Design, syntheses and antitumor activities evaluation of 1,5-diaryl substituted pyrazole secnidazole ester derivatives

Article abstract of DOI:10.1002/jhet.4302

According to the drug hybridization principle, a series of novel 1,5-diaryl substituted pyrazole secnidazole ester derivatives (6aa–6gc) have been synthesized by the combinations of various 1,5-diarylpyrazole-3-carboxylic acids with secnidazole. The in vi

Full text of DOI:10.1002/jhet.4302

Design, syntheses and antitumor activities evaluation of 1,5-
diaryl substituted pyrazole secnidazole ester derivatives
Qing-Hu Teng, Gui-Xia Sun, Shu-Ying Luo, Kai Wang*, Fu-Pei Liang*
Guangxi Key Laboratory of Electrochemical and Magnetochemical Functional Materials, College of Chemistry and
Bioengineering, Guilin University of Technology, Guilin, 541004, China.
*Corresponding author: e-mail: kaiwang2011@yahoo.com, fliangoffice@yahoo.com.
((Additional Supporting Information may be found in the online version of this article.))((Please delete if not appropriate))
which could behavior as the bifunctional conjugates
Abstract. According to the drug hybridization principle, a
with
potent
inhibitory
activity
towards
series of novel 1,5-diaryl substituted pyrazole secnidazole
ester derivatives (6aa-6gc) have been synthesized by the
combinations of various 1,5-diarylpyrazole-3-carboxylic
acids with secnidazole. The in vitro antitumor/ cytotoxicities
activities against tumor and normal cell lines, including
NCI-H460 (lung tumor cell), MCG-803 (gastric tumor cell),
Skov-3 (ovarian tumor cell), BEL-7404 (liver tumor cell)
and HL-7702 (normal liver cell), have been evaluated using
MTT assay. All compounds showed promising inhibitory
activities against four tumor cell lines. The IC50 of 6bc
against the BEL-7404 cell was 2.03 μM, and those of 6fc
against the NCI-H460, MCG-803 and Skov-3 were 1.34,
0.14 and 0.87 μM, respectively. All these values were much
lower than those of the cisplatin. Furthermore, 6fc and 6bc
were also verified to be considerable safe for normal human
liver cell, since the lower IC50 values than cisplatin. Based
on these results, the cell cycle analysis, apoptosis ratio
detection, and mitochondrial membrane potential assay of
6fc and 6bc were further performed aiming to investigate
their inhibition mechanism of BEL-7404 cells. It is revealed
that they have effectively inhibited the cell growth by
arresting the BEL-7404 cells at S phase and induced
apoptosis through the mitochondria-mediated pathway.
cyclooxygenase (COX) and histone deacetylase
(HDAC), could arrest the cell cycle progression of
androgen dependent prostate tumor cell line (LNCaP)
in the S- and G0/G1 phases.11 Several hybridization
of pyrazoles and substituted coumarins/morpholines
groups have also found to be potent dual COX-2/5-
LOX inhibitors and antitumor agents. Their lowest
IC50 have reached to 0.16 μM.12 These results
suggested that more extensive and prominent anti-
tumor activities could be expected when such
pyrazole parents were hybridized with other novel
active groups.
As
a
next-generation of 5-nitroimidazole,
secnidazole has been approved for a single-dose (2 g)
treatment of bacterial vaginosis (BV), as well as
commercially available for more than three decades
in 31 countries.13 Moreover, secnidazole can also
undergo a bioreduction of the nitro group to generate
toxic reactive oxygen species (ROS), which leading
to DNA helix damage, disruption of bacterial protein
synthesis and replication, and ultimately, cell death.14
On this basis, special attention have been paid to its
structural modifications. Several secnidazole
derivatives, including esters and metal-complexes,
have been reported as potential drug in tumor growth
inhibition.15 It should be noted that, its derivatives that
stem from the combination of pyrazoles with
secnidazoles was rarely investigated yet. Up to now,
only series of compounds synthesized form the
coupling reaction of 4-(1-acetyl-3-phenyl-4,5-
Keywords: Drug hybridization principle; 1,5-Diaryl
substituted pyrazole; Secnidazoles; Antitumor activities
Introduction
As one type of the core frameworks in various natural
products
and
medicines,
nitrogen-containing
dihydro-1H-pyrazol-5-yl)benzoic
acid
with
heterocycles have played important roles in drug
design and development.1 The pyrazole is just one of
such species. They have shown extensive
bioactivities,2 such as anti-tumor,3 antimicrobial,4
antifungal,5 hypoglycemic,6 anti-inflammatory,7
analgesic,8 against oxidative stress and diabetes,9 and
secnidazole has been developed. And it is gratifying
that these compounds have been determined to bear
excellent antitumor activities.16
In recent years, our group have been interested in the
various enzyme inhibiting activities,10 etc. In particular, development of new methodologies for the syntheses
promising potentials against the malignant tumor have
been demonstrated in some of modified pyrazole
derivatives when they were grafted with other active
groups according to the drug hybridization principle.
