =
2.12, H4). IR νmax cm–1 (KBr): 3193 (N-H, lactam), 1780 (C=O,
lactam), 1666 (C=O, ketone), 1609, 1589 (C=C, aromatic).
Anal.: C14H7Cl2NO3.
The solution of 3-(7-benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-
yl)propionitrile derivative (3 mmol) in a previously prepared and
cooled mixture of ethanol (20 mL) and sulphuric acid (10 mL)
was refluxed for 1 h, then cooled to room temperature. Water

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Novel analgesic and anti-inflammatory benzoxazole derivatives 319
Ethyl 2-(7-benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)ace-
tate (5 a)
Ethyl 4-[5-chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-
yl]butanoate (7 b)
1H-NMR (CDCl3):δ 8.02–7.70 (m, 5 H, H2Ј-H6Ј), 7.60 (d, 1 H, J =
2.02, H6), 7.23 (d, 1 H, J = 2.02, H4), 4.76 (s, 2 H, N-CH2-CO),
4.49 (q, 2 H, J = 7,14, O-CH2-CH3), 1.52 (t, 3 H, J = 7,14, O-
CH2-CH3). IR νmax cm–1 (KBr): 2992 (C-H, aliphatic), 1789
(C=O, lactam), 1732 (C=O, ester), 1659 (C=O, ketone), 1614,
1597 (C=C, aromatic). Anal.: C18H14ClNO5.
1H-NMR (CDCl3): δ 7.42 (d, 1 H, J = 2.07, H6), 7.39–7.28 (m,
4 H, H3Ј-H6Ј), 7.15 (d, 1 H, J = 2.08, H4), 4.06 (q, 2 H, J = 7,13,
-O-CH2-CH3), 3.79 (t, 2 H, N-CH2-CH2), 2.31 (t, 2 H, CH2-CH2-
CO), 1.99 (m, 2 H, CH2-CH2-CH2), 1.17 (t, 3 H, J = 7,13,
-O-CH2-CH3). IR νmax cm–1 (KBr): 3078 (C-H, aromatic), 2981,
2920, 2890 (C-H, aliphatic), 1786 (C=O, lactam), 1724 (C=O,
ester), 1670 (C=O, ketone), 1604, 1589 (C=C, aromatic).Anal.:
C20H17Cl2NO5.
Ethyl 2-[5-chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-
yl]acetate (5 b)
Ethyl 4-[5-chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-
yl]butanoate (7 c)
1H-NMR (DMSO-d6): δ 7.81 (d, 1 H, J = 2.1, H6), 7.55–7.42 (m,
4 H, H3Ј-H6Ј), 7.28 (d, 1 H, J = 2.1, H4), 4.70 (s, 2 H, -N-CH2-
CO), 4.13 (q, 2 H, J = 7,10, -O-CH2-CH3), 1.15 (t, 3 H, J = 7,10,
-O-CH2-CH3). IR νmax cm–1 (KBr): 3070, 3059 (C-H, aromatic),
2987, 2937 (C-H, aliphatic), 1793 (C=O, lactam), 1741 (C=O,
ester), 1659 (C=O, ketone), 1612, 1590 (C=C, aromatic).Anal.:
C18H13Cl2NO5.
1H-NMR (CDCl3):δ 7.68–7.40 (m, 4 H, H2Ј-H6Ј), 7.29 (d, 1 H, J =
2.03, H6), 7.17 (d, 1 H, J = 2.03, H4), 4.07 (q, 2 H, J = 7,15, O-
CH2-CH3), 3.85 (t, 2 H, N-CH2-CH2), 2.37 (t, 2 H, -CH2-CH2-
CO), 2.04 (m, 2 H, -CH2-CH2-CH2), 1.20 (t, 3 H, J = 7,15, O-
CH2-CH3). IR νmax cm–1 (KBr): 3100 (C-H, aromatic), 2985,
2936 (C-H, aliphatic), 1782 (C=O, lactam), 1729 (C=O, ester),
1647 (C=O, ketone), 1609, 1587 (C=C, aromatic). Anal.:
C20H17Cl2NO5.
