Synthesis of some newer derivatives of 2-amino benzoic acid as potent anti-inflammatory and analgesic agents

Article abstract of DOI:10.1016/S0968-0896(03)00529-7

Diazotization of N-benzylidene anthranilic acids 1a-1n at pH 9 yielded N-[α-(phenylazo) benzylidene] anthranilic acids 2a-2n and at pH 3 yielded N-benzylidene-5-(phenylazo) anthranilic acids 3a-3n. When compounds 3a-3n were treated with thioglycolic/thiolactic acid in the presence of anhydrous ZnCl ₂, 2-(4-oxo-2-phenylthiazolidin-3-yl)-5-(phenylazo) benzoic acids 4a-4n were afforded. The newly synthesized compounds were screened for their anti-inflammatory and analgesic activities and were compared with standard drugs, aspirin and phenylbutazone. Out of the compounds studied, the most active compound 4n showed more potent activity than the standard drugs at all doses tested.

Full text of DOI:10.1016/S0968-0896(03)00529-7

Bioorganic & Medicinal Chemistry 11 (2003) 5281–5291
Synthesis of Some Newer Derivatives of 2-Amino Benzoic Acid as
Potent Anti-inflammatory and Analgesic Agents
Ashok Kumar,* Deepti Bansal, Kiran Bajaj, Shalabh Sharma,
Archana and V. K. Srivastava
Medicinal Chemistry Division, Department of Pharmacology, L.L.R.M. Medical College, Meerut (U.P) 250004, India
Received 17 March 2003; accepted 28 July 2003
Abstract—Diazotization of N-benzylidene anthranilic acids 1a–1n at pH 9 yielded N-[a-(phenylazo) benzylidene] anthranilic acids
2a–2n and at pH 3 yielded N-benzylidene-5-(phenylazo) anthranilic acids 3a–3n. When compounds 3a–3n were treated with thio-
glycolic/thiolactic acid in the presence of anhydrous ZnCl2, 2-(4-oxo-2-phenylthiazolidin-3-yl)-5-(phenylazo) benzoic acids 4a–4n
were afforded. The newly synthesized compounds were screened for their anti-inflammatory and analgesic activities and were
compared with standard drugs, aspirin and phenylbutazone. Out of the compounds studied, the most active compound 4n showed
more potent activity than the standard drugs at all doses tested.
# 2003 Elsevier Ltd. All rights reserved.
Introduction
Chemistry
Anthranilic acids (2-aminobenzoic acids) constitute an
important group of non-steriodal antiinflammatory
agents. Some of these anti-inflammatory derivatives
were synthesized by several scientists of the world and
have been reported as potent anti-inflammatory
agents.1ꢀ5 Anthranilic acid, an aromatic nucleus, has
gained prominence after the discovery of mefenamic
acid and meclofenamate,6 which are currently useful
drugs for the treatment of various inflammatory dis-
orders. Heterocyclic/aliphatic functionalized systematic
variation at the 2-position of anthranilic acid nucleus
remarkably increases the anti-inflammatory activity.
Formazan7 and thiazolidine8 derivatives are also well
known for their pronounced anti-inflammatory activity.
In view of these observations, it was thought worthwhile
to synthesize some newer more potent derivatives of
anthranilic acid by incorporating formazanyl and thi-
azolidinyl moieties at the 2-position of anthranilic acid.
The structure of these newly synthesised compounds
was confirmed by elemental and spectral analysis. Then,
these compounds were subjected to screens for their
anti-inflammatory, analgesic, antipyretic, cyclo-oxygen-
ase inhibition activity, and acute toxicity.
The desired anthranilic acid (2-aminobenzoic acid)
derivatives, that is 1a–4n were synthesized as shown in
Scheme 1. The reaction of different substituted benz-
aldehydes and 2-aminobenzoic acid yielded N-benzyli-
dene anthranilic acids 1a–1n. Diazotization of 1a–1n at
pH adjusted to 9 afforded N-[a-(phenylazo) benzyli-
dene] anthranilic acids 2a–2n. When diazonium salt
solution was added to 1a–1n at pH adjusted to 3 with
constant stirring on mechanical stirrer for 72 h below
0 ꢁC gave N-benzylidene-5-(phenylazo) anthranilic acids
3a–3n. 2-(4-Oxo-2-phenylthiazolidin-3-yl)-5-(phenylazo)
benzoic acids 4a–4n were synthesized by refluxing 3a–3n
with thioglycolic/thiolactic acid in the presence of
anhydrous ZnCl2.
Results and Discussion
Anti-inflammatory, analgesic, ulcerogenic, cyclooxy-
genase activities, and acute toxicity of these new
anthranilic acid derivatives 1a–1n, 2a–2n, 3a–3n and 4a–
4n are represented in Table 1.
Anti-inflammatory and analgesic activity
The characteristic feature of the first stage compounds is
the presence of carbon–nitrogen double bond between
two substitued phenyl rings. All the fourteen N-benzyl-
*Corresponding author. E-mail: rajput_ak@indiatimes.com
0968-0896/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/S0968-0896(03)00529-7

5282
A. Kumar et al. / Bioorg. Med. Chem. 11 (2003) 5281–5291
Scheme 1.
idene anthranilic acids have shown moderate degree
(17.3–32.7%) of anti-inflammatory activity while these
compounds unexpectedly showed mild degree of
analgesic activity. Out of the 14 N-benzylidene anthra-
nilic acids, compound 1n which was substituted with
NO2 group at the p-position of phenyl ring elicited
maximum inhibition of oedema at a dose of 50 mg/kg
po. Being the most active, it was tested at three graded
doses (25, 50 and 100 mg/kg po) and was also compared
with standard drugs, phenylbutazone and aspirin. The
compound showed more potent activity than phenylbu-
tazone and aspirin. However, this compound exhibited
very low analgesic activity (16%). Interestingly, com-
pounds 1a, 1c, 1g, 1j and 1k which were substituted with
chloro atom on positions 2 and 4, fluoro atom on posi-
tion 2, chloro group on position 4, bromo atom on
position 4 and bromo atom on position 2 of phenyl ring,
respectively, showed potent anti-inflammatory activity.
On the contrary, when phenyl group is substituted with
chloro atom on position 3 and ethyl group on position 4
showed lesser degree of activity (i.e., 17.4% in both the
compounds).
