Synthesis and biological activities of novel ethers of quinolinone linked with coumarins

Article abstract of DOI:10.1007/s00706-011-0460-3

A series of new ethers of quinolinone linked with different substituted coumarins and benzofurans weresynthesized from 4-(bromomethyl)quinolinones. All newly synthesized comp ounds were screened for their in vitro antibacterial and antifungal activities.

Full text of DOI:10.1007/s00706-011-0460-3

Monatsh Chem (2011) 142:305–315
DOI 10.1007/s00706-011-0460-3
ORIGINAL PAPER
Synthesis and biological activities of novel ethers of quinolinone
linked with coumarins
•
Rajesh G. Kalkhambkar G. Aridoss Geeta M. Kulkarni R. M. Bapset
•
•
•
•
T. Y. Mudaraddi N. Premkumar Yeon Tae Jeong
•
Received: 18 September 2010 / Accepted: 4 February 2011 / Published online: 11 March 2011
Ó Springer-Verlag 2011
Abstract A series of new ethers of quinolinone linked
with different substituted coumarins and benzofurans were
synthesized from 4-(bromomethyl)quinolinones. All newly
synthesized compounds were screened for their in vitro
antibacterial and antifungal activities. Most of the com-
pounds with chloro substitution at the C-6 or C-7 position
in quinolinone showed potent antibacterial and antifungal
activities. In pharmacological evaluations, some of these
chloroquinolinones also showed 70–77% inhibition of
inflammation after 8 h, whereas the other compounds
showed 51–55% inhibition. Most of the compounds
showed potent analgesic activity compared to the standard
and control. The structures of all newly synthesized com-
Keywords Quinolinones ꢀ Coumarin ꢀ Benzofuran ꢀ
Antibacterial ꢀ Antifungal ꢀ Analgesic Anti-inflammatory
Introduction
Coumarins and their nitrogen analogues 1-azacoumarins
(also known as quinolinones and carbostyrils) are hetero-
cyclic units, lactones and lactams, respectively, which are
indispensable to both chemists and biochemists. The nat-
ural occurrence, antimicrobial, anti-inflammatory, anti-
cancer, and miscellaneous other properties of these two
systems were recently reviewed and compared [1]. Many
coumarin derivatives are known for their ability to scav-
enge free radicals, especially reactive oxygen species
(ROS), and they have been used as inhibitors of cyclooxy-
genase and lipoxygenase in the arachidonic acid pathway of
inflammation suppression [2]. Nonsteroidal anti-inflamma-
tory drugs (NSAIDs) have a broad spectrum of effects in
acute pain management and target the cyclooxygenase
enzyme. Several coumarin derivatives have significant anti-
inflammatory activities and inhibit various enzymes
involved in modulating inflammation [3–5]. The potential
of coumarin derivatives as anti-inflammatory agents has
been explored in our laboratory, by incorporating biocom-
patible pharmacophores like p-vanillyl, cyanoester, and
paracetamol at position 4 of bromomethylcoumarin [6, 7].
We recently reported the synthesis and preliminary bio-
logical evaluation of an array of novel angularly fused
polycyclic heterocycles with coumarin, benzofuran, and
pyridine [8].
1
pounds were characterized by elemental analysis, IR, H
NMR, 13C NMR, and EI-MS.
R. G. Kalkhambkar ꢀ G. M. Kulkarni
Department of Chemistry, Karnatak Science College,
Karnatak University, Dharwad, Karnataka 580001, India
R. M. Bapset
Department of Chemistry, D.M.S Mandal’s Bhaurao
Kakatkar Science College, Club Road, Belgaum,
Karnataka 590001, India
T. Y. Mudaraddi
Department of Biotechnology and Microbiology,
Karnatak University, Dharwad, Karnataka 580003, India
N. Premkumar
Department of Pharmacology, Krupanidhi College of Pharmacy,
Bangalore, Karnataka 560034, India
G. Aridoss ꢀ Y. T. Jeong (&)
Quinolinone derivatives are metabolized to the corre-
sponding 8-hydroxycoumarins in biological systems
and were therefore found to be very good anti-inflamma-
tory and analgesic agents [9]. The triheterocyclic
Department of Image Science and Engineering,
Pukyong National University, Busan 608-739,
Republic of Korea
e-mail: ytjeong@pknu.ac.kr
123

306
R. G. Kalkhambkar et al.
thiazoles synthesized from 4-(aminomethyl)quinolinone
and 3-(bromoacetyl)coumarins [10] in our laboratory were
found to exhibit promising anti-inflammatory and analgesic
activities even after 24 h. We also reported that the intro-
duction of halogens in general, and fluorine in particular, at
position 4 in the aryloxy and arylamino moieties of coumarin
and quinolinone [11–13] enhances their antimicrobial,
analgesic, and anti-inflammatory activities. Interest in cou-
marins and quinolinones as antibiotics is due to the
observation that they are potent inhibitors of bacterial DNA
gyrase, which is involved in cell growth [14, 15].
Escherichia coli at 100 lg cm-3 and 10–27% inhibition at
25 lg cm-3. Similar compounds showed 68–79% inhibition
of Bacillus cirrhosis at 100 lg cm-3 and up to 20–35%
inhibition at 25 lg cm-3, and the remaining compounds
showed moderate activity against both the bacteria.
Compounds 2b–2d, 3b–3d, 4b–4d, 5b, 5c, and 6c
showed very good antifungal activity of 52% (3d) to 74%
(2b) inhibition of Aspergillus niger at 100 lg cm-3 and
12% (2d) to 41% (2b) inhibition at 25 lg cm-3 (Table 2).
Similar compounds showed 53% (3d) to 78% (4b) inhibi-
tion of Rhizoctonia bataticola at 100 lg cm-3 and 14%
We have documented the very good antibacterial activity
associated with many coumarin and quinolinone derivatives
bearing a variety of substituents at position 4 [16, 17]. In view
of the biological importance of coumarin and quinolinone, we
have designed new templates linking these two biolabile
heterocycles together through an ether linkage in order to
studythebiologicalpropertiesofthese heterocyclesasasingle
entity. This paper reports the synthesis and preliminary bio-
logical evaluation of ethers derived from quinolinone linked
with different substituted coumarins and benzofurans.
(2d) to 44% (4b) inhibition at 25 lg cm-3
.
