Utility of novel 2-furanones in synthesis of other heterocyclic compounds having anti-inflammatory activity with dual COX2/LOX inhibition

Article abstract of DOI:10.1080/14756366.2021.1908277

Inflammation is associated with the development of several diseases comprising cancer and cardiovascular disease. Agents that suppress cyclooxygenase (COX) and lipoxygenase (LOX) enzymes, besides chemokines have been suggested to minimise inflammation. Here, a variety of novel heterocyclic and non-heterocyclic compounds were prepared from novel three furanone derivatives. The structures of all synthesised compounds were confirmed by elemental and spectral analysis including mass, IR, and ¹H-NMR spectroscopy. Anti-inflammatory activities of these synthesised compounds were examined in?vitro against COX enzymes, 15-LOX, and tumour necrosis factor-α (TNF-α), using inhibition screening assays. The majority of these derivatives showed significant to high activities, with three pyridazinone derivatives (5b, 8b, and 8c) being the most promising anti-inflammatory agents with dual COX-2/15-LOX inhibition activities along with high TNF-α inhibition activity.

Full text of DOI:10.1080/14756366.2021.1908277

Journal of Enzyme Inhibition and Medicinal Chemistry
ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/ienz20
Utility of novel 2-furanones in synthesis of other
heterocyclic compounds having anti-inflammatory
activity with dual COX2/LOX inhibition
Rania H. Abd El-Hameed, Shahenda Mahgoub, Hend M. El-Shanbaky,
Mosaad S. Mohamed & Sahar A. Ali
To cite this article: Rania H. Abd El-Hameed, Shahenda Mahgoub, Hend M. El-Shanbaky,
Mosaad S. Mohamed & Sahar A. Ali (2021) Utility of novel 2-furanones in synthesis of other
heterocyclic compounds having anti-inflammatory activity with dual COX2/LOX inhibition, Journal of
Enzyme Inhibition and Medicinal Chemistry, 36:1, 977-986, DOI: 10.1080/14756366.2021.1908277
To link to this article: https://doi.org/10.1080/14756366.2021.1908277
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JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY
2021, VOL. 36, NO. 1, 977–986
https://doi.org/10.1080/14756366.2021.1908277
RESEARCH PAPER
Utility of novel 2-furanones in synthesis of other heterocyclic compounds having
anti-inflammatory activity with dual COX2/LOX inhibition
Rania H. Abd El-Hameeda, Shahenda Mahgoubb, Hend M. El-Shanbakya, Mosaad S. Mohameda and Sahar A. Alib
b
aPharmaceutical Organic Chemistry Department, Faculty of Pharmacy, Helwan University, Cairo, Egypt; Department of Biochemistry and
Molecular Biology, Faculty of Pharmacy, Helwan University, Cairo, Egypt
ABSTRACT
ARTICLE HISTORY
Received 8 January 2021
Revised 7 February 2021
Accepted 18 March 2021
Inflammation is associated with the development of several diseases comprising cancer and cardiovascular
disease. Agents that suppress cyclooxygenase (COX) and lipoxygenase (LOX) enzymes, besides chemokines
have been suggested to minimise inflammation. Here, a variety of novel heterocyclic and non-heterocyclic
compounds were prepared from novel three furanone derivatives. The structures of all synthesised com-
KEYWORDS
1
pounds were confirmed by elemental and spectral analysis including mass, IR, and H-NMR spectroscopy.
Pyridazinone; selective COX-
2 inhibitor; 15-LOX
inhibitors; TNF-a inhibitor;
anti-inflammatory
Anti-inflammatory activities of these synthesised compounds were examined in vitro against COX enzymes,
15-LOX, and tumour necrosis factor-a (TNF-a), using inhibition screening assays. The majority of these
derivatives showed significant to high activities, with three pyridazinone derivatives (5b, 8b, and 8c) being
the most promising anti-inflammatory agents with dual COX-2/15-LOX inhibition activities along with high
TNF-a inhibition activity.
1. Introduction
inflammatory mediators; playing an important role in the proin-
flammatory PGs biosynthesis7.
Inflammation is a protective physiological defence mechanism
provided by the body immune system in response to toxins, infec-
tious pathogens, and local injury1. It occurs as a result of biosyn-
thesis of pro-inflammatory mediators (leukotrienes [LTs] and
prostaglandins [PGs]) from arachidonic acid (AA) by the action of
the enzymes lipoxygenase (LOX) and cyclooxygenase (COX),
respectively. Although inflammation is a normal defence mechan-
ism, persistent/untreated inflammation leads to complicated
events with release of many mediators that can turn the condition
to be harmful and may lead to the development of certain dis-
eases, as asthma, rheumatoid arthritis, atherosclerosis, diabetes,
and cancer2. Therefore, anti-inflammatory agents may be helpful
in the management of inflammatory disorders3. Anti-inflammatory
effects of various compounds may result mainly from their ability
to inhibit some of the key enzymes involved in inflammation and/
or cell signalling pathways such as COX and LOX4.
Thus, inhibition of these enzymes may be valuable treatment
for inflammatory conditions. In the human system, COX occurs in
two isoforms: COX-1 and COX-25. Both isoforms catalyse a COX
reaction in which they act on AA as substrate. COX isoforms are
haem containing enzymes that demonstrate distinctive expression
profiles and roles in numerous physiological processes.6 COX-1 is
constitutive isoform and is found in the gastrointestinal (GI)-tract,
renal collecting tubules, and platelets and is believed to be
responsible for the maintenance of physiological homeostasis
such as renal function and GI integrity through production of gas-
troprotective PGs. On the other hand, the inducible isoform; COX-
Inhibition of both isoforms of COX by classical nonsteroidal
anti-inflammatory drugs (NSAIDs) leads to inhibition of gastropro-
tective PGs produced via the COX-1 pathway; which occurs along
with suppression of the pathological COX effects; resulting in the
GI toxicities accompanying the use of numerous NSAIDs as GI irri-
tation, bleeding, and ulceration8. Several studies revealed that
COX-2 is highly expressed in a wide range of cancer tissues, such
as colon, breast, and prostate, suggesting that it may control sev-
eral cellular processes. Thus, selective COX-2 inhibitors have been
extensively investigated for the treatment and prevention of a var-
iety of cancers9.
Also, COX-2 is found in wide range of tissues e.g. brain, spinal
cord, and kidneys, as well as many cells like vascular endothelium,
suggesting that this isoenzyme may play a more complex physio-
logical role than was expected10,11
.
