Synthesis and evaluation of the analgesic activity of some new isoxazolo[4,5-d]pyridazin-4(5H)-one derivatives

Article abstract of DOI:10.3906/kim-1004-584

Some isoxazolo[4,5-d]pyridazin4(5H)-one derivatives were synthesized and tested for their analgesic activity. The analgesic activities of the compounds were determined by hot-plate and acetic acid writhing tests using morphine and diclofenac as references

Full text of DOI:10.3906/kim-1004-584

Turk J Chem
35 (2011) , 121 – 130.
¨
˙
c
ꢀ TUBITAK
doi:10.3906/kim-1004-584
Synthesis and evaluation of the analgesic activity of some
new isoxazolo[4,5-d]pyridazin-4(5H)-one derivatives
1,∗
1
2
¨
¨
¨
˙
Oya UNSAL TAN , Keriman OZADALI , Ozgu¨r YES¸ILYURT ,
Hakan KAYIR2, Tayfun UZBAY2 and Ayla BALKAN1
1Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Hacettepe University,
Ankara-TURKEY
e-mail: oyaunsal@hacettepe.edu.tr
2Psychopharmacology Research Unit, Department of Medical Pharmacology,
Gu¨lhane Military Medical Academy, Ankara-TURKEY
Received 27.04.2010
Some isoxazolo[4,5-d]pyridazin4(5H)-one derivatives were synthesized and tested for their analgesic
activity. The analgesic activities of the compounds were determined by hot-plate and acetic acid writhing
tests using morphine and diclofenac as references. Compounds 4a, 4f, 4g, and 4i (25 mg/kg) showed
analgesic profiles similar to that of morphine (5 mg/kg).
Key Words: Isoxazolo[4,5-d]pyridazin-4(5H)-one, hot-plate test, writhing test, analgesic effect
Introduction
3(2H)-Pyridazinones are heterocyclic systems that have been widely studied due to their different biological
activities.1−7 Condensed 3(2H)-pyridazinones have become interesting structures for medicinal chemists.8,9
However, the biological properties of isoxazolo[4,5-d]pyridazin-4(5H)-ones, which are condensed 3(2H)-pyridazi-
none derivatives, are not well known even though they were described by Erichomovitch and Desimoni in the
1960s.10,11
One of the most extensive studies on biological activities of isoxazolopyridazinones was done by Giovan-
noni and co-workers.12 They evaluated the analgesic activities of a series of isoxazolo[3,4-d]- and isoxazolo[4,5-
d]pyridazinone derivatives. They reported that 5-{[4-(3-chlorophenyl)piperazine-1-yl]propyl}-3-methyl-7-pheny-
lisoxazolo[4,5-d]pyridazin-4(5H)-one (Figure 1) showed higher analgesic activity than morphine. This interest-
∗Corresponding author
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Synthesis and evaluation of the analgesic activity of some new..., O. UNSAL TAN, et al.,
ing result prompted us to synthesize and screen the analgesic activity of a series of isoxazolo[4,5-d]pyridazine-
4(5H)-ones.
Cl
O
CH3
N
N
(CH2)3
N
N
N
O
Figure 1. Structure of 5-{[4-(3-chlorophenyl)-piperazine-1-yl]propyl}-3-methyl-7-phenylisoxazolo[4,5-d]pyridazin-4(5H)-
one.
Experimental
Chemistry
All chemicals used in this study were supplied by Aldrich (Steinheim, Germany). Melting points were determined
by a Thomas Hoover capillary melting point apparatus (Philadelphia, PA, USA) and are uncorrected. IR spectra
(KBr disc) were recorded on a Bruker Vector 22 IR (Beaconsfield, UK). The 1 H-NMR spectra (DMSO-d6) were
recorded on a Varian Mercury 400 FT NMR spectrophotometer using TMS as an internal reference (chemical
shift represented in δ ppm). The ESI-MS spectra were measured on a Micromass ZQ-4000 single quadruple
mass spectrometer.