For instances, the methyl sulfone pyrazole derivatives,
of new nitrogen heterocycles molecules, as well as the
searching of novel antitumor targeted drug
candidates.17 Based on these works and combined with
excellent antitumor potentials of the pyrazole and
secnidazole groups (Scheme 1), we have designed and
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which may lead to differences between this version and the Version of Record. Please
cite this article as doi: 10.1002/jhet.4302
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Journal of Heterocyclic Chemistry
Page 2 of 28
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R2
R2
synthesized various 1,5-diaryl substituted pyrazole
3:
O
O
OH
CO2CH3
DMO
CH3ONa
NHNH2
MeOH, HCl, reflux
secnidazole ester derivatives. The antitumor activities
of all obtained compounds have been evaluated using
MTT assay. Several compounds have been found to
bear much higher inhibiting activities against most of
selected tumor cells than the cisplatin. Furthermore,
they were also verified to be much safer for normal
human liver cell than the cisplatin. In order to reveal
their inhibition mechanism, the cell cycle analysis,
apoptosis ratio detection, and mitochondrial
membrane potential assay of selected compounds were
also investigated.
O
N
N
R1
MeOH
reflux
R1
O
R1
4
1
2
R2
R2
N
N
i: EDC, HOBT,
DMAP, DCE, 45 o
OH
O
N
i: KOH, MeOH, reflux
ii: HCl
C
N
O
O
N
N
ii: Secnidazole
O2N
R1
R1
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5
6
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53
54
55
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59
60
Scheme 2. Synthesis route of compounds 6.
The molecular structures and yields (%) of these
targeted compounds were summarized in Table 1. All
targeted compounds were first reported and
N
FG1
=
Ar2
Ar1
O
or
N
1
FG1
structurally characterized by H NMR spectroscopy,
N
X
13C NMR spectroscopy and ESI-MS. Taking 6aa as
the example, its 1H NMR spectrum showed a doublet
of Me proton of propane at N1-secnidazole at δ 1.44
(d, J = 6.1 Hz, 3H), three singlets of Me protons at
para-5-phenyl, para-1-phenyl, and C2 of secnidazole
appeared at δ 2.29 (s, 3H), 2.35 (s, 3H), 2.55 (s, 3H),
respectively. The methylene (CH2) protons of propane
at N1-imidazole showed a multiplet at δ 4.54 (m, 1H),
and a doublet at δ 4.69 (d, J = 14.1 Hz, 1H),
respectively. There was a broad singlet of methylene
(CH) proton of propane at N1-imidazole at δ 5.45 (br,
1H). The protons of parazole and imidazole ring
displayed at δ 6.96 (s, 1H) and 8.02 (s, 1H) as singlets,
respectively. The protons of 1, 5-diphenyl showed a
multiplet at δ 7.11-7.19 (m, 6H), and a doublet at δ
7.26 (d, J = 7.8, 2H). Nearly similar patterns were
noticed in 1H and 13C NMR spectra of all the
compounds (6aa-6gc). The HRMS (ESI) of
compounds (6aa-6gc) showed characteristic [M+H]+
corresponding peaks equivalent to their molecular
formulae. Relevant spectral data is furnished in
supporting information.
