Synthesis, Characterization, and Anticancer Activity (MCF-7) of Some Acetanilide-based Heterocycles

Article abstract of DOI:10.1002/jhet.3294

A series of some novel 4-(N-substituted amino)-acetanilide scaffolds was synthesized through the nucleophilic substitution reactions of the highly versatile N-(4-acetamidophenyl)-2-chloroacetamide (3) with various types of nucleophilic reagents such as be

Full text of DOI:10.1002/jhet.3294

Month 2018
Synthesis, Characterization, and Anticancer Activity (MCF-7) of Some
Acetanilide-based Heterocycles
Ehab Abdel-Latif,*
Eman M. Keshk, Abdel-Galil M. Khalil, Ali Saeed, and Heba M. Metwally
Department of Chemistry, Faculty of Science, Mansoura University, 35516 Mansoura, Egypt
*
E-mail: ehabattia00@gmx.net
Received April 8, 2018
DOI 10.1002/jhet.3294
Published online 00 Month 2018 in Wiley Online Library (wileyonlinelibrary.com).
A series of some novel 4-(N-substituted amino)-acetanilide scaffolds was synthesized through the
nucleophilic substitution reactions of the highly versatile N-(4-acetamidophenyl)-2-chloroacetamide (3) with
various types of nucleophilic reagents such as benzothiazole-2-thiol, ethyl 2-mercaptoacetate, 4,6-dimethyl-
2
-mercapto-nicotinonitrile, various thiocarbamoyl derivatives, ammonium thiocyanate, 3-cyano-4,6-
dimethyl-5-arylazopyridin-2-ones, and malononitrile. The synthesized 4-(N-substituted amino) acetanilide
scaffolds were characterized by spectral analyses and assayed in vitro for breast anticancer activity. 2-(4-
Acetamidophenylaminocarbonyl)-3-amino-thiophenes 11, 13a, and 13b showed the highest cytotoxic
activity.
J. Heterocyclic Chem., 00, 00 (2018).
INTRODUCTION
compounds [18] (Fig. 1) were found to have a potent effect
against A549 and C6 tumor cell lines. Histone deacetylase
inhibitors (HDACi) such as suberoylanilide hydroxamic
acid [19,20] (Fig. 1) has shown potent cytotoxic effects
against several tumor types with low toxicity towards
normal cells.
Chloroacetamides are valuable synthetic intermediates
as they allow the incorporation of unmined heteroatoms
to afford a wide range of heterocycles. In this paper, we
describe the synthesis and anticancer activity of many
acetanilides incorporating thiophene, thiazole, pyridine,
and/or pyrrole moieties through different linkages.
Since the discovery of acetanilide as analgesic under the
name of Antifebrin [1], acetanilide derivatives have gained
much attention in medicinal chemistry. Literature studies
expose that acetanilide derivatives are an important
structural backbone of biologically active molecules.
They showed various activities such as antimicrobial
2,3], antiviral [4,5], anthelmintic [6], anti-inflammatory
7], cytotoxic [8,9], antifungal, and antibacterial [10,11].
Moreover, they possess anti-arthritic [12], analgesic [13],
antileishmanial [14], antitumor [15], and insecticidal [16]
properties.
[
[
Away from the famous activity of acetanilides, literature
studies reveal a potent anticancer activity for those
compounds. For example, AZD1152 [17] (Fig. 1)
inhibited cell division and cell proliferation in Aurora
kinase B overexpressed tumor cells. 2-[Benzimidazole-2-
yl)sulfanyl]-N-[4-(2-methylthiazol-4-yl)phenyl]acetamide
RESULTS AND DISCUSSION
Chemistry. The starting scaffold N-(4-acetamidophenyl)-
2-chloroacetamide (3) has been prepared by the
reported experimental conditions through treatment of
©
2018 Wiley Periodicals, Inc.

E. Abdel-Latif, E. M. Keshk, A.-G. M. Khalil, A. Saeed, and H. M. Metwally
Vol 000
Figure 1. Selected representative examples for anticancer activity of acetanilide derivatives.