Ethyl 2-[5-chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-
yl]acetate (5 c)
1H-NMR (CDCl3):δ 7.70–7.41 (m, 4 H, H2Ј-H6Ј), 7.32 (d, 1 H, J =
2.0, H6), 7.00 (d, 1 H, J = 2.0, H4), 4.49 (s, 2 H, N-CH2-CO), 4.22
(q, 2 H, J = 7,15, O-CH2-CH3), 1.24 (t, 3 H, J = 7,15, O-CH2-
CH3). IR νmax cm–1 (KBr): 3067, 3062 (C-H, aromatic), 2998,
2980 (C-H, aliphatic), 1782 (C=O, lactam), 1742 (C=O, ester),
1664 (C=O, ketone), 1600, 1586 (C=C, aromatic). Anal.:
C18H13Cl2NO5.
4-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)butanoic acid
(8 a)
1H-NMR (DMSO-d6): δ 11.81 (s, 1 H, -COOH), 7.69–7.43 (m,
5 H, H2Ј-H6Ј), 7.61 (d, 1 H, J = 2.13, H6), 7.18 (d, 1 H, J = 2.07,
H4), 3.73 (t, 2 H, N-CH2-CH2), 2.35 (t, 2 H, CH2-CH2-CO), 1.79
(m, 2 H, CH2-CH2-CH2). IR νmax cm–1 (KBr): 3300–2400 (O-H,
carboxylic acid), 3110 (C-H, aromatic), 2928 (C-H, aliphatic),
1786 (C=O, lactam), 1731 (C=O, carboxylic acid), 1652 (C=O,
ketone), 1610, 1597 (C=C, aromatic). Anal.: C18H14ClNO5.
2-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)acetic acid
(6 a)
1H-NMR (CDCl3):δ 7.87–7.57 (m, 5 H, H2Ј-H6Ј), 7.44 (d, 1 H, J =
2.01, H6), 7.11 (d, 1 H, J = 2.03, H4), 4.65 (s, 2 H, N-CH2-CO).
IR νmax cm–1 (KBr): 3300–2500 (O-H, carboxylic acid), 1744
(C=O, lactam), 1734 (C=O, carboxylic acid), 1663 (C=O,
ketone), 1608, 1594 (C=C, aromatic). Anal.: C16H10ClNO5.
4-[5-Chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]bu-
tanoic acid (8 b)
1H-NMR (CDCl3): δ 7.41 (d, 1 H, J = 2.06, H6), 7.40–7.28 (m,
4 H, H3Ј-H6Ј), 7.13 (d, 1 H, J = 2.06, H4), 3.82 (t, 2 H, N-CH2-
CH2-), 2.42 (t, 2 H, CH2-CH2-CO), 2.02 (m, 2 H, CH2-CH2-CH2).
IR νmax cm–1 (KBr):3300–2750 (O-H, carboxylic acid), 3031 (C-
H, aromatic), 2968, 2953 (C-H, aliphatic), 1782 (C=O, lactam),
1733 (C=O, carboxylic acid), 1664 (C=O, ketone), 1608, 1589
(C=C, aromatic). Anal.: C18H13Cl2NO5.
2-[5-Chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]ace-
tic acid (6 b)
1H-NMR (DMSO-d6): δ 7.81 (d, 1 H, J = 2.1, H6), 7.55–7.42 (m,
4 H, H3Ј-H6Ј), 7.27 (d, 1 H, J = 2.1, H4), 4.58 (s, 2 H, N-CH2-CO).
IR νmax cm–1 (KBr): 3300–2500 (O-H, carboxylic acid), 3098
(C-H, aromatic), 2987, 2956 (C-H, aliphatic), 1773 (C=O,
lactam), 1748 (C=O, carboxylic acid), 1676 (C=O, ketone),
1613, 1500 (C=C, aromatic). Anal.: C16H9Cl2NO5.