However, potent selective COX-2 inhibitors, which were used
instead of NSAIDs; also showed disadvantages as incidence of vas-
cular-diseases12. AA; which is the substrate of COX enzymes, is
also converted by LOX enzyme to several lipid mediators recog-
nised as eicosanoids13. LOXs are an exceptional group of non-
haem iron-containing enzymes that catalyse the peroxidation of
polyunsaturated fatty acids viz. AA and linoleic acid to their hydro-
peroxides14. 15-LOXs are implicated in a variety of human dis-
eases, like the oxidative alteration of low-density lipoproteins and
thus, the progression of atherosclerosis15. In addition to many
neurodegenerative diseases as Alzheimer’s disease16, 15-LOX-1
inhibition has been reported to be a focal point to decrease the
2 is released during tissue injury and induced by many kinds of biosynthesis of eoxines, which are known to be pro-inflammatory
CONTACT Rania H. Abd El-Hameed
zeiadomar@yahoo.com
Pharmaceutical Organic Chemistry Department, Faculty of Pharmacy, Helwan University, Ain-
Helwan, Helwan, Cairo, Egypt
Supplemental data for this article can be accessed here.
ß 2021 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use,
distribution, and reproduction in any medium, provided the original work is properly cited.

978
R. H. A. EL-HAMEED ET AL.
mediators17 and promotors for cancer disease18. Thus, some litera- activity over celecoxib besides compounds VIIIa,b; which were
reported to be comparable with ibuprofen as dual COX/LOX inhib-
ture work has been targeting 15-LOX-1.
ition activity42,43
.
Yet, the use of LOX-inhibitors might represent an insufficient
single therapeutic model in inflammatory diseases other than
asthma19,20. It was discovered that dual inhibition of the COX and
LOX pathways could produce a wider spectrum of anti-inflamma-
tory effects and can limit the vascular-changes seen during inflam-
mation and leukocyte-induced GI damage21.
The anti-inflammatory activity shown by the above derivatives
has drawn our interest to continue our research44–46 for develop-
ment of new anti-inflammatory agents. The synthesis and bio-
logical evaluation of new 2-furanone derivatives are reported
herein. Also 2-furanone derivatives were used for synthesis of
other heterocyclic and non-heterocyclic derivatives, which were
tested as for their potential as anti-inflammatory agents against
COX enzymes, LOX, and tumour necrosis factor-a (TNF-a).
2-Furanones; well-known heterocyclic derivatives; had attracted
a great attention during the last decade due to facile ring open-
ing and conversion to other heterocycles; pyrrolones, pyridazi-
nones, pyrazoles, and oxadizoles. These heterocycles acquired an
obvious medicinal interest as antimicrobial22–24, antiviral25–27, anti-
2. Materials and methods
mycobacterial28,29
, . Literature is
and anti-cancer agents30–32
2.1. Synthesis of lead compounds
enriched with different 2-furanones subjected to ring opening to
form 2-pyrrolone, pyridazinone, and oxadiazole derivatives, all
have high activity as anti-inflammatory24,33–41. As shown in Figure
1, 2-furanone derivatives Ia–c were reported to exhibit compar-
able anti-inflammatory activity to that of diclofenac33,35,36, while
2-pyrrolone derivatives II, IIIa–b, and IV were reported to have
comparable anti-inflammatory activity to that of indomethacin,
All commercial chemicals used as starting materials and reagents
in this study were purchased from Merck (Darmstadt, Germany)
and were of reagent grade. All melting points were uncorrected
and measured using Electro-thermal IA 9100 apparatus (Shimadzu,
Japan); IR spectra were recorded as potassium bromide pellets on
a Perkin-Elmer 1650 spectrophotometer (Waltham MA, Faculty of
ibuprofen, and diclofenac, respectively24,34–36
.
1
Science, Cairo University, Cairo, Egypt. H-NMR spectra were deter-
Many pyridazinone derivatives were also reported as Va,b;
which have superior anti-inflammatory activities over celecoxib
and indomethacin, while derivatives VIa–c were more potent than
aminopyrine, mebirizole, phenylbutazone, and mefenamic
mined on a Varian Mercury (300 MHz) spectrometer (Varian,
Crawley, UK) and chemical shifts were expressed as ppm against
TMS as internal reference (The Main Chemical warfare
Laboratories, Almaza, Cairo, Egypt). Mass spectra were recorded
acid37,38. Some of oxadiazole derivatives VIIa,b showed superior on 70 eV (EI Ms-QP 1000 EX, Shimadzu, Japan), Faculty of Science,
Ia: Ar1= 4-CH3-C6H5
Ar2=3-OC2H5,4-OCOCH3-C6H3
II Ar1=4-OPh-C6H4, Ar2= 3,4,5-OCH3-C6H2
Ib: Ar1=2-naphthyl, Ar2=2,7-Cl-3-quinolinyl
X=CH2-Ph
Ic: Ar1=2-naphthyl, Ar2=2-Cl,7-NO2-3-quinolinyl
IIIa: Ar1=4-Cl-C6H4, Ar2=4-Cl-C6H4
IIIb: Ar1=4-F-C6H4, Ar2=4-Cl-C6H4
X=CH2-Ph
IV: Ar1=2-naphthyl, Ar2=2,7-Cl-3-quinolinyl
X=H
Va: R1=H, R2=3-CH3-C6H4, X=H
VIIa: R1=C6H4SO2CH3, R2= 4-NO2-C6H4
VIIb: R1=C6H4SO2CH3, R2=4-tBu-C6H4
Vb: R1=4-F-C6H4, R2=2-CH3OC6H4, X=H
VIa: R1=CH3, R2=n-C3H7O, X=N-morpholinyl
VIb: R1=CH3, R2=n-C4H9O, X=N-morpholinyl
VIc: R1=C2H5, R2=C2H5O, X=N-morpholinyl
VIIIa: R1=4-Cl-C6H4, R2= Chromon-3-yl
VIIIb: R1=2,4-Cl2C6H3, R2= Chromon-3-yl
Figure 1. 2-furanones, 2-pyrrolones, pyridazinones, and oxadiazoles as potent anti-inflammatory agents.

JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY
979
Cairo University, Cairo, Egypt. Microanalyses were operated using 2.1.2.3. 5-(3,4-Dichlorophenyl)-3-(4-dimethylaminobenzylidene)-1H-
pyrrol-2-one (3c). Yield: 56%; m.p.: >300 ꢀC; IR (KBr) ꢀ (cmꢁ1):
Vario, Elmentar apparatus (Shimadzu, Japan), Organic
Microanalysis Unit, Faculty of Science, Cairo University, Cairo,
Egypt. Column Chromatography was performed on (Merck) Silica
3697 (NH), 1673 (C¼O); MS (EI) m/z: 360 (M þ 2, 43.58%), 358 (Mþ,
62%); 1H-NMR (DMSO-d6, 300 MHz) d (ppm): 2.98 (s, 6H, N(CH3)2),
6.46–8.37 (m, 9H, Ar-H þ CH methyne), 10.37 (s, 1H, NH, D2O-
gel 60 (particle size 0.06–0.20 mm). All the listed compounds are
new except compound 1 was previously reported47,48
.
exchangable); Anal. Calcd. for C19H16Cl2N2O (358.05): C, 63.52; H,
4.49; N, 7.80%. Found: C, 63.21; H, 4.56; N, 7.34%.
