Choline chloride based thiourea catalyzed highly efficient, Eco-Friendly synthesis and anti-bacterial evaluation of some new 6-Amino-4-Aryl-2,4-Dihydro-3-Phenyl pyrano [2,3-C] pyrazole-5-Carbonitrile derivatives

Article abstract of DOI:10.1007/s11164-016-2810-6

In this study, a simple and highly efficient synthesis of some new 6-amino-4-aryl-2,4-dihydro-3-phenyl pyrano [2,3-c] pyrazole-5-carbonitrile derivatives by one-pot, four-component reaction with aryl aldehydes, hydrazine hydrate, ethyl benzoylacetate and malonitrile has been achieved. This method provides many advantages such as shorter reaction time with high yields, mild reaction conditions and environmental friendliness. Furthermore, all compounds were subsequently evaluated for their in-vitro antibacterial activity compared with cefazolin by minimum inhibitory concentration, all of these compounds were more active than cefazolin. The characterization of the synthesized compounds was established by melting point, IR, ¹H NMR and ¹³C NMR spectra.

Full text of DOI:10.1007/s11164-016-2810-6

Res Chem Intermed
DOI 10.1007/s11164-016-2810-6
Choline chloride based thiourea catalyzed highly
efficient, eco-friendly synthesis and anti-bacterial
evaluation of some new 6-amino-4-aryl-2,4-dihydro-3-
phenyl pyrano [2,3-c] pyrazole-5-carbonitrile
derivatives
Masoumeh Gholami Dehbalaei1 Naser Foroughifar1
•
•
Hoda Pasdar1 Alireza Khajeh-Amiri2
Neda Foroughifar3 Mohammad Alikarami4
•
•
•
Received: 3 September 2016 / Accepted: 8 November 2016
Ó Springer Science+Business Media Dordrecht 2016
Abstract In this study, a simple and highly efficient synthesis of some new
6-amino-4-aryl-2,4-dihydro-3-phenyl pyrano [2,3-c] pyrazole-5-carbonitrile
derivatives by one-pot, four-component reaction with aryl aldehydes, hydrazine
hydrate, ethyl benzoylacetate and malonitrile has been achieved. This method
provides many advantages such as shorter reaction time with high yields, mild
reaction conditions and environmental friendliness. Furthermore, all compounds
were subsequently evaluated for their in-vitro antibacterial activity compared with
cefazolin by minimum inhibitory concentration, all of these compounds were more
active than cefazolin. The characterization of the synthesized compounds was
1
established by melting point, IR, H NMR and 13C NMR spectra.
Keywords Pyranopyrazoles Á Multicomponent reactions Á Green chemistry Á Deep
eutectic solvents Á MIC
&
Naser Foroughifar
n_foroughifar@yahoo.com
1
2
Department of Chemistry, Tehran North Branch, Islamic Azad University, Tehran, Iran
Young Researchers and Elites Club, Yadegar-e Imam Khomeini (RAH) Branch, Islamic Azad
University, Tehran, Iran
3
4
Division of Diabetes, Endocrinology and Metabolism, Faculty of Medicine, Imperial College,
London W12 ONN, UK
Department of Chemistry, Ilam Branch, Islamic Azad University, Ilam, Iran
123

M. G. Dehbalaei et al.
Introduction
Multicomponent reactions (MCRs) consist of two or more starting materials
reacting simultaneously to give a single product without isolating the intermediates
which are one-step processes [1]. These reactions are an efficient green tool for the
synthesis molecular complexity and diversity coupled with shorter reaction time,
atom economy, low cost and minimum waste production [2–4].
During the last fer years, pyrano pyrazoles have interested synthetic organic
chemists and biochemists because of their biological [5] and pharmacological
activities [6] such as antibacterial activity compared with cefazolin and
ciprofloxacin [7, 8], biodegradable agrochemicals [8], antimicrobial [9], antifungal
[10], antitumor, anticancer [11], analgesic and anti-inflammatory properties
[12, 13], anti-Alzheimer’s disease [14], anti-oxidant agents [15], and also as
potential inhibitors of human Chk1 kinase [16].
The pyran derivatives can couple DNA through an intercalative mode wherein
the planar polycyclic aromatic molecules insert and stack between the base pairs of
DNA which are known to intercalate into DNA [17].
DNA intercalation with exogenous molecules leads to the making of DNA-
specific drugs. The intercalation of small molecules into DNA is used in therapeutic
approaches in which the suppression of DNA iteration and gene transcription is
engaged to destroy tumor cells or infected tissue. Some of the studies have been
performed to gain more intense insight into different aspects of the interaction of
small molecules with DNA because of acquiring highly selective and efficient DNA
intercalators [18]. A large number of drugs with a pyrazole structure that are
affected in the treatment of trigeminal neuralgia, valproate, and topiramate are
useful for migraine prophylaxis, for example, carbamazepine, phenytoin, lamotrig-
ine and felbamate. This indicates a strong linkage between anti-epileptic activity
and antinociception which is already proven for many established anticonvulsant
drugs which have long been used in pain management, particularly in chronic
neuropathic pain [19, 20], with gabapentin and carbamazepine as well as phenytoin
being used in the treatment of post-herpetic neuralgia and diabetic neuropathy,
respectively [21–23].
A number of methods have been reported for the synthesis of dihydro pyrano
[2,3-c] pyrazoles employing different catalysts, such as organic bases [24, 25],
magnetized water [26], c-alumina [27], TrCl (trityl chloride) [28], DABCO [29, 30],
-
[Dsim]AlCl4 [31], ionic liquid Bmim-BF4 [32], sodium bisulfite under ultrasound
irradiation and solvent-free conditions [33], [DMDBSI].2HSO4 as ionic liquid
catalyst [34], isonicotinic acid [35], magnetic Fe3O4 nanoparticles [36], PS-PSTA
[37], TEBAC (triethyl benzyl ammonium chloride) [38], catalyst-free [39], L-
proline [40, 41], nano-TiO2 [42], and bleaching earth clay (pH 12.5) [43, 44], using
a one-pot four-component reaction in deep eutectic solvent, DES (choline
chloride:urea).
Many chemical reactions have used ionic liquids such as green alternative media
for volatile organic solvents because of their low vapor pressures, chemical and
thermal stability, nonflammability, high ionic conductivity, and
a wide
123