4-[5-Chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]-
butanoic acid (8 c)
1H-NMR (DMSO-d6): δ 11.77 (s, 1 H, -COOH), 7.55-7.26 (m,
4 H, H2Ј-H6Ј), 7.29 (d, 1 H, J = 2.1, H6), 7.04 (d, 1 H, J = 2.1, H4),
3.66 (t, 2 H, N-CH2-CH2-), 2.15 (t, 2 H, CH2-CH2-CO), 1.76 (m,
2 H, CH2-CH2-CH2). IR νmax cm–1 (KBr): 3300–2500 (O-H,
carboxylic acid), 3058 (C-H, aromatic), 2950 (C-H, aliphatic),
1790 (C=O, lactam), 1703 (C=O, carboxylic acid), 1651 (C=O,
ketone), 1607, 1589 (C=C, aromatic). Anal.: C18H13Cl2NO5.
2-[5-Chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]ace-
tic acid (6 c)
1H-NMR (CDCl3 + DMSO-d6): δ 7.89–7.61 (m, 4 H, H2Ј-H6Ј),
7.47 (d, 1 H, J = 1.98, H6), 7.42 (d, 1 H, J = 1.98, H4), 4.70 (s,
2 H, N-CH2-CO). IR νmax cm–1 (KBr): 3300–2700 (O-H, carb-
oxylic acid), 3100, 3075 (C-H, aromatic), 2980, 2939 (C-H, ali-
phatic), 1763 (C=O, lactam), 1758 (C=O, carboxylic acid), 1666
(C=O, ketone), 1609, 1586 (C=C, aromatic). Anal.:
C16H9Cl2NO5.
3-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)propionitrile
(9 a)
1H-NMR (DMSO-d6): δ 7.68 (d, 1 H, J = 2.08, H6), 7.62–7.37
(m, 5 H, H2Ј-H6Ј), 7.15 (d, 1 H, J = 2.08, H4), 3.97 (t, 2 H, N-CH2-
CH2), 2.81 (t, 2 H, -CH2-CH2-CN). IR νmax cm–1 (KBr): 3062
(C-H, aromatic), 2958, 2937 (C-H, aliphatic), 2250 (CN), 1783
(C=O, lactam), 1651 (C=O, ketone), 1609 (C=C, aromatic).
Anal.: C17H11ClN2O3.
Ethyl 4-(7-benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)bu-
tanoate (7 a)
1H-NMR (CDCl3):δ 7.75–7.45 (m, 5 H, H2Ј-H6Ј), 7.31 (d, 1 H, J =
2.04, H6), 7.19 (d, 1 H, J = 2.06, H4), 4.12 (q, 2 H, J = 7,15, O-
CH2-CH3), 3.86 (t, 2 H, N-CH2-CH2), 2.38 (t, 2 H, -CH2-CH2-
CO), 2.07 (m, 2 H, CH2-CH2-CH2), 1.23 (t, 3 H, J = 7,15,
-O-CH2-CH3). IR νmax cm–1 (KBr): 3110 (C-H, aromatic), 2984,
2810 (C-H, aliphatic), 1790 (C=O, lactam), 1729 (C=O, ester),
1652 (C=O, ketone), 1611, 1500 (C=C, aromatic). Anal.:
C20H18ClNO5.
3-[5-Chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]pro-
pionitrile (9 b)
1H-NMR (DMSO-d6): δ 7.88 (d, 1 H, J = 2.1, H6), 7.55–7.43 (m,
4 H, H3Ј-H6Ј), 7.27 (d, 1 H, J = 2.1, H4), 4.09 (t, 2 H, N-CH2-CH2),

320 Ünlü et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
2.93 (t, 2 H, CH2-CH2-CN).IR νmax cm–1 (KBr):3099, 3054 (C-H,
aromatic), 2970, 2937 (C-H, aliphatic), 2250 (CN), 1787 (C=O,
lactam), 1677 (C=O, ketone), 1608, 1591 (C=C, aromatic).
