Synthesis and biological evaluation of some new pyridazinone derivatives

Article abstract of DOI:10.3109/14756366.2010.548810

A series of pyridazinone derivatives (1934) were synthesized with an aim to synthesize safer anti-inflammatory agents. The compounds were evaluated for their anti-inflammatory, analgesic, ulcerogenic and lipid peroxidation (LPO) actions. The percentage inhibition in edema at different time intervals indicated that compounds 20, 26, 28 and 34 exhibited good anti-inflammatory potential, comparable with that of ibuprofen (85.77%) within a range of 67.4877.23%. The results illustrate that 5-(4-fluoro-benzyl)-3-(4-chloro-phenyl) -1,6-dihydro-6-pyridazinone (26) and 5-(4-chloro-benzyl)-3-(4-chloro-phenyl)-1, 6-dihydro-6-pyridazinone (20) showed best anti-inflammatory activity. Furthermore, activity is more in case of chloro substitution as compared with methyl-substitution. The compounds synthesized were also evaluated for their ulcerogenic and LPO action and showed superior gastrointestinal safety profile along with reduction in LPO as compared with that of the ibuprofen.

Full text of DOI:10.3109/14756366.2010.548810

Journal of Enzyme Inhibition and Medicinal Chemistry, 2011; 26(5): 742–748
© 2011 Informa UK, Ltd.
ISSN 1475-6366 print/ISSN 1475-6374 online
DOI: 10.3109/14756366.2010.548810
Short CoMMunICatIon
Synthesis and biological evaluation of some new pyridazinone
derivatives
Asif Husain1, Sushma Drabu1, Nitin Kumar1, M. Mumtaz Alam1, and Aftab Ahmad2
1Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Hamdard University, New Delhi, India and 2Faculty of
Pharmaceutical Sciences, Jodhpur National University, Jodhpur, Rajasthan, India
abstract
A series of pyridazinone derivatives (19–34) were synthesized with an aim to synthesize safer anti-inflammatory
agents. The compounds were evaluated for their anti-inflammatory, analgesic, ulcerogenic and lipid peroxidation
(LPO) actions. The percentage inhibition in edema at different time intervals indicated that compounds 20, 26, 28
and 34 exhibited good anti-inflammatory potential, comparable with that of ibuprofen (85.77%) within a range of
67.48–77.23%. The results illustrate that 5-(4-fluoro-benzyl)-3-(4-chloro-phenyl)-1,6-dihydro-6-pyridazinone (26) and
5-(4-chloro-benzyl)-3-(4-chloro-phenyl)-1,6-dihydro-6-pyridazinone (20) showed best anti-inflammatory activity.
Furthermore, activity is more in case of chloro substitution as compared with methyl-substitution. The compounds
synthesized were also evaluated for their ulcerogenic and LPO action and showed superior gastrointestinal safety
profile along with reduction in LPO as compared with that of the ibuprofen.
Keywords: Pyridazinone, anti-inflammatory, analgesic, lipid peroxidation
Introduction
(emorfazone) and its derivatives have emerged as being of
Non-steroidal anti-inflammatory drugs (NSAIDs) are
the most commonly prescribed medications in the
world for the treatment of pain, fever and inflamma-
tion and disorders particularly arthritis1,2. Long-term
use of the NSAIDs such as ibuprofen, indomethacin and
naproxen exhibit gastric toxicity (gastrointestinal (GI)
ulceration, bleeding) and nephrotoxicity3. e search for
safer NSAIDs continues with the failure of anticipated
‘Ideal’ anti-inflammatory agents, the coxibs, on long-
term usage4,5. erefore, synthetic approaches based on
chemical modification of NSAIDs have been taken with
the aim of improving their safety profile.
Pyridazinones have been the subject of intensive syn-
theticinvestigations,becausetheypossessawidespectrum
ofpharmacologicalactivities.Various3-(2H)-pyridazinone
derivatives have been described and their cardiotonic6,7
anti-secretory and antiulcer8 as well as analgesic and
anti-inflammatory9,10 activities have been investigated.
4-Ethoxy-2-methyl-5-morpholino-3(2H)-pyridazinone
particular interest. Derivatives of N-substituted-4,6-diaryl-
3-pyridazinone have emerged as potential analgesic anti-
inflammatory and antipyretic agents11.
As part of the continuing study for new
anti-inflammatory and analgesics in our laboratories12,13
,
a new series of pyridazinone derivatives were synthe-
sized with an aim to obtain safer NSAIDs. e analgesic
and anti-inflammatory activities were investigated for
the title compounds by the acetic acid-induced writhing
test and the carrageenan-induced rat paw edema test,
respectively. Lipid peroxidation (LPO) and ulcerogenic
actions of some of the selected compounds were also
investigated and correlated.