The purity of the compounds was controlled by thin layer chromatography (Merck, silicagel, HF254+366
,
type 60, 0.25 mm, Darmstadt, Germany). The elemental analyses (C, H, N) were performed using a Leco
CHNS 932 (Leco Coop, St. Joseph, MI, USA) analyzer by the Scientific and Technological Research Council of
Turkey Instrumental Analysis Laboratories (Ankara, Turkey). The elemental analysis results were within 0.4%
of theoretical values.
Methyl 2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 1
This compound was prepared according to the literature method.13
General procedure for the preparation of methyl 5-substituted benzylidene-2-methyl-4-oxo-4,5-
dihydrofuran-3-carboxylates 2a-i
1 (3.25 mmol) and an equimolar amount of appropriate benzaldehyde were dissolved in 25 mL of anhydrous
benzene. p-Toluensulfonic acid (0.08 g) was added as a catalytic reagent. The mixture was then heated with
a Dean-Stark separator for 2 h. Benzene was evaporated and residue recrystallized from an ethyl acetate and
n-hexane mixture (1:1).
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Synthesis and evaluation of the analgesic activity of some new..., O. UNSAL TAN, et al.,
Methyl 5-benzylidene-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2a
Yield 64%. Mp 118-9 ◦ C. IR (KBr); 1707 (C=O, ester), 1655 (C=O, ring). 1 H-NMR (DMSO-d6 , 400 MHz); δ
2.75 (3H, s, -CH3), 3.76 (3H, s, -OCH3), 6.90 (1H, s, -CH=), 7.5-7.52 (3H, m, phenyl-H3 , H4 , H5), 7.92-7.95
(2H, m, phenyl-H2 , H6). ESI-MS (m/z); 267 [M+Na]+ (100%), 245 [M+H]+
.
Methyl 5-(2-chlorobenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2b
Yield 73%. Mp 131-2 ◦ C. IR: 1711 (C=O, ester), 1654 (C=O, ring). ESI-MS (m/z); 303 [M+Na+2]+ , 301
[M+Na]+ (100%), 279 [M+H]+ , 247, 102, 73.
Methyl 5-(4-chlorobenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2c
Yield 55%. Mp 148-9 ◦ C. IR: 1714 (C=O, ester), 1646 (C=O, ring), ESI-MS (m/z); 303 [M+Na+2]+ , 301
[M+Na]+ (100%), 279 [M+H]+ , 247, 102, 73.
Methyl 5-(2-methylbenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2d
Yield 21%. Mp 146 ◦ C (dec.). IR: 1703 (C=O, ester), 1646 (C=O, ring), ESI-MS (m/z); 281 [M+Na]+ (100%),
259 [M+H]+ , 188, 160, 102, 73.
Methyl 5-(4-methylbenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2e
Yield 31%. Mp 149-50 ◦ C. IR: 1708 (C=O, ester), 1649 (C=O, ring), ESI-MS (m/z); 281 [M+Na]+ (100%),
259 [M+H]+ , 188, 102, 73.
Methyl 5-(2-nitrobenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2f
Yield 60%. Mp 122-3 ◦ C. IR: 1714 (C=O, ester), 1653 (C=O, ring), 1522, 1343 (N=O), ESI-MS (m/z); 312
[M+Na]+ (100%), 290 [M+H]+ , 243, 217, 188, 160, 102, 73.
Methyl 5-(4-nitrobenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2g
Yield 62%. Mp 202 ◦ C (dec.). IR: 1706 (C=O, ester), 1655 (C=O, ring), 1512, 1341 (N=O), ESI-MS (m/z);
312 [M+Na]+ (100%), 290 [M+H]+ , 243, 217, 188, 160, 102, 73.
Methyl 5-(2-methoxybenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2h
Yield 70%. Mp 161 ◦ C (dec.). IR: 1704 (C=O, ester), 1659 (C=O, ring), ESI-MS (m/z); 297 [M+Na]+ (100%),
275 [M+H]+ , 102, 73.
Methyl 5-(4-methoxybenzylidene)-2-methyl-4-oxo-4,5-dihydrofuran-3-carboxylate 2i
Yield 52%. Mp 153 ◦ C (dec.). IR: 1712 (C=O, ester), 1658 (C=O, ring). ESI-MS (m/z); 297 [M+Na]+ (100%),
275 [M+H]+ , 102, 73.
