Synthesis and evaluation of 1-hydroxy/methoxy-4-methyl-2-phenyl-1 H-imidazole-5-carboxylic acid derivatives as non-purine xanthine oxidase inhibitors

Article abstract of DOI:10.1016/j.ejmech.2015.08.056

Xanthine oxidase is a key enzyme that catalyses hypoxanthine and xanthine to uric acid, whose overproduction leads to the gout-causing hyperuricemia. In this study, a series of 1-hydroxy/methoxy-4-methyl-2-phenyl-1H-imidazole-5-carboxylic acid derivatives (4ae4k and 6ae6k) was synthesized and evaluated for their inhibitory potency against xanthine oxidase. The 1-hydroxyl substituted derivatives 4ae4k showed excellent inhibitory potency with IC₅₀ values ranging from 0.003 μM to 1.2 μM, with compounds 4d (IC₅₀= 0.003 μM), 4e (IC₅₀ = 0.003 μM), and 4f (IC₅₀ = 0.006 μM) manifesting the most potent xanthine oxidase inhibitory potency that were comparable with that of Febuxostat (IC₅₀ = 0.01 μM). LineweavereBurk plot analysis revealed that representative compound 4f acted as a mixed-type inhibitor for xanthine oxidase. The basis of significant inhibition of xanthine oxidase by 4f was rationalized by its molecular docking into the active site of xanthine dehydrogenase.

Full text of DOI:10.1016/j.ejmech.2015.08.056

European Journal of Medicinal Chemistry 103 (2015) 343e353
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Synthesis and evaluation of 1-hydroxy/methoxy-4-methyl-2-phenyl-
1H-imidazole-5-carboxylic acid derivatives as non-purine xanthine
oxidase inhibitors
,
,
Shaolei Chen a 1, Tingjian Zhang b 1, Jian Wang a, Fangyang Wang c, Handong Niu a,
Chunfu Wu c, Shaojie Wang a
,
*
a Key Laboratory of Structure-Based Drugs Design & Discovery of Ministry of Education, School of Pharmaceutical Engineering, Shenyang Pharmaceutical
University, 103 Culture Road, Shenhe District, Shenyang 110016, China
b School of Pharmacy, China Medical University, 77 Puhe Road, North New Area, Shenyang 110122, China
c Department of Pharmacology, Shenyang Pharmaceutical University, 103 Culture Road, Shenhe District, Shenyang 110016, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Xanthine oxidase is a key enzyme that catalyses hypoxanthine and xanthine to uric acid, whose over-
production leads to the gout-causing hyperuricemia. In this study, a series of 1-hydroxy/methoxy-4-
methyl-2-phenyl-1H-imidazole-5-carboxylic acid derivatives (4ae4k and 6ae6k) was synthesized and
evaluated for their inhibitory potency against xanthine oxidase. The 1-hydroxyl substituted derivatives
Received 4 February 2015
Received in revised form
27 August 2015
Accepted 28 August 2015
Available online 3 September 2015
4ae4k showed excellent inhibitory potency with IC50 values ranging from 0.003
compounds 4d (IC50 ¼ 0.003 M), 4e (IC50 ¼ 0.003 M), and 4f (IC50 ¼ 0.006 M) manifesting the most
potent xanthine oxidase inhibitory potency that were comparable with that of Febuxostat
M). LineweavereBurk plot analysis revealed that representative compound 4f acted as a
mM to 1.2 mM, with
m
m
m
Keywords:
4-Methyl-2-phenyl-1H-imidazole-5-
carboxylic acid
(IC50 ¼ 0.01
m
mixed-type inhibitor for xanthine oxidase. The basis of significant inhibition of xanthine oxidase by 4f
was rationalized by its molecular docking into the active site of xanthine dehydrogenase.
© 2015 Elsevier Masson SAS. All rights reserved.
Gout
Xanthine oxidase inhibitor
1. Introduction
StevenseJohnson syndrome [4]. Therefore, identifying some novel
non-purine XO inhibitors with potent XO inhibitory potency and
Gout is a common metabolic disorder characterized by gouty
arthritis and caused by the deposition of monosodium urate crys-
tals within joints. Gout affects millions worldwide and is increasing
in prevalence [1]. As a key enzyme in the last two steps of the
purine metabolic pathway, xanthine oxidase (XO) could catalyze
the oxidation of hypoxanthine to xanthine, and then to uric acid.
The overproduction of uric acid leads to hyperuricemia, which is a
key cause of gout. Therefore, XO is considered the most promising
target for treating this condition [2,3]. Allopurinol (Fig. 1), a pro-
totypical inhibitor of XO, has been the main therapy for the man-
agement of gout and conditions associated with hyperuricemia for
several decades. However, in some cases, severe life-threatening
side effects of allopurinol and its analogs with purine backbone
have been reported, such as fulminant hepatitis, renal failure, and
fewer side effects is extremely necessary. In fact, some excellent
non-purine XO inhibitors have been reported in the recent litera-
ture, including carboxyl moiety containing inhibitors, such as
Febuxostat [5], Y-700 [6], selenazoles [7], isoxazoles [8] and 2-
(indol-5-yl)thiazoles [9]; and non-carboxyl moiety containing in-
hibitors, like Topiroxostat [10], isocytosines [11e13], N-(1,3-diaryl-
3-oxopropyl)amides [14], N-1-acetyl-3,5-diaryl-4,5-dihydro- (1H)
pyrazoles [15], naphthoflavones [16], 2-amino-5-alkylidene-thia-
zol-4-ones [17], azaflavones [18], naphthopyrans [19], 4,6-diaryl/
heteroarylpyrimidin-2(1H)-ones [20], and 2,4-diaryl- pyrano[3,2-c]
chromen-5(4H)-one [21] (Fig. 1). Among them, Febuxostat and
Topiroxostat were successfully introduced into the market in 2009
in the USA and 2013 in Japan, respectively. Indeed, non-purine XO
inhibitors have drawn extensive attention from pharmaceutical
industries worldwide.
In continuation of our previous efforts on finding novel non-
purine XO inhibitors [8], we had designed and synthesized a
series of 1-hydroxy/methoxy-4-methyl-2-phenyl-1H-imidazole-5-
* Corresponding author.
E-mail address: sjwang_99@163.com (S. Wang).
1
These two authors contributed equally to this work.
http://dx.doi.org/10.1016/j.ejmech.2015.08.056
0223-5234/© 2015 Elsevier Masson SAS. All rights reserved.

344
S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
Fig. 1. Chemical structures of Allopurinol, non-purine XO inhibitors, and designed compounds 4ae4k and 6ae6k.
carboxylic acid derivatives (Fig. 1) and tested their XO inhibitory
potency.
formylbenzonitriles with ethyl 2-hydroxyimino-3-oxobutanoate
in the presence of ammonium acetate [22]. 2-Hydroxyimino-3-
oxobutanoate was obtained by nitrosation of ethyl 3-oxobutanoate
with sodium nitrite in acetic acid [23].
2. Results and discussion
Key intermediates 3ae3k were hydrolyzed with sodium hy-
droxide followed by acidification to obtain 1-hydroxyimidazole
series compounds 4ae4k. The methylation of compounds 3ae3k
with dimethyl sulfate to give compounds 5ae5k [24], which were
then hydrolyzed with sodium hydroxide and then acidified to
provide 1-methoxyimidazole series compounds 6ae6k [8].
Structures of the synthesized target compounds were eluci-
dated by ESI-HRMS, IR, 1H NMR, and 13C NMR analyses. All spectral
data were in accordance with assumed structures. In the ESI-HRMS
analysis, 4a showed an [M þ Na]þ ion peak, whereas the other
compounds showed [M þ H]þ ion peaks. The IR spectra of target
compounds displayed cyano group stretching vibrations at
2223e2236 cmꢀ1. In 1H NMR spectra, the 4-methyl group of
imidazole was observed as a singlet at around 2.51 and 2.39 ppm
2.1. Chemistry
The synthesis of the target 1-hydroxy/methoxy-4-methyl-2-
phenyl-1H-imidazole-5-carboxylic acid derivatives 4ae4k and
6ae6k was performed as outlined in Scheme 1. The commercially
available 3-bromo-4-hydroxybenzaldehyde was alkylated with
dimethyl sulfate or appropriate alkyl bromides in DMF in the
presence of potassium carbonate to provide 3-bromo-4-
alkyloxybenzaldehydes 1ae1k, which were treated with cuprous
cyanide in DMF to obtain 2-alkyloxy-5-formylbenzonitriles 2ae2k
[8]. The ethyl 2-(4-alkyloxy-3-cyanophenyl)-1- hydroxy-4-methyl-
1H-imidazole-5-carboxylates 3ae3k, as key intermediates, were
prepared through cyclization of the corresponding 2-alkyloxy-5-

S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
345
Scheme 1. Reagents and conditions: (i) Me2SO4, K2CO3, 0 ꢁC, 2 h or RBr, K2CO3, KI, DMF, 8 h; (ii) CuCN, DMF, 150 ꢁC, 6 h; (iii) NaNO2, AcOH, 0 ꢁC, 3 h; (iv) NH4OAc, AcOH, 50 ꢁC, 24 h;
(v) NaOH, THF, EtOH, H2O, 50 ꢁC, 8 h, then 1 M HCl; (vi) Me2SO4, K2CO3, 0 ꢁC, 1.5 h; (vii) NaOH, THF, EtOH, H2O, 50 ꢁC, 8 h, then 1 M HCl.
for most of the 1-hydroxy substituted compounds 4ae4k and 1-
methoxy substituted compounds 6ae6k, respectively. Moreover,
the 1-methoxy group of imidazole was observed as a singlet at
around 3.95 ppm for 1-methoxy substituted compounds 6ae6k.
magnitude according to their pIC50 values.
As shown in Table 1, the XO inhibitory potency for the 1-
hydroxyl substituted compounds was influenced by the size of
the carbon chain at the 40-position. Potencies decreased when
butoxy groups (n-butoxy, sec-butoxy, and iso-butoxy) were
replaced with methoxy (4a, IC50 ¼ 0.55
m
M), n-propoxy (4b,
2.2. Biological activity
IC50 ¼ 0.13 M), or iso-propoxy (4c, IC50 ¼ 0.17
m
mM). Moreover, it
was also found that the inhibitory potency gradually declined with
the increased size of 40-position groups from butoxy to octyloxy.
The exceptions were compounds 4g and 4h, bearing iso-pentyloxy
and n-hexyloxy, respectively, which had the same inhibitory po-
tency. In addition, to discuss the potency of benzyloxy substitution
at the 40-position, the 4-methoxybenzyloxy-substituted 4k
In vitro bovine XO inhibitory potency for target compounds
4ae4k and 6ae6k were spectrophotometrically measured by
following uric acid levels at 295 nm [25]. Febuxostat was included
as a reference compound. Results are shown in Table 1.
