4,6-Diaryl/heteroarylpyrimidin-2(1H)-ones as a new class of xanthine oxidase inhibitors

Article abstract of DOI:10.1002/ardp.201400031

Summary A series of 4,6-diaryl/heteroarylpyrimidones was synthesized employing silica-supported fluoroboric acid under solvent-free conditions in a microwave reactor. The catalytic influence of HBF₄-SiO₂ was investigated in detail to

Full text of DOI:10.1002/ardp.201400031

Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
1
Full Paper
4,6-Diaryl/heteroarylpyrimidin-2(1H)-ones as a New Class of
Xanthine Oxidase Inhibitors
Shiwani Shukla1, Dinesh Kumar1, Ritu Ojha1, Manish K. Gupta2, Kunal Nepali1, and Preet M. S. Bedi1
1
Department of Pharmaceutical Sciences, Guru Nanak Dev University, Amritsar, Punjab, India
Department of Pharmaceutical Chemistry, ISF College of Pharmacy, Moga, Punjab, India
2
A series of 4,6-diaryl/heteroarylpyrimidones was synthesized employing silica-supported fluoroboric
acid under solvent-free conditions in a microwave reactor. The catalytic influence of HBF4-SiO2 was
investigated in detail to optimize the reaction conditions. The synthesized compounds were evaluated
for in vitro xanthine oxidase (XO) inhibitory activity for the first time. Structure-activity relationship
analyses are also presented. Among the synthesized compounds, VA-5, -9, -10, -12, -22, -23, and -25 were
the active inhibitors with IC50 values ranging from 6.45 to 13.46 mM. Compound VA-25 with a pyridinyl
ring as ring A and a thiophenyl ring as ring B emerged as the most potent XO inhibitor (IC50 ¼ 6.45 mM)
in comparison to allopurinol (IC50 ¼ 12.24 mM). Some of the important interactions of VA-25 with the
amino acid residues of the active site of XO were figured out by molecular modeling studies.
Keywords: Catalyst / Fluoroboric acid / Inhibitors / Microwave radiation / Pyrimidones / Silica / Xanthine oxidase
Received: January 27, 2014; Revised: February 27, 2014; Accepted: March 12, 2014
DOI 10.1002/ardp.201400031
Introduction
renal function induced in some of the patients [2–4] has led to
the continuous search for novel scaffolds as XO inhibitors.
We have been actively involved in the design, synthesis, and
evaluation of XO inhibitors during the recent past [4, 8–10].
Recently our research group explored the XO inhibitory
potential of a series of N-acetyl pyrazolines [8] and N-(1,3-diaryl-
3-oxopropyl)amides [9] (Fig. 1). The structure–activity rela-
tionship established on the basis of in vitro evaluation and the
molecular modeling studies of both the classes revealed
some important structural features required for inhibiting
the enzyme such as (i) aryl/heteroaryl rings linked via a 3-carbon
open chain or a cyclic linker with heteroatoms on 1- and 3-
carbon, (ii) sites for hydrophobic interaction, arene–arene
interaction, electrostatic interactions (Fig. 1) and (iii)
appropriately placed carbonyl functionality for hydrogen
bonding interactions (Fig. 1). The results also indicated that
the dependence of inhibitory potential on the electronic
and steric effects varies with the nature of the linker (a cyclic
linker as in case of N-acetyl pyrazolines or a open chain
linker as in case of N-(1,3-diaryl-3-oxopropyl)amides. With
this background, the present study explores the potential
of a series of 4,6-diarylpyrimidin-2(1H)-ones (target com-
pounds, Fig. 1) as a new class of XO inhibitors. The target
compounds employ pyrimidone ring as the linker for the
two aryl rings that allows similar arrangement of the two aryl
Xanthine oxidase (XO), a versatile molybdoflavoprotein that
catalyzes oxidative hydroxylation of purine substrates to
produce uric acid and subsequent reduction of oxygen at the
flavin center with the generation of reactive oxygen species
(ROS), either as superoxide anion radical or hydrogen [1–3]
peroxide, leads to many diseases such as gout and at least
symptoms of diseases such as oxidative damage to the
tissue [2]. Involvement of reactive oxygen species in
pathological events including inflammation, metabolic dis-
orders, cellular ageing, atherosclerosis, and carcinogenesis is
well reported. Increased XO serum level in pathological states
such as hepatitis, inflammation, cancer, etc. indicates that XO
inhibitor may result in broad-spectrum therapeutics for gout,
cancer, inflammation, and oxidative damage [4]. Despite the
potential of allopurinol [3, 5], 2-alkyl hypoxanthines [6],
pterin, and 6-formylpterin [7] as XO inhibitors, their use
associated with Steven Johnson syndrome and worsening of
Correspondence: Dr. Kunal Nepali, Department of Pharmaceutical
Sciences, Guru Nanak Dev Univeristy, Amritsar, Punjab-143005, India
E-mail: kunal_8855@rediffmail.com, kunalbvk3@gmail.com
Fax: þ91-1832258819
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
O
O
O
HN
N
N
Aryl/
hetero
aryl
Aryl/
hetero
aryl
3-Carbon
linker
N
O
ring
ring
Br
3-Carbon linker
3-Carbon linker
N-Acetyl pyrazoline
Scaffold for xanthine oxidase inhibitors
N-(1,3-Diaryl-3-oxopropyl)amide
Hydrogen bonding interaction
with Ser876
O
O
N
N
NH
N
Positioned in the cavity
formed by Leu648,
Leu873 and involved in
weak electrostatic
interactions
A
B
Arene-arene interaction
with Phe914
N
O
Target compounds
Hydrophobic interactions
withThr803
Hydrogen bonding
interaction with
Asn768
Hydrogen bonding
interaction with
Glu802
O
NH
O
Arene–arene interaction
with Phe914 and Phe1009
Arene–cation interactions
with protonated amino
group of Lys771
Br
Hydrophobic interactions with
isopropyl group of Leu101
Figure 1. Structural features and interactions for XO inhibition.