p-aminoacetanilide with chloroacetyl chloride in dry
acetone in the presence of anhydrous potassium
carbonate [21] (Scheme 1). The chemical behavior of
Refluxing equimolar amounts of 3 and 2-mercapto-4,6-
dimethylnicotinonitrile (10) in ethanolic solution of
sodium ethoxide furnished the corresponding 3-
aminothieno[2,3-b] pyridine scaffold 11, the reaction
starts through nucleophilic substitution of the chlorine
atom from chloroacetamide 3 followed by intramolecular
cyclization at the nitrile function (Scheme 2). The IR
spectrum of 11 indicated the absence of any absorption
2
-chloroacetamide derivative 3 was tested towards
the reaction with several sulfur nucleophiles. Thus,
heating of chloroacetamide derivative with 2-
mercaptobenzothiazole, 2-mercaptoethanol, and/or ethyl
-mercaptoacetate in absolute ethanol and sodium acetate
3
2
afforded the corresponding sulfide derivatives 5, 7, and 9,
respectively. The sulfide derivatives were isolated in 67
to 90% yields, and their structures were established based
on spectral and elemental analyses. In the infrared
spectrum of sulfide 5, the absorption bands at 3286,
due to nitrile function and showed sharp bands of NH
2
ꢀ
1
1
group at 3490 and 3290 cm . The H NMR spectrum of
11 clearly secured the disappearance of any signal due to
the methylene function and showed the recoupment
singlet signal at 6.92 ppm for the protons of amino
function.
ꢀ
1
3
150, and 1654 cm clearly indicated the presence of
1
NH and carbonyl functions. The H NMR spectrum of 5
displayed the protons of methyl and methylene groups as
two singlet signals at 2.02 and 4.37 ppm. The aromatic
protons resonated as multiplet and doublet signals in the
region 7.34–8.02 ppm. The protons of NH functions
resonated as two singlet signals at 9.87 and 10.35 ppm.
Stirring of 2-chloroacetamide derivative
3
with
thiocarbamoyl derivatives 12 that are derived from ethyl
cyanoacetate and cyanoacetamide [22,23] in ethanolic
solution of sodium ethoxide followed by refluxing at
60–70°C
yielded
the
corresponding
2-(4-
acetamidophenylaminocarbonyl)-3-aminothiophenes 13a
Scheme 1. Synthesis of sulfide derivatives 5, 7, and 9.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

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Acetanilide-based Heterocycles
Scheme 2. Synthesis of 2-(4-acetamidophenylaminocarbonyl)-3-amino
hydroxyl)-thiophenes 11, 13, and 15.
Scheme 3. Synthesis of 2-(4-acetamidophenyl-imino)thiazolidin-4-ones
18 and 20.
(
and 13b. Elemental and spectral analyses were utilized to
secure the chemical structure of compounds 13, the IR
absorption bands of 13b at 3432, 3397, 3268, 3161, and
through intramolecular cyclization of the thiocyanate
intermediate 17 and the Dimroth-like rearrangements
[26]. The amino–imino tautomerism of the 2-arylimino-
thiazolidin-4-one 18 was established through the analysis
1
661 referred to functional groups (NH, NH , and C¼O),
2
1
1
and the absence of nitrile function. In addition, the H
NMR signals of the same compound resonated as singlet
of IR and H NMR spectral data. The tautomeric
structure 18 finds support through the appearance of a
i
1
at 2.01 ppm for three protons (CH ), singlet at 4.98 for
lactam proton (-CONH-) in the H NMR spectrum as
3
two protons (NH ), multiplet in the region 6.93–
singlet at lower field (11.12 ppm) rather than the imine
proton of 18ii which should appears at higher field
(≈9.70 ppm). The IR absorption of the lactam NH group
2
7
9
.52 ppm for the aromatic protons, three singlet at 8.80,
.32, and 10.24 ppm for the protons of three NH groups,
ꢀ
1
ꢀ1
and singlet at 9.89 ppm for two protons (NH2).
Treatment of 2-chloroacetamide derivative
at 3109 cm , together with a strong band at 1662 cm ,
3
confirms the γ-lactam tautomeric form 18 in the solid
i
with 2-(arylhydrazono)-2-ethoxycarbonyl-thioacetanilide
derivatives 14 [24] in ethanolic sodium ethoxide yielded
the corresponding 2-(4-acetamidophenylaminocarbonyl)-
state. The possibility of the tautomeric structure 18iii
involving a hydroxylic group was diminished due to the
1
absence of typical signals for OH group in IR and H
4-arylazo-3-hydroxythiophenes 15a–c. The reaction
NMR spectra.
starts via nucleophilic substitution of the chlorine atom
from chloroacetamide 3 followed by intramolecular
elimination of ethanol molecule to furnish the target 3-
hydroxythiophene products. The chemical structures of
The methylene group in this thiazolidin-4-one derivative
18 proved to be reactive towards Knoevenagel
condensation reaction with substituted benzaldehyde
derivatives. The condensation proceeded readily by
heating in glacial acetic acid and fused sodium acetate to
furnish the corresponding 5-(substituted benzylidene)-
thiazolidin-4-one derivatives 20a–d with 70–85% yield.