The diazotisation of the N-benzylidene anthranilic acids
at pH 9 converts them into the corresponding N-[a-
(phenylazo) benzylidene] anthranilic acids 2a–2n which

Table 1. Anti-inflammatory, analgesic, ulcerogenic, cyclooxygenase, and toxicity data of compounds 1a–4n (Scheme 1)
Compd
R
R’
X
Dose
(mg/kg po)
Anti-inflammatory
activity
Dose
(mg/kg po)
Analgesic
activity
Dose
(mg/kg po)
Ulcerogenic activity
Cyclo-oxygenase
activity assay
ED50
(mg/kg po)
ALD50
(mg/kg po)
% oedema
inhibition
relative to
control
% decrease of
writhes in 25 min
after treatment
relative to control
% of
animal with
hyperemia
% of
animal
with ulcer
Inhibitory action
of some selected
compound%
inhibition 10 m M
1a
2,4-Cl2
—
—
25
50
100
50
25
50
100
50
50
50
50
50
50
50
50
50
50
25
50
100
25
50
100
50
25
50
100
50
25
50
100
25
50
100
50
50
25
50
100
25
50
100
50
50
50
29
50
100
50
50
14.8
28.6
54.6
17.3
15.3
29.3
61.3
19.0
21.1
20.6
28.6
12.3
30.6
24.7
25.1
18.4
17.4
16.4
32.7
62.6
14.2
22.2
42.4
24.6
12.8
27.4
52.4
21.2
13.7
22.8
56.8
10.8
21.7
44.8
21.3
20.8
19.6
28.6
62.3
14.5
23.5
46.5
24.9
18.2
15.3
19.7
29.7
59.7
23.5
19.2
25
50
100
50
25
50
100
50
50
50
50
50
50
50
50
50
50
25
50
100
25
50
100
50
25
50
100
50
25
50
100
25
50
100
50
50
25
50
100
25
50
100
50
50
50
25
50
100
50
50
9.0
14.0
30.0
30
10
15
125
250
500
250
125
250
500
250
250
250
250
250
250
250
250
250
250
125
250
500
125
250
500
250
125
250
500
250
125
250
500
125
250
500
250
250
125
250
500
125
250
500
250
250
250
125
250
500
250
250
40
60
100
50
30
70
100
60
90
10
20
30
100
90
80
70
100
50
70
90
30
80
100
70
20
30
50
10
20
60
80
20
50
70
60
70
70
90
100
20
40
60
30
60
50
30
60
90
100
50
10
20
40
10
10
20
30
20
30
60
10
20
30
10
20
30
10
5
10
15
10
20
40
20
15
30
45
40
15
30
60
20
40
90
50
30
10
20
40
40
60
80
40
70
80
5
20
79.4
79.4
>1000
>1000
1b
1c
3-Cl
2-F
—
—
—
—
ni
30
30
1d
1e
1f
1g
1h
1I
1j
1k
1l
1m
1n
2-CH3
2-OCH3
4-OCH3
4-Cl
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
12.0
10.1
9.9
15.0
7.5
16.1
12.3
15.2
9.2
ni
ni
ni
ni
ni
ni
ni
ni
ni
ni
70
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
H
2,6-Cl2
4-Br
2-Br
N-(CH3)2
4-C2H5
4-NO2
8.5
10.0
16.0
40.0
3.0
79.4
2a
2, 4-Cl2
3-Cl
—
30
128.8
>1000
9.8
20.2
11.9
10.2
15.3
30.6
16.2
5.0
10.1
20.1
6.7
2b
2c
3-Cl
2-F
4-Cl
H
—
—
ni
20
>1000
>1000
79.4
2d
2e
2-CH3
2-OCH3
3-Cl
4-Cl
—
—
ni
20
>1000
>800
79.4
2f
4-OCH3
3-Cl
—
20
125.9
>1000
9.7
20.1
10.3
8.9
2g
2h
2I
4-Cl
H
2,6-Cl2
H
4-Cl
H
—
—
—
ni
ni
40
>1000
>1000
>1000
9.7
63.1
14.3
32.6
10.6
12.6
24.6
12.2
9.0
7.3
9.6
15.3
30.0
13.0
10.3
2j
4-Br
4-OCH3
—
ni
177.8
>1000
2k
2l
2m
2n
2-Br
N-(CH3)2
4-C2H5
4-NO2
2-OCH3
H
3-Cl
4-Cl
—
—
—
—
ni
ni
>1000
>1000
70
79.4
>1000
10
20
40
60
3a
3b
2, 4-Cl2
3-Cl
3-Cl
4-Cl
—
—
ni
ni
>1000
>1000
(Continued on next page.)

Table 1 (continued)
Compd
R
R’
X
Dose
(mg/kg po)
Anti-inflammatory
activity
Dose
(mg/kg po)
Analgesic
activity
Dose
(mg/kg po)
Ulcerogenic activity
Cyclo-oxygenase
activity assay
ED50
(mg/kg po)
ALD50
(mg/kg po)
% oedema
inhibition
relative to
control
% decrease of
writhes in 25 min
after treatment
relative to control
% of
animal with
hyperemia
% of
animal
with ulcer
Inhibitory action
of some selected
compound%
inhibition 10 m M
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
2-F
2-CH3
2-OCH3
4-OCH3
4-Cl
H
3-Cl
4-Cl
3-Cl
H
4-Cl
H
4-OCH3
2-OCH3
H
—
—
—
—
—
—
—
—
—
—
—
—
50
50
50
50
50
50
50
50
50
50
50
25
50
100
50
50
50
50
25
50
100
25
50
100
50
50
25
50
100
50
50
50
50
25
50
100
25
50
100
25
50
100
25
50
100
24.5
17.2
15.7
14.6
12.2
22.9
21.7
19.9
10.1
9.2
50
50
50
50
50
50
50
50
50
50
50
25
50
100
50
50
50
50
25
50
100
25
50
100
50
50
25
50
100
50
50
50
50
25
50
100
25
50
100
25
50
12.0
8.0
7.0
6.8
250
250
250
250
250
250
250
250
250
250
250
125
250
500
250
250
250
250
125
250
500
125
250
500
250
250
125
250
500
250
250
250
250
125
250
500
125
250
500
125
250
500
125
250
500
100
40
30
60
20
80
70
50
40
60
100
30
30
30
60
60
60
50
80
20
30
60
80
5
10
20
40
50
50
40
10
30
50
10
20
30
20
10
20
30
40
10
30
40
4
ni
ni
ni
ni
ni
ni
ni
ni
ni
ni
ni
ni
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
>1000
6.2
H
11.3
10.6
9.7
5.0
8.6
2, 6-Cl2
4-Br
2-Br
N-(CH3)2
4-C2H5
4-NO2
3m
3n
3-Cl
4-Cl
26.8
28.6
33.6
66.7
32.2
19.6
31.5
23.2
16.3
28.2
56.0
17.2
29.1
54.2
19.2
36.5
21.8
44.8
76.8
22.8
28.7
31.6
22.8
29.1
47.2
93.2
30.42
34.4
56.4
29.5
34.4
58.8
—
9.2
7.5
90.3
13.2
28.2
16.0
11.2
15.3
11.10
6.0
12.0
25.0
8
15
45
10.10
18.2
14.1
70
100
100
60
90
60
20
60
100
30
70
100
80
100
50
70
100
100
100
90
100
30
60
90
30
60
4a
4b
4c
4d
4e
2, 4-Cl2
3-Cl
2-F
2-CH3
2-OCH3
3-Cl
4-Cl
H
3-Cl
4-Cl
H
H
H
H
H
ni
ni
ni
ni
ni
>1000
>1000
>1000
>1000
>1000
79.4
79.4
4f
4-OCH3
3-Cl
H
ni
>1000
4g
4h
4i
4-Cl
H
2, 6-Cl2
H
4-Cl
H
H
CH3
CH3
ni
ni
90
>1000
>1000
>1000
61.6
4j
4k
4l
4m
4n
4-Br
2-Br
N-(CH3)2
4-C2H5
4-NO2
4-OCH3
2-OCH3
H
3-Cl
4-Cl
CH3
CH3
CH3
CH3
CH3
13.4
14.3
15.0
22.2
10.0
21.3
40.4
15.25
28.36
39.43
28.36
43.30
54.30
—
ni
ni
ni
ni
ni
>1000
>1000
>1000
>1000
>1000
10
20
11
30
60
90
80
90
90
—
50.1
—
Phenylbutazone
Aspirin
—
—
—
—
—
—
—
—
—
90
100
ni
—
—
—
90
30
60
90
99.6
—
100
25
50
100
Control
—
ni, no inhibition.

A. Kumar et al. / Bioorg. Med. Chem. 11 (2003) 5281–5291
5285
showed more or less similar degree of anti-inflamma-
tory, analgesic and other activities, while the diazotisa-
tion at 5-position of 1a–1n at pH 3 gave N-benzylidene-
5-(phenylazo)anthranilic acids 3a–3n. These com-
pounds, however, exhibited varying degree of anti-
inflammatory (9.2–33.6%) and analgesic activity (5–
13.2%).
diazotisation at pH 9. Furthermore, incorporation of
thiazolidine ring in compounds 3a–3n was found to
increase the potency of these compounds and resulted
into the formation of 4a–4n with high anti-inflamma-
tory and analgesic activity with low ulcerogenic activity.
These compounds were found to have no suppersive
effect on cyclooxygenase which is the prime mechanism
of anti-inflammatory activity of anthranilic acid deriva-
tives. Since the analgesic, anti-inflammatory and
ulcerogenic activities of 4n corresponds mostly to the
activity shown by 1n which has got 70% inhibition of
cyclooxygenase activity (Table 1). It seems likely that 4n
which is devoid of cyclooxygenase inhibition is con-
verted to 1n in the body thereby exhibiting the anti-
inflammatory, analgesic and ulcerogenic activities
through inhibition of cyclooxygenase by 1n. It is quiet
likely that the other anthranilic acid derivatives which
do not show cylooxygenase inhibition may be acting for
anti-inflammatory, analgesic and ulcerogenic activities
by their conversion to metabolites which possess cyclo-
oxygenase inhibition activity.