Choline chloride based thiourea catalyzed highly efficient…
electrochemical potential window, but the high costs and toxicity of some aquatic
species are the main disadvantages of these green solvents, which has limited their
applications in the laboratory and industry [45, 46]. DESs, such as homogeneous
catalysts, eutectic mixtures of an ammonium salt and a hydrogen-bond donor
compound such as urea, acid, amine, and salts, developed by Abbott and co-
workers, are alternatives to ionic liquids [47–49]. These eutectic mixtures are
attractive alternatives to room-temperature ionic liquids, as DESs can be less
expensive, more synthetically available, nontoxic, and biodegradable [47, 50–54].
In continuation of our work on the synthesis of biologically important
compounds using simple, efficient, nontoxic and readily available catalysts, we
have used catalysts such as alumina, DABCO, L-proline, benzyl triphenyl
phosphonium chloride (BTPPC) and choline chloride:thiourea (DES) and choline
chloride:urea as ionic liquid catalysts for the synthesis of new pyranopyrazole
derivatives from aryl aldehydes, malonitrile, hydrazine hydrate, and ethyl benzoyl
acetate under reflux and ultrasonic irradiation conditions in short reaction times with
high yields (Scheme 1). In this work, the best results according to the reaction
conditions in the presence of catalyst choline were with chloride:thiourea.
Experimental
General
All melting points were uncorrected and measured using capillary tubes on an
electrothermal digital apparatus. IR spectra were recorded on a Shimadzo (FT)-IR
300 spectrophotometer in KBr. NMR spectra were recorded on Brucker 500 and
400 MHz spectrometers in DMSO-d6 as solvent with TMS as an internal standard.
Chemical shifts (d) for 1H and 13C NMR was reported in parts per million (ppm) and
was referenced to the solvent peak; DMSO-d6 (2.50 ppm for 1H and 39.70 ppm for
13C). Multiplicities are represented by s (singlet), d (doublet), t (triplet), q (quartet)
and m (multiplet). The progress of the reaction was monitored by TLC (thin-layer
chromatography) using n–hexane/EtOAc (1:1) as an eluent.
4
R
CN
5
6
3
2
O
H
CN
O
2
1
Ph
CO2Et
Reaction Condition
CN
*
+
HN
R
N
O
NH2
NH2NH2 . H2O
1a-p
3
4
5a-p
Scheme 1 Synthesis of compounds 5a–p
123

M. G. Dehbalaei et al.
S
S
N
100 0c
5h
N
H
H
2
+
Cl
N
Cl
N
H2N
NH2
H
HO
H
HO
H
H
H
N
N
H
S
Scheme 2 Synthesis of DES based on choline chloride and thiourea
Synthesis of deep eutectic solvent
According to Ref. [44], we have synthesized choline chloride:thiourea ionic liquid
catalyst. A mixture of choline chloride (14 g) and thiourea (16 g) was heated in thae
flask at 100 °C with stirring for 5 h (Scheme 2). The resulting eutectic solvent was
then allowed to cool to room temperature and was used for the synthesis of new
pyranopyrazoles derivatives.
General procedure for the synthesis of pyranopyrazole derivatives 5a–p
To a mixture of aryl aldehyde (1 mmol), malonitrile (1 mmol) and ethanol (10 mL)
was added a catalyst (10% mol), and the reaction mixture was stirred magnetically
for 5 min at room temperature. After 5 min of stirring, hydrazine hydrate
(1.5 mmol) and ethyl benzoyl acetate (1 mmol) were added. The mixture was
ground under reflux conditions at 80 °C or ultrasonic irradiation until completion of
the reaction. The progress of the reaction was followed by TLC (thin-layer
chromatography). After completion of the reaction, the mixture was cooled to room
temperature and then water was added to it for washing, and filtered through the
filtration flask to afford the pure product without further purification.
Spectroscopic data for selected products
6-amino-2,4-dihydro-3-phenyl-4-p-tolylpyrano [2,3-c] pyrazole-5-carbonitrile (5b)
Yellow solid; m.p: 237–238 °C; Yield 98%; IR (KBr, cm-1) mmax: 3480 (NH), 3212
and 3123 (NH2), 2196 (C:N). 1H-NMR (DMSO-d6, 400 MHz) d ppm: 2.18 (s, 3H,
CH3); 4.94 (s, 1H, CH); 6.90 (brs, 2H, NH2, D2O-exchangable); 6.98–7.024 (dd,
3
J = 7.7 Hz, 4H, C6H4): 6.98–7.00 (d1, 2H, JH2,3 = 8.0 Hz, H2,3); 7.02–7.00 (d2,
2H, 3JH4,5 = 8.0 Hz, H4,5, C6H4); 7.24–7.48 (m, 5H, C6H5); 12.88 (s, 1H, NH, D2O-
exchangable). 13C-NMR (DMSO-d6, 100 MHz) d ppm: 160.4, 156.5, 142.2, 138.3,
123