In general, the 1-hydroxyl substituted compounds 4ae4k
exhibited excellent inhibitory potency with IC50 values ranging
(IC50 ¼ 0.11
mM) was synthesized and found to have only moderate
potency. Therefore, the presence of a butoxy group (n-butoxy, sec-
from 0.003
mM
to 1.2
m
M. In particular, compounds 4d
(IC50 ¼ 0.003
m
M, pIC50 ¼ 8.5), 4e (IC50 ¼ 0.003
mM, pIC50 ¼ 8.5),
butoxy, or iso-butoxy) at the 40-position benefited the XO inhibitory
and 4f (IC50 ¼ 0.006
m
M, pIC50 ¼ 8.2), bearing a n-butoxy, sec-
butoxy, and iso-butoxy substituent at the 40-position of phenyl
moiety, respectively, emerged as the most potent XO inhibitors, and
their potencies were 3.3-fold, 3.3-fold, and 1.7-fold higher than that
potency
of
1-hydroxy-4-methyl-2-phenyl-1H-imidazole-5-
carboxylic acid derivatives. This finding was also true for 1-
phenylpyrazoles [6] and 5-phenylisoxazoles [8] XO inhibitors. A
dose-dependent inhibition of representative compound 4f on XO
was exhibited (Fig. 2) and LineweavereBurk plot analysis revealed
of Febuxostat (IC50 ¼ 0.01 M, pIC50 ¼ 8.0), respectively. However,
m
compounds 4d, 4e, 4f and Febuxostat were in the same order of
Table 1
In vitro XO inhibitory potency of designed compounds.
Compound
Inhibition rate at 10
m
g/mL (%)
IC50
(m
M)a
pIC50
Compound
Inhibition rate at 10
m
g/mL (%)
IC50
(
m
M)a
pIC50
4a
4b
4c
4d
4e
4f
4g
4h
90.14 2.01
89.77 1.19
91.04 0.23
89.57 0.54
88.33 1.44
94.08 0.89
91.25 0.73
92.51 0.71
89.93 1.37
88.17 0.11
91.65 0.88
90.89 0.16
0.55
0.13
0.17
0.003
0.003
0.006
0.03
0.03
0.11
1.2
6.3
6.9
6.8
8.5
8.5
8.2
7.5
7.5
7.0
5.9
7.0
8.0
6a
6b
6c
6d
6e
6f
6g
6h
6i
10.68 0.09
18.34 0.63
32.91 0.98
41.19 2.00
83.13 0.70
31.91 0.97
59.55 0.84
80.34 1.26
26.80 1.97
19.00 0.75
86.72 1.59
n.a.b
n.a.
n.a.
n.a.
1.5
n.a.
8.0
3.9
n.a.b
n.a.
n.a.
n.a.
5.8
n.a.
5.1
5.4
4i
4j
4k
n.a.
n.a.
1.1
n.a.
n.a.
5.9
6j
6k
0.11
0.01
Febuxostat
a
IC50 values: the concentration of inhibitor required to produce 50% inhibition of xanthine oxidase (mean SD; n ¼ 4).
b
n.a.: not active (<50% inhibition at 10 mg/mL).

346
S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
obviously decreased compared with the corresponding compounds
4ae4k. The majority of these compounds were inactive (<50% in-
hibition at 10 M), 6g (IC50 ¼ 8.0 M),
g/mL), except 6e (IC50 ¼ 1.5
6h (IC50 ¼ 3.9 M) and 6k (IC50 ¼ 1.1 M), their IC50 values were
497-fold, 268-fold, 129-fold and 10-fold lower than the corre-
sponding 4e (IC50 0.003 M), 4g (IC50 0.03 M), 4h
M). These results suggested
m
m
m
m
m
¼
m
¼
m
(IC50 ¼ 0.03
m
M) and 4k (IC50 ¼ 0.11
m
that 1-hydroxy group was much acceptable than 1-methoxyl group.
2.3. Molecular modeling
XO is a form of molybdoflavin protein xanthine oxidoreductase
[26], and bovine and human sequences of xanthine oxidoreductase
are characterized by 90% identity [27]. The crystal structure of the
complex of bovine xanthine dehydrogenase (XDH) with Febuxostat
shows a highly specific binding pocket, presenting a long, narrow
cavity leading toward the molybdenum-pterin center [5]. More-
over, molybdenum-pterin sites of both XO and XDH are structurally
equivalent [27].
To rationalize the structureeactivity relationships observed and
to gain an insight into the binding mode of synthesized compounds
with XO, docking simulations of 4f and 6f into the binding pocket of
the bovine milk XDH/Febuxostat complex (PDB entry code: 1N5X)
were performed [5]. The simulations were carried out with Auto-
Dock 4 software package [28], and carboxyl groups of 4f and 6f
were calculated in dissociated form. The energy-minimized struc-
ture of Febuxostat was preliminarily docked into XDH to examine
how closely the AutoDock 4 algorithm can reproduce the binding
modes observed in the crystallographic structure. A superposition
of docked Febuxostat onto the crystallographic geometry yielded a
root mean square deviation (RMSD) of 0.95 Å. The hydrogen bonds
predicted by AutoDock 4 were virtually identical to those found in
the crystal structure.
Fig. 2. The inhibition of compound 4f on XO. Values are means SD, n ¼ 3.
that 4f acted as a mixed-type inhibitor for XO (Fig. 3).
To explore the influence of bulk at the 1-position of imidazole
moiety [5], the 1-methoxyl-substituted compounds 6ae6k were
designed and synthesized. Biological data presented in Table 1
showed that the inhibitory potency of all these compounds
As shown in Fig. 4, compound 4f interacted very closely with
original ligand (Febuxostat) by hydrogen bonds that involved the
carboxylate moiety interacting with Arg880 and Thr1010, the ni-
trogen atom of imidazole forming hydrogen bond with Glu802, and
the nitrile group of phenyl unit forming hydrogen bond with
Asn768 [5]. More importantly, 1-hydroxyl group of 4f exhibited an
extra hydrogen bond interaction with Thr1010, which may help 4f
to anchor onto the binding site more tightly. Furthermore, the
calculated pKa of 4f was 2.01, lower than that of Febuxostat (2.39),
which may benefit the interaction of carboxylate group with gua-
nidinium side chain of Arg880 [5,13]. These observations could
provide a rational explanation for the inhibitory potency difference
between 4f and Febuxostat and also indicated that introducing a
hydroxyl group into the position between phenyl unit and carboxyl
moiety may be favorable for Febuxostat analogues to display better
XO inhibitory potency.
Meanwhile, 6f, which may have bumped with protein residues
for the presence of 1-methoxy group, interacted in a distinct way
with XDH by inserting the iso-butoxy moiety into the channel
instead of carboxylate segment. Consequently, carboxylate moiety
showed an interaction with Lys771, and the nitrile group of phenyl
unit formed a hydrogen bond with Thr1010. These two interactions
may help fix the molecule at the binding site of XO. However, in-
teractions between 6f and XO may be too weak compared with
those of 4f and Febuxostat to produce any apparent inhibitory
potency in the experimental test. As exemplified by 6f, other
compounds in the 1-methoxy series may interact with XO in a
similar pattern, which could reasonably explain their inferior po-
tencies compared with those of their counterparts in the 1-hydroxy
series.
Fig. 3. LineweavereBurk plot of the inhibition of xanthine oxidase by compound 4f.

S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
347
Fig. 4. Binding modes of 4f (A) and 6f (B) within the protein binding pocket.
3. Conclusions
carried out using silica gel plates (Qingdao Makall Group Co., Ltd.,
Qingdao, China), and spots were visualized under UV light at
254 nm.
We report the synthesis and in vitro XO inhibitory potency of 1-
hydroxy/methoxy-4-methyl-2-phenyl-1H-imidazole-5-carboxylic
acid derivatives. 1-Hydroxyl substituted derivatives 4a-4k were
found to have excellent inhibitory potency, with IC50 values ranging
4.1.1. Synthesis of 3-bromo-4-methoxybenzaldehyde (1a)
Dimethyl sulfate (6.6 g, 0.053 mol) was added dropwise at a rate
that maintained <0 ꢁC reaction temperature to a stirred solution of
3-bromo-4-hydroxybenzaldehyde (10 g, 0.05 mol) and anhydrous
potassium carbonate (10.28 g, 0.075 mol) in DMF (40 mL). Upon
completion of addition, the mixture was stirred at 25 ꢁC for another
2 h. The reaction mixture was slowly poured into cold water
(100 mL) and stirred for 20 min. The solid was separated by
filtration as a gray powder (9.7 g) in 90.6% yield, mp 40.1e41.0 ꢁC.
from 0.003
mM
to 1.2
m
M. Among them, compounds 4d
(IC50 ¼ 0.003
m
M), 4e (IC50 ¼ 0.003
m
M), and 4f (IC50 ¼ 0.006 M)
m
emerged as the most potent XO inhibitors comparable with
Febuxostat (IC50 ¼ 0.01
mM).1-Hydroxy group was much acceptable
than 1-methoxyl group. Molecular docking studies rationalized the
structureeactivity relationships observed and provided insight into
the binding mode of the synthesized compounds with XO. Line-
weavereBurk plot analysis revealed that representative compound
4f acted as a mixed-type inhibitor for xanthine oxidase. The most
potent compounds 4d, 4e, and 4f appeared to be promising can-
didates for treating hyperuricemia, gout, and other diseases
involving XO. Moreover, our research indicated that introducing a
hydroxyl group into the position between phenyl unit and carboxyl
moiety may be favorable for Febuxostat analogues to display better
XO inhibitory potency.
1H NMR (300 MHz, CDCl3):
d 9.84 (s, 1H, CHO), 8.08 (d, 1H,
J ¼ 2.0 Hz, AreH), 7.80 (dd, 1H, J ¼ 2.0 & 8.5 Hz, AreH), 6.98 (d, 1H,
J ¼ 8.5 Hz, AreH), 3.86 (s, 3H, OCH3).
4.1.2. General procedure for the synthesis of 3-bromo-4-
alkoxybenzaldehydes 1be1k
A solution of 3-bromo-4-hydroxybenzaldehyde (50 mmol),
anhydrous potassium carbonate (65 mmol), potassium iodide
(1 mmol), and alkyl bromide (75 mmol) in DMF (40 mL) was stirred
at 50 ꢁC under nitrogen atmosphere for 8 h. After filtering the re-
action mixture, the filtrate was concentrated in a vacuum. Ethyl
acetate was added to the residue, and the ethyl acetate layer was
sequentially washed with water and brine. After drying with
anhydrous sodium sulfate and evaporation, the crude product was
purified by column chromatography (petroleum ether) to provide
the corresponding 4-alkoxy-3-bromobenzaldehydes 1be1k.
4. Experimental protocols
4.1. Chemistry
Reagents and solvents were purchased from commercial sour-
ces, and some were redistilled under a positive pressure of dry
nitrogen atmosphere in the presence of an appropriate desiccant
when necessary. Melting points were obtained using an YRT-3
melting apparatus and were uncorrected. 1H NMR and 13C NMR
spectra were recorded on a Bruker 300, 400, or 600 MHz spec-
trometer. Chemical shifts were expressed in parts per million using
tetramethylsilane as an internal reference and DMSO-d6 or CDCl3 as
solvent. IR spectra were determined as KBr pellets on a Bruker IFS-
55 spectrometer and expressed in per centimeter. ESI-MS data were
obtained using an Agilent 1100 instrument, and the ESI-HRMS data
were obtained using a Bruker microTOF-Q instument. TLC was
4.1.2.1. 3-Bromo-4-propoxybenzaldehyde (1b). A yellow powder,
yield 80.5%, mp 38.3e39.5 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.83 (s,
1H, CHO), 8.07 (d, 1H, J ¼ 1.9 Hz, AreH), 7.79 (dd, 1H, J ¼ 2.0 &
8.5 Hz, AreH), 6.98 (d, 1H, J ¼ 8.5 Hz, AreH), 4.08 (t, 2H, J ¼ 6.4 Hz,
OCH2), 1.90 (m, 2H, CH2), 1.10 (t, 3H, J ¼ 7.4 Hz, CH3).