Furthermore, cyclocondensation of 3a–3n with thiogly-
colic/thiolactic acids resulted into the corresponding
five-membered ring compounds, 4a–4n, which have
shown more potent activities than their parent com-
pounds 3a–3n. The most active compound of 4a–4n is
compound 4n. This compound 4n along with other
promising compounds 4e and 4i were studied in details
at three graded doses and have shown dose-dependent
activity. Anti-inflammatory activity of potent com-
pounds 4e, 4i and 4n and their comparison with stan-
dard drugs, aspirin and phenylbutazone, is given in
Scheme 1.
Ulcerogenic activity
Compounds 1a–1n showed significant lesser degree (10–
40%) of ulcer production activity. Moreover, the most
active compound 1n showed lesser ulcerogenic activity
than standard drugs, acetylsalicyclic and phenylbuta-
zone. Futhermore, compounds 2a–3n also exihibited
low ulcer production activity ranging between 10 and
50% at 250 mg/kg po. However, compounds 4e, 4i and
4n showed very low degree of ulcerogenic activity.
Experimental
Chemistry
Melting points were determined in open capillaries with
the help of thermonic melting point apparatus and are
uncorrected. IR spectra (KBr/Nujol) were recorded on
1
Backmann Acculab-10-FTIR spectrometers. H NM R
spectra were recorded by Bruker WM400 FT MHz
instrument using CDCl3 or DMSO-d6 as solvent and
tetramethylsilane (TMS) as internal reference standard.
All chemical shifts (d) are in ppm. The purity of the
compounds was checked by thin layer chromatography
(TLC) on silica gel-G plates of 0.5 mm thickness. The
elemental analysis (C, H, N) of the compounds were
performed on Heracus Carlo Erba 1108 analyser.
Cyclooxygenase assay activity
None of the N-benzylidene anthranilic acids inhibited
the cyclooxygenase activitiy. They, therefore, seems to
act through some other mechanism rather than inhibit-
ing prostaglandin synthesis. Cyclooxygenase activity of
most of the compounds 2a–3n did not show any activity
suggesting that they might be acting through some other
mechanism of their anti-inflammatory action, except
compounds 2e, 2f, 2n, 3n and 4i which showed good
cyclooxygenase activity indicating that these com-
pounds reduces inflammatory response by inhibition of
prostaglandins.
N-(p-Nitrobenzylidene)anthranilic acid (1n). A mixture
of p-nitrobenzaldehyde (0.01 mol), anthranilic acid (0.01
mol) and anhydrous ZnCl2 heated on free flame for 10
min. The reaction mixture was cooled, cocentrated and
recrystallised from DMF and water to give 1n (70%):
mp 213 ꢁC; IR (Nujol) 1610 (C¼N), 3060 (C–C aro-
matic), 1720 (C¼O), 3010 (O–H) cmꢀ1
;
1H NM R
ALD50 studies
(CDCl3) d 8.70 (1H, s), 12.40 (1H, s), 8.55–7.90 (8H, m)
(ppm); ms: m/z (6.7%) M+ 270. Anal. calcd for
C14H10N2O4; C, 62.22; H, 3.70; N, 10.37. Found: C,
62.51; H, 3.41; N, 10.00.
The toxicity study of these compounds indicate their
good safety margin.
N-(2,4-Dichlorobenzylidene)anthranilic acid (1a). 90%,
mp 176 ꢁC (ethanol); IR (Nujol) 1600 (C¼N), 3065 (C–
Conclusion
1
From the results of the biological activities, we may
conclude that diazotisation of 1a–1n is fruitful as it was
found to increase their anti-inflammatory and analgesic
activities with decreased ulcer production activity.
Moreover, diazotisation of 1a–1n at pH 3 was found to
be more beneficial then their diazotization at pH 9, as
compounds 3a–3n which are formed by the diazotisa-
tion at pH 3 were found to be comparatively more
potent than compounds 2a–2n which are formed by
C aromatic), 1715 (C¼O), 3000 (O–H) cmꢀ1; H NM R
(CDCl3) d 8.68 (1H, s), 12.42 (1H, s), 8.45–7.85 (7H, m)
(ppm): ms: m/z (6.57%) M+ 294. Anal. calcd for
C14H9NO2Cl2: C, 57.14; H, 3.06 ;N, 4.76. Found: C,
57.44; H, 3.16; N, 4.80.
N-(m-Chlorobenzylidene)anthranilic acid (1b). 80%, mp
158 ꢁC (methanol); IR (Nujol) 1615 (C¼N), 3068 (C–C
1
of aromatic C–H), 1717 (C¼O), 3012 (O–H) cmꢀ1; H

5286
A. Kumar et al. / Bioorg. Med. Chem. 11 (2003) 5281–5291
1
NMR (CDCl3) d 8.70 (1H, s), 12.44 (1H, s), 8.40–7.80
(7H, m) (ppm): ms: m/z (7.5%) M+ 259.5. Anal. calcd
for C14H10NO2Cl: C, 64.73; H, 3.85; N, 5.39; Found: C,
64.75; H, 3.76; N, 5.40.
of aromatic C–H), 1714 (C¼O), 3015 (O–H) cmꢀ1; H
NMR (CDCl3) d 8.61 (1H, s), 12.43 (1H, s), 8.30–7.72
(8H, m) (ppm): ms: m/z (5.67%) M+ 304. Anal. calcd
for C14H10NO2Br: C, 55.26; H, 3.28; N, 4.60; Found: C,
55.23; H, 3.33; N, 4.68.
N-(o-Fluorobenzylidene)anthranilic acid (1c). 90%, mp
223 ꢁC (DMF/water); IR (Nujol) 1612 (C¼N), 3065 (C–
N-(o-Bromobenzylidene)anthranilic acid (1k). 80%, mp
192 ꢁC (ethanol/water); IR (Nujol) 1618 (C¼N), 3061
1H NMR (CDCl3) d 8.60 (1H, s), 12.42 (1H, s), 8.30–
7.70 (8H, m) (ppm): ms: m/z (4.30%) M+ 304. Anal.
calcd for C14H10NO2Br: C, 55.26; H, 3.28; N, 4.60;
Found: C, 55.30; H, 3.39; N, 4.23.
C of aromatic), 1716 (C¼O), 3010 (O–H) cmꢀ1
;
1H
NMR (CDCl3) d 8.68 (1H, s), 12.41 (1H, s), 8.35–7.75
(7H, m) (ppm): ms: m/z (8.50%) M+ 243. Anal. calcd
for C14H10NO2F: C, 69.13; H, 4.11; N, 5.76; Found: C,
69.23; H, 3.99; N, 5.81.
(C-C of aromatic C–H), 1715 (C¼O), 3012 (O–H) cmꢀ1
;
N-(o-Methylbenzylidene)anthranilic acid (1d). 70%, mp
186 ꢁC (ethanol); IR (Nujol) 1610 (C¼N), 3066 (C–C of
N - (p - N,N - Dimethylbenzylidene)anthranilic acid (1l).
90%, mp 178 ꢁC (ethanol/water); IR (Nujol) 1620
(C¼N), 3068 (C–C of aromatic C–H), 3015 (O–H)
1
aromatic), 1717 (C¼O), 3014 (O–H) cmꢀ1; H NM R
(CDCl3) d 2.30 (3H, s), 8.66 (1H, s), 12.42 (1H, s), 8.38–
7.76 (7H, m) (ppm): ms: m/z (9.69%) M+ 239. Anal.
calcd for C15H13NO2: C, 75.31; H, 5.43; N, 5.85; Found:
C, 75.33; H, 5.95; N, 5.90.
1
cmꢀ1; H NMR (CDCl3): d 1.25 (6H, s), 8.68 (1H, s),
12.45 (1H, s), 8.35–7.80 (8H, m) (ppm): ms: m/z (5.01%)
M
+ 268. Anal. calcd for C16H16N2O2: C, 71.64; H, 5.97;
N, 10.44; Found: C, 71.90; H, 5.66; N, 10.61.
N - (o - Methoxybenzylidene)anthranilic acid (1e). 90%,
mp 177 ꢁC, (ethanol/water); IR (Nujol) 1611 (C¼N),
3064 (C–C of aromatic), 1715 (C¼O), 3011 (O–H)
N-(p-Ethylbenzylidene)anthranilic acid (1m). 80%, mp
155 ꢁC (ethanol); IR (Nujol) 1615 (C¼N), 3066 (C–C of
1
1
cmꢀ1; H NMR (CDCl3) d 3.39 (3H, s), 8.64 (1H, s),
aromatic C–H), 3015 (O–H) cmꢀ1; H NMR (CDCl3) d
12.40 (1H, s), 8.40–7.70 (8H, m) (ppm): ms: m/z (9.01%)
M
8.66 (1H, s), 12.46 (1H, s), 8.38–7.75 (8H, m) (ppm): ms:
m/z (6.32%) M+ 253. Anal. calcd for C16H15NO2: C,
75.88; H, 5.92; N, 5.53; Found: C, 76.13; H, 5.63; N,
5.51.