Acute toxicity studies
The acute toxicity studies of the test compounds were per-
formed on albino mice fasted for 24 h. The test compounds
were administered orally and intraperitoneally. The animals
were watched for mortality and symptoms for 8 days [23,
24]. All the compounds possess a good safety profile and no
mortality of animals was observed even after 24 h.
Analgesic activity
Results and discussion
Abdominal constriction response induced by acetic acid is
a sensitive procedure to establish efficacy of peripherally
acting analgesics. Intraperitoneal (i.p.) administration of
acetic acid causes an increase in the level of PGE2 and
PGF 2a [25]. The results of the analgesic activity tests
(Table 3) indicate that compounds 3b, 2b, 4b, 5c, and 6c
with chloro substitution in the quinolinone moiety showed
the highest analgesic activity, even greater than that of the
standard. Compounds 3c, 2c, 4c, 5d, and 6d
showed moderate analgesic activity. A graphical repre-
sentation of the analgesic activity of selected compounds
is given in Fig. 1. Compounds 2b, 3b, 4b, 5c, and 6c with
chloro substitution in the quinolinone moiety were found
to be potent analgesic agents as compared to the standard.
Syntheses
The basic synthetic route employed for the construction of
various ethers is outlined in Scheme 1. The 4-(bromo-
methyl)quinolinones 1a–1d required for the construction of
the target molecules were synthesized by the bromination of
acetoacetanilides and cyclizing the intermediate x-bromo-
acetoacetanilides in sulfuric acid [11, 18]. The ethers 2a–2d,
3a–3d, 4a–4d, and 5a–5d were synthesized by the reaction
of 1a–1d and anhydrous K2CO3 with 6-hydroxy-4-methyl-
coumarin, 7-hydroxy-4-methylcoumarin, 7-hydroxy-4,5-
dimethylcoumarin, and 2,4-dihydroxyacetophenone, respec-
tively, in dry ethanol [19]. Further reaction of 5d with various
4-(bromomethyl)coumarins and anhydrous K2CO3 in dry
ethanol afforded the benzofuranyl ethers 6a–6e via an intra-
molecular aldol condensation followed by dehydration. All
the products gave satisfactory analytical and spectroscopic
data, which are in full accordance with their assigned
structures.
Anti-inflammatory activity
The anti-inflammatory activity of the test compounds was
assessed by the formalin-induced rat-paw edema method
[26]. The results in Table 3 indicate that most of the
compounds showed an extremely significant anti-inflam-
matory activity when compared to the control and the
standard group and their onset of action was much quicker
than the standard, because they showed significant anti-
inflammatory activity at 2 h after administration. Com-
pounds 5b, 5c, and 6b showed a maximum of 77%
inhibition of inflammation after 8 h. Compound 5c was
even more active than the other compounds after 12 and
24 h. The metabolites of compound 5c might also show
Antimicrobial activity
The in vitro antibacterial and antifungal activities of all
newly synthesized compounds were assessed by the cup-
plate method [20–22], and the results are shown in Table 1.
Compounds 2b, 2c, 3b, 3c, 4b, 4c, 5d, 5c, and 6c with
chloro substitution at the C-6 or C-7 position in quinolinone
showed potent antibacterial activity of 62–74% inhibition of
123

Synthesis and biological activities of novel ethers of quinolinone linked with coumarins
307
O
Compd
3a
3b
3c
3d
R
H
6-Cl
7-Cl
8-CH3
R
O
N
H
O
O
3a-3d
Reflux 7-Hydroxy-4-methylcoumarin
K2CO3 / EtOH
O
Br
O
6-Hydroxy-4-
methylcoumarin
7-Hydroxy-4,5-di-
methylcoumarin
O
O
R
R
R
O
K2CO3 / EtOH
K2CO3 / EtOH
Reflux
N
H
-
O
N
H
O
N
H
O
Reflux
O
2a 2d
1a-1d
4a-4d
Compd
R
H
6-Cl
7-Cl
8-CH3
Compd
4a
4b
4c
4d
R
H
6-Cl
7-Cl
2a
2b
2c
2d
K2CO3 / EtOH
Reflux
2,4-Dihydroxy-
acetophenone
8-CH3
O
OH
O
R
Subst. 4-(bromo-
methyl)coumarins
N
H
O
K2CO3 / EtOH
Reflux
5a-5d
O
Compd
5a
R
H
5b
5c
5d
6-Cl
7-Cl
8-CH3
R'
O
N
H
O
6a-6e
O
O
Compd R'
6a
6b
6c
6d
6e
6-CH3
7-CH3
6-Cl
5,6-Benzo
7,8-Benzo
Scheme 1
some anti-inflammatory activity. Compound 5c may be
more potent than the standard. Similarly compounds 6c and
5d showed a maximum of 74% inhibition of inflammation
and compound 6d showed a maximum of 70% inhibition of
inflammation after 8 h, whereas the rest of the compounds
showed moderate activity of 51–55% inhibition of
inflammation after 8 h. A graphical representation of the
anti-inflammatory activity of selected compounds is given
in Figs. 2 and 3. Compounds 4c, 5c, and 6b were active
even after 24 h.
123

308
R. G. Kalkhambkar et al.
Table 1 Antibacterial activity of selected compounds
Compd.
Compd. conc. (lg cm-3
100 50
)
Compd. conc. (lg cm-3
100 50
)
25
25
E. coli (gram negative)
B. cirrhosis (gram positive)
Zone of inhibition
Zone of inhibition
mm
%
mm
%
mm
%
mm
%
mm
%
mm
%
2b
184
178
102
172
177
98
74
98
88
51
81
78
44
81
72
58
62
84
92
43
61
51
80
40
200
20
43
41
32
–
27
210
202
138
198
188
122
190
184
136
132
186
192
128
130
138
190
140
260
20
79
128
122
72
54
71
64
–
35
2c
67
37
69
62
35
68
65
42
44
64
67
42
37
42
65
36
100
–
38
18
34
27
14
34
29
21
24
36
40
13
24
18
34
12
100
–
14
–
75
49
74
70
42
70
68
48
46
69
71
45
46
49
71
47
100
–
51
26
49
44
24
45
37
26
32
39
42
26
25
27
45
24
100
–
30
–
2d
3b
28
25
–
12
10
–
118
108
68
58
52
–
26
23
–
3c
3d
4b
170
164
114
118
162
178
113
102
114
165
100
240
20
32
26
–
14
10
–
110
94
58
40
–
26
20
–
4c
4d
72
5a
–
–
84
32
48
62
–
10
20
29
–
5b
32
44
–
14
22
–
98
5c
104
72
5d
6a
–
–
70
–
–
6b
–
–
74
–
–
6c
30
–
13
–
110
68
40
–
20
–
6d
Standard
Control
160
20
100
–
220
20
170
20
100
–
Standard, norfloxacin (100% inhibition at each concentration); control, dimethylformamide (DMF)
Conclusion
administration, i.e., these compounds can act as slow-
release anti-inflammatory agents. Our findings will have a
big impact on chemists and biochemists conducting further
investigations in this field in search of potent antimicrobial,
analgesic, and anti-inflammatory agents.