Materials and methods
Chemistry
Melting points were determined in open capillary tubes
and are uncorrected. in-layer chromatography (TLC)
Address for Correspondence: Asif Husain, Asst. Professor, Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Hamdard
University, New Delhi-110 062, India. Tel.: +91-11-26059681, 26059688, ext. 5647, +91–989-1116086 (Mobile); Fax: +91-11-16988874. E-mail:
drasifhusain@yahoo.com, ahusain@jamiahamdard.ac.in
(Received 05 August 2010; revised 13 December 2010; accepted 13 December 2010)
742

Synthesis and biological evaluation of new pyridazinone derivatives 743
was carried out to monitor the reactions using silica
5-(4-Nitrobenzyl)-3-(4-chlorophenyl)-1,6-dihydro-6-
pyridazinone (21)
Yield: 53%, m.p. 197°C. H NMR (CDCl3) δ 3.71 (s, 2H,
CH2), 6.78 (s, 1H, H-4, pyridazinone ring), 7.37 (m, 4H,
H-2,3,5,6, benzyl ring), 7.53 and 7.62 (d each, 2xA2B2,
p-substituted phenyl), 10.93 (s, 1H, NH). MS: m/z
341(M+). IR (cm−1, KBr): 3173 (NH), 2936 (CH), 1672 (CO),
707 (CCl). Anal calcd. for C17H12ClN3O3: C, 59.75; H, 3.54;
N, 12.30. Found: C, 59.63; H, 3.57; N, 12.31.
gel (Merck No. 5554). e IR spectra were measured as
potassium bromide pellets using a Perkin-Elmer 1725X
1
1
spectrophotometer. H NMR spectra were recorded on
Bruker spectropsin DPX-300 MHz in CDCl3; chemical
shift (δ) values are reported in parts per million (ppm).
e splitting pattern abbreviations are as follows: s, sin-
glet; d, doublet; m, multiplet. Mass spectroscopic analy-
ses for compounds were performed on a JEOL JMS-D
300 instrument. Spectral data are consistent with the
assigned structures. e molecular ion for compounds
containing chloro-group was calculated according to
35Cl isotope. Elemental analyses were performed on a
Perkin-Elmer analyzer and were in range of 0.4% for
each element analyzed (C, H, N). Dry solvents were used
throughout.
5-(4-Hydroxybenzyl)-3-(4-chlorophenyl)-1,6-dihydro-
6-pyridazinone (22)
1
Yield: 57%, m.p. 192°C. H NMR (CDCl3) δ 3.59 (s, 2H,
CH2), 6.11 (m, 1H, OH), 7.06 (m, 2H, H-2,6, benzyl ring),
7.31 (s, 1H, H-4, pyridazinone ring), 7.47 and 7.71 (d
each, 2xA2B2, p-substituted phenyl), 7.49 (m, 2H, H-3,5,
benzyl ring), 9.41 (s, 1H, NH). MS: m/z 312(M+). IR (cm−1,
KBr): 3178 (NH), 2942 (CH), 1686 (CO), 722 (C–Cl). Anal
calcd. for C17H13ClN2O2: C, 65.29; H, 4.19; N, 8.96. Found:
C, 65.39; H, 4.17; N, 8.97.
3-(4-Chloro/methyl benozyl)propionic acid (1,2)
e compounds, 1 and 2, were synthesized according to
the reported method12.
5-(4-Methylbenzyl)-3-(4-chlorophenyl)-1,6-dihydro-6-
pyridazinone (23)
Yield: 59%, m.p. 186°C. H NMR (CDCl3) δ 2.27 (s, 3H,
CH3), 3.51 (s, 2H, CH2), 6.60 (s, 1H, H-4, pyridazinone
ring), 7.13 and 7.36 (d each, 2xA2B2, p-substituted benzyl
ring), 7.34 (m, 2H, H-3,5, phenyl ring), 7.61 (m, 2H, H-2,4,
phenyl ring), 10.93 (s, 1H, NH). MS: m/z 310(M+). IR (cm−1,
KBr): 3173 (NH), 2939 (CH), 1684 (CO), 708 (C–Cl). Anal
calcd. for C18H15ClN2O: C, 69.57; H, 4.86; N, 9.01. Found:
C, 69.53; H, 4.87; N, 9.03.
3-Arylidene-5-(4-chloro/methyl phenyl)-2(3H)-
furanones (3–18)
Compounds (3–18) were synthesized from 3-(4-chloro/
methyl benozyl)propionic acid (1,2) following the litera-
ture method12.