rings as in case of the earlier reported inhibitors by us [8, 9]
(Fig. 1).
reaction has been well explored for the synthesis of 3,4-
dihydropyrimidinones [11], three-component cyclocondensa-
tion of an aldehyde, acetophenone, and urea resulting in the
formation of 4,6-diarylpyrimidin-2(1H)-ones (Fig. 1) still
requires an efficient protocol. The existing methodologies
for the synthesis of 4,6-diarylpyrimidin-2(1H)-ones lack in
one or more of the following – two-step synthesis [12a],
As a part of our continued search for efficient methodolo-
gies for XO inhibitors [10], a series of 4,6-diaryl/heteroaryl
pyrimidones as a new class of XO inhibitors was designed and
synthesized via silica-supported fluoroboric acid under neat
conditions in a microwave reactor. Though the Biginelli
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
4,6-Diaryl/heteroarylpyrimidin-2(1H)-ones
3
applicabilty to limited substrates (not investigated for
heteroaryl ketones/aldehydes and bicyclic ketones/aldehydes)
[12b, 12c], and use of hazardous solvents [12d]. This tempted
us to investigate the reaction under microwave conditions
employing a silicated fluoroboric acid and a variety of
substrates such as (i) aryl/heteroaryl ketones, (ii) aryl/
heteroaryl aldehydes, (iii) bicyclic ketones, (iv) bicyclic
aldehydes and (v) aldehydes and ketones with electronically
diverse substituents. The use of catalysts immobilized on solid
supports has received considerable attention [13] as adsorp-
tion of reagent onto an insoluble inorganic or organic support
improves activity and selectivity of reagent by increasing
effective surface area of reagent dispersed on a support up to
100 times and also helps in preventing the release of reaction
residues into the environment for achieving environment-
friendly organic synthesis [14]. Moreover multicomponent
synthesis by Lewis acids/Bronsted acids have gained enough
attention in the recent past [15]. Fluoroboric acid was
employed in the present study as it is a weak protic acid
that when adsorbed on silica would circumvent problem of
side reaction for acid-sensitive substrate [13c].
irradiation from 10 to 15 min and further to 20 min. The
optimum reaction conditions involve the use of 5 mol% of
the catalyst for 10 min in
a microwave reactor. The
percentage yield of the pyrimidones was generally high
ranging from 62 to 94%. In general pyrimidones with
halogen-substituted phenyl rings (at both 4 and 6 positions)
were obtained in excellent percentage yield (>90%). The
excellent yields with halogens-substituted phenyl rings could
be attributed to the dominant ꢀI effect of halogens in
comparison to þR effect. Pyrimidones with phenyl ring
possessing OCH3 groups were also obtained in good yield (80–
90%). However, the yields were slightly less in comparison to
pyrimidones with halogen-substituted phenyl groups. This
might be due to dominant þR effect of methoxy groups in
comparison to –I effect. Pyrimidones with heteroaryl rings
and bicyclic rings were obtained in moderate yields (62–71%),
indicating the versatility of the supported catalyst towards
electronically and sterically diverse substitutents. Applicabil-
ity to a variety of substrates (bicyclic/aromatic/heteroaro-
matic ketones and aldehydes), short reaction time, high
yields, solvent-free synthesis, and utilization of silica-
supported cataylst, which makes the overall methodology
cost-effective, are the highlights of the methodology devel-
oped. The structures of the synthesized compounds were
elucidated by spectral data. All spectral data were in
accordance with assumed structures.