The structures of these targeted thiazolidine-4-ones were
assigned based on spectroscopic and analytical data. The
characteristic stretching absorptions for AcN–H, N–H,
and C¼O bonds were observed at 3218–3214, 3162–
1
5a–c were established based on their spectral and
1
elemental analyses. The H NMR spectrum of 15b (as an
example) displayed signals that agree with the designed
structure, the two methyl substituents appeared at 2.03
and 2.35 ppm as two singlet signals. Aromatic protons
were observed in the region 7.28 to 7.75 ppm as
multiplet and doublet. The protons of NH and OH
functions were observed as four singlet signals at 9.41,
ꢀ
1
1
3103, and 1676–1661 cm , respectively. The H NMR
spectra of 20a–c showed singlet for the proton of
methine function (CH¼C) deshielded to the range 7.72–
7.77 ppm.
9.90, 13.24, and 14.35 ppm.
Heterocyclization of 2-chloroacetamide
3
upon
treatment with ammonium thiocyanate to generate the
-(p-acetamidophenylimino)-thiazolidin-4-one (18) has
2
In an attempt to prepare the furo[2,3-b] pyridine
derivatives 24 via intramolecular cyclization of the O-
alkylated compounds 23, heating of 2-chloroacetamide
derivative 3 with 5-arylazo-3-cyano-4,6-dimethylpyridin-
been achieved by heating in ethyl alcohol under reflux
for 4 h (Scheme 3). The formation of this lactam
structure 18 proceeded as mentioned by Vicini et al. [25]
Journal of Heterocyclic Chemistry
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E. Abdel-Latif, E. M. Keshk, A.-G. M. Khalil, A. Saeed, and H. M. Metwally
Vol 000
Scheme 4. Reaction of chloroacetamide 3 with 5-arylazo-3-cyano-4,6-dimethylpyridin-2-ones.
2
-ones 21 in dimethylformamide containing potassium
carbonate furnished the corresponding N-alkylated
product, 2-(3-cyano-2-oxo-5-arylazopyridin-1-yl)-N-(p-
designed and identified as the 1-acetamidophenyl-2-
aminopyrrole scaffold 27 because of its correct elemental
and spectral analyses. IR absorptions for the NH and NH
2
ꢀ
1
acetamidophenyl)-acetamide derivatives 22a–d, as a sole
product (Scheme 4). The reaction failed to substitute the
chlorine atom by the nucleophilic oxygen (O-alkylation),
and the substitution proceeded by the more powerful
nucleophilic nitrogen of the pyridine ring. Elemental and
spectral analyses were utilized to secure the chemical
structure of these synthesized pyridones 22a–d,
characteristic stretching infrared absorptions for N–H and
CꢁN bonds were observed at 3315–3275 and 2224–
were observed at 3328, 3225, and 3185 cm . The
ꢀ
1
stretching absorptions at 2175, 1728, and 1647 cm were
1
recorded for CꢁN and C¼O bonds. The H NMR
spectrum clearly indicated the protons of methylene and
amino functions as singlet signals at 3.36 and 6.78 ppm.
In vitro anticancer screening. In vitro cytotoxic action
of the synthesized 4-(N-substituted amino)-acetanilide
scaffolds was assessed against human breast cancer cell
line, MCF7. Doxorubicin, which is a standout among the
best anticancer agents, was utilized as the reference
sedate as a part of this work. The relationship between
relative viability of cells (%) and concentration (μg/mL)
was plotted to acquire the survival curve of MCF7. The
results of statistical analysis of drug in vitro anticancer
revealed a significant rapprochement between the control
and the tested 2-(4-acetamidophenylaminocarbonyl)-3-
amino-thiophene compounds 11, 13a, and 13b that
proved to be the most active members in our study. They
showed very strong potency towards MCF7.