This study has shown that the presence of a chlorine sub-
stituent in the quinolinone or coumarin moiety enhances
the antimicrobial, analgesic, and anti-inflammatory activi-
ties. Most of the compounds, viz. 2b, 2c, 3b, 3c, 4b, 4c, 5b,
5c, and 6c with chloro substitution at the C-6 or C-7
position in quinolinone, showed potent antibacterial and
antifungal activities. In the pharmacological evaluation,
most of the compounds with chloro substitution showed
potent analgesic activity as compared to the standard and
control. Compounds 5b, 5c, and 5d showed 74–77%
inhibition of inflammation after 8 h. The introduction of a
benzofuran moiety between the quinolinone and coumarin
also enhanced the biological properties: compound 6c was
very active against both the bacteria and fungi. In phar-
macological studies, compounds 6b, 6c, and 6d showed
70–77% inhibition of inflammation after 8 h, and com-
pounds 2b, 2c, 3b, 3c, 4b, and 4c showed 51–55%
inhibition of inflammation after 8 h. In conclusion, nearly
all of the compounds with chloro substitution in quinoli-
none or coumarin showed the highest activity after 8 h and
some of them were active even at 12 and 24 h after
Experimental
The melting points of the products were determined in
open capillaries on a Buchi apparatus. The IR spectra were
recorded on a Nicolet Impact-410 FT-IR spectrophotome-
ter using KBr pellets. 1H NMR and 13C NMR spectra were
recorded on a Bruker AC-300F spectrometer in CDCl3,
DMSO-d6, or a mixture of CDCl3 and CF3COOH using
TMS as an internal standard at 300 MHz (1H) and 75 MHz
(13C) resonance frequency. D2O exchange was applied to
confirm the assignment of the signals of NH protons. The
mass spectra were recorded on an Autospec ESI-MS. The
elemental analysis was carried out using a Heraus CHN
rapid analyzer. C, H, and N values of all compounds were
found to be within 0.4% of the theoretical values. The
homogeneity of the compounds was assessed by TLC on
123

Synthesis and biological activities of novel ethers of quinolinone linked with coumarins
309
Table 2 Antifungal activity of some selected compounds
Compd.
Compd. conc. (lg cm-3
)
Compd. conc. (lg cm-3
)
100
50
25
100
50
25
A. niger
R. bataticola
Zone of inhibition
Zone of inhibition
mm
%
mm
%
mm
%
mm
%
mm
%
mm
%
2b
192
188
148
184
187
140
187
170
150
110
170
164
128
150
165
172
155
250
20
74
126
120
78
55
82
78
38
78
69
42
75
70
40
–
41
180
172
142
173
168
138
192
178
148
100
162
158
118
118
102
178
110
240
20
72
120
112
70
55
74
63
40
74
65
40
82
74
48
–
38
31
14
38
32
14
44
38
20
–
2c
73
55
71
72
52
72
65
56
39
65
62
46
56
62
66
57
100
–
52
30
51
47
30
51
47
30
22
44
41
26
22
32
51
28
100
–
38
12
38
32
14
36
33
13
–
69
55
69
67
53
78
71
58
36
64
62
44
44
37
71
39
100
–
50
27
54
50
26
60
55
30
–
2d
3b
118
110
78
118
110
68
3c
3d
4b
118
110
78
128
120
74
4c
4d
5a
62
–
5b
104
98
64
60
–
29
26
–
110
104
60
50
46
18
26
–
68
65
–
34
32
–
5c
5d
70
6a
62
–
–
68
–
–
6b
80
38
70
–
12
33
–
–
–
–
6c
118
74
105
–
46
–
66
–
32
–
6d
Standard
Control
210
20
170
20
100
–
200
20
100
–
160
20
100
–
Standard, griseofulvin (100% inhibition at each concentration); control, DMF
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.56 (s, 3H, C40-CH3 of coumarin), 5.50 (s, 2H, C4-
CH2 of quinolinone), 6.26 (s, 1H, C3-H of coumarin), 6.56
(s, 1H, C3-H of quinolinone), 7.34–7.94 (m, 7H, Ar–H),
13.03 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ?
CF3COOH): d = 163.50, 160.22, 157.28, 156.34, 154.28,
136.89, 133.48, 132.80, 127.08, 124.60, 119.40, 116.30,
116.21, 114.47, 114.30, 112.65, 110.98, 109.10, 65.98,
18.34 ppm; EI-MS: m/z = 345 (M ? H).
aluminum silica gel 60 F254 (Merck) detected by UV light
(254 nm) and iodine vapors. The reagents were all ana-
lytical reagent grade or chemically pure. All solvents were
dried, deoxygenated, and redistilled before use by standard
procedures [19].
General procedure for the synthesis of compounds
2a–2e
A mixture of substituted 4-(bromomethyl)quinolinone 1a–1d
(4.0 mmol), 6-hydroxy-4-methylcoumarin (4.0 mmol), and
anhydrous potassium carbonate (4.0 mmol) in 20 cm3 dry
ethanol was refluxed on a water bath for 8 h. The separated
solid was filtered, washed with 20% HCl and with an excess
of cold water, dried, and crystallized from a suitable solvent.
6-Chloro-4-[(4-methyl-2-oxo-2H-chromen-6-yloxy)-
methyl]quinolin-2(1H)-one (2b, C20H14ClNO4)
Yield 84%; colorless crystals (acetic acid); m.p.:
284–286 °C; IR (KBr): vꢀ = 3,428 (N–H stretching),
1,720 (C=O stretching, lactone), 1,660 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.50 (s, 3H, C40-CH3 of coumarin), 5.56 (s, 2H, C4-
CH2 of quinolinone), 6.30 (s, 1H, C3-H of coumarin), 6.48
(s, 1H, C3-H of quinolinone), 7.30–7.90 (m, 6H, Ar–H),
13.08 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ?