Results
Chemistry
In an attempt to investigate the catalytic efficiency of
fluoroboric acid adsorbed on silica, a model reaction
(Scheme 1) was performed exposing a mixture of aromatic
aldehyde (5.0 mmol, 1 eq), aromatic ketone (5.0 mmol, 1 eq),
urea (5.0 mmol, 1 eq), and varying mol% of silicated
fluoroboric acids to microwave radiations (varying time) in
a microwave synthesizer operating at 150°C with the
maximum microwave power of 400 W for the synthesis of
target compound (Table 1).
In order to examine the substrate scope of this reaction,
various aromatic ketones/aldehydes were used (Tables 1
and 2), and from the results, we could see that all reactions
with aromatic aldehydes/ketones possesing electronically
diverse substituents as well as heteroaryl aldehydes/ketones
proceeded smoothly to afford the corresponding pyrimidones
in good to excellents yields. Careful observation of Table 1
indicates a decrease in percentage yield of the target
compounds on increasing the time of exposure of microwave
In vitro XO assay
All the synthetic compounds were evaluated for in vitro XO
inhibitory activity. In vitro screening of the pyrimidones using
bovine milk XO (grade 1, ammonium sulfate suspension)
enzymatic assay was performed as described in the litera-
ture [16]. Allopurinol [17] was employed as reference
inhibitors. The molecules exhibiting % age inhibition of
more than 50% at 50 mM were further tested in triplicate for
the XO inhibitory activity. The pyrimidones VA-2, VA-5, VA-7,
VA-8, VA-9, VA-10, VA-12, VA-14, VA-15, VA-21, VA-22, VA-23,
and VA-25 displayed percentage inhibition of more than 50%
at 50 mM (Table 3) and were further considered for the
calculation of IC50 values. The pyrimidones with heteroaryl
ring (VA-12, VA-21, VA-22, VA-25) at position 4/6 or both
displayed significant % inhibition of >80%. Among them, VA-
12 and VA-25 possesing heteroaryl rings at both 4 and 6
O
O
O
N
NH
O
HBF4-SiO2
H
B
A
H2N
NH2
A
B
MW, solvent-free
Scheme 1. Microwave-assisted synthesis of 4,6-diaryl/heteroarylpyrimidones.
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
Table 1. Percentage yields of the 4,6-diaryl/heteroarylpyrimi-
dones with varying mol% of catalyst and time of exposure to
microwave irradiation.
Table 2. Synthesized 4,6-substituted pyrimidones.
Code
Product
Yield (%)
O
O
N
NH
O
N
NH
VA-1
82
O
R'
O
R
O
O
Loading
(mol%)
Time
Entry
R
R0
(MW)a) Yield
N
NH
NH
VA-2
VA-3
92
80
1.
H
H
1
10
15
20
10
15
20
10
15
20
72
67
59
82
73
68
78
70
64
Cl
5
O
N
10
O
O
O
2.
3.
4.
4-Cl
4-CH3
H
H
1
5
10
15
20
10
15
20
10
15
20
79
72
68
92
84
79
87
79
73
O
N
O
NH
VA-4
VA-5
69
82
10
N
NH
O
H
1
5
10
15
20
10
15
20
10
15
20
76
69
63
84
76
68
77
71
63
10
N
NH
VA-6
VA7
84
93
4-OCH3
1
5
10
15
20
10
15
20
10
15
20
73
67
62
85
75
69
79
71
63
H3C
O
N
NH
10
F
O
O
5.
1
5
10
15
20
10
15
20
10
15
20
56
52
47
68
62
54
61
54
50
N
NH
VA8
91
94
N
NH
O
N
Br
10
O
N
NH
VA-9
a)
Exposing the reaction mixture to microwave radiations for
5 min produced unsatisfactory yileds, i.e. <40%.
Cl
Br
(Continued)
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
Table 2. (Continued)
4,6-Diaryl/heteroarylpyrimidin-2(1H)-ones
5
Table 2. (Continued)
Code
Product
Yield (%)
Code
Product
Yield (%)
O
O
N
NH
N
NH
VA-10
93
VA-18
86
O
O
O
Cl
Cl
O
O
N
NH
NH
N
NH
VA-11
68
VA-19
87
94
O
HO
Br
O
O
O
O
N
NH
VA-20
VA-21
N
N
VA-12
VA-13
68
67
O
Cl
Cl
N
O
N
NH
NH
NH
NH
71
62
O
O
S
O
O
O
O
O
N
NH
N
N
VA-22
O
O
VA-14
VA-15
84
83
N
Br
O
N
NH
O
O
O
O
VA-23
81
Cl
O
N
NO2
O
O
N
NH
N
NH
O
VA-16
VA-17
85
VA-24
VA-25
80
69
O
OCH3
O
N
NH
NH
82
S
O
O
N
(Continued)
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
Table 3. In vitro screening of the synthesized compounds for XO
inhibitory activity.