2.1.1. General procedure for the synthesis of compounds 2a–c
A mixture of compound 1 (0.03 mol) and equimolar amount of
aromatic aldehyde was refluxed in acetic anhydride (15 ml) with
triethylamine (3–4 drops) for 4 h. After completion of reaction, the
product was filtered, washed with ethanol, and recrystallised from
ethanol to obtain compounds 2a–c.
2.1.3. General procedure for the synthesis of compounds 4a–c
To a solution of the furanone 2a–c (0.01 mol) in absolute ethanol
(20 ml), benzylamine (1.07 ml, 0.01 mol) was added and the reac-
tion mixture was refluxed for 5 h. The product was filtered off,
washed with ethanol, and finally recrystallised from ethanol to
give the amides 4a–c.
2.1.1.1.
5-(3,4-Dichlorophenyl)3-(2-nitrobenzylidene)-furan-2-one
(2a). Yield: 57%; m.p.: 259–261 ꢀC; IR (KBr) ꢀ (cmꢁ1): 1769 (C¼O),
1515,1337 (NO2); MS (EI) m/z: 363 (M þ 2, 9.35%), 361 (M, 14.24%);
1H-NMR (DMSO-d6, 300 MHz) d (ppm): 7.51–8.20 (m, 9H, Ar-H þ CH
methyne); Anal. Calcd. for C17H9Cl2NO4 (361.05): C, 56.51; H, 2.49;
N, 3.88%. Found: C, 56.69; H, 2.81; N, 3.54%.
2.1.3.1.
N-benzyl-2-(2-nitrobenzylidene)-4-(3,4-dichlorophenyl)-4-
oxobutanamide (4a). Yield: 30%; m.p.: 250–252 ꢀC; IR (KBr) ꢀ
(cmꢁ1): 3310 (NH), 1681, 1647 (2 C¼O), 1559, 1336 (NO2); MS (EI)
m/z: 468 (Mþ, 0.33%), 91 (tropylium, 100%); 1H-NMR (DMSO-d6,
300 MHz) d (ppm): 3.40 (s, 2H, CH2–C¼O), 4.36 (dd, 2H, CH2–NH),
7.10 (s, 1H, NH, D2O-exchangable), 7.11–8.80 (m, 13H, Ar-H þ CH-
methyne); Anal. Calcd. for C24H18Cl2N2O4 (468.07): C, 61.42; H,
3.87; N, 5.97%. Found: C, 61.32; H, 3.51; N, 5.82%.
2.1.1.2.
5-(3,4-Dichlorophenyl)3-(3-nitrobenzylidene)-furan-2-one
(2b). Yield: 90%; m.p.: 230–232 ꢀC; IR (KBr) ꢀ (cmꢁ1): 1781 (C¼O),
1519,1345 (NO2); MS (EI) m/z: 363 (M þ 2, 23.28%), 361 (M,
34.48%), 173 (benzoyl, 100%); 1H-NMR (DMSO-d6, 300 MHz) d
(ppm): 7.23–8.67 (m, 9H, Ar-H þ CH methyne); Anal. Calcd. for
C17H9Cl2NO4 (361.05): C, 56.51; H, 2.49; N, 3.88%. Found: C, 56.26;
H, 2.35; N, 3.76%.
2.1.3.2.
N-benzyl-2-(3-nitrobenzylidene)-4-(3,4-dichlorophenyl)-4-
oxobutanamide (4b). Yield: 70%; m.p.: 124–126 ꢀC; IR (KBr) ꢀ
(cmꢁ1): 3312 (NH), 1676, 1648 (2 C¼O), 1530, 1350 (NO2); MS (EI)
m/z: 468 (Mþ, 0.29%); 1H-NMR (DMSO-d6, 300 MHz) d (ppm): 3.61
(s, 2H, CH2-CO), 4.21 (dd, 2H, CH2–NH), 7.05–8.35 (m, 12H, Ar-
H þ CH-methyne þ s, 1H, NH, D2O-exchangable); Anal. Calcd. for
C24H18Cl2N2O4 (468.07): C, 61.42; H, 3.87; N, 5.97%. Found: C,
61.35; H, 3.74; N, 5.82%.
2.1.1.3.
5-(3,4-Dichlorophenyl)3-(4-dimethylaminobenzylidene)-
furan-2-one (2c). Yield: 73%; m.p.: 204–206 ꢀC; IR (KBr) ꢀ (cmꢁ1):
1
1748 (C¼O); MS (EI) m/z: 361 (M þ 2, 12.99%), 359 (M, 19.98%); H-
NMR (DMSO-d6, 300 MHz) d (ppm): 3.05 (s, 6H, N(CH3)2), 6.75–8.11
(m, 9H, Ar-H þ CH methyne); Anal. Calcd. for C19H15Cl2NO2
(359.05): C, 63.51; H, 4.18; N, 3.90%. Found: C, 63.29; H, 3.96;
N, 3.98%.
2.1.3.3. N-benzyl-2-(4-dimethylaminobenzylidene]-4-(3,4-dichloro-
phenyl)-4-oxobutanamide (4c). Yield: 40%; m.p.: 241–243 ꢀC; IR
(KBr) ꢀ (cmꢁ1): 3300 (NH), 1756, 1670 (2 C¼O); MS (EI) m/z: 468
(M þ 2, 1.73%), 466 (M, 3.15%), 448 (Mþ-H2O, 56.15%), 91 (tropy-
lium, 100%); 1H-NMR (DMSO-d6, 300 MHz) d (ppm): 2.88 (s,6H,
N(CH3)2), 3.26 (s, 2H, CH2-CO), 4.20 (dd, 2H, CH2–NH), 6.93 (s, 1H,
NH, D2O-exchangable), 6.69–7.43 (m, 13H, Ar-H þ CH-methyne);
Anal. Calcd. for C26H24Cl2N2O2 (466.09): C, 66.81; H, 5.18; N, 5.99%.
Found: C, 66.75; H, 5.24; N, 5.86%.
2.1.2. General procedure for the synthesis of compounds 3a–c
A solution of the furanone derivatives 2a–c (0.01 mol) and ammo-
nium acetate (7.7 g, 0.1 mol) in acetic acid (10 ml) was refluxed for
3 h. The reaction mixture was left to cool at room temperature
and the product obtained was filtered off, recrystallised from etha-
nol to give compounds 3a–c.