CF3COOH): d = 164.30, 161.20, 158.23, 156.74, 155.88,
4-[(4-Methyl-2-oxo-2H-chromen-6-yloxy)methyl]-
quinolin-2(1H)-one (2a, C20H15NO4)
Yield 74%; colorless crystals (acetic acid); m.p.:
308–310 °C; IR (KBr): vꢀ = 3,433 (N–H stretching),
1,707 (C=O stretching, lactone), 1,657 (C=O stretching,
123

310
R. G. Kalkhambkar et al.
Table 3 Anti-inflammatory and analgesic activities of some selected compounds
Compd.
Anti-inflammatory activitya
Analgesic activity
(number of writhings)
Edema volume at different time intervals (% of inhibition)
0.5 h
1 h
2 h
4 h
8 h
0.63* (42)
12 h
0.81 (16)
24 h
Standard
Control
2b
0.66 (4)
0.71
0.81 (9)
0.91
0.88 (12)
0.68* (36)
0.98 (14)
1.01
22*
35
1.01
1.06
1.07
0.89
0.64 (5)
0.60 (5)
0.73 (0)
0.71 (0)
0.78 (0)
0.65 (5)
0.61 (7)
0.60 (7)
0.58 (11)
0.60 (8)
0.63 (5)
0.64 (4)
2.86
0.78 (8)
0.80 (9)
1.19 (0)
1.04 (0)
1.10 (0)
0.77 (10)
0.76 (10)
0.78 (8)
0.75 (12)
0.77 (10)
0.78 (8)
0.78 (8)
8.16
0.69* (32)
0.66* (31)
0.88 (12)
0.70 (30)
0.66* (31)
0.88 (12)
0.70 (30)
0.78 (18)
0.75 (21)
0.55* (42)
0.78 (18)
0.75 (21)
0.59* (45)
0.59* (45)
0.57* (44)
0.58* (46)
0.43* (60)
0.58* (46)
0.28* (72)
0.75*** (25)
0.58* (42)
0.28* (72)
0.75*** (25)
0.58* (42)
0.53* (51)
0.53* (51)
0.48* (55)
0.52* (51)
0.48* (55)
0.52* (51)
0.23* (77)
0.23* (77)
0.26* (74)
0.23* (77)
0.26* (74)
0.29* (70)
0.77 (12)
1.14 (0)
1.14 (0)
1.00 (0)
1.13 (0)
0.88 (7)
0.98 (14)
0.86 (9)
0.98 (14)
0.88 (7)
0.96 (12)
0.86 (9)
0.88 (7)
2.54
28*
19*
29*
20*
27*
18*
13*
27*
18*
13*
26*
18*
86.44
2c
0.76 (12)
3b
0.54** (39)
0.77 (11)
3c
4b
0.76 (12)
4c
0.54** (39)
0.50* (39)
0.45* (45)
0.56*** (32)
0.50* (39)
0.56*** (32)
0.45* (45)
5b
5c
5d
6b
6c
6d
F value
12.3
21.7
27.3
10.6
Standard error of the mean (SEM) 0.05–0.15. Standard, indomethacin at a dose of 10 mg kg-1 (i.p.). Test compounds at a dose of 100 mg kg-1
(i.p.). Data were analyzed by one-way ANOVA followed by Tukey comparison of all pairs
* P \ 0.05, ** P \ 0.01, *** P \ 0.001 when compared to control
a
Assessed in a formalin-induced (3.5%) acute inflammation model in male Wistar rats (n = 6)
30
25
20
15
10
5
60
50
40
30
20
10
0
2b
2c
3b
3c
4b
4c
5b
5c
5d
6b
6c
6d
std
0.5h
1h
2h
4h
8h
12h
24h
2b
2c
3b
3c
4b
4c
std
Compounds
Fig. 2 Anti-inflammatory activity of selected compounds and
standard
0
80
Compounds
70
60
50
40
30
20
10
0
0.5h
1h
Fig. 1 Analgesic activity of selected compounds and standard
2h
4h
8h
136.89, 134.48, 132.80, 127.08, 124.60, 118.40, 116.33,
116.21, 115.47, 114.30, 111.60, 110.98, 108.14, 64.18,
19.14 ppm; EI-MS: m/z = 368 (M ? H).
12h
24h
5b
5c
5d
6b
6c
6d
std
7-Chloro-4-[(4-methyl-2-oxo-2H-chromen-6-yloxy)-
methyl]quinolin-2(1H)-one (2c, C20H14ClNO4)
Compounds
Yield 80%; colorless crystals (acetic acid); m.p.:
320–322 °C; IR (KBr): vꢀ = 3,434 (N–H stretching),
Fig. 3 Anti-inflammatory activity of selected compounds and
standard
123

Synthesis and biological activities of novel ethers of quinolinone linked with coumarins
311
1,707 (C=O stretching, lactone), 1,669 (C=O stretching,
1
6-Chloro-4-[(4-methyl-2-oxo-2H-chromen-7-yloxy)-
methyl]quinolin-2(1H)-one (3b, C20H14ClNO4)
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.58 (s, 3H, C40-CH3 of coumarin), 5.60 (s, 2H, C4-
CH2 of quinolinone), 6.20 (s, 1H, C3-H of coumarin), 6.58
(s, 1H, C3-H of quinolinone), 7.37–7.95 (m, 6H, Ar–H),
13.02 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ?
CF3COOH): d = 164.10, 160.40, 158.23, 156.44, 155.58,
136.85, 134.45, 132.80, 127.08, 124.60, 118.40, 116.30,
116.21, 115.47, 114.10, 111.67, 110.98, 109.10, 65.10,
18.34 ppm; EI-MS: m/z = 368 (M ? H).
Yield 70%; colorless crystals (acetic acid); m.p.:
315–317 °C; IR (KBr): vꢀ = 3,440 (N–H stretching),
1,720 (C=O stretching, lactone), 1,660 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.48 (s, 3H, C40-CH3 of coumarin), 5.65 (s, 2H, C4-
CH2 of quinolinone), 6.22 (s, 1H, C3-H of coumarin), 6.40
(s, 1H, C3-H of quinolinone), 7.10–8.08 (m, 6H, Ar–H),
12.98 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ?
