Synthesis of new pyridazinones substituted by 4-arylpiperazin-1-yl-carbonylalkyl moieties and their analgesic properties in mice

Article abstract of DOI:10.1211/146080800128736259

A new series of 4,6-diaryl pyridazin-3-ones substituted on the nitrogen atom in the 2-position have been prepared and evaluated for their analgesic activity in the phenylbenzoquinone-induced abdominal constriction test. Most of the eighteen compounds disp

Full text of DOI:10.1211/146080800128736259

Pharm. Pharmacol. Commun. 2000, 6: 387±396
Received April 18, 2000
Accepted June 22, 2000
# 2000 Pharm. Pharmacol. Commun.
Synthesis of New Pyridazinones Substituted by 4-Arylpiperazin-
1-yl-carbonylalkyl Moieties and their Analgesic Properties
in Mice
PASCAL COUDERT, CATHERINE RUBAT*, FLORENCE ROHET, FERNAND LEAL, JOSEPH FIALIP*
AND JACQUES COUQUELET
Â
Groupe de recherche en Pharmacochimie ± Laboratoire de Chimie Therapeutique and *Laboratoire de
Pharmacologie, Faculte de Pharmacie, Universite d' Auvergne, 63001 Clermont-Ferrand Cedex, France
Â
Â
Abstract
A new series of 4,6-diaryl pyridazin-3-ones substituted on the nitrogen atom in the 2-
position have been prepared and evaluated for their analgesic activity in the phenylbenzo-
quinone-induced abdominal constriction test.
Most of the eighteen compounds displayed signi®cant antinociceptive effects with ED50
1
ranging from 22Á2 to 76Á7 mg kg , intraperitoneally. Pyridazinones 4f and 5f were among
the most interesting derivatives with marked analgesic properties associated to weak
1
neurosedative activities (ED50 ˆ 83Á3 and 247Á2 mg kg , i.p., respectively).
Further investigations provided support for the hypothesis that the antinociceptive action
of 4f and 5f probably resulted from interactions with opioidergic, 5-hydroxytryptaminergic
and noradrenergic pathways.
The search for new analgesic drugs devoid of irri-
tant side effects on gastric mucosa or profound
respiratory depression, nausea, constipation and
physical dependence found with non-steroidal anti-
in¯ammatory drugs and opiates, respectively, con-
tinues to be an active area of investigation in
medicinal chemistry. There have been many reports
on the antinociceptive activity of several pyrid-
azinones (Loriga et al 1979; Rubat et al 1992; Dal
Piaz et al 1996). Previously we have synthesized
some 4,6 diarylpyridazines substituted on the 3-
position by arylpiperazinyl moieties and described
their analgesic effects (Rohet et al 1997). Thus, we
were prompted to take an active interest in new 4,6
diarylpyridazin-3-ones substituted on the 2-position
by arylpiperazinyl moieties.
5-hydroxytryptaminergic (5-HTergic) and=or nor-
adrenergic pathways in their biological response.
Materials and Methods
Chemistry
Melting points were determined on a Reichert
apparatus and were uncorrected. The infrared (IR)
spectra were recorded on a Beckman 4240
spectrophotometer (Beckman Instruments, Gagny,
France) in KBr pellets. The proton and carbon
nuclear magnetic resonance (1H NMR and 13C
NMR) spectra were run on a BruÈker AC-400
(400 MHz) spectrometer (Wissembourg, France)
with tetramethylsilane as the internal standard. The
purity of the compounds was determined by TLC
on precoated plates (silica gel 60 F 254, Merck,
Darmstadt, Germany) and spots were visualized
with UV light or iodine. Elemental analyses were
performed at the Service Central d'Analyses
(Vernaison, France). Starting materials were pur-
chased from Acros (Noisy Le Grand, France).
We report here the synthesis of these original
derivatives and their analgesic activities in the
phenylbenzoquinone-induced abdominal constric-
tion test in mice. The most potent pyridazinones
were selected to investigate possible opioidergic,
Â
Correspondence: P. Coudert, Laboratoire de Chimie Thera-
Â
peutique, Faculte de Pharmacie, 28 place H. Dunant, 63001
Clermont-Ferrand Cedex, France.

388
PASCAL COUDERT ET AL
Figure 1. Reaction of N-substituted pyridazinones with arylpiperazines led to compounds 4 and 5.
4-Phenyl-6-aryl-2-[(4-arylpiperazin-1-yl)-
carbonylalkyl]-pyridazin-3-ones (4±5)
different groups was randomized and the experi-
ments were carried out under blind conditions. The
IASP Committee for Research and Ethical Issues
Guidelines (Zimmermann 1983) were followed.
The starting esters (2 or 3; Figure 1) were prepared
as described by Coudert et al (1989). A suspension
of the appropriate ester (0Á005 mol) 2 or 3 in an
excess of suitable arylpiperazine (20 mL) was
heated at 160ꢀC for 24 h in a bomb apparatus. After
cooling, the piperazine was distilled under vacuum.
