Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 17 (2007) 6439–6443
Synthesis and anti-inﬂammatory/antioxidant activities of some
new ring substituted 3-phenyl-1-(1,4-di-N-oxide
quinoxalin-2-yl)-2-propen-1-one derivatives and of their
Asuncio´n Burguete,a Eleni Pontiki,b Dimitra Hadjipavlou-Litina,b,* Raquel Villar,a
Esther Vicente,a Beatriz Solano,a Saioa Ancizu,a Silvia Pe´rez-Silanes,a
aUnidad de Investigacio´n y Desarrollo de Medicamentos, Centro de Investigacio´n en Farmacobiologı´a Aplicada (CIFA),
Universidad de Navarra, c/Irunlarrea s/n, 31080 Pamplona, Spain
bDepartment of Pharmaceutical Chemistry, School of Pharmacy, Aristotle University of Thessaloniki, Thessaloniki 54124, Greece
Received 7 September 2007; revised 1 October 2007; accepted 1 October 2007
Abstract—We report the synthesis, anti-inﬂammatory and antioxidant activities of novel ring substituted 3-phenyl-1-(1,4-di-N-oxidequinoxalin-2-yl)-2-propen-1-one derivatives and of their 4,5-dihydro-(1H)-pyrazole analogues. The tested compounds inhibit thecarrageenin-induced rat paw edema (4.5–56.1%) and present important scavenging activities. Compound 2a is the most potent(56.1%) in the in vivo experiment and exhibits promising in vitro inhibition of soybean lipoxygenase (IC50 < 1 lM). Ó 2007 Elsevier Ltd. All rights reserved.
Quinoxalines, including their fused-ring derivatives, dis-
Systemic hypoxia increases reactive oxygen species gen-
play diverse pharmacological activities (antiviral, anti-
eration and promotes leukocyte-endothelial adherence
cancer, and antibacterial).Scientists in Belgium and
via reactive oxidant generation. Thus, antioxidants pre-
the United Kingdom have found that quinoxaline is a
vent the increase in leukocyte-endothelial adhesive inter-
potential treatment for HIV infection, and works well
actions observed in hypoxia.Bioreductive drugs have
with lamivudine, abacavir, and efavirenz.Oxidation
been designed to take advantage of the particular meta-
of both nitrogens of the quinoxaline ring dramatically
bolic characteristics of hypoxic cells.
increased the diversity of certain biological properties,such as antibacterial activitand hypoxia-selective
The formation of reactive oxygen species (ROS) is char-
anticancer activityMonge et are involved in the
acteristic of aerobic organisms that normally defend
synthesis and biological evaluation of new agents de-
themselves against these highly reactive species using en-
rived from quinoxaline 1,4-di-N-oxide and related com-
zymes, like superoxide dismutase and glutathione perox-
pounds that have proved to be eﬃcient cytotoxic agents
idase and naturally occurring antioxidantsROS, like
for hypoxic cells in solid tumors. The poor tumor vascu-
superoxide radical anion, hydrogen peroxide, and hy-
lar structure, the ineﬃcient blood supply along with a
droxyl radical, are produced during the inﬂammation
high interstitial pressure generate a variable proportion
process by phagocytic leukocytes (e.g., neutrophils,
of viable hypoxic cells in solid tumors which is one of
monocytes, macrophages, eosinophils) that invade the
the causes of cell resistance to anticancer treatments.
tissue. Moreover, these reactive species are involved inthe biosynthesis of prostaglandins and in the cycloxy-genase- and lipoxygenase-mediated conversion of ara-
Keywords: Quinoxaline 1,4-di-N-oxide; a,b-Unsaturated ketone; 4,5-
chidonic acid into proinﬂammatory intermedi
Persistently high levels of ROS may involve pathological
* Corresponding author. Tel.: +30 2310997627; fax: +30 2310997679;
conditions, as the active species can modify essentially
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmcl.2007.10.002
A. Burguete et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6439–6443
biological molecules, such as lipids, proteins, and DNA.
