Synthesis, density functional theory, and cytotoxic activity of some heterocyclic systems derived from 3-(3-(1,3-diphenyl-1H-pyrazol-4-yl)acryloyl)-2H-chromen-2-one

Article abstract of DOI:10.1007/s13738-021-02298-6

The behavior and reactivity of a coumarin-based chalcone bearing a pyrazole core 3 toward malononitrile, hydrazine, thiourea, hydrazinecarbothioamide, 2-aminoaniline, and 6-aminothiouracil were investigated as an attempt to design and synthesize a novel series of pyrazole-based heterocycles, viz. pyranocoumarin, diazepine, pyrimidochromene, triazepine, benzodiazepine, and pyrimidopyrimidine derivatives. DFT based on quantum chemical computation outlines the structure optimization of the intermediate that reacted to achieve the desired product. The antitumor activity screening against HePG-2 and MCF-7 cancer cell lines disclosed that the most potent compounds against two cell lines were 9 and 17 as compared to doxorubicin which may be attributed to their presence in more tautomeric structures. Also, the minimized energy, dipole moment, ionization potential, transferred electrons, and charge density distribution disclosed that the greater value of 0.126 and 0.8 for pyrazole derivatives 9 and 17, respectively, indicates the maximum transfer of electron and hence greater tendency of scavenging radicals and rapidly reduce oxygen to superoxide.

Full text of DOI:10.1007/s13738-021-02298-6

Journal of the Iranian Chemical Society
https://doi.org/10.1007/s13738-021-02298-6
ORIGINAL PAPER
Synthesis, density functional theory, and cytotoxic activity of some
heterocyclic systems derived from 3‑(3‑(1,3‑diphenyl‑1H‑pyrazol‑4‑yl)
acryloyl)‑2H‑chromen‑2‑one
Sayed K. Ramadan1 · Sameh A. Rizk1
Received: 16 January 2021 / Accepted: 29 May 2021
© Iranian Chemical Society 2021
Abstract
The behavior and reactivity of a coumarin-based chalcone bearing a pyrazole core 3 toward malononitrile, hydrazine,
thiourea, hydrazinecarbothioamide, 2-aminoaniline, and 6-aminothiouracil were investigated as an attempt to design and
synthesize a novel series of pyrazole-based heterocycles, viz. pyranocoumarin, diazepine, pyrimidochromene, triazepine,
benzodiazepine, and pyrimidopyrimidine derivatives. DFT based on quantum chemical computation outlines the structure
optimization of the intermediate that reacted to achieve the desired product. The antitumor activity screening against HePG-2
and MCF-7 cancer cell lines disclosed that the most potent compounds against two cell lines were 9 and 17 as compared to
doxorubicin which may be attributed to their presence in more tautomeric structures. Also, the minimized energy, dipole
moment, ionization potential, transferred electrons, and charge density distribution disclosed that the greater value of 0.126
and 0.8 for pyrazole derivatives 9 and 17, respectively, indicates the maximum transfer of electron and hence greater tendency
of scavenging radicals and rapidly reduce oxygen to superoxide.
Keywords Chromene · Chromenodiazepine · Computational chemical study · Pyrimidopyrimidine · Triazepine
Introduction
and seven-membered ring systems [22]. Indeed, as an exten-
sion of our study of the reaction products and synthesis of
some important biological heterocycles [23–41], this work
aimed to investigate the behavior and reactivity of the chal-
cone bearing a coumarin moiety 3 toward some carbon and
nitrogen nucleophiles under diferent aspects. It was found
that chalcone 3 sufers from four electron-defcient centers
attacked by the nucleophiles depending on the nature of the
nucleophile and the reaction conditions (Fig. 1).
Chalcones (1,3-diaryl-2-propane-1-ones) are secondary
metabolites obtained from the main precursors of favo-
noids and isofavonoids biosynthesis occurred in terrestrial
plants and associated with diverse biological efect [1].