ꢀ
1
2
223 cm , respectively. The stretching absorption at
ꢀ
1
about 1666–1661 cm was recorded for C¼O double
1
bonds. In the H NMR spectra, the methylene of
acetamide moiety (NHCOCH ) was observed as singlet at
2
5.01–5.09 ppm.
The reaction of 2-chloroacetamide derivative 3 with
malononitrile as an example carbon nucleophile proceeded
by heating in ethyl alcohol in the presence of base such as
triethylamine (Scheme 5). The reaction product was
In vitro cytotoxic activities of the synthesized scaffolds
were recognized in (Table 1). IC50 values refer to the
concentration required for 100% inhibition of cell
viability. Analysis of the data in Table 1 indicated that
compounds 11, 13a, and 13b containing the thiophene
moiety exhibited the lowest IC , which means that they
Scheme 5. Synthesis of 1-(p-acetamidophenyl)-2-amino-3-cyano-5-
oxopyrrole 27.
50
are the most effective cytotoxic drugs. Accordingly,
compounds 15a–c and 27 can be used as very potent
cytotoxic drugs for breast carcinoma cell, while
compounds 3, 20b, and 22d have moderate IC , which
50
means that they are lower effective cytotoxic drugs for
breast carcinoma cell. On the other hand, the remaining
compounds 5, 7, 9, 18, 20a, 20c, 20d, 22a, and 22b are
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2018
Acetanilide-based Heterocycles
1
Table 1
iS10 FTIR spectrometer (Waltham, MA). The H NMR
spectra were recorded on a Varian XL 300 MHz
Cytotoxic activity of the synthesized scaffolds against MCF-7 cell line.
apparatus (Palo Alto, CA) using DMSO-d as a solvent.
6
In vitro cytotoxicity IC50 (μg/mL)
The mass spectra were recorded on a Quadrupole GC/MS
Thermo Scientific Focus/DSQII (Waltham, MA) at 70 eV.
Elemental analyses (C, H, and N) were determined on
Perkin-Elmer 2400 analyzer (PerkinElmer Instruments,
Shelton, CT). The NMR spectra of the products 15c, 20d,
and 22d are not provided due to their limited solubility in
common NMR solvents.
Compound
DOX
MCF-7
4.17 ± 0.2
28.94 ± 1.9
81.65 ± 3.3
90.66 ± 4.3
67.04 ± 3.0
8.96 ± 0.7
9.45 ± 0.9
10.4 ± 0.88
11.71 ± 1.0
19.39 ± 1.6
17.34 ± 1.4
52.70 ± 2.8
88.97 ± 3.8
46.83 ± 2.5
72.43 ± 3.2
94.26 ± 4.7
59.84 ± 2.9
84.72 ± 3.6
<100
3
5
7
9
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
1
3a
3b
5a
5b
5c
8
0a
0b
0c
0d
2a
2b
2c
2d
7
Synthesis of N-(4-acetamidophenyl)-2-chloroacetamide
(
3).
To a stirred suspension of 4-aminoacetanilide
(10 mmol, 1.5 g) and anhydrous potassium carbonate
(10 mmol, 1.38 g) in 40-mL acetone, 1.2 mL
chloroacetyl chloride (15 mmol) has been added drop by
drop while the stirring is continued for 2 h. The solid that
obtained on dilution with cold water was filtered and
recrystallized from ethyl alcohol.
White crystals, yield 72%, mp 248–250°C; lit. mp 240–
ꢀ1
2
43°C [21]; IR (ν/cm ): 3269, 3171 (NH), 1668 (C¼O);
1
H NMR (DMSO-d ): δ/ppm = 2.01 (s, 3H, CH ), 3.47
38.92 ± 2.1
13.80 ± 1.2
6
3
(s, 2H, CH ), 7.49 (s, 4H, Ar–H), 9.84 (s, 1H, NH),
2
1
0.04 (s, 1H, NH). Anal. Calcd. for C H ClN O
10 11 2 2
IC50 (μg/mL): 1–10 (very strong); 11–20 (strong); 21–50 (moderate); 51–
00 (weak); above 100 (non-cytotoxic). DOX, Doxorubicin.
1
(226.05): C, 52.99; H, 4.89; N, 12.36%. Found: C, 52.91;
H, 4.85; N, 12.38%.
very weak cytotoxic drugs compared with the reference
drug, Doxorubicin.