CF3COOH): d = 165.14, 160.11, 158.18, 155.34, 154.28,
137.89, 134.18, 132.10, 127.08, 125.60, 120.40, 116.30,
116.20, 114.47, 114.30, 110.65, 109.18, 107.13, 65.28,
18.18 ppm; EI-MS: m/z = 368 (M ? H).
8-Methyl-4-[(4-methyl-2-oxo-2H-chromen-6-yloxy)-
methyl]quinolin-2(1H)-one (2d, C21H17NO4)
Yield 88%; colorless crystals (acetic acid); m.p.:
296–298 °C; IR (KBr): vꢀ = 3,440 (N–H stretching),
1,708 (C=O stretching, lactone), 1,658 (C=O stretching,
7-Chloro-4-[(4-methyl-2-oxo-2H-chromen-7-yloxy)-
methyl]quinolin-2(1H)-one (3c, C20H14ClNO4)
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.48 (s, 3H, C8-CH3 of quinolinone), 2.56 (s, 3H, C40-
CH3 of coumarin), 5.55 (s, 2H, C4-CH2 of quinolinone),
6.32 (s, 1H, C3-H of coumarin), 6.50 (s, 1H, C3-H of
quinolinone), 7.35–7.88 (m, 6H, Ar–H), 13.08 (s, 1H, NH)
ppm; 13C NMR (75 MHz, CDCl3 ? CF3COOH):
d = 166.00, 161.20, 158.23, 156.48, 154.28, 136.85,
134.45, 130.80, 128.08, 124.60, 118.40, 116.30, 116.21,
114.47, 114.10, 111.67, 110.98, 108.10, 65.10, 20.30,
18.10 ppm; EI-MS: m/z = 348 (M ? H).
Yield 72%; colorless crystals (acetic acid); m.p.:
292–294 °C; IR (KBr): vꢀ = 3,436 (N–H stretching),
1,719 (C=O stretching, lactone), 1,667 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.50 (s, 3H, C40-CH3 of coumarin), 5.45 (s, 2H,
C4-CH2 of quinolinone), 6.30 (s, 1H, C3-H of coumarin),
6.48 (s, 1H, C3-H of quinolinone), 7.05–7.96 (m, 6H,
Ar–H), 12.88 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3
? CF3COOH): d = 164.34, 161.20, 159.18, 154.14,
153.18, 137.89, 134.18, 132.10, 128.08, 125.60, 119.40,
116.30, 115.20, 114.47, 114.20, 110.65, 109.18, 108.13,
64.22, 19.28 ppm; EI-MS: m/z = 368 (M ? H).
General procedure for the synthesis of compounds
3a–3d
8-Methyl-4-[(4-methyl-2-oxo-2H-chromen-7-yloxy)-
methyl]quinolin-2(1H)-one (3d, C21H17NO4)
A mixture of substituted 4-(bromomethyl)quinolinone 1a–1d
(4.0 mmol), 7-hydroxy-4-methylcoumarin (4.0 mmol), and
anhydrous potassium carbonate (4.0 mmol) in 20 cm3
dry ethanol was refluxed on a water bath for 8 h. The sepa-
rated solid was filtered, washed with 20% HCl and with an
excess of cold water, dried, and crystallized from a suitable
solvent.
Yield 82%; colorless crystals (acetic acid); m.p.:
288–290 °C; IR (KBr): vꢀ = 3,442 (N–H stretching),
1,710 (C=O stretching, lactone), 1,658 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.48 (s, 3H, C8-CH3 of quinolinone), 2.60 (s, 3H, C40-
CH3 of coumarin), 5.65 (s, 2H, C4-CH2 of quinolinone),
6.32 (s, 1H, C3-H of coumarin), 6.50 (s, 1H, C3-H of
quinolinone), 7.10–7.98 (m, 6H, Ar–H), 12.98 (s, 1H, NH)
ppm; 13C NMR (75 MHz, CDCl3 ? CF3COOH):
d = 165.24, 160.10, 159.28, 155.10, 152.18, 138.80,
134.18, 132.10, 128.08, 126.66, 119.40, 116.30, 115.20,
114.47, 114.10, 110.65, 109.18, 108.30, 64.22, 20.22,
19.28 ppm; EI-MS: m/z = 348 (M ? H).
4-[(4-Methyl-2-oxo-2H-chromen-7-yloxy)methyl]-
quinolin-2(1H)-one (3a, C20H15NO4)
Yield 78%; colorless crystals (acetic acid); m.p.:
310–312 °C; IR (KBr): vꢀ = 3,435 (N–H stretching),
1,717 (C=O stretching, lactone), 1,657 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.57 (s, 3H, C40-CH3 of coumarin), 5.55 (s, 2H, C4-
CH2 of quinolinone), 6.28 (s, 1H, C3-H of coumarin), 6.43
(s, 1H, C3-H of quinolinone), 7.08–7.96 (m, 7H, Ar–H),
13.07 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ?
CF3COOH): d = 164.54, 161.02, 158.18, 156.34, 154.28,
136.89, 133.18, 132.10, 127.08, 124.60, 119.40, 116.30,
116.21, 114.47, 114.30, 111.65, 110.18, 108.10, 64.38,
19.32 ppm; EI-MS: m/z = 334 (M ? H).
General procedure for the synthesis of compounds
4a–4d
A mixture of substituted 4-(bromomethyl)quinolinone 1a–1d
(4.0 mmol), 7-hydroxy-4,5-dimethylcoumarin (4.0 mmol),
and anhydrous potassium carbonate (4.0 mmol) in 20 cm3
123

312
R. G. Kalkhambkar et al.
114.40, 114.10, 111.65, 110.10, 108.10, 65.40, 20.12,
19.22 ppm; EI-MS: m/z = 304 (M?).
dry ethanol was refluxed on a water bath for 8 h. The sepa-
rated solid was filtered, washed with 20% HCl and with an
excess of cold water, dried, and crystallized from a suitable
solvent.