+ 255. Anal. calcd for C15H13NO3: C, 70.61; H, 5.19;
N, 5.49; Found: C, 70.58; H, 5.09; N, 5.40.
N-(p-Methoxybenzylidene)anthranilic acid (1f). 80%, mp
104 ꢁC, (benzene); IR (Nujol) 1612 (C¼N), 3062 (C–C
N-[ꢀ-(p-Chlorophenylazo)-p-nitrobenzylidene] anthranilic
acid (2n). Concd HCl (3 mL) was added to a solution
of p-chloroaniline (0.02 mol) in glacial acetic acid (5
mL). A solution of sodium nitrite (1 gm in 5 mL of
water) was then added drop by drop in this solution.
The diazonium salt solution of N-(p-nitrobenzylidene)
anthranilic acid (0.02 mol) was added dropwise with
constant stirring in methanol (50–70 mL) at below 0 ꢁC.
The reaction mixture was kept at room temperature for
3–6 days and then poured into cold water. The solid
thus obtained was washed with water and recrystallised
from methanol to afford 2n. 70%, mp 260 ꢁC (metha-
nol); IR (Nujol) 1600 (C¼N), 3090 (C–C of aromatic C–
1
of aromatic C–H), 1711 (C¼O), 3010 (O–H)cmꢀ1; H
NMR (CDCl3) d 3.40 (3H, s), 8.60 (1H, s), 12.43 (1H,
s), 8.38–7.65 (8H, m) (ppm): ms: m/z (8.60%) M+ 255.
Anal. calcd for C15H13NO3: C, 70.58; H, 5.09; N, 5.49;
Found: C, 70.63; H, 5.23; N, 5.33.
N-(p-Chlorobenzylidene)anthranilic acid (1g). 60%, mp
196 ꢁC (methanol); IR (Nujol) 1608 (C¼N), 3060 (C–C
1
of aromatic C–H), 1710 (C¼O), 3015 (O–H) cmꢀ1; H
NMR (CDCl3) d 8.62 (1H, s), 12.41 (1H, s), 8.35–7.70
(8H, m) (ppm): ms: m/z (8.95%) M+ 259.5. Anal. calcd
for C14H10NO2Cl: C, 64.73; H, 3.85; N, 5.39; Found: C,
64.77; H, 3.99; N, 5.17.
H), 1720 (C¼O), 3010 (OH), 1430 (N¼N) cmꢀ1
;
1H
NMR (CDCl3+DMSO-d6) d 12.45 (1H, s), 8.60–7.00
(12H, m) (ppm): ms: m/z (4.35%) M+ 408.5. Anal.
calcd for C20H13N4O4Cl: C, 58.75; H, 3.18; N, 13.70;
Found: C, 58.63; H, 3.26; N, 13.86.
N-benzylidene anthranilic acid (1h). 70%, mp 201 ꢁC
(ethanol); IR (Nujol) 1600 (C¼N), 3015 (C–C of aro-
1
matic C–H), 1710 (C¼O), 3015 (O–H) cmꢀ1; H NM R
(CDCl3) d 8.60 (1H, s), 12.38 (1H, s), 8.30–7.70 (9H, m)
(ppm): ms: m/z (9.22%) M+ 225. Anal. calcd for
C14H11NO2: C, 74.46; H, 4.88; N, 6.22; Found: C,
74.76; H, 4.90; N, 6.31.
N-[ꢀ-(m-Chlorophenylazo)-2,4-dichlorobenzylidene] an-
thranilic acid (2a). 70%, mp 223 ꢁC (methanol); IR
(Nujol) 1610 (C¼N), 3092 (C–C of aromatic C–H),
1
1723 (C¼O), 3008 (OH), 1432 (N¼N) cmꢀ1; H NM R
N - (2,6 - Dichlorobenzylidene)anthranilic acid (1i). 90%,
mp 203 ꢁC (ethanol); IR (Nujol) 1612 (C¼N), 3061 (C–
C of aromatic C–H), 1712 (C¼O), 3016 (O–H) cmꢀ1; 1H
NMR (CDCl3) d 8.60 (1H, s), 12.43 (1H, s), 8.33–7.73
(7H, m) (ppm): ms: m/z (8.76%) M+ 294. Anal. calcd
for C14H9NO2Cl2: C, 57.14; H, 3.06; N, 4.76. Found: C,
57.43; H, 3.16; N, 4.77.
(CDCl3+DMSO-d6) d 12.44 (1H, s), 8.62–7.05 (12H,
m) (ppm): ms: m/z (3.67%) M+ 432.5. Anal. calcd for
C20H12N3O2Cl3: C, 55.49; H, 2.77; N, 9.71; Found: C,
55.50; H, 3.01; N, 10.03.
N-[m-Chloro-ꢀ-(p-Chlorophenylazo) benzylidene] anthra-
nilic acid (2b). 50%, mp 240 ꢁC (benzene); IR (Nujol)
1602 (C¼N), 3095 (C–C of aromatic C–H), 1722
N-(p-Bromobenzylidene)anthranilic acid (1j). 70%, mp
198 ꢁC (methanol); IR (Nujol) 1615 (C¼N), 3063 (C–C
(C¼O), 3013 (OH), 1432 (N¼N) cmꢀ1
;
1H NM R
(CDCl3+DMSO-d6) d 12.44 (1H, s), 8.62–7.00 (12H,

A. Kumar et al. / Bioorg. Med. Chem. 11 (2003) 5281–5291
5287
m) (ppm): ms: m/z (4.12%) M+ 398. Anal. calcd for
C20H13N3O2Cl2: C, 60.30; H, 3.26; N, 10.55; Found: C,
60.41; H, 3.31; N, 10.60.
ms: m/z (4.95%) M+ 398. Anal. calcd for
C20H13N3O2Cl2: C, 60.30; H, 3.26; N, 10.55; Found: C,
60.45; H, 3.25; N, 10.57.
N-[o-Fluoro-ꢀ-(phenylazo) benzylidene] anthranilic acid
(2c). 60%, mp 246 ꢁC (ethanol); IR (Nujol) 1605
(C¼N), 3096 (C–C of aromatic C–H), 1725 (C¼O),
N-[o-Bromo-ꢀ-(p-methoxyphenylazo)benzylidene]anthra-
nilic acid (2j). 70%, mp 229 ꢁC (benzene); IR (Nujol)
1608 (C¼N), 3094 (C–C of aromatic C–H), 1724
3015 (OH), 1434 (N¼N) cmꢀ1
;
1H NM
R
(C¼O), 3020 (OH), 1442 (N¼N) cmꢀ1
;
1H NM R
(CDCl3+DMSO-d6) d 12.43 (1H, s), 8.60–7.05 (12H,
m) (ppm): ms: m/z (5.95%) M+ 347. Anal. calcd for
C20H14N3O2F: C, 69.16; H, 4.03; N, 12.10; Found: C,
69.21; H, 4.13; N, 12.19.
(CDCl3+DMSO-d6) d 3.39 (3H, s) 12.40 (1H, s), 8.72–
7.10 (12H, m) (ppm): ms: m/z (7.01%) M+ 438. Anal.
calcd for C21H16N3O3Br: C, 57.53; H, 3.65; N, 9.58;
Found: C, 57.13; H, 3.70; N, 9.66.
N-[ꢀ-(m-Chlorophenylazo)-o-methoxybenzylidene] an-
thranilic acid (2d). 55%, mp 211 ꢁC (methanol); IR
(Nujol) 1608 (C¼N), 3097 (C–C of aromatic C–H),
N-[o-Bromo-ꢀ-(o-methoxyphenylazo)benzylidene]anthra-
nilic acid (2k). 70%, mp 236 ꢁC (benzene/hexane); IR
(Nujol) 1606 (C¼N), 3092 (C–C of aromatic C–H),
1
1
1725 (C¼O), 3015 (OH), 1435 (N¼N) cmꢀ1; H NM R
1720 (C¼O), 3021 (OH), 1444 (N¼N) cmꢀ1; H NM R
(CDCl3+DMSO-d6) d 2.40 (3H, s),12.46 (1H, s), 8.62–
7.02 (12H, m) (ppm): ms: m/z (6.11%) M+ 377.5. Anal.
calcd for C21H16N3O2Cl: C, 66.75; H, 4.23; N, 11.12;
Found: C, 66.80; H, 4.26; N, 11.22.