Choline chloride based thiourea catalyzed highly efficient…
136.1, 129.4, 129.2, 129.1, 128.7, 127.6, 126.6, 121.1, 97.9, 58.9, 36.9, and 21.3.
Anal.Calc for C20H16N4O (328.4); C, 73.17; H, 4.87; N, 17.07%.
6-amino-2,4-dihydro-4-(3-nitrophenyl)-3-phenylpyrano [2,3-c] pyrazole-5-carbonitrile
(5c)
Yellow solid; m.p: 254–255 °C; Yield 93%; IR (KBr, cm-1) mmax: 3472 (NH), 3247
and 3096 (NH2), 2190 (C:N), 1530 and 1349 (NO2). 1H-NMR (DMSO-d6,
400 MHz) d ppm: 5.33 (s, 1H, CH); 7.11 (brs, 2H, NH2, D2O-exchangable);
3
7.23–7.48 (m, 5H, C6H5); 7.492–7.514 (dd, 1H, JH6,5 = 1.2 Hz, H6, C6H4);
3
7.576–7.602 (dt, H5): 7.576–7.579 (dt1, 1H, JH5,4 = 1.2 Hz, H5, C6H4);
3
7.599–7.602 (dt2, 1H, JH5,6 = 1.2 Hz, H5, C6H4); 7.96–7.969 (dd, 1H,
4JH2,4 = 1.6 Hz, H2, C6H4); 7.978–7.986 (dd): 7.978–7.980 (d1, 1H,
4
3JH4,5 = 0.8 Hz, H4, C6H4); 7.984–7.986 (d2, 1H, JH4,2 = 0.8 Hz, H4, C6H4);
12.96 (s, 1H, NH, D2O-exchangable). 13C-NMR (DMSO-d6, 100 MHz) d ppm:
160.9, 156.1, 147.9, 147.1, 138.9, 134.7, 130.4, 129.0, 128.9, 128.7, 126.9, 122.3,
122.2, 120.8, 96.9, 57.5, and 36.5. Anal. Calc for C19H13N5O3 (359.4): C, 63.51; H,
3.62; N, 19.50%.
6-amino-2,4-dihydro-4-(3-hydroxyphenyl)-3-phenylpyrano [2,3-c] pyrazole-5-
carbonitrile (5d)
White solid; m.p: 258–260 °C; Yield 94%; IR (KBr, cm-1) mmax: 3833 (OH),
1
3472(NH), 3331 and 3205 (NH2), 2193 (C:N). H-NMR (DMSO-d6,400 MHz) d
ppm: 4.85 (s, 1H, CH); 6.91 (brs, 2H, NH2, D2O-exchangable); 6.473–6.482 (dd,
4
1H, JH2,4 = 2 Hz, H2, C6H4); 6.5–6.528 (dd, 1H, J = 7.7 Hz); 6.5–6.502 (d1, 1H,
4
3JH4,5 = 0.8 Hz, H4); 6.526–6.528 (d2, 1H, JH4,6 = 0.8 Hz, H4); 6.571–6.59 (dd,
1H, 3JH6,5 = 7.6 Hz, H6); 6.98–7.02 (t, 1H, 3JH5,6 = 7.6 Hz, H5); 7.25–7.48 (m, 5H,
C6H5); 9.26 (1H, s, OH, D2O-exchangable); 12.89 (s, 1H, NH, D2O-exchangable).
13C-NMR (DMSO-d6, 100 MHz) d ppm: 160.5, 157.8, 156.5, 146.7, 138.3, 129.7,
129.2, 129.1, 128.8, 126.6, 121.1, 118.5, 114.4, 114.3, 97.9, 58.9 and 37.2. Anal.
Calc for C19H14N4O2 (330.37): C, 69.09; H, 4.24; N, 16.97%.
6-amino-2,4-dihydro-4-(4-methoxyphenyl)-3-phenylpyrano[2,3-c]pyrazole-5-carbonitrile
(5i)
1
White solid; m.p: 241–243 °C; Yield 98%; H-NMR (DMSO-d6,400 MHz) d ppm:
3.34 (s, 3H, OCH3); 5.05 (s, 1H, CH); 6.98 (brs, 2H, NH2, D2O-exchangable);
3
7.116–7.267 (dd, 4H, C6H4, J = 7.7 Hz): 7.116–7.137 (d1, 2H, JH2,3 = 8.4 Hz,
3
H
2,3): 7.240–7.261 (d2, 2H, JH5,6 = 8.4 Hz, H5,6); 7.274–7.466 (m, 5H, C6H5);
12.90 (s, 1H, NH, D2O-exchangable).13C-NMR (DMSO-d6, 100 MHz) d ppm:
161.06, 160.6, 156.3, 144.04, 138.5, 136.5, 133.07, 131.5, 130.5, 129.6, 129.5,
129.06, 128.9, 128.8, 128.7, 126.7, 120.9, 97.4, 58.2, 36.5. Anal. Calc for
C20H16N4O2 (344.6): C, 69.64; H, 4.64, N, 16.25%.
123

