Design and synthesis of aza-flavones as a new class of xanthine oxidase inhibitors

Article abstract of DOI:10.1002/ardp.201200296

In an attempt to develop non-purine-based xanthine oxidase (XO) inhibitors, keeping in view the complications reported with the use of purine-based XO inhibitors, the flavone framework (a class possessing XO inhibitory potential) was used as lead structure for further optimization. By means of structure-based classical bioisosterism, quinolone was used as an isoster for chromone (a bicyclic unit present in flavones), owing to the bioactive potential and drug-like properties of quinolones. This type of replacement does not alter the shape and structural features required for XO inhibition, and also provides some additional interaction sites, without the loss of hydrogen bonding and hydrophobic and arene-arene interactions. In the present study, a series of 2-aryl/heteroaryl-4-quinolones (aza analogs of flavones) was rationally designed, synthesized and evaluated for in vitro XO inhibitory activity. Some notions about structure-activity relationships are presented indicating the influence of the nature of the 2-aryl ring on the inhibitory activity. Important interactions of the most active compound 3l (IC₅₀ = 6.24 μM) with the amino acid residues of the active site of XO were figured out by molecular modeling. To develop non-purine-based xanthine oxidase inhibitors, the flavone framework was used as lead structure for further optimization. By means of structure-based classical bioisosterism, quinolone was used as an isoster for chromone. This type of replacement does not alter the shape and structural features required for xanthine oxidase inhibition. The rationally designed and synthesized series of 2-aryl/heteroaryl-4-quinolones (aza analogs of flavones) was evaluated for in vitro xanthine oxidase inhibitory activity. Copyright

Full text of DOI:10.1002/ardp.201200296

Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
7
Full Paper
Design and Synthesis of Aza-Flavones as a New Class
of Xanthine Oxidase Inhibitors
Rajni Dhiman1, Sahil Sharma1, Gagandip Singh2, Kunal Nepali1, and Preet Mohinder Singh Bedi1
1 Department of Pharmaceutical Sciences, Guru Nanak Dev University, Amritsar, Punjab, India
2 Laboratory of Drug Design, Department of Pharmaceutical Chemistry, ISF College of Pharmacy, Moga,
Punjab, India
In an attempt to develop non-purine-based xanthine oxidase (XO) inhibitors, keeping in view the
complications reported with the use of purine-based XO inhibitors, the flavone framework (a class
possessing XO inhibitory potential) was used as lead structure for further optimization. By means of
structure-based classical bioisosterism, quinolone was used as an isoster for chromone (a bicyclic unit
present in flavones), owing to the bioactive potential and drug-like properties of quinolones. This type of
replacement does not alter the shape and structural features required for XO inhibition, and also provides
some additional interaction sites, without the loss of hydrogen bonding and hydrophobic and arene–
arene interactions. In the present study, a series of 2-aryl/heteroaryl-4-quinolones (aza analogs of flavones)
was rationally designed, synthesized and evaluated for in vitro XO inhibitory activity. Some notions about
structure–activity relationships are presented indicating the influence of the nature of the 2-aryl ring on
the inhibitory activity. Important interactions of the most active compound 3l (IC50 ¼ 6.24 mM) with the
amino acid residues of the active site of XO were figured out by molecular modeling.
Keywords: Bioisosterism / Flavones / Isosteric / Quinolones / Xanthine oxidase
Received: July 24, 2012; Revised: August 29, 2012; Accepted: August 31, 2012
DOI 10.1002/ardp.201200296
Supporting information available online
:
Introduction
including inflammation, metabolic disorders, cellular age-
ing, atherosclerosis, and carcinogenesis is well reported [2, 3].
Increased XO serum level in pathological states like hepatitis,
inflammation, cancer, etc. indicates that XO inhibition may
result in broad spectrum therapeutics for gout, cancer,
inflammation, and oxidative damage [2–5]. Despite the poten-
tial of purine based compounds as xanthine oxidative inhibi-
tors such as allopurinol [3, 4], 2-alkyl hypoxanthines [6],
pterin, and 6- formylpterin [7], there is a continuous search
for non-purine based XO inhibitors. The revived interest
among the researchers toward the XO inhibitors with struc-
turally diverse and non-purine isosters [8] such as feboxustat
[9], flavones [10], FYX, a 1,3-diaryltriazole [11] derivative, and
curcumin [12] (Fig. 2) can be attributed to the interactions of
purine analogs XO inhibitors on activities of purine and
pyrimidine metabolizing enzymes like guanine deaminase,
HGPRT (hypoxanthine–guanine phosphoribosyl transferase),
PNP (purine nucleoside phosphorylase), OPRT (orotate phos-
pho-ribosyl transferase) and OMPDC (orotidine-5-monophos-
phate decarboxylase) leading to Steven Johnson syndrome
Xanthine oxidase (XO) is a versatile molybdoflavoprotein,
widely distributed, occurring in milk, kidney, lung, heart,
and vascular endothelium. In humans, the highest specific
activity for XO is found in the liver and intestine. The enzyme
is involved in the metabolism of purines, catalyzing the
oxidative hydroxylation of hypoxanthine and xanthine to
produce uric acid and reduction of oxygen at the flavin
center generating reactive oxygen species either as super-
oxide anion radical or hydrogen peroxide (Fig. 1) [1–3].