(CDCl3+DMSO-d6) d 3.40 (3H, s) 12.42 (1H, s), 8.70-
7.08 (12H, m) (ppm): ms: m/z (6.89%) M+ 438. Anal.
calcd for C21H16N3O3Br: C, 57.53; H, 3.65; N, 9.58;
Found: C, 57.61; H, 3.55; N, 9.67.
N-[ꢀ-(Chlorophenylazo)-o-methoxybenzylidene] anthra-
nilic acid (2e). 45%, mp 233 ꢁC (DMF/water); IR
(Nujol) 1610 (C¼N), 3095 (C–C of aromatic C–H),
N-[p-N,N-Dimethyl-ꢀ-(phenylazo)benzylidene]anthranilic
acid (2l). 60%, mp 217 ꢁC (ethanol); IR (Nujol) 1606
(C¼N), 3092(C–C of aromatic C–H), 1722 (C¼O), 3020
1
1
1728 (C¼O), 3018 (OH), 1436 (N¼N) cmꢀ1; H NM R
(OH), 1440 (N¼N) cmꢀ1; H NMR (CDCl3+DMSO-
(CDCl3+DMSO-d6) d 3.45 (3H, m), 12.48 (1H, s),
8.68–7.10 (12H, m) (ppm): ms: m/z (5.87%) M+ 393.5.
Anal. calcd for C21H16N3O3Cl: C, 64.04; H, 4.06; N,
10.67; Found: C, 64.13; H, 4.16; N, 10.66.
d6) d 1.30 (6H, s), 12.38 (1H, s), 8.74–7.12 (12H, m)
(ppm): ms: m/z (5.25%) M+ 372. Anal. calcd for
C22H20N4O2: C, 70.96; H, 5.37; N, 15.05; Found: C,
71.20; H, 5.22; N, 15.19.
N-[ꢀ-(m-Chlorophenylazo)-p-methoxybenzylidene] an-
thranilic acid (2f). 40%, mp 217 ꢁC (benzene/hexane);
IR (Nujol) 1608 (C¼N), 3090 (C–C of aromatic C–H),
N-[p-Ethyl-ꢀ-(m-chlorophenylazo)benzylidene]anthranilic
acid (2m). 50%, mp 198 ꢁC (ethanol); IR (Nujol) 1605
(C¼N), 3090 (C–C of aromatic C–H), 1718 (C¼N),
1
1730 (C¼O), 3020 (OH), 1432 (N¼N) cmꢀ1; H NM R
3020 (OH), 1441 (N¼N) cmꢀ1
;
1H NMR (CDCl3+
(CDCl3+DMSO-d6) d 3.39 (3H, m), 12.50 (1H, s),
8.70–7.10 (12H, m) (ppm): ms: m/z (6.01%) M+ 393.5.
Anal. calcd for C21H16N3O3Cl: C, 64.04; H, 4.06; N,
10.67; Found: C, 64.00; H, 4.26; N, 10.59.
DMSO-d6) d 2.75 (2H,q), 2.05 (3H, t), 12.38 (1H, s),
8.67–7.05 (12H, m) (ppm): ms: m/z (4.87%) M+ 391.5.
Anal. calcd for C22H18N3O2Cl: C, 67.43; H, 4.59; N,
10.72; Found: C, 67.42; H, 4.60; N, 11.58.
N-[p-Chloro-ꢀ -(phenylazo)benzylidene]anthranilic acid
(2g). 60%, mp 222 ꢁC (methanol); IR (Nujol) 1605
(C¼N), 3098 (C–C of aromatic C–H), 1728 (C¼O),
5-(p-Chlorophenylazo)-N-(p-nitrobenzylidene)anthranilic
acid (3n). To a solution of N-(p-nitrobenzylidene)an-
thranilic acid (0.15 mol) in ethanol (abs. 50 mL) con-
taining glacial acetic acid; pH adjusted to 3, a
diazonium salt solution of p-chloroaniline was added
dropwise with constant vigorous stirring on mechanical
stirrer for 72 h below 0 ꢁC. The reaction mixture was
kept at room temperature for 24 h and the solvent was
removed and the solid thus obtained was recrystallized
from acetone to yield 3n (45%): mp 278 ꢁC; IR (Nujol)
1580 (C¼N), 3060 (C–C of aromatic C–H), 1700
3018 (OH), 1435 (N¼N) cmꢀ1
;
1H NMR (CDCl3+
DMSO-d6) d 12.44 (1H, s), 8.65–7.10 (12H, m) (ppm):
ms: m/z (5.54%) M+ 363.5. Anal. calcd for
C20H14N3O2Cl: C, 66.02; H, 3.85; N, 11.55; Found: C,
66.12; H, 3.87; N, 11.75.
N-[ꢀ - (p-Chlorophenylazo)benzylidene] anthranilic acid
(2h). 60%, mp 186 ꢁC (Ethanol) ; IR (Nujol) 1608
(C¼N), 3095 (C–C of aromatic C–H), 1726 (C¼O),
1
(C¼O), 3020 (OH), 1420 (N¼N), 765 (C–Cl) cmꢀ1; H
3020 (OH), 1438 (N¼N) cmꢀ1
;
1H NMR (CDCl3+
NMR (CDCl3) d 12.40 (1H, s), 8.80 (1H, s) 8.60–7.80
(11H, m) (ppm): ms: m/z (7.65%) M+ 409. Anal. calcd
for C20H13N4O4Cl: C, 58.75; H, 3.18; N, 13.70; Found:
C, 58.63; H, 3.22; N, 13.51.
DMSO-d6) d 12.45 (1H, s), 8.68–7.12 (12H, m) (ppm):
ms: m/z (5.17%) M+ 363.5. Anal. calcd for
C20H14N3O2Cl: C, 66.02; H, 3.85; N, 11.55; Found: C,
66.32; H, 3.40; N, 11.26.
5-(m-Chlorophenylazo)-N-(2,4-dichlorobenzylidene)an-
thranilic acid (3a). 50%, mp 246 ꢁC (ethanol); IR
(Nujol) 1578 (C¼N), 3059 (C–C of aromatic C–H),
N - [2,6 - Dichloro - ꢀ - (phenylazo)benzylidene]anthranilic
acid (2i). 60%, mp 254 ꢁC (ethanol); IR (Nujol) 1605
(C¼N), 3096 (C–C of aromatic C–H), 1727 (C¼O),
1708 (C¼O), 3018 (OH), 1422 (N¼N), 762 (C–Cl) cmꢀ1
;
3022 (OH), 1440 (N¼N) cmꢀ1
;
1H NMR (CDCl3+
1H NMR (CDCl3) d 12.36 (1H, s), 8.76 (1H, s), 8.62–
DMSO-d6) d 12.42 (1H, s), 8.70–7.08 (12H, m) (ppm):
7.84 (10H, m) (ppm): ms: m/z (4.66%) M+ 432.5. Anal.

5288
A. Kumar et al. / Bioorg. Med. Chem. 11 (2003) 5281–5291
calcd for C20H12N3O2Cl3: C, 55.49; H, 2.77; N, 9.71;
Found: C, 55.60; H, 2.63; N, 9.76.
C20H14N3O2Cl: C, 66.02; H, 3.85; N, 11.55; Found:
C,66.22; H, 3.70; N, 11.29.
N-(m-Chlorobenzylidene)-5-(p-chlorophenylazo)anthrani-
lic acid (3b). 30%, mp 266 ꢁC (benzene); IR (Nujol)
1582 (C¼N), 3056 (C–C of aromatic C–H), 1706
N - (2,6 - Dichlorobenzylidene) - 5 - (phenylazo)anthranilic
acid (3i). 50%, mp 274 ꢁC (hexane); IR (Nujol) 1580
(C¼N), 3060 (C–C of aromatic C–H), 1701 (C¼O), 3018
(OH), 1420 (N¼N), 752 (C–Cl) cmꢀ1; 1H NMR (CDCl3)
d 8.81 (1H, s), 8.60–7.82 (11H, m) (ppm): ms: m/z (6.67%)
M+ 398. Anal. calcd for C20H13N3O2Cl2: C, 60.30; H,
3.26; N, 10.55; Found: C, 60.31; H, 3.06; N, 10.65.