measure of the overall lipophilic/hydrophilic balance
It is therefore evident that the treatment of the above-
of these molecules. We could attribute this to the diﬀer-
mentioned pathophysiological conditions could beneﬁt
ent nature of the hydrophilic and lipophilic phases in the
from the use of drugs that combine antioxidant and
two systems and to the presence of basic nitrogen atoms
anti-inﬂammatory activity, as has already been proven
in the examined compounds, which could disturb the
for a number of commercially available non-steroidal
anti-inﬂammatory drugs (NSAIDs), for example, tolfe-namic acid which simultaneously possess radical scav-
In acute toxicity experiments, the studied compounds
did not present in vivo toxic eﬀects at doses up to0.5 mmol/ml/kg body weight. The in vivo anti-inﬂam-
There is increasing evidence from animal models and
matory eﬀects of the tested compounds were assessed
by using the functional model of carrageenin-induced
(LOX) and their products may play a role in tumor for-
rat paw edemaand are presented in as per-
mation and cancer metastasis.Recently the concept
centage inhibition of weight increase at the right hind
has been put forward that LOX activation may be in-
paw in comparison to the uninjected left hind paw
volved in both pro- and anti-tumorigenic eﬀects.Ara-
chidonate lipoxygenase pathway appears to play a rolein brain tumor growth as well as inhibition of apoptosis
All the tested compounds (dose ip 0.01 mmol/ml/kg
in in vitro studies. Emerging reports now indicate alter-
body weight), after 3.5 induced protection (ran-
ations of arachidonic acid metabolism related to carci-
ged from 4.5% to 56.1%) against carrageenin induced
nogenesis and many anti-inﬂammatory drugs are being
paw edema while the reference drug indomethacin
investigated as potential anticancer drugs.
(IMA) induced 47% protection at an equivalent dose.Compounds 2a and 4a were the most potent (56.1% and
So, on the basis of these results, it seemed interesting to
55.4%, and presented almost equipotent eﬀect.
synthesize some novel ring substituted 3-phenyl-1-(1,4-
Among derivatives 2a, 2b, and 2c, compound 2a was
di-N-oxide quinoxalin-2-yl)-2-propen-1-one derivatives
found to be the most potent followed by 2b and 2c
and their 4,5-dihydro-(1H)-pyrazole analogues. Repre-
(2b > 2c). The existence of the pyrazolyl ring decreases
sentative compounds have been tested, in order to study
the biological response (compounds 5a and 5b) whereas
their scavenging activities, their role in inﬂammation,
the condensed ring –OCH2O– (compound 3a) is corre-
and their inhibition of LOX since LOX inhibitors are
lated with a very signiﬁcant loss in inhibition.
able to induce the anti-carcinogenic and/or to inhibitthe pro-carcinogenic enzymes responsible for polyunsat-
Compounds 2a and 4a, the most potent in vivo, as well
as compounds 2b, 2c, 3a, 5a, and 5b were further evalu-ated for inhibition of soybean lipoxygenase LOX by the
Synthesis of the derivatives 2a–c, 3a–c, and 4a–c
() was carried out by a base-catalyzed Clais-
oxidize certain fatty acids at speciﬁc positions to hydro-
en–Schmidt condenestablishing a required
peroxides that are the precursors of leukotrienes, which
temperature of À10 °C. The synthesis of compounds
contain a conjugated triene structure. It is known that
5a–c was carried out by dissolution of derivatives 4a–c
soybean lipoxygenase, which converts linoleic to 13-
in absolute ethanol and subsequent addition of hydra-
hydroperoxylinoleic acid, is inhibited by NSAIDs in a
zine hydrate.The starting reagents used (1a–c) were
qualitatively similar way to that of the rat mast cell
lipoxygenase and may be used in a reliable screen forsuch activity. Perusal of % inhibition values or IC50 val-
All the synthesized compoundswere characterized by
ues () shows that compound 2a (IC50 < 1 lM) is
infrared, proton nuclear magnetic resonance, elemental
the most active, within the set, followed by compounds
analysis of C, H, and N, and melting point.