Anal.: C17H10Cl2N2O3.
Ethyl 3-[5-chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-
yl]propanoate (11 c)
1H-NMR (CDCl3):δ 7.68–7.40 (m, 4 H, H2Ј-H6Ј), 7.29 (d, 1 H, J =
2, H6), 7.26 (d, 1 H, J = 2, H4), 4.10 (m, 4 H, O-CH2-CH3, N-CH2-
CH2), 2.77 (t, 2 H, -CH2-CH2-CO), 1.19 (t, 3 H, J = 7,15,
-O-CH2-CH3). IR νmax cm–1 (KBr): 3095 (C-H, aromatic), 2991
(C-H, aliphatic), 1787 (C=O, lactam), 1728 (C=O, ester), 1647
(C=O, ketone), 1608, 1588 (C=C, aromatic). Anal.:
C19H15Cl2NO5.
3-[5-Chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]pro-
pionitrile (9 c)
1H-NMR (DMSO-d6):δ 8.06–7.78 (m, 4 H, H2Ј-H6Ј), 7.81 (d, 1 H,
J = 2.03, H6), 7.66 (d, 1 H, J = 2.03, H4), 4.51 (t, 2 H, N-CH2-
CH2), 3.23 (t, 2 H, CH2-CH2-CN).IR νmax cm–1 (KBr):3085, 3072
(C-H, aromatic), 2983, 2968, 2937 (C-H, aliphatic), 2250 (CN),
1771 (C=O, lactam), 1660 (C=O, ketone), 1608, 1584 (C=C,
aromatic). Anal.: C17H10Cl2N2O3.
Pharmacology
Animals
Male Swiss albino mice (The Animal Breeding Laboratories of
Refik Saydam Hifzisihha Institute Ankara, Turkey) weighing
20–25 g were used for all experiments.The animals were kept
in colony cages (6 mice each), maintained on standard pellet
diet, water ad libitum, and left for two days for acclimatization
before the experimental sessions. The food was withdrawn on
the day before the experiment, but the animals were allowed
free access to water. All experiments were carried out accord-
ing to the suggested ethical guidelines for the care of laboratory
animals.
3-(7-Benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)propanoic
acid (10 a)
1H-NMR (DMSO-d6): δ 12.17 (s, 1 H, -COOH), 7.60–7.34 (m,
5 H, H2Ј-H6Ј), 7.58 (d, 1 H, J = 2.11, H6), 7.10 (d, 1 H, J = 2.11,
H4) 3.82 (t, 2 H, N-CH2CH2), 2.52 (t, 2 H, -CH2CH2-CO). IR νmax
cm–1 (KBr): 3300-2500 (O-H, carboxylic acid), 2926 (C-H,
aliphatic), 1789 (C=O, lactam), 1698 (C=O, carboxylic acid),
1648 (C=O, ketone), 1612, 1598 (C=C, aromatic). Anal.:
C17H12ClNO5.
Preparation of test samples for bioassay
3-[5-Chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]pro-
panoic acid (10 b)
Test samples were given orally to test animals after suspending
them in a mixture of distilled H2O and 0.5 % sodium carboxy-
methyl cellulose (CMC). The animals of the control group re-
ceived the same experimental handling, except that the drug
treatment was replaced with appropriate volumes of the dosing
vehicle. Either indomethacin (10 mg/kg) or aspirin (100 mg/kg)
in 0.5 % CMC was used as reference drug.