2.1.4. General procedure for the synthesis of compounds 5a–c
A solution of the furanone 2a–c (0.01 mol) and phenyl hydrazine
3 ml in Na ethoxide (10 ml) was refluxed for 3 h. The product
obtained was filtered, washed with water, and recrystallised from
ethanol to give compounds 5a–c.
2.1.2.1. 5-(3,4-Dichlorophenyl)-3-(2-nitrobenzylidene)-1H-pyrrol-2-
one (3a). Yield: 80%; m.p.: >300 ꢀC; IR (KBr) ꢀ (cmꢁ1): 3140 (NH),
1698 (C¼O), 1514, 1350 (NO2); MS (EI) m/z: 362 (M þ 2, 10.78%),
360 (Mþ, 16.29%); H-NMR (DMSO-d6, 300 MHz) d (ppm): 6.80–8.17
1
(m, 9H, Ar-H þ CH methyne), 10.72 (s, 1H, NH, D2O-exchangable);
Anal. Calcd. for C17H10Cl2N2O3 (360.02): C, 56.67; H, 2.78; N, 7.78%.
Found: C, 56.81; H, 2.55; N, 7.46%.
2.1.4.1. 6-(3,4-Dichlorophenyl)-4-(2-nitrobenzylidene)-1-phenyl-1,4-
dihydropyridazin-3(2H)-one (5a). Yield: 72%; m.p.: >300 ꢀC; IR (KBr)
ꢀ (cmꢁ1): 3317 (NH), 1629 (C¼O), 1556, 1320 (NO2); MS (EI) m/z:
451 (Mþ, 0.49%); 1H-NMR (DMSO-d6, 300 MHz) d (ppm): 7.46–8.77
(m, 15H, Ar-H þ CH-methyne þ NH-D2O exchangeable); Anal. Calcd.
for C23H15Cl2N3O3 (451.04): C, 61.08; H, 3.34; N, 9.29%. Found: C,
61.29; H, 3.74; N, 9.35%.
2.1.2.2. 5-(3,4-Dichlorophenyl)-3-(3-nitrobenzylidene)-1H-pyrrol-2-
one (3b). Yield: 70%; m.p.: >300 ꢀC; IR (KBr) ꢀ (cmꢁ1): 3143 (NH),
1704 (C¼O), 1517, 1355 (NO2); MS (EI) m/z: 362 (M þ 2, 67.3%),
360 (Mþ, 100%); 1H-NMR (DMSO-d6, 300 MHz) d (ppm): 7.05–8.57
(m, 9H, Ar-H þ CH methyne), 10.68 (s, 1H, NH, D2O-exchangable);
Anal. Calcd. for C17H10Cl2N2O3 (360.02): C, 56.67; H, 2.78; N, 7.78%.
Found: C, 56.52; H, 2.45; N, 7.43%.
2.1.4.2. 6-(3,4-Dichlorophenyl)-4-(3-nitrobenzylidene)-1-phenyl-1,4-
dihydropyridazin-3(2H)-one (5b). Yield: 69%; m.p.: >300 ꢀC; IR (KBr)
ꢀ (cmꢁ1): 3456 (NH), 1671 (C¼O), 1549, 1328 (NO2); MS (EI) m/z:

980
R. H. A. EL-HAMEED ET AL.
451 (Mþ, 1.56%); 1H-NMR (DMSO-d6, 300 MHz) d (ppm): 7.16–8.49 2.1.6.1. 3-(3,4-Dichlorophenyl)-5-[(2-nitropheny)l-methyl]-1H-pyri-
dazin-6-one (7a). Yield: 85%; m.p.: 224–226 ꢀC; IR (KBr) ꢀ (cmꢁ1):
(m, 15H, Ar-H þ CH-methyne þ NH-D2O exchangeable); Anal. Calcd.
for C23H15Cl2N3O3 (451.04): C, 61.08; H, 3.34; N, 9.29%. Found: C,
61.28; H, 3.61; N, 9.35%.
3187 (NH), 1655 (C¼O), 1517 þ 1343 (NO2); MS (EI) m/z: 377
(M þ 2, 2.63%), 375 (Mþ, 3.8%), 330 (M þ 2-NO2, 63.21%), 329
(M – NO2, 100%); 1H-NMR (DMSO-d6, 300 MHz) d (ppm): 4.13 (s,
2H, CH2), 7.49–8.00 (m, 8H, Ar-H), 13.32 (s, 1H, NH-pyridazinone,
D2O-exchangable); Anal. Calcd. for C17H11Cl2N3O3(375.04): C, 54.40;
H, 2.93; N, 11.20%. Found: C, 54.72; H, 3.05; N, 11.08%.
2.1.4.3. 6-(3,4-Dichlorophenyl)-4-(4-dimethylaminobenzylidene)-1-
phenyl-1,4-dihydropyridazin-3(2H)-one (5c). Yield: 43%; m.p.:
>300 ꢀC; IR (KBr) ꢀ (cmꢁ1): 3300 (NH), 1655 (C¼O); MS (EI) m/z:
1
449 (Mþ, 1.63%); H-NMR (DMSO-d6, 300 MHz) d (ppm): 2.86 (s,6H,
2.1.6.2. 3-(3,4-Dichlorophenyl)-5-[(3-nitropheny)l-methyl]-1H-pyri-
dazin-6-one (7b). Yield: 43%; m.p.: 257–259 ꢀC; IR (KBr) ꢀ (cmꢁ1):
3203 (NH), 1653 (C¼O), 1527 þ 1346 (NO2); MS (EI) m/z: 377
(M þ 2, 6.56%), 375 (Mþ, 10.06%); 1H-NMR (DMSO-d6, 300 MHz) d
(ppm): 4.01 (s, 2H, CH2), 7.59–8.25 (m, 8H, Ar-H), 13.34 (s, 1H, NH-
N(CH3)2), 6.55–8.51 (m, 15H, Ar-H þ CH-methyne þ NH-D2O
exchangeable); Anal. Calcd. for C25H21Cl2N3O (449.04): C, 66.67; H,
4.70; N, 9.33%. Found: C, 66.46; H, 5.12; N, 9.17%.
2.1.5. General procedure for the synthesis of compounds 6a–c
To a solution of the furanones 2a–c (0.01 mol) in absolute ethanol
(20 ml), hydrazine hydrate (3.5 ml, 0.11 mol) was added. The reac-
tion mixture was left at room temperature with occasional shaking
until complete dissolving and poured onto ice water. The product
obtained 6a–c was filtered off, washed with hexane.
pyridazinone,
D2O-exchangable);
Anal.
Calcd.
for
C17H11Cl2N3O3(375.04): C, 54.40; H, 2.93; N, 11.20%. Found: C,
54.32; H, 2.83; N, 11.32%.
2.1.6.3.