M. G. Dehbalaei et al.
6-amino-4-(4-bromophenyl)-2,4-dihydro-3-phenylpyrano [2,3-c] pyrazole-5-
carbonitrile (5k)
White solid; m.p: 255–256 °C; Yield 89%; IR (KBr, cm-1) mmax: 3431 (NH), 3287
1
and 3134 (NH2), 2185 (C:N), 1064 (Br). H-NMR (DMSO-d6, 400 MHz) d ppm:
5.05 (s, 1H, CH); 6.99 (brs, 2H, NH2, D2O-exchangable); 7.052v7.085 (dd, 4H,
3
C6H4, J = 7.7 Hz): 7.052–7.063 (d1, 1H, JH3,2 = 2.4 Hz, H3); 7.075–7.085 (d2,
3
1H, JH2,3 = 2.4 Hz, H2); 7.25–7.40 (m, 5H, C6H5); 7.449–7.472 (d1, 1H,
3
3JH5,6 = 1.6 Hz, H5); 7.075–7.085 (d2, 1H, JH6,5 = 1.6 Hz, H6); 12.93 (s, 1H,
NH, D2O-exchangable). 13C-NMR (DMSO-d6, 100 MHz) d ppm: 160.6, 156.3,
144.5, 138.5, 131.7, 130.1, 129.0, 128.9, 126.7, 121.0, 120.1, 97.3, 58.2 and 36.6.
Anal. Calc for C19H13BrN4O (393.3); C, 58.01; H, 3.31; N, 14.25%.
6-amino-2,4-dihydro-4-(4-isopropylphenyl)-3-phenylpyrano [2,3-c] pyrazole-5-
carbonitrile (5m)
White solid; m.p: 250–251 °C; Yield 97%; IR (KBr, cm-1) mmax: 3481 (NH), 3222
1
and 3108 (NH2), 2196 (C:N). H-NMR (DMSO-d6, 500 MHz) d ppm:1.094–1.09
= 2.4 Hz, CH3); 1.1084–1.1132 (d, 3H,
CH) Isopropyl
3
(d, 3H, JH(CH3,
3JH(CH3, CH) Isopropyl = 2.4 Hz, CH3); 2.48–3.34 (m, 1H, CHIsopropyl); 4.92 (s, 1H,
CH); 6.89 (brs, 2H, NH2, D2O-exchangable); 7.0102–7.0822 (dd, 4H, C6H4):
3
7.0102–7.0265 (d1, 2H, JH2,3 = 8.15 Hz, H2,3); 7.0659–7.0822 (d2, 2H, JH5,6
3
=
8.15 Hz, H5,6); 7.228–7.455 (m, 5H, C6H5); 12.87 (s, 1H, NH, D2O-exchangable).
13C-NMR (DMSO-d6, 125 MHz) d ppm: 160.1, 156.1, 146.5, 142.2, 137.6, 128.7,
128.6, 128.5, 128.2, 127.1, 126.9, 126.3, 126.2, 126.1, 126.1, 120.7, 97.6, 58.5, 36.4,
32.8, 23.8 and 23.6. Anal. Calc for C22H20N4O (356.46); C, 74.12; H, 5.67; N, 15.72%.
6-amino-4-(2-chlorophenyl)-2,4-dihydro-3-phenylpyrano [2,3-c]pyrazole-5-carbonitrile
(5o)
White solid; m.p: 271–273 °C; Yield 91%; IR (KBr, cm-1) mmax: 3442 (NH), 3268
1
and 3120 (NH2), 2190 (C:N), 1058 (Cl). H-NMR (DMSO-d6, 400 MHz) d ppm:
5.43 (s, 1H, CH); 6.99 (brs, 2H, NH2, D2O-exchangable); 7.086–7.274 (m, 5H,
3
C6H5); 7.294–7.298 (dd, 1H, JH6,5 = 1.6 Hz, H6, C6H4); 7.305–7.314 (dt, 1H,
JH4 = 3.6 Hz, H4(5), C6H4); 7.361–7.385(dd, 1H, 3JH3,4 = 1.6 Hz, H3, C6H4); 12.86
(s, 1H, NH, D2O-exchangable). 13C-NMR (DMSO-d6, 100 MHz) d ppm: 160.9,
156.6, 141.5, 138.5, 132.5, 131.1, 129.8, 128.9, 128.8, 128.0, 126.5, 120.5, 97.1,
56.9 and 34.5. Anal. Calc for C19H13ClN4O (348.81); C, 65.36; H, 3.73; N, 16.05%.
Results and discussion
Chemistry
First, we began our study with the optimization of the four-component reaction in
ethanol between arylaldehyde, malonitrile, hydrazine hydrate and ethylbenzylacetate.
123