1
(C¼O), 3021 (OH), 1418 (N¼N), 760 (C–Cl) cmꢀ1; H
NMR (CDCl3) d 12.34 (1H, s), 8.77 (1H, s), 8.60–7.80
(11H, m) (ppm): ms: m/z (6.39%) M+ 398. Anal. calcd
for C20H13N3O2Cl2: C, 60.30; H, 3.26; N, 10.55; Found:
C,60.33; H, 2.99; N, 10.35.
N-(p-Bromobenzylidene)-5-(p-methoxyphenylazo)anthra-
nilic acid (3j). 60%, mp 284 ꢁC (ethanol); IR (Nujol)
1578 (C¼N), 3063 (C–C of aromatic C–H), 1698
N - (o - Fluorobenzylidene) -5-(phenylazo)anthranilic acid
(3c). 40%, mp 280 ꢁC (methanol); IR (Nujol) 1581
(C¼N), 3063 (C–C of aromatic C–H), 1769 (C¼O),
(C¼O), 3017 (OH), 1422 (N¼N) cmꢀ1
;
1H NM R
1
3016 (OH), 1425 (N¼N), 760 (C–Cl) cmꢀ1; H NM R
(CDCl3) d 3.40 (3H, s), 8.79 (1H, s), 8.58–7.80 (11H, m)
(ppm): ms: m/z (3.65%) M+ 438. Anal. calcd for
C21H16N3O3Br: C, 57.53; H, 3.65; N, 9.58; Found:
C,57.77; H, 3.80; N, 9.76.
(CDCl3) d 12.41 (1H, s), 8.78 (1H, s), 8.62–7.85 (11H,
m) (ppm): ms: m/z (7.03%) M+ 347. Anal. calcd for
C20H14N3O2F: C, 69.16; H, 4.03; N, 12.10; Found:
C,69.26; H, 3.95; N, 12.11.
N-(o-Bromobenzylidene)-5-(o-methoxyphenylazo)anthra-
nilic acid (3k). 60%, mp 263 ꢁC (ethanol); IR (Nujol)
1580 (C¼N), 3061 (C–C of aromatic C–H), 1699
5-(m-Chlorophenylazo)-N-(o-methylbenzylidene)anthrani-
lic acid (3d). 35%, mp 282C (DMF/water); IR (Nujol)
1580 (C¼N), 3060 (C–C of aromatic C–H), 1710
(C¼O), 3016 (OH), 1421 (N¼N) cmꢀ1
;
1H NM R
1
(C¼O), 3020 (OH), 1420 (N¼N), 756 (C–Cl) cmꢀ1; H
(CDCl3) d 3.42 (3H, s), 8.80 (1H, s), 8.60–7.80 (11H, m)
(ppm): ms: m/z (4.00%) M+ 438. Anal. calcd for
C21H16N3O3Br: C, 57.53; H, 3.65; N, 9.58; Found: C,
57.69; H, 3.80; N, 9.23.
NMR (CDCl3) d 2.36 (3H, s), 12.38 (1H, s), 8.82 (1H,
s), 8.60–7.82 (11H, m) (ppm): ms: m/z (6.79%) M+
377.5. Anal. calcd for C21H16N3O2Cl: C, 66.75; H, 4.23;
N, 11.12; Found: C,66.77; H, 4.46; N, 11.23.
N-(p-N,N-Dimethylbenzylidene)-5-(phenylazo)anthranilic
acid (3l). 40%, mp 260 ꢁC (ethanol); IR (Nujol) 1583
(C¼N), 3062 (C–C of aromatic C–H), 1700 (C¼O),
5-(p-Chlorophenylazo)-N-(o-methoxybenzylidene)anthra-
nilic acid (3e). 20%, mp 276 ꢁC (benzene/pet.ether); IR
(Nujol) 1585 (C¼N), 3061 (C–C of aromatic C–H),
1H NMR (CDCl3) d 3.45 (3H, s), 12.40 (1H, s), 8.86
(1H, s), 8.62–7.82 (11H, m) (ppm): ms: m/z (6.98%) M+
393.5. Anal. calcd for C21H16N3O3Cl: C, 64.04; H, 4.06;
N, 10.67; Found: C,64.08; H, 4.03; N, 11.00.
3018 (OH), 1425 (N¼N) cmꢀ1
;
1H NMR (CDCl3) d
1700 (C¼O), 3017 (OH), 1423 (N¼N), 758 (C–Cl) cmꢀ1
;
1.45 (6H, s) 8.81 (1H, s), 8.64–7.81 (12H, m) (ppm): ms:
m/z (5.35%) M+ 372. Anal. calcd for C22H20N4O2: C,
70.96; H, 5.37; N, 15.05; Found: C, 70.77; H, 5.40; N,
15.01.
5-(m-Chlorophenylazo)-N-(p-ethylbenzylidene)anthranilic
acid (3m). 30%, mp 266 ꢁC (DMF/water); IR (Nujol)
1584 (C¼N), 3065 (C–C of aromatic C–H), 1703
5-(m-Chlorophenylazo)-N-(p-methoxybenzylidene)anthra-
nilic acid (3f). 25%, mp 278 ꢁC (chloroform); IR (Nujol)
1581 (C¼N), 3058 (C–C of aromatic C–H), 1712
1
(C¼O), 3020 (OH), 1422 (N¼N), 760 (C–Cl) cmꢀ1; H
1
(C¼O), 3017 (OH), 1419 (N¼N), 757 (C–Cl) cmꢀ1; H
NMR (CDCl3) d 12.40 (5H, s), 8.80 (1H, s), 8.61–7.80
(11H, m) (ppm): ms: m/z (6.71%) M+ 391.5. Anal.
calcd for C22H18N3O2Cl: C, 67.43; H, 4.59; N, 10.72;
Found: C, 67.63; H, 4.71; N, 10.63.
NMR (CDCl3) d 3.43 (3H, s), 12.41 (1H, s), 8.85 (1H,
s), 8.61–7.85 (11H, m) (ppm): ms: m/z (7.11%) M+
393.5. Anal. calcd for C21H16N3O3Cl: C, 64.04; H, 4.06;
N, 10.67; Found: C,64.13; H, 4.11; N, 10.93.
5-(p-Chlorophenylazo)-2-[4-oxo-2-(p-nitrophenyl)thiazoli-
din-3-yl]benzoic acid (4n). To a solution of 5-(p-chloro-
phenylazo)-N-(p-nitrobenzylidine)anthranilic acid (0.01
mol) in benzene (dry, 50 mL) thiolactic acid/thioglycolic
acid (0.01 mol) was added separately and the reaction
mixture was refluxed for 48 h and the completion of the
reaction was monitored by TLC. The excess of solvent
was distilled off, and the product was recrystallised from
DMF/water to afford 4n. 50%; mp 196 ꢁC; IR (Nujol)
1510 (C–N), 3040 (C–C of aromatic C–H), 1700 (C¼O),
3000 (O–H), 1420 (N¼N), 760 (C–Cl), 1760 (C¼O of b-
N-(p-Chlorobenzylidene)-5-(phenylazo)-anthranilic acid
(3g). 40%, mp 263C (ethanol); IR (Nujol) 1583 (C¼N),
3060 (C–C of aromatic C–H), 1705 (C¼O), 3015 (OH),
1
1420 (N¼N), 759 (C–Cl) cmꢀ1; H NMR (CDCl3) d
12.42 (1H, s), 8.84 (1H, s), 8.60–7.84 (11H, s) (ppm): ms:
m/z (8.75%) M+ 263. Anal. calcd for C20H14N3O2Cl: C,
66.02; H, 3.85; N, 11.55; Found: C, 66.22; H, 3.90; N,
11.32.
5-(p-Chlorophenylazo)-N-(benzylidene)anthranilic acid
(3h). 30%, mp 268 ꢁC (methanol); IR (Nujol) 1582
(C¼N), 3061 (C–C of aromatic C–H), 1702 (C¼O),
1
thialactum) cmꢀ1, H NMR (CDCl3) d 12.40 (1H, s),
6.95 (1H, s), 8.65–7.20 (11H, m), 2.15 (3H, d), 5.95 (1H,
q) (ppm): ms: m/z (3.75%) M+ 469. Anal. calcd for
C23H17N4O5SCl: C, 55.59; H, 3.42; N, 11.28; Found: C,
55.53; H, 3.55; N, 11.53.