Catalysis by XO to produce uric acid and reactive oxygen
species leads to many diseases like gout and at least symp-
toms of diseases like oxidative damage to the tissue [2].
Involvement of reactive oxygen species in pathological events
Correspondence: Dr. Kunal Nepali, Department of Pharmaceutical
Sciences, Guru Nanak Dev University, Amritsar-143005, Punjab, India.
E-mail: kunal_8855@rediffmail.com
Fax: þ91-1832258819
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R. Dhiman et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
Figure 1. Role of XO.
Figure 2. Some XO inhibitors.
and worsening of renal function induced in some of the
patients [2–4].
Bioisosterism, a lead optimization technique to enhance
the biological activity or to attenuate the toxicity has proved
to be an effective methodology to create therapeutically
equivalent surrogates [15–18]. Flavonoids represent a class
of secondary metabolites possessing a diverse array of bio-
logical activities such as anti-inflammatory [19], antiallergic
[19, 20], antioxidant [20], CNS [21], vascular [22], antitumor
[22], and XO inhibitors [23–25]. This family of secondary
metabolites possesses a diverse array of biological activities
but numerous reports on the XO inhibitory potential of
flavones particularly highlights their potential as XO inhibi-
tors. The structures of flavones with potent XO inhibitory
activity are shown in Fig. 4. The pharmacophoric features
responsible for XO inhibition by this class of secondary
metabolites possessing the bicyclic benzopyrone as a core
structural unit is well reported [23–25]. The rational design of
2-aryl-4-quinolones involving the isosteric replacement of
chromones with quinolones (two being isoelectric analogues
of one another) was thought worthwhile, keeping in view
the shape and structural features of flavones responsible for
their inhibitory potential (Fig. 5).
Recently our research group reported the XO inhibitory
potential of N-acetyl pyrazolines and N-(1,3-diaryl-3-oxo-
propyl) [13, 14] amides designed via chemical modification
of 1,3-diaryl triazoles (Fig. 3). In continuation of our search
for non-purine based XO inhibitors, the present study utilizes
the concept of classical bioisosterism employing 2-phenyl-
chromone (flavone framework) as the lead structure.
Figure 3. Chemical modification of 1,3-diaryl triazoles.
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Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
Aza-Flavones as Xanthine Oxidase Inhibitors
9
Figure 4. Flavones with XO inhibitory activity.
action; (ii) an olefinic center at C-2,3 maintaining the pla-
narity of the designed molecules; (iii) an appropriately placed
carbonyl function for hydrogen bonding interaction; (iv) an
appropriately placed aryl/heteroaryl ring for hydrophobic
interactions; (v) an appropriate torsional angle; (vi) a sub-
stituent free C-3 center earlier reported to lower the binding
affinity of flavones [23–25]. As the designed molecules are
derived from non-purine based skeletons, it also excludes
the possibility of them being converted like allopurinol to
natural nucleotides.
Figure 5. Isosteric replacement of the chromone nucleus with
In the present paper, we describe the synthesis of non-
purine 2-aryl/heteroaryl-4-quinolones without ignoring
shape and structural features as a new class of XO inhibitors
for the first time. The results of the biological evaluation have
been further rationalized by the aid of computational
studies.
quinolone.
Quinolones as privileged building blocks allow the pro-
duction of large libraries of bioactive molecules owing to
their diversity and drug like properties and are considered
as central scaffolds to build promising bioactive chemical
libraries [26].