Choline chloride based thiourea catalyzed highly efficient…
This reaction was initially carried out in the presence of the choline chloride/thiourea
catalyst (10% mol DES) under reflux for 40 min or under ultrasonic irradiation for
8 min and afforded 6-amino-2,4-dihydro-3-phenyl-4-p-tolylpyrano[2,3-c]pyrazole-5-
carbonitrile, 5a (Fig. 1).
It was also studied under several conditions. Several catalysts (choline chloride/
urea, choline chloride/thiourea, DABCO, Al2O3, L-proline, and BTPPC) and various
environmentally benign solvents (water, water–EtOH and EtOH) were investigated
to find the best conditions (Scheme 3). After optimization, the best results in terms
of yield were achieved by using 10% mol choline chloride/thiourea (DES) in EtOH
under reflux or ultrasonic irradiation conditions (isolated yields were, respectively,
98 and 95%). The results of this comparative experiment are shown in Table 1. The
recovery and reusability of the catalyst are very important for commercial and
industrial applications as well as green process considerations. Thus, after
completion of the reaction, the ChCl.2thiourea ionic liquid catalyst was recycled
by simple extraction of the product with diethyl ether from the reaction mixture. The
viscous ionic liquid that remains in the reaction test tube was thoroughly washed
with ether and reused in subsequent reactions without further purifications.
Moreover, the catalyst is reusable for the next catalytic cycles after activating the
ionic liquid at 80 °C under vacuum in each cycle. To optimize the reaction
conditions, we also verified the amount of catalyst needed for the preparation of 5a–
p, and the best result was obtained using 10% mol ChCl.2thiourea catalyst
(Table 2). The results compare the effect of the catalyst on the reaction conditions
shown in the Tables 3 and 4.
The structure of compounds (5a–p) was established via IR, 1HNMR, and
13CNMR spectroscopic data for compound 5b as a representative example. The
1HNMR spectrum of 5b has a singlet at 2.19 ppm, ascribable to methyl hydrogens;
a singlet at 4.94 ppm, ascribable to hydrogen attached to the pyran ring, a broad
singlet at 6.90 ppm for NH2 hydrogens, and a part of the aromatic has a doublet of
doublet for C6H4, a multiplet for C6H5, and a singlet at 12.88 ppm, for N–H
hydrogen.
We proposed a possible mechanism for the synthesis of pyrano [2,3-c] pyrazole,
in a one-pot reaction in the presence of a catalyst. Firstly, there is Knoevenagel
condensation between the aryl aldehyde and the malonitrile to from an alkene
intermediate in situ. The intermediate undergoes Michael addition with pyrazolone
Fig. 1 Structure of compound
5a, synthesized during the
optimization studies
*
Ph
H
CN
HN
N
O
NH2
123

M. G. Dehbalaei et al.
CN
CN
O
H
O
2
Ph
CO2Et
Reaction Condition
CN
+
HN
10 ml solvent
N
O
NH2
NH2NH2 . H2O
1a
3
4
5a
Scheme 3 Synthesized compound 5a during the optimization studies
Table 1 Comparison the effect of catalyst for the optimization of reaction conditions (5a)
Entry
Catalyst (10% mol)
Solvent (10 ml)
Reflux (80 °C)
Sonication (r.t.)
Time
Yielda (%)
Time
Yielda (%)
1
2
3
4
5
6
7
8
a
–
EtOH
EtOH
EtOH
Water
EtOH
EtOH–water
EtOH
–
12 h
40 min
1 h
20
98
87
80
50
45
90
78
1 h
25
95
80
ChCl.2thiourea
ChCl.2urea
ChCl.2thiourea
DABCO
8 min
12 min
70 min
3 h
Al2O3
4 h
L-Proline
1 h
10 min
1 h
88
25
BTPPC (20% mol)
4 h
Isolated yield
Table 2 Optimization of
amount of catalyst (% mol of
ChCl.2thiourea)
Catalyst
(%mol)
Reflux
Time (min)
Sonication (r.t.)
Time (min)
Yielda (%)
Yielda (%)
2
55
40
30
30
35
70
96
98
20
15
12
10
40
60
93
96
5
7
10
a
Isolated yield
(formed in situ from hydrazine hydrate and ethylbenzoylacetate) followed by an
intra-molecular cyclization to give polyfunctionalized pyrano [2,3-c] pyrazoles. The
possible mechanism is shown in Scheme 4.
123

Choline chloride based thiourea catalyzed highly efficient…
Table 3 Effect of catalyst for the reaction conditions (5a–p)
Entry
Benzaldehyde
(R)
Catalyst
(10% mol)
Reflux (80 °C)
Sonication (r.t.)
Time
Yielda
(%)
Time
Yielda
(%)
1
5a
5b
5b
5c
5c
5d
5d
5e
5f
H
ChCl.2thiourea
ChCl.2thiourea
ChCl.2urea
40 min
40 min
1 h
98
94
80
93
83
94
89
97
97
88
93
88
69
98
73
91
80
89
96
75
89
80
91
89
89
96
8 min
95
90
76
92
80
2
4-Me
11 min
15 min
10 min
17 min
3
4-Me
4
3-NO2
3-NO2
3-OH
ChCl.2thiourea
ChCl.2urea
30 min
40 min
50 min
30 min
45 min
35 min
1 h
5
6
ChCl.2thiourea
ChCl.2urea
7
3-OH
8
2-Me
ChCl.2thiourea
ChCl.2thiourea
ChCl.2urea
9
4-OH
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
a
5f
4-OH
5g
5h
5h
5i
2-NO2
2-OH
ChCl.2thiourea
ChCl.2thiourea
ChCl.2urea
50 min
1 h
2-OH
2 h
4-OMe
4-OMe
4-Cl
ChCl.2thiourea
ChCl.2urea
30 min
45 min
20 min
25 min
15 min
5 min
5i
5j
ChCl.2thiourea
ChCl.2urea
4 min
90
82
5j
4-Cl
11 min
5k
5l
4-Br
ChCl.2thiourea
ChCl.2thiourea
ChCl.2urea
4-N(Me)2
4-N(Me)2
4-CH(Me)2
4-CH(Me)2
4-NO2
4-NO2
2-Cl
5l
12 min
10 min
30 min
23 min
90 min
20 min
15 min
5m
5m
5n
5n
5o
5p
ChCl.2thiourea
ChCl.2urea
ChCl.2thiourea
ChCl.2urea
ChCl.2thiourea
ChCl.2thiourea
3-Br
Isolated yield
Antibacterial studies
All the synthesized compounds were assessed for their antibacterial activity against
two Gram-positive bacteria (Staphylococcus saprophyticus and S. aureus) and two
Gram-negative bacteria (Esherichia coli and Pseudomonas aeruginosa). Normal
saline was used for preparation of inoculants having 0.5 McFarland standards. The
compounds were dissolved in dimethylsolfuxide (DMSO) for bioassay. The micro-
plates were incubated at 37 °C for 24 h. Values of minimum inhibitor concentration
(MIC) were recorded as the lowest concentration of substance, which gives no
growth of inoculated bacteria. All the compounds showed antibacterial activity
against both Gram-positive and Gram-negative standard strains and their MICs
123