1
General procedure for the synthesis of
5-(substituted benzyl)-3-aryl-1,6-dihydro-6-
pyridazinones (19–34)
2(3H)-Furanones (3–18) (0.005 mole) and hydrazine
hydrate (1–2 mL) in n-propanol (5–6 mL) were refluxed
for 3 h. After refluxing, reaction mixture was poured
onto crushed ice, a precipitate was obtained, which was
filtered, dried and recrystallized from methanol to give
TLC pure 5-(substituted benzyl)-3-aryl-1,6-dihydro-6-
pyridazinone derivatives.
5-(4-Methoxybenzyl)-3-(4-chlorophenyl)-1,6-dihydro-
6-pyridazinone (24)
Yield: 61%, m.p. 169°C. H NMR (CDCl3) δ 3.42 (s, 2H,
CH2), 3.71 (s, 3H, OCH3), 6.81 (s, 1H, H-4, pyridazinone
ring), 7.13 and 7.36 (d each, 2xA2B2, p-substituted benzyl
ring), 7.59 (m, 2H, H-3,5, phenyl ring), 7.68 (m, 2H, H-2,4,
phenyl ring), 10.73 (s, 1H, NH). MS: m/z 326(M+). IR (cm−1,
KBr): 3167 (NH), 3002 (CH), 1675 (CO), 717 (C–Cl). Anal
calcd. for C18H15ClN2O2: C, 66.16; H, 4.63; N, 8.57. Found:
C, 66.23; H, 4.61; N, 8.55.
1
5-Benzyl-3-(4-chlorophenyl)-1,6-dihydro-6-
pyridazinone (19)
1
Yield: 59%, m.p. 174°C. H NMR (CDCl3) δ 3.78 (s, 2H,
CH2), 7.16 (s, 1H, H-4, pyridazinone ring), 7.14–7.48
(m, 5H, benzyl ring), 7.42 and 7.73 (d each, 2xA2B2,
p-substituted phenyl ring), 10.72 (s, 1H, NH). MS: m/z
296(M+). IR (cm−1, KBr): 3186 (NH), 2949 (CH), 1683 (CO),
718 (C–Cl). Anal calcd. for C17H13ClN2O: C, 68.81; H, 4.42;
N, 9.44. Found: C, 68.93; H, 4.45; N, 9.43.
5-(4-Hydroxy-3-methoxybenzyl)-3-(4-chlorophenyl)-
1,6-dihydro-6-pyridazinone (25)
Yield: 53%, m.p. 191°C. H NMR (DMSO) δ 3.48 (s, 2H,
CH2), 3.64 (s, 3H, OCH3), 6.53 (s, 1H, H-4, pyridazinone
ring), 7.07–7.29 (m, 3H, H-2,5,6, disubstituted benzyl
ring), 7.46 and 7.71 (d each, 2xA2B2, p-substituted phe-
nyl ring), 10.92 (s, 1H, NH). MS: m/z 342(M+). IR (cm−1,
KBr): 3173 (NH), 2951 (CH), 1680 (CO), 713 (C–Cl). Anal
calcd. for C18H15ClN2O3: C, 63.07; H, 4.41; N, 8.17. Found:
C, 62.97; H, 4.39; N, 8.19.
1
5-(4-Chlorobenzyl)-3-(4-chlorophenyl)-1,6-dihydro-6-
pyridazinone (20)
1
Yield: 58%, m.p. 196°C. H NMR (CDCl3) δ 3.95 (s, 2H,
CH2), 7.11 (s, 1H, H-4, pyridazinone ring), 7.23 (m, 4H,
H-2,3,5,6, benzyl ring), 7.32 and 7.41 (d each, 2xA2B2,
p-substituted phenyl), 10.97 (s, 1H, NH). MS: m/z
330(M+). IR (cm−1, KBr): 3179 (NH), 2942 (CH), 1688 (CO),
726 (CCl). Anal calcd. for C17H12Cl2N2O: C, 61.65; H, 3.65;
N, 8.46. Found: C, 61.55; H, 3.67; N, 8.47.
5-(4-Fluorobenzyl)-3-(4-chlorophenyl)-1,6-dihydro-6-
pyridazinone (26)
Yield:57%,m.p.190°C.1HNMR(CDCl3)δ3.61(s,2H,CH2),
7.05 (s, 1H, H-4, pyridazinone ring), 7.10 and 7.26 (d each,
© 2011 Informa UK, Ltd.