1H-NMR (CDCl3): δ 7.42 (d, 1 H, J = 2, H6), 7.4–7.30 (m, 4 H,
H3Ј-H6Ј), 7.21 (d, 1 H, J = 2, H4), 4.02 (t, 2 H, N-CH2-CH2), 2.81
(t, 2 H, N-CH2-CH2). IR νmax cm–1 (KBr): 3390–3100 (O-H,
carboxylic acid), 3086 (C-H, aromatic), 2979, 2950 (C-H,
aliphatic), 1767 (C=O, lactam), 1745 (C=O, carboxylic acid),
1678 (C=O, ketone), 1607, 1592 (C=C, aromatic). Anal.:
C17H11Cl2NO5.
p-Benzoquinone-induced writhing test [25]
60 Minutes after the oral administration of test samples, the
mice were intraperitoneally injected with 0.1 mL/10 g body
weight of 2.5 % (v/v) p-benzoquinone (PBQ) solution in distilled
H2O (PBQ; Merck, Darmstadt, Germany). Control animals re-
ceived an appropriate volume of dosing vehicle.The mice were
then kept individually for observation and the total number of
abdominal contractions (writhing movements) was counted for
the next 15 min, starting on the 5th min after the PBQ injection.
The data represent the average of the total number of writhes
observed. The analgesic activity was expressed as the per-
centage change from writhing controls.
3-[5-Chloro-7-(4-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-yl]pro-
panoic acid (10 c)
1H-NMR (CDCl3 + DMSO-d6): δ 12.02 (s, 1 H, COOH), 7.58–
7.30 (m, 4 H, H2Ј-H6Ј), 7.37 (d, 1 H, J = 2, H6), 7.08 (d, 1 H, J = 2,
H4), 3.92 (t, 2 H, N-CH2-CH2), 2.55 (t, 2 H, CH2-CH2-CO). IR
ν
max cm–1 (KBr): 3300–2500 (O-H, carboxylic acid), 3071 (C-H,
aromatic), 2930 (C-H, aliphatic), 1786 (C=O, lactam), 1706
(C=O, carboxylic acid), 1653 (C=O, ketone), 1610, 1588 (C=C,
aromatic). Anal.: C17H11Cl2NO5.
Carrageenan-induced hind paw edema test [30]
Ethyl 3-(7-benzoyl-5-chloro-2-oxo-3H-benzoxazol-3-yl)pro-
The method of Kasahara et al.[30] was applied.The difference
in footpad thickness between the right and left foot was meas-
ured with a pair of dial thickness gauge callipers (Ozaki Co.,
Tokyo, Japan). Mean values of treated groups were compared
with mean values of a control group and analyzed using statisti-
cal methods. 60 min after the oral administration of test sample
or dosing vehicle each mouse was injected with freshly pre-
pared (0.5 mg/25 µL) suspension of carrageenan (Sigma,
St. Louis, Mo, USA) in physiological saline (154 mM NaCl) into
subplantar tissue of the right hind paw and 25 µL of saline solu-
tion into that of the left as secondary control. Measurements
were done and evaluated every 90 min during 360 min as de-
scribed above.
panoate (11 a)
1H-NMR (DMSO-d6):δ 7.61–7.37 (m, 5 H, H2Ј-H6Ј), 7.58 (d, 1 H,
J = 2, H6), 7.11 (d, 1 H, J = 2, H4), 3.88 (m, 4 H, N-CH2-CH2, O-
CH2-CH3), 2.59 (t, 2 H, -CH2-CH2-CO), 0.92 (t, 3 H, J = 7,13,
-O-CH2-CH3). IR νmax cm–1 (KBr): 3062, 3060 (C-H, aromatic),
2997, 2910 (C-H, aliphatic), 1780 (C=O, lactam), 1719 (C=O,
carboxylic acid), 1651 (C=O, ketone), 1608, 1597 (C=C,
aromatic). Anal.: C19H16ClNO5.
Ethyl 3-[5-chloro-7-(2-chlorobenzoyl)-2-oxo-3H-benzoxazol-3-
yl]propanoate (11 b)
1H-NMR (CDCl3): δ 7.41 (d, 1 H, J = 2.08, H6), 7.40–7.30 (m,
4 H, H3Ј-H6Ј), 7.22 (d, 1 H, J = 2.08, H4), 4.07 (m, 4 H, OCH2CH3,
N-CH2-CH2), 2.73 (t, 2 H, N-CH2-CH2), 1.15 (t, 3 H, J = 7,14,
-OCH2CH3). IR νmax cm–1 (KBr): 3060, 3058 (C-H, aromatic),
2985, 2980, 2974, 2937 (C-H, aliphatic), 1778 (C=O, lactam),
1743 (C=O, ester), 1678 (C=O, ketone), 1604, 1590 (C=C, aro-
matic). Anal.: C19H15Cl2NO5.