M. G. Dehbalaei et al.
Table 4 The preparation of pyranopyrazole (5a–p) derivatives using ChCl.2thiourea as catalyst, under
both ultrasonic irradiation and reflux conditions
Entry
1
Product
no.
Product
M.P (°C)
found
M.P (°C)
reported (Ref.)
Yielda
(%)
Yielda (%)
(Ref.)
5a
242–244
–
98
–
Ph
CN
HN
N
O
NH2
CH3
2
3
4
5
5b
237–238
–
94
–
Ph
CN
HN
N
O
NH2
NO2
5c
5d
5e
254–255
258–260
238–240
255–256 [7]
96
94
97
95 [7]
Ph
CN
HN
N
O
NH2
OH
–
–
–
–
Ph
CN
HN
N
O
NH2
Me
CN
Ph
HN
N
O
NH2
OH
6
5f
240–242
–
97
–
Ph
CN
HN
N
O
NH2
7
8
9
5g
5h
5i
255–257
208–210
241–243
–
–
–
93
88
98
–
–
–
NO2
CN
Ph
HN
N
O
NH2
OH
CN
Ph
HN
N
O
NH2
OMe
Ph
CN
HN
N
O
NH2
Cl
10
5j
244–245
244–246 [7]
91
90 [7]
Ph
CN
HN
N
O
NH2
123

Choline chloride based thiourea catalyzed highly efficient…
Table 4 continued
Entry
Product
no.
Product
M.P (°C)
found
M.P (°C)
reported (Ref.)
Yielda
(%)
Yielda (%)
(Ref.)
Br
11
5k
5l
255–256
233–234
–
–
89
–
Ph
CN
HN
N
O
NH2
Me
Me
N
12
13
14
96
–
Ph
CN
HN
N
O
NH2
Me
Me
5m
5n
250–251
249–250
–
–
89
91
–
–
Ph
CN
HN
N
O
NH2
NO2
Ph
CN
HN
N
O
NH2
15
16
5o
5p
271–273
261–262
–
89
96
–
Cl
Ph
CN
HN
N
O
NH2
Br
262–263 [7]
94 [7]
Ph
CN
HN
N
O
NH2
a
Isolated yield
ranged between 15 and 45 lg/ml. The MICs of these compounds and cefazolin were
determined by using the standard protocol of the NCCLS Broth Microdilution MIC
method [55] and the results are presented in Tables 5 and 6.
Conclusion
The structures of some new 6-amino-4-aryl-2,4-dihydro-3-phenyl pyrano [2,3-c]
pyrazole-5-carbonitrile derivatives were assigned on the basis on their spectral data
and reported compounds by comparing them with earlier literature. This one-pot
green chemistry protocol is advantageous in requiring shorter reaction times with
high yields, mild reaction conditions, easily available and cheap catalyst, and a
simple and practical procedure without any special handing technique and requiring
routine reagents.
123

M. G. Dehbalaei et al.
S
R
H
N
H
H
(1)
Cl
N
CN
CN
H
N
O
H
OH
H
Knovenageal
H
Condensation
H
H
H
N
-H2O
N
H
R
S
Ylidenemalonitrile
(2)
H
CN
CN
S
N
Ph
H
H
Cyclization
H
N
Cl
O
N
H
-H2O, -EtOH
N
NH
OH
H
OEt
O
H
Pyrazolone
H
N
N
NH2
NH2
(3)
O
H
S
S
N
H
H
N
Ph
Cl
(4)
N
H
H
OH
H
H
H
O
N
CN
C
Ph
H
NH
S
N
N
N
H
Michael
N
OH
H
Addition Reaction
N
N
NH
S
H
H
H
N
H
Cl
N
H
R
N
S
OH
H
OH
N
l
C
H
H
R
H
H
R
H
N
H
N
H
N
S
N
CN
N
R
H
H
S
H
H
S
Cl
N
Ph
N
Ph
N
CN
H
OH
H
O
H
H
N
Ph
N
HN
N
H
H
N
H
N
O
NH2
H
Cyclization
CN
N
S
N
O
NH
(5)
H
H
H
S
N
N
H
s
H
N
N
H
S
H
C
l
OH
N
Scheme 4 Plausible mechanism for the synthesis of substituted pyrano [2,3-c] pyrazole
Screening of the compounds for their antibacterial activity compared with
cefazolin were performed at different concentrations: 5, 10, 15, 20, 25, 30, 35, 40,
45 and 50 lg/ml by using the Broth Microdilution MIC method.
123