Results and discussion
Chemistry
2-Aryl-4-quinolones were synthesized (Scheme 1) via
With this background, 2-aryl/heteroaryl-4-quinolones have
been designed incorporating the quinolone nucleus within
the chemical architecture of flavones based on the consider-
ations earlier reported to be essential for the XO inhibitory
activity of flavones [23–25] (Fig. 6) such as: (i) a bicyclic
nucleus appropriate for hydrophobic and arene–arene inter-
a
sequence of reactions starting from 2-amino benzoic acid
which was converted to 2-aminoacetophenone employing
methyl lithium in dimethyl ether. The 2-amino acetophe-
none was further acetylated to prevent any imine formation
on condensation with different substituted aldehydes. All the
diaryl propenones were synthesized by base catalyzed Claisen
Schmidt condensation. The method is attractive as it selec-
1
tively yields E-isomers. From the H NMR spectra, all diaryl
propenones were geometrically pure and with trans configur-
ation (J ¼ 15.50–15.60 Hz). Diaryl propenones were further
cyclized to yield the tetrahydroquinolones by acid catalyzed
deprotection of the acetylated amine to undergo simul-
taneous b-additions to the enone system. The 2-aryl tetra-
hydroquinolones showed a classical ABX system in the
1H NMR and were further dehydrogenated to yield the di-
hydroquinolones employing molecular iodine. The dehydro-
Figure 6. Key Interactions and structural features.
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Scheme 1. Reagents and conditions: (i) CH3Li/DME, aq NH4Cl, stirring, 08C, 1 h; (ii) (CH3CO)2O, C6H5N, MeOH, stirring, 708C, 6 h;
(iii) MeOH, 5% NaOH, stirring, rt, 6 h; (iv) EtOH, 5%HCl, reflux, 12 h; (v) DMSO, I2, reflux, 12 h.
genated products were confirmed by the appearance of a
singlet of the proton at positions at 6.1–6.5 ppm.
The synthesis and biological evaluation of the series of
compounds with diverse substitutions at the aryl ring
(2nd position) such as halo, alkyl, alkoxy groups, and place-
ment of bicyclic aryl/heteroaryl, cinnamyl, and heteroaryl
at the 2nd position enabled us to correlate the dependence of
the inhibitory activity of these molecules on the electronic
and steric factors and thus a structure–activity relationship
could be established. Careful investigation of Table 2
revealed interesting observations and indicated the influence
of electronic and steric factors linked with the aryl ring
at the 2nd position on the inhibitory potential of the
molecules. In general, electronic factors such as introduction
of deactivating groups, chloro and fluoro on the 2-phenyl
ring resulted in a decrease in inhibitory activity as compared
to the activating groups such as methoxy (compare 3h, 3i
with 3b, 3c, 3d, 3e, 3g, respectively). The presence of
methoxy substituent and its position on the aryl ring played
an important part in the inhibitory potential. Placement of
a methoxy group on the aryl ring at the para position (3g)
remarkably enhances the inhibitory activity. This can be
attributed to the electron drift due to the methoxy group
leading to quinoid formation (see Supporting Information)
which enhances the rigidity of the molecules toward the
planar structure. A methoxy group at the ortho position
(3d) did not show any enhancement of the XO inhibitory
activity and the activity profile was similar to the compound
with unsubstituted phenyl ring (2nd position) (3a). The
presence of an additional methoxy group apart from one
3
However in case of aryl rings (at the 2nd position) having
oxygenated functionality at the ortho position, the signal for
the proton at C-3 was found to be deshielded. The deshielded
signal for the proton at C-3 can be attributed to intra-
molecular H bonding with the oxygen functionality at the
ortho position of the 2-aryl ring. The dehydrogenation
proceeded at C-2 and C-3. All the synthesized products
(Table 1) were characterized by spectroscopic techniques.
Biological evaluation of the synthesized compounds
for XO inhibitory activity
In vitro screening of the 2-aryl-4-quinolones (3a–3o) using
bovine milk XO (grade 1, ammonium sulfate suspension)
enzymatic assay was performed. Each synthesized compound
was tested in triplicate for the XO inhibitory activity.
Apigenin (earlier reported most active flavone) [19] and
allopurinol were used as standards for the study. Among
the series, compounds 3l, 3k, 3g were found to be most
active against XO with IC50 ranging from 6.24 to 12.0 mM
(Table 2). The most active compound 3l possessing the
pyridinyl ring at the 2nd position exhibited significant
inhibition (IC50 ¼ 6.24 mM) in comparison to apigenin
(IC50 ¼ 10.21 mM) as well as allopurinol (IC50 ¼ 8.31 mM).
Compound 3k with the thiophenyl ring also displayed
significant inhibition with an IC50 ¼ 11.23 mM followed by
3g with 4-methoxy phenyl ring (IC50 ¼ 12.02 mM).