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Synthesis and evaluation of the analgesic activity of some new..., O. UNSAL TAN, et al.,
General procedure for the preparation of methyl 5-(1-hydroxy-2-(substituted phenyl)vinyl)-3-
methylisoxazole-4-carboxylates 3a-i
A mixture of appropriate 2 (0.01 mol), sodium acetate (0.012 mol), hydroxylamine hydrochloride (0.011 mol),
water (10 mL), and ethanol (50 mL) was refluxed for 2 h. After cooling, yellow crystals were collected and
recrystallized from ethanol to give pure compound.
Methyl 5-(1-hydroxy-2-phenylvinyl)-3-methylisoxazole-4-carboxylate 3a
Yield 66%. Mp 135-6 ◦ C. IR (KBr); 1672 (C=O). 1 H-NMR (DMSO-d6 , 400 MHz); δ 2.40 (3H, s, -CH3),
3.85 (3H, s, -OCH3), 6.27 (1H, s, -CH=), 7.33-7.39 (3H, m, phenyl-H3 , H4 , H5), 7.72-7.82 (2H, m, phenyl-H2 ,
H6), 10.53 (1H, s, -OH). ESI-MS (m/z); 282 [M+Na]+ (100%), 260 [M+H]+ , 102.
Methyl 5-(1-hydroxy-2-(2-chlorophenyl)vinyl)-3-methylisoxazole-4-carboxylate 3b
Yield 36%. Mp 131 ◦ C. IR: 1676 (C=O), ESI-MS (m/z); 318 [M+Na+2]+ , 316 [M+Na]+ (100%), 294 [M+H]+
262, 130, 102, 82, 73.
,
Methyl 5-(1-hydroxy-2-(4-chlorophenyl)vinyl)-3-methylisoxazole-4-carboxylate 3c
Yield 39%. Mp 150-1 ◦ C IR: 1677 (C=O), ESI-MS (m/z); 318 [M+Na+2]+ , 316 [M+Na]+ (100%), 294
[M+H]+ , 262, 130, 102, 82, 73.
Methyl 5-(1-hydroxy-2-(2-methylhenyl)vinyl)-3-methylisoxazole-4-carboxylate 3d
The compound could not be isolated. The oily residue was directly used for the synthesis of compound 4d.
Methyl 5-(1-hydroxy-2-(4-methylhenyl)vinyl)-3-methylisoxazole-4-carboxylate 3e
Yield 58%. Mp 162-3 ◦ C IR: 1668 (C=O), ESI-MS (m/z); 296 [M+Na]+ (100%), 274 [M+H]+ , 214, 102, 73.
Methyl 5-(1-hydroxy-2-(2-nitrophenyl)vinyl)-3-methylisoxazole-4-carboxylate 3f
Yield 60%. Mp 110-1 ◦ C IR: 1677 (C=O), 1523, 1332 (N=O), ESI-MS (m/z); 327 [M+Na]+ (100%), 305
[M+H]+ , 273, 122, 102, 82, 73.
Methyl 5-(1-hydroxy-2-(4-nitrophenyl)vinyl)-3-methylisoxazole-4-carboxylate 3g
Yield 63%. Mp 181 ◦ C IR: 1674 (C=O), 1507, 1328 (N=O), ESI-MS (m/z); 327 [M+Na]+ (100%), 305
[M+H]+ , 273, 163, 122, 102, 82, 73.
Methyl 5-(1-hydroxy-2-(2-methoxyphenyl)vinyl)-3-methylisoxazole-4-carboxylate 3h
Yield 49%. Mp 133 ◦ C IR: 1674 (C=O), ESI-MS (m/z); 312 [M+Na]+ (100%), 290 [M+H]+ , 273, 82.
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Synthesis and evaluation of the analgesic activity of some new..., O. UNSAL TAN, et al.,
Methyl 5-(1-hydroxy-2-(4-methoxyphenyl)vinyl)-3-methylisoxazole-4-carboxylate 3i
Yield 65%. Mp 156 ◦ C IR: 1673 (C=O), ESI-MS (m/z); 312 [M+Na]+ (100%), 290 [M+H]+ , 273, 102, 82.