Coumarins (naturally occurring oxygen heterocyclic com-
pounds) and pyrazoles (nitrogen heterocyclic compounds)
are of great importance due to their pharmacological efects
like antioxidant, antiviral, antiproliferative, antimicrobial,
and anti-infammatory activities [2–16]. Recently, pyrimi-
dines (an integral part of DNA and RNA) and their related
fused heterocyclic systems exhibit antiviral, anti-infuenza,
antioxidant, anti-infammatory, antimalarial, and antitumor
properties [6–8, 17–21]. It is well known that chalcones are
highly reactive due to α,β-unsaturated carbonyl functional-
ity, which was a good synthon for the synthesis of fve-, six-,
Results and discussion
Chemistry
Claisen–Schmidt condensation of 3-acetylcoumarin 1 with
pyrazole aldehyde 2 [42] achieved the chalcone derivative
3 as yellow crystals (Scheme 1). The IR spectrum displayed
coumarinyl C = O at ν 1722 cm−1 and ketonic C = O at ν
1655 cm−1. Its 1H NMR spectrum disclosed C5-H of pyra-
zole at δ 9.29 ppm, C4-H of coumarin at δ 8.65 ppm as well
as the aromatic and olefnic protons. Further support for the
assigned structure 3 was acquired from its mass spectrum
* Sayed K. Ramadan
sayed.karam2008@sci.asu.edu.eg
1
Department of Chemistry, Faculty of Science, Ain Shams
University, Cairo 11566, Egypt
Vol.:(0123456789)
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with malononitrile at room temperature [43] (Scheme 1).
The reaction pathway can be visualized via two routes as
displayed in Scheme 2 (according to the plausible reaction
mechanistic pathway) and Fig. 2 (according to the stability
of both intermediates and fnal products). Noteworthy, the
pyranochromene derivative 6 was obtained upon cyclocon-
densation of 5 with 4-hydroxycoumarin in boiling ethanol
and triethylamine. The 1H NMR spectrum of 6 revealed an
exchangeable singlet signal for NH2 and singlet for methine
proton in addition to the aromatic protons (cf. Experimen-
tal). In turn, cyclocondensation reaction of 5 with thiourea
in boiling ethanol including anhydrous potassium carbonate
furnished 6-aminopyrimidin-5-carbonitrile derivative 7. Its
IR spectrum disclosed bands characteristic for NH, NH2,
C≡N, and C=N groups.
Fig. 1 Nucleophilic-attacking sides in coumarin-based chalcone 3
which showed the correct molecular ion peak at m/z 418
(22%) and the base peak at m/z 77 (100%) corresponding
to phenyl cation, in addition to some important fragments
(cf. Experimental). Our research team was intrigued by
studying the regioselectivity of the reaction products under
diferent conditions [27–41]. Therefore, treating chalcone
3 with malononitrile in boiling ethanol and piperidine
failed to produce 2-amino-3-cyanopyran derivative 4 and
achieved the arylidene malononitrile 5 as a sole product
which was identical in all respects with an authentic sam-
ple prepared from condensation of pyrazole aldehyde 2
Otherwise, when chalcone 3 was treated with hydra-
zine in refuxing dioxane, chromenodiazepine derivative 9
was obtained instead of 3-coumarinyl-5-pyrazolylpyrazole
derivative 8. IR spectrum of 9 revealed the carbonyl func-
tionalities at ν 1722 and 1676 cm−1. In contrast, hydrazinol-
ysis of chalcone 3 in boiling glacial acetic acid achieved
the diazepinedione derivative 10 (Scheme 3). The chemical
Scheme 1. Synthesis of chalcone 3 and compounds 5–7
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Journal of the Iranian Chemical Society
Scheme 2. Plausible pathways for the reaction of chalcone 3 with malononitrile
structures were substantiated from their analytical and spec-
tral properties (cf. Experimental). The plausible pathways
of hydrazinolysis reaction can proceed via two routes as
depicted in Scheme 4.
with chalcone 3 in boiling dioxane to provide benzodiaz-
epine derivative 15. In contrast, pyridopyrimidine derivative
17 was obtained instead of pyridopyrimidine derivative 16
upon treating chalcone 3 with 6-aminothiouracil in boiling
dioxane including piperidine (Scheme 7). The IR spectra
conserved the coumarin carbonyl functionality. The reac-
tions can proceed via aza-Michael addition followed by ring
closure by loss of water.
The behavior of hydrazinolysis of chalcone 3 using dif-
ferent solvents to achieve compounds 9 and 10 instead of
the pyrazole derivative 8 may be attributed to the electro-
philicity of the C20 followed by C10 and C8 that are more
electrophilic center than C12, respectively (cf. Fig. 3).
The reaction of chalcone 3 with thiourea was not suc-
cessful in boiling dioxane while in boiling sodium ethoxide
acquired the chromenopyrimidine derivative 12 instead of
pyrimidinethione derivative 11 (Scheme 5). The IR spec-
trum was devoid of C=O of coumarin moiety and displayed
ν C=O of amide at 1689 cm−1. The reaction pathway can be
extrapolated via Scheme 6.