The resulting solid was puri®ed by chromatography
on a silica gel column (35±70 m; eluent, ethyl
acetate±hexane).
Acute toxicity studies. The compounds were admi-
nistered intraperitoneally at graded doses of 100,
1
200, 400, 600 and 800 mg kg . Animals were kept
under observation for eight days to detect any sign
of toxicity.
Phenylbenzoquinone-induced abdominal constric-
tion test. A 0Á02% solution (ethanol : water, 5 : 95)
of phenylbenzoquinone (Eastman Kodak, Roches-
ter, NY), maintained at 37ꢀC, was intraperitoneally
injected into mice 30 min after the intraperitoneal
administration of drugs. The number of abdominal
constrictions of each animal was counted between
the ®fth and the ®fteenth minute after the injection
of the irritant agent (Siegmund et al 1957; Linee
1972).
Behavioural studies
All compounds were administered intraperitoneally
in saline (0Á9% NaCl). Swiss male mice (18±22 g;
 Â
Depre, Saint-Doulchard, France) were used in all
experiments and were kept in groups of 10 in a
temperature-controlled room with a 12-h light=dark
cycle. Food and water were freely available during
the experiment. The allocation of animals in the

SYNTHESIS AND ANALGESIC PROPERTIES OF NEW PYRIDAZINONES
389
Locomotor activity. The number of crossed photo-
cell beams was recorded 30 min after drug admin-
istration (i.p.) in mice individually placed for
10 min in a photocell actimeter (Apelex, Massy,
France) (Boissier & Simon 1965).
animals to maximal electroshock through corneal
electrodes. Protection against seizures was de®ned
as the abolition of the hind limb tonic extensor
component of seizures (Krall et al 1978).
Hot-plate test. Animals were placed on a copper
plate (Apelex, Massy, France) maintained at a
constant temperature of 56ꢀC. The time necessary
to induce the licking re¯ex of the forepaws was
then recorded. Measurements were carried out 30-
min later (Bell et al 1991; Viaud et al 1995). A
standard 40-s cut-off time was used as a maximal
effect.
Binding study
Preparation of membranes. Male Sprague±Dawley
rats (325±375 g) were decapitated and whole brains
were removed rapidly and placed on ice. Brains
were homogenized in 10 vols (w=v) ice-cold Tris-
HCl buffer (50 mM, pH 7Á7; Sigma, MontlucËon,
France) with an ultra Turrax homogenizer (Janke
and Kunkel). The homogenate was centrifuged at
48Á340 g (Sorvall RC2-B) for 15 min at 4ꢀC, the
pellet resuspended in 10 vols fresh Tris-HCl buffer
and incubated at 37ꢀC for 40 min to dissociate any
receptor-bound endogenous opioid peptides. The
homogenate was centrifuged again as described
above. The ®nal pellet was used immediately and
resuspended in fresh Tris-HCl buffer to yield a ®nal
Potentiation of morphine analgesia. The protocol
used was the same as for the phenylbenzoquinone
1
Â
test. Morphine (0Á15 mg kg , s.c.) (Cooperation
Pharmaceutique FrancËaise, Melun, France) was
injected at the same time as the drugs, 30 min
before the test (Fialip et al 1989).
1
protein concentration of 0Á5 mg mL . Protein con-
Antagonism of drug antinociception by naloxone.
The protocol used for the evaluation of the effect of
naloxone (Narcan, Du Pont de Nemours, Paris,
France) on drug-induced analgesia was similar to
that described for the phenylbenzoquinone test.
centrations were determined according to Lowry
et al (1951) with bovine serum albumin as standard.
Competition experiment. Each assay tube contained
Tris-HCl buffer (50mM, pH 7Á7) or buffer plus test
drug (50 mL), [3H]DAMGO (50 mL, ®nal concen-
tration: 1Á17 nM; Amersham, Les Ullis, France)
and tissue (1000 mL; ®nal assay concentration,
1
Naloxone (1 mg kg , s.c.) was injected 5 min
before intraperitoneal administration of the phenyl-
benzoquinone solution (Nevinson et al 1991).
1
0Á5 mg mL ) to a ®nal volume of 2000 mL.
Potentiation of drug antinociception by 5-hydroxy-
tryptophan (5-HTP). The protocol used was
adapted from the technique of Vonvoigtlander et
al (1984). Experiments were carried out in a similar
manner as in the phenylbenzoquinone test. Carbi-
Total binding was determined in the absence of
the test drugs, non-speci®c binding was determined
with naloxone (1 mM; Sigma, Saint-Quentin Fala-
vier, France) and speci®c binding was the differ-
ence between the two values. Incubation at 25ꢀC
was terminated after 1 h by rapid ®ltration over
Whatman GF=B glass ®bre ®lters wetted with
buffer. The ®lters were washed twice with 5 mL
Tris-HCl buffer (50 mM) and incubated overnight
in ultima gold M.V. (Packard) (4Á5 mL). Radio-
activity was measured using a Packard 1900
Tricarb counter (counting ef®ciency by external
standardisation 54±57%).