1
3017 (OH), 1421 (N¼N), 760 (C–Cl) cmꢀ1; H NM R
(CDCl3) d 12.40 (1H, s), 8.82 (1H, s), 8.62–7.86 (12H,
m) (ppm): ms: m/z (8.70%) M+ 263. Anal. calcd for

A. Kumar et al. / Bioorg. Med. Chem. 11 (2003) 5281–5291
5289
5-(p-Chlorophenylazo)-2-[4-oxo-2-(2,4-dichlorophenyl)-
thiazolidin-3-yl]benzoic acid (4a). 80%; mp 222 ꢁC
(C¼O), 3020 (OH), 1425 (N¼N), 763 (C–Cl), 1772
(C¼O of b-thialactum) cmꢀ1, 1H NMR (CDCl3) d 12.32
(1H, s), 6.92 (1H, s), 8.58–7.10 (12H, m), 3.63 (2H, s),
5.90 (1H, q) (ppm); ms: m/z (3.26%) M+ 437.5. Anal.
calcd for C22H16N3O3SCl: C, 60.34; H, 3.65; N, 9.60 ;
Found: C, 60.48; H, 3.71; N, 10.00.
...
(methanol); IR (Nujol) 1509 (C–N), 3042 (C C of aro-
matic C–H), 1701 (C¼O), 3010 (OH), 1420 (N¼N), 761
1
(C–Cl), 1762 (C¼N of b-thialactum) cmꢀ1, H NM R
(CDCl3) d 12.38 (1H, s), 6.96 (1H, s), 8.68–7.28 (10H,
m), 3.68 (2H, s) (ppm); ms: m/z (2.60%) M+ 506.5.
Anal. calcd for C22H14N3O3SCl3: C, 52.12; H, 2.76; N,
8.29; Found: C, 52.32; H, 2.90; N, 8.41.
5-(m-Chlorophenylazo)-2-[5-methyl-4-oxo-2-phenylthi-
azolidin-3-yl]benzoic acid (4h). 40% mp 195 ꢁC (metha-
...
nol); IR (Nujol) 1512 (C–N), 3044 (C C of aromatic
5-(p-Chlorophenylazo)-2-[2-(m-chlorophenyl)-4-oxo-thi-
azolidin-3-yl]benzoic acid (4b). 70% ; mp 210 ꢁC (etha-
C–H), 1702 (C¼O), 3002 (OH), 1422 (N¼N), 761 (C–
Cl), 1761 (C¼O of b-thialactum) cmꢀ1
,
1H NM R
...
nol); IR (Nujol) 1507 (C–N), 3041 (C C of aromatic
(CDCl3) d 12.41 (1H, s), 6.94 (1H, s), 8.68–7.24 (12H,
m), 2.14 (3H, d), 5.98 (1H, q) (ppm); ms: m/z (2.67%)
M
+ 451.5. Anal. calcd for C23H18N3O3SCl: C, 61.12; H,
3.98; N, 9.30 ; Found: C, 61.42; H, 3.71; N, 9.20.
C–H), 1705 (C¼O), 3012 (OH), 1425 (N¼N), 763 (C–
Cl), 1764 (C¼N of b-thialactum) cmꢀ1
,
1H NM R
(CDCl3) d 12.36 (1H, s), 6.92 (1H, s), 8.65–7.26 (11H,
m), 3.65 (2H, s) (ppm); ms: m/z (3.21%) M+ 472. Anal.
calcd for C22H15N3O3SCl2: C, 55.93; H, 3.17; N, 8.89;
Found: C, 55.71; H, 3.10; N, 9.01.
2-[2-(2,3-Dichlorophenyl)-5-methyl-4-oxo-thiazolidin-3-
yl]-5-(phenylazo)benzoic acid (4i). 50%; mp 178 ꢁC
(methanol); IR (Nujol) 1511 (C–N), 3043 (C–C of aro-
matic C–H), 1703 (C¼O), 3005 (OH), 1425 (N¼N), 762
2-[2-(o-Fluorophenyl)-4-oxo-thiazolidin-3-yl]-5-(phenyl-
azo)benzoic acid (4c). 60%; mp 264 ꢁC (benzene/pet.e-
ther); IR (Nujol) 1505 (C–N), 3040 (C–C of aromatic
C–H), 1702 (C¼N), 3015 (OH), 1426 (N¼N), 1768
(C¼O of b-thialactum) cmꢀ1, 1H NMR (CDCl3) d 12.36
(1H, s), 6.90 (1H, s), 8.68–7.30 (12H, m), 3.62 (2H, s)
(ppm); ms: m/z (2.95%) M+ 421. Anal. calcd for
C22H16N3O3SF: C, 62.70; H, 3.80; N, 9.97 ; Found: C,
62.83; H, 3.93; N, 10.02.
1
(C–Cl), 1760 (C¼O of b-thialactum) cmꢀ1, H NM R
(CDCl3) d 12.44 (1H, s), 6.92 (1H, s), 8.66–7.22 (11H,
m), 2.15 (3H, d), 5.95 (1H, q) (ppm); ms: m/z (2.30%)
M+ 487. Anal. calcd for C23H17N3O3SCl2: C, 56.79; H,
3.50; N, 8.64 ; Found: C, 56.73; H, 3.85; N, 8.56.
2-[2-(p-Bromophenyl)-5-methyl-4-oxo-thiazolidin-3-yl]-5-
(p-Methoxyphenylazo)benzoic acid (4j). 65%; mp 203 ꢁC
(ethanol); IR (Nujol) 1514 (C–N), 3042 (C–C of aro-
matic C–H), 1705 (C¼N), 3002 (OH), 1424 (N¼N),
1762 (C¼O of b-thialactum) cmꢀ1, 1H NMR (CDCl3) d
12.42 (1H, s), 6.95 (1H, s), 8.64–7.20 (11H, m), 3.41(3H,
s) 2.12 (3H, d), 5.96 (1H, q) (ppm); ms: m/z (1.67%)
M+ 526. Anal. calcd for C24H20N3O4SBr: C, 54.75; H,
3.80; N, 7.98 ; Found: C, 54.80; H, 3.72; N, 8.02.
5-(m-Chlorophenylazo)-2-[2-(o-methoxyphenyl)-4-oxo-
thiazolidin-3-yl]benzoic acid (4d). 50%; mp 256 ꢁC
...
(methanol); IR (Nujol) 1508 (C–N), 3045 (C C of aro-
matic C–H), 1705 (C¼O), 3018 (OH), 1428 (N¼N), 765
1
(C–Cl), 1770 (C¼O of b-thialactum) cmꢀ1, H NM R
(CDCl3) d 12.30 (1H, s), 6.95 (1H, s), 8.60–7.10 (11H,
m), 2.35 (3H, s), 3.60 (2H, s) (ppm); ms: m/z (4.26%)
M
+ 451.5. Anal. calcd for C23H18N3O3SCl: C, 61.12; H,
3.98; N, 9.30; Found: C, 61.28; H, 3.91; N, 9.56.
2-[2-(o-Bromophenyl)-5-methyl-4-oxo-thiazolidin-3-yl]-5-
(o-Methoxyphenylazo)benzoic acid (4k). 70%; mp
ꢁ
...
226 C (methanol); IR (Nujol) 1510 (C–N), 3041 (C C
5-(p-Chlorophenylazo)-2-[2-(o-methoxyphenyl)-4-oxo-
thiazolidin-3-yl]benzoic acid (4e). 60%; mp 223 ꢁC (etha-
of aromatic C–H), 1708 (C¼O), 3005 (OH), 1426
1
(N¼N), 1765 (C¼O of b-thialactum) cmꢀ1, H NM R
...
nol); IR (Nujol) 1510 (C–N), 3042 (C C of aromatic
(CDCl3) d 12.40 (1H, s), 6.90 (1H, s), 8.66–7.26 (11H,
m), 3,43 (3H, s) 2.14 (3H, d), 5.98 (1H, q) (ppm); ms: m/z
(2.01%) M+ 526. Anal. calcd for C24H20N3O4SBr: C,
54.75; H, 3.80; N, 7.98 ; Found: C, 54.94; H, 3.67; N,
7.70.