Density functional theory study
It is well known that high EHOMO are likely to indicate a
strong tendency of the molecule to donate electrons. The
low values of the energy gap (ΔE=ELUMO–EHOMO) will ren-
der good inhibition efciencies because the energy needed
to remove an electron from the last occupied orbital will
be low [44]. Density functional theory (DFT) based on
quantum chemical computation using Quantum espresso
version 6.7 outlines the structure optimization of the inter-
mediate that reacted to aford the desired product. In the
presence of reaction of the coumarin chalcone with car-
bon nucleophile, e.g., malononitrile, the HOMO energy
(−14.54 eV) rather than HOMO of nitrogen nucleophiles,
e.g., hydrazine hydrate (−10.83 eV), thiourea (−8.96 eV)
Interaction of chalcone 3 with thiosemicarbazide in boil-
ing dioxane failed to furnish the pyrazole derivative 13 and
aforded the triazepinethione derivative 14 (Scheme 7). Its
IR and 1H NMR spectra were devoid of NH2 bands. Further,
the highest recorded peak in its mass spectrum was at m/z
489 (5%) corresponding to the correct molecular ion peak
and the base peak at m/z 77 (100%) which is attributable to
phenyl cation. In a similar manner, 2-aminoaniline reacted
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Journal of the Iranian Chemical Society
Fig. 2 Stability of the intermediates and fnal products
Scheme 3. Reactions of chal-
cone 3 with hydrazine under
diferent conditions
and 6-aminothiouracil (−9.05 eV) and so good matching
to LUMO energy (−6.37 eV) of the carbonyl electrophilic
site of chalcone to form the corresponding new heterocyclic
via addition reaction followed by cyclization. In the case
of using thiosemicarbazide (HOMO −5.32 eV) is more
preferred than HOMO of o-phenylenediamine (−7.72 eV)
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Journal of the Iranian Chemical Society
Scheme 4. Plausible pathways for formation of compounds 9 and 10
proceeding via mechanism of Scheme 7 to form thermo-
dynamically intermediate. This means in the case of malo-
nonitrile, it is a thermodynamic stable intermediate (cf.
Figure 4).
were agreed with an excellent explanation for the synthe-
sized compounds and their anticancer efciency. Also, the
Ionization potential (I, eV), transferred electrons, and charge
density distribution (ΔN) indicate the greater value pyrazole
derivatives 5, 9, 12, 14, and 17 have the maximum transfer
of electron and hence, greater tendency of adsorption and
inhibition for the cancer cell. The optimization structures of
the synthesized pyrazole 3–17 are outlined in Fig. 5 (supple-
mentary material). The coordinates of the optimized struc-
tures are given in Table 2 (cf. the supporting information).
The calculated ΔE was compared with the theoretical ref-
erence data based on the corresponding experimental results
in gas phase reaction. With ΔE being a criterion, three most
typical and popular exchange–correlation functionals, e.g.,
PW91 were systematically compared in terms of the typi-
cal pyrazole, and diazepine synthesis in gas phase via reac-
tions of chalcone of coumarin with hydrazine hydrate, thio-
urea, thiosemicarbazide and 2-aminouracil in the presence
of diferent solvents, e.g., dioxane, acetic acid and ethanol
So, DFT simulation helped us to know why the reac-
tions of chalcone 3 with malononitrile, hydrazine hydrate,
thiourea, thiosemicarbazide, and 6-aminothiouracil did not
aford the expected heterocyclic compounds 4, 8, 11, 13,
and 16 and they yielded newly heterocyclic compounds 5,
9, 12, 14 and 17, respectively. The structures have been sup-
ported by full spectral analysis and microanalytical data.
Quantum chemical parameters calculation is using the den-
sity functional theory (DFT) method for the calculations
of the newly synthesized compounds (Table 1). The high
EHOMO are likely to indicate a strong tendency of the mol-
ecule to donate electrons. The low values of the energy gap
(ΔE=ELUMO–EHOMO) will render good efciencies [45]. The
dipole moment, hardness, softness, and surface area (nm2)
for newly pyrazole derivatives carrying hydrophobic groups
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Journal of the Iranian Chemical Society
Fig. 3 Electrophilicity of carbon atoms in chalcone 3
motorized via piperidine and sodium ethoxide. In addition,
the verifcations of geometrical and electronic properties of
modeling the anticancer products, as well as the adsorption
behaviors of typical probe reactants and solvents are also
suggested for further screening proper functionalization. The
present work shows general implications for how to choose
a reliable exchange–correlation functional in the computa-
tional solvents and catalyst on reactant surface.