744 A. Husain et al.
2xA2B2, p-substituted benzyl ring), 7.41 and 7.60 (d each,
2xA2B2, p-substituted phenyl ring), 11.14 (s, 1H, NH). MS:
m/z 314(M+). IR (cm−1, KBr): 3191 (NH), 2944 (CH), 1682
(CO), 719 (C–Cl). Anal calcd. for C17H12ClFN2O: C, 64.87;
H, 3.84; N, 8.90. Found: C, 64.93; H, 3.85; N, 8.91.
5-(4-Methoxybenzyl)-3-(4-methylphenyl)-1,6-dihydro-
6-pyridazinone (32)
Yield: 43%, m.p. 192°C. H NMR (CDCl3) δ 2.37 (s, 3H,
CH3), 3.67 (s, 3H, OCH3), 3.97 (s, 2H, CH2), 6.79 (s, 1H,
H-4, pyridazinone ring), 7.37 and 7.82 (d each, 2xA2B2,
p-substituted benzyl ring), 7.49 (m, 2H, H-3,5, phenyl
ring), 7.72 (m, 2H, H-2,6, phenyl ring), 11.15 (s, 1H, NH).
MS: m/z 305(M+). IR (cm−1, KBr): 3186 (NH), 2944 (CH),
1682 (CO). Anal calcd. for C19H18N2O2: C, 74.49; H, 5.92; N,
9.14. Found: C, 74.53; H, 5.91; N, 9.13.
1
5-Benzyl-3-(4-methylphenyl)-1,6-dihydro-6-
pyridazinone (27)
1
Yield: 53%, m.p. 168°C. H NMR (CDCl3) δ 2.36 (s, 3H,
CH3), 3.59 (s, 2H, CH2), 6.86 (s, 1H, H-4, pyridazinone
ring), 7.02–7.43 (m, 5H, benzyl ring), 7.46 and 7.79 (d
each, 2xA2B2, p-substituted phenyl ring), 8.92 (s, 1H, NH).
MS: m/z 276(M+). IR (cm−1, KBr): 3185 (NH), 2952 (CH),
1676 (CO). Anal calcd. for C18H16N2O: C, 78.24; H, 5.84; N,
10.14. Found: C, 78.23; H, 5.87; N, 10.17.
5-(4-Hydroxy-3-methoxy-benzyl)-3-(4-methylphenyl)-
1,6-dihydro-6-pyridazinone (33)
1
Yield: 51%, m.p. 180°C. H NMR (DMSO) δ 2.39 (s, 3H,
CH3), 3.25 (s, 3H, OCH3), 3.92 (s, 2H, CH2), 6.82 (s, 1H,
H-4, pyridazinone ring), 7.04 (m, 1H, H-6, benzyl ring),
7.25 (m, 2H, H-3,5, phenyl ring), 7.53 (m, 1H, H-2, ben-
zyl ring), 7.68 (m, 2H, H-2,6, phenyl ring), 7.75 (m, 1H,
H-5, benzyl ring), 10.73 (s, 1H, NH). MS: m/z 321(M+). IR
(cm−1, KBr): 3189 (NH), 2952 (CH), 1687 (CO). Anal calcd.
for C19H18N2O3: C, 70.79; H, 5.63; N, 8.69. Found: C, 70.83;
H, 5.65; N, 8.67.
5-(4-Chlorobenzyl)-3-(4-methylphenyl)-1,6-dihydro-6-
pyridazinone (28)
Yield: 48%, m.p. 188°C. 1H NMR (CDCl3) δ2.38 (s, 3H, CH3),
3.94 (s, 2H, CH2), 7.24 (s, 1H, H-4, pyridazinone ring), 7.26
and 7.56 (d each, 2xA2B2, p-substituted benzyl ring), 7.28–
7.35 (m, 4H, H-2,3,5,6, phenyl ring), 10.69 (s, 1H, NH). MS:
m/z 310(M+). IR (cm−1, KBr): 3174 (NH), 2939 (CH), 1683
(CO), 718 (C–Cl). Anal calcd. for C18H15ClN2O: C, 69.57; H,
4.86; N, 9.01. Found: C, 69.53; H, 4.87; N, 8.99.
5-(4-Fluorobenzyl)-3-(4-methylphenyl)-1,6-dihydro-6-
pyridazinone (34)
1
Yield: 49%, m.p. 174°C. H NMR (CDCl3) δ 2.36 (s, 3H,
5-(4-Nitrobenzyl)-3-(4-methylphenyl)-1,6-dihydro-6-
pyridazinone (29)
CH3), 3.95 (s, 2H, CH2), 7.01 (s, 1H, H-4, pyridazinone
ring), 7.05 and 7.55 (d each, 2xA2B2, p-substituted
benzyl ring), 7.21–7.29 (m, 4H, H-2,3,6,5, phenyl ring),
11.42 (s, 1H, NH). MS: m/z 293(M+). IR (cm−1, KBr): 3183
(NH), 2948(CH), 1672 (CO). Anal calcd. for C18H15FN2O:
C, 73.45; H, 5.14; N, 9.52. Found: C, 73.43; H, 5.17; N,
9.53.