1
dopa (25 mg kg , i.p.) (ICN Biomedicals, Orsay,
France) was administered, followed 30 min later by
1
5-HTP (50 mg kg , i.p.) (ICN Biomedicals, Orsay,
France), followed 15 min after that by the drugs.
Twenty minutes later, the analgesic test was
performed with administration of phenylbenzo-
quinone solution.
Antagonism of drug antinociception by yohimbine.
Thirty minutes after simultaneous administration of
1
drugs (i.p.) and yohimbine (1 mg kg , i.p.; Sigma,
Data analysis
MontlucËon, France), phenylbenzoquinone solution
was given intraperitoneally. Five minutes after
injection of the irritant, mice were observed for
abdominal constriction for a 10-min period (Lut-
tinger et al 1985).
Behavioural studies. Statistical analysis of the
results was performed using the method of
Schwartz (1984). The ED50 values were deter-
mined by the method of Carmines et al (1980).
The signi®cance of pharmacological data expressed
as meanÆ s.e. were analysed by the Student's t-test.
Data from the maximal electroshock seizure test
Effects on maximal electroshock seizures. Test
drugs were administered 30 min before subjecting

390
PASCAL COUDERT ET AL
Table 1. Physicochemical data of compounds 4 and 5.
Compound
R1
R2
n
Yield
mp (ꢀC)
Molecular formula (MW)
Elemental analysisa
C
H
N
F
Cl
4a
4b
4c
4d
4e
4f
H
H
H
F
H
3Cl
3CF3
H
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
42
40
32
34
39
36
38
30
39
26
41
33
28
33
34
34
24
25
156
162
130
156
148
154
164
158
148
132
142
166
172
146
130
148
136
120
C28H26N4O2 (450Á6)
74Á63
74Á65
69Á32
69Á30
67Á16
67Á05
71Á76
71Á82
66Á84
66Á70
64Á91
65Á17
69Á32
68Á97
64Á72
64Á80
62Á97
62Á89
74Á96
74Á54
69Á78
69Á95
67Á65
68Á02
72Á17
71Á98
67Á35
67Á03
65Á44
65Á35
69Á78
69Á53
65Á27
65Á02
63Á54
63Á67
5Á83
6Á06
5Á21
5Á35
4Á87
5Á10
5Á39
5Á52
4Á82
4Á94
4Á53
4Á75
5Á21
5Á38
4Á67
4Á92
4Á38
4Á52
6Á09
6Á47
5Á46
5Á34
5Á12
4Á90
5Á65
5Á73
5Á08
5Á21
4Á77
4Á83
5Á46
5Á62
4Á92
4Á67
4Á63
4Á75
12Á44
12Á15
11Á55
11Á44
10Á81
10Á51
11Á96
11Á58
11Á14
11Á53
10Á44
10Á32
11Á55
11Á35
10Á79
10Á57
10Á13
10Á25
12Á06
11Á66
11Á23
11Á07
10Á52
10Á37
11Á61
11Á77
10Á84
10Á62
10Á18
10Á04
11Á23
11Á15
10Á50
10Á72
9Á88
C28H25N4O2Cl (485Á1)
C29H25N4O2F3 (518Á6)
C28H25N4O2F (468Á6)
C28H24N4O2FCl (503Á1)
C29H24N4O2F4 (536Á6)
C28H25N4O2Cl (485Á1)
C28H24N4O2Cl (519Á6)
C29H24N4O2F3Cl (553Á1)
C29H28N4O2 (464Á6)
7Á32
7Á28
10Á99
11Á18
4Á06
4Á20
F
3Cl
3CF3
H
3Á78
7Á06
6Á87
3Á56
F
14Á16
13Á77
4g
4h
4i
Cl
Cl
Cl
H
H
H
F
7Á32
7Á67
3Cl
3CF3
H
13Á66
13Á53
6Á42
10Á31
10Á03
6Á55
5a
5b
5c
5d
5e
5f
3Cl
3CF3
H
C29H27N4O2Cl (499Á1)
C30H27N4O2F3 (532Á6)
C29H27N4O2F (482Á6)
C29H26N4O2FCl (517Á1)
C30H26N4O2F4 (550Á6)
C29H27N4O2Cl (499Á1)
C29H26N4O2Cl2 (533Á6)
C30H26N4O2ClF3 (567Á1)
7Á12
7Á20
10Á70
10Á75
3Á94
3Á87
F
3Cl
3CF3
H
3Á67
6Á87
7Á18
3Á73
F
13Á80
13Á92
5g
5h
5i
Cl
Cl
Cl
7Á12
7Á28
3Cl
3CF3
13Á12
13Á37
6Á26
10Á05
10Á17
9Á62
6Á17
aUpper values: calculated; lower ones: found.
were analysed by means of the chi-square test with
Yate's correction.
lished from analytical and spectral data (Tables 1, 2
and 3).
The compounds were ®rst evaluated for their
toxicity. With the exception of a decrease of
locomotor activity that disappeared after 4 h, no
serious behavioural effects were observed even at
Binding studies. Data were analysed by the
program EBDA (McPherson 1983, 1985) and
LIGAND (Munson & Rodbard 1980).