Quantum chemical parameter calculations using den-
sity functional theory (DFT) method used for the calcu-
lations of the synthesized compounds are in good agree-
ment with the anticancer efciency (Table 1). The listed
results indicated that the values of gap energy (ΔE), where
ΔE=ELUMO−EHOMO follow the order: heterocyclic deriva-
tives 9 < 17 < doxorubicin. Compounds having small ΔE
values are generally referred to as soft compounds that are
more reactive toward radical surface interactions, being
capable of donating electrons easily to hole surface [46].
Not only the scavenging ability toward positive hole, tumor,
radical, and oxygen removable depended upon EHOMO val-
ues, but also the number of heteroatoms, electron distribu-
tions, surface area, and lipophilicity must be considered. The
dipole moment, softness (ϭ, eV−1), and surface area (nm2)
Scheme 5. Reactions of Chalcone 3 with thiourea
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Journal of the Iranian Chemical Society
Scheme 6. A suggested pathway for compound 12
for most potent synthesized compounds carrying hydropho-
bic groups were agreed to an excellent correlation between
oxidation inhibition efciencies.
for phenolic diazepine 9, pyrimidinethiol 17, and doxoru-
bicin cardiotoxicity because the myocardial tissues lack
sufcient antioxidant mechanisms [49]. Compounds 9 and
17 are the stronger anticancer than the other synthesized
compounds which may be due to their presence in more
tautomeric structures (Table 2, Fig. 6).
On the other hand, the minimized energy (kcal/mol),
dipole moment, ionization potential (I, eV), transferred
electrons, and charge density distribution (ΔN) revealed
that the greater value of 0.126 and 0.8 for pyrazole 9 and
17, respectively, indicates the maximum transfer of electron
and, hence, greater tendency of scavenging radicals and rap-
idly reduces oxygen to superoxide (O2•–) [47] (see more in
Table 1). The comparison between the Mulliken total atomic
charges of frst 20 atoms for compounds (11, 12, 13, 14, 16
and 17) is depicted in Fig. 4.
Pharmacology
Antiproliferative screening
The antitumor activity screening of the synthesized com-
pounds was measured against two cell lines, namely hepa-
tocellular carcinoma (HePG-2) and mammary gland breast
cancer (MCF-7), using MTT assay [50, 51]. The results in
Table 3 and Fig. 7 disclosed that the most potent compounds
against two cell lines were 9 and 17 as compared to doxoru-
bicin. The compounds 6, 12, and 15 were moderately potent
against HePG-2 cell line. Moderate activity was displayed
against MCF-7 cell line by the compounds 12, 14, and 15.
The rest of compounds were of weak activity.
Biological screening
It was shown that several enzymes including the cytochrome
P450 reductase, NADH dehydrogenase, and xanthine oxi-
doreductase could reduce doxorubicin via a one-electron
reduction mechanism giving rise to the phenate radical inter-
•–
mediate that can rapidly reduce oxygen to superoxide (O2
)
via a futile redox-cycling mechanism [48]. In the presence
of a redox-active metal ion such as iron (III), hydroxyl radi-
cals are formed via the Fenton mechanism. Chelators like
desferrioxamine inhibit the formation of hydroxyl radicals
through inhibition of the Haber–Weiss mechanism (Fig. 5).
Based on these in vitro studies, redox activation of newly
synthesized compounds 9 and 17 to O2•–, hydrogen peroxide
(H2O2), and iron-catalyzed hydroxyl radical formation was
suggested to be a similar mechanism of doxorubicin toxicity
[47]. Oxidative stress is thought to be primarily responsible
Conclusion
The behavior and reactivity of coumarin-based chalcone bear-
ing a pyrazole core were investigated toward some carbon and
nitrogen nucleophiles under diferent reaction conditions to
synthesize a new series of fused and pyrazole-based hetero-
cyclic systems like chromenopyran, chromenodiazepine, diaz-
epine, chromenopyrimidine, triazepinethione, benzodiazepine,
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Journal of the Iranian Chemical Society
Scheme 7. Reactions of chal-
cone 3 with thiosemicarbazide,
2-aminoaniline, and 6-ami-
nothiouracil
and pyrimidopyrimidine derivatives. The antitumor activity
screening disclosed that the chromenodiazepine and pyrimi-
dopyrimidine derivatives were the most potent against the
two tumor cells, HepG-2 and MCF-7. DFT based on quantum
chemical computation outlines the structure optimization of
the intermediate that reacted to aford the desired product.