and Ring B (VA-4, 11, 13) also displayed a decreased inhibitory
profile. However, placement of heteroaryl ring (Ring A) and
naphthyl ring (Ring B) in the chemical architecture of the
designed pyrimidones regenerated the inhibitory potential
(VA-22). 4) Placement of heteroaryl ring either on Ring A or
Ring B proved to be extremely beneficial for the activity (VA-
21, 22). 5) Placement of heteroaryl rings at both 4th and 6th
positions (Ring A and Ring B) proved to be even more potent
than compounds with monoheteroaryl ring. (Compare VA-12,
VA-25 with VA-21, 22). On the basis of the results of in vitro XO
inhibitors, a basic pharmacophore (Fig. 2) was formulated
with the structural features required for XO inhibitory
activity.
Percentage
inhibition (%)
IC50
Compounds
(mM)a)
VA-1
VA-2
VA-3
40
60
39
–
20.25
–
VA-4
VA-5
VA-6
35.56
90.32
48
–
10.21
VA-7
VA-8
VA-9
60.70
79.50
80.03
76.26
30.90
95
40.70
69.13
68.48
45.11
36.57
25.90
28.14
81.85
85.55
68.90
40.56
97
21.24
15.67
13.35
12
–
7.23
–
18.87
18.57
–
–
–
VA-10
VA-11
VA-12
VA-13
VA-14
VA-15
VA-16
VA-17
VA-18
VA-19
VA-21
VA-22
VA-23
VA-24
VA-25
Allopurinol
Molecular modeling studies
In order to theoretically investigate the recognition process of
the most potent XO inhibitor VA-25, flexible docking
experiment using GOLD software was performed assuming
that it gets accommodated into the salicylic acid XO active
site [18]. The docking study revealed some non-covalent
interactions of the VA-25 at salicylic acid binding site of XO.
The inhibitor VA-25 completely occupies the salicylic acid
binding cavity of XO. In the binding conformation, VA-25
indicates some interaction, which may be crucial for the
inhibition of XO. The thiophene ring in VA-25 is found to be
involved in hydrogen bonding with side chain of Glu802. The
p-electrons of thiophene ring also get engaged in face-to-face
stacking with p-electrons of Phe914. This provides stability to
the thiophene ring in the binding cavity. A strong hydrogen
bonding was observed between the carbonyl group of VA-25
and hydroxyl group of the Ser876, which additionally help in
stabilization of VA-25 in the binding cavity. The nitrogen of
pyridine ring interacts with NH2 group of Asn768. Thus, the
docking result provides the rationale behind the XO
inhibition (Fig. 3).
–
13.46
11.23
18.93
–
6.45
12.24
a)
Values are mean of three experiments.
position were the most potent inhibitors with % inhibition of
95 and 97% (Table 3). This could be due to the hydrogen
bonding ability of both the heteroatoms. Both VA-12 and -25
displayed low IC50 values of 7.23 and 6.45 mM as compared to
allopurinol (IC50 ¼ 12.24 mM). VA-25 with a pyridinyl ring as
Ring A and thiophenyl as Ring B was the most potent inhibitor
of the series. The slight difference in the inhibitory profile of
VA-12 and VA-25 (both possesing a pyridinyl ring as Ring A)
could be attributed to the higher aromatic character of 2-
thiophenyl ring (Ring B) in VA-25 as compared to 2-furanyl
ring (Ring B) in VA-12. Rings with higher aromatic character
generally display stronger interactions with the amino acids
posessing phenyl rings. This was further investigated by
docking of VA-25 with in the active site of the enzyme.
Calculation of molecular properties
The drug-like properties of the active compounds were
calculated, which were found to be in accordance to Lipinski’s
rule of five (Table 4) [19]. The molecular properties of the most
potent pyrimidones were found to be within the specified
limits, which indicate their drug likeness.
Structure–activity relationship
From the data shown in Table 3, some generalizations about
the structure–activity relationship were made such as: 1) Any
substitutions on the phenyl rings (Ring A or Ring B or both)
resulted in a decline in the inhibitory potential (compare V-5
with other pyrimidones except VA-12 and VA-25). 2) The
decline in activity for the halo-substituted phenyl rings (at
both 4th and 6th positions) was less as compared to other
substituents. 3) Substitution of naphthyl ring either as Ring A
O
Heteroaryl >
Heteroaryl >
phenyl > substituted
phenyl > substituted
N
NH
phenyls/naphthyl
phenyls/naphthyl
A
B
Figure 2. Pharmacophore.