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Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
Aza-Flavones as Xanthine Oxidase Inhibitors
11
Table 1. % Yield and melting points of various products.
Compound
R1
R2
R3
R4
R5
% Yield
mp (8C)
3a
3b
3c
3d
3e
3f
3g
3h
3i
H
H
OCH3
OCH3
OCH3
H
H
H
H
H
H
OCH3
H
H
OCH3
H
H
OCH3
H
H
H
OCH3
OCH3
Cl
H
OCH3
H
H
H
H
H
H
H
H
H
H
75
76
78
77
76
79
75
81
80
250–252
252–254
233–235
234–236
260–262
231–221
288–290
339–340
250–252
H
OCH3
OCH3
H
H
H
H
H
F
O
3j
72
76
280–281
300–302
N
H
O
3k
S
N
H
O
3l
72
74
234–235
234–235
N
H
N
O
3m
O
N
H
O
O
3n
3o
65
80
217–219
280–290
N
H
H
H
CH3
H
H
at the para position was not found to enhance the inhibitory
activity (3b, 3f). The placement of other activating groups
such as methyl did not enhance the inhibitory activity which
might be due to the absence of oxygen functionality and
thus quinoid formation is not possible (3o). The presence of
two oxygen functions (3rd and 4th position) at the phenyl
ring (2nd position) forms ortho quinoid which does not con-
tribute in bringing planarity of phenyl ring and thus no
enhancement was observed on placing additional methoxy
functionalities. A single methoxy group at the ortho position
induces steric compression directing the molecule toward
non-planarity. Apart from the electronic factors, steric
factors also influenced the inhibitory activity such as change
of the 2-phenyl ring with naphthyl (3j) leads to a drastic
decrease in inhibitory potential which could be attributed
to the steric bulk of the naphthyl. The steric bulk of naphthyl
makes the molecule non-planar and planarity is a pre-
requisite for the inhibitory activity. Change of 2-phenyl
with monocyclic heteroaryl rings such as thiophenyl (3k)
and pyridinyl (3l) significantly enhanced the XO inhibitory
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R. Dhiman et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
activity. The enhanced activity of 3k and 3l can be explained
due to an increase in rigidity toward planar structure
and by reducing the effect of rotamer formation through
entirely different modes (see Supporting Information).
Lengthening of the linker between the two rings (3n) led
to the dilution of the XO inhibitory activity which could
be due to the increase in the flexibility of the molecule
and also indicates that increasing the distance between
the two aryl rings (quinolone and the aryl) diminishes the
activity.
Leu648, Leu1014, and Pro1076. Some energetically favorable
arene–arene interactions were observed between 3l and
the receptor similar to salicylate in co-crystal of XO. The
p-electrons of the quinolone ring were interacting with
p-electrons of Phe914. The nitrogen lone pair of the pyridinyl
is involved in van der Waals interactions with the residues
in close proximity (Fig. 7a). A comparison study was done
to get a clear vision of the binding mode of apigenin and
the synthesized compound (3l) to rationalize their affinities
for XO (Fig. 7b). Apigenin and 3l show similar binding con-
formation in the binding site. In the salicylic acid binding
site, apigenin get stabilized by the two hydroxyl groups
attached to chromone ring which are involved in hydrogen
bonding with Arg880, Phe914 and Glu1261. From the dock-
ing study, it can be deduced that the 2-aryl-4-quinolone
molecule makes stronger interaction with the binding
residues of XO because of additional hydrogen bonding
between the –NH– in the quinolone ring of 3l and Glu802.
This additional hydrogen bond enhances the binding
affinity for the inhibition of XO. The molecular modeling
study concludes that our vision of isosteric replacement
enhances the affinity for the XO enzyme due to the additional
hydrogen bond between the quinoline ring and residues
of XO.
Molecular modeling
The inhibition profile of synthesized 2-aryl-4-quinolones
was investigated at the molecular level with the help of
molecular docking. GOLD program was used to predict the
binding pose of the potent XO inhibitor 3l (Table 2), at the
salicylic acid binding site of XO. The scoring function of
the GOLD software was used to predict the strength of
interaction between inhibitor (3l) and receptor. In each
run, ten different conformations were generated out of
which the conformation with the highest Gold score was
selected for the study of ligand–receptor interaction. The
salicylic acid binding cavity easily accommodates 3l which
shares hydrophobic, hydrogen bonding, and arene–arene
interactions with the receptor. In the binding mode, the
quinolone ring was found to be sandwiched in the hydro- Conclusion
phobic pocket created by Leu873, Phe914, and Phe1009. 3l
forms two strong hydrogen bonds with binding residues,
the first one is found between the hydroxyl group of
Thr1010 with the carbonyl group of 3l and another with
the hydroxyl group of Glu802 with the nitrogen of 3l. The
pyridinyl ring fits in the hydrophobic cavity formed by
Bioisosterism has proved to be a useful lead optimization
technique to improve the pharmacological activity, gain
selectivity, and optimize the pharmacokinetics of the lead
compounds. In continuation of our search for non-purine
XO inhibitors, we designed 2-aryl-4-quinolones, also known
as aza analogs of flavones as a new class of XO inhibitors.