Non-steroidal anti-inﬂammatory drugs (NSAIDs) are
Most of the LOX inhibitors are antioxidants or free rad-
widely used for the treatment of pain, fever, and inﬂam-
ical scavengers, since lipoxygenation occurs via a carbon-
mation. All of the NSAIDs are approximately equiva-
centered radical. Although lipophilicity is referredto
lent in terms of anti-inﬂammatory eﬃcacy but also
as an important physicochemical property for LOX
cause untoward side eﬀects (such as gastrointestinal
inhibitors, all the above tested derivatives do not follow
ulcers, hemorrhages) in a signiﬁcant fraction of treated
patients and this fact frequently limits therapy. The vari-ations in both eﬃcacy and their tolerability are partly
Many non-steroidal anti-inﬂammatory drugs have been
due to diﬀerences in their physicochemical properties,
reported to act either as inhibitors of free radical pro-
which determine their distribution in the body and their
duction or as radical scavengers. Compounds with anti-
ability to pass through and to enter cells.Thus par-
oxidant properties could be expected to oﬀer protection
tition coeﬃcients such as RM values are determined
in rheumatoid arthritis and inﬂammation and to lead to
experimentallyand compared with the corresponding
potentially eﬀective drugs. Thus, we tested the new
theoretically calculated clog P in n-octanol-buf-
derivatives with regard to their antioxidant ability and
fer. From our results () it can be concluded that
in comparison to well-known antioxidant agents, for
RM values could not be used as a successful relative
example, nordihydriguaiaretic acid (NDGA), trolox,
A. Burguete et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6439–6443
1b: R7=F 1c: R7=CH O 2c: R7=CH O 4c: R7=CH O 3c: R7=CH O 5a: R7=H 5b: R7=F 5c: R7=CH O
Scheme 1. Reagents and conditions: (a) 3-methoxy-4-(tetrahydro-pyran-2-yloxy)-benzaldehyde, 3% NaOH/methanol, À10 °C; HCl 35%; (b)benzo[1,3]dioxole-5-carbaldehyde, 3% NaOH/methanol, À10 °C; (c) 3,4,5-trimethoxy-benzaldehyde, 3% NaOH/methanol, À10 °C; (d) NH2NH2,ethanol, rt.
Table 1. Experimentally determined R values and theoretically
and caﬀeic acid. The interaction of the examined com-
calculated clog P17 values; inhibition% of induced carrageenin rat paw
pounds with the stable free radical DPPH was studied
edema (CPE%) at 0.01 mmol/ml/kg; in vitro inhibition of soybean
by the use of the stable 1,1-diphenyl-2-picrylhydrazyl
radical DPPH at 0.05 and 0.1 mM after 20 and 60 min
). This interaction indicates their radical scav-
enging ability in an iron-free system. Compounds 2b
and 2c interact with DPPH in a concentration and timedependent manner, whereas compounds 2a, 3a, and 4a
do not present any interaction at 0.05 mM. Slight diﬀer-
ences are observed between the compounds 2c and 5b
with the time and the concentration whereas compound
2a presents reducing ability at 0.1 mM. The presence of
the condensed ring –OCH2O– (compound 3a) dimin-
ishes the reducing activity. Preliminary QSAstudies
on the values of DPPH interaction have shown that the
molar refractivity (MR) of substituents R7 plays a sig-
niﬁcant role. Molar refractivity (MR) is related not only
to the volume of the substituents but also to the London
dispersive forces. Thus, high molar refractivity values of
substituent R7– (MR for R7–OCH3 = 0.78, MR for R7–
RM values are the average of at least 10 measurements; each experi-
H = 0.103, and MR for R7–F = 0.092) aﬀect the reduc-
ment was performed at least in triplicate and the standard deviation
ing ability. No role for lipophilicity is deﬁned.
of absorbance was less than 10% of the mean.