4.1.2.2. 3-Bromo-4-iso-propoxybenzaldehyde (1c). A pale yellow oil,
yield 77.0%. 1H NMR (300 MHz, CDCl3):
d 9.83 (s,1H, CHO), 8.08 (d,

348
S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
1H, J ¼ 2.0 Hz, AreH), 7.78 (dd, 1H, J ¼ 2.0 & 8.5 Hz, AreH), 7.00 (d,
1H, J ¼ 8.6 Hz, AreH), 4.71 (m, 1H, OCH), 1.43 (d, 6H, J ¼ 6.1 Hz,
2CH3).
washed with ammonia water, water, and brine. After drying with
anhydrous sodium sulfate and evaporation, the crude product was
purified by column chromatography (petroleum ether) to give the
corresponding 2-alkoxy-5-formylbenzonitriles 2ae2k.
4.1.2.3. 3-Bromo-4-butoxybenzaldehyde (1d). A white waxy solid,
yield 90.0%, mp 37.2e38.6 ꢁC. 1H NMR (300 MHz, CDCl3):
d
9.83 (s,
4.1.3.1. 5-Formyl-2-methoxybenzonitrile (2a). A white powder,
1H, CHO), 8.08 (d, 1H, J ¼ 2.0 Hz, AreH), 7.79 (dd, 1H, J ¼ 2.0 &
8.5 Hz, AreH), 6.98 (d, 1H, J ¼ 8.5 Hz, AreH), 4.12 (t, 2H, J ¼ 6.4 Hz,
OCH2), 1.86 (m, 2H, CH2), 1.56 (m, 2H, CH2), 1.01 (t, 3H, J ¼ 7.4 Hz,
CH3).
yield 56.5%, mp 70.1e72.0 ꢁ 1H NMR (300 MHz, CDCl3):
C d 9.88 (s,
1H, CHO), 8.10 (d, 1H, J ¼ 2.0 Hz, AreH), 8.06 (dd, 1H, J ¼ 2.1 &
8.67 Hz, AreH), 7.10 (d, 1H, J ¼ 8.7 Hz, AreH), 3.85 (s, 3H, OCH3).
4.1.3.2. 5-Formyl-2-propoxybenzonitrile (2b). A yellow powder,
4.1.2.4. 3-Bromo-4-sec-butoxybenzaldehyde (1e). A yellow oil, yield
yield 91.6%, mp 51.1e54.0 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.89 (s,
80.5%. 1H NMR (300 MHz, CDCl3):
d
9.82 (s, 1H, CHO), 8.07 (d, 1H,
1H, CHO), 8.09 (d, 1H, J ¼ 1.6 Hz, AreH), 8.06 (dd, 1H, J ¼ 1.6 &
8.76 Hz, AreH), 7.10 (d, 1H, J ¼ 8.7 Hz, AreH), 4.15 (t, 2H, J ¼ 6.5 Hz,
OCH2), 1.92 (m, 2H, CH2), 1.11 (t, 3H, J ¼ 7.4, CH3).
J ¼ 2.0 Hz, AreH), 7.77 (dd, 1H, J ¼ 2.0 & 8.5 Hz, AreH), 6.97 (d, 1H,
J ¼ 8.5 Hz, AreH), 4.48 (m, 1H, OCH), 1.78 (m, 2H, CH2), 1.38 (d, 3H,
J ¼ 6.1 Hz, CH3), 1.02 (t, 3H, J ¼ 7.4 Hz, CH3).
4.1.3.3. 5-Formyl-2-iso-propoxybenzonitrile (2c). A pale yellow
powder, yield 85.3%, mp 51.5e52.6 ꢁC. 1H NMR (300 MHz, CDCl3):
4.1.2.5. 3-Bromo-4-iso-butoxybenzaldehyde (1f). A yellow oil, yield
64.7%. 1H NMR (300 MHz, CDCl3):
d
9.83 (s, 1H, CHO), 8.07 (d, 1H,
d
9.88 (s, 1H, CHO), 8.09 (d, 1H, J ¼ 2.0 Hz, AreH), 8.05 (dd, 1H,
J ¼ 1.9 Hz, AreH), 7.79 (dd, 1H, J ¼ 2.0 & 8.5 Hz, AreH), 6.97 (d, 1H,
J ¼ 8.5 Hz, AreH), 3.88 (d, 2H, J ¼ 6.4 Hz, OCH2), 2.12 (m, 1H, CH),
1.09 (d, 6H, J ¼ 6.7 Hz, 2CH3).
J ¼ 2.1 & 8.7 Hz, AreH), 7.10 (d, 1H, J ¼ 8.8 Hz, AreH), 4.80 (m, 1H,
OCH), 1.46 (d, 6H, J ¼ 6.1 Hz, 2CH3).
4.1.3.4. 2-Butoxy-5-formylbenzonitrile (2d). A white waxy solid,
4.1.2.6. 3-Bromo-4-iso-pentyloxybenzaldehyde (1g). A pale yellow
powder, yield 98.0%, mp 35.5e35.9 ꢁC. 1H NMR (300 MHz, CDCl3):
yield 88.0%, mp 21.8e23.0 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.89 (s,
1H, CHO), 8.09 (d, 1H, J ¼ 2.0 Hz, AreH), 8.06 (dd, 1H, J ¼ 2.1 &
8.6 Hz, AreH), 7.10 (d, 1H, J ¼ 8.7 Hz, AreH), 4.20 (t, 2H, J ¼ 6.4 Hz,
OCH2), 1.87 (m, 2H, CH2), 1.56 (m, 2H, CH2), 1.03 (t, 3H, J ¼ 7.4, CH3).
d
9.84 (s, 1H, CHO), 8.08 (d, 1H, J ¼ 2.0 Hz, AreH), 7.80 (dd, 1H,
J ¼ 2.0 & 8.5 Hz, AreH), 6.99 (d, 1H, J ¼ 8.5 Hz, AreH), 4.15 (t, 2H,
J ¼ 6.5 Hz, OCH2), 1.90 (m, 1H, CH), 1.78 (m, 2H, CH2), 0.98 (d, 6H,
J ¼ 6.5 Hz, 2CH3).
4.1.3.5. 2-sec-Butoxy-5-formylbenzonitrile (2e). A yellow oil, yield
83.9%. 1H NMR (300 MHz, CDCl3):
d 9.88 (s, 1H, CHO), 8.09 (d, 1H,
4.1.2.7. 3-Bromo-4-hexyloxybenzaldehyde (1h). A pale yellow
powder, yield 90.9%, mp 36.8e37.4 ꢁC. 1H NMR (300 MHz, CDCl3):
J ¼ 1.8 Hz, AreH), 8.04 (dd, 1H, J ¼ 1.8 & 8.8 Hz, AreH), 7.09 (d, 1H,
J ¼ 8.8 Hz, AreH), 4.56 (m, 1H, OCH), 1.79 (m, 2H, CH2), 1.41 (d, 3H,
J ¼ 6.1 Hz, CH3), 1.01 (t, 3H, J ¼ 7.4, CH3).
d
9.84 (s, 1H, CHO), 8.08 (d, 1H, J ¼ 2.0 Hz, AreH), 7.79 (dd, 1H,
J ¼ 2.0 & 8.5 Hz, AreH), 6.98 (d, 1H, J ¼ 8.5 Hz, AreH), 4.12 (t, 2H,
J ¼ 6.5 Hz, OCH2), 1.88 (m, 2H, CH2), 1.52 (m, 2H, CH2), 1.39e1.34 (m,
4H, 2CH2), 0.92 (t, 3H, J ¼ 7.0 Hz, CH3).
4.1.3.6. 2-iso-Butoxy-5-formylbenzonitrile (2f). A white powder,
yield 77.8%, mp 52.8e53.9 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.89 (s,
1H, CHO), 8.10 (d, 1H, J ¼ 2.0 Hz, AreH), 8.06 (dd, 1H, J ¼ 2.1 &
8.7 Hz, AreH), 7.09 (d, 1H, J ¼ 8.7 Hz, AreH), 3.95 (t, 2H, J ¼ 6.5 Hz,
OCH2), 2.22 (m, 1H, CH), 1.01 (d, 6H, J ¼ 6.8, 2CH3).
4.1.2.8. 3-Bromo-4-heptyloxybenzaldehyde (1i). A white powder,
yield 98.0%, mp 49.3e50.1 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.84 (s,
1H, CHO), 8.08 (d, 1H, J ¼ 1.9 Hz, AreH), 7.79 (dd, 1H, J ¼ 1.9 &
8.5 Hz, AreH), 6.98 (d, 1H, J ¼ 8.5 Hz, AreH), 4.12 (t, 2H, J ¼ 6.5 Hz,
OCH2),1.87 (m, 2H, CH2), 1.49 (m, 2H, CH2),1.42e1.34 (m, 6H, 3CH2),
0.90 (t, 3H, J ¼ 6.5 Hz, CH3).
4.1.3.7. 5-Formyl-2-iso-pentyloxybenzonitrile (2g). A white powder,
yield 81.3%, mp 46.1e47.8 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.90 (s,
1H, CHO), 8.09 (s, 1H, AreH), 8.06 (d, 1H, J ¼ 8.8 Hz, AreH), 7.11 (d,
1H, J ¼ 8.8 Hz, AreH), 4.21 (t, 2H, J ¼ 6.5 Hz, OCH2), 1.91 (m, 1H, CH),
1.80 (m, 2H, CH2), 1.00 (d, 6H, J ¼ 6.6, 2CH3).
4.1.2.9. 3-Bromo-4-octyloxybenzaldehyde (1j). A yellow powder,
yield 97.4%, mp 44.3e45.2 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.83 (s,
1H, CHO), 8.08 (d, 1H, J ¼ 2.0 Hz, AreH), 7.80 (dd, 1H, J ¼ 2.0 &
8.5 Hz, AreH), 6.98 (d, 1H, J ¼ 8.5 Hz, AreH), 4.12 (t, 2H, J ¼ 6.5 Hz,
OCH2),1.88 (m, 2H, CH2),1.54 (m, 2H, CH2), 1.42e1.34 (m, 8H, 4CH2),
0.89 (t, 3H, J ¼ 6.5 Hz, CH3).
4.1.3.8. 5-Formyl-2-hexyloxybenzonitrile (2h). A white waxy solid,
yield 86.0%, mp 20.7e21.1 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.89 (s,
1H, CHO), 8.10 (d, 1H, J ¼ 1.6 Hz, AreH), 8.06 (dd, 1H, J ¼ 2.0 &
8.70 Hz, AreH), 7.10 (d, 1H, J ¼ 8.7 Hz, AreH), 4.12 (t, 2H, J ¼ 6.5 Hz,
OCH2), 1.90 (m, 2H, CH2), 1.50 (m, 2H, CH2), 1.37e1.34 (m, 4H, 2CH2),
0.92 (t, 3H, J ¼ 6.9 Hz, CH3).