O
O
O
N
dual inhibitors of COX-2 and 5-LOX
potent antiproliferative activities
N
O
Ar1
Ar2
O
O
N
N
this work
NO2
Tcm99O
FG2
=
[Au],
higher activities against cancer
N
cells and less toxic to healthy cells
O
N
FG2
O
O2N
Ph
reduction of NO2 generating ROS (NO2•- ) to
damage of DNA and other biomolecules
Scheme 1. Design philosophy of pyrazole secnidazole
derivatives as antitumor agents.
Results and discussion
Chemistry
The syntheses of targeted compounds 6 are illustrated
in Scheme 2. Their structural diversity stem from
precursor compounds of 1,5-diaryl substituted 1H-
pyrazole-3-carboxylic acid (5), in which various
substituted benzene moieties have been introduced
into the 1- and 5- positions of pyrazole moieties
according to previously reported mature method.3,12b
Originally, methyl (Z)-2-hydroxy-4-oxo-4-phenylbut-
2-enoate (2) were synthesized from substituted
acetophenones (1) and dimethyl oxalate (DMO)
through aldol type condensation reaction. The reaction
of 2 with para-substituted phenylhydrazines (3)
Table 1. Molecular structures and the yields of compounds
6aa-6gc.
Cl
Cl
N
N
N
N
N
N
O
O
O
O
O
O
N
N
N
N
N
N
O2
N
O2
N
O2N
I
afforded
methyl
1,5-diphenyl-1H-pyrazole-3-
6aa
6ab
, 85%
, 81%
6bb
, 82%
carboxylates (4) via an intermolecular cyclization
mechanism. Various 1,5-diphenyl-1H-pyrazole-3-
carboxylic acid (5) were subsequently synthesized
through successively implemented hydrolysis and
acidification processes of 4. Finally, secnidazole
active groups were introduced by simple esterification
reactions, which yielded target compounds 1-(2-
methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-1,5-
diphenyl-1H-pyrazole-3-carboxylate (6) in moderate
to good yields.
Cl
Cl
N
N
N
N
N
N
O
O
O
O
N
N
O
O
N
N
N
N
O2
N
O2
N
O2
N
I
6bc
6dc
, 79%
, 77%
6cb
, 84%
N
N
N
N
O2
N
N
O2
O
O
N
O
O
N
O
O
N
N
N
N
N
O2
N
N
O
O
6ec
6gc
, 78%
, 75%
6fc
, 78%
Antiproliferation assays and MTT assays
The antiproliferative effects of all targeted compounds
were evaluated against four human tumor cell lines
and a normal cell line using MTT assay. These cell
2
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Page 3 of 28
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lines include human lung tumor cell NCI-H460,
investigated by an annexin V–FITC/propidium iodide
assay using flow cytometry. Considering the fact that
in apoptosis process the phosphatidylserine (PS)
exposure usually occurs before the loss of plasma
membrane integrity, the presence of annexin V+/PI-
cells was regarded as an indicator of apoptosis. As
shown in Fig. 1, the presence of both 6fc and 6bc have
led to the obvious increasing of the population of
annexin V+/PI- cells (26.1% for 6fc and 23.96% for
6bc) in comparison with that of control (9.56%). These
results suggested that apoptotic death of BEL-7404
cells was induced by two tested compounds.
human gastric tumor cell MCG-803, human ovarian
tumor cell Skov-3, human liver tumor cell BEL-7404,
and normal liver cell HL-7702. Cisplatin, a well-
known marketed antitumor drug, was employed as the
positive control in the MTT assay. As shown in Table
2, all compounds exhibited potent inhibitory activity
against four tumor cells of NCI-H460, MCG-803,
Skov-3, and BEL-7404. The IC50 values of all
compounds against four tumor cells range from 0.14
to 14.08 μM. Obviously, these values were all much
lower than the corresponding IC50 values of the
cisplatin. Among these, 6fc showed the highest
inhibiting effects on tumor cell lines of NCI-H460,
MCG-803, and Skov-3, with the IC50 values of 1.34,
0.14, and 0.87 μM, respectively. While for BEL-7404
tumor cell lines, the 6bc possessed the most significant
inhibiting activity, whose IC50 value was determined to
be 2.03 μM. It should be noted that the IC50 values of
6fc and 6bc against the human normal liver cell HL-
7702 were 8.97 and 5.77 μM, respectively, all of which
were closed to that of the cisplatin (7.93 μM). This
meant that both of them displayed comparable
cytotoxic activities as the cisplatin.
