Synthesis, characterization and antitumor potential of cinnamoyl urea derivatives

Article abstract of DOI:10.14233/ajchem.2016.19403

Cinnamoyl ureas have been identified as novel compounds with their various biological activities. Some novel cinnamoyl ureas were synthesized successfully by these substitutions, at the phenyl ring, at α,β -unsaturated carbon and the substitution at the c

Full text of DOI:10.14233/ajchem.2016.19403

Asian Journal of Chemistry; Vol. 28, No. 2 (2016), 410-414
A
SIAN
J
OURNAL OF HEMISTRY
C
http://dx.doi.org/10.14233/ajchem.2016.19403
Synthesis, Characterization and Antitumor Potential of Cinnamoyl Urea Derivatives
1,*
2
3
4
RAKHI CHAUDHARY , MOHD. SHUAIB , S. RIAZ HASHIM and PREM SHANKAR MISHRA
1
2
3
4
Department of Pharmaceutical Chemistry, Translam Institute of Pharmaceutical Education and Research, Meerut-250 001, India
Pharmacy Department, Kalka Institute of Research and Advanced Studies, Meerut-250 006, India
Indian Institute of Chemical Technology, Hyderabad-500 007, India
Department of Pharmaceutical Chemistry, Meerut Institute of Engineering and Technology, Meerut-250 005, India
*Corresponding author: E-mail: rakhi.misra84@rediffmail.com
Received: 8 June 2015; Accepted: 3 August 2015;
Published online: 3 November 2015;
AJC-17619
Cinnamoyl ureas have been identified as novel compounds with their various biological activities. Some novel cinnamoyl ureas were
synthesized successfully by these substitutions, at the phenyl ring, at α,β-unsaturated carbon and the substitution at the carbonyl functionality,
which further were studied for their antitumor activity. Thus in this research work, we aimed to use all these active moieties with phenyl
urea substitutions at carbonyl moiety. Molecular docking study was used for confirming their interaction with tubulin protein for their
antitumor activity. Through in silico molecular docking study, the result showed that all the synthesized compounds (3a-3e) have minimum
binding energy and good affinity toward their active pocket, thus they were considered as good antitumor agents.
Keywords: Cinnamoyl ureas, Tubulin, Antitumor, Molecular docking study.
Various cinnamic acid analogs have been discovered
which acts as cytotoxic or microtubule destabilizing agents
[7]. Most of the cinnamic acid derivatives are substituted
with electron donating hydroxy or methoxy groups at various
positions [8]. This create the interest in the development of
cinnamoyl derivatives as tubulin inhibitors, for the design and
synthesis of novel antitumor agents with various substitution
[9].
Thus we aimed to synthesize some novel cinnamoyl ureas
by substitutions at various position. Molecular docking study
was used for confirming their interaction with tubulin protein
for their antitumor activity.
INTRODUCTION
Tumor is an abnormal growth of tissue resulting from
uncontrolled, progressive multiplication of cells and serving no
physiological function. Various factors causing this uncont-
rolled multiplication are genetic factor, radiation chemicals/
toxins, sun exposure and some causes are unknown [1]. The
tumor cells division occurs due to complex cellular signaling
involving hormones and growth factors. Latest evidences
suggests that tubulin and microtubule-associated proteins may
play an important role in a range of cellular stress responses,
thus conferring survival advantage to tumor cells [2]. Tubulin
is the target of some of the most widely used and time-honored
anticancer tubulin-binding agents (TBAs). A major chemo-
therapeutic approach to the treatment of malignant tumors has
been to disrupt the organization of microtubules in order to
prevent mitotic spindle formation and hence progression to
mitotic division [3].