General procedure for the preparation of 7-(substituted benzyl)-3-methylisoxazolo[4,5-d]pyridazin-
4(5H )-ones 4a-i
The mixture of appropriate 3 (0.005 mol), hydrazine hydrate (0.01 mol), and ethanol (10 ml) was refluxed for
2 h. After cooling, the white precipitate was collected and recrystallized from ethanol to give pure compound.
7-Benzyl-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4a)
Yield 91%. Mp 182 ◦ C (179 ◦ C).14 IR (KBr); 3176-2955 (CON-H), 1682 (C=O). 1 H-NMR (DMSO-d6 , 400
MHz); δ 2.53 (3H, s, -CH3), 4.20 (2H, s, -CH2 -), 7.23-7.34 (5H, m, phenyl), 13.07 (1H, br, NH). ESI-MS (m/z);
264 [M+Na]+ (100%), 242 [M+H]+ , 104, 82.
7-(2-Chlorobenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4b)
Yield 44%. Mp 206 ◦ C. IR: 1701 (C=O), 1562 (C=N), 1 H-NMR: 2.55 (3H, s, -CH3), 4.34 (2H, s, -CH2 -), 7.31-
7.48 (4H, m, ar.), 13.04 (1H, br, NH). ESI-MS (m/z); 300 [M+Na+2]+ , 298 [M+Na]+ (100%), 276 [M+H]+
,
240, 225, 125, 82.
7-(4-Chlorobenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4c)
Yield 39%. Mp 206 ◦ C (202 ◦ C).14 IR: 1689 (C=O), 1567 (C=N), 1 H-NMR: 2.54 (3H, s, CH3), 4.22 (2H, s,
-CH2 -), 7.34 (2H, d, ar., H2 , H6 , J: 8.4 Hz), 7.39 (2H, d, ar., H3 , H5 , J: 8.4 Hz), 13.10 (1H, br, NH). ESI-MS
(m/z); 300 [M+Na+2]+ , 298 [M+Na]+ (100%), 276 [M+H]+ , 240, 225, 125, 82.
7-(2-Methylbenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4d)
Mp 191-2 ◦ C. IR: 1688 (C=O), 1559 (C=N), 1 H-NMR: 2,28 (3H, s, -ar.-CH3), 2.52 (3H, s, CH3), 4.19 (2H, s,
-CH2 -), 7.13-7.19 (4H, m, ar.), 13.05 (1H, br, NH), ESI-MS (m/z); 278 [M+Na]+ (100%), 256 [M+H]+ , 151,
136, 102, 82, 73.
7-(4-Methylbenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4e)
Yield 51%. Mp 202-3 ◦ C. IR: 1680 (C=O), 1565 (C=N), 1 H-NMR: 2.22 (3H, s, -ar.-CH3), 2.50 (3H, s, CH3),
4.11 (2H, s, -CH2 -), 7.09 (2H, d, ar., J: 8.4 Hz), 7.15 (2H, d, ar., J: 8.4 Hz), 13.02 (1H, br, NH). ESI-MS (m/z);
278 [M+Na]+ (100%), 256 [M+H]+ , 240, 225, 212, 82.
7-(2-Nitrobenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4f)
Yield 35%. Mp 190-1 ◦ C. IR: 1686 (C=O), 1579 (C=N), 1523, 1344 (N=O), 1 H-NMR: 2.57 (3H, s, CH3), 4.60
(2H, s, -CH2 -), 7.57-7.65 (2H, m, ar., H4 ,H6), 7.75 (1H, t, ar., H5), 8.10 (1H, d, ar., H3), 12.95 (1H, br, NH).
ESI-MS (m/z); 309 [M+Na]+ (100%), 287 [M+H]+ , 240, 102, 73.
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7-(4-Nitrobenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4g)
Yield 31%. Mp>250 ◦ C. IR: 1691 (C=O), 1571 (C=N), 1514, 1349 (N=O), 1 H-NMR: 2.55 (3H, s, CH3), 4.40
(2H, s, -CH2 -), 7.61 (2H, d, ar., H2 , H6 , J: 8.8 Hz), 8.20 (2H, d, ar., H3 , H5 , J: 8.8 Hz), 13.13 (1H, br, NH).