b Theoretically calculated values of lipophilicity. c Statistical studies were done with Student’s t-test, *p < 0.01,
The insertion of a pyrazolyl ring (compounds 5a and 5b)
increases the reducing ability. The R7 = –F substitution
A. Burguete et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6439–6443
Table 2. Interaction % with DPPH at 0.05 mM and at 0.1 mM
Competition % with DMSO for hydroxyl radical (HOÅ %); superoxide radical scavenging activity (PMS %). No, no result under the experimental conditions; NDGA, nordihydroguaiaretic acid; CA acid, caﬀeic acid; each experiment was performed at least intriplicate and the standard deviation of absorbance was less than 10% of the mean.
in compound 5b is correlated with an increase compared
Further investigation is in progress to delineate the
to compound 5a. Further investigations are in progress
physicochemical properties implicated in the in vivo re-
in order to have a detailed structure–activity relation-
sponse. Regression analysis was performed to discover
ship study on their interaction with DPPH.
whether any correlation existed between anti-inﬂamma-tory activity and several physicochemical parameters
It is consistent that rates of reactive oxygen species
(lipophilicity, steric and electronic variables). Unfortu-
nately the conﬁdence limits were found to be poor.
mainly due to their transformation into ÅOH, reactive
For the in vivo results the following equation was
radical metal complexes, and 1O2. During the inﬂamma-
tory process, phagocytes generate the superoxide anion
log %ðCPEÞ ¼ À1:312ðÆ0:797ÞclogP þ 1:902ðÆ0:367Þ
radical at the inﬂamed site and this is connected to otheroxidizing species as ÅOH. Hydroxyl radicals are among
the most reactive oxygen species and are considered to
be responsible for some of the tissue damage occurring
in inﬂammation. It has been claimed that hydroxyl rad-ical scavengers could serve as protectors, thus increasing
Hydrophilicity (lipophilicity with negative sign) is the
most signiﬁcant parameter. Compound 5b, the morelipophilic, is not included in the regression. This fact
The competition of compounds with dimethylsulfoxide
proceeds in parallel to the observation that low lipophil-
(DMSO) for OH radicals,generated by the Fe3+/
icity is highly involved to the biological response. At-
ascorbic acid system, expressed as the inhibition of
tempts to correlate the in vivo/in vitro expressions of
formaldehyde production, was used for the evaluation
of their hydroxyl radical scavenging activity. All the
M values in a linear or non-linear regres-
sion analysis gave statistically non-signiﬁcant correla-
tested derivatives show high inhibition of DMSO
tions. Unfortunately the number of compounds is not
(33 mM) oxidation at 0.1 mM Lipophilicity
enough to calculate a combination of all the eﬀects.
is not well correlated with the results. Antioxidantsof hydrophilic or lipophilic character are both neededto act as radical scavengers in the aqueous phase
The authors are grateful to Drs. C. Hansch and A. Leoand to Biobyte Corp. for the free access to the C-QSAR
Non-enzymatic superoxide anion radicals were gener-
program. A. Burguete was awarded a Ph.D. fellowship
The superoxide producing system was set up by
supported by the ‘‘Gobierno de Navarra’’.
mixing phenazine methosulfate (PMS), nicotinamideadenine dinucleotide NADH and air–oxygen. The pro-duction of superoxide was estimated by the nitroblue
tetrazolium method. The majority of the compoundsdoes not present scavenging activity at 0.1 mM
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Analysis of Susanna Roxman’s poem “Cornelia” Introduction An Example of Roman Virtues Cornelia was a highly educated Roman woman who lived in the second century B.C. She was the daughter of a Roman hero, Publius Cornelius Scipio Africanus Major, who defeated Hannibal in the second Punic War. She married a young, most honourable plebeian, Tiberius Sempronius Gracchus an
la santé au travail RESAAD 72 _ 15 mai 2012 _ Dr Yves Rouault la santé au travail l'équipe médicale du travail est astreinte au secret médical et peut être contactée par le salarié, ses collègues de travail ou son employeur. RESAAD 72 _ 15 mai 2012 _ Dr Yves Rouault la santé au travail le médecin du travail peut aborder les consommations en tabac, a