3-(3,4-Dichlorophenyl)-5-[(4-dimethylaminopheny)l-
methyl]-1H-pyridazin-6-one (7c). Yield: 87%; m.p.: 229–231 ꢀC; IR
(KBr) ꢀ (cmꢁ1): 3164 (NH), 1651 (C¼O); MS (EI) m/z: 377 (M þ 4,
12.06%), 375 (M þ 2, 67.79%), 373 (Mþ, 100%); 1H-NMR (DMSO-d6,
300 MHz) d (ppm): 2.81 (s, 6H, N(CH3)2), 3.70 (s, 2H, CH2), 6.62–8.06
(m, 8H, Ar-H), 13.19 (s, 1H, NH-pyridazinone, D2O-exchangable);
Anal. Calcd. for C19H17Cl2N3O (373.09): C, 61.13; H, 4.56; N, 11.26%.
Found: C, 61.33; H, 4.54; N, 10.99%.
2.1.5.1. 4-(3,4-Dichlorophenyl)-2-(2-nitrobenzylidene)-4-oxo-buta-
nehydrazide (6a). Yield: 65%; m.p.: 164–166 ꢀC; IR (KBr) ꢀ (cmꢁ1):
3322 (NH), 3250 (NH2), 1703,1670 (2 C¼O), 1521, 1343 (NO2); MS
(EI) m/z: 395 (M þ 2, 0.37%), 393 (Mþ, 0.47%), 364 (M þ 2 – NHNH2,
21.29%), 362 (M – NHNH2, 31.7%); 1H-NMR (DMSO-d6, 300 MHz) d
(ppm): 3.17 (s, 2H, CH2), 4.55 (s, 2H, NH2, D2O-exchangable), 6.87
(s, 1H, NHCO, D2O-exchangable), 7.32–8.06 (m, 8H, Ar-H þ CH-
methyne); Anal. Calcd. for C17H13Cl2N3O4 (393.04): C, 51.91; H,
4.33; N, 10.69%. Found: C, 51.77; H, 4.51; N, 10.58%.
2.1.7. General procedure for the synthesis of compounds 8a–c
To a solution of the hydrazides 6a–c (0.01 mol) in dry benzene
(20 ml), benzoyl chloride (1.4 ml, 0.01 mol) was added. The reaction
mixture was heated under reflux for 2 h. The solvent was evapo-
rated, and the solid obtained was washed thoroughly with etha-
nol, drained, and recrystallised from hexane to give
compounds 8a–c.
2.1.5.2. 4-(3,4-Dichlorophenyl)-2-(3-nitrobenzylidene)-4-oxo-buta-
nehydrazide (6b). Yield: 58%; m.p.: 160–162 ꢀC; IR (KBr) ꢀ (cmꢁ1):
3315 (NH), 3257 (NH2), 1697,1656 (2 C¼O), 1528, 1360 (NO2); MS
(EI) m/z: 393 (Mþ, 0.27%), 375 (M – H2O, 100%); 1H-NMR (DMSO-
d6, 300 MHz) d (ppm): 3.24 (s, 2H, CH2), 4.57 (s, 2H, NH2, D2O-
exchangable), 6.92 (s, 1H, NHCO, D2O-exchangable), 7.35–8.32 (m,
8H, Ar-H þ CH-methyne); Anal. Calcd. for C17H13Cl2N3O4 (393.04):
C, 51.91; H, 4.33; N, 10.69%. Found: C, 51.75; H, 4.55; N, 10.74%.
2.1.7.1. 2-Benzoyl-5-(2-nitrobenzylidene)-3-(3,4-dichlorophenyl)-1H-
pyridazin-6-one (8a). Yield: 84%; m.p.: 222–224 ꢀC; IR (KBr) ꢀ
(cmꢁ1): 3275 (NH), 1740,1660 (2 C¼O),1512, 1343 (NO2); MS (EI)
m/z: 481 (M þ 2, 1.17%), 479 (Mþ, 1.69%), 105 (benzoyl, 100%);
1H-NMR (DMSO-d6, 300 MHz)
d
(ppm): 6.69 (s, 1H, CH-
2.1.5.3. 4-(3,4-Dichlorophenyl)-2-(4-dimethylaminobenzylidene)-4-
oxo-butanehydrazide (6c). Yield: 56%; m.p.: 154–156 ꢀC; IR (KBr) ꢀ
(cmꢁ1): 3310 (NH), 3258 (NH2), 1751,1672 (2 C¼O); MS (EI) m/z:
393 (M þ 2, 9.38%), 391 (Mþ, 14.69%), 364 (M þ 2 – NHNH2,
methyne),7.48–8.20 (m, 13H, Ar-H), 11.30 (s, 1H, NH, D2O-exchang-
able); Anal. Calcd. for C24H15Cl2N3O4 (479.04): C, 60.02; H, 3.15; N,
8.75%. Found: C, 60.34; H, 3.01; N, 8.94%.
1
32.60%), 360 (M – NHNH2, 47.87%); H-NMR (DMSO-d6, 300 MHz) d
2.1.7.2. 2-Benzoyl-5-(3-nitrobenzylidene)-3-(3,4-dichlorophenyl)-1H-
pyridazin-6-one (8b). Yield: 40%; m.p.: 150–152 ꢀC; IR (KBr) ꢀ
(cmꢁ1): 3250 (NH), 1717,1677 (2 C¼O),1523, 1350 (NO2); MS (EI) m/
z: 479 (Mþ, 2.82%), 105 (benzoyl, 100%); 1H-NMR (DMSO-d6,
300 MHz) d (ppm): 7.03–8.59 (m, 13H, Ar-H þ CH-methyne), 11.34
(s, 1H, NH, D2O-exchangable); Anal. Calcd. for C24H15Cl2N3O4
(479.04): C, 60.02; H, 3.15; N, 8.75%. Found: C, 60.12; H, 3.56;
N, 8.91%.
(ppm): 2.82 (s, 2H, CH2), 2.92 (s, 6H, N(CH3)2), 4.39 (s, 2H, NH2,
D2O-exchangable), 6.73 (s, 1H, NHCO, D2O-exchangable), 6.67–7.61
(m, 8H, Ar-H þ CH-methyne); Anal. Calcd. for C19H19Cl2N3O2
(391.01): C, 58.31; H, 4.86; N, 10.74%. Found: C, 58.66; H, 5.05;
N, 10.77%.
2.1.6. General procedure for the synthesis of compounds 7a–c
Method 1: A solution of hydrazides 6a–c (0.01 mol) in HCl/AcOH
(1:3) was refluxed for 3 h. The solid that separated after concentra-
tion and cooling was recrystallised from ethanol to obtain com-
pounds 7a–c.