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Table 2. IC50 (µM) values of compounds 6aa-6gc against
four tumor cell lines and a normal cell line.a
Compound NCI-H460 MCG-803 Skov-3 BEL-7404 HL-7702
6aa
6bb
14.08
2.43
4.68
6.42
7.88
6.67
3.85
1.34
3.54
48.52
0.32
1.27
0.68
1.05
2.47
3.52
0.79
0.14
1.86
4.19
7.34
2.67
1.47
3.38
6.59
3.17
9.69
0.87
10.37
84.21
4.56
2.78
6.45
7.66
2.03
8.59
4.65
5.28
11.24
24.87
8.46
6.25
5.34
7.81
5.77
11.76
16.52
8.97
57.78
7.93
6ab
Fig. 1 Annexin-V/propidium iodide assay of the BEL-7404
cells treated with 6fc and 6bc (20.0 μM).
6cb
6bc
6dc
Cell cycle arrest
6ec
6fc
Meanwhile, the flow cytometry analysis was also
employed to determine the apoptosis ration of BEL-
7404 cell line induced by 6fc and 6bc, thereby the
relationships between inhibiting activities of above
compounds and the cell cycle arrest of corresponding
BEL-7404 cells could be illustrated. In this assay, the
BEL-7404 cells were treated with 6fc and 6bc at their
IC50 concentrations, following which the cell cycle
phase was assayed by evaluating the DNA content of
cells stained with propidium iodide. As shown in Fig.
2, with the extending of treating time, compound 6fc
has caused increased accumulation of BEL-7404 cells
in the S phases but a decrease in the accumulation in
the G1 phase. The percentage of S phase for control
and compound 6fc at 24 and 48 h were 31.39, 37.40
and 55.26%, respectively. As for the cells treated with
6bc, the percentage of G1 phase among them also
decreased along with the treating time extending. And
the percentage of S phase also underwent a
substantially rising. It has increased from 26.21% for
control to the 33.57% at 24 h, and finally reached to
the 45.85% at 48 h. Therefore, the 6fc and 6bc could
arrest cell cycle of BEL-7404 cells at S phase. And
based on these results it could be concluded that DNA
6gc
Cisplatin
aIC50: Concentration inhibits 50% of cell growth. Data are
shown as the mean of three independent experiments run in
triplicate.
The above results confirmed the validity of our design,
the hybridization of pyrazole and secnidazole could
result in compounds with excellent antitumor activities.
In order to further probe antitumor mechanism of
targeted compounds, compounds 6fc and 6bc were
selected to determine the cell cycle analysis, apoptosis
ratio detection, and mitochondrial membrane potential
assay using BEL-7404 cell line.
Apoptosis study by flow cytometry and confocal
microscopy
In order to verify whether the compounds induced cell
death was caused by apoptosis or necrosis, the
interactions of BEL-7404 cells with 6fc or 6bc was
3
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Journal of Heterocyclic Chemistry
Page 4 of 28
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may be one of the possible intracellular targets for 6fc
or 6bc, because most DNA replication occurs within
this S stage.
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(c) control
(d) 6bc (20 μM)
Fig. 3 Loss of ΔΨm induced by 6fc and 6bc.
Conclusion
In summary, nine novel hybrid compounds containing
both diaryl substituted pyrazole and secnidazole
groups have been designed and synthesized. They
have been determined to showed potent in vitro
inhibiting activities against selected four human tumor
cell lines, which were all superior to those of the
cisplatin. Among them, compound 6fc displayed the
most significant inhibiting activities against human
tumor cell lines of NCI-H460, MCG-803 and Skov-3.
While for BEL-7404 cell, compound 6bc did the best.