Aza analogs of flavones were designed after considerations
of the pharmacophoric features and structural requisites
for XO inhibition by one of the most privileged bioactive
classes of secondary metabolites known as flavones. Thus
the flavone framework was employed as the lead structure
for further optimization. The design strategy involved
isosteric replacement of chromone (a bicyclic unit present
as a core structural unit in flavones) with quinolones
after careful study of the enzyme’s structural topography
providing appropriate sites for interaction with the
amino acid residues of the enzyme. On this basis, a series
of 2-aryl/heteroaryl-4-quinolones was prepared and eval-
uated for in vitro XO inhibition. The results of the inhibitory
evaluation indicated the strong influence of the electronic
and steric factors linked with the aryl ring. The placement
of heteroaryl rings proved to be critical for potentiating
the inhibitory activity. SAR study revealed that: (i) the
nature of the aryl ring, (ii) the nature of the substituent(s)
on the aryl ring and (iii) the length of the linker greatly
Table 2. In vitro screening of the synthesized compounds (3a–3o)
for XO inhibitory activity.
COMPOUNDS
XO inhibitory activity (IC50, mM)
3a
3b
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
3m
3n
3o
18.0
12.7
12.5
19.5
18.2
15.4
12.0
25.2
26.3
31.2
11.23
6.24
30.8
100
18.3
10.21
8.31
Apigenin
Allopurinol
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Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
Aza-Flavones as Xanthine Oxidase Inhibitors
13
Figure 7. (a) Binding interactions of 3l at the salicylic acid binding site of XO. (b) Superimposition of apigenin and 3l at the salicylic acid
binding site of XO.
CDCl3 and DMSO-d6 relative to TMS (0.00 ppm). IR (KBr pellets)
spectra were recorded on a Fourier transform infrared (FT-IR)
Thermo spectrophotometer. Melting points were determined in
open capillaries and were uncorrected.
affect the XO inhibitory activity. The factor enhancing the
rigidity toward the planar structure remarkably enhanced
the inhibitory activity and this feature is in agreement
with the previously reported studies claiming the planarity
to be an important structural feature for flavones. The
Experimental procedure for the synthesis of 2-amino
acetophenone
results of the in vitro assay were further rationalized by
molecular modeling studies. Docking simulations were
performed to position the most active compound 3l into
the XO active site to determine the probable binding con-
formation, and the results confirmed that the compound
was a potential inhibitor of XO. The docking studies further
clarified the role of the heteroaryl ring responsible for the
enhancement of the inhibition. The docking simulations
also indicated that the bioisosteric replacement of chro-
mones with quinolones in the design of aza analogs of
flavones as non-purine based XO inhibitors preserves the
key interactions with the enzyme and also provide some
additional interaction sites. Rationalization of the in vitro
inhibition results by the computational studies justified
our design strategy. Further lead modification on 3l such
as synthesis of compounds with diverse permutation and
combinations on ring A and B is under progress and will be
published in due course.
To a solution of 2-aminobenzoic acid (6.0 mmol) in DME (42 mL),
methyl lithium (1.5 M in Et2O, 13.2 mL, 19.8 mmol) was slowly
added at 08C. After being stirred for 1 h, the resulting solution
containing white precipitates was quenched with saturated
NH4Cl (5 mL) and DME was evaporated under reduced pressure.
The mixture was poured into saturated NH4Cl (40 mL), extracted
with methylene chloride (3 ꢀ 25 mL), and washed with satu-
rated NaHCO3 (40 mL). The combined organic phases were dried
over MgSO4, filtered, and concentrated under reduced pressure.
The residues were purified by column chromatography to give
2-amino acetophenone.