Chemicals and solvents used were obtained from com-
mercial sources and used as received or dried by stand-
ard procedures. IR spectra (ν, cm−1) were recorded using
potassium bromide disks on Fourier Transform Infrared
Thermo Electron Nicolet iS10 Spectrometer (Thermo
Fisher Scientifc Inc., Waltham, MA, USA) at Faculty of
Science, Ain Shams University. The 1H NMR spectra (δH,
ppm) were recorded at 300 MHz on a Varian GEMINI
(GEMINI, Manufacturing & Engineering Inc., Anaheim,
CA, USA) utilizing tetramethyl silane (TMS) as an inter-
nal standard in deuterated dimethyl sulfoxide (DMSO-d6)
at Faculty of Science, Cairo University. The mass spec-
tra (MS) were recorded on Shimadzu GC-MS-QP-1000
EX mass spectrometer (Shimadzu Scientifc Instruments,
Experimental
General
All melting points were measured on a GALLENKAMP
electric melting point apparatus and are uncorrected.
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Journal of the Iranian Chemical Society
Fig. 4 Outline of the EHOMO of
nucleophiles toward ELUMO of
the starting material
Table 1 Global reactivity
indices and energy level
Compound EHOMO ELUMO ΔE Ia
Ab
−7.66 −6.14 1.52 7.09 5.38 7.29
−7.95 −5.70 2.25 8.08 5.93 16.39 1.13 21.56 0.046
µc
ηd
ωe
ΔNf
Sg
Amolec (nm2)
distribution of frontier orbitals
3
4
0.76 35.60 0.028
1.34 232.246
0.88 364.594
5
6
7
8
−7.67 −4.40 3.27 7.36 4.36 6.62
−7.80 −3.53 4.27 8.03 3.29 7.77
−8.88 −2.67 6.21 8.69 2.78 6.13
−9.38 −4.53 4.85 9.65 4.85 7.03
−4.754 −3.604 1.15 4.96 3.82 4.23
−8.41 −2.96 5.45 7.36 4.36 5.86
−7.75 −4.98 2.77 7.66 4.66 4.04
1.63 13.366 0.0748 0.61 863.013
2.14 13.88 0.0720 0.46 761.829
3.16 102.6 0.023
2.51 62.02 0.017
0.39 232.246
0.398 265.246
1.67 609.594
0.349 493.013
0.719 687.625
0.645 756.888
0.64 488.744
9
0.60 5.36
0.126
10
11
12
13
14
15
16
17
2.83 48.59 0.095
1.39 12.94 0.077
−7.67 −4.56 3.11 7.63 4.24 −3.51 1.55 11.65 0.086
−7.46 −4.35 3.11 7.49 4.38 3.25
−5.40 −5.15 0.25 5.88 5.23 8.24
−7.25 −4.67 2.58 7.26 4.66 5.14
−6.88 −4.69 2.19 6.73 4.39 2.67
−7.51 −4.62 2.89 7.62 4.38 0.43
1.56 11.49 0.087
0.13 113.78 0.0087 7.69 377.146
1.29 10.24 0.09776 0.775 491.093
1.10 13.62 0.073
1.45 12.42 0.80
0.909 344.641
0.689 411.210
aIonization potential
bElectron afnity
cChemical potential
dHardness
eElectrophilicity index
f nucleophilicity index
gSoftness
Inc., USA) operating at 70 eV at the Regional Center for
Mycology and Biotechnology (RCMB) of Al-Azhar Uni-
versity, Cairo, Egypt. The reactions were monitored by
thin-layer chromatography (TLC) using Merck Kiesel gel
60 F254 analytical sheets obtained from Fluka, Switzerland.
The starting pyrazole-4-carbaldehyde derivative 2 was pre-
viously prepared [42].
3‑(3‑(1,3‑Diphenyl‑1H‑pyrazol‑4‑yl)
acryloyl)‑2H‑chromen‑2‑one (3)
A mixture of 3-acetylcoumarin 1 (0.05 mol) and pyrazole
aldehyde 2 (0.05 mol) in absolute ethanol (20 mL) including
piperidine (0.1 mL) was refuxed for 2 h. The solid obtained
while hot was fltered of and recrystallized from n-butanol
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Journal of the Iranian Chemical Society
Fig. 5 Comparison between the Mulliken total atomic charges of frst 20 atoms for compounds (11, 12, 13, 14, 16 and 17)
to furnish chalcone 3 as yellow crystals, mp. 260–262 °C,
9.29 (s, 1H, C5-H pyrazole), 8.65 (s, 1H, C4-H coumarin),
8.00–7.40 (m, 16H, Ar–H). MS, m/z (%): 418 (M+., 22), 341
(4), 268 (9), 245 (89), 173 (25), 115 (26), 77 (100). Anal.