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
4,6-Diaryl/heteroarylpyrimidin-2(1H)-ones
7
purification. All yields refer to isolated products after purifica-
tion. All the reactions were carried out in Biotage Microwave
Synthesizer (Model: Initiator) operating at 150°C with the micro-
wave power maximum level of 400 W. Products were characterized
by comparison by spectroscopic data. 1H NMR spectra were
recorded on Bruker Advance II 400 NMR spectrometer and JEOL AL
300 NMR spectrometer. The spectra were measured in DMSO
relative to TMS (0.00 ppm). Melting points were determined in
open capillaries and were uncorrected.
Preparation of tetrafluoroboric acid adsorbed on silica
gel (HBF4-SiO2)
The catalyst system HBF4-SiO2 was prepared following the originally
reported method [20]. A mixture of silica gel (26.7 g, 300–400 mesh)
and 40% aq. HBF4 (3.3 g, 8.25 mL, 15 mmol) in diethyl ether (75 mL)
was stirred for 3h. The mixture was concentrated, and the residue
dried under vacuum at 100°C for 72 h to afford HBF4-SiO2 (0.5 mmol
HBF4/g) as a free-flowing powder.
Procedure for the synthesis of 4,6-diaryl/heteroaryl-
pyrimidin-2(1H)-one
Figure 3. Binding mode of VA-25 at salicylic binding site of XO.
To a 50mL dried conical flask differently substituted benzaldehydes
(1 mmol, 1 eq), acetophenones (5 mmol, 1 eq), urea (5 mmol, 1 eq),
and 5 mol% of silicated flouroboric acid were added. The resulting
mixture was placed into the microwave reactor for 10min. On
completion of the reaction, the crude mixture was dissolved in
methanol and adsorbed on silica (60–120 #). The desired product
was purified by column chromatography with increasing percent-
age of ethyl acetate in hexane as eluting solvent.
Conclusion
The present study provides an efficient methodology for the
synthesis of 4,6-diarylpyrimidin-2(1H)-ones as a new class
of XO inhibitors. HBF4-SiO2 at 5 mol% for 10 min under
microwave conditions efficiently catalyzes all the reactions
employing electronically and sterically diverse aldehydes/
ketones. The synthesized compounds were evaluated for
in vitro XO inhibitory potential. SAR study revealed that the
nature of aryl ring and nature of substituent(s) on the aryl
rings at positions 4 and 6 greatly affect the XO inhibitory
activity. Pyrimidones with heteroaryl rings (both at 4th and
6th positions) as Ring A and Ring B were the most potent
inhibitors. Docking simulations were performed to position
most active compound VA-25 into the XO active site to
determine the probable binding mode, and the results
confirmed that the compound was a potential inhibitor of
XO. Further detailed investigation will be carried out on VA-
25, which appears to be a good hit among the series.
Characterization of synthesized compounds
4-(3,4-Dimethoxy-phenyl)-6-phenyl-1H-pyrimidin-2-one
(VA-1)
Yellow powder; yield 78%; mp: 215–217°C; 1H NMR (CDCl3 þ
DMSO-d6, 300 MHz, d, TMS ¼ 0): 8.04 (2H, d, J ¼ 8.1 Hz), 7.54–7.85
(5H, m), 7.02 (1H, d, J ¼ 7.5 Hz), 6.84 (1H, d, J ¼ 7.2 Hz), 3.97 (3H, s,
OCH3), 3.93 (3H, s, OCH3). Anal. calcd. for C18H16N2O3: C, 70.12; H,
5.23; N, 9.09. Found: C, 70.41; H, 5.04; N, 9.13.
4-(4-Chloro-phenyl)-6-phenyl-1H-pyrimidin-2-one (VA-2)
White powder; yield 78%; mp: 230–232°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.23 (2H, d, J ¼ 8 Hz), 8.16 (2H, d, J ¼ 8 Hz),
7.48–7.65 (6H, m). Anal. calcd. for C16H11ClN2O: C, 67.97; H, 3.92;
N, 9.91. Found: C, 67.75; H, 4.03; N, 9.81.
Experimental
6-Phenyl-4-(3,4,5-trimethoxy-phenyl)-1H-pyrimidin-2-one
(VA-3)
Brown powder; yield 80%; mp: 249–251°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.17 (2H, bs), 7.49–7.80 (4H, m), 7.45 (2H, s),
The reagents were purchased from Sigma–Aldrich, Merck, CDH,
Loba Chem., Spectro Chem., India and used without further
Table 4. Molecular properties of some potent compounds.