4.1.2.10. 3-Bromo-4-(4-methylbenzyloxy)benzaldehyde
white powder, yield 88.0%, mp 49.3e50.1 ꢁC. 1H NMR (300 MHz,
CDCl3):
(1k). A
d
9.85 (s, 1H, CHO), 8.08 (d, 1H, J ¼ 1.9 Hz, AreH), 7.78 (dd,
4.1.3.9. 5-Formyl-2-heptyloxybenzonitrile (2i). A yellow oil, yield
1H, J ¼ 1.9 & 8.5 Hz, AreH), 7.38 (d, 2H, J ¼ 8.0 Hz, AreH), 7.24 (d,
2H, J ¼ 7.8 Hz, AreH), 6.98 (d, 1H, J ¼ 8.5 Hz, AreH), 5.32 (s, 2H,
CH2), 2.32 (s, 3H, CH3).
64.6%. 1H NMR (300 MHz, CDCl3):
d 9.89 (s, 1H, CHO), 8.09 (s, 1H,
AreH), 8.06 (d, 1H, J ¼ 8.7 Hz, AreH), 7.10 (d, 1H, J ¼ 8.6 Hz, AreH),
4.13 (t, 2H, J ¼ 6.5 Hz, OCH2), 1.88 (m, 2H, CH2), 1.51e1.32 (m, 8H,
4CH2), 0.90 (t, 3H, J ¼ 6.3 Hz, CH3).
4.1.3. General procedure for the synthesis of 2-alkoxy-5-
formylbenzonitriles 2ae2k
4.1.3.10. 5-Formyl-2-octyloxybenzonitrile (2j). A yellow powder,
A
mixture of 4-alkoxy-3-bromobenzaldehyde (100 mmol),
yield 75.0%, mp 36.2e36.8 ꢁC. 1H NMR (300 MHz, CDCl3):
d 9.89 (s,
CuCN (110 mmol), and DMF (100 mL) was stirred at 150 ꢁC under
nitrogen atmosphere for 8 h. The reaction mixture was cooled to
room temperature and diluted with CH2Cl2. The insoluble sub-
stance was filtered off, and the organic layer was sequentially
1H, CHO), 8.09 (d, 1H, J ¼ 1.4 Hz, AreH), 8.06 (dd, 1H, J ¼ 1.4 &
8.8 Hz, AreH), 7.09 (d, 1H, J ¼ 8.8 Hz, AreH), 4.18 (t, 2H, J ¼ 6.5 Hz,
OCH2), 1.90 (m, 2H, CH2), 1.50 (m, 2H, CH2),1.38e1.25 (m, 8H, 4CH2),
0.90 (t, 3H, J ¼ 6.6 Hz, CH3).

S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
349
4.1.3.11. 5-Formyl-2-(4-methylbenzyloxy)benzonitrile (2k). A gray
powder, yield 81.0%, mp 146.1e147.8 ꢁC. 1H NMR (300 MHz, CDCl3):
4.00 (d, 2H, J ¼ 6.5 Hz, CH2), 2.38 (s, 3H, CH3), 2.08 (m, 1H, CH), 1.32
(t, 3H, J ¼ 7.1 Hz, CH3), 1.02 (d, 6H, J ¼ 6.7 Hz, 2CH3). ESI-MS m/z
344.4 [M þ H]þ.
d
9.90 (s, 1H, CHO), 8.08 (s, 1H, AreH), 8.06 (d, 1H, J ¼ 8.8 Hz, AreH),
7.38 (d, 2H, J ¼ 7.9 Hz, AreH), 7.25 (d, 2H, J ¼ 7.9 Hz, AreH), 7.10 (d,
1H, J ¼ 8.8 Hz, AreH), 5.32 (s, 2H, CH2), 2.32 (s, 3H, CH3).
4.1.4.7. Ethyl 2-(3-cyano-4-iso-pentyloxyphenyl)-1-hydroxy-4-
methyl-1H-imidazole-5- carboxylate (3g). A white crystalline pow-
der, yield 69.7%, mp 148.8e150.1 ꢁC. 1H NMR (300 MHz, DMSO-d6):
4.1.4. General procedure for the synthesis of ethyl 2-(4-alkoxy- 3-
cyanophenyl)-1-hydroxy-4-methyl-1H-imidazole-5-carboxylates
3ae3k
d
8.41 (s,1H, AreH), 8.25 (dd,1H, J ¼ 2.1 &8.7 Hz, AreH), 7.21 (d,1H,
J ¼ 9.0 Hz, AreH), 4.20 (q, 2H, J ¼ 6.9 Hz, CH2), 4.18 (t, 2H, J ¼ 6.3 Hz,
CH2), 2.24 (s, 3H, CH3), 1.85 (m, 1H, CH), 1.68 (q, 2H, J ¼ 6.6 Hz, CH2),
1.27 (t, 3H, J ¼ 6.9 Hz, CH3), 0.98 (d, 6H, J ¼ 6.6 Hz, 2CH3). ESI-MS m/
z 358.4 [M þ H]þ.
A mixture of 2-alkoxy-5-formylbenzonitrile (43 mmol), ethyl 2-
hydroxyimino-3-oxobutanoate (52 mmol), ammonium acetate
(430 mmol), and acetic acid (176 mL) was stirred at 50 ꢁC under
nitrogen atmosphere for 24 h. The reaction mixture was cooled to
room temperature and then slowly poured into cold water
(500 mL). The resulting precipitate was filtered, dried, and washed
with ethyl acetate to obtain the corresponding ethyl 2-(4-alkoxy- 3-
cyanophenyl)-1-hydroxy-4-methyl-1H-imidazole-5-carboxylates
3ae3k.
4.1.4.8. Ethyl
1H-imidazole-5-carboxylate (3h). A white crystalline powder, yield
64.1%, mp 109.3e112.8 ꢁC. 1H NMR (300 MHz, DMSO-d6):
12.14 (s,
2-(3-cyano-4-hexyloxyphenyl)-1-hydroxy-4-methyl-
d
1H, OH), 8.29 (s, 1H, AreH), 8.27 (s, 1H, AreH), 7.38 (d, 1H,
J ¼ 8.6 Hz, AreH), 4.29 (q, 2H, J ¼ 7.1 Hz, CH2), 4.20 (t, 2H, J ¼ 6.4 Hz,
CH2), 2.50 (s, 3H, CH3), 1.77 (m, 2H, CH2), 1.46 (m, 2H, CH2),
1.35e1.29 (m, 7H, 2CH2 & CH3), 0.87 (t, 3H, J ¼ 6.9 Hz, CH3). ESI-MS
m/z 372.4 [M þ H]þ.
4.1.4.1. Ethyl
1H-imidazole-5-carboxylate (3a). A white crystalline powder, yield
65.8%, mp 173.2e175.1 ꢁC. 1H NMR (300 MHz, DMSO-d6):
8.32 (d,
2-(3-cyano-4-methoxyphenyl)-1-hydroxy-4-methyl-
d
1H, J ¼ 2.1 Hz, AreH), 8.21 (dd, 1H, J ¼ 2.1 & 9.0 Hz, AreH), 7.14 (d,
1H, J ¼ 8.1 Hz, AreH), 4.41 (q, 2H, J ¼ 6.9 Hz, CH2), 3.94 (s, 3H, CH3),
2.19 (s, 3H, CH3), 1.26 (t, 3H, J ¼ 6.9 Hz, CH3). ESI-MS m/z 302.3
[M þ H]þ.
4.1.4.9. Ethyl 2-(3-cyano-4-heptyloxyphenyl)-1-hydroxy-4-methyl-
1H-imidazole-5-carboxylate (3i). A white crystalline powder, yield
66.8%, mp 139.5e142.2 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.27 (d,
1H, J ¼ 2.0 Hz, AreH), 8.24 (dd, 1H, J ¼ 2.2 & 8.6 Hz, AreH), 7.39 (d,
1H, J ¼ 8.9 Hz, AreH), 4.24 (q, 2H, J ¼ 7.1 Hz, CH2), 4.18 (t, 2H,
J ¼ 6.4 Hz, CH2), 2.31 (s, 3H, CH3), 1.75 (m, 2H, CH2), 1.48e1.28 (m,
11H, 4CH2 & CH3), 0.88 (t, 3H, J ¼ 6.7 Hz, CH3). ESI-MS m/z 386.5
[M þ H]þ.
4.1.4.2. Ethyl
1H-imidazole-5-carboxylate (3b). A white crystalline powder, yield
62.1%, mp 152.2e154.1 ꢁC. 1H NMR (300 MHz, DMSO-d6):
8.29 (dd,
2-(3-cyano-4-propoxyphenyl)-1-hydroxy-4-methyl-
d
1H, J ¼ 2.1 &8.9 Hz, AreH), 8.26 (s, 1H, AreH), 7.40 (d,1H, J ¼ 9.0 Hz,
AreH), 4.38 (q, 2H, J ¼ 7.2 Hz, CH2), 4.22 (t, 2H, J ¼ 6.6 Hz, CH2), 2.25
(s, 3H, CH3),1.84 (m, 2H, CH2),1.35 (t, 3H, J ¼ 7.3 Hz, CH3),1.03 (t, 3H,
J ¼ 7.1 Hz, CH3). ESI-MS m/z 330.4 [M þ H]þ.
4.1.4.10. Ethyl 2-(3-cyano-4-octyloxyphenyl)-1-hydroxy-4-methyl-
1H-imidazole-5-carboxylate (3j). A white crystalline powder, yield
72.1%, mp 143.2e145.1 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.35 (d,
1H, J ¼ 1.8 Hz, AreH), 8.19 (dd, 1H, J ¼ 2.1 & 9.0 Hz, AreH), 7.26 (d,
1H, J ¼ 8.9 Hz, AreH), 5.24 (s, 2H, CH2), 4.14 (q, 2H, J ¼ 7.2 Hz, CH2),
2.33 (s, 3H, CH3), 1.79 (m, 2H, CH2), 1.45e1.20 (m, 13H, 5CH2 & CH3),
0.90 (t, 3H, J ¼ 6.7 Hz, CH3). ESI-MS m/z 400.4 [M þ H]þ.
4.1.4.3. Ethyl 2-(3-cyano-4-iso-propoxyphenyl)-1-hydroxy-4-
methyl-1H-imidazole-5-carboxylate (3c). A white crystalline pow-
der, yield 70.1%, mp 150.2e151.9 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d
8.26 (dd, 1H, J ¼ 2.1 & 8.8 Hz, AreH), 8.23 (d, 1H, J ¼ 2.1 Hz, AreH),
7.40 (d, 1H, J ¼ 8.9 Hz, AreH), 4.88 (m, 1H, CH), 4.35 (q, 2H,
J ¼ 7.2 Hz, CH2), 2.42 (s, 3H, CH3), 1.38 (d, 6H, J ¼ 6.3 Hz, 2CH3), 1.35
(t, 3H, J ¼ 7.0 Hz, CH3). ESI-MS m/z 330.4 [M þ H]þ.