Experimental procedure for the synthesis
of N-(2-acetylphenyl)acetamide
A mixture of 2-amino acetophenone (0.1 mol, 13.5 g), 15 mL of
acetic anhydride and a pinch of DMAP was stirred at 708C
in an oil bath for 6 h. The completion of reaction was monitored
by TLC. The reaction mixture on completion was poured onto
ice and the precipitates were collected and crystallized from
ethanol.
Experimental
Experimental procedure for the synthesis of diaryl
propenones (1a–1o)
The reagents were purchased from Sigma–Aldrich, Loba and
CDH, India, and used without further purification. All yields
refer to isolated products after purification. Products were
characterized by comparison with authentic samples and by
spectroscopic data (IR, 1H NMR). The spectra were measured in
An aqueous solution of NaOH (5%, 4 mL) was added slowly to the
stirring solution of appropriate aryl aldehyde (1 mmol) and N-(2-
acetylphenyl)acetamide (1 mmol) in methanol (20 mL) in a
100 mL conical flask. The stirring was continued at room
temperature for 6 h and the completion of reaction was moni-
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R. Dhiman et al.
Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
tored by TLC. The reaction on completion was poured onto ice
cold water; a yellow solid was obtained after filtration which was
recrystallized from ethanol. The physical data for the character-
istic compound is shown below.
J ¼ 8.7 Hz), 8.11 (1H, d, J ¼ 7.5 Hz), 7.78 (1H, m), 7.26–7.28
(3H, m), 7.17 (2H, m), 3.95 (3H, s), 3.80 (3H, s). Anal. calcd.
for C17H15NO3: C, 72.58; H, 5.37; N, 4.98. Found: C, 72.34;
H, 5.22; N, 5.11.
N-[2-(3-Phenylacryloyl)-phenyl]-acetamide (1a)
2-(2-Methoxyphenyl)-quinolin-4(1H)-one (3d)
Yield 77%; mp: 54–568C (lit. 54–568C) [27]; IR (KBr, cmꢁ1): 3321,
1660, 1652, 1590, 1240. 1H NMR (CDCl3, d, TMS ¼ 0): 8.71 (1H,
d, J ¼ 8.31 Hz), 7.98 (1H, d, J ¼ 7.55 Hz), 7.86 (1H, d, J ¼ 15.51 Hz),
7.65–7.38 (6H, m), 7.40 (1H, d, J ¼ 15.51 Hz), 7.17 (1H, dd, J ¼ 7.81
and 7.42 Hz), 2.26 (3H, s). Anal. calcd. for C17H15NO2: C, 76.96;
H, 5.70; N, 5.28. Found C, 77.21; H, 5.39; N, 5.34.
mp: 234–2368C (lit. 233–2358C) [29]. IR (KBr, cmꢁ1): 3341, 2969,
1663, 1596, 1311, 1229. 1H NMR (DMSO-d6, 400 MHz, d, TMS ¼ 0):
8.34–8.36 (1H, m), 8.03–8.07 (1H, m), 7.76–7.80 (1H, m), 7.67–7.71
(2H, m), 7.35 (1H, m), 7.29 (1H, s), 7.26 (1H, m), 3.90 (3H, s). Anal.
calcd. for C16H13NO2: C, 76.48; H, 5.21; N, 5.57. Found: C, 76.14;
H, 5.12; N, 5.77.
Experimental procedure for the synthesis
of 2,3-dihydro-2-aryl quinolones (2a–2o)
A mixture of 1 (1 mmol) and 5% HCl (10 mL) was refluxed for
12 h. The reaction mixture was kept at room temperature and
then basified with NH4OH. The precipitated solid was filtered and
recrystallized from ethanol. The physical data for the character-
istic compound is shown below.
2-(2,5-Dimethoxyphenyl)-quinolin-4(1H)-one (3e)
mp: 260–2628C. IR (KBr, cmꢁ1): 3329, 2981, 1659, 1588, 1303,
1224. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 8.09 (2H, m), 7.83
(1H, d, J ¼ 8.70 Hz), 7.18–7.33 (4H, m), 7.02 (1H, s), 3.85 (6H, s).
Anal. calcd. for C17H15NO3: C, 72.58; H, 5.37; N, 4.98. Found:
C, 72.74; H, 5.43; N, 4.77.
2-(3,4,5-Trimethoxyphenyl)-quinolin-4(1H)-one (3f)
mp: 231–2218C (lit. 232–2348C) [30]. IR (KBr, cmꢁ1): 3335, 2973,
1664, 1585, 1305, 1222. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0):
8.11 (1H, d, J ¼ 7.21), 7.75 (1H, d, J ¼ 8.40 Hz), 7.64 (1H, m), 7.31
(1H, m), 7.08 (2H, s), 6.41 (1H, s), 3.92 (9H, s). Anal. calcd.
for C18H17NO4: C, 69.44; H, 5.50; N, 4.50. Found: C, 69.64;
H, 5.53; N, 4.37.