1
doses up to 800 mg kg intraperitoneally.
The analgesic activity of pyridazinones 4 and 5
was evaluated using the phenylbenzoquinone test.
They were compared with the classical analgesic
drugs paracetamol, aspirin, noramidopyrine, and
morphine. Trazodone (Figure 2) was also used as a
reference compound because of its closely related
chemical structure to the pyridazinones (Riblet &
Taylor 1981). Tramadol (Figure 2) was chosen as a
reference compound because of its original and
non-classical mechanism of analgesic activity
(Raffa et al 1992; Desmeules et al 1996). As can be
seen from Table 4, most of the compounds, with
Results and Discussion
Preparation of pyridazinones 2 and 3 was achieved
by a nucleophilic substitution of the hydrogen atom
attached in the 2-position of the pyridazine ring by
ethyl bromoacetate or ethyl 3-bromopropionate,
respectively. Reaction of these resulting N-sub-
stituted pyridazinones with arylpiperazines led to
compounds 4 and 5 (Figure 1). The structure of
derivatives 4a±i and 5a±i (Table 1) was estab-

SYNTHESIS AND ANALGESIC PROPERTIES OF NEW PYRIDAZINONES
391
1
Table 2. Characteristic IR frequencies and H NMR chemical shifts of compounds 4 and 5.
1
IR (KBr) n (cm
)
Compound
CˆO
CˆC, CˆN
1H NMR (CDCl3) d ppm
4a
4b
4c
4d
4e
4f
1640
1630
1649
1640
1640
1645
1640
1640
1640
1650
1640
1640
1640
1640
1645
1645
1640
1647
1590, 1480, 1440
1580, 1480, 1450
1595, 1480, 1446
1600, 1500, 1440
1580, 1500, 1440
1600, 1500, 1450
1590, 1490, 1460
1580, 1480, 1440
1590, 1485, 1440
1590, 1490, 1440
1590, 1485, 1440
1590, 1500, 1440
1600, 1500, 1440
1585, 1505, 1440
1590, 1480, 1440
1598, 1493, 1446
1590, 1500, 1445
1600, 1493, 1447
7Á86±6Á91 (m, 16H, 3 C6H5 ‡ HCˆ), 5Á17 (s, 2H, 2 H-5), 3Á82±3Á67 (m, 4H, 2 H-
8 ‡ 2 H-80), 3Á25±3Á17 (m, 4H, 2 H-7 ‡ 2 H-70)
7Á90±6Á70 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 5Á20 (s, 2H, 2 H-5), 3Á85±3Á65 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á25±3Á15 (m, 4H, 2 H-7 ‡ 2 H-70)
7Á90±7Á05 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 5Á20 (s, 2H, 2 H-5), 3Á85±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á35±3Á20 (m, 4H, 2 H-7 ‡ 2 H-70)
7Á90±6Á95 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 5Á20 (s, 2H, 2 H-5), 3Á85±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á30±3Á20 (m, 4H, 2 H-7 ‡ 2 H-70)
7Á85±6Á80 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HCˆ), 5Á15 (s, 2H, 2 H-5), 3Á75±3Á60 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á25±3Á15 (m, 4H, 2 H-7 ‡ 2 H-70)
7Á85±7Á05 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HCˆ), 5Á20 (s, 2H, 2 H-5), 3Á85±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á35±3Á25 (m, 4H, 2 H-7 ‡ 2 H-70)
4g
4h
4i
7Á85±6Á90 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 5Á17 (s, 2H, 2 H-5), 3Á83±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á28±3Á19 (m, 4H, 2 H-7 ‡ 2 H-70)
7Á85±6Á80 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HCˆ), 5Á20 (s, 2H, 2 H-5), 3Á85±3Á75 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á30±3Á20 (m, 4H, 2 H-7 ‡ 2 H-70)
7Á84±7Á05 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HCˆ), 5Á15 (s, 2H, 2 H-5), 3Á82±3Á69 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á30±3Á22 (m, 4H, 2 H-7 ‡ 2 H-70)
5a
5b
5c
5d
5e
5f
7Á85±6Á90 (m, 16H, 3 C6H5 ‡ HCˆ), 4Á70 (t, 2H, 2 H-5), 3Á80±3Á65 (m, 4H, 2 H-
8 ‡ 2 H-80), 3Á15±3Á10 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á00 (t, 2H, 2 H-50)
7Á85±6Á75 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 4Á70 (t, 2H, 2 H-5), 3Á80±3Á65 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á15±3Á10 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á00 (t, 2H, 2 H-50)
7Á85±7Á05 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 4Á70 (t, 2H, 2 H-5), 3Á85±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á25±3Á20 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á05 (t, 2H, 2 H-50)
7Á85±6Á90 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 4Á70 (t, 2H, 2 H-5), 3Á80±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á20±3Á15 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á05 (t, 2H, 2 H-50)
7Á85±6Á75 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HC ˆ ), 4Á65 (t, 2H, 2 H-5), 3Á80±3Á65 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á20±3Á15 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á05 (t, 2H, 2 H-50)
7Á85±7Á05 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HCˆ), 4Á70 (t, 2H, 2 H-5), 3Á85±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á25±3Á20 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á05 (t, 2H, 2 H-50)
7Á85±6Á90 (m, 15H, 2 C6H5 ‡ C6H4 ‡ HCˆ), 4Á70 (t, 2H, 2 H-5), 3Á85±3Á70 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á20±3Á15 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á05 (t, 2H, 2 H-50)
7Á85±6Á75 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HCˆ), 4Á70 (t, 2H, 2 H-5), 3Á80±3Á65 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á15±3Á10 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á00 (t, 2H, 2 H-50)
7Á80±6Á95 (m, 14H, C6H5 ‡ 2 C6H4 ‡ HCˆ), 4Á75 (t, 2H, 2 H-5), 3Á75±3Á60 (m,
4H, 2 H-8 ‡ 2 H-80), 3Á15±3Á10 (m, 4H, 2 H-7 ‡ 2 H-70), 3Á00 (t, 2H, 2 H-50)
5g
5h
5i
the exception of pyridazinones 4d and 5g, exhibited
marked antinociceptive effects with ED50 values
were more powerful than compounds 4a±i. Thus,
the lengthening of the phenylpiperazinyl side chain
increased antinociceptive activity.