4-[(4,5-Dimethyl-2-oxo-2H-chromen-7-yloxy)methyl]-8-
methylquinolin-2(1H)-one (4d, C22H19NO4)
Yield 88%; colorless crystals (acetic acid); m.p.:
244–246 °C; IR (KBr): vꢀ = 3,415 (N–H stretching),
1,721 (C=O stretching, lactone), 1,659 (C=O stretching,
4-[(4,5-Dimethyl-2-oxo-2H-chromen-7-yloxy)methyl]-
quinolin-2(1H)-one (4a, C21H17NO4)
Yield 80%; colorless crystals (acetic acid); m.p.:
285–287 °C; IR (KBr): vꢀ = 3,535 (N–H stretching),
1,720 (C=O stretching, lactone), 1,666 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.47 (s, 3H, C40-CH3 of coumarin), 2.55 (s, 3H, C50-
CH3 of coumarin), 2.62 (s, 3H, C8-CH3 of quinolinone),
5.58 (s, 2H, C4-CH2 of quinolinone), 6.25 (s, 1H, C3-H of
coumarin), 6.68 (s, 1H, C30-H of quinolinone), 6.96 (s, 1H,
C60-H of coumarin), 7.12 (s, 1H, C80-H of coumarin),
7.34–7.90 (m, 3H, Ar–H), 13.02 (s, 1H, NH) ppm; 13C
NMR (75 MHz, CDCl3 ? CF3COOH): d = 165.10,
160.22, 159.10, 158.24, 154.48, 136.80, 134.15, 130.10,
125.08, 120.60, 119.40, 117.30, 115.19, 114.44, 114.10,
111.66, 110.10, 108.10, 66.40, 20.12, 19.22, 18.30 ppm;
EI-MS: m/z = 362 (M ? H).
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.48 (s, 3H, C40-CH3 of coumarin), 2.54 (s, 3H, C50-
CH3 of coumarin), 5.56 (s, 2H, C4-CH2 of quinolinone),
6.27 (s, 1H, C3-H of coumarin), 6.75 (s, 1H, C30-H of
quinolinone), 6.98 (s, 1H, C60-H of coumarin), 7.16 (s, 1H,
C80-H of coumarin), 7.55–7.90 (m, 4H, Ar–H), 13.04 (s,
1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ? CF3COOH):
d = 163.44, 160.22, 159.18, 156.34, 154.28, 136.89,
134.10, 132.10, 127.08, 124.60, 119.40, 116.30, 116.12,
114.47, 114.10, 111.65, 110.10, 108.10, 65.40, 20.12,
18.32 ppm; EI-MS: m/z = 348 (M ? H).
General procedure for the synthesis of compounds
5a–5d
6-Chloro-4-[(4,5-dimethyl-2-oxo-2H-chromen-7-yloxy)-
methyl]quinolin-2(1H)-one (4b, C21H16ClNO4)
Yield 78%; colorless crystals (acetic acid); m.p.:
282–284 °C; IR (KBr): vꢀ = 3,434 (N–H stretching),
1,721 (C=O stretching, lactone), 1,668 (C=O stretching,
A mixture of substituted 4-(bromomethyl)quinolinone 1a–
1d (4.0 mmol), 2,4-dihydroxyacetophenone (4.0 mmol),
and anhydrous potassium carbonate (4.0 mmol) in 20 cm3
dry ethanol was refluxed on a water bath for 8 h. The
separated solid was filtered, washed with 20% HCl and
with an excess of cold water, dried, and crystallized from a
suitable solvent.
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.47 (s, 3H, C40-CH3 of coumarin), 2.55 (s, 3H, C50-
CH3 of coumarin), 5.54 (s, 2H, C4-CH2 of quinolinone),
6.28 (s, 1H, C3-H of coumarin), 6.76 (s, 1H, C30-H of
quinolinone), 6.96 (s, 1H, C60-H of coumarin), 7.15 (s, 1H,
C80-H of coumarin), 7.54–7.90 (m, 3H, Ar–H), 13.02 (s,
1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ? CF3COOH):
d = 165.14, 160.20, 158.18, 157.34, 153.28, 135.89,
133.10, 131.10, 126.08, 120.60, 118.40, 116.30, 115.12,
114.47, 114.00, 111.65, 110.10, 109.10, 64.40, 20.82,
19.22 ppm; EI-MS: m/z = 382 (M ? H).
4-[(4-Acetyl-3-hydroxyphenoxy)methyl]quinolin-2(1H)-
one (5a, C18H15NO4)
Yield 68%; colorless crystals (acetic acid); m.p.:
280–282 °C; IR (KBr): vꢀ = 3,430 (N–H stretching),
1,718 (C=O stretching, lactone), 1,664 (C=O stretching,
amide) cm-1; 1H NMR (300 MHz, DMSO-d6): d = 2.58 (s,
3H, CH3 of acetyl), 5.49 (s, 2H, C4-CH2 of quinolinone),
6.61–7.90 (m, 8H, Ar–H), 10.88 (s, 1H, NH), 12.62 (s, 1H,
OH) ppm; 13C NMR (75 MHz, DMSO-d6): d = 166.44,
162.22, 159.18, 156.34, 154.28, 136.89, 134.10, 132.10,
127.08, 124.60, 119.40, 116.30, 116.12, 114.10, 111.65,
108.10, 66.40, 23.12 ppm; EI-MS: m/z = 310 (M ? H).
7-Chloro-4-[(4,5-dimethyl-2-oxo-2H-chromen-7-yloxy)-
methyl]quinolin-2(1H)-one (4c, C21H16ClNO4)
Yield 75%; colorless crystals (acetic acid); m.p.:
281–282 °C; IR (KBr): vꢀ = 3,428 (N–H stretching),
1,718 (C=O stretching, lactone), 1,660 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.47 (s, 3H, C40-CH3 of coumarin), 2.54 (s, 3H, C50-
CH3 of coumarin), 5.57 (s, 2H, C4-CH2 of quinolinone),
6.26 (s, 1H, C3-H of coumarin), 6.75 (s, 1H, C30-H of
quinolinone), 6.97 (s, 1H, C60-H of coumarin), 7.16 (s, 1H,
C80-H of coumarin), 7.56–7.91 (m, 3H, Ar–H), 13.08 (s,
1H, NH) ppm; 13C NMR (75 MHz, CDCl3 ? CF3COOH):
d = 164.14, 161.20, 159.18, 158.34, 154.28, 136.89,
134.10, 130.10, 125.08, 120.60, 119.40, 117.30, 115.12,
4-[(4-Acetyl-3-hydroxyphenoxy)methyl]-6-chloroquinolin-
2(1H)-one (5b, C18H14ClNO4)
Yield 65%; colorless crystals (acetic acid); m.p.:
294–296 °C; IR (KBr): vꢀ = 3,434 (N–H stretching),
1,710 (C=O stretching, lactone), 1,650 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, DMSO-d6): d = 2.68
(s, 3H, CH3 of acetyl), 5.58 (s, 2H, C4-CH2 of quinoli-
none), 6.60–7.80 (m, 7H, Ar–H), 11.34 (s, 1H, NH), 12.43
123

Synthesis and biological activities of novel ethers of quinolinone linked with coumarins
313
(s, 1H, OH) ppm; 13C NMR (75 MHz, DMSO-d6):
d = 165.45, 161.32, 158.18, 156.34, 154.28, 136.89,
134.10, 132.10, 127.08, 124.60, 119.40, 116.30, 116.10,
114.10, 111.64, 108.10, 65.42, 25.32 ppm; EI-MS:
m/z = 344 (M ? H).