EXPERIMENTAL
Most of the solvents used were ofA.R. grade and purified
before use in different reactions. Chemicals used were obtained
from Merck, Central Drug House Pvt. Ltd. (CDH), India and
Rankem Pvt. Ltd, India. Melting points of synthesized
compounds were determined by melting point apparatus and
uncorrected. All the infrared spectra were recorded on the
Perkin Elmer and Shimadzu FTIR-spectrophotometer using
Nowadays, docking methodology is used for finding the
binding sites of synthesized derivatives to its biological
receptor [4]. This technique can be used for the study of drug
receptor interaction, design and synthesis of new molecules
1
KBR pellets. H NMR spectra were recorded on 400 MHz
[
5]. Along with binding site, the favourable conformation of
spectrophotometer using DMSO as solvent and TMS as internal
standard. The mass spectra were recorded on Waters, Q-TOF
Micromass (LC-MS).All the reactions were monitored on thin
molecule can also be identified by any bond interaction either
hydrophilic or hydrophobic [6].

Vol. 28, No. 2 (2016)
Synthesis, Characterization and Antitumor Potential of Cinnamoyl Urea Derivatives 411
layer chromatography prepared by using silica gel G, petro-
leum ether and ethyl acetate in various ratio were used as
mobile phase.
3-(4-Hydroxyphenyl)-2-methyl acryloyl chloride (2b):
White crystalline solid; m.f.: C10 Cl;Yield: 67 %; R : 0.86;
m.p.: 140-141 °C; IR (neat, νmax, cm ): 1715, 3143, 1637, 2854,
H
9
O
2
f
-1
1
General procedure for the synthesis of compounds (1a-
3380; H NMR (300 MHz, CDCl
3
): 6.69 (2H C-H aromatic),
,
1
e): Substituted benzaldehyde, propionic anhydride and
7.23 (2H, C-H aromatic), 1.92 (1H, C-H), 4.79 (1H, OH).
freshly fused and finely powdered potassium acetate were
heated in an oil bath at 160 °C for 1 h and at 180 °C for 3 h.
Mixture was then poured into 100 mL of water and steam
distilled. Filtrate was acidified by conc. HCl until the evolution
of carbon dioxide cease [10]. The solids so obtained were
recrystallized from mixture of 3 vol. of water and 1 vol. of rectified
spirit. The purity of compounds was checked by the TLC.
3-(2,3,4-Trimethoxyphenyl)-2-methyl acryloyl chloride
(2c): White crystalline solid; m.f.: C13
R : 0.67; m.p.: 123-124 °C; IR (neat, νmax, cm ): 1727 (C=O,
acid) 3137 (CH, aromatic), 1658 (C=C, alkene), 2754 (C-H,
H
15
O Cl; Yield: 69 %;
4
-1
f
1
methyl), 1028, 1036 (C-O, methoxy); H NMR (300 MHz,
CDCl
4.73 (3H, OCH
3-(3-Methoxyphenyl)-2-methyl acryloyl chloride (2d):
White crystalline solid; m.f.: C11 Cl;Yield: 65 %; R : 0.74;
3
): 6.27 (1H
,
C-H aromatic), 6.64 (1H, C-H aromatic),
3
), 3.73 (3H, OCH
3
), 3.73 (3H, OCH ).
3
3
-(4-Aminophenyl)-2-methyl acrylic acid (1a): White
crystalline solid; m.f.: C10 ;Yield: 82 %; R : 0.80; m.p.:
21-123 °C; IR (neat, νmax, cm ): 1720, 2950, 3050, 1640, 2950,
H
11NO
2
f
H
11
O
2
f
-
1
-1
1
3
7
m.p.: 127-128 °C; IR (neat, νmax, cm ): 1726 (C=O, acid), 3039
(C-H, aromatic), 1656 (C=C, alkenes), 2756 (C-H, methyl),
1122 (C-O, methoxy); H NMR (300 MHz, CDCl
1
200 H NMR (300 MHz, CDCl
.0 (2H, C-H aromatic), 11 (1H, OH acid), 4 (2H, NH
-(4-Hydroxyphenyl)-2-methyl acrylic acid (1b):White
crystalline solid; m.f.: C10 Yield: 77 %; R : 0.72; m.p.:
37-138 °C; IR (neat, νmax, cm ): 1713, 3100, 3144, 1637, 2850,
3
): 6.41 (2H
,
C-H aromatic),
1
2
).