General procedure for the synthesis of sulfide derivatives 5,
7
(
, and 9. To a solution of chloroacetamide derivative 3
1.13 g, mmol) in 30-mL ethyl alcohol, 2-
mercaptobenzothiazole, 2-mercaptoethanol, and/or ethyl
-mercaptoacetate (5 mmol) in addition to 0.5 g sodium
5
Based on these preliminary screening results,
aminothiophene compounds 11, 13a, and 13b showed the
highest cytotoxic activity in the tested cancer cell. In
addition, the newly synthesized thiophene scaffolds 15a–c
exhibited good efficacy relative to the standard anticancer
drug Doxorubicin. They could be important leads in
continuing development against anticancer disease.
2
acetate was added. The mixture was heated under reflux
for 3 h and then poured into ice water. The precipitate
that formed was isolated by filtration and purified by
recrystallization from the ethyl alcohol to furnish sulfide
derivatives 5, 7, and/or 9, respectively.
N-(4-Acetamidophenyl)-2-(benzothiazol-2-thio)acetamide
(
5). White crystals, yield 90%, mp 208–210°C; IR (ν/
CONCLUSION
ꢀ
1
1
cm ): 3286, 3150 (NH), 1654 (C¼O); H NMR
(
DMSO-d ): δ/ppm = 2.02 (s, 3H, CH ), 4.37 (s, 2H,
6
3
The present study has been focused on the synthesis of
new thiophene, thiazolidine-4-one, pyridine, and pyrrole
derivatives containing biologically active acetanilide
nucleus and evaluation of their anticancer activity. Some
of the newly synthesized compounds 11, 13a, and 13b
exhibited significant activity (very strong) compared with
the control drug, Doxorubicin. Compounds 15a–c and 27
are strong cytotoxic drugs while compounds 3, 20b, and
CH ), 7.34–7.44 (m, 2H, Ar–H), 7.50 (s, 4H, Ar–H),
2
7
.83 (d, J = 8.1 Hz, 1H, Ar–H), 8.02 (d, J = 7.8 Hz, 1H,
Ar–H), 9.87 (s, 1H, NH), 10.35 (s, 1H, NH). Anal.
Calcd. for C H N O S (357.06): C, 57.12; H, 4.23; N,
17 15 3 2 2
1
1.76%. Found: C, 57.26; H, 4.25; N, 11.84%.
N-(4-Acetamidophenyl)-2-((2-hydroxyethyl)thio)acetamide
(
7). White powder, yield 70%, mp 143–145°C; IR (ν/
ꢀ1
1
cm ): 3259, 3169 (NH and OH), 1649 (C¼O); H NMR
22d exhibited a moderate activity.
(
DMSO-d ): δ/ppm = 2.01 (s, 3H, CH ), 2.70 (t,
6
3
J = 6.75 Hz, 2H, CH ), 3.28 (s, 2H, CH ), 3.56 (t,
2
2
J = 6.75 Hz, 2H, CH ), 4.85 (s, 1H, OH), 7.48 (s, 4H,
Ar–H), 9.86 (s, 1H, NH), 9.98 (s, 1H, NH). Anal. Calcd.
2
EXPERIMENTAL
for C H N O S (268.09): C, 53.71; H, 6.01; N,
12 16 2 3
All melting points (uncorrected) were measured on
Gallenkamp electric melting point apparatus. The infrared
1
0.44%. Found: C, 53.81; H, 6.04; N, 10.36%.
N-(4-Acetamidophenyl)-2-(ethoxycarbonyl-methylthio)
spectra were determined on a Thermo Scientific Nicolet
acetamide (9). White powder, yield 67%, mp 148–150°C;
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

E. Abdel-Latif, E. M. Keshk, A.-G. M. Khalil, A. Saeed, and H. M. Metwally
Vol 000
ꢀ
1
1
IR (ν/cm ): 3293, 3155 (NH), 1742, 1657 (C¼O); H
C H N O S (409.12): C, 58.67; H, 4.68; N, 17.10%.
20 19 5 3
NMR (DMSO-d ): δ/ppm = 1.18 (t, J = 7.20 Hz, 3H,
CH ), 2.01 (s, 3H, CH ), 3.40 (s, 2H, CH ), 3.49 (s, 2H,
CH ), 4.08 (q, J = 7.20 Hz, 2H, CH ), 7.48 (s, 4H, Ar–
H), 9.84 (s, 1H, NH), 10.00 (s, 1H, NH). Anal. Calcd. for
Found: C, 58.78; H, 4.71; N, 17.16%.