Method 2: To a solution of the furanones 2a–c (0.01 mol) in
absolute ethanol (20 ml), hydrazine hydrate (3.5 ml, 0.11 mol) was
added. The reaction mixture was refluxed for 4 h, then cooled and
2.1.7.3. 2-Benzoyl-5-(4-dimethylaminobenzylidene)-3-(3,4-dichloro-
phenyl)-1H-pyridazin-6-one (8c). Yield: 37%; m.p.: 229–231 ꢀC; IR
(KBr) ꢀ (cmꢁ1): 3225 (NH), 1687,1664 (2 C¼O); MS (EI) m/z: 479
(M þ 2, 33.09%), 474 (Mþ, 49.44%), 105 (benzoyl, 100%); 1H-NMR
(DMSO-d6, 300 MHz) d (ppm): 3.03 (s, 6H, N(CH3)2), 6.76–8.01 (m,
14H, Ar-H þ CH-methyne), 11.17 (s, 1H, NH, D2O-exchangable);
poured onto ice water. The product obtained 7a–c was filtered Anal. Calcd. for C26H21Cl2N3O2 (477.09): C, 65.28; H, 4.42; N, 8.78%.
off, washed with hexane.
Found: C, 65.31; H, 4.68; N, 8.23%.

JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY
981
2.1.8. General procedure for the synthesis of compounds 9a–c
pre-incubated with 90 ml of 15-LOX enzyme. The reaction was
A solution of hydrazides 6a–c (0.01 mol) and carbon disulphide started by addition of 10 ml of substrate (AA) and the plate was
(3 ml) in pyridine (10 ml) was refluxed for 3 h. The reaction mixture shaken for at least 5 min. Then, 100 ml of chromogen supplied
was left to cool at room temperature and poured onto ice water; with the kit was added to each well to stop the enzymatic reac-
the product obtained was filtered, washed with water and recrys-
tallised from ethanol to give compounds 9a–c.
tion and develop the colour. The absorbance was measured at
490 nm using microplate reader.
2.1.8.1.
1-(3,4-Dichlorophenyl)-4-(2-nitrophenyl)-3-(2-thioxo-4,5-
2.2.3. TNF-a [biotinylated] inhibition assay
dihydro-1,3,4-oxadiazol-5-yl)but-3-en-1-one (9a). Yield: 40%; m.p.:
214–216 ꢀC; IR (KBr) ꢀ (cmꢁ1): 3196 (NH), 1653 (C¼O),1504, 1340
(NO2), 1254 (C¼S); MS (EI) m/z: 435 (Mþ, 0.31%); 1H-NMR (DMSO-
d6, 300 MHz) d (ppm): 4.13 (s, 2H, CH2), 7.36–8.00 (m, 8H, Ar-
H þ CH-methyne), 13.32 (s, 1H, NH, D2O-exchangable); Anal. Calcd.
for C18H11Cl2N3O4S (435.04): C, 49.56; H, 2.54; N, 9.63%. Found: C,
49.38; H, 2.76; N, 9.54%.
The assay was performed in triplicates using TNFR2: TNF-a [bioti-
nylated] Inhibitor Screening Assay Kit (BPS bioscience, San Diego,
CA, Catalog #79756), following the manufacturer’s protocol. First,
TNFR2 is coated on a 96-well plate. Next, biotinylated TNF-a is
incubated with TNFR2 on the plate. Finally, the plate was treated
with streptavidin-HRP followed by addition of an HRP substrate to
produce chemiluminescence, which can be measured using a
chemiluminescence reader.
2.1.8.2.
1-(3,4-Dichlorophenyl)-4-(3-nitrophenyl)-3-(2-thioxo-4,5-
dihydro-1,3,4-oxadiazol-5-yl)but-3-en-1-one (9b). Yield: 58%; m.p.:
242–244 ꢀC; IR (KBr) ꢀ (cmꢁ1): 3195 (NH), 1653 (C¼O),1514, 1346
(NO2), 1238 (C¼S); MS (EI) m/z: 435 (Mþ, 0.07%); 1H-NMR (DMSO-
d6, 300 MHz) d (ppm): 3.99 (s, 2H, CH2), 7.38–8.57 (m, 8H, Ar-
3. Results and discussion
3.1. Chemistry
H þ CH-methyne), 13.32 (s, 1H, NH, D2O-exchangable); Anal. Calcd. Overall, 22 new compounds and two reported compounds were
for C18H11Cl2N3O4S (435.04): C, 49.56; H, 2.54; N, 9.63%. Found: C, synthesised as revealed in Schemes A and B. 2-Furanone deriva-
49.37; H, 2.34; N, 9.41%.
tives 2a–c were synthesised from 4-oxobutanoic derivative 1 by
reacting with aromatic aldehydes in acetic anhydride following
modified Perkin reaction conditions23,27–29,35
.
2.1.8.3. 1-(3,4-Dichlorophenyl)-4-(4-dimethylaminophenyl)-3-(2-thi-
oxo-4,5-dihydro-1,3,4-oxadiazol-5-yl)but-3-en-1-one (9c). Yield:
89%; m.p.: 162–164 ꢀC; IR (KBr) ꢀ (cmꢁ1): 3200 (NH), 1655 (C¼O),
1230 (C¼S); MS (EI) m/z: 433 (Mþ, 1.4%); 1H-NMR (DMSO-d6,
300 MHz) d (ppm): 2.84 (s, 6H, N(CH3)2), 3.72 (s, 2H, CH2), 6.65–8.59
(m, 8H, Ar-H þ CH-methyne), 13.22 (s, 1H, NH, D2O-exchangable);
Anal. Calcd. for C20H17Cl2N3O2S (433.04): C, 55.31; H, 3.95; N,
9.67%. Found: C, 55.64; H, 3.47; N, 9.21%.
The required 4-oxobutanoic derivative 1 was prepared by con-
densing dry dichlorobenzene with succinic anhydride in presence
of anhydrous aluminium chloride, following Friedel–Crafts acyl-
ation reaction conditions.
As revealed in Scheme A, 2-furanone derivatives 2a–c proved
to be useful precursors in the synthesis of several heterocyclic and
non-heterocyclic derivatives. When they were allowed to react,
separately, with ammonium acetate; pyrrol-2-one derivatives 3a–c
were produced. 4-oxobutanamides 4a–c were prepared by reflux-
ing 2-furanones 2a–c with benzylamine and 1-phenylpyridazinone
5a–c were also prepared from 2-furanones by refluxing with phe-
2.2. Biological evaluation of anti-inflammatory activity
2.2.1. Cyclooxygenase (COX-1 and COX-2) inhibition assay
nyl hydrazine according to the reported procedure29,31
.