3
): 6.81 (1H
,
3
C-H aromatic), 6.55 (1H, C-H aromatic), 7.30 (1H, C-H
aromatic), 6.86 (1H, C-H aromatic), 7.81 (1H, C-H aromatic),
H
10
O
3
f
-1
1
3
7
3.53 (3H, OCH
3-(2,3-Dihydroxyphenyl)-2-methyl acryloyl chloride
(2e): White crystalline solid; m.f.: C10 Cl; Yield: 71 %;
3
).
1
380; H NMR (300 MHz, CDCl
.13 (2H, C-H aromatic), 10.93 (1H, OH acid), 5 (1H, OH).
-(2,3,4-Trimethoxyphenyl)-2-methyl acrylic acid (1c):
White crystalline solid; m.f.: C13 Yield: 66 %; R : 0.64;
m.p.: 119-120 °C; IR (neat, νmax, cm ): 1717, 3026, 3039, 1648,
3
): 6.68 (2H
,
C-H aromatic),
H O
9 3
-1
3
R : 0.66; m.p.: 136-137 °C; IR (neat, νmax, cm ): 1710 (C=O,
f
H
16
O
5
f
acid), 3049 (C-H, aromatic), 1669 (C=C, alkenes), 2845 (C-H,
-1
methyl), 1214 (C-O, hydroxy), 1225 (C-O, hydroxyl), 3339
1
1
2
854, 1012, 1018, 1038; H NMR (300 MHz, CDCl
1H C-H aromatic), 6.64 (1H, C-H aromatic), 11.10 (1H, OH
acid), 4.73 (3H, OCH ), 3.73 (3H, OCH ), 3.73 (3H, OCH ).
-(3-Methoxyphenyl)-2-methyl acrylic acid (1d): White
crystalline solid; m.f.: C11 Yield: 70 %; R : 0.76; m.p.:
20-121 °C; IR (neat, νmax, cm ): 1716, 3126, 3049, 1646, 2856,
3
): 6.17
(OH, alcohol), 3403 (OH, alcohol); H NMR (300 MHz, CDCl
3
):
(
,
6.69 (1H,
,
C-H aromatic), 6.58 (1H, C-H aromatic), 6.44 (1H,
3
3
3
C-H aromatic), 4.73 (1H, OH), 5.00 (1H, OH).
3
General procedure for the synthesis of compounds (3a-
3e): Acyl ureas were prepared by reacting various cinnamoyl
chloride derivatives with phenyl urea. Commercially available
phenyl urea was used for the reaction. Required amount of
phenyl urea in 5 % NaOH and small amount of cinnamoyl
chloride prepared in previous step was added one at a time,
with constant shaking and cooling in water (if necessary) until
odor of cinnamoyl chloride had disappeared. It was made sure
that the reaction was alkaline in nature [12]. The solid obtained
was collected by filtration and washed with cold water (Scheme-
I). The product was recrystallized from ethanol or dilute ethanol
and purity of the compound was checked by TLC.
H
12
O
3
f
-
1
1
1
6
1
120; H NMR (300 MHz, CDCl
.65 (1H, C-H aromatic), 7.10 (1H, C-H aromatic), 10.90 (1H,
).
-(2,3-Dihydroxyphenyl)-2-methyl acrylic acid (1e):
White crystalline solid; m.f.: C10 4; Yield: 75 %; R : 0.75;
m.p.: 122-123 °C; IR (neat, νmax, cm ): 1700, 3120, 3050, 1659,
3
): 6.81 (1H
,
C-H aromatic),
OH acid), 3.73 (3H, OCH
3
3
H
10
O
f
-1
1
2
855, 1210, 1223, 3340, 3400; H NMR (300 MHz, CDCl
3
):
6
.44 (1H C-H aromatic), 6.60 (1H, C-H aromatic), 6.69 (1H,
,
C-H aromatic) 10.80 (1H, OH acid), 4.73 (1H,OH), 5.00 (1H,
OH).