Choline chloride based thiourea catalyzed highly efficient…
Table 5 MIC values of
Comp no. MIC (lg ml-1
compounds (5a–p)
)
E. coli P. aeruginosa S. saprophyticus S. aureus
5a
5b
5c
5d
5e
5f
25
30
30
35
35
25
30
40
40
30
35
30
30
30
25
15
20
40
40
20
20
40
20
20
45
35
30
25
45
45
30
20
30
35
25
35
30
30
35
40
30
35
30
25
35
15
35
35
35
30
40
35
40
25
30
35
30
35
30
35
35
35
15
25
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
Table 6 Antibacterial activity of different compounds by the Broth Microdilution MIC method
Antibacterial activity
against standard
strain-compounds
Concentration (lg/ml)
5
10
15
20
25
30
35
40
45
50
5a
1
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
-
-
?
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5b
1
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
-
-
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5c
1
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
-
?
-
?
-
?
-
?
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5d
1
?
?
?
?
?
?
?
?
?
?
-
?
?
-
?
?
-
?
-
-
-
-
-
-
-
-
-
-
-
-
2
3
123

M. G. Dehbalaei et al.
Table 6 continued
Antibacterial activity
against standard
strain-compounds
Concentration (lg/ml)
5
10
15
20
25
30
35
40
45
50
4
5e
1
?
?
?
?
?
?
-
-
-
-
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
?
-
?
?
?
-
-
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5f
1
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
-
-
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5g
1
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
?
-
?
?
-
-
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5h
1
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
?
-
?
?
?
-
?
?
?
-
?
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5i
1
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
-
-
?
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
2
3
4
5j
1
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5k
1
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5l
1
?
?
?
?
?
?
?
?
?
?
?
?
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
123

Choline chloride based thiourea catalyzed highly efficient…
Table 6 continued
Antibacterial activity
against standard
strain-compounds
Concentration (lg/ml)
5
10
15
20
25
30
35
40
45
50
4
5m
1
?
?
?
?
?
?
-
-
-
-
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
?
-
?
?
?
-
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
2
3
4
5n
1
?
?
?
?
?
?
?
?
?
?
-
?
?
?
-
?
?
?
-
?
-
?
-
?
-
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
2
3
4
5o
1
?
?
?
?
?
?
?
?
?
?
?
-
?
?
?
-
-
?
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
5p
1
?
?
?
?
?
?
?
?
-
?
?
?
-
-
?
?
-
-
?
-
-
-
?
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
2
3
4
Cefazolin is taken as a standard drug and its MIC is [35 lg/mL against all the four strains
1 Esherichia coli, 2 Pseudomonas aeruginosa, 3 Staphylococcus saprophyticus, 4 Staphylococcus aureus
? Resistant, - susceptible
In this work, the catalytic activity was greater for choline chloride:thiourea
compared with choline chloride:urea because of the acidic strength of thiourea
compared with urea. All the synthesized compounds showed antibacterial activity
against both Gram-positive and Gram-negative standard strains, showing that the
antibacterial activity of these compounds is greater than cefazolin.
Acknowledgements We gratefully acknowledge the financial support from the Research Council of
Islamic Azad University-Tehran North Branch.
References
1. R.M. Armstrong, A.P. Combs, P.A. Tempest, S.D. Brown, T.A. Keating, Acc. Chem. Res. 29, 123
(1996)
2. I. Ugi, Pure Appl. Chem. 73, 187 (2001)
3. Y. Gu, Green Chem. 14, 2091 (2012)
123