1
Yield: 47%, m.p. 189°C. H NMR (CDCl3) δ 2.37 (s, 3H,
CH3), 3.41 (s, 2H, CH2), 7.13 (s, 1H, H-4, pyridazinone
ring), 7.31 and 8.01 (d each, 2xA2B2, p-substituted ben-
zyl ring), 7.48 (m, 2H, H-3,5, phenyl ring), 7.81 (m, 2H,
H-2,4, phenyl ring), 11.13 (s, 1H, NH). MS: m/z 321(M+).
IR (cm−1, KBr): 3183 (NH), 2948 (CH), 1679 (CO). Anal
calcd. for C18H15N3O3: C, 67.28; H, 4.70; N, 13.08. Found:
C, 67.23; H, 4.71; N, 13.07.
Pharmacology
e protocol of the animal experiments was approved
by the Institutional Animal Ethics Committee. e
in-vivo anti-inflammatory activity of the synthesized
compounds (19–34) was evaluated by carrageenan-
induced rat paw edema method as described by Winter
et al.14. Ibuprofen was taken as standard drug for com-
parison. All the compounds were administered orally
and assayed at a dose of 20 mg/kg of body weight. e
compounds that exhibited good anti-inflammatory
activity (>65%) were further evaluated for their anal-
gesic activity using acetic acid-induced writhing
method15. e compounds that were screened for the
analgesic activity were also tested for their ulcerogenic
activity by the method reported by Cioli et al.16 and LPO
according to the method of Okhawa et al.17. e results
of the pharmacological evaluation have been listed in
Tables 1 and 2.
5-(4-Hydroxybenzyl)-3-(4-methylphenyl)-1,6-dihydro-
6-pyridazinone (30)
Yield: 47%, m.p. 196°C. H NMR (CDCl3) δ 2.54 (s, 3H,
CH3), 3.43 (s, 2H, CH2), 6.40 (s, 1H, H-4, pyridazinone
ring), 6.63 (m, 2H, H-2,6, phenyl ring), 6.66 (m, 2H, H-2,6,
benzyl ring), 6.79 (m, 2H, H-3,5, phenyl ring), 6.81 (m, 2H,
H-3,5, benzyl ring), 12.25 (s, 1H, NH). MS: m/z 291(M+).
IR (cm−1, KBr): 3173 (NH), 2957 (CH), 1685 (CO). Anal
calcd. for C18H16N2O2: C, 73.96; H, 5.52; N, 9.58. Found: C,
73.98; H, 5.51; N, 9.57.
1
5-(4-Methylbenzyl)-3-(4-methylphenyl)-1,6-dihydro-6-
pyridazinone (31)
1
Yield: 48%, m.p. 182°C. H NMR (CDCl3) δ 2.29 and 2.31
(s, each, 6H, 2xCH3), 3.67 (s, 2H, CH2), 6.51 (s, 1H, H-4,
pyridazinone ring), 7.31 and 7.85 (d each, 2xA2B2, p-sub-
stituted benzyl ring), 7.37 (m, 2H, H-3,5, phenyl ring),
7.59 (m, 2H, H-2,6, phenyl ring), 10.51 (s, 1H, NH). MS:
m/z 289(M+). IR (cm−1, KBr): 3182 (NH), 2940 (CH), 1676
(CO). Anal calcd. for C19H18N2O: C, 78.59; H, 6.25; N, 9.65.
Found: C, 78.53; H, 6.27; N, 9.67.
Anti-inflammatory activity
e synthesized compounds were evaluated for their
anti-inflammatory activity using carrageenan-induced
paw edema method of Winter et al.14. e experiment was
performed on Albino rats of Wistar strain of either sex,
Journal of Enzyme Inhibition and Medicinal Chemistry

Synthesis and biological evaluation of new pyridazinone derivatives 745
Table 1. Anti-inflammatory activity of the pyridazinone derivatives. 19–34.