ESI-MS (m/z): 309 [M+Na]+ (100%), 287 [M+H]+ , 102, 73.
7-(2-Methoxybenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4h)
Yield 61%. Mp 186 ◦ C. IR: 1688 (C=O), 1559 (C=N), 1 H-NMR: 2.52 (3H, s, CH3), 3.79 (3H, s, OCH3), 4.19
(2H, s, -CH2 -), 7.13-7.19 (4H, m, ar.), 13.05 (1H, br, NH), ESI-MS (m/z): 294 [M+Na]+ (100%), 272 [M+H]+
,
225, 116, 82.
7-(4-Methoxybenzyl)-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-one (4i)
Yield 70%. Mp 179 ◦ C (176 ◦ C).14 IR: 1693 (C=O), 1565 (C=N), 1 H-NMR: 2.50 (3H, s, CH3), 3.77 (3H, s,
OCH3), 4.11 (2H, s, -CH2 -), 7.09 (2H, d, ar.,H3 , H5 , J: 8.0 Hz), 7.15 (2H, d, ar., H2 , H6 , J: 8.0 Hz), 13.02
(1H, br, NH). ESI-MS (m/z): 294 [M+Na]+ (100%), 272 [M+H]+ , 240, 225, 82.
Pharmacology
Animals: Adult male albino Swiss-Webster mice (25-30 g) were used. They were housed in a quiet and
temperature and humidity controlled room (22
2
◦ C and 60
5%, respectively) in which a 12-h (light/dark)
cycle was maintained (0700-1900 light). All experiments were performed at the same time of the day and in the
light period (0900-1130).
All procedures in the present study are in accordance with the Guide for the Care and Use of Laboratory
Animals as adapted by the National Institutes of Health (Washington, DC, USA, 1996). Local ethical committee
approval was also obtained (March 21 2005; No: 05/22). All efforts were made to minimize animal suffering to
reduce the number of animals used. Each animal was used only once and the animals were killed immediately
after the termination of tests.
Evaluation of the analgesic effect
Analgesia was assessed by using acetic acid-induced writhing test and hot-plate test in mice. Each group
consisted of 6-8 mice.
Writhing test: Writhing syndrome was elicited by ip injection of 0.3% acetic acid in a volume of 10
mL/kg. The number of stretching movements was counted for 10 min, starting 5 min after acetic acid injection.
Injections of compounds and vehicle (0.5% CMC Na) to mice were done 60 min before testing. The degree of
analgesia was expressed as the percentage decrease in the number of writhes and calculated according to the
formula: % inhibitions of writhes= [(Control – Test)/Control] × 100 where control is the number of writhes in
CMC Na-treated animals and test is the number of writhes in compound-treated animals.15
Hot-Plate test: The hot-plate test (52.5 0.4 ◦ C) (MAYCOM-9601) was used to assess paw withdrawal
latency to a thermal pain stimulus. Each mouse was placed individually on a hot plate and the reaction time
was measured starting from time the mouse was placed on the plate until the mouse either demonstrated hind
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Synthesis and evaluation of the analgesic activity of some new..., O. UNSAL TAN, et al.,
paw licking or jumping. We used the cut-off latency time as 90 s to prevent tissue damage. The percentage of
the maximum possible effect (MPE%) was calculated for each animal using the following formula Eq. (1):
ꢀ
ꢁ
ꢀ
ꢁ
Pre-drug
latency
Pre-drug
latency
Test
−
latency
%MPE =
ꢀ
ꢁ
(Cut-off) −
where predrug latency is time elapsed from when the mouse was placed on the plate until the mouse either
demonstrated hind paw licking or jumping before injection of any drugs and test latency is time elapsed during
the same procedure after drug injection in the same mouse.16
Drugs: We used 4a-i, morphine HCl, and diclofenac. Morphine HCl, diclofenac, and the compounds
were suspended in 0.5% CMC Na (vehicle) and injected into the mice intraperitoneally. We used the doses of
compounds according to the locomotor activity test results that did not inhibit locomotor activity (data not
shown).