C–H), 1708 (C¼O), 3016 (OH), 1430 (N¼N), 760 (C–
Cl), 1772 (C¼O of b-thialactum) cmꢀ1
,
1H NM R
(CDCl3) d 12.32 (1H, s), 6.98 (1H, s), 8.62–7.20 (11H,
m), 3.41 (3H, s), 3.65 (2H, s) (ppm); ms: m/z (3.23%)
M
+ 467.5. Anal. calcd for C23H18N3O4SCl: C, 59.03; H,
3.85; N, 8.98; Found: C, 59.13; H, 3.45; N, 8.64.
2-[2-(p-N,N-Dimethylphenyl)-5-methyl-4-oxo-thiazolidin-
3-yl]-5-(phenylazo)benzoic acid (4l). 50%; mp 203 ꢁC
(ethanol); IR (Nujol) 1515 (C–N), 3045 (C–C of aro-
matic C–H), 1700 (C¼O), 3006 (OH), 1427 (N¼N),
1768 (C¼O of b-thialactum) cmꢀ1, 1H NMR (CDCl3) d
12.41 (1H, s), 6.88 (1H, s), 8.65–7.25 (12H, m), 1.48
(6H, s), 2.15 (3H, d), 5.96 (1H, q) (ppm); ms: m/z
(3.56%) M+ 447. Anal. calcd for C24H23N4O3S: C,
64.42; H, 5.12; N, 12.52; Found: C, 64.62; H, 5.13; N,
12.56.
5-(m-Chlorophenylazo)-2-[2-(p-methoxyphenyl)-4-oxo-
thiazolidin-3-yl]benzoic acid (4f). 65%; mp 221 ꢁC
...
(chloroform); IR (Nujol) 1505 (C–N), 3040 (C C of
aromatic C–H), 1700 (C¼O), 3014 (OH), 1428 (N¼N),
762 (C–Cl), 1771 (C¼O of b-thialactum) cmꢀ1
,
1H
NMR (CDCl3) d 12.30 (1H, s), 6.95 (1H, s), 8.65–7.20
(11H, m), 3.44 (3H, s), 3.64 (2H, s) (ppm); ms: m/z
(2.75%) M+ 467.5. Anal. calcd for C23H18N3O4SCl: C,
59.03; H, 3.85; N, 8.98; Found: C, 59.29; H, 3.76; N, 8.76.
5-(m-Chlorophenylazo)-2-[2-(p-ethylphenyl)5-methyl-4-
oxo-thiazolidin-3-yl]benzoic acid (4m). 35%; mp 212 C
ꢁ
(benzene); IR (Nujol) 1512 (C–N), 3044 (C C of aro-
2-[2-(p-Chlorophenyl)-4-oxo-thiazolidin-3-yl]-5-(phenyl-
azo)benzoic acid (4g). 55%; mp 210 ꢁC (ethanol); IR
(Nujol) 1502 (C–N), 3048 (C–C of aromatic C–H), 1700
...
matic C–H), 1705 (C¼O), 3002 (OH), 1426 (N¼N),

5290
A. Kumar et al. / Bioorg. Med. Chem. 11 (2003) 5281–5291
1769 (C¼O of b-thialactum), 764 (C–Cl) cmꢀ1
,
1H
Cyclooxygenase activity. This test was carried out in
vitro on the microsomal fraction of mucosal pre-
parations of rabbit distal colon in order to search out
the possible mechanism of the compounds. The pre-
paration was based on the method of Calderano et
al.13 Colonic mucosa ( ffi 2–3 g), stripped as pre-
viously described was minced and homogenized Pot-
ter homogenizer in 3 vols of Tris buffer 0.1, pH 8.0.
The homogenate was centrifused for 30 min at
10,000g. The resulting supernatent was centrifused for
1 h at 100,000g. The precipitate was suspended in
Tris buffer 0.1 M, pH 8.0, and recentrifused for 1 h
at 100,000g. The microsomal pellet was used imme-
diately for enzyme assay cyclooxygenase activity was
assayed by measuring the rate of conversion of ara-
chidonic acid to PGE2. Microsomal fractions (50 mL)
were incubated with test agents for 5 min at 37 ꢁC in
30 mL Tris–HCl, pH 8.0 containing 2 mMreduced
glutathione, 5 mM l-tryptophan, 1 mMhematin. The
substrate 20 mMarachidonic acid with tracer amount
of [1-14C] arachidonic acid [ ffi 200 (xx) cpm] was then
added and the reaction proceeded for 3 min at 37 ꢁC.
The reaction was stopped by addition of 0.2 mL of
ethyl ether/methanol/citric acid 0.2 M(30:4:1), which
was precooled at ꢀ25 ꢁC PGE2, was extracted twice
into the same mixture. The solvent was evaporated
under an N2 stream and radiolabelled arachidonic
acid was separated from radiolabelled acid was sepa-
rated from radiolabelled PGE3 by RP-HBLC. HPLC
NMR (CDCl3) d (5H, s), 12.38 (1H, s), 6.90 (1H, s),
8.66–7.28 (11H, m), 2.18 (3H, d), 5.98 (1H, q)
(ppm); ms: m/z (3.52%) M+ 479.5. Anal. calcd for
C25H22N3O3SCl: C, 62.56; H, 4.58; N, 8.75; Found: C,
62.63; H, 4.92; N, 8.80.
Pharmacological evaluation
Anti-inflammatory activity. Paw edema inhibition test
was used on albino rats of charles foster species by
adopting the method of Winter et al.9 Groups of five
rats of both sexes (body weight 120–160 g), pregnant
females excluded, were given a dose of a test com-
pound. 30 min later, 0.2 mL of 1% carrageenan
suspension in 0.9% NaCl solution was injected
subcutaneously, into the plantar aponeurosis of the
hind paw and the paw volume was measured by a
water plethysmometer socrel and then measured again
3 h later. The mean increase of paw volume at each
time interval was compared with that of control
group (five rats treated with carrageenan, but not
treated with test compounds) at the same time inter-
vals and percent inhibition values were calculated by
the formula given below:
Dt
% anti-inflammatory activity ¼ 1 À
 100
Dc
where Dt and Dc are tested and control groups, respec-
tively.
analysis was performed on
a
Hitachi spectro-
photometer equipped with a flow cell. the sample was
injected on an ultra sphere column (Beckman) ODS
5 mmꢂ4.6 mmꢂ25 cm. with 2 nmol unlabelled PGE2
as an interval standard, PG chromatographic profile
was obtained by isocratic elution with 150 mM
H3PO4 in water, pH 3.5, containing 30% acetonitrile,
a flow rate of 1 mL/min monitoring the uv absorp-
tion at 214 nm. Radioactivity that co-eluted with
autheutic PGE2 was quantified by liquid scintillation
spectrometry. Test samples were compared to paired
control incubations. The percentage of inhibition was
calculated as follows.
Analgesic activity. Acetic acid writhing test was per-
formed on mice by following the method of Davis et
al.10 Groups of five mice body weight (20–30 g) of both
sexes, pregnant female excluded, were given a dose of a
test compound. 30 min later, the animals were injected
intraperitoneally with 0.25 mL/mouse of 0.5% acetic
acid during the following 25 min. The mean number of
writhes for each experimental groups and percentage
decrease compared with the control group (five mice not
treated with test compounds) were calculated after 60 min.
Ulcerogenic activity. Groups of 10 rats (body weight
200–250 g) of both sexes, pregnant females excluded,
fasted for 24 h, were treated with an oral dose of a
test compound, except control group. All animals
were sacrificed 6 h after dosing and their stomachs
and small intestines were microscopically examined to
assess the incidence of hyperaemia, shedding of epi-
thelium, petechial and frank haemorrhages and erosion
or discrete ulceration with or without perforation. The
presence of any one of these criteria was considered to
be an evidence of ulcerogenic activity.11
½ðcpm control À cpm test= ðcpm controlÞÞ Â 100ꢀ
Statistical analysis. This was carried out using Student
t-test.
Acknowledgements
The authors are thankful to Central Drug Research
Institute, Lucknow, India (CDRI) for elemental and
spectral analysis.
Acute toxicity study. Approximate 50% lethal dose
(ALD50) of the compounds were determined in albino
mice. The mice of either sex 20–25 g were used. The test
compounds were injected intraperitoneally at different
dose levels in groups of 10 animals. After 24 h of drug
administration, percent mortality in each group was
observed from the data obtained ALD50 was calculated
by the method of Smith.12
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