1-[(E)-3-(4-Aminophenyl)-2-methyl acryloyl]-3-
General procedure for the synthesis of compounds (2a-
e): These compounds were prepared by reacting initially
formed acrylic acid derivatives with thionyl chloride. Mixture
of 0.2 mol of substituted acrylic acid formed in the first step
and 0.84 mol of thionyl chloride was stirred under reflux until
the disappearance of starting material for about 4 h [11].After
phenylurea (3a): White crystalline solid; m.f.: C17
Yield: 72 %; R : 0.76; m.p.: 143-144 °C; IR (neat, νmax, cm ):
1724, 1920, 3030, 1644, 2878, 3189, 3203; H NMR (400
MHz, CDCl ): 6.41 (2H d, J = 14 Hz), 7.12 (2H, d, J = 13
H
17
N
3
O ;
2
-
1
2
f
1
3
,
Hz), 4 (2H, s), 1.78 (3H, d, J = 8.01), 5.98 (1H, s), 7.24 (5H,
+
s), 10 (1H, s) ESI-MS: m/z 295.13 (M+H ).
the reaction, excess SOCl
2
was removed in vacuum and yellow
1-[(E)-3-(4-Hydroxyphenyl)-2-methyl acryloyl]-3-
residue was directly used for further reaction without any
purification.
Stationary phase used in TLC was silica gel and mobile
phase used were acetone/petroleum ether or hexane/ethyl
acetate in 3:1 ratio.
phenylurea (3b): White crystalline solid; m.f.: C17
Yield: 73 %; R : 0.88; m.p.: 140-141 °C; IR (neat, νmax, cm ):
1688, 1927, 3055, 1643, 2949, 3196, 3327; H NMR (400
MHz, CDCl ) 6.68 (2H d, J = 14 Hz), 7.13 (2H, d, J = 14 Hz),
H
16
N
2
O ;
3
-
1
f
1
3
,
5 (2H, s), 1.93 (3H, d, J = 8), 6 (1H, s), 7.64 (5H, s), 10 (1H,
+
3
-(4-Aminophenyl)-2-methyl acryloyl chloride (2a):
s); ESI-MS: m/z 296 (M+H ).
White crystalline solid; m.f.: C10 10NOCl; Yield: 72 %; R
H
f
:
1-[(E)-3-(2,3,4-Trimethoxyphenyl)-2-methyl acryloyl]-
-1
0
2
.87; m.p.: 132-133 °C; IR (neat, νmax, cm ): 1723, 3052, 1643,
3-phenylurea (3c): White crystalline solid; m.f.: C20
Yield: 70 %; R : 0.69; m.p.: 144-145 °C; IR (neat, νmax, cm ):
1726, 1928, 3054, 1642, 2957, 3192, 3200; H NMR (400
MHz,CDCl ): 6.16-6.18 (1H m), 6.63 (1H, d, J = 15 Hz), 3.70
H
22
N
2
O ;
5
1
-1
850, 3200; H NMR (300 MHz, CDCl
3
): 6.51 (2H, C-H
), 4.32 (2H,
f
1
aromatic), 7.05 (2H, C-H aromatic), 1.93 (3H, CH
NH ).
3
2
3
,

412 Chaudhary et al.
Asian J. Chem.
R
6
.69 (1H
,
d, J = 13 Hz), 1.90 (3H, d, J = 8), 6 (1H, s), 7.64
+
(
5H, s), 10 (1H, s); ESI-MS: m/z 313 (M+H ).
CHO
) base
(R'CH CO) O
in silico Molecular docking studies: The ligands were
2
2
drawn in Marvin Sketch assigned with proper 2-D orientation
[13] in silico virtual screening of receptors is however, a daunting
task, for both of the receptor based approaches (docking) and
ligand based approaches. To perform the docking model, the
Auto Dock 4.0 suite molecular-docking tool was used and the
methodology was followed [14]. The anticancer compounds
were manually docked into sites of the enzymes and the docking
energy was monitored to achieve a minimum value [5]. The
default parameters of the automatic settings were used. Each
docking experiment consisted of 10 docking runs and the
search was conducted in a grid of 40 points per dimension.