Tested compounds were dissolved in DMSO. Each compound was
tested in triplicates using a COX inhibitory screening assay kit
according to the manufacturer (Cayman test kit-560131, Cayman
Chemical, Ann Arbor, MI). The COX inhibitor screening assay
depends on direct measurement of the amount of PG2a produced
in the COX reaction. Celecoxib, rofecoxib, and indomethacin were
used as the positive control for inhibition of COX-1 and COX-2. An
aliquot of 20 ml of each test compound or solvent (100% initial
activity) was added to 950 ml of Reaction Buffer (0.1 M Tris–HCl,
pH 8.0, containing 5 mM EDTA and 2 mM phenol), 10 ml of haem,
and 10 ml of COX-1 or COX-2, then incubated with the enzymes at
37 ꢀC for 10 min. The reaction was initiated by addition of 10 ml
AA to all the test tubes and incubation at 37 ꢀC for an additional
2 min. Enzyme catalysis was terminated by addition of 50 ml 1 M
HCl and 100 ml of the saturated stannous chloride solution. The
PGs are quantified by enzyme immunoassay (EIA) at 410 nm.
Pyridazinone derivatives 7a–c can be prepared by first stirring
of 2-furanone derivatives 2a–c with hydrazine hydrate to obtain
hydrazide derivatives 6a–c which can be cyclised by refluxing in
HCl to obtain the desired pyridazinone derivatives 7a–c. They can
be prepared directly by refluxing 2-furanone derivatives 2a–c with
hydrazine hydrate.
Hydrazide derivatives 6a–c were refluxed with benzoyl chloride
to obtain 2-benzoylpyridazinone derivatives 8a–c.
Finally, oxadiazole-thione derivatives 9a–c were prepared via
refluxing hydrazide derivatives 6a–c with carbon disulphide in
pyridine2,22,23,27,49 as revealed in Scheme B.
3.2. Anti-inflammatory activity results
Inflammation is a protective defence response of the body.
However, during inflammation, several pathological changes
occur, which involve the release of common mediators of inflam-
mation like PGs, histamine, nitric oxide, leukotrienes (LTB4), plate-
let-activation factor, lipoxins, and cytokines50. The inflammatory
response must be terminated; using anti-inflammatory com-
pounds, when no more needed to prevent avoidable harmful bio-
logical processes51. Inhibition of eicosanoids generation, in
2.2.2. Lipoxygenase (15-LOX) inhibition assay
The experiment was performed in triplicates using Cayman’s LOX
inhibitor screening assay kit (Cayman test kit-760700, Cayman
Chemical, Ann Arbor, MI) according to the manufacturer’s proto-
col. DMSO was used as 100% initial activity and Quercetin was
used as the positive control. Briefly, in a 96-well plate, 10 ml addition to the release of the pro-inflammatory cytokine; TNF-a
of each test compound (dissolved in DMSO) or vehicle were from macrophages are used for in vitro inflammation tests52.

982
R. H. A. EL-HAMEED ET AL.
Succinic anhydride
anh. AlCl3
ArCHO
Ac2O
1
Ar; a= 2-NO2-C6H4
b= 3-NO2-C6H4
2a-c
c= 4-N(CH3)2-C6H4
CH3COONH4
reflux
PhCH2NH2
reflux
reflux
PhNHNH2
3a-c
4a-c
5a-c
Scheme A. Synthesis of compounds 2a–c to 5a–c.
NH2NH2
stirring
6a-c
2a-c
NH2NH2
reflux
HCl
reflux
CS2
reflux
PhCOC
Pyridin
e
l
reflux
7a-c
9a-c
8a-c
Scheme B. Synthesis of compounds 6a–c to 9a–c.
Thus, the newly synthesised compounds were tested for their
Compound 2c was the most potent COX-1 inhibitor (IC50
anti-inflammatory activity against isozymes COX-1 and COX-2, 3.93 0.06 mM) among test compounds, being 39 times less active
which were determined by the COX-catalysed transformation of than the reference drug indomethacin (IC50¼0.10 mM) indicating
AA into PGH2 that was reduced to PGF2a and detected by the the selective COX-2 inhibition activity of all test compounds.
EIA53, as shown in Table 1.
Pyridazinone derivatives 5b, 8b, and 8c were the most potent

JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY
983
Table 1. COX-1, COX-2, and 15-LOX enzymes inhibition activities of the synthes-
Table 2. Inhibitory effects of tested compounds against TNF-a.
ised compounds.
TNF-a
COX1
IC50 (mm)
COX 2
IC50 (mm)
Selectivity indexes
(SI)
LOX
IC50 (mm)
Compound number
IC50 (nM)
Compound number
2a
7.47 0.12
7.77 1.68
8.20 0.10
5.93 0.15
8.27 0.12
6.77 0.12
4.77 0.12
3.53 0.12
5.47 0.12
4.93 0.06
3.27 0.12
3.57 0.12
5.87 0.12
4.10 0.10
6.17 0.06
8.53 0.12
8.17 0.06
7.57 0.06
3.83 1.44
3.47 0.06
2.90 0.10
5.57 0.06
4.33 0.12
6.27 0.15
6.70 0.12
2a
2b
2c
3a
3b
3c
4a
4b
4c
5a
5b
5c
6a
6b
6c
7a
7b
7c
8a
8b
8c
9a
9b
9c
4.53 0.12 0.35 0.01
6.97 0.12 0.28 0.01
3.93 0.06 0.42 0.02
8.37 0.12 0.19 0.01
6.87 0.12 0.39 0.01
5.93 0.15 0.34 0.01
11.07 0.15 0.05 0.00
13.33 0.15 0.06 0.00
11.90 0.10 0.05 0.00
12.73 0.12 0.06 0.00
13.00 0.10 0.04 0.00
11.73 0.12 0.09 0.00
9.67 0.06 0.09 0.00
10.07 0.15 0.08 0.00
8.83 0.06 0.10 0.01
7.03 0.15 0.17 0.01
6.67 0.12 0.25 0.01
8.33 0.06 0.12 0.01
10.77 0.15 0.07 0.00
12.67 0.12 0.04 0.00
13.07 0.12 0.04 0.00
9.43 0.12 0.28 0.01
10.43 0.12 0.08 0.00
8.93 0.06 0.22 0.01
14.70 0.06 0.05 0.00
14.50 0.10 0.03 0.00
0.10 0.00 0.08 0.00
12.94
24.89
9.36
44.05
17.62
5.10 0.10
4.40 0.10
6.13 0.12
2.33 0.12
1.87 0.15
2.21 0.98
5.47 0.12
5.87 0.06
4.67 0.06
2.70 0.95
2.43 0.12
2.97 0.12
2.83 0.06
2.47 0.06
3.17 0.06
5.47 0.12
5.87 0.06
4.67 0.06
2.33 0.51
2.47 0.06
3.17 0.06
2.07 0.15
1.63 0.15
2.00 0.17
ND
2b
2c
3a
3b
3c
4a
4b
4c
5a
5b
5c
6a
6b
6c
7a
7b
7c
8a
8b
8c
9a
9b
9c
17.44
221.40
222.17
238.00
212.17
325.00
130.33
107.44
125.88
88.30
41.35
26.68
69.42
153.86
316.75
326.75
33.68
130.38
40.59
326.67
580.00
1.25
Certolizumab
Celecoxib
Rofecoxib
Indomethacin
Quercetin
Data are presented as mean SD of three experiments.