M. G. Dehbalaei et al.
´
4. P. Prasanna, S. Perumal, Menendez JC. Green Chem 15, 1292–1299 (2013)
5. A. Mobinikhaledi, N. Foroughifar, T. Mosleh, A. Hamta, Iranian J of Pharm Res 13, 873 (2013)
6. N. J. Parmar, B. R. Pansuriya, B. D. Parmar, H. A. Barad, Med. Chem. Res., doi:10.1007/s00044-
013-0608-2
7. S.R. Mandha, S. Siliveri, M. Alla, V.R. Bommena, M.R. Bommineni, S. Balasubramanian, Bioorg.
Med. Chem. Lett. 22, 5272 (2012)
8. M. Shahi, N. Foroughifar, A. Mobinikhaledi, Iran J Pharm Res. 14, 757 (2014)
9. H. Junek, H. Aigner, Chem. Ber. 106, 914 (1973)
10. A.H. Abdel-Rahman, E.M. Keshk, M.A. Hanna, S.H.M. El-Bady, Bioorg Med Chem. 12, 2483
(2004)
11. S.M. Bensaber, H.A. Allafe, N.B. Ermeli, S.B. Mohamed, A.A. Zetrini, S.G. Alsabri, M. Erhuma, A.
Hermann, M.I. Jaeda, A.M. Gbaj, Med. Chem. Res. 23, 5120 (2014)
12. D. Kaushik, R. Kumar, S. Ahmed Khan, G. Chawla, Med. Chem. Res. 21, 3646 (2011)
13. M.E.A. Zaki, H.A. Soliman, O.A. Hiekal, A.E. Rashad, Naturforsch. 61C, 1 (2006)
14. N.R. Dighore, P. Anandgaonker, S.T. Gaikwad, A.S. Rajbhoj, Green Process. Synth. 5, 139 (2016)
15. A. Anshu Dandia, D. Saini, S. Bhaskaran, D.K. Saini, Med. Chem. Res. (2013). doi:10.1007/s00044-
013-0671-8
16. N. Foloppe, L.M. Fisher, R. Howes, A. Potter, A.G.S. Robertson, A.E. Surgenor, Bioorg Med Chem.
14, 4792 (2006)
17. J.K. Barton, Science 233, 727 (1986)
18. R.D. Snyder, Mutat. Res. 623, 72 (2007)
19. M.A. Rogawski, Epilepsy Res. 68, 22 (2006)
20. M.A. Rogawski, W. Loscher, Nat. Med. 10, 685 (2004)
21. P. Wiffen, S. Collins, H. McQuary, D. Carroll, A. Jadad, Cochrane Database Syst. Rev. 3:CD001133
(2005)
22. C.P. Taylor, Curr. Pharm. Des. 2, 375 (1996)
23. S.L. Collins, R.A. Moore, H.J. McQuary, P. Wiffen, J Pain Symp Manage. 20, 449 (2000)
24. G. Vasuki, K. Kumaravel, Tetrahedron Lett. 49, 5636 (2008)
25. S.H.S. Azzam, M.A. Pasha, Tetrahedron Lett. 53, 6834 (2012)
26. M. Bakherad, A. Keivanloo, M. Gholizadeh, R. Doosti, M. Javanmardi, Res. Chem. Intermed.
(2016). doi:10.1007/s11164-016-2680-y
27. H. Mecadon, M.D.R. Rohman, M. Rajbangshi, B. Myrboh, Tetrahedron Lett. 52, 2523 (2011)
28. A.R. Moosavi-Zare, M.A. Zolfigol, A. Mousavi-Tashar, Res. Chem. Intermed. (2016). doi:10.1007/
s11164-016-2537-4
29. A. Hasaninejad, M. Shekouhy, N. Golzar, A. Zare, M. M. Doroodmand, Appl. Catal, A: General. 402,
11 (2011)
30. Z. Li, H. Zheng, Green Process Synth. 3, 447 (2014)
31. A. R. Moosavi-Zare, M. A. Zolfigol, E. Noroozizadeh, M. Tavasoli, V. Khakyzadeh, A. Zare, New J
Chem. 4089 (2013)
32. J.M. Khurana, B. Nand, S. Kymar, Synthetic Commun. 41, 405 (2011)
33. S.N. Darandale, J.N. Sangshetti, D.B. Shinde, J of the Korean Chem Soc. 56, 2512 (2012)
34. M. Zakeri, M.M. Nasef, T. Kargaran, A. Ahmad, E. Abouzari-Lotf, J. Asadi, Res. Chem. Intermed.
(2016). doi:10.1007/s11164-016-2648-y
35. M.A. Zolfigol, M. Tavasoli, A.R. Moosavi-Zare, P. Moosavi, H.G. Kruger, M. Shiri, V. Khakyzadeh,
RSC Adv. 3, 25681 (2013)
36. M.A.E.A.A. El-Remaily, Tetrahedron 70, 2971 (2014)
37. M.A. Chaudhari, J.B. Guijar, D.S. Kawade, N.R. Jogdand, M.S. Shingare, Cogent Chemistry (2015).
doi:10.1080/23312009.1063830
38. A. Mobinikhaledi, T. Mosleh, N. Foroughifar, Res. Chem. Intermed. 41, 2985 (2015)
39. A. Mobinikhaledi, N. Foroughifar, H. Moghanian, R. Mozaffari, Res. Chem. Intermed. 41, 6523
(2015)
40. H. Mecadon, M.D.R. Rohman, I. Kharbangar, B.M. Laloo, I. Kharkongor, M. Rajbangshi, B.
Myrboh, Tetrahedron Lett. 52, 3228 (2011)
41. S. Ambethkar, V. Padmini, N. Bhuvanesh, J Adv Res. 6, 975 (2015)
42. H.R. Shaterian, K. Azizi, Res. Chem. Intermed. 40, 661 (2014)
43. R.D. Kamble, B.S. Dawane, O.S. Yemul, A.B. Kale, S.D. Patil, Res. Chem. Intermed. 39, 3859
(2013)
123

Choline chloride based thiourea catalyzed highly efficient…
44. M.R. Bhosle, L.D. Khillare, S.T. Dhumal, R.A. Mane, Chin. Chem. Lett. (2015). doi:10.1016/j.cclet.
12.005
45. N.V. Plechkova, K.R. Seddon, Chem. Soc. Rev. 37, 123 (2008)
46. N.V. Plechkova, K.R. Seddon, P. Tundo, A. Perosa, F. Zecchini, Methods Reag. Green Chem. 105,
853 (2007)
47. C. Burda, X.B. Chen, R. Narayanan, M.A. El-Sayed, Chem. Rev. 105, 1025 (2005)
48. A.P. Abbott, R.C. Harris, K.S. Ryder, C. D’Agostino, L.F. Gladden, M.D. Mantle, Green Chem. 13,
82 (2011)
49. I.T. Horvath, Green Chem. 10, 1024 (2008)
50. A.P. Abbott, G. Capper, D.L. Davies, R.K. Rasheed, V. Tambyrajah, Chem. Commun. 7, 70 (2003)
51. B. Singh, H. Lobo, G. Shankarling, Catal. Lett. 141, 178 (2011)
52. I. Mamajanov, A.E. Engelhart, H.D. Bean, N.V. Hud, Angew. Chem. Int. Ed. 36, 6310 (2010)
53. A. Khajeh-Amiri, A. Mobimikhaledi, Res. Chem. Intermed. 39, 1491 (2013)
54. A. Mobimikhaledi, A. Khajeh-Amiri, Res. Chem. Intermed. 41, 2063 (2015)
55. P. A. Wayne, NCCLS, Performance Standards for Antimicrobial Susceptibility Testing: Twelfth
Information Supplement M, In National Committee for Clinical Laboratory Standards, 100–512
(2002)
123