Further studies revealed the possible antitumor
mechanism of 6fc and 6bc. Both of them could arrest
the BEL-7404 cell cycle at S phase, and induced
apoptosis of BEL-7404 cells through mitochondrial-
mediated pathway.
Fig. 2. Cell cycle analysis by flow cytometry for BEL-7404
cells treated with 6fc and 6bc.
Detection of mitochondrial membrane potential
It is known that mitochondria play significant roles in
the process of cell apoptosis. They could act as a point
of integration for apoptotic signals, which stem from
both the extrinsic and intrinsic apoptotic pathways.
Particularly, the loss of mitochondrial membrane
potential (MMP, ΔΨm) is involved in apoptotic cell
death due to the cytotoxicity of the antitumor drugs.
Because of this, it is regarded as an important indicator
of mitochondrial dysfunction. To investigate these
roles of mitochondria in BEL-7404 apoptosis induced
by 6fc or 6bc, the change of the mitochondrial
membrane potential was evaluated. From Fig. 3 it
could be observed that the BEL-7404 cells were
obviously changed into green by treating with
compounds 6fc and 6bc in the concentration of 20 μM,
which indicated that the MMP was declined
comparing with the control (Fig. 3.). That meant the
mitochondrial-mediated pathways were involved in
the apoptosis of BEL-7404 cells induced by 6fc and
6bc.
Experimental Section
Synthesis and characterization of 1-(2-methyl-5-nitro-
1H-imidazol-1-yl)propan-2-yl
1,5-diphenyl-1H-
pyrazole-3-carboxylate (6)
Conventional procedure:
Synthesis of compounds 2
The reaction mixture of 1 (8 mmol), dimethyl oxalate (16
mmol), and dry CH3OH (20 mL) was slowly added into the
mixture of CH3ONa/CH3OH (16 mmol/5 mL). The reaction
mixture was stirred at reflux for 5 h, and monitored
periodically by TLC. Upon completion, the reaction mixture
was cooled to room temperature and poured into water (50
mL). Then, the mixture was stirred and added HCl (1 M)
until pH = 3-4. After neutralization, the mixture was
extracted with ethyl acetate (3 × 100 mL). The combined
organic layers were washed with water and brine, dried over
NaSO4 and filtered. The solvent was removed under vacuum.
The residue was purified by flash column chromatography
to afford 2.
Synthesis of compounds 4
The diluted HCl (1 mL/1 M) was added into the reaction
mixture of 2 (1 mmol), phenylhydrazines 3 (2 mmol), and
CH3OH (12 mL). The reaction mixture was reflux for 6 h,
and monitored periodically by TLC. Upon completion, the
reaction mixture was cooled to room temperature and
extracted with ethyl acetate (3 × 20 mL). The combined
(a) control
(b) 6fc (20 μM)
4
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Page 5 of 28
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organic layers were washed with water and brine, dried over
C23H20ClIN5O4 [M+H]+ 592.02 and 594.02, found 592.10
NaSO4 and filtered. The solvent was removed under vacuum. and 594.10.
The residue was purified by flash column chromatography
to afford 4.
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-1,5-
bis(4-chlorophenyl)-1H-pyrazole-3-carboxylate (6cb)
：
Synthesis of compounds 5
yellow solid, mp: 224-226 oC; 1H NMR (500 MHz, DMSO-
d6): δ = 1.44 (d, J = 6.4 Hz, 3H), 2.55 (s, 3H), 4.54 (dd, J =
15.0, 9.7 Hz, 1H), 4.69 (dd, J = 15.0, 2.6 Hz, 1H), 5.46 (ddd,
J = 9.4, 6.4, 2.8 Hz, 1H), 7.08 (s, 1H), 7.26-7.31 (m, 2H),
7.33-7.38 (m, 2H), 7.44-7.50 (m, 2H), 7.53-7.60 (m, 2H),
8.02 (s, 1H) ppm; 13C NMR (125 MHz, DMSO-d6): δ =
14.58, 17.74, 49.99, 60.23, 70.33, 100.00, 110.57, 127.75,
127.90, 129.33, 129.89, 131.03, 133.66, 133.80, 134.42,
138.14, 138.98, 143.48, 143.92, 152.35, 160.93 ppm. m/z
(MS): calcd for C23H20Cl2N5O4 [M+H]+ 500.09 and 502.09,
found 500.10 and 502.10.