% Inhibition SEMa
Compound
Control
Ibuprofen
19
R
R`
—
After 2 h
—
After 3 h
—
—
—
—
75.23 1.08
23.09 2.32***
54.52 2.16***
29.99 2.95***
85.77 0.75
36.99 2.03***
71.95 1.69***
44.51 1.80***
4-Cl
4-Cl
4-Cl
H
20
4-Cl
4-NO2
21
22
23
24
25
26
27
28
29
30
31
32
33
34
4-Cl
4-Cl
4-OH
4-CH3
27.62 2.26***
22.14 3.08***
27.62 2.35***
30.24 2.84***
58.57 1.04***
20.71 1.97***
48.81 2.25***
22.85 2.33***
21.19 1.93***
31.67 1.70***
38.09 2.14***
30 2.50***
41.05 1.65***
35.56 2.54***
42.47 1.73***
44.51 2.17***
77.23 0.68***
36.58 1.83***
67.48 1.68***
40.85 2.61***
38.41 2.63***
53.86 1.73***
60.57 1.65***
46.13 1.76***
68.90 1.12***
4-Cl
4-OCH3
4-OH; 3-OCH3
4-F
4-Cl
4-Cl
4-CH3
4-CH3
4-CH3
4-CH3
4-CH3
4-CH3
4-CH3
4-CH3
H
4-Cl
4-NO2
4-OH
4-CH3
4-OCH3
4-OH; 3-OCH3
4-F
46.19 1.50***
*p<0.05; **p<0.01; ***p<0.001.
aRelative to the standard (ibuprofen) and data were analyzed by one-way ANOVA followed by Turkey test for n=6.
Table 2. Analgesic activity along with ulcerogenic and lipid peroxidation effect of the pyridazinone derivatives (20, 26, 28 and 34).
Analgesic activity
Compound
Control
Ibuprofen
20
R
—
R`
—
Number of writhingsb
Inhibitory ratioa (%) Lipid peroxidationc
Severity indexb
0.239 0.007a***
42.34 1.45
—
0.00 0.00
16.34 0.67***
22.34 0.67***
21.5 0.76***
0.597 0.007ab***
0.833 0.105*
0.083 0.083
0.083 0.083
0.25 0.118
—
—
61.03 2.56
46.83 2.78***
48.97 2.33***
42.12 3.10***
44.22 2.42***
0.362 0.008ab***
0.342 0.004ab***
0.318 0.018ab***
0.418 0.005ab***
4-Cl
4-Cl
4-CH3
4-Cl
4-F
4-Cl
26
28
24.34 0.88***
34
4-CH3
4-F
0.25 0.118
23.5 0.76***
*p<0.05; **p<0.01; ***p<0.001.
aRelative to the standard (ibuprofen) and data were analyzed by one-way ANOVA followed by Turkey test for n=6.
bRelative to their respective control and data were analyzed by one-way ANOVA followed by Turkey test for n=6.
cLipid peroxidation activity is expressed as nanomoles of MDA per milligram of protein.
weighing 180–200 g. e animals were randomly divided
into two groups of six each. Group I was kept as control
and received only 0.5% carboxymethyl cellulose (CMC)
solution. Groups II was kept as standard and received
ibuprofen (20 mg/kg, p.o.). Carrageenan solution (0.1%
in sterile 0.9% NaCl solution) in a volume of 0.1 mL was
injected subcutaneously into the sub-plantar region of
the right hind paw of each rat, 30 min after the adminis-
tration of the test compounds (20 mg/kg) and standard
drugs. e paw volume was measured by saline displace-
ment shown on screen of digital Plethysmometer (Ugo
Basile) at 2 and 3 h after carrageenan injection. us, the
edema volume in control group (Vc) and edema volume
in groups treated with test compounds (Vt) was measured
and the percentage inhibition of edema was calculated
using the formula:
Analgesic activity
Compounds that showed good anti-inflammatory
activity (>65%) were also screened for analgesic activ-
ity. Analgesic activity was done by acetic acid induce
writhing method15. Swiss albino mice (25–30 g) of either
sex were divided into different groups of six each. A 1%
aqueous acetic acid solution (intraperitoneal injection in
a volume of 0.1 mL) was used as writhing induced agent.
Mice were kept individually in the test cage, before acetic
acid injection and habituated for 30 min. Screening of
analgesic activity was performed after oral administra-
tion of test drugs at a dose of 20mg/kg. Group I was taken
as control and received CMC suspension only, group II
received reference drug ibuprofen and rest of the groups
were treated with test drugs (20 mg/kg) suspended in
1.0% CMC orally. After 1 h of drug administration 0.10 mL
of 1% acetic acid solution was given to mice intraperito-
neally. Stretching movements consisting of arching of
Anti-inflammatory activity (% inhibition) = {(Vc
− Vt)/Vc} × 100
© 2011 Informa UK, Ltd.

746 A. Husain et al.
the back, elongation of body and extension of hind limbs
were counted for 5–15 min of acetic acid injection. e
analgesic activity was expressed in terms of percentage
inhibition.
compounds,
1,6-dihydro-6-pyridazinone
i.e.