Entry
Compound
TPSA
Mol. wt.
ClogP
nOHNH
nON
nrotb
1
2
3
4
VA-12
VA-22
VA-23
VA-25
71.78
45.75
58.68
58.64
239
254
299
255
1.29
3.00
3.14
1.71
1
1
1
1
5
3
4
4
2
2
2
2
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
3.92 (6H, s, OCH3), 3.78 (3H, s, OCH3). Anal. calcd. for C19H18N2O4:
C, 67.44; H, 5.36; N, 8.28. Found: C, 67.55; H, 5.12; N, 8.49.
m), 7.06–7.18 (3H, m), 3.86 (6H, s, OCH3). Anal. calcd. for
C22H18N2O3: C, 73.73; H, 5.06; N, 7.82. Found: C, 74.00; H, 4.77;
N, 8.18.
4-Naphthalen-2-yl-6-phenyl-1H-pyrimidin-2-one (VA-4)
Brown powder; yield 81%; mp: 267–269°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 11.25 (1H, s, NH, D2O exchangeable proton),
8.10–8.20 (4H, m), 7.77–7.78 (1H, d, J ¼ 8 Hz), 7.58–7.78 (8H, m).
Anal. calcd. for C20H14N2O: C, 80.52; H, 4.73; N, 9.39. Found: C,
80.33; H, 4.92; N, 9.21.
6-Furan-2-yl-4-pyridin-4-yl-1H-pyrimidin-2-one (VA-12)
Light brown powder, yield 89%; mp: 272–274°C; 1H NMR (CDCl3
þDMSO-d6, 300 MHz, d, TMS ¼ 0): 8.45–8.76 (3H, m), 7.66 (2H, d,
J ¼ 4.8 Hz), 7.21–7.40 (3H, m). Anal. calcd. for C13H9N3O2: C, 65.27;
H, 3.79; N, 17.56. Found: C, 65.38; H, 3.54; N, 17.69.
4,6-Diphenyl-1H-pyrimidin-2-one (VA-5)
6-Naphthalen-2-yl-4-(3,4,5-trimethoxy-phenyl)-1H-
Yellow powder; yield 89%; mp: 270–272°C; 1H NMR (DMSO-d6,
300 MHz, d, TMS ¼ 0): 11.13 (1H, s, NH, D2O exchangeable
protons), 8.15–8.16 (4H, m), 7.54–7.56 (7H, m). Anal. calcd. for
C16H12N2O: C, 77.40; H, 4.87; N, 11.28. Found: C, 77.35; H, 4.98; N,
11.03.
pyrimidin-2-one (VA-13)
Yellow powder; yield 81%; mp: 267–269°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.82(1H, bs), 8.29 (1H, bs), 8.02–8.12 (3H, m),
7.30–7.61 (5H, m), 3.94 (6H, s, OCH3), 3.77 (3H, s, OCH3). Anal.
calcd. for C23H20N2O4: C, 71.12; H, 5.19; N, 7.21. Found: C, 70.95;
H, 4.91; N, 7.38.
6-Phenyl-4-p-tolyl-1H-pyrimidin-2-one (VA-6)
White powder; yield 79%; mp: 255–257°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 11.32 (1H, s, NH, D2O exchangeable
protons), 8.16 (2H, d, J ¼ 8 HZ), 8.08 (2H, d, J ¼ 8 Hz), 7.53–7.61
(4H, m), 7.37 (2H, d, J ¼ 8 Hz), 2.49 (3H, s, CH3). Anal. calcd. for
C17H14N2O: C, 77.84; H, 5.38; N, 10.68. Found: C, 78.01; H, 5.41; N,
10.55.
6-(4-Bromo-phenyl)-4-(3,4,5-trimethoxy-phenyl)-1H-
pyrimidin-2-one (VA-14)
White powder; yield 79%; mp: 278–280°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.16 (2H, d, J ¼ 8 Hz), 7.77–7.79 (3H, m), 7.44
(2H, d, J ¼ 8 Hz), 3.92 (6H, s, OCH3), 3.75 (3H, s, OCH3). Anal. calcd.
for C19H17BrN2O4: C, 54.69; H, 4.11; N, 6.71. Found: C, 54.97; H,
3.88; N, 6.94.
4-(4-Fluoro-phenyl)-6-phenyl-1H-pyrimidin-2-one (VA-7)
Light yellow powder; yield-90%; mp: 265–267°C; 1H NMR (DMSO-
d6, 400 MHz, d, TMS ¼ 0): 11.20 (1H, s, NH, D2O exchangeable
protons), 8.29 (2H, d, J ¼ 8 Hz), 8.16 (2H, d, J ¼ 8 Hz), 7.56–7.71 (3H,
m), 7.39–7.47 (3H, m). Anal. calcd. for C16H11FN2O: C, 71.17; H,
4.16; N, 10.52. Found: C, 71.99; H, 3.89; N, 10.64.