H2O (1 mL) was slowly added into the reaction mixture of
4 (0.5 mmol), KOH (1.75 mmol) and CH3OH (10 mL). The
mixture was reflux for 2 h, and monitored periodically by
TLC. Upon completion, the reaction mixture was cooled to
room temperature and poured into H2O (15 mL). Then, HCl
(1 M) was added into the reaction mixture until pH = 3-4,
followed by extracted with ethyl acetate (3 × 20 mL). The
combined organic layers were washed with water and brine,
dried over NaSO4 and filtered. The solvent was removed
under vacuum. The residue was purified by flash column
chromatography to afford 5.
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1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-5-(4-
iodophenyl)-1-phenyl-1H-pyrazole-3-carboxylate (6bc) ：
yellow solid, mp: 182-183 oC; 1H NMR (500 MHz, DMSO-
d6): δ = 1.44 (d, J = 6.4 Hz, 3H), 2.55 (s, 3H), 4.54 (dd, J =
15.0, 9.7 Hz, 1H), 4.69 (dd, J = 15.0, 2.7 Hz, 1H), 5.46 (ddd,
J = 9.4, 6.4, 2.8 Hz, 1H), 7.02 (s, 1H), 7.04 (s, 1H), 7.07 (s,
1H), 7.32-7.34 (m, 2H), 7.48-7.49 (m, 3H), 7.73 (d, J = 8.4
Hz, 2H), 8.02 (s, 1H) ppm; 13C NMR (125 MHz, DMSO-d6):
δ = 14.58, 17.75, 50.01, 70.22, 96.31, 100.00, 110.23,
126.03, 128.75, 129.39, 129.85, 131.04, 133.66, 137.95,
138.98, 139.38, 143.25, 144.02, 152.35, 161.00. m/z (MS):
calcd for C23H21IN5O4 [M+H]+ 558.06, found 558.10.
Synthesis of compounds 6
The reaction mixture of
5
(0.4 mmol), 1-(3-
dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride
(EDC, 0.48 mmol), 1-Hydroxybenzotriazole hydrate (HOBt,
0.48 mmol), 4-dimethylaminopyridine (DMAP, 0.2 mmol),
o
and DCE (10 mL) was stirred at 45 C for 0.5 h. Then,
sycnidazole (0.52 mmol) was added into the reaction
mixture and heated to reflux overnight. Upon completion,
the reaction mixture was cooled to room temperature and
extracted with DCM (3 × 20 mL). The combined organic
layers were washed with water and brine, dried over NaSO4
and filtered. The solvent was removed under vacuum. The
residue was purified by flash column chromatography or
recrystallized in EtOH to afford 6.
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-1,5-
diphenyl-1H-pyrazole-3-carboxylate (6dc) ：yellow solid,
mp: 183-185 oC; 1H NMR (500 MHz, DMSO-d6): δ = 1.45
(d, J = 6.4 Hz, 3H), 2.56 (s, 3H), 4.54 (dd, J = 15.0, 9.7 Hz,
1H), 4.70 (dd, J = 15.0, 2.7 Hz, 1H), 5.46 (ddd, J = 9.3, 6.3,
2.8 Hz, 1H), 7.03 (s, 1H), 7.22-7.27 (m, 2H), 7.33–7.29 (m,
2H), 7.34-7.39 (m, 3H), 7.44-7.51 (s, 3H), 8.02 (s, 1H) ppm;
13C NMR (125 MHz, DMSO-d6): δ = 14.59, 17.75, 50.01,
70.19, 110.05, 126.01, 129.08, 129.13, 129.23, 129.39,
Characterization of the compounds
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-1,5-di-
p-tolyl-1H-pyrazole-3-carboxylate (6aa)：yellow solid, mp: 129.74, 133.67, 138.96, 139.54, 143.13, 144.93, 152.35,
171–172 oC; 1H NMR (500 MHz, DMSO-d6): δ =1.44 (d, J
= 6.1 Hz, 3H), 2.29 (s, 3H), 2.35 (s, 3H), 2.55 (s, 3H), 4.54
(m, 1H), 4.69 (d, J = 14.1 Hz, 1H), 5.45 (br, 1H), 6.96 (s,
1H), 7.11-7.19 (m, 6H), 7.26 (d, J = 7.8, 2H), 8.02 (s, 1H)
ppm; 13C NMR (125 MHz, DMSO-d6): δ = 14.58, 17.75,
21.12, 21.23, 50.01, 70.11, 109.66, 125.78, 126.45, 128.93,
129.70, 130.14, 133.67, 137.26, 138.76, 138.94, 142.91,
144.90, 152.37, 161.13 ppm. m/z (MS): calcd for
C25H26N5O4 [M+H]+ 460.20, found 460.25.