yield 90%. IR: 3059 (Arom-H), 1722 (C = O coumarin),
1
1655 (C = O ketone). H NMR (300 MHz, DMSO-d6):
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Table 2 Outline of the
Compound
ECal/mol
ΔEeV
µDipole
eCharge density diaz- EHOMO
ELUMO
eCharge density
pyrimidine
N6 bridge head
minimized energy and electron
density of the tautomers of
the more efective anticancer
compounds 9 and 17
epine N11, N12
9a
9b
17a
17b
17c
78.70
67.67
69.5
90.1
102.0
1.21
1.28
2.89
2.20
6.6
15.55
4.08
0.43
3.6
0.246, 0.256
0.807, 0.434
0.753
−2.74
−7.51
−7.0
1.746
−4.02
−4.62
−4.8
0.468
0.616
6.6
−6.70 −4.50 0.587
Calcd. for C27H18N2O3 (418.45): C, 77.50; H, 4.34; N, 6.69.
found: C, 77.32; H, 4.27; N, 6.71%.
C5-H pyrazole), 7.98–7.22 (m, 10H, Ar–H), 7.10 (br.s, 2H,
NH2, exchangeable), 6.42 (br.s, 1H, NH, exchangeable).
Anal. Calcd. for C20H14N6S (370.43): C, 64.85; H, 3.81; N,
22.69. found: C, 64.71; H, 3.75; N, 22.72%.
Reaction of chalcone 3 with malononitrile
A mixture of Chalcone 3 (0.01 mol) and malononitrile
(0.01 mol) containing piperidine (0.1 mL) in dioxane
(15 mL) was refuxed for 8 h. The solid was fltered of and
crystallized from petroleum ether (80–100) and found to
be 2-((1,3-diphenyl-1H-pyrazol-4-yl)methylene)malononi-
trile (5) as yellow crystals, mp. 208–210 °C, (Lit. [43] mp.
210–212 °C), yield 88%.
3‑(1,3‑Diphenyl‑1H‑pyrazol‑4‑yl)‑1,2‑dihydrochrom
eno[4,3‑c][1,2]diazepine‑5,6‑dione (9)
A solution of Chalcone 3 (0.01 mol) and hydrazine hydrate
(0.012 mol, 80%) in dioxane (15 mL) was refuxed for 8 h
(TLC). The deposited solid was fltered of and of-white
crystals were obtained after recrystallization from ethanol,
mp. 240–242 °C, yield 52%. IR: 3142 (NH), 1722, 1676
(C=O). 1H NMR (300 MHz, DMSO-d6): 11.19 (br.s, 1H,
NH, exchangeable), 9.23 (br.s, 1H, NH, exchangeable), 9.13
(s, 1H, C5-H pyrazole), 8.68 (s, 1H, =CH-CO), 8.00–7.39
(m, 14H, Ar–H). MS, m/z (%): 446 (M+., 11), 433 (21), 387
(26), 355 (31), 312 (46), 246 (34), 138 (16), 88 (32), 77
(100). Anal. Calcd. for C27H18N4O3 (446.47): C, 72.64; H,
4.06; N, 12.55. found: C, 72.51; H, 3.97; N, 12.51%.
2‑Amino‑4‑(1,3‑diphenyl‑1H‑pyrazol‑4‑yl)‑5‑oxo‑4H,
5H‑pyrano[3,2‑c]chromene‑3‑carbonitrile (6)
A mixture of arylidine malononitrile 5 (0.01 mol) and
4-hydroxycoumarin (0.01 mol) in absolute ethanol (20 mL)
including triethylamine (0.1 mL) was refluxed for 10 h
(TLC), the contents were poured onto ice-cold water and
then acidifed by dil. HCl (10%). The deposited solid was
fltered of and gray crystals were obtained after crystalliza-
tion from ethanol, mp. 228–230 °C, yield 57%. IR: 3392,
1
3318 (NH2), 2193 (C≡N), 1710 (C = O coumarin). H
2‑((7‑(1,3‑Diphenyl‑1H‑pyrazol‑4‑yl)‑3,5‑di‑
oxo‑1,2‑diazepan‑4‑ylidene)methyl)phenyl acetate
(10)
NMR (300 MHz, DMSO-d6): 8.58 (s, 1H, C5-H pyrazole),
7.87–7.29 (m, 14H, Ar–H), 7.26 (br.s, 2H, NH2, exchange-
able), 4.70 (s, 1H, C4-H pyran). MS, m/z (%): 458 (M+., 22),
419 (8), 393 (59), 392 (100), 315 (19), 270 (30), 243 (21),
120 (14), 92 (41), 77 (55). Anal. Calcd. for C28H18N4O3
(458.48): C, 73.35; H, 3.96; N, 12.22. found: C, 73.22; H,
3.89; N, 12.25%.