6-(4-Chloro-phenyl)-4-(3,4,5-trimethoxy-phenyl)-1H-
pyrimidin-2-one (VA-15)
Yellow powder; yield 85%; mp: 266–267°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.25 (2H, d, J ¼ 8 Hz), 7.64–7.71 (3H, m), 7.45
(2H, d, J ¼ 8 Hz), 3.92 (6H, s, OCH3), 3.75 (3H, s, OCH3). Anal. calcd.
for C19H17ClN2O4: C, 61.21; H, 4.60; N, 7.51. Found: C, 61.38; H,
4.76; N, 7.32.
6-(4-Bromo-phenyl)-4-phenyl-1H-pyrimidin-2-one (VA-8)
White powder; yield 70%; mp: 279–280°C; 1H NMR (DMSO-d6,
300 MHz, d, TMS ¼ 0): 8.04 (4H, m), 7.66 (2H, d, J ¼ 7.2 Hz), 7.52–
7.55 (4H, m). Anal. calcd. for C16H11BrN2O: C, 58.74; H, 3.39; N,
8.56. Found: C, 58.82; H, 3.49; N, 8.42.
6-(4-Methoxy-phenyl)-4-phenyl-1H-pyrimidin-2-one (VA-
16)
White powder; yield 80%; mp: 237–239°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 11.20 (1H, s, NH, D2O exchangeable
protons), 8.09–8.15 (4H, m), 7.53–7.61 (3H, m), 7.44 (1H, s),
7.09–7.10 (2H, d, J ¼ 8.00 Hz), 3.84 (3H, s, OCH3). Anal. calcd. for
6-(4-Bromo-phenyl)-4-(4-chloro-phenyl)-1H-pyrimidin-2-
one (VA-9)
Yellow powder; yield 85%; mp: 278–280°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.22 (2H, d, J ¼ 8 Hz, Ar-H), 8.16 (2H, d,
J ¼ 8.00 Hz), 7.78 (2H, d, J ¼ 8 Hz), 7.68 (1H, m), 7.64 (2H, d, J ¼ 8 Hz).
Anal. calcd. for C16H10BrClN2O: C, 53.14; H, 2.79; N, 7.75. Found:
C, 52.93; H, 3.03; N, 8.02.
C17H14N2O2: C, 73.37; H, 5.07; N, 10.07. Found: C, 73.22; H, 4.76; N,
9.86.
6-(4-Methoxy-phenyl)-4-(2-methoxy-phenyl)-1H-pyrimidin-
2-one (VA-17)
Brown powder; yield 90%; mp: 240–242°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.11 (2H, bs), 7.21–7.71 (4H, m), 7.15 (1H, m),
7.09 (2H, d, J ¼ 8Hz), 3.81 (6H, s, OCH3). Anal. calcd. for
C18H16N2O3: C, 70.12; H, 5.23; N, 9.09. Found: C, 69.84; H, 5.46;
N, 8.78.
4,6-Bis-(4-chloro-phenyl)-1H-pyrimidin-2-one (VA-10)
Yellow powder; yield 88%; mp: 274–276°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 12.12 (1H, s, NH, D2O exchangeable
protons), 8.24 (4H, bs), 7.63–7.65 (5H, m). Anal. calcd. for
C16H10Cl2N2O: C, 60.59; H, 3.18; N, 8.83. Found: C, 60.83; H,
2.88; N, 9.10.
6-(3,4-Dimethoxy-phenyl)-4-(4-methoxy-phenyl)-1H-
pyrimidin-2-one (VA-18)
Yellow powder, yield 90%; mp: 255–257°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.16 (2H, d, J ¼ 6 Hz), 7.80 (1H, bs), 7.70 (1H,
bs), 7.42 (1H, s), 7.09–7.12 (3H, m), 3.89 (3H, s, OCH3), 3.86 (6H, s,
OCH3). Anal. calcd. for C19H18N2O4: C, 67.44; H, 5.36; N, 8.28.
Found: C, 67.66; H, 5.42; N, 8.19.