161.07 ppm. m/z (MS): calcd for C23H22N5O4 [M+H]+
432.17, found 432.20.
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-5-(4-
methoxyphenyl)-1-phenyl-1H-pyrazole-3-
carboxylate
(6ec)：yellow solid, mp: 218-220 oC; 1H NMR (500 MHz,
DMSO-d6): δ = 1.45 (d, J = 6.1 Hz, 3H), 2.55 (s, 3H), 3.75
(s, 3H), 4.54 (dd, J = 14.5, 10.0 Hz, 1H), 4.69 (d, J = 14.5
Hz, 1H), 5.45 (br, 1H), 6.97-6.85 (m, 3H), 7.17 (d, J = 8.4,
2H), 7.32 (d, J = 5.6, 2H), 7.46 (s, 3H), 8.02 (s, 1H) ppm;
13C NMR (125 MHz, DMSO-d6): δ = 14.58, 17.76, 50.03,
55.69, 70.11, 109.47, 114.60, 121.49, 126.00, 129.15,
129.72, 130.49, 133.64, 138.99, 139.72, 143.06, 144.86,
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-1-(4-
chlorophenyl)-5-(4-iodophenyl)-1H-pyrazole-1-(2-Methyl-
5-nitro-1H-imidazol-1-yl)propan-2-yl-1-(4-chlorophenyl)-
5-(p-tolyl)-1H-pyrazole-3- carboxylate (6ab)：yellow solid, 152.36, 160.08, 161.15. m/z (MS): calcd for C24H24N5O5
mp: 169-171 oC; 1H NMR (500 MHz, DMSO-d6): δ = 1.44
(d, J = 6.4 Hz, 3H), 2.30 (s, 3H), 2.55 (s, 3H), 4.55 (dd, J =
15.0, 9.7 Hz, 1H), 4.69 (dd, J = 15.0, 2.7 Hz, 1H), 5.45 (ddd,
J = 9.4, 6.4, 2.8 Hz, 1H), 6.99 (s, 1H), 7.14 (d, J = 8.2, 2H),
7.20 (d, J = 8.1, 2H), 7.34 (m, 2H), 7.54 (m, 2H), 8.02 (s,
1H) ppm; 13C NMR (125 MHz, DMSO-d6): δ = 14.58, 17.74,
19.91, 21.26, 48.48, 70.24, 80.47, 109.99, 126.15, 127.70,
129.05, 129.78, 129.82, 133.67, 139.20, 141.38, 143.39,
145.16, 148.84, 152.36, 156.64, 161.01 ppm. m/z (MS):
calcd for C24H23ClN5O4 [M+H]+ 480.14 and 482.14, found
480.20 and 482.20.
[M+H]+ 462.20, found 462.30.
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-1-
phenyl-5-(m-tolyl)-1H-pyrazole-3-carboxylate (6fc)
：
yellow solid, mp: 158-160 oC; 1H NMR (500 MHz, DMSO-
d6): δ = 1.45 (d, J = 6.4 Hz, 3H), 2.25 (s, 3H), 2.55 (s, 3H),
4.54 (dd, J = 15.0, 2.7 Hz, 1H), 4.70 (dd, J = 15.0, 2.7 Hz,
1H), 5.46 (ddd, J = 9.3, 6.4, 2.8 Hz, 1H), 6.96 (m, J =7.2,
1H), 7.01 (s, H), 7.14 (s, H), 7.18-7.24 (m, 2H), 7.30-7.33
(m, 2H), 7.50-7.48 (m, 3H), 8.02 (s, 1H) ppm; 13C NMR
(125 MHz, DMSO-d6): δ = 14.60, 17.74, 50.02, 70.22,
110.35, 126.03, 128.10, 129.21, 129.39, 129.84, 130.91,
133.69, 134.24, 139.32, 143.18, 143.74, 161.00 ppm. m/z
(MS): calcd for C24H24N5O4 [M+H]+ 446.18, found 446.19.