A solution of Chalcone 3 (0.01 mol) and hydrazine hydrate
(0.012 mol, 80%) in glacial acetic acid (10 mL) was refuxed
for 4 h (TLC). After cooling, the reaction mixture was
poured onto ice-cold water. The solid obtained was fltered
of and yellow crystals were obtained after crystallization
from ethanol, mp. 296–298 °C, yield 69%. IR: 3127 (NH),
6‑Amino‑4‑(1,3‑diphenyl‑1H‑pyrazol‑4‑yl)‑2‑thi‑
oxo‑1,2‑dihydropyrimidine‑5‑carbonitrile (7)
1
1734 (C = O ester), 1657 (C = O). H NMR (300 MHz,
DMSO-d6): 8.54 (s, 1H, C5-H pyrazole), 8.34 (s, 1H, CH=),
7.89–7.27 (m, 16H, Ar–H + 2NH), 5.76–5.70 (dd, 1H,
CHNH), 3.98–3.88 (dd, 1H, 1H of CH2), 3.28–3.22 (dd, 1H,
1H of CH2), 2.33 (s, 3H, CH3). MS, m/z (%): 492 (M+., 29),
491 (47), 474 (97), 431 (82), 353 (22), 241 (26), 220 (100),
155 (17), 104 (26), 77 (28). Anal. Calcd. for C29H24N4O4
(492.54): C, 70.72; H, 4.91; N, 11.38. found: C, 70.92; H,
4.49; N, 11.38%.
A solution of 5 (0.01 mol) and thiourea (0.01 mol) in
absolute ethanol (20 mL) containing anhydrous potassium
carbonate (0.01 mol) was heated under refux for 6 h. The
deposited solid was collected and recrystallized from ethanol
to aford compound 7 as yellow crystals, mp. 170–172 °C,
yield 68%. IR: 3375, 3250, 3175 (NH,NH2), 2195 (C≡N),
1621 (C=N). 1H NMR (300 MHz, DMSO-d6): 8.73 (s, 1H,
1 3

Journal of the Iranian Chemical Society
Fig. 6 Structures of compounds 9 and 17
1 3

Journal of the Iranian Chemical Society
Table 3 Cytotoxic activity of some compounds against human tumors
(474.54): C, 70.87; H, 3.82; N, 11.81. found: C, 70.72; H,
3.74; N, 11.84%.
Compound
In vitro cytotoxicity IC50 (µM) •
HePG-2
MCF-7
3‑(7‑(1,3‑Diphenyl‑1H‑pyrazol‑4‑yl)‑3‑thioxo‑3,4‑di‑
hydro‑2H‑1,2,4‑triazepin‑5‑yl)‑2H‑chromen‑2‑one
(14)
Doxorubicin
4.50 0.3
4.17 0.2
81.57 4.1
70.18 3.6
9.16 0.7
51.72 2.8
26.28 1.9
38.91 2.2
45.05 2.5
14.34 1.2
3
68.96 3.3
36.50 2.3
12.68 1.0
56.24 3.0
43.07 2.6
59.12 3.1
30.61 2.1
19.32 1.5
6
9
10
12
14
15
17
A solution of 3 (0.01 mol) and thiosemicarbazide (0.01 mol)
in dioxane (15 mL) was refuxed for 8 h (TLC). The reac-
tion mixture was poured onto ice-cold water, the separated
solid was fltered of and yellow crystals were obtained after
crystallization from ethanol, mp. 218–220 °C, yield 67%.
IR: 3150 (NH), 1721 (C=O), 1656 (C=N), 1231 (C=S).
1H NMR (300 MHz, DMSO-d6): 11.38 (br.s, 2H, 2NH,
exchangeable), 9.27 (s, 1H, C5-H pyrazole), 8.64 (s, 1H,
C4-H coumarin), 8.23 (s, 1H, C6-H triazepin), 7.99–7.36
(m, 14H, Ar–H). MS, m/z (%): 489 (M+., 5), 453 (7), 419
(12), 245 (51), 195 (10), 115 (22), 89 (34), 77 (100). Anal.
Calcd. for C28H19N5O2S (489.55): C, 68.70; H, 3.91; N,
14.31. found: C, 68.59; H, 3.85; N, 14.35%.