1
ranging from 22Á2 to 76Á7 mg kg , intraperi-
toneally. Thus, these derivatives possess a more
potent activity than paracetamol (ED50 ˆ 231Á3 mg
Compounds 4d, 4g and 5d, 5g were not more
potent than 4a and 5a, respectively, suggesting that
substitution of the phenyl ring of the pyridazinone
did not improve their activity. In contrast, sub-
stituents on the phenyl ring of the phenylpiperazine
moiety markedly increased activity vs non-
substituted derivatives. Precisely, substitution by an
m-tri¯uoromethyl group (4c, 4f, 4i, 5c, 5f, and 5i)
1
kg , i.p.). The less active derivatives were equi-
1
potent to noramidopyrine (ED50 ˆ 68Á5 mg kg ,
i.p.). Pyridazinones which possessed the strongest
analgesic properties presented ED50 values close to
1
that of trazodone (ED50 ˆ 10Á2 mg kg , i.p.). The
results showed that, on the whole, compounds 5a±i

392
PASCAL COUDERT ET AL
Table 3. 13C NMR chemical shifts of compounds 4 and 5.
13C NMR (CDCl3) d ppm
Compound
C1
C3
C4
C5
C50
C6
C7=C70
4a
4b
4c
4d
4e
4f
4g
4h
4i
5a
5b
5c
5d
5e
5f
5g
5h
5i
145Á3
145Á3
145Á4
144Á3
142Á2
144Á3
144Á1
144Á3
144Á1
144Á8
144Á9
144Á9
143Á9
143Á9
144Á0
143Á7
143Á7
143Á8
139Á8
139Á8
139Á9
139Á9
139Á8
140Á0
140Á0
140Á2
140Á0
139Á9
139Á9
140Á0
140Á0
140Á0
140Á1
140Á1
140Á1
140Á1
159Á6
159Á6
159Á6
159Á5
159Á4
159Á4
159Á4
159Á6
159Á4
159Á8
159Á4
159Á5
159Á3
159Á3
159Á3
159Á3
159Á3
159Á4
54Á1
54Á1
54Á1
54Á0
54Á0
54Á0
54Á1
54Á1
54Á0
50Á0
49Á4
50Á0
49Á4
50Á0
50Á0
50Á0
50Á0
50Á1
±
±
±
±
±
±
±
±
±
31Á7
31Á7
31Á8
31Á7
31Á7
31Á7
31Á7
31Á7
31Á7
164Á6
164Á7
164Á9
164Á7
164Á7
164Á8
164Á6
164Á7
164Á7
168Á8
168Á8
168Á9
168Á7
168Á8
168Á8
168Á8
168Á8
168Á8
42Á1±44Á9
41Á9±44Á7
42Á0±44Á8
42Á1±45Á0
41Á8±44Á7
41Á9±44Á8
42Á2±45Á0
42Á0±44Á9
41Á9±44Á7
41Á6±45Á5
41Á4±45Á3
41Á4±45Á4
41Á6±45Á5
41Á3±45Á3
41Á4±45Á3
41Á6±45Á5
41Á4±45Á3
41Á4±45Á4
Compound
C8=C80
C9
C10
C11
C12
C13
CF3
C2 ‡ Ar
4a
4b
4c
4d
4e
4f
4g
4h
4i
5a
5b
5c
5d
5e
5f
5g
5h
5i
49Á3±49Á6
48Á8±49Á0
48Á9±49Á0
49Á4±49Á7
48Á8±48Á7
48Á8±49Á0
49Á4±49Á7
48Á9±49Á1
48Á8±48Á9
49Á8±49Á2
48Á9±49Á2
49Á0±49Á3
49Á8±50Á0
48Á5±48Á8
48Á9±49Á2
49Á0±49Á4
48Á9±49Á2
49Á0±49Á3
150Á9
151Á9
151Á1
150Á8
151Á8
151Á0
150Á7
151Á9
151Á0
150Á9
151Á9
151Á1
150Á9
151Á9
151Á0
150Á9
151Á9
151Á1
135Á1
135Á1
135Á1
131Á3
131Á2
131Á0
133Á5
133Á2
133Á8
135Á0
135Á0
135Á1
131Á2
131Á1
131Á2
133Á4
133Á4
133Á5
134Á0
134Á0
134Á1
133Á9
133Á8
133Á9
133Á9
133Á9
133Á5
134Á1
134Á1
134Á2
134Á0
134Á0
134Á0
133Á9
133Á9
134Á0
±
±
±
±
±
±
116Á8±129Á6
114Á5±130Á2
112Á9±129Á8
115Á8±129Á7
114Á4±130Á2
112Á9±129Á8
116Á9±129Á8
114Á6±130Á1
112Á9±129Á8
116Á7±129Á7
114Á5±130Á2
112Á8±129Á8
115Á6±131Á2
114Á4±130Á8
112Á8±129Á8
116Á7±129Á8
114Á5±130Á2
112Á8±129Á8
135Á2
131Á5
±
135Á0
131Á0
±
135Á2
131Á0
±
135Á1
131Á0
±
122Á9±125Á3
162Á4±164Á8
162Á3±164Á7
162Á3±164Á8
135Á5
±
±
122Á9±125Á6
±
±
135Á6
135Á5
122Á8±125Á6
±
±
±
±
±
122Á9±125Á7
161Á4±164Á8
162Á3±164Á8
162Á4±164Á9
135Á6
±
±
135Á0
131Á0
±