4.1.4.11. Ethyl 2-[3-cyano-4-(4-methylbenzyloxy)phenyl]-1-hydroxy-
4-methyl-1H- imidazole-5-carboxylate (3k). A white crystalline
powder, yield 70.0%, mp 150.3e151.5 ꢁC. 1H NMR (300 MHz, DMSO-
d6):
d
8.27 (dd,1H, J ¼ 2.1 &7.2 Hz, AreH), 8.26 (s,1H, AreH), 7.51 (d,
4.1.4.4. Ethyl 2-(4-butoxy-3-cyanophenyl)-1-hydroxy-4-methyl-1H-
1H, J ¼ 7.2 Hz, AreH), 7.39 (d, 2H, J ¼ 8.1 Hz, AreH), 7.24 (d, 2H,
J ¼ 7.8 Hz, AreH), 5.32 (s, 2H, CH2), 4.31 (q, 2H, J ¼ 7.2 Hz, CH2), 2.40
(s, 3H, CH3), 2.32 (s, 3H, CH3), 1.33 (t, 3H, J ¼ 7.2 Hz, CH3). ESI-MS m/
z 392.4 [M þ H]þ.
imidazole-5-carboxylate (3d). A white crystalline powder, yield
71.0%, mp 149.8e150.5 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 12.16 (s,
1H, OH), 8.28 (dd, 1H, J ¼ 2.1 & 8.7 Hz, AreH), 8.26 (d, 1H, J ¼ 2.4 Hz,
AreH), 7.39 (d, 1H, J ¼ 9.6 Hz, AreH), 4.30 (q, 2H, J ¼ 7.2 Hz, CH2),
4.21 (t, 2H, J ¼ 6.3 Hz, CH2), 2.37 (s, 3H, CH3), 1.77 (m, 2H, CH2), 1.48
(m, 2H, CH2), 1.32 (t, 3H, J ¼ 6.9 Hz, CH3), 0.96 (t, 3H, J ¼ 7.5 Hz, CH3).
ESI-MS m/z 344.4 [M þ H]þ.
4.1.5. General procedure for the synthesis of 2-(4-alkoxy- 3-
cyanophenyl)-1-hydroxy-4-methyl-1H-imidazole-5-carboxylic
acids 4ae4k
A mixture of ethyl 2-(4-alkoxy-3-cyanophenyl)-1-hydroxy-4-
4.1.4.5. Ethyl 2-(4-sec-butoxy-3-cyanophenyl)-1-hydroxy-4-methyl-
methyl-1H-imidazole-5- carboxylate (2.9 mmol), 1 M NaOH
1H-imidazole-5-carboxylate (3e). A white crystalline powder, yield
aqueous (11 mL), THF (5 mL), and ethanol (5 mL) was stirred at
50 ꢁC for 8 h. The solvent was concentrated in a vacuum, and the
residue was acidified with dilute hydrochloric acid to pH 1. The
resulting precipitate was filtered, dried, and recrystallized with a
mixture of methanol and ethyl acetate (2:1) to yield the corre-
sponding 2-(4-alkoxy- 3-cyanophenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acids 4ae4k.
69.8%, mp 159.8e162.3 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 12.11 (s,
1H, OH), 8.28 (s, 1H, AreH), 8.24 (d, 1H, J ¼ 9.1 Hz, AreH), 7.41 (d,
1H, J ¼ 9.0 Hz, AreH), 4.67 (m, 1H, CH), 4.29 (q, 2H, J ¼ 9.1 Hz, CH2),
2.38 (s, 3H, CH3), 1.77 (m, 2H, CH2), 1.30 (m, 6H, 2CH3), 0.96 (t, 3H,
J ¼ 7.4 Hz, CH3). ESI-MS m/z 344.4 [M þ H]þ.
4.1.4.6. Ethyl 2-(4-iso-butoxy-3-cyanophenyl)-1-hydroxy-4-methyl-
1H-imidazole-5-carboxylate (3f). A white crystalline powder, yield
4.1.5.1. 2-(3-Cyano-4-methoxyphenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4a). A white crystalline powder, yield
52.5%. HPLC: 98.15%. mp 214.1e214.5 ꢁC. 1H NMR (300 MHz, DMSO-
69.4%, mp 147.4e148.3 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 12.12 (s,
1H, OH), 8.30 (d, 1H, J ¼ 2.2 Hz, AreH), 8.28 (dd,1H, J ¼ 2.2 & 9.6 Hz,
AreH), 7.39 (d, 1H, J ¼ 9.6 Hz, AreH), 4.29 (q, 2H, J ¼ 7.1 Hz, CH2),
d6):
d
8.60 (d, 1H, J ¼ 2.0 Hz, AreH), 8.44 (dd, 1H, J ¼ 2.2 & 8.8 Hz,

350
S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
AreH), 7.27 (d,1H, 8.9 Hz, AreH), 3.88 (s, 3H, CH3), 2.42 (s, 3H, CH3).
13C NMR (150 MHz, DMSO-d6):
160.95, 160.37, 142.98, 139.84,
132.55, 131.20, 117.28, 116.67, 115.58, 113.69, 101.00, 70.70, 21.72.
ESI-HRMS calcd. for C13H11N3NaO4 [MþNa]þ 296.0642, found:
296.0645. IR (KBr, cmꢀ1): 3459.7, 2967.7, 2230.2, 1611.0, 1502.3,
1376.5, 1304.5, 1001.4.
4.1.5.7. 2-(3-Cyano-4-iso-pentyloxyphenyl)-1-hydroxy-4-methyl-
1H-imidazole-5-carboxylic acid (4g). A white crystalline powder,
yield 64.6%. HPLC: 97.36%. mp 194.7e194.9 ꢁC. 1H NMR (300 MHz,
d
DMSO-d6):
d 8.54 (s, 1H, AreH), 8.48 (s, 1H, AreH), 7.50 (s, 1H,
AreH), 4.25 (s, 2H, CH2), 2.51 (s, 3H, CH3), 1.82 (s, 1H, CH), 1.70 (s,
2H, CH2), 0.97 (s, 6H, 2CH3). 13C NMR (150 MHz, DMSO-d6):
d
160.89, 160.33, 142.94, 139.80, 132.34, 131.03, 117.27, 116.59,
115.54, 113.57, 100.96, 69.33, 30.86, 28.21, 24.91, 22.04. ESI-HRMS
calcd. for C17H20N3O4 [M þ H]þ 330.1148, found: 330.1149. IR
(KBr, cmꢀ1): 3458.6, 2930.4, 2230.1, 1610.7, 1495.4, 1373.9, 1294.4.
4.1.5.2. 2-(3-Cyano-4-propoxyphenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4b). A white crystalline powder, yield
62.5%. HPLC: 96.12%. mp 191.9e193.0 ꢁC. 1H NMR (300 MHz, DMSO-
d6):
d
13.69 (s, 1H, COOH), 8.54 (s, 1H, AreH), 8.46 (d, 1H, J ¼ 8.7 Hz,
AreH), 7.45 (d, 1H, J ¼ 8.8 Hz, AreH), 4.18 (t, 2H, J ¼ 6.3 Hz, CH2),
2.50 (s, 3H, CH3), 1.80 (m, 2H, CH2), 1.03 (t, 3H, J ¼ 7.4 Hz, CH3). 13
C
4.1.5.8. 2-(3-Cyano-4-hexyloxyphenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4h). A white crystalline powder, yield
75.0%. HPLC: 99.74%. mp 190.8e191.1 ꢁC. 1H NMR (300 MHz, DMSO-
NMR (150 MHz, DMSO-d6):
d 160.00, 159.78, 142.97, 139.78, 133.65,
132.22, 120.34, 116.42, 115.75, 114.48, 101.88, 66.81, 21.54, 15.75,
10.96. ESI-HRMS calcd. for C15H16N3O4 [MþH]þ 302.1135, found:
302.1132. IR (KBr, cmꢀ1): 3460.1, 2983.5, 2231.5, 1610.7, 1493.3,
1376.5, 1294.8, 1102.8.
d6):
d
8.53 (s, 1H, AreH), 8.45 (d, 1H, J ¼ 8.7 Hz, AreH), 7.45 (d, 1H,
J ¼ 8.9 Hz, AreH), 4.21 (t, 2H, J ¼ 5.9 Hz, CH2), 2.50 (s, 3H, CH3), 1.78
(m, 2H, CH2), 1.46 (m, 2H, CH2), 1.34 (m, 4H, 2CH2), 0.89 (s, 3H, CH3).
13C NMR (150 MHz, DMSO-d6):
d 160.92, 160.31, 142.94, 139.80,
132.40, 131.08, 120.54, 117.28, 115.54, 113.61, 100.98, 69.34, 31.22,
28.34, 25.23, 22.03, 13.92, 10.92. ESI-HRMS calcd. for C18H22N3O4
[M þ H]þ 344.1605, found: 344.1596. IR (KBr, cmꢀ1): 3485.4, 2928.1,
2229.8, 1610.8, 1502.3, 1374.0, 1302.6.
4.1.5.3. 2-(3-Cyano-4-iso-propoxyphenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4c). A white crystalline powder, yield
64.6%. HPLC: 96.25%. mp 190.8e191.2 ꢁC. 1H NMR (300 MHz,
DMSO-d6):
d 13.65 (s, 1H, COOH), 8.54 (s, 1H, AreH), 8.48 (d, 1H,
J ¼ 8.7 Hz, AreH), 7.52 (d, 1H, J ¼ 8.8 Hz, AreH), 4.91 (m, 1H, CH),
2.51 (s, 3H, CH3), 1.37 (d, 6H, J ¼ 5.9 Hz, 2CH3). 13C NMR (150 MHz,
4.1.5.9. 2-(3-Cyano-4-heptyloxyphenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4i). A white crystalline powder, yield
70.8%. HPLC: 94.12%. mp 194.3e194.9 ꢁC. 1H NMR (600 MHz, DMSO-
DMSO-d6):
d 160.37, 160.09, 142.95, 139.82, 132.48, 131.44, 117.28,
116.43, 115.76, 114.54, 101.68, 72.10, 21.55, 10.99. ESI-HRMS calcd.
for C15H16N3O4 [M þ H]þ 302.1135, found: 302.1139. IR (KBr, cmꢀ1):
3433.3, 2983.3, 2231.4, 1610.4, 1490.8, 1376.2, 1293.2, 1103.8.
d6):
d
13.75 (s, 1H, COOH), 8.55 (s, 1H, AreH), 8.48 (d, 1H, J ¼ 8.9 Hz,
AreH), 7.47 (d, 1H, J ¼ 9.1 Hz, AreH), 4.22 (t, 2H, J ¼ 6.4 Hz, CH2),
2.51 (s, 3H, CH3), 1.78 (m, 2H, CH2), 1.45 (m, 2H, CH2), 1.35 (m, 2H,
CH2), 1.29 (m, 4H, 2CH2), 0.88 (t, 3H, J ¼ 6.6 Hz, CH3). 13C NMR
4.1.5.4. 2-(4-Butoxy-3-cyanophenyl)-1-hydroxy-4-methyl-1H-imid-
azole-5-carboxylic acid (4d). A white crystalline powder, yield
68.7%. HPLC: 98.15%. mp 193.7e195.3 ꢁC. 1H NMR (300 MHz, DMSO-
(150 MHz, DMSO-d6):
d 160.90, 159.82, 142.99, 139.77, 133.74,
132.04, 120.55, 116.45, 115.74, 113.56, 101.23, 69.17, 31.26, 28.38,
28.32, 25.28, 22.07, 15.76, 13.93. ESI-HRMS calcd. for C19H24N3O4
[M þ H]þ 358.1761, found: 358.1758. IR (KBr, cmꢀ1): 3432.3, 2929.1,
2230.8, 1610.4, 1494.5, 1373.9, 1293.8.
d6):
d
8.56 (s, 1H, AreH), 8.49 (d, 1H, J ¼ 7.0 Hz, AreH), 7.45 (d, 1H,
J ¼ 8.9 Hz, AreH), 4.22 (t, 2H, J ¼ 6.2 Hz, CH2), 2.51 (s, 3H, CH3), 1.77
(m, 2H, CH2), 1.48 (m, 2H, CH2), 0.97 (t, 3H, J ¼ 7.3 Hz, CH3). 13C NMR
(150 MHz, DMSO-d6): d 160.94, 160.32, 142.88,139.81, 132.44, 131.11,
117.27, 116.65, 115.56, 113.63, 100.97, 69.06, 30.32, 18.55, 13.62,
10.95. ESI-HRMS calcd. for C16H18N3O4 [M þ H]þ 316.1292, found:
316.1291. IR (KBr, cmꢀ1): 3430.4, 2959.0, 2938.1, 2229.9, 1610.3,
1294.1.