IC50 (µM): 1–10 (very strong), 11–20 (strong), 21–50 (moderate),
51–100 (weak), and above 100 (non-cytotoxic)
5‑(2‑(1,3‑Diphenyl‑1H‑pyrazol‑4‑yl)vinyl)‑2‑thi‑
oxo‑2,3‑dihydro‑4H‑chromeno[4,3‑d]pyrimi‑
din‑4‑one (12)
3‑(4‑(1,3‑Diphenyl‑1H‑pyrazol‑4‑yl)‑1H‑benzo[b][1,4]
diazepin‑2‑yl)‑2H‑chromen‑2‑one (15)
A solution of Chalcone 3 (0.01 mol), thiourea (0.01 mol) and
sodium ethoxide (0.01 mol Na/10 mL ethanol) in absolute
ethanol (10 mL) was refuxed for 6 h. After cooling, the
contents were poured onto ice-cold water and then acidifed
with dil. HCl (10%). The solid obtained was fltered of and
pale yellow crystals were obtained after crystallization from
ethanol, mp. 182–184 °C, yield 88%. IR: 3200 (NH), 1689
(C=O), 1622 (C=N), 1222 (C=S). 1H NMR (300 MHz,
DMSO-d6): 12.20 (br.s, 1H, NH, exchangeable), 9.20 (s,
1H, C5-H pyrazole), 7.94–6.41 (m, 16H, Ar–H+CH=CH).
MS, m/z (%): 474 (M+., 12), 431 (27), 427 (52), 401 (19),
331 (100), 302 (22), 248 (50), 170 (32), 135 (24), 103 (43),
92 (39), 91 (28), 77 (94). Anal. Calcd. for C28H18N4O2S
A solution of Chalcone 3 (0.01 mol) and 2-aminoaniline
(0.01 mol) in dioxane (15 mL) was refuxed for 8 h (TLC).
The reaction mixture was poured onto ice-cold water. The
precipitated solid was fltered of and dark yellow crys-
tals were obtained after crystallization from ethanol, mp.
212–214 °C, yield 84%. IR: 3160 (NH), 1721 (C=O cou-
1
marin), 1651 (C = N). H NMR (300 MHz, DMSO-d6):
12.50 (br.s, 1H, NH, exchangeable), 9.27 (s, 1H, C5-H
pyrazole), 8.64 (s, 1H, C4-H coumarin), 7.99–7.00 (m,
19H, Ar–H+ C3-H diazepin). MS, m/z (%): 506 (M+., 8),
505 (10), 467 (9), 335 (16), 307 (23), 258 (18), 184 (33),
Fig. 7 IC50 of the tested com-
doxorubicin
3
6
9
10
12
14
15
17
pounds
Compounds
1 3

Journal of the Iranian Chemical Society
103 (33), 91 (38), 77 (100). Anal. Calcd. for C33H22N4O2
(506.57): C, 78.25; H, 4.38; N, 11.06. found: C, 78.12; H,
4.30; N, 11.00%.
Dimethyl sulfoxide (DMSO) in volume of 100 µl is added
into each well to dissolve the purple formazan formed. The
colorimetric assay is measured and recorded at absorbance
of 570 nm using a plate reader (EXL 800, USA). The relative
cell viability in percentage was calculated as (A570 of treated
samples/A570 of untreated sample)×100.
2‑(1,3‑Diphenyl‑1H‑pyra‑
zol‑4‑yl)‑4‑(2‑oxo‑2H‑chromen‑3‑yl)‑6‑thioxo‑6,7‑dihy‑
dro‑8H‑pyrimido[1,6‑a]pyrimidin‑8‑one (17)
Supplementary Information The online version contains supplemen-
tary material available at https://doi.org/10.1007/s13738-021-02298-6.
An equimolar mixture of 3 and 6-aminothiouracil
(0.01 mol) in dioxane (15 mL) containing piperidine
(0.1 mL) was refuxed for 8 h (TLC). The solid was fl-
tered of and yellow crystals were obtained after crystal-
lization from ethanol, mp. 218–220 °C, yield 88%. IR:
Declarations
Conflict of interest The authors declare no confict of interest.
1
3316 (NH), 1716, 1690 (C = O), 1277 (C = S). H NMR
(300 MHz, DMSO-d6): 12.10 (br.s, 1H, NH, exchange-
able), 9.28 (s, 1H, C5-H pyrazole), 8.65 (s, 1H, C4-H cou-
marin), 8.08–7.25 (m, 15H, Ar–H + C-H pyrimidinone),
5.49 (s, 1H, C-H pyrimidine). MS, m/z (%): 541 (M+.,
45), 538 (41), 457 (33), 377 (50), 310 (82), 270 (94), 232
(100), 197 (81), 146 (99), 118 (91), 94 (37), 67 (70). Anal.
Calcd. for C31H19N5O3S (541.59): C, 68.75; H, 3.54; N,
12.93. found: C, 68.60; H, 3.47; N, 12.90%.
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