ND
ND
3.34 0.05
ND
ND
ND
Data are presented as mean SD of three experiments.
ND: not determined; SI ¼ IC50 COX-1/COX-2 ratios.
interest in the development of anti-inflammatory agents60. This
declares the expected activity of N-phenyl and N-benzoylpyridazi-
nones derivatives as reported for some other derivatives with the
same scaffold61 . On the other hand, compounds 5a, 5c, 6a, 6b,
8a, and 9b exhibited a little lower COX-2 inhibition but still show
high dual COX-2/15-LOX inhibition activity.
Moreover, the ability to inhibit TNF-a, which is a potent pro-
inflammatory chemokine was tested; results are shown in Table 2.
The 15-LOX pathway was reported to induce inflammation
through increased expression of IL-6, IFN-c, IL-12, and TNF-a62.
Sixteen compounds of our newly synthesised tested com-
pounds; pyrrolone 3a, amides 4a–c, N-phenylpyridazinones 5a–c,
hydrazides 6a–c, N-benzoylpyridazinones 8a–c, and oxadiazoles
9a–c showed higher activities (IC50 ¼ 2.90 ꢁ 6.27 mM range) than
the used reference inhibitor Certolizumab (IC50¼6.70 mm). While
compound 3c showed comparable activity with IC50¼6.77 mM.
Compound 8c showed the lowest IC50 value for TNF-a inhibition,
selective COX-2 inhibitors (IC50¼0.04 mM for the three com-
pounds), being superior to the reference drug (celecoxib
IC50¼0.05 mM). While amide derivatives; 4a–c, hydrazides; 6a, 6b,
N-phenyl and N-benzoylpyridazinones; 5a, 5c, 8a and oxadiazole;
9b (IC50¼0.05–0.09 mM range), showed comparable activity to cel-
ecoxib as selective COX-2 inhibitors. These results indicate the
high activity of hydrazides, pyridazinones, and oxadiazole; which
is in consistent with previously reported activities of similar scaf-
folds as selective COX-2 inhibitors37,42,54–57
.
Studies showed that soybean LOX enzyme has a wide sub-
strate specificity, and that the oxygenation sites for the soybean
LOX has been demonstrated to be at C-13 of a-linolenic acid and
C-15 of AA58. Therefore, the soybean LOX is most like the mam-
malian 15-LOX59.
Consequently, the in vitro inhibitory effect of all synthesised
compounds against 15-LOX was determined using a lipoxygena-
tion reaction to transform AA into hydroperoxides, which were
detected by the addition of a chromogen, whose results are dis-
played in Table 160.
Results also revealed that all the synthesised derivatives of pyr-
rolones 3a–c, hydrazides 6a–c, N-phenyl and N-benzoylpyridazi-
nones; 5a–c, 8a–c, and oxadiazoles 9a–c inhibited 15-LOX (IC50
¼1.63 ꢁ 3.17 mM range), 9b being the most potent 15-LOX inhibi-
tor (IC50¼1.63 0.15 mM), compared to the positive control
Quercetin (IC50¼3.34 mM). However, furanones 2a–c, amides 4a–c,
and pyridazinones 7a–c exhibited lower 15-LOX inhibitory effect.
By evaluating the COX and 15-LOX inhibition results, three
compounds namely 6–(3,4-dichlorophenyl)-4–(3-nitrobenzylidene)-
1-phenyl-1,4-dihydropyridazin-3(2H)-one (5b), 2-benzoyl-5–(3-nitro-
benzylidene)-3–(3,4-dichlorophenyl)-1H-pyridazin-6-one (8b), and
2-benzoyl-5–(4-dimethylaminobenzylidene)-3–(3,4-dichlorophenyl)-
1H-pyridazin-6-one (8c) seemed to be the best candidates as a
indicating that this activity may be
LOX inhibition.
a consequence of 15-
To analyse the structure–activity relationship (SAR) of the
tested compounds from the previous results, it is obvious that our
three newly synthesised furanones are totally inactive towards all
the tested enzymes. But upon their conversion to other heterocyc-
lic and non-heterocyclic compounds the biological activities
appear as revealed in Figure 2 and explained as follows:
3 Pyrrolone derivatives 3a–c acquired high activity as LOX-
inhibitors, only one of them having 3-nitrobenzylidene substituent
(3b) showed high activity as TNF-a inhibitor. They still have no
significant activities as COX-2 inhibitors.
Upon ring opening and formation of amide derivatives 4a–c, a
high TNF-a inhibition appears with significant selective COX-2
inhibition, but no significant LOX-inhibition observed.
Also, ring opening and hydrazides formation in 6a–c showed
activity against 15-LOX and TNF-a, while 6a,b having 2-nitroben-
zylidene and 3-nitrobenzylidene substituents showed high activity
COX-2/15-LOX dual inhibitors, which is a current subject of as selective COX-2 inhibitors.

984
R. H. A. EL-HAMEED ET AL.
3a,3c have higher activity as LOX and TNF-
3b has higher activity as LOX inhibitor
α
inhibitors
6a,6b have higher activity against 3 enzymes
6c has higher activity as LOX and TNF-αinhibitors
2a-c
Inactive compounds
4a,4b,4c have higher activity as
5a-c and 8a-c have higher activity
against 3 enzymes
7a-c are inactive compounds
9a,9c have higher activity as LOX and TNF-
9b has higher activity against 3 enzymes
α
inhibitors
TNF-α
inhibitor andsignificant
selective COX2 inhibitor
Figure 2. Structure–activity relationship of the tested compounds against the tested enzymes.
Conversion of furanones to pyridazinines 7a–c did not affect
their biological activities as they were still inactive. But formation
of N-phenylpyridazinones 5a–c and N-benzoylpyridazinones 8a–c
was a perfect pathway for highly potent derivatives against all the
tested enzymes with desirable dual COX-2/15-LOX inhibition activ-
ities for compounds 5b, 8b, and 8c indicating the important influ-
ence of presence of 3-nitrobenzylidene substituent in both
pyridazinones (5b, 8b) and 4-dimethylaminobenzylidene in N-ben-
zoylpyridazinone (8c).
Finally, three oxadiazole derivatives 9a–c showed high activity
against 15-LOX and TNF-a, while two derivatives, having 2-nitro-
benzylidene and 4-dimethylaminobenzylidene substituents, 9a
and 9c, respectively, showed high activity as selective COX-
2 inhibitors.
Funding
Authors are grateful to Science and Technology Development
Fund (STDF) for financial support of this research.
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