4-(3,4-Dimethoxy-phenyl)-6-naphthalen-2-yl-1H-
pyrimidin-2-one (VA-11)
Dark yallow powder; yield 76%; mp: 279–281°C; 1H NMR (DMSO-
d6, 400 MHz, d, TMS ¼ 0): 11.17 (1H, s, NH, D2O exchangeable
protons), 8.11 (1H, d, J ¼ 8 Hz), 8.04 (1H, d, J ¼ 8 Hz), 7.22–7.61 (6H,
ß 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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Arch. Pharm. Chem. Life Sci. 2014, 347, 1–10
4,6-Diaryl/heteroarylpyrimidin-2(1H)-ones
9
6-(4-Bromo-phenyl)-4-(4-hydroxy-phenyl)-1H-pyrimidin-2-
mixture contained 50 mM potassium phosphate buffer (pH 7.6),
75 mM xanthine, and 0.08 units of XO. Inhibition of XO activity by
various inhibitors was measured by following the decrease in the
uric acid formation at 293 nm at 25°C. The enzyme was
preincubated for 5 min with the test compound dissolved in
DMSO (1% v/v), and the reaction was started by the addition
of xanthine. Final concentration of DMSO (1% v/v) did not
interfere with the enzyme activity. All the experiments were
performed in triplicate and values were expressed as means of
three experiments.
one (VA-19)
Yellow powder; yield 78%; mp: 239–241°C; 1H NMR (DMSO-d6,
300 MHz, d, TMS ¼ 0): 8.00 (2H, d, J ¼ 8Hz), 7.89–7.93 (3H, m), 7.64
(2H, d, J ¼ 7.5Hz), 6.92 (2H, d, J ¼ 7.5 Hz). Anal. calcd. for
C16H11BrN2O2: C, 56.00; H, 3.23; N, 8.16. Found: C, 56.38; H,
3.11; N, 7.95.
6-(4-Chloro-phenyl)-4-(2-chloro-phenyl)-1H-pyrimidin-2-
one (VA-20)
Yellow powder; yield 84%; mp: 274–276°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 11.17 (1H, s, NH, D2O exchangeable proton),
8.18 (3H, m), 7.71–7.72 (6H, m). Anal. calcd. for C16H10Cl2N2O: C,
60.59; H, 3.18; N, 8.83. Found: C, 60.73; H, 2.81; N, 9.09.
Molecular modeling
A flexible docking study of most active compound at the salicylic
acid binding site of XO was performed with the help of GOLD
software [22]. To validate the docking procedure for the
prediction of the correct binding mode of inhibitor at the
salicylic acid binding site, salicylic acid was extracted from
the original X-ray structure (1FIQ) [18] and re-docked. The
highest scoring conformation was selected and compared with
X-ray structure conformation. The docked conformation of
salicylic acid was found to be similar to the original X-ray
structure. The root mean square deviation between the best
6-Phenyl-4-thiophen-2-yl-1H-pyrimidin-2-one (VA-21)
Brown powder; yield 87%; mp: 277–279°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 11.21 (1H, s, NH, D2O exchangeable proton),
8.23 (1H, d, J ¼ 4 Hz), 8.03–8.00 (2H, m), 7.88 (1H, d, J ¼ 4 Hz), 7.56–
7.61 (4H, m), 7.28 (1H, m). Anal. calcd. for C14H10N2OS: C, 66.12; H,
3.96; N, 11.02; S, 12.61 Found: C, 66.12; H, 3.96; N, 11.02; S, 12.61.
scored conformers from docking and X-ray structure was found to
be 0.21 A .
ꢁ
6-Naphthalen-1-yl-4-pyridin-4-yl-1H-pyrimidin-2-one
(VA-22)
Brown powder; yield 80%; mp: 239–241°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 11.19 (1H, s, NH, D2O exchangeable proton),
8.20–8.35 (4H, m), 7.50–7.62 (8H, m). Anal. calcd. for C19H13N3O: C,
76.24; H, 4.38; N, 14.04. Found: C, 76.05; H, 4.62; N, 13.89.
The authors are grateful to the vice chancellor Dr. A. S. Brar and
Dr. Palwinder Singh, Associate Professor, Department of Chemistry,
Guru Nanak Dev University, Amritsar for providing the facilities
necessary for the present study.
6-(3,4-Dimethoxy-phenyl)-4-(3-nitro-phenyl)-1H-pyrimidin-
The authors have declared no conflict of interest.
2-one (VA-23)
Yellow powder; yield 89%; mp: 248–250°C; 1H NMR (DMSO-d6,
400 MHz, d, TMS ¼ 0): 8.69 (1H, s), 8.43 (1H, d, J ¼ 8 Hz), 7.75–7.89
(5H, m), 7.14 (1H, d, J ¼ 8 Hz), 3.90 (3H, s, OCH3), 3.85 (3H, s, OCH3).
Anal. calcd. for C18H15N3O5: C, 61.19; H, 4.28; N, 11.89. Found: C,
60.82; H, 4.17; N, 12.15.
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