Serotonin-induced edema test
The method of Kasahara et al. [30] was used. Briefly, 60 min af-
ter the oral administration of test sample or dosing vehicle each
mouse was injected with serotonin (serotonin creatinin sulfate,
Merck, Art.7768) in Tyrode’s solution (0.5 µg/5 µL) into sub-

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 310–321
Novel analgesic and anti-inflammatory benzoxazole derivatives 321
plantar tissue of the right hind paw and 5 µL ofTyrode's solution
into that of the left as secondary control.The difference in foot-
pad thickness between the right and left foot was measured
every 6 min during 30 min with a pair of dial thickness gauge
callipers (Ozaki).
[7] Z. Moussavi, M. O. Plancke, P. Olivier, D. Lesieur, J. C.
Fruchart, Clin. Chim. Acta 1989, 180, 35–44.
[8] W. J. Close, B. D. Tiffany, M. A. Spielman, J. Am. Chem.
Soc. 1949, 71, 1265–1268.
[9] J.P.Bonte, D.Lesieur, C.Lespagnol, J.C.Cazin, M.Cazin,
Acetic acid-induced peritoneal capillary permeability test
Eur. J. Med. Chem. Chim. Ther. 1974, 9, 491–496.
The influence of the test samples on the increased vascular
permeability induced by acetic acid in mice was determined ac-
cording to Whittle method with some modifications [33]. Each
test sample was administered orally to a group of 10 mice in
0.2 mL per 20 g body weight. 30 min after the administration
each mouse was injected with 0.1 mL of 4 % Evans blue
(Sigma) in saline solution (i.v.) in the tail.Then, 10 min after the
i.v.injection of the dye solution, 0.4 mL of 0.5 % (v/v) AcOH was
injected ip. After 20 min, the mice were killed by dislocation of
the neck, and the viscera were exposed and irrigated with dis-
tilled water, which was then poured into 10 mL volumetric flasks
through glass wool. Each flask was filled up to 10 mL with dis-
tilled water, 0.1 of 0.1 N NaOH solution was added to the flask,
and the absorption of the final solution was measured at
590 nm (Beckmann Dual Spectrometer; Beckmann, Beck-
mann Instruments Inc., Fullerton, California, USA). In the con-
trol animals a mixture of distilled water and 0.5 % CMC was
given orally, otherwise they were treated in the same manner as
described above.
[10] P. Renard, D. Lesieur, C. Lespagnol, Eur. J. Med. Chem.,
Chim. Ther. 1980, 15, 453–456.
[11] S. Ünlü, H. Erdogƒ an, R. Sunal., H.U. J. Faculty of Pharma-
cy 1992, 12, 23–31.
[12] G.Pilli, H.Erdogan, R.Sunal, Arzneim.-Forsch./Drug Res.
1993, 43, 1351–1354.
[13] S.Ünlü, H.Erdogƒ an, R.Sunal., B.Gümü¸sel, J.Fac.Pharm.
Gazi 1992, 9, 75–80.
[14] D. S. Dogruer, S. Ünlü, E.Yes¸ ilada, M. F. S¸ ahin, Farmaco
1997, 52, 745–750.
[15] D. S. Dogruer, S. Ünlü, M. F. S¸ ahin, E.Ye¸silada, Farmaco
1998, 53, 80–84.
[16] T. Önkol, S. Ito, E. Yildirim, K. Erol, M. F. S¸ ahin, Archiv.
Pharm. Pharm. Med. Chem. 2001, 334, 17–20.
[17] G. Mairesse, J. C. Boivin, D. J.Thomas, M. C. Bermann, J.