122Á9±125Á6
±
±
135Á6
135Á1
135Á7
131Á0
122Á9±125Á6
gave rise to a better activity than substitution by the
3-chloro group (4b, 4e, 4h, 5b, 5e, 5h).
Compounds 4f and 5f were among the most
active and so they were selected to evaluate inter-
actions with the opioidergic, 5-HTergic and nor-
adrenergic pathways.
When administrated simultaneously with mor-
phine 30 min before testing, 4f and 5f failed to
potentiate the analgesia produced by the opioid
agonist (Table 5). Thus, they behaved like ¯uox-
etine, a 5-hydroxytryptamine-uptake inhibitor, and
interacted to give only a simple addition.
The potent antinociceptive effects of these two
derivatives exhibited in the phenylbenzoquinone test
were not repeated in the hot-plate test (Table 4).
Moreover, the diminished antinociceptive effect
of compounds 4f and 5f by a low dose of naloxone
provided good evidence that this effect was partly

SYNTHESIS AND ANALGESIC PROPERTIES OF NEW PYRIDAZINONES
393
Considering the high af®nity of the arylpiper-
azinyl moiety for 5-HTergic receptors (Caccia &
Garattini 1990; Kuipers et al 1995; Dukic et al
1997; Wustrow et al 1998; Corsano et al 1999;
Lopez-Rodriguez et al 1999), we examined the
effect of the 5-HTergic agonist 5-hydroxy-
tryptophan (5-HTP) combined with carbidopa, a
peripheral decarboxylase inhibitor, on analgesia
induced by pyridazinones. As demonstrated in
Table 7, co-administration of 4f, 5f as well as 5-HT
uptake inhibitors citalopram and ¯uoxetine with
5-HTP, resulted in a signi®cant potentiation of
analgesia in the phenylbenzoquinone test. 5-HTP
also greatly enhanced the activity of tramadol,
which is in accordance with its mechanism of
action: a weak opioid mechanism probably related
to its ability to inhibit neuronal re-uptake 5-HT
(Raffa et al 1992; Desmeules et al 1996). These
results provide support for the hypothesis that the
antinociceptive action of 4f and 5f is partly medi-
ated through 5-hydroxytryptamine.
Figure 2. The structures of trazodone and tramadol.
To investigate the role of the noradrenergic
system in the pyridazinone antinociceptive effects,
we examined the capacity of the a2-adrenergic
antagonist yohimbine to prevent the analgesic
effects of 4f and 5f in the phenylbenzoquinone test.
It is now well known that the activation of des-
cending 5-HTergic and noradrenergic inhibitory
mediated by opioid m-receptors (Table 6). This is
in accordance with the binding assay, which con-
®rmed a weak af®nity of 5f for m-receptor, similar
to that of tramadol (Ki values in the micromolar
range while Ki values of naloxone and DAMGO
were 1Á38 and 0Á32 nM, respectively).
Table 4. Results from phenylbenzoquinone test, hot-plate test and locomotor activity.
Compound
Phenylbenzoquinone
test (analgesic activity)
ED50 (mg kg 1, i.p.)b
Decrease of locomotor activity
ED50 (mg kg 1, i.p.)b
Hot-plate test
(% of analgesia
100 mg kg 1, i.p.)