4.1.5.10. 2-(3-Cyano-4-octyloxyphenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4j). A white crystalline powder, yield
70.8%. HPLC: 97.38%. mp 186.5e187.5 ꢁC. 1H NMR (300 MHz, DMSO-
d6):
d
13.76 (s, 1H, COOH), 8.57 (s, 1H, AreH), 8.51 (d, 1H, J ¼ 9.2 Hz,
4.1.5.5. 2-(4-sec-Butoxy-3-cyanophenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4e). A white crystalline powder, yield
65.6%. HPLC: 96.17%. mp 190.9e191.5 ꢁC. 1H NMR (300 MHz, DMSO-
AreH), 7.50 (d, 1H, J ¼ 9.1 Hz, AreH), 4.23 (t, 2H, J ¼ 6.4 Hz, CH2),
2.52 (s, 3H, CH3), 1.78 (m, 2H, CH2), 1.44 (m, 2H, CH2), 1.41e1.27 (m,
8H, 4CH2), 0.86 (t, 3H, J ¼ 6.4 Hz, CH3). 13C NMR (150 MHz, DMSO-
d6):
d
8.55 (s, 1H, AreH), 8.48 (d, 1H, J ¼ 8.8 Hz, AreH), 7.52 (d, 1H,
d6):
d 160.96, 160.38, 142.95, 139.85, 132.52, 131.19, 117.30, 116.69,
J ¼ 9.1 Hz, AreH), 4.72 (m, 1H, CH), 2.51 (s, 3H, CH3), 1.71 (m, 2H,
115.57, 113.68, 101.02, 69.35, 31.21, 28.65, 28.62, 28.24, 25.26, 22.09,
13.95, 10.99. ESI-HRMS calcd. for C20H26N3O4 [M þ H]þ 372.1918,
found: 372.1914. IR (KBr, cmꢀ1): 3430.8, 2927.0, 2855.9, 2230.3,
1610.5, 1495.2, 1374.3, 1294.2.
CH2), 1.33 (d, 3H, J ¼ 5.9 Hz, CH3), 0.97 (t, 3H, J ¼ 7.3 Hz, CH3). 13
C
NMR (150 MHz, DMSO-d6): d 160.38, 159.75, 142.95, 139.84, 132.45,
131.40, 120.54, 117.27, 115.71, 114.53, 101.69, 76.71, 28.36, 18.76,
10.98, 9.20. ESI-HRMS calcd. for C16H18N3O4 [M þ H]þ 316.1292,
found: 316.1292. IR (KBr, cmꢀ1): 3432.1, 2959.9, 2934.0, 2230.7,
1610.5, 1498.4, 1383.5, 1294.1.
4.1.5.11. 2-[3-Cyano-4-(4-methylbenzyloxy)phenyl]-1-hydroxy-4-
methyl-1H-imidazole-5- carboxylic acid (4k). A white crystalline
powder, yield 71.9%. HPLC: 99.14%. mp 199.2e199.8 ꢁC. 1H NMR
4.1.5.6. 2-(4-iso-Butoxy-3-cyanophenyl)-1-hydroxy-4-methyl-1H-
imidazole-5-carboxylic acid (4f). A white crystalline powder, yield
65.2%. HPLC: 95.24%. mp 191.6e192.2 ꢁC. 1H NMR (300 MHz, DMSO-
(300 MHz, DMSO-d6): d 13.81 (s, 1H, COOH), 8.58 (s, 1H, AreH), 8.51
(d, 1H, J ¼ 8.9 Hz, AreH), 7.60 (d, 1H, J ¼ 9.1 Hz, AreH), 7.39 (d, 2H,
J ¼ 8.0 Hz, AreH), 7.24 (d, 2H, J ¼ 7.9 Hz, AreH), 5.33 (s, 2H, CH2),
2.51 (s, 3H, CH3), 2.32 (s, 3H, CH3). 13C NMR (150 MHz, DMSO-d6):
d6):
d
8.55 (s, 1H, AreH), 8.47 (d, 1H, J ¼ 8.8 Hz, AreH), 7.46 (d, 1H,
J ¼ 9.0 Hz, AreH), 4.00 (d, 2H, J ¼ 6.4 Hz, CH2), 2.51 (s, 3H, CH3), 2.10
(m, 1H, CH), 1.03 (d, 6H, J ¼ 6.7 Hz, 2CH3). 13C NMR (150 MHz,
d
160.63, 160.36, 140.45, 139.39, 137.74, 132.52, 132.48, 131.33,
DMSO-d6):
d
160.47, 159.02, 141.95, 140.75, 133.91, 132.09, 121.34,
129.18, 127.96, 117.31, 116.95, 115.61, 114.23, 101.31, 70.63, 20.80,
13.87. ESI-HRMS calcd. for C20H18N3O4 [MþH]þ 364.1296, found:
364.1295. IR (KBr, cmꢀ1): 3433.3, 3052.7, 2923.5, 2230.5, 1610.0,
1494.3, 1375.8, 1279.3.
117.94, 115.92, 113.30, 100.71, 67.65, 24.62, 22.37, 15.71. ESI-HRMS
calcd. for C16H18N3O4 [MþH]þ 316.1292, found: 316.1291. IR (KBr,
cmꢀ1): 3219.9, 2957.5, 2229.6, 1711.5, 1469.7, 1293.8, 1104.3.

S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
351
4.1.6. General procedure for the synthesis of ethyl 2-(4-alkoxy-3-
cyanophenyl)-1-methoxy-4-methyl-1H-imidazole-5-carboxylates
5ae5k
z 358.4 [M þ H]þ.
4.1.6.7. Ethyl 2-(3-cyano-4-iso-pentyloxyphenyl)-1-methoxy-4-
methyl-1H-imidazole-5- carboxylate (5g). A white crystalline pow-
der, yield 73.7%, mp 138.1e139.6 ꢁC. 1H NMR (300 MHz, DMSO-d6):
Dimethyl sulfate (53 mmol) was added dropwise at a rate that
maintained <0 ꢁC reaction temperature to a stirred mixture of ethyl
2-(4-alkoxy-3-cyanophenyl)-1-hydroxy-4-methyl-1H- imidazole-
5-carboxylate (50 mmol), anhydrous potassium carbonate
(75 mmol) and DMF (40 mL). Upon completion of addition, the
mixture was stirred at 25 ꢁC for another 1.5 h, and then the reaction
mixture was slowly poured into cold water (100 mL). The resulting
precipitate was filtered, dried, and recrystallized with ethyl acetate
to yield the corresponding ethyl 2-(4-alkoxy- 3-cyanophenyl)-1-
methoxy-4-methyl-1H-imidazole-5-carboxylates 5ae5k.
d
8.27 (dd, 1H, J ¼ 2.1 &8.8 Hz, AreH), 8.24 (d, 1H, J ¼ 2.0 Hz, AreH),
7.45 (d, 1H, J ¼ 8.9 Hz, AreH), 4.33 (q, 2H, J ¼ 7.1 Hz, CH2), 4.26 (t,
2H, J ¼ 6.6 Hz, CH2), 3.96 (s, 3H, CH3), 2.40 (s, 3H, CH3), 1.82 (m, 1H,
CH), 1.69 (m, 2H, CH2), 1.33 (t, 3H, J ¼ 7.1 Hz, CH3), 0.96 (d, 6H,
J ¼ 6.6 Hz, 2CH3). ESI-MS m/z 372.4 [M þ H]þ.
4.1.6.8. Ethyl 2-(3-cyano-4-hexyloxyphenyl)-1-methoxy-4-methyl-
1H-imidazole-5-carboxylate (5h). A white crystalline powder, yield
71.8%, mp 118.7e120.6 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.26 (dd,
4.1.6.1. Ethyl 2-(3-cyano-4-methoxyphenyl)-1-methoxy-4-methyl-
1H, J ¼ 2.3 & 8.7 Hz, AreH), 8.24 (d, 1H, J ¼ 2.0 Hz, AreH), 7.42 (d,
1H, J ¼ 8.9 Hz, AreH), 4.32 (q, 2H, J ¼ 7.1 Hz, CH2), 3.96 (s, 3H, CH3),
2.40 (s, 3H, CH3), 1.78 (m, 2H, CH2), 1.49e1.31 (m, 9H, 3CH2 & CH3),
0.89 (t, 3H, J ¼ 6.8 Hz, CH3). ESI-MS m/z 386.5 [M þ H]þ.
1H-imidazole-5-carboxylate (5a). A white crystalline powder, yield
84.7%, mp 135.1e137.4 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.27 (dd,
1H, J ¼ 2.3 & 8.8 Hz, AreH), 8.24 (d, 1H, J ¼ 1.9 Hz, AreH), 7.41 (d,
1H, J ¼ 8.9 Hz, AreH), 4.31 (m, 2H, J ¼ 7.2 Hz, CH2), 3.96 (s, 3H, CH3),
3.88 (s, 3H, CH3), 2.40 (s, 3H, CH3), 1.33 (t, 3H, CH3). ESI-MS m/z
316.4 [M þ H]þ.
4.1.6.9. Ethyl 2-(3-cyano-4-heptyloxyphenyl)-1-methoxy-4-methyl-
1H-imidazole-5-carboxylate (5i). A white crystalline powder, yield
76.8%, mp 166.2e166.6 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.26
4.1.6.2. Ethyl
1H-imidazole-5-carboxylate (5b). A white crystalline powder, yield
78.8%, mp 143.2e144.5 ꢁC. 1H NMR (300 MHz, DMSO-d6):
8.27
(dd, 1H, J ¼ 2.3 & 8.7 Hz, AreH), 8.24 (s, 1H, AreH), 7.42 (d, 1H,
J ¼ 8.8 Hz, AreH), 4.33 (q, 2H, J ¼ 7.2 Hz, CH2), 4.19 (t, 2H, J ¼ 6.4 Hz,
CH2), 3.96 (s, 3H, CH3), 2.40 (s, 3H, CH3), 1.80 (m, 2H, CH2), 1.33 (t,
3H, J ¼ 7.1 Hz, CH3), 1.03 (t, 3H, J ¼ 7.3 Hz, CH3). ESI-MS m/z 344.4
[M þ H]þ.
2-(3-cyano-4-propoxyphenyl)-1-methoxy-4-methyl-
(dd, 1H, J ¼ 2.3 & 8.7 Hz, AreH), 8.24 (d, 1H, J ¼ 2.0 Hz, AreH), 7.42
(d, 1H, J ¼ 8.8 Hz, AreH), 4.32 (q, 2H, J ¼ 7.1 Hz, CH2), 4.22 (t, 2H,
J ¼ 6.4 Hz, CH2), 3.96 (s, 3H, CH3), 2.34 (s, 3H, CH3), 1.78 (m, 2H,
CH2), 1.48e1.28 (m, 11H, 4CH2 & CH3), 0.87 (t, 3H, J ¼ 6.7 Hz, CH3).