6
2
-(4-Acetamidophenylaminocarbonyl)-3-hydroxy-5-
3
3
2
phenylamino-4-phenylazo-thiophene (15a). Reddish brown
2
2
ꢀ
1
crystals, yield 58%, mp 224–225°C; IR (ν/cm ): 3364
1
(
NH and OH), 1664, 1628 (C¼O); H NMR (DMSO-d ):
C H N O S (310.10): C, 54.18; H, 5.85; N, 9.03%.
6
1
4 18 2 4
δ/ppm = 2.08 (s, 3H, CH ), 7.32–7.89 (m, 14H, Ar–H),
Found: C, 54.07; H, 5.90; N, 9.08%.
General procedure for the synthesis of 2-(4-
acetamidophenylaminocarbonyl)-3-amino (hydroxyl)-thiophenes
3
9
.46 (s, 1H, NH), 9.94 (s, 1H, NH), 13.32 (s, 1H, NH),
1
4.28 (s, 1H, OH). Anal. Calcd. for C H N O S
25 21
5 3
1
3
1, 13, and 15. N-(4-Acetamidophenyl)-2-chloroacetamide
(5 mmol, 1.13 g) was stirred for 10 min in sodium
(471.14): C, 63.68; H, 4.49; N, 14.85%. Found: C, 63.82;
H, 4.43; N, 14.95%.
2
-(4-Acetamidophenylaminocarbonyl)-3-hydroxy-5-
ethoxide solution (previously prepared by dissolving small
granules of sodium, 0.23 g, in 30-mL absolute ethanol)
and then the thiol derivatives (5 mmol) [namely, 2-
mercapto-4,6-dimethylnicotinonitrile (10), thiocarbamoyl
derivatives (12), or 2-(arylhydrazono)-2-ethoxycarbonyl-
thioacetanilide derivatives (14)] were added. The reaction
mixture was heated on a steam bath for 4 h and then
allowed to pour into ice water. The solid that formed after
neutralization by dilute HCl was filtered and recrystallized
from EtOH/DMF mixture (4:1) to afford the thiophene
phenylamino-4-(4-tolylazo)-thiophene (15b).
brown crystals, yield 80%, mp 235–237°C; IR (ν/cm ):
Reddish
ꢀ
1
1
3
364, 3297 (NH and OH), 1662, 1631 (C¼O); H NMR
(DMSO-d ): δ/ppm = 2.03 (s, 3H, CH ), 2.35 (s, 3H,
6
3
CH ), 7.28–7.53 (m, 11H, Ar–H), 7.75 (d, J = 8.7 Hz,
3
2
1
5
H, Ar–H), 9.41 (s, 1H, NH), 9.90 (s, 1H, NH), 13.24 (s,
H, NH), 14.35 (s, 1H, OH); MS m/z (%): 485 (M ,
6.40), 428 (45.44), 381 (73.22), 337 (51.68), 150
+
(48.81), 80 (100), 64 (97.98). Anal. Calcd. for
C H N O S (485.15): C, 64.31; H, 4.77; N, 14.42%.
26 23 5 3
products 11, 13, or 15, respectively.
N-(4-Acetamidophenyl)-3-amino-4,6-dimethylthieno[2,3-b]
pyridine-2-carboxamide (11). Orange powder, yield 88%,
Found: C, 64.38; H, 4.79; N, 14.38%.
2
-(4-Acetamidophenylaminocarbonyl)-4-(4-bromophenylazo)-
ꢀ
1
3-hydroxy-5-phenyl amino-thiophene (15c). Brown powder,
mp 278–280°C; IR ( ν /cm ): 3490, 3333, 3290, 3251
ꢀ
1
1
yield 76%, mp 290–292°C; IR (ν/cm ): 3298 (NH and
(
NH and NH ), 1684 (C¼O); H NMR (DMSO-d6):
2
+
OH), 1657 (br, C¼O); MS m/z (%): 551 (M , Br-81,
δ/ppm = 2.02 (s, 3H, CH ), 2.74 (s, 3H, CH ), 2.88 (s,
3
3
+
9
4
1
.94), 549 (M , Br-79, 6.08), 495 (32.6), 479 (61.26),
47 (100), 330 (28.7), 174 (45.04), 134 (25.8),
08 (29.8), 91 (16.85), 77 (56.10), 65 (43.70), 43 (49.4).