coumarin), 2.60 (s, 3H, C8-CH3 of quinolinone), 5.35 (s,
2H, C4-CH2 of quinolinone), 6.53–7.72 (m, 11H, Ar–H),
9.23 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3):
d = 166.44, 160.22, 159.18, 158.90, 156.34, 154.28,
138.87, 136.89, 134.10, 132.10, 130.30, 128.83, 127.08,
125.87, 124.60, 123.45, 120.56, 119.40, 116.30, 116.12,
115.54, 114.10, 111.65, 110.90, 108.10, 63.40, 23.12,
21.12, 18.90 ppm; EI-MS: m/z = 478 (M ? H).
4-[(4-Acetyl-3-hydroxyphenoxy)methyl]-7-chloroquinolin-
2(1H)-one (5c, C18H14ClNO4)
Yield 60%; colorless crystals (acetic acid); m.p.:
290–292 °C; IR (KBr): vꢀ = 3,431 (N–H stretching),
1,711 (C=O stretching, lactone), 1,652 (C=O stretching,
8-Methyl-4-[[3-methyl-2-(7-methyl-2-oxo-2H-chromen-4-yl)-
benzofuran-6-yloxy]methyl]quinolin-2(1H)-one
(6b, C30H23NO5)
1
amide) cm-1; H NMR (300 MHz, CDCl3 ? CF3COOH):
d = 2.69 (s, 3H, CH3 of acetyl), 5.56 (s, 2H, C4-CH2 of
quinolinone), 6.67–7.90 (m, 7H, Ar–H), 11.54 (s, 1H, NH),
13.14 (s, 1H, OH) ppm; 13C NMR (75 MHz,
CDCl3 ? CF3COOH): d = 166.75, 160.22, 159.18,
156.34, 154.28, 136.89, 134.10, 132.10, 127.08, 124.60,
120.40, 116.30, 116.10, 114.10, 111.64, 109.10, 64.42,
24.32 ppm; EI-MS: m/z = 344 (M ? H).
Yield 68%; colorless crystals (acetic acid); m.p.:
252–254 °C; IR (KBr): vꢀ = 3,433 (N–H stretching),
1,727 (C=O stretching, lactone), 1,675 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3): d = 2.47 (s,
3H, C3-CH3 of benzofuran), 2.54 (s, 3H, C7-CH3 of
coumarin), 2.62 (s, 3H, C8-CH3 of quinolinone), 5.42 (s,
2H, C4-CH2 of quinolinone), 6.60–7.78 (m, 11H, Ar–H),
9.33 (s, 1H, NH) ppm; 13C NMR (75 MHz, CDCl3):
d = 167.40, 162.12, 159.88, 158.90, 156.34, 154.28,
138.87, 136.80, 134.10, 132.10, 131.30, 128.83, 127.08,
125.87, 124.60, 123.45, 121.56, 119.40, 118.30, 116.12,
115.54, 114.10, 111.65, 110.90, 109.10, 64.40, 24.12,
21.10, 18.92 ppm; EI-MS: m/z = 478 (M ? H).
4-[(4-Acetyl-3-hydroxyphenoxy)methyl]-8-methylquinolin-
2(1H)-one (5d, C19H17NO4)
Yield 75%; colorless crystals (acetic acid); m.p.:
270–272 °C; IR (KBr): vꢀ = 3,445 (N–H stretching),
1,700 (C=O stretching, lactone), 1,660 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3): d = 2.49 (s,
3H, C8-CH3 of quinolinone), 2.60 (s, 3H, CH3 of acetyl),
5.35 (s, 2H, C4-CH2 of quinolinone), 6.53–7.72 (m, 7H,
Ar–H), 8.94 (s, 1H, NH), 12.75 (s, 1H, OH) ppm; 13C NMR
(75 MHz, CDCl3): d = 165.70, 162.12, 159.18, 156.34,
154.28, 136.89, 134.10, 132.10, 127.08, 124.60, 120.40,
116.30, 116.10, 113.10, 111.64, 108.18, 65.42, 25.32,
18.30 ppm; EI-MS: m/z = 324 (M ? H).
8-Methyl-4-[[2-(6-chloro-2-oxo-2H-chromen-4-yl)-3-
methylbenzofuran-6-yloxy]methyl]quinolin-2(1H)-one
(6c, C29H20ClNO5)
Yield 63%; colorless crystals (acetic acid); m.p.:
240–242 °C; IR (KBr): vꢀ = 3,428 (N–H stretching),
1,730 (C=O stretching, lactone), 1,662 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, DMSO-d6): d = 2.63
(s, 3H, C3-CH3 of benzofuran), 2.64 (s, 3H, C8-CH3 of
quinolinone), 5.46 (s, 2H, C4-CH2 of quinolinone),
7.28–7.75 (m, 11H, Ar–H), 11.49 (s, 1H, NH) ppm; 13C
NMR (75 MHz, DMSO-d6): d = 165.40, 160.12, 159.00,
158.90, 156.34, 154.28, 138.87, 135.80, 134.10, 132.10,
131.30, 128.83, 127.08, 125.87, 124.60, 123.45, 121.56,
120.40, 118.30, 116.12, 115.54, 114.10, 111.65, 110.90,
108.10, 65.40, 24.12, 18.92 ppm; EI-MS: m/z = 498
(M ? H).