Statistical analysis: Results were expressed as mean
S.E.M. The analgesic effects of the compound
for each observation time were evaluated by one way analysis of variance (a one-way ANOVA) followed by
Dunnet’s test for post-hoc comparisons. The level of significance was set at the P < 0.05 level.
Results and discussion
Chemistry
The route used to synthesize 7-substituted benzyl-3-methylisoxazolo[4,5-d]pyridazin-4(5H)-ones 4a-i is outlined
in Figure 2. Some of these compounds (4a, 4c, and 4i) were synthesized by Chantegrel and co-workers14 earlier.
In our synthetic procedure, to use methyl acetoacetate instead of ethyl acetoacetate was the only exception. 13
We observed that the yield of 1 increased using methyl acetoacetate. The structures of our target compounds
were confirmed using IR, 1 H-NMR, mass spectra, and elemental analysis (except 4a, 4c, and 4i) data.
O
O
R
Mg(OC2H5)2
ClCH2COCl
CH3OOC
CH3
CH3OOC
CH3
CHO
H
CH3COCH2COOCH3
C
O
O
1
2a-i
R
COOCH3
O
CH3
CH3
H
NH
NH2OH
NH2NH2
N
N
O
N
OH
3a-i
O
R
4a-i
R
Figure 2. Synthetic route of 4a-i.
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Analgesic activity
The analgesic activities of the synthesized compounds in this study were measured using hot-plate and acetic
acid induced writhing tests in mice.
Generally it is well known that opiates have analgesic effects in both writhing and hot-plate tests but
non-steroidal antiinflammatory drugs (NSAIDs) have analgesic activity only in the writhing test. For this reason
we evaluated the analgesic effect of morphine in both tests but we evaluated diclofenac only in the writhing
test.
In the writhing test, it was observed that all compounds tested showed at least equal analgesic activity
to diclofenac. However, these compounds had less activity in comparison with mice that received morphine 5
mg/kg. Compounds 4h and 4i (25 mg/kg), which had electron donating substituents on the phenyl ring, were
the most potent compounds. In addition, 4a, which had no substituent on the phenyl ring, showed moderate
analgesic activity (Figure 3).
100
90
80
70
60
50
40
30
20
10
0
Compounds (25mg/kg)
Figure 3. Analgesic effects of compounds 4a-i in acetic acid-induced writhing test.
90
80
70
60
50
40
30
20
10
0
Compounds (25mg/kg)
Figure 4. Analgesic effects of compounds 4a-i in hot-plate test.
The results of the hot-plate test of all compounds are shown in the Table. Morphine increased paw
withdrawal latency 47.47%. It was observed that 4a, 4f, 4g, and 4i (65.38%, 56.67%, 61.56%, and 36.82%,
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Synthesis and evaluation of the analgesic activity of some new..., O. UNSAL TAN, et al.,
respectively) had an analgesic effect similar to that of morphine. Among them 4a was the most effective
compound in the series. In addition to these results 4b-e and 4h did not demonstrate a significant effect
(Figure 4).
Table. Effects of the compounds in acetic acid induced abdominal writhing and hot-plate tests.
Doses (mg/kg)
(mg/kg)
Inhibition on
writhing test
SEM
% MPE on
Compounds
R
Hot-plate test
Control
-
-
-
0.1 8.93
1.11 6.49
47.47 9.56*
-
Morphine
5
97.97 1.67*
63.25 8.54*#
Diclofenac
-
8
4a
4b
4c
4d
4e
4f
H
25
25
25
25
25
25
25
25
25
73.86 8.29*# 65.38 10.88*
2-Cl
58.33 8.92*#
61.36 5.42*#
62.5 6.13*#
66.66 3.62*#
12.33 6.01
6.25 3.81
18.70 10.22
3.46 6.00
4-Cl
2-CH3
4-CH3
2-NO2
4-NO2
2-OCH3
4-OCH3
62.12 5.75*# 56.67 16.75*
60.22 6.36*# 61.56 12.10*
4g
4h
4i
78.78 5.46*#
17.99 8.28
78.40 5.85*# 36.82 13.18*
*P < 0.05 significantly different from control. # P < 0.05 significantly different from morphine. n = 6-8.