The binding position and bound conformation of the peptide
and the rough estimate of its interactions were examined with
the Auto Dock results [15]. To analyze the mode of binding
docked conformation with minimum binding energy was
selected. In the present study, the binding site was selected
based on the amino acid residues, which are involved in binding
with tubulin protein [14].
1
2
+
) H O
3
R
R
R
CH C COOH
R'
1
a-1e
SOCl2
CH C COCl
R'
2
a-2e
NH CONHZ
2
RESULTS AND DISCUSSION
CH
C
CONHCONHZ
Synthesis of cinnamoyl ureas derivatives and charac-
terization: 3-Substituted-2-methyl acrylic acid (1a-1e) were
synthesized by reaction of substituted aromatic aldehydes with
propionic anhydride by refluxing in oil bath at 160 °C for 1 h
and at 180 °C for 3 h. Further all substituted-2-methyl acrylic
acid (2a-2e) were converted to corresponding acid chlorides
by reaction with thionyl chlorides. Finally cinnamoyl ureas
were obtained in good yield by reacting with phenyl urea in
basic media. All the synthesized compounds (1a-1e, 2a-2e)
R'
R = a:
p
p
-NH2
3a-3e
b:
-OH
c: 2,3,4-Trimethoxy
d: 3-Methoxy
e: 2,3-Dihydroxy
R' = CH3
Z = C H5
6
1
Scheme-I
were characterized by IR, H NMR analysis. Final compounds
(
3a-3e) were characterized and confirmed by recording their
1
(
(
3H, s), 3.73 (3H, s), 3.73 (3H, s), 1.93 (3H, d, J = 9), 5.97
1H, s), 7.64 (5H, s), 10 (1H, s) ESI-MS: m/z 371 (M+H ).
IR, H NMR and mass spectra.All compounds were characterized
after recrystallization from appropriate solvents. IR spectrum
of most final compound showed absorption at 3200-3100 cm
+
-1
1
-[(E)-3-(3-Methoxyphenyl)-2-methyl acryloyl]-3-
phenylurea (3d): White crystalline solid; m.f.: C18
Yield: 72 %; R : 0.75; m.p.: 149-150 °C; IR (neat, νmax, cm ):
723, 1937, 3053, 1645, 2953, 3189, 3204; H NMR (400
MHz, CDCl ): 6.65-6.67 (1H m), 6.80-6.81 (1H, m) 7.10 (1H,
H
18
N
2
O
3
;
which is due to the NH stretching. Bands near 1726, 1928,
3054, 1642, 2957 cm are observed due to C=O, C=O (urea),
-
1
-1
f
1
1
C-H (aromatic), C=C and C-H (methyl) respectively in
1
3
,
compounds 3a-3e. The H NMR spectrum of all synthesized
t, J = 14 Hz), 6.86 (1, H, d, J = 12 Hz), 3.73 (3H, s), 1.93 (3H,
compounds confirms their structure showing singlet near 6.00
due to 1H of NH and a doublet near 4.00.
d, J = 8), 6 (1H, s), 7.24 (5H, s), 9.96 (1H, s) ESI-MS: m/z 310
+
(
M+H ).
Molecular docking studies: Considering the well obtained
experimental results, it was thought worthy to perform mole-
cular docking studies, hence screening the compounds,
inculcating the compounds for in silico. Native crystal structure
of tubulin protein was obtained from Protein Data Bank.
Considering tubulin protein as the target receptor, automated
1
-[(E)-3-(2,3-Dihydroxyphenyl)-2-methyl acryloyl]-3-
phenylurea (3e): White crystalline solid; m.f.: C17
Yield: 80 %; R : 0.67; m.p.: 141-142 °C; IR (neat, νm 1a x, cm ):
160, 3204, 3040, 1727, 1653, 1932, 1223, 1255, 1255; H NMR
400 MHz, CDCl ) 5 (1H, s), 5 (1H, s), 6.44 (1H, t, J = 9),
H
16
N
2
O ;
2
-
1
f
3
(
3
TABLE-1
DOCKING STUDY DATA SHOWING BINDING AFFINITY BETWEEN SYNTHESIZED COMPOUNDS AND TUBULIN PROTEIN
Compound
no.