3
7
H, CH ), 6.92 (s, 2H, NH ), 7.05 (s, 1H, pyridine-H ),
3 2 5
.50 (d, J = 9 Hz, 2H, Ar–H), 7.56 (d, J = 9 Hz, 2H, Ar–
H), 9.33 (s, 1H, NH), 9.88 (s, 1H, NH); MS m/z (%): 355
+
+
Anal. Calcd. for C H BrN O S (549.05): C, 54.55;
25 20 5 3
H, 3.66; N, 12.72%. Found: C, 54.55; H, 3.61; N, 12.80%.
Synthesis of 2-(4-acetamidophenylimino)-thiazolidin-4-one
(M
+1, 16.15), 354 (M , 71.93), 205 (82.62), 177
(19.46), 150 (100), 133 (26.56), 108 (40.74). Anal.
Calcd. for C H N O S (354.12): C, 61.00; H, 5.12; N,
1
18 18 2 3
(
1
3
18).
A suspension of chloroacetamide 3 (5 mmol,
.13 g) and ammonium thiocyanate (10 mmol, 0.76 g) in
0-mL ethyl alcohol was heated under reflux for 4 h. The
5.81%. Found: C, 61.16; H, 5.06; N, 15.90%.
Ethyl
5-((4-acetamidophenyl)carbamoyl)-4-amino-2-
(13a).
(phenylamino)thiophene-3-carboxylate
Green
ꢀ
1
precipitate that obtained on cooling was picked up by
powder, yield 66%, mp 208–210°C; IR (ν/cm ): 3301,
3
1
filtration and then recrystallized by heating in ethyl alcohol.
267, 3173 (NH and NH ), 1668, 1653 (C¼O); H NMR
2
ꢀ
1
Beige crystals, yield 70%, mp 280–282°C; IR (ν/cm ):
(DMSO-d ): δ/ppm = 1.21 (t, J = 7.20 Hz, 3H, CH ),
6
3
1
3
248, 3109 (NH), 1662 (br, C¼O); H NMR (DMSO-d ):
6
2
.04 (s, 3H, CH ), 4.16 (q, J = 7.20 Hz, 2H, CH ), 6.07
3
2
δ/ppm = 2.08 (s, 3H, CH ), 4.00 (s, 2H, CH ), 7.56–7.59 (s,
3
2
(s, 2H, NH ), 7.14–7.72 (m, 9H, Ar–H), 9.84 (s, 1H,
2
4H, Ar–H), 9.95 (s, 1H, NH), 11.12 (s, 1H, NH). Anal.
NH), 10.17 (s, 1H, NH), 11.53 (s, 1H, NH). MS m/z (%):
4
+
+
Calcd. for C H N O S (249.06): C, 53.00; H, 4.45; N,
11 11 3 2
39 (M +1, 38.50), 438 (M , 53.00), 428 (52.50), 408
16.86%. Found: C, 53.12; H, 4.42; N, 16.79%.
(
(
(
56.00), 398 (46.50), 390 (50.50), 368 (46.50), 342
43.00), 323 (54.50), 314 (42.00), 288 (58.39), 215
91.26), 187 (47.89), 143, (30.75), 77 (100.00). Anal.
General
acetamidophenylimino)-5-arylidene-thiazolidin-4-ones 20a–
d.
procedure
for
synthesis
of
2-(4-
To a suspension of 2-(4-acetamidophenylimino)
Calcd. for C H N O S (438.14): C, 60.26; H, 5.06; N,
22 22 4 4
thiazolidin-4-one (18) (2 mmol, 0.5 g) and 0.5 g fused
sodium acetate in 15-mL glacial acetic acid, the
appropriate aromatic aldehyde derivative (2 mmol) was
added. The reaction mixture was refluxed for 4 h and
then allowed to cool to 25°C. The solid that formed, after
dilution with cold water, was isolated by filtration. The
resulting crude product was purified by recrystallization
from EtOH/DMF mixture (1:1) to yield 20a–d.
1
2.78%. Found: C, 60.12; H, 5.00; N, 12.70%.
N -(4-Acetamidophenyl)-3-amino-5-(phenylamino)
2
thiophene-2,4-dicarboxamide (13b).
Beige powder, yield
ꢀ
1
7
3
0%, mp 230–232°C; IR ( ν /cm ): 3432, 3397, 