4a
4b
4c
4d
4e
4f
4g
4h
4i
5a
5b
5c
5d
5e
76Á7 (45Á9±128Á1)
45Á9 (22Á0±95Á7)
26Á7 (16Á7±42Á7)
> 100
72Á9 (41Á8±127Á2)
77Á1 (46Á7±127Á4)
77Á7 (45Á4±133Á1)
75Á7 (48Á1±119Á2)
54Á9 (31Á1±97Á0)
83Á3 (47Á1±147Á2)
> 100
55Á6 (34Á1±90Á7)
54Á2 (34Á1±86Á2)
69Á9 (43Á2±113Á0)
76Á3 (57Á6±101Á1)
> 100
37Á6 (6Á2±227Á5)
24Á9 (13Á8±44Á8)
76Á2 (56Á3±103Á2)
54Á6 (36Á7±81Á1)
50Á6 (28Á1±91Á1)
66Á4 (37Á3±118Á2)
52Á6 (27Á8±99Á4)
44Á3 (24Á0±79Á7)
58Á7 (33Á1±104Á2)
23Á5 (11Á8±47Á0)
22Á2 (10Á7±46Á2)
> 100
8Á9Æ 2Á7 (NS)
85Á5 (49Á1±148Á8)
87Á5 (53Á0±144Á4)
247Á2 (148Á4±411Á7)
74Á2 (41Á7±132Á1)
68Á8 (36Á6±129Á3)
> 100
5f
5g
5h
5i
7Á8Æ 3Á2 (NS)
23Á8 (10Á3±54Á9)
25Á9 (13Á6±49Á2)
231Á3 (147Á3±363Á2)
3Á0 (1Á5±6Á1)
Paracetamol
Aspirin
Morphinea
Noramidopyrine
Tramadol
Trazodone
11Á7Æ 3Á6d (NS)
5Á8Æ 11Á6d (NS)
9Á6 (8Á0±11Á5)c
60Á7Æ 4Á7*d
1Á0Æ 1Á0 (NS)
27Á5Æ 2Á9e*
0Á6 (0Á3±1Á1)
68Á5 (22Á8±205Á3)
10Á0Æ 6Á0 (NS)
82Á0Æ 6Á7*
8Á6 (4Á3±17Á3)
40Á4Æ 13Á9*d
10Á2 (8Á1±12Á9)
6Á0Æ (5Á7±6Á4)c
7Á4Æ 4Á9e (NS)
aSubcutaneous route. b95% con®dence intervals in parentheses. cIncrease of motor activity. dIncrease of motor activity at
1
e
1
100 mg kg (i.p.). 10 mg kg (i.p.). *P < 0Á05. NS, not signi®cant.

394
PASCAL COUDERT ET AL
Table 8. Effect of yohimbine (1 mg kg 1, i.p.) on pyridazi-
none 4f- and 5f-induced analgesia in the phenylbenzoquinone
test.
Table 5. Analgesic effect of morphine in the phenylbenzo-
quinone test after acute intraperitoneal administration of pyr-
idazinones 4f and 5f.
Compound
(i.p. route)
% analgesia
Compound
(i.p. route)
% analgesia
Drug plus
saline
Drug plus
morphine
Drug plus
saline
Drug plus
yohimbine
1 a
)
1
1
75Á4Æ 8Á2*b
75Á0Æ 7Á1*b
99Á4Æ 1Á1*b
85Á8Æ 6Á1*b
47Á1Æ 11Á8*c
33Á9Æ 10Á7*c
27Á5Æ 16Á1*c
55Á3Æ 12Á9*c
Morphine (0Á15 mg kg
11Á2Æ 6Á6 (NS)
4f (50 mg kg
5f (50 mg kg
)
)
1
4f (5 mg kg
5f (5 mg kg
)
27Á6Æ 8Á9 (NS) 40Á8Æ 10Á0 (NS)
30Á3Æ 9Á5 (NS) 42Á6Æ 7Á1 (NS)
1
1 a
)
)
Citalopram (10 mg kg
Clonidine (0Á05 mg kg
1
)
32Á4Æ 6Á1*b
72Á9Æ 6Á2*c
1
)
Tramadol (15 mg kg
1
Fluoxetine (3 mg kg
Tramadol (2Á5 mg kg
Trazodone (2 mg kg
)
20Á2Æ 3Á5 (NS) 30Á5Æ 4Á2 (NS)
23Á7Æ 9Á5 (NS) 48Á8Æ 9Á0*c
10Á1Æ 6Á0 (NS) 43Á3Æ 12Á0*c
1
)
aSubcutaneous route. *bP < 0Á05 compared with saline.
1
)
*cP < 0Á05 compared with drug plus saline.
aSubcutaneous route. *bP < 0Á05 compared with saline.
*cP < 0Á05 compared with saline plus morphine. NS, not
signi®cant.
Table 9. Results from maximal electroshock seizure test.