2,3-Dihydro-2-phenylquinolin-4(1H)-one (2a)
Yield 56%; mp: 149–1508C (lit. 150–1518C) [28]; IR (KBr, cmꢁ1):
1
3321, 1663, 1593, 1237. H NMR (CDCl3, 300 MHz, d, TMS ¼ 0):
7.87 (1H, dd, J ¼ 8.10 and 1.51 Hz), 7.40–7.48 (4H, m), 7.31–7.35
(3H, m), 6.80 (1H, d, J ¼ 8.11 Hz), 4.76 (1H, dd, J ¼ 3.91 and
13.51 Hz), 4.49 (1H, s, NH, D2O exchangeable proton), 2.90
(2H, dd, J ¼ 13.50 and 16.21 Hz), 2.78 (2H, dd, J ¼ 4.20 and
16.21 Hz). Anal. calcd. for C15H13NO: C, 80.69; H, 5.87; N, 6.27.
Found C, 80.83; H, 5.64; N, 6.01.
2-(4-Methoxyphenyl)-quinolin-4(1H)-one (3g)
mp: 287–2888C (lit. 288–2908C) [28]. IR (KBr, cmꢁ1): 3329, 2978,
1662, 1587, 1306, 1225. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0):
11.37 (1H, s), 8.27 (1H, d, J ¼ 7.21 Hz), 7.72 (3H, m), 7.59 (1H,
t, J ¼ 8.10 Hz), 7.31(1H, t, J ¼ 7.51 Hz), 7.04 (1H, d, J ¼ 8.70 Hz),
6.45 (1H, s). Anal. calcd. for C16H13NO2: C, 76.48; H, 5.21; N, 5.57.
Found: C, 76.84; H, 5.32; N, 5.26.
Experimental procedure for the synthesis
of 2-aryl-4-quinolones (3a–3o)
A mixture of 2 (1 mmol) and iodine (1.5 mmol) in DMSO was
warmed at 808C in an oil bath for 12 h. On completion of
the reaction, the reaction mixture was poured onto a saturated
solution of sodium thiosulfate. The precipitated solid was
collected and the desired product was purified by column
chromatography using silica gel (60 ꢀ 120 mesh) with increas-
ing percentage of ethyl acetate in hexane as eluting solvent. The
physical data of the compounds are provided below.
2-(4-Chlorophenyl)-quinolin-4(1H)-one (3h)
mp: 339–3408C (lit. 340–3428C) [28]. IR (KBr, cmꢁ1): 3343, 2971,
1665, 1586, 1222, 751. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0):
11.77 (1H, s), 8.11 (2H, dd, J ¼ 8.11 and 1.20 Hz), 7.88 (2H,
d, J ¼ 2.80 Hz), 7.76 (1H, J ¼ 8.12 Hz), 7.68–7.71 (1H, m), 7.66
(2H, d, J ¼ 8.61 Hz), 7.33–7.38 (1H, m), 6.38 (1H, s). Anal. calcd.
for C15H10ClNO: C, 70.46; H, 3.94; Cl, 13.87; N, 5.48. Found: C,
70.24; H, 3.62; Cl, 13.99; N, 5.16.
2-Phenylquinolin-4(1H)-one (3a)
mp: 250–2528C (lit. 252–2548C) [28]. IR (KBr, cmꢁ1): 3321, 2978,
1666, 1591, 1233. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 11.45
(1H, s), 8.29 (1H, d, J ¼ 8.41 Hz), 7.62–7.77 (5H, m), 7.60–7.62 (2H,
m), 7.32 (1H, t, J ¼ 8.11 Hz), 6.48 (1H, s). Anal. calcd. for C15H11NO:
C, 81.43; H, 5.01; N, 6.33. Found: C, 81.66; H, 4.99; N, 6.64.
2-(4-Fluorophenyl)-quinolin-4(1H)-one (3i)
mp: 250–2528C. IR (KBr, cmꢁ1): 3339, 2966, 1662, 1583, 1228,
911. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 8.09 (1H,
d, J ¼ 7.60 Hz), 7.91 (2H, m), 7.75 (1H, d, J ¼ 8.42 Hz), 7.67
(1H, t, J ¼ 7.62 Hz), 7.43 (2H, m), 7.33 (1H, t, J ¼ 7.21 Hz), 6.34
(1H, s). Anal. calcd. for C15H10FNO: C, 75.30; H, 4.21; F, 7.94;
N, 5.85. Found: C, 75.49; H, 4.52; Cl, 7.79; N, 5.46.