5-(substituted-benzyl)-3-aryl-
derivatives (19–34).
2(3H)-Furanones (3–18) were prepared using 3-(4-sub-
stituted-benzoyl)propionic acid (1–2) following the pre-
viously reported methods of modified Perkin’s reaction
in higher yield12. e 3-(4-substituted-benzoyl)propionic
acid (1, 2) was synthesized according to Friedal Craft’s
acylation reaction condition using substituted benzene
(Scheme 1).
In the IR spectral data, all the compounds showed
peaks each around 3180 (NH), 2940 (CH), 1685 (CO)
cm−1. e 1H NMR spectra of the synthesized compounds
showed characteristic peaks at appropriate δ values.
e CH proton of fourth position of pyridazinone ring
appeared as singlet around δ 6.60–7.26. e CH2 proton
appeared around δ 3.23–4.47 in aliphatic region. e NH
proton appeared as a broad singlet appeared around δ
9.23–12.89. e structure of the compounds was fur-
ther supported by mass spectral data. e synthesized
compounds gave M+ peak in reasonable intensities.
e molecular ion or other related ions produced the
appropriate isotopic abundances due to the presence of
chlorine atom(s).
% Analgesic activity = {(n − n′)/n} × 100
where n is mean number of writhes of control group and
n′ the mean number of writhes of test group.
Acute ulcerogenesis
Acute ulcerogenesis test was done according to the
study of Cioli et al.16 Albino rats (150–200 g) were divided
into different groups consisting of six animals in each
group. Ulcerogenic activity was evaluated after oral
administration of test compounds or ibuprofen at the
dose of 60 mg/kg. Control rats received oral administra-
tion of vehicle (suspension of 1% methyl cellulose). Food
but not water was removed 24 h before administration
of the test compounds. After the drug treatment, the rats
were fed with normal diet for 17h and then killed. e
stomach was removed and opened along the greater cur-
vature, washed with distilled water and cleaned gently
by dipping in normal saline. e mucosal damage was
examined by means of a magnifying glass. For each stom-
ach, the mucosal damage was assessed according to the
following scoring system: 0.5: redness, 1.0: spot ulcers,
1.5: hemorrhagic streaks, 2.0: ulcers > 3 but <5, 3.0: ulcers
> 5. e mean score of each treated group minus the
mean score of control group was regarded as severity
index of gastric mucosal damage.
Pharmacological evaluation
e pharmacological evaluation showed that the com-
pounds are capable anti-inflammatory (Table 1) and
analgesic agents with lesser GI toxicity and reduced LPO
(Table 2).
e results illustrate that 5-(4-fluoro-benzyl)-3-(4-
chloro-phenyl)-1,6-dihydro-6-pyridazinone (26) and
5-(4-chloro-benzyl)-3-(4-chloro-phenyl)-1,6-dihydro-6-
pyridazinone (20) showed best anti-inflammatory activ-
ity, having maximum percentage inhibition in edema
at different time intervals. eir percentage inhibition
in edema at different time intervals, i.e. 77.23% (com-
pound 26) and 71.95% (compound 20) was comparable
with that of ibuprofen (85.77%). In addition, two more
compounds, 28 and 34, showed 67.48 and 68.90% inhibi-
tion, respectively. e remaining derivatives too possess
Lipid peroxidation
LPO in the gastric mucosa was determined according to
the method of Ohkawa et al.17. After screening for ulcero-
genic activity, the gastric mucosa was scraped with two
glass slides and 10% of that tissue was homogenized at
10,000g in 1.8 mL of 1.15% ice-cold KCl solution. One
millilitre of suspension medium was taken from the
supernatant, 0.5 mL of 30% trichloroacetic acid fol-
lowed by 0.5 mL of 0.8% thiobarbituric acid reagent were
added to it. e tubes were covered with aluminium
foil and kept in a shaking water bath for 30 min at 80°C.
After 30 min, tubes were taken out and kept in ice cold
water for 10 min. ese were then centrifuged at 3000g
for 15 min. e absorbance of supernatant was read at
540 nm at room temperature against the blank on UV
spectrophotometer.
0.5
0.45
0.4
0.35
0.3
y = 0.0734x − 0.0047
R2 = 0.9962
0.25
0.2
0.15
0.1
0.05
0
e standard curve was used for estimating the con-
centration of malondialdehyde (MDA) prepared by using
1,1,3,3-tetraethoxypropane (Figure 1). e results are
presented as nanomoles of MDA per milligram of protein
(Table 2).