General procedure for the synthesis of compounds
6a–6e
A mixture of 4-[(4-acetyl-3-hydroxyphenoxy)methyl]-8-
methylquinolin-2(1H)-one 5d (4.0 mmol), the appropriate
substituted 4-(bromomethyl)coumarin (4.0 mmol), and
anhydrous potassium carbonate (8.0 mmol) in 20 cm3 dry
ethanol was refluxed on a water bath for 14 h. The sepa-
rated solid was filtered, washed with 20% HCl and with an
excess of cold water, dried, and crystallized from a suitable
solvent.
8-Methyl-4-[[3-methyl-2-(3-oxo-3H-naphtho[2,1-b]pyran-
1-yl)benzofuran-6-yloxy]methyl]quinolin-2(1H)-one
(6d, C34H23NO5)
8-Methyl-4-[[3-methyl-2-(6-methyl-2-oxo-2H-chromen-4-
yl)benzofuran-6-yloxy]methyl]quinolin-2(1H)-one
(6a, C30H23NO5)
Yield 62%; colorless crystals (acetic acid); m.p.:
248–250 °C; IR (KBr): vꢀ = 3,425 (N–H stretching),
1,718 (C=O stretching, lactone), 1,659 (C=O stretching,
amide) cm-1; 1H NMR (300 MHz, DMSO-d6): d = 2.54 (s,
3H, C3-CH3 of benzofuran), 2.66 (s, 3H, C8-CH3 of
quinolinone), 5.48 (s, 2H, C4-CH2 of quinolinone),
7.20–7.89 (m, 14H, Ar–H), 11.38 (s, 1H, NH) ppm; 13C
NMR (75 MHz, DMSO-d6): d = 166.80, 161.10, 160.12,
Yield 58%; colorless crystals (acetic acid); m.p.:
270–272 °C; IR (KBr): vꢀ = 3,430 (N–H stretching),
1,722 (C=O stretching, lactone), 1,665 (C=O stretching,
1
amide) cm-1; H NMR (300 MHz, CDCl3): d = 2.42 (s,
3H, C3-CH3 of benzofuran), 2.51 (s, 3H, C6-CH3 of
123

314
R. G. Kalkhambkar et al.
100 mg kg-1, i.e., 1/10 of the cutoff value, was taken as
the screening dose for the evaluation of anti-inflammatory
activity.
158.98, 157.40, 156.30, 154.22, 144.09, 138.80, 135.18,
133.90, 132.10, 132.23, 128.86, 127.14, 125.80, 124.67,
123.55, 123.50, 122.60, 122.40, 120.41, 118.37, 117.45,
116.10, 115.50, 114.14, 111.75, 110.87, 109.20, 66.38,
24.10, 18.82 ppm; EI-MS: m/z = 526 (M?H).
Analgesic activity
8-Methyl-4-[[3-methyl-2-(2-oxo-2H-naphtho-
[1,2-b]pyran-4-yl)benzofuran-6-yloxy]methyl]-
quinolin-2(1H)-one (6e, C34H23NO5)
The analgesic activity of the test compounds was assessed
in vivo using the abdominal constriction test induced by
acetic acid 0.6% (0.1 cm3/10 g) in mice [25]. Albino mice
of both sexes (18–22 g) were used. Compounds were
administered orally (10 mg kg-1) as a suspension in 5%
carbethoxymethyl cellulose (CMC, vehicle). Indomethacin
(10 mg kg-1) was used as the standard drug under the
same conditions.
Yield 60%; colorless crystals (acetic acid); m.p.:
260–262 °C; IR (KBr): vꢀ = 3,435 (N–H stretching),
1,723 (C=O stretching, lactone), 1,675 (C=O stretching,
amide) cm-1; 1H NMR (300 MHz, DMSO-d6): d = 2.58 (s,
3H, C3-CH3 of benzofuran), 2.60 (s, 3H, C8-CH3 of
quinolinone), 5.55 (s, 2H, C4-CH2 of quinolinone),
7.16–7.88 (m, 14H, Ar–H), 11.52 (s, 1H, NH) ppm; 13C
NMR (75 MHz, DMSO-d6): d = 166.47, 161.00, 159.44,
158.76, 157.44, 156.37, 155.32, 144.10, 138.96, 135.78,
134.76, 130.70, 129.73, 128.66, 128.14, 125.71, 124.22,
124.10, 123.80, 123.60, 122.60, 120.11, 118.17, 117.67,
116.18, 115.54, 114.55, 111.95, 110.70, 109.25, 65.40,
24.50, 18.20 ppm; EI-MS: m/z = 526 (M ? H).
Anti-inflammatory activity
The anti-inflammatory activity of the test compounds was
assessed by the formalin-induced rat paw edema inhibition
method according to Winter et al. [26], by employing 3.5%
of formalin as the phlogistic agent. All test compounds
were administered orally as suspensions in 2% CMC,
30 min before the injection of the phlogistic agent, at a
dose of 100 mg kg-1 body weight. Indomethacin was used
as a standard at a dose of 10 mg kg-1 body weight. Groups
of six Sprague–Dawley rats of either sex were used in each
experiment. Plain CMC (2%) served as a control. The paw
edema volume was measured with the help of a plethys-
mograph by the mercury displacement method at 0 h
(immediately after injection of formalin), 1, 2, 3, 4, and
5 h.
Antimicrobial assay
The in vitro antimicrobial activity of the target compounds
was tested against the two bacterial microorganisms E. coli
(gram negative) and B. cirrhosis (gram positive) and the
two fungal microorganisms A. niger and R. bataticola.
DMF was used as a solvent control, and the reference drugs
used were norfloxacin and griseofulvin. The tests were
carried out by the cup-plate method [20–22], at concen-
trations of 100, 50, and 25 lg cm-3. After 48 h of
incubation at 37 °C the zone of inhibition was measured.
The percentage inhibition of test compounds was calcu-
lated by relating the zone of inhibition of the test
compound (ZOItest compound) to those of the standard
(ZOIstandard, taken as 100%) and control (ZOIcontrol) as
follows:
Acknowledgments The authors thank the University Sophisticated
Instrument Center, KUD, for IR, 1H NMR, and 13C NMR. Thanks are
also due to SAIF-CDRI Lucknow for ESI-MS data. One of the authors
(RGK) is grateful to Karnatak University and Karnatak Science
College Dharwad for the University Research Studentship. GA and
YTJ are grateful for the financial support provided by the second
stage of the BK21 program.
À
Á
ZOItest compound ꢁ ZOIcontrol
% inhibition =
ꢂ 100%:
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