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-1-(4-
chlorophenyl)-5-(4-iodophenyl)-1H-3-carboxylate (6bb)：
yellow solid, mp: 156-158 oC; 1H NMR (500 MHz, DMSO-
d6): δ = 1.44 (d, J = 6.4 Hz, 3H), 2.54 (s, 3H), 4.54 (dd, J =
15.0, 9.7 Hz, 1H), 4.69 (dd, J = 15.0, 2.7 Hz, 1H), 5.45 (ddd,
J = 9.4, 6.4, 2.8 Hz, 1H), 7.05 (s, 1H), 7.07 (d, J = 1.2, 2H),
7.36 (d, J = 8.7, 2H), 7.56 (d, J = 8.7, 2H), 7.77 (d, J = 8.4,
2H), 8.02 (s, 1H) ppm; 13C NMR (125 MHz, DMSO-d6): δ
= 14.60, 17.73, 67.70, 70.31, 96.60, 110.45, 117.46, 127.72,
129.88, 131.11, 131.15, 133.78, 138.03, 138.13, 143.48,
143.64, 144.15, 155.17, 160.89 ppm. m/z (MS): calcd for
1-(2-Methyl-5-nitro-1H-imidazol-1-yl)propan-2-yl-5-(3-
methoxyphenyl)-1-phenyl-1H-pyrazole-3-
carboxylate
o
1
(6gc): yellow solid, mp: 175-176 C; H NMR (500 MHz,
DMSO-d6): δ = 1.44 (d, J = 6.4 Hz, 3H), 2.55 (s, 3H), 3.75
(s, 3H), 4.54 (dd, J = 15.0, 2.7 Hz, 1H), 4.69 (dd, J = 15.0,
2.6 Hz, 1H), 5.45 (br, 1H), 6.91-6.95 (m, 3H), 7.17 (d, J =
8.8, 2H), 7.31-7.33 (m, 2H), 7.45-7.48 (m, 3H), 8.03 (s, 1H)
ppm; 13C NMR (125 MHz, DMSO-d6): δ = 14.59, 17.75,
21.34, 50.02, 70.15, 109.96, 125.97, 126.14, 128.93, 129.15,
5
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Journal of Heterocyclic Chemistry
Page 6 of 28
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129.22, 129.66, 129.70, 130.02, 133.67, 138.46, 139.59,
[5] a) M. Samet, K. Rahmi, I. Tuba, Ç. Ferdag, A. Ahmet,
143.09, 145.02, 152.35, 161.07 ppm. m/z (MS): calcd for
O. Salim, Eur. J. Med. Chem. 2014, 78, 86-96;
C24H24N5O5 [M+H]+ 462.18, found 462.20.
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Samuel, H.-L. Francisco, Eur. J. Med. Chem. 2013, 69,
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Acknowledgements
This work was financially supported by the National Natural
Science Foundation of China (Nos. 21961008 and 21771043) and
the
Guangxi
Natural
Science
Foundation
(No.
9
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Conflict of interest
The authors declare no conflict of interest.
[9] H.-V. Eduardo, C.-A. Romina, J. R.-E. Juan, N. M.-C.
Omar, H.-L. Francisco, P. C. Jose, E.-S. Samuel, Eur.
J. Med. Chem. 2015, 100, 106-118.
Data availability statement
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X. M. Wu, E. Mak, P. Gallant and D. Keith, Bioorg.
Med. Chem. Lett. 2003, 13, 2231–2234; b) M. A.
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Data available in article supplementary material.
The authors declare no conflict of
interest.
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