No. of
conformation
Binding affinity
(kcal/mol)
S. No.
Protein residue name, number and distance (Å)
1
2
3
4
5
3a
3b
3c
3d
3e
First
First
Second
Second
First
-8.00
-8.60
-8.60
-7.90
-7.40
Gly 134 (3.5), Leu 122 (2.7), Arg 158 (3.0) and Ser 160 (3.2)
Ile 121 (3.1), Leu 122 (2.8), Phe 124 (2.6), Asp 157 (2.5) and Ser 160 (3.4)
Arg 132 (3.4), Gly 136 ((3.4), Arg 137 (3.4)
Gly 134 (3.4)
Arg 132 (3.5), Lys 165 (2.5)

Vol. 28, No. 2 (2016)
Synthesis, Characterization and Antitumor Potential of Cinnamoyl Urea Derivatives 413
docking studies with newly synthesized candidates lead
compounds was performed to determine the best in silico
conformation. The docking of tubulin with newly synthesized
candidates ligands (3a-3e) exhibited well established bonds
with one or more amino acids in the receptor active pocket.
The synthesized ligand molecules having 2D structure were
converted to energy minimized 3D structures. All the five
synthesized molecules were docked. Figs. 1-5 shows the
docked images of selected candidate ligands with tubulin
protein. Table-1 shows the binding affinity, protein residue
name, number and distance covered by binding to ligands. in
silico studies revealed all the synthesized molecules showed
good binding energy toward the target protein ranging from
-1
-
8.60 to -7.40 kcal mol .
Fig. 3. in silico Binding of 3c with Tubulin (Binding energy = -8.60 kcal/
mole, receptor contacts- Arg 132, Gly 136, Arg 137
Fig. 1. in silico Binding of 3a with Tubulin (Binding energy = -8 kcal/
mole, receptor contacts- Gly-134, Leu-122, Arg-158 and Ser-160)
Fig. 4. in silico Binding of 3d with Tubulin (Binding energy= -7.90 kcal/
mole, receptor contacts- Gly 134
Fig. 2. in silico Binding of 3b with Tubulin (Binding energy = -8.60 kcal/
mole, receptor contacts- Ile 121, Leu 122, Phe 124, Asp 157 and
Ser 160
Fig. 5. in silico Binding of 3e with Tubulin (Binding energy= -7.40 kcal/
mole, receptor contacts- Arg 132, Lys 165
Conclusion
newly synthesized compounds were tested for antitumor
activity. Finally the molecular docking studies of the synthe-
sized compounds were carried out and the results of such
studies were reported by in silico docking studies. in silico
studies revealed that among the synthesized compounds 3b
and 3c showed high affinity with low energy of (-8.60) Kcal/
These new cinnamoyl urea derivatives were synthesized
in reasonably good yields.All the synthesized compounds were
purified by recrystallization using appropriate solvents and
monitored by TLC They were further characterized by H NMR,
Mass spectrometry, IR studies and elemental analyses. All the
1

414 Chaudhary et al.
Asian J. Chem.
7
8
.
.
.
F.S.K. Barar, Essentials of Pharmacotherapeutics, S. Chand & Company
Ltd., edn 3, p. 119 (2000).
H.V. Sanghani, S.H. Ganatra and R. Pande, J. Comput. Sci. Syst. Biol.,
mol with employed tubulin protein. Hence, this study has
widened the scope of developing these cinnamoyl urea
derivatives as promising antitumor agents.
5, 12 (2012).
9
1
J. Chenault and J.-F. E. Dupin, J. Heterocycl. Chem., 20, 2401 (1983).
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