Acute toxicity
P. Bonte, D. Lesieur, Acta Cryst. 1984, C 40, 1019–1021.
Animals employed in the carrageenan-induced paw edema ex-
periment were observed during 24 h and mortality, if present,
was recorded for each group at the end of the observation
period.
[18] T. Takao, T. Hisashi, European Patent, 84-301068, 1984.
[19] P. J. Breinagel, G. Ridge, US Patent, 3,161,649, 1964.
[20] H. Aichaoui, I. Lesieur, J. P. Henichart, Synthesis 1990,
679–680.
Gastric-ulcerogenic effect
[21] V.M.Guran, U.K.Jagwany, J.Indian Chem.Soc.1979, 56,
After the analgesic activity experiment, mice were killed under
deep ether anesthesia and stomachs were removed.Then the
abdomen of each mouse was opened through great curvature
and examined under the dissecting microscope for lesions or
bleedings.
325–329.
[22] T. Sandmeyer, Chem. Ber. 1886, 19, 8656.
[23] R. J. Nachman, J. Heterocyclic Chem. 1985, 22, 279–280.
[24] A. Ostaszynski, H. Plenkiewicz, T. Urbanski, Bull. Acad.
Pol. Sci., Ser. Sci. Chim. 1971, 19, 15–20.
Statistical analysis of data
Data obtained from the animal experiments were expressed as
the mean standard error ( SEM). Statistical differences be-
tween the treatments and the control were tested by ANOVA
test. Data with p < 0.05 value was considered to be significant.
[25] R.Okun, Sc.Liddon, L.Lasagnal, J.Pharmacol.Exp.Ther.
1963, 139, 107–114.
[26] E.A.Seigmund, R.A.Cadmus, G.Lu, Proc.Soc.Exp.Biol.
1957, 95, 729–731.
[27] E. T. Eckhardt, F. Cheplovitz, M. Lipo, W. M. Govier, Proc.
Soc. Exp. Biol. 1958, 98, 186–191.
References
[28] L. C. Hendershot, J. Forsaith, J. Pharmacol. Exp. Ther.
1959, 125, 237–240.
[1] K. Kolasa, Z. Kleinrok, Acta Pol. Pharm. 1979, 36, 383–
391.
[29] B. A. Whittle, Brit. J. Pharmacol. 1964, 22, 246–253.
[2] D. Paskov, A. Bakurdzhiev, V. Kalcheva, D. Simov, L. Ka-
[30] Y. Kasahara, H. Hikino, S. Tsurufuji, M. Watanabe, K.
menova, K. H. Boicheva, Farmatsiya 1975, 25, 61–64.
Ohuchi, Planta Medica 1985, 51, 325–331.
[3] C. F. Turk, J. Krapcho, I. M. Michel, I. Weinryb, J. Med.
Chem. 1977, 20, 729–732.
[31] R. Vinegar, W, Schreiber, R. Hugo, J. Pharmacol. Exp.
Ther. 1969, 166, 96–103.
[4] J. A. Orcutt, J. B. Prytherch, M. Konikov, S. M. Michaelson,
[32] P. Patrignani, M. R. Panara, A. Greco, O. Fusco, C. Natoli,
S. Iacobelli, F. Chipollone, A. Ganci, C. Creminon, J. Mac-
louf, C. Patrono, J. Pharmacol. Exp. Ther. 1994, 271,
1705–1712.
Arch. Intern. Pharmacodyn. 1964, 152, 121–126.
[5] D. Lesieur, H. Aichaoui, C. Lespagnol, J. Bonnet, French
patent 89-04129 1989.
[6] Tacquet, C. Lespagnol, H. Beerens, D. Lesieur, B. De-
[33] E.Ye¸silada, S. Tanaka, E. Sezik, M. Tabata, J. Natr. Prod-
ucts, 1988, 51, 504–508.
vulder, Ann. Inst. Pasteur Lille 1971, 22, 189–200.