Conclusion
In the present study, 9 (6 of them are new compounds) isoxazolo[4,5-d]pyridazin-4(5H)-one derivatives were
synthesized in order to evaluate their analgesic activities. According to the results we can conclude the following:
(i) All compounds have an analgesic effect in the acetic acid-induced writhing test. (ii) The compounds (4h
and 4i) that have a methoxy substituent on the phenyl ring are the most potent compounds. (iii) Substitution
of highly electron donating groups such as OH or NH2 may be useful to increase activity in the writhing test.
(iv) In the hot-plate test, the compounds carrying NO2 groups (4f and 4g) are the most potent ones among
the substituted derivatives. (v) When the results of the 2 tests are considered together, it can be suggested
that 4a, 4f, 4g, and 4i have analgesic profiles similar to that of morphine in acetic acid-induced writhing and
hot-plate tests. However, 4b-4e and 4h showed an analgesic effect only in the writhing test, which means they
do not have the same analgesic profile as morphine and the analgesic profiles of these compounds appear to be
more similar to those of NSAIDs. (vi) Among the compounds, 4a showed remarkable activity in all tests.
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Synthesis and evaluation of the analgesic activity of some new..., O. UNSAL TAN, et al.,
Acknowledgements
The authors gratefully acknowledge the financial support provided by the Scientific Research Fund of Hacettepe
University though Project 0501301001. The authors would also like to thank Mr. Selami Alan for his technical
assistance during the study.
References
1. Curran, W. V.; Ross, A. J. Med. Chem. 1974, 17, 273-281.
2. Takaya, M.; Sato, M.; Terashima, K.; Tanizawa, H. J. Med. Chem. 1979, 22, 53-58.
3. Rubat, C.; Courdert, P.; Albuisson, E.; Bastide, J.; Couquelet, J.; Tronche, P. J. Pharm. Sci. 1992, 81, 1084-1089.
4. Dal Piaz, V.; Ciciani, G.; Giovannoni, M. Il Farmaco, 1997, 52, 173-178.
¨
5. Balkan, A.; Ozkanlı, F.; Unsal, O.; Go¨ren, Z.; Terziog˘lu, B. FABAD J. Pharm. Sci. 2003, 28, 19-25.
¨
¨
6. Unsal, O.; Balkan, A.; Aypak, C.; Terziog˘lu, B.; G¨oren, Z. FABAD J. Pharm. Sci. 2004, 29, 185-194.
7. Betti, L.; Botta, M.; Corelli, F.; Floridi, M.; Fossa, P.; Giannaccini, G.; Manetti, F.; Strappaghetti, G.; Corsano,
S. Bioorg. Med. Chem. Lett. 2002, 12, 437-440.
8. Strappaghetti, G.; Brodi, C.; Giannaccini, G.; Betti, L. Bioorg. Med. Chem. Lett. 2006, 16, 2575-2579.
9. Pakulska, W.; Malinowski, Z.; Szczesniak, A. K.; Czarnecka, E.; Epsztajn, J. Arch. Pharm. Chem. Life Sci. 2009,
342, 41-47.
10. Erichomovitch, L.; Chubb, F. Can. J. Chem. 1966, 44, 2095-2100.
11. Desimoni, G.; Finzi, P. Tetrahedron 1967, 23, 681-685.
12. Giovannoni, M. P.; Vergelli, C.; Gherlardini, C.; Galeotti, N.; Bartolini, A.; Dal Piaz, V. J. Med. Chem. 2003, 46,
1055-1059.
¨
13. Unsal-Tan, O.; Ozden, K.; Rauk, A.; Balkan, A. Eur. J. Med. Chem. 2010, 45, 2345-2352.
¨
14. Chantegrel, B.; Deshayes, C.; Pujol, B. J. Heterocycl. Chem. 1990, 27, 927-34.
¨
15. Balkan, A.; G¨oren, Z.; Urgun, H.; C¸ alı¸s, U.; C¸ akar, A. N.; Atilla, P.; Uzbay, I. T. Arzneim.-Forsch./Drug Res.
˙
2002, 52, 462-467.
˙
16. Uzbay, I. T.; C¸ınar, M. G.; Aytemir, M.; Tu˘glular, I. Life Sciences 1999, 64, 1313-1319.
˙
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