ESI-MS m/z 400.5 [M þ H]þ.
d
4.1.6.10. Ethyl 2-(3-cyano-4-octyloxyphenyl)-1-methoxy-4-methyl-
1H-imidazole-5-carboxylate (5j). A white crystalline powder, yield
78.1%, mp 165.8e167.3 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.27 (dd,
4.1.6.3. Ethyl 2-(3-cyano-4-iso-propoxyphenyl)-1-methoxy-4-
methyl-1H-imidazole-5- carboxylate (5c). A white crystalline pow-
der, yield 80.8%, mp 170.4e172.8 ꢁC. 1H NMR (300 MHz, DMSO-d6):
1H, J ¼ 2.0 & 8.8 Hz, AreH), 8.24 (d, 1H, J ¼ 2.0 Hz, AreH), 7.45 (d,
1H, J ¼ 8.9 Hz, AreH), 4.32 (q, 2H, J ¼ 7.1 Hz, CH2), 4.22 (t, 2H,
J ¼ 6.4 Hz, CH2), 3.96 (s, 3H, CH3), 2.33 (s, 3H, CH3), 1.79 (m, 2H,
CH2), 1.45e1.20 (m, 13H, 5CH2 & CH3), 0.90 (t, 3H, J ¼ 6.7 Hz, CH3).
ESI-MS m/z 414.5 [M þ H]þ.
d
8.27 (dd,1H, J ¼ 2.3 & 8.5 Hz, AreH), 8.24 (d, 1H, J ¼ 2.3 Hz, AreH),
7.45 (d, 1H, J ¼ 8.6 Hz, AreH), 4.90 (m, 1H, CH), 4.32 (q, 2H,
J ¼ 7.1 Hz, CH2), 3.96 (s, 3H, CH3), 2.40 (s, 3H, CH3), 1.36 (d, 6H,
J ¼ 5.9 Hz, 2CH3), 1.31 (t, 3H, J ¼ 7.1 Hz, CH3). ESI-MS m/z 344.4
[M þ H]þ.
4.1.6.11. Ethyl 2-[3-cyano-4-(4-methylbenzyloxy)phenyl]-1-
methoxy-4-methyl-1H-imidazole-5-carboxylate (5k). A white crys-
talline powder, yield 70.1%, mp 147.5e149.5 ꢁC. 1H NMR (300 MHz,
4.1.6.4. Ethyl 2-(4-butoxy-3-cyanophenyl)-1-methoxy-4-methyl-1H-
DMSO-d6):
d
8.27 (dd,1H, J ¼ 2.1 &7.2 Hz, AreH), 8.26 (s, 1H, AreH),
imidazole-5-carboxylate (5d). A white crystalline powder, yield
7.51 (d, 1H, J ¼ 7.2 Hz, AreH), 7.39 (d, 2H, J ¼ 8.1 Hz, AreH), 7.24 (d,
2H, J ¼ 7.8 Hz, AreH), 5.32 (s, 2H, CH2), 4.31 (q, 2H, J ¼ 7.2 Hz, CH2),
3.96 (s, 3H, CH3), 2.40 (s, 3H, CH3), 2.32 (s, 3H, CH3), 1.33 (t, 3H,
J ¼ 7.2 Hz, CH3). ESI-MS m/z 405.5 [M þ H]þ.
86.5%, mp 135.4e136.2 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.26
(dd, 1H, J ¼ 2.3 & 8.8 Hz, AreH), 8.24 (d, 1H, J ¼ 1.9 Hz, AreH), 7.42
(d, 1H, J ¼ 8.9 Hz, AreH), 4.32 (q, 2H, J ¼ 7.1 Hz, CH2), 4.23 (t, 2H,
J ¼ 6.4 Hz, CH2), 3.96 (s, 3H, CH3), 2.34 (s, 3H, CH3), 1.77 (m, 2H,
CH2), 1.48 (m, 2H, CH2), 1.33 (t, 3H, J ¼ 7.1 Hz, CH3), 0.96 (t, 3H,
J ¼ 7.3 Hz, CH3). ESI-MS m/z 358.4 [M þ H]þ.
4.1.7. General procedure for the synthesis of 2-(4-alkoxy-3-
cyanophenyl)-1-methoxy-4 -methyl-1H-imidazole-5-carboxylic
acids 6ae6k
4.1.6.5. Ethyl 2-(4-sec-butoxy-3-cyanophenyl)-1-methoxy-4-methyl-
A mixture of ethyl 2-(4-alkoxy -3-cyanophenyl)-1-methoxy-4-
1H-imidazole-5-carboxylate (5e). A white crystalline powder, yield
methyl-1H-imidazole-5- carboxylate (2.8 mmol), 1 M NaOH
88.5%, mp 122.9e123.1 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d
8.26
aqueous (7 mL), THF (5 mL), and ethanol (5 mL) was stirred at 50 ꢁC
for 2 h. The solvent was concentrated in vacuum and the residue
was acidified with dilute hydrochloric acid to pH 1, The resulting
precipitate was filtered, dried, and recrystallized with mixture of
methanol and ethyl acetate (1:2) to yield the corresponding 2-(4-
(dd, 1H, J ¼ 2.1 & 8.8 Hz, AreH), 8.24 (s, 1H, AreH), 7.51 (d, 1H,
J ¼ 9.0 Hz, AreH), 4.69 (m, 1H, CH), 4.31 (q, 2H, J ¼ 6.9 Hz, CH2), 3.96
(s, 3H, CH3), 2.40 (s, 3H, CH3), 1.70 (m, 2H, CH2), 1.34 (t, 3H,
J ¼ 6.9 Hz, CH3), 1.31 (d, 3H, J ¼ 6.0 Hz, CH3), 0.96 (t, 3H, J ¼ 7.2 Hz,
CH3). ESI-MS m/z 358.4 [M þ H]þ.
alkoxy-
3-cyanophenyl)-1-methoxy-4-methyl-1H-imidazole-5-
carboxylic acids 6ae6k.
4.1.6.6. Ethyl 2-(4-iso-butoxy-3-cyanophenyl)-1-methoxy-4-methyl-
1H-imidazole-5-carboxylate (5f). A white crystalline powder, yield
4.1.7.1. 2-(3-Cyano-4-methoxyphenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6a). A white crystalline powder, yield
50.4%. HPLC: 98.64%. mp 185.4e186.3 ꢁC. 1H NMR (300 MHz,
92.9%, mp 123.8e124.9 ꢁC. 1H NMR (300 MHz, DMSO-d6):
d 8.30 (d,
1H, J ¼ 2.2 Hz, AreH), 8.28 (dd, 1H, J ¼ 2.2 & 9.6 Hz, AreH), 7.39 (d,
1H, J ¼ 9.6 Hz, AreH), 4.31 (q, 2H, J ¼ 6.9 Hz, CH2), 4.01 (d, 2H,
J ¼ 6.4 Hz, CH2), 3.96 (s, 3H, CH3), 2.39 (s, 3H, CH3), 2.10 (m, 1H, CH),
1.34 (t, 3H, J ¼ 6.9 Hz, CH3), 1.03 (d, 6H, J ¼ 6.7 Hz, 2CH3). ESI-MS m/
DMSO-d6):
d
13.01 (s, 1H, COOH), 8.26 (dd, 1H, J ¼ 2.0 & 9.0 Hz,
AreH), 8.24 (s, 1H, AreH), 7.40 (d, 1H, J ¼ 8.9 Hz, AreH), 3.95 (s, 3H,
CH3), 3.88 (s, 3H, CH3),2.40 (s, 3H, CH3). 13C NMR (100 MHz, DMSO-

352
S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
d6):
d
160.80, 159.79, 142.98, 139.84, 133.85, 132.16, 120.53, 116.43,
4.1.7.7. 2-(3-Cyano-4-iso-pentyloxyphenyl)-1-methoxy-4-methyl-
1H-imidazole-5-carboxylic acid (6g). A white crystalline powder,
yield 63.0%. HPLC: 97.24%. mp 191.3e191.9 ꢁC. 1H NMR (300 MHz,
115.85, 113.62, 101.16, 70.70, 66.84, 21.72. ESI-HRMS calcd. for
C
14H14N3O4 [M þ H]þ 288.2710, found: 288.2715. IR (KBr, cmꢀ1):
3393.9, 2966.3, 2223.5, 1704.1, 1611.2, 1527.8, 1443.2, 1287.0, 1262.5,
1100.6.
DMSO-d6):
d
13.06 (s, 1H, COOH), 8.27 (dd, 1H, J ¼ 2.2 & 8.9 Hz,
AreH), 8.24 (d, 1H, J ¼ 2.0 Hz, AreH), 7.44 (d, 1H, J ¼ 8.9 Hz, AreH),
4.25 (t, 2H, J ¼ 6.5 Hz, CH2), 3.95 (s, 3H, CH3), 2.40 (s, 3H, CH3), 1.86
(m, 1H, CH), 1.70 (m, 2H, CH2), 0.96 (d, 6H, J ¼ 6.6 Hz, 2CH3). 13C
4.1.7.2. 2-(3-Cyano-4-propoxyphenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6b). A white crystalline powder, yield
63.0%. HPLC: 97.42%. mp 193.1e194.0 ꢁC. 1H NMR (300 MHz, DMSO-
NMR (150 MHz, DMSO-d6):
d 160.90, 159.77, 142.94, 139.80, 133.77,
132.05, 120.54, 116.44, 115.71, 113.62, 101.21, 69.17, 66.79, 30.82,
28.21, 24.91, 22.02. ESI-HRMS calcd. for C18H22N3O4 [M þ H]þ
344.1605, found: 344.1603. IR (KBr, cmꢀ1): 3383.3, 2956.4, 2924.0,
2231.0, 1702.2, 1606.7, 1461.1, 1397.6.
d6):
d
13.06 (s, 1H, COOH), 8.28 (dd, 1H, J ¼ 2.1 & 9.8 Hz, AreH), 8.24
(s, 1H, AreH), 7.41 (d, 1H, J ¼ 8.9 Hz, AreH), 4.19 (t, 2H, J ¼ 6.4 Hz,
CH2), 3.95 (s, 3H, CH3), 2.39 (s, 3H, CH3), 1.79 (m, 2H, CH2), 1.03 (t,
3H, J ¼ 7.3 Hz, CH3). 13C NMR (150 MHz, DMSO-d6):
d 160.92, 159.77,
142.95, 139.81, 133.79, 132.07, 120.54, 116.44, 115.73, 113.63, 101.20,
68.89, 66.80, 18.55, 15.77, 13.59. ESI-HRMS calcd. for C16H18N3O4
[M þ H]þ 316.1292, found: 316.1297. IR (KBr, cmꢀ1): 3442.5, 2980.4,
2936.9, 2231.5, 1661.8, 1464.4, 1297.6, 1108.6.