0
1
2
3
4
5
6
7
result and discussion
TEP (µg)
Chemistry
2(3H)-Furanones
Figure 1. Calibration standard curve for TBARS concentration.
(3–18)
on
reaction
with
hydrazine hydrate in n-propanol gave the title
Journal of Enzyme Inhibition and Medicinal Chemistry

Synthesis and biological evaluation of new pyridazinone derivatives 747
R'
O
(i)
(ii)
.
OH
O
O
R
R
O
R
Substituted Benzene
3-(4-Substituted-benzoyl)propionic acid (1-2)
2(3H)-Furanone (3-18)
(19) R=4-Cl; R'=H, (20) R=4-Cl; R'=4-Cl, (21) R=4-Cl; R'=4-NO,
(222) R=4-Cl; R'=4-OH, (23) R=4-Cl; R'=4-CH3 , (24) R=4-Cl; R'=4-OCH3,
(25) R=4-Cl; R'=4-OH-3-OCH3, (26) R=4-Cl; R'=4-F, (27) R=4-CH3; R'=H,
(28) R=4-CH3; R'=4-Cl, (29) R=4-CH3; R'=4-NO2 , (30) R=4-CH3; R'=4-OH,
(31) R=4-CH3; R'=4-CH3, (32) R=4-CH3; R'=4-OCH3 ,
(iii)
R'
(33) R=4-CH3; R'=4-OH-3-OCH3, (34) R=4-CH3; R'=4-F
R
N
N
H
O
Pyridazinone (19-34)
Reagents and Condition: (i) Succinic anhydride, Anhyd. AlCl3; (ii) Aryl Aldehyde, Triethylamine
Acetic anhydride, reflux; (iii) Hydrazine hydrate, Propanol, reflux
Scheme 1. Protocol for synthesis of title compounds.
anti-inflammatory activity but less than of the standard,
ibuprofen.
high LPO 0.597 0.007, whereas control group showed
0.239 0.007.
e anti-inflammatory activity enhanced in the
presence of electronegative substitution (chloro) and
diminished in the presence of electropositive substitu-
tion (methyl) indicating that the activity declines with
replacement of electronegative group to electropositive
group.
us, these compounds showed superior GI safety
profile along with reduction in LPO in comparison with
ibuprofen. e other tested compounds also exhibited
better GI safety profile as compared with the standard
drug ibuprofen.
In
summary,
5-(4-fluoro-benzyl)-3-(4-chloro-
(26) and
Test
compounds
that
exhibited
good
phenyl)-1,6-dihydro-6-pyridazinone
anti-inflammatory activity 20, 26, 28 and 34 were fur-
ther evaluated for their analgesic, ulcerogenic and LPO
actions. e results of analgesic activity indicated that
compounds 20 and 26 showed 46.83% and 48.97% pro-
tection, respectively, against acetic acid-induced writh-
ingsandthispercentageprotectionwascomparablewith
that of ibuprofen (61.03%). Compounds 28 and 34 also
showed good analgesic activity with 42.12% and 44.22%
protection, respectively. According to structure–activity
relationship (SAR), it is clear that 5-(4-fluoro-benzyl)-
3-(4-chloro-phenyl)-1,6-dihydro-6-pyridazinone (26)
5-(4-chloro-benzyl)-3-(4-chloro-phenyl)-1,6-dihydro-6
-pyridazinone (20) were found to have dual functional,
i.e. anti-inflammatory and analgesic properties and
hence, can be a promising class of compounds with an
interesting pharmacological profile. It is conceivable that
these derivatives could be further modified to develop
potent and safer anti-inflammatory and analgesic agents.
Further studies to acquire more information about quan-
titative SAR are in progress in our laboratory.
acknowledgements
and
5-(4-chloro-benzyl)-3-(4-chloro-phenyl)-1,6-
dihydro-6-pyridazinone (20) were found to be a good
anti-inflammatory agents with significant analgesic
activity.
The tested compounds showed low ulcerogenic
activity ranging from 0.1 0.1 to 0.2 0.12, whereas
the standard drug ibuprofen showed high severity
index of 0.8 0.1. The maximum reduction in ulcero-
genic activity (0.1 0.1) was found in the compounds
26 and 20. The LPO was measured as nanomoles of
MDA per milligram of protein. Ibuprofen exhibited
e authors thank UGC for financial assistance. anks
are also due to In-charge animal house, Jamia Hamdard,
New Delhi, for providing animals for pharmacological
studies.
Declaration of interest
e authors report no conflicts of interest. e authors
alone are responsible for the content and writing of the
paper.
© 2011 Informa UK, Ltd.

748 A. Husain et al.
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