2-(3,4-Dimethoxyphenyl)-quinolin-4(1H)-one (3b)
mp: 252–2548C. IR (KBr, cmꢁ1): 3328, 2972, 1665, 1594, 1313, 1237.
1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 8.11 (1H, d, J ¼ 7.21 Hz),
7.75 (1H, d, J ¼ 8.40 Hz), 7.64 (1H, m), 7.31 (1H, m), 7.01–7.06 (3H,
m), 6.41 (1H, s), 3.92 (6H, s). Anal. calcd. for C17H15NO3: C, 72.58;
H, 5.37; N, 4.98. Found: C, 72.66; H, 5.48; N, 4.78.
2-(Naphthalen-1-yl)-quinolin-4(1H)-one (3j)
mp: 280–2818C (lit. 284–2858C) [30]. IR (KBr, cmꢁ1): 3344, 2972,
1660, 1585, 1230. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 12.01
(1H, s), 8.20 (2H, m), 8.03–8.11 (3H, m), 7.89 (1H, d, J ¼ 6.90 Hz),
7.55–7.70 (4H, m), 7.33 (1H, m), 6.20 (1H, s). Anal. calcd.
2-(2,3-Dimethoxyphenyl)quinolin-4(1H)-one (3c)
mp: 233–2358C. IR (KBr, cmꢁ1): 3333, 2975, 1669, 1597, 1309,
1232. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 8.20 (1H, d,
ß 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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Arch. Pharm. Chem. Life Sci. 2013, 346, 7–16
Aza-Flavones as Xanthine Oxidase Inhibitors
15
for C19H13NO: C, 84.11; H, 4.83; N, 5.16. Found: C, 84.44; H, 4.42;
N, 5.36.
Molecular modeling
A flexible docking study of the most active compound at the
salicylic acid binding site of XO was performed with the help of
GOLD software [32]. To validate the docking procedure for the
prediction of the correct binding mode of the inhibitor at the
salicylic acid binding site, the salicylic acid was extracted from
the original X-ray structure (1FIQ) [33] and re-docked using GOLD
[32]. The highest scoring conformation was selected and com-
pared with the X-ray structure conformation. The docked con-
formation of salicylic acid was found to be similar to the original
X-ray structure conformation. The root mean square deviation
(RMSD) between the best scored conformers from docking and
2-(Thiophen-2-yl)quinolin-4(1H)-one (3k)
mp: 300–3028C (lit. 3008C) [29]. IR (KBr, cmꢁ1): 3343, 2985, 1659,
1577, 1313, 1225. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 8.10
(1H, d, J ¼ 8.80 Hz), 8.05 (1H, s), 7.70 (1H, d, J ¼ 6.40 Hz), 7.54
(1H, t, J ¼ 8.00 Hz), 7.32 (1H, m), 6.87 (1H, s), 6.55 (1H, s), 6.37 (1H,
s). Anal. calcd. for C13H9NO2: C, 73.92; H, 4.29; N, 6.63. Found:
C, 73.74; H, 4.52; N, 6.96.
2-(Pyridin-4-yl)-quinolin-4(1H)-one (3l)
˚
X-ray structure was found to be 0.21 A.
mp: 234–2358C. IR (KBr, cmꢁ1): 3335, 2977, 1658, 1583, 1233.
1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0): 12.19 (1H, s), 9.05 (1H,
s), 8.69 (1H, m), 8.34 (1H, d, J ¼ 7.2 Hz), 8.24 (1H, m), 7.70 (2H, m),
7.62 (1H, m), 7.45 (1H, m). Anal. calcd. for C14H10N2O: C, 75.66;
H, 4.54; N, 12.60. Found: C, 75.44; H, 4.82; N, 12.46.
Special thanks to Dr. K. A. Suri, Ex-Director Grade Scientist, IIIM (CSIR),
Jammu, for providing pure sample of apigenin.
The authors have declared no conflict of interest.
2-(3,4-Methylenedioxyphenyl)-1,4-dihydro-4-
oxoquinoline (3m)
References
mp: 234–2358C (lit. 284–2858C) [30]. IR (KBr, cmꢁ1): 3464, 1636,
2981, 1580, 1315, 1221. 1H NMR (DMSO-d6, 300 MHz, d, TMS ¼ 0):
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