4.1.7.8. 2-(3-Cyano-4-hexyloxyphenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6h). A white crystalline powder, yield
64.5%. HPLC: 98.26%. mp 192.8e193.9 ꢁC. 1H NMR (300 MHz,
DMSO-d6):
d
13.02 (s, 1H, COOH), 8.26 (dd, 1H, J ¼ 2.2 & 9.0 Hz,
AreH), 8.23 (d, 1H, J ¼ 2.0 Hz, AreH), 7.41 (d, 1H, J ¼ 8.9 Hz, AreH),
4.22 (t, 2H, J ¼ 6.4 Hz, CH2), 3.95 (s, 3H, CH3), 2.39 (s, 3H, CH3), 1.78
(m, 2H, CH2), 1.46 (m, 2H, CH2), 1.32 (m, 4H, 2CH2), 0.89 (t, 3H,
4.1.7.3. 2-(3-Cyano-4-iso-propoxyphenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6c). A white crystalline powder, yield
65.2%. HPLC: 96.98%. mp 189.0e189.3 ꢁC. 1H NMR (300 MHz,
J ¼ 6.8 Hz, CH3). 13C NMR (150 MHz, DMSO-d6):
d 160.90, 159.77,
DMSO-d6):
d
13.06 (s, 1H, COOH), 8.27 (dd, 1H, J ¼ 2.2 & 9.1 Hz,
142.94, 139.80, 133.76, 132.05, 120.54, 116.44, 115.71, 113.62, 101.21,
69.17, 66.78, 31.19, 28.29, 25.22, 21.99, 15.72, 13.89. ESI-HRMS calcd.
for C19H24N3O4 [M þ H]þ 358.1761, found: 358.1757. IR (KBr, cmꢀ1):
3435.2, 2955.2, 2934.3, 2229.4, 1663.7, 1612.5, 1465.9, 1296.9.
AreH), 8.24 (s,1H, AreH), 7.40 (d,1H, J ¼ 9.0 Hz, AreH), 4.86 (m,1H,
CH), 3.93 (s, 3H, CH3), 2.39 (s, 3H, CH3), 1.37 (d, 6H, J ¼ 6.0 Hz, 2CH3).
13C NMR (150 MHz, DMSO-d6):
d 160.01, 159.78, 142.95, 139.82,
133.70, 132.25, 120.37, 116.43, 115.90, 114.53, 101.89, 71.92, 66.82,
21.57, 15.73. ESI-HRMS calcd. for C16H18N3O4 [M þ H]þ 316.1292,
found: 316.1295. IR (KBr, cmꢀ1): 3455.2, 2957.0, 2223.0, 1704.8,
1611.5, 1443.0, 1287.4, 1100.9.
4.1.7.9. 2-(3-Cyano-4-heptyloxyphenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6i). A white crystalline powder, yield
63.4%. HPLC: 98.10%. mp 193.9e194.2 ꢁC. 1H NMR (600 MHz, DMSO-
d6):
d
13.05 (s, 1H, COOH), 8.26 (dd, 1H, J ¼ 2.2 & 9.0 Hz, AreH), 8.23
4.1.7.4. 2-(4-Butoxy-3-cyanophenyl)-1-methoxy-4-methyl-1H-imid-
azole-5-carboxylic acid (6d). A white crystalline powder, yield
72.8%. HPLC: 95.96%. mp 193.1e194.0 ꢁC. 1H NMR (300 MHz, DMSO-
(d, 1H, J ¼ 2.2 Hz, AreH), 7.41 (d, 1H, J ¼ 9.0 Hz, AreH), 4.21 (t, 2H,
J ¼ 6.4 Hz, CH2), 3.95 (s, 3H, CH3), 2.40 (s, 3H, CH3), 1.79 (m, 2H,
CH2),1.45 (m, 2H, CH2),1.36 (m, 2H, CH2),1.28 (m, 4H, 2CH2), 0.87 (t,
d6):
d
8.28 (s, 1H, AreH), 8.24 (s, 1H, AreH), 7.41 (d, 1H, J ¼ 8.9 Hz,
3H, J ¼ 6.7 Hz, CH3). 13C NMR (100 MHz, DMSO-d6):
d 160.94,
AreH), 4.23 (t, 2H, J ¼ 6.3 Hz, CH2), 3.95 (s, 3H, CH3), 2.39 (s, 3H,
159.80, 142.98, 139.84, 133.83, 132.09, 120.54, 116.44, 115.76, 113.67,
101.21, 69.18, 66.83, 31.23, 28.32, 28.28, 25.25, 22.03, 15.76, 13.95.
ESI-HRMS calcd. for C20H26N3O4 [M þ H]þ 372.1918, found:
372.1914. IR (KBr, cmꢀ1): 3439.8, 2931.0, 2229.8, 1663.8, 1613.5,
1466.3, 1298.2, 1108.6.
CH3), 1.78 (m, 2H, CH2), 1.50 (m, 2H, CH2), 0.96 (t, 3H, J ¼ 7.3 Hz,
CH3). 13C NMR (150 MHz, DMSO-d6):
d 160.31, 159.77, 142.95,
139.84, 133.71, 132.24, 120.37, 116.42, 115.85, 114.53, 101.88, 76.57,
66.82, 28.38, 18.80, 15.73, 9.20. ESI-HRMS calcd. for C17H20N3O4
[M þ H]þ 330.1448, found: 330.1447. IR (KBr, cmꢀ1): 3429.4, 2959.0,
2938.1, 2229.9, 1664.5, 1613.3, 1465.6, 1295.8, 1108.4.
4.1.7.10. 2-(3-Cyano-4-octyloxyphenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6j). A white crystalline powder, yield
66.7%. HPLC: 99.17%. mp 188.6e189.8 ꢁC. 1H NMR (300 MHz, DMSO-
4.1.7.5. 2-(4-sec-Butoxy-3-cyanophenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6e). A white crystalline powder, yield
67.4%. HPLC: 98.55%. mp 181.5e182.3 ꢁC. 1H NMR (300 MHz, DMSO-
d6):
d
13.06 (s, 1H, COOH), 8.26 (dd, 1H, J ¼ 2.2 & 8.9 Hz, AreH), 8.23
(d, 1H, J ¼ 2.0 Hz, AreH), 7.41 (d, 1H, J ¼ 8.9 Hz,AreH), 4.21 (t, 2H,
J ¼ 6.4 Hz, CH2), 3.95 (s, 3H, CH3), 2.39 (s, 3H, CH3), 1.78 (m, 2H,
CH2),1.45e1.27 (m,10H, 5CH2), 0.86 (t, 3H, J ¼ 6.3 Hz, CH3). 13C NMR
d6):
d 13.06 (s, 1H, COOH), 8.27 (s, 1H, AreH), 8.24 (s, 1H, AreH),
7.44 (d, 1H, J ¼ 8.8 Hz, AreH), 4.69 (m, 1H, CH), 3.97 (s, 3H, CH3),
2.39 (s, 3H, CH3), 1.67 (m, 2H, CH2), 1.33 (d, 3H, J ¼ 5.9 Hz, CH3), 0.97
(100 MHz, DMSO-d6):
d 160.94, 159.78, 142.95, 139.85, 133.85,
(t, 3H, J ¼ 7.3 Hz, CH3). 13C NMR (150 MHz, DMSO-d6):
d
160.91,
132.10, 120.54, 116.45, 115.75, 113.70, 101.20, 69.18, 66.83, 31.18,
28.62, 28.57, 28.24, 25.25, 22.07, 15.75, 13.95. ESI-HRMS calcd. for
159.75, 142.95, 139.84, 133.80, 132.08, 120.54, 116.42, 115.73, 113.66,
101.19, 67.74, 66.81, 36.95, 24.60, 22.35, 15.72. ESI-HRMS calcd. for
C
21H28N3O4 [M þ H]þ 386.2704, found: 386.2706. IR (KBr, cmꢀ1):
C
17H20N3O4 [M þ H]þ 330.1448, found: 330.1449. IR (KBr, cmꢀ1):
3432.3, 2929.0, 2229.6, 1610.4, 1464.5, 1293.8.
3440.0, 2954.8, 2229.6, 1663.0, 1465.6, 1296.1, 1107.8.
4.1.7.11. 2-[3-Cyano-4-(4-methylbenzyloxy)phenyl]-1-methoxy-4-
methyl-1H-imidazole-5- carboxylic acid (6k). A white crystalline
powder, yield 58.1%. HPLC: 96.75%. mp 189.0e190.1 ꢁC. 1H NMR
4.1.7.6. 2-(4-iso-Butoxy-3-cyanophenyl)-1-methoxy-4-methyl-1H-
imidazole-5-carboxylic acid (6f). A white crystalline powder, yield
69.6%. HPLC: 99.10%. mp 190.8e191.3 ꢁC. 1H NMR (300 MHz, DMSO-
(300 MHz, DMSO-d6):
d
8.31 (s, 1H, AreH), 8.30 (dd, 1H, J ¼ 2.2 &
d6):
d
13.06 (s, 1H, COOH), 8.27 (dd, 1H, J ¼ 2.2 & 8.6 Hz, AreH), 8.24
8.1 Hz, AreH), 7.55 (d, 1H, J ¼ 9.8 Hz, AreH), 7.40 (d, 2H, J ¼ 8.0 Hz,
AreH), 7.25 (d, 2H, J ¼ 7.9 Hz, AreH), 5.34 (s, 2H, CH2), 3.98 (s, 3H,
CH3), 2.50 (s, 3H, CH3), 2.32 (s, 3H, CH3). 13C NMR (100 MHz, DMSO-
(s, 1H, AreH), 7.41 (d, 1H, J ¼ 8.8 Hz, AreH), 4.01 (d, 2H, J ¼ 6.4 Hz,
CH2), 3.96 (s, 3H, CH3), 2.39 (s, 3H, CH3), 2.10 (m, 1H, CH), 1.03 (d,
6H, J ¼ 6.7Hz, 2CH3). 13C NMR (150 MHz, DMSO-d6):
d
160.97,
d6): d 161.26, 159.22, 140.45, 139.39, 137.77, 134.39, 133.03, 132.54,
159.79, 142.88, 139.81, 133.84, 132.02, 120.59, 116.51, 115.67, 113.73,
101.22, 74.97, 66.80, 27.57, 18.70, 15.71. ESI-HRMS calcd. for
129.22, 127.96, 118.46, 116.91, 115.59, 114.34, 101.62, 70.68, 67.37,
20.83, 14.34. ESI-HRMS calcd. for C21H20N3O4 [MþH]þ 378.1448,
found: 378.1446. IR (KBr, cmꢀ1): 3405.1, 2659.0, 2235.6, 1719.9,
1609.7, 1499.7, 1456.6, 1306.3, 1288.3, 1266.7, 1115.3.
C
17H20N3O4 [M þ H]þ 330.1448, found: 330.1445. IR (KBr, cmꢀ1):
3440.3, 2969.6, 2932.7, 2230.8, 1663.6, 1460.3, 1288.0, 1107.2.

S. Chen et al. / European Journal of Medicinal Chemistry 103 (2015) 343e353
(2001) 879e882.
353
4.2. Assay of the in vitro XO inhibitory potency
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Acknowledgments
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This work was supported by Program for Liaoning Excellent
Talents in University (LR2012035) and the National Natural Science
Foundation of China (J1210029). The authors are grateful to Wen Li,
Aihua Song, and Dr. Ying Peng (Analytical Department of Shenyang
Pharmaceutical University, Shenyang, China) for assistance in
gathering 1H NMR, 13C NMR, IR, and ESI-HRMS spectroscopy data.
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