Compound
Anticonvulsant activity (%)
(100 mg kg 1, i.p.)
Table 6. Effect of naloxone (1 mg kg 1, s.c.) on pyridazinone
4f- and 5f-induced analgesia in the phenylbenzoquinone test.
4a
4b
4c
4d
4e
4f
4g
4h
4i
5a
5b
5c
5d
5e
10.(NS)
10.(NS)
10.(NS)
50*.
40*.
50*.
10.(NS)
50*.
10.(NS)
50*.
20.(NS)
40*.
50*.
20.(NS)
10.(NS)
30.(NS)
30.(NS)
Compound (i.p. route)
% analgesia
Drug plus saline Drug plus naloxone
1
4f (50 mg kg
5f (50 mg kg
)
75Á4Æ 8Á2*b
75Á0Æ 7Á1*b
77Á4Æ 5Á2*b
75Á1Æ 6Á9*b
68Á1Æ 7Á4*b
85Á8Æ 6Á1*b
41Á2Æ 6Á5*b
51Á1Æ 7Á8*c
1
)
27Á5Æ 13Á9*c
15Á8Æ 9Á5*c
1 a
)
Morphine (1Á5 mg kg
Citalopram (25 mg kg
Fluoxetine (30 mg kg
Tramadol (15 mg kg
1
)
)
66Á3Æ 7Á6 (NS)
67Á5Æ 6Á9 (NS)
45Á8Æ 14Á1*c
58Á6Æ 9Á1 (NS)
1
1
)
1
Trazodone (5 mg kg
)
5f
5g
5h
5i
aSubcutaneous route. *bP < 0Á05 compared with saline.
*cP < 0Á05 compared with drug plus saline. NS, not signi®cant.
50*.
Carbamazepine
Diazepam
Phenytoin
Phenobarbital
Valproate
Lamotrigine
15Á6.(10Á6±23Á0)a
4Á5 (2Á3±8Á7)a
10Á0 (4Á8±20Á8)a
12Á9 (10Á4±15Á9)a
216Á7 (158Á9±295Á6)a
4Á0 (1Á9±8Á5)a
Table 7. Potentiation of pyridazinone 4f- and 5f-induced
analgesia by 5-HTP plus carbidopa in the phenylbenzoquinone
test.
1
aED50 mg kg
(i.p.) with 95% con®dence intervals in
parentheses. *P < 0Á05. NS, not signi®cant.
Compound
(i.p. route)
% analgesia
systems results in antinociception (Stamford 1995).
Otherwise, some arylpiperazinylpyridazinones
have been described as a1-adrenergic and=or 5-
HT1A-ergic receptor ligands (Gleason et al 1998;
Montesano et al 1998; Barlocco et al 1999). The
results presented in Table 8 show that yohimbine
clearly attenuated the antinociceptive effects of 4f
and 5f suggesting that both compounds might exert
a part of their action through activation of the
central inhibitory noradrenergic pathway. Yohim-
bine also signi®cantly blocked analgesic response
produced by injection of tramadol, which is strong
Drug plus
saline
Drug plus 5-HTP
plus carbidopa
1
5-HTP (50 mg kg
plus carbidopa
)
16Á6Æ 5Á8 (NS)
1
(25 mg kg
4f (5 mg kg
5f (5 mg kg
)
1
)
27Á6Æ 8Á9 (NS)
30Á3Æ 9Á5 (NS)
32Á4Æ 6Á1*a
60Á8Æ 7Á6*b
64Á5Æ 5Á8*b
73Á1Æ 9Á6*b
51Á4Æ 5Á6*b
46Á4Æ 8Á7*b
47Á1Æ 9Á5*b
1
)
1
Citalopram (10 mg kg
)
1
Fluoxetine (3 mg kg
Tramadol (2Á5 mg kg
Trazodone (2 mg kg
)
20Á2Æ 3Á5 (NS)
23Á7Æ 9Á5 (NS)
10Á1Æ 6Á0 (NS)
1
)
1
)
*aP < 0Á05 compared with saline. *bP < 0Á05 compared with
drug plus saline. NS, not signi®cant.

SYNTHESIS AND ANALGESIC PROPERTIES OF NEW PYRIDAZINONES
395
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evidence that the tramadol analgesic effect may be
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central noradrenergic inhibitory system (Kayser
et al 1992).
To complete our investigations we evaluated 4f
and 5f as anticonvulsants. We previously reported
the synthesis and the anticonvulsant activity of a
series of 3-N-substituted diaryl-4,6-pyridazinones
(Coudert et al 1989). Other new pyridazinones have
been reported to exert potent anticonvulsant
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1
pounds, neither 4f nor 5f at 100 mg kg (i.p.) was
able to protect mice ef®ciently against maximal
electroshock seizure (Table 9). We have synthe-
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substituted in the 2-position. Two of the tested
compounds (4f, 5f) showed marked antinociceptive
activity dependent upon opioidergic, 5-HTergic
and=or noradrenergic systems.
Acknowledgements
The authors wish to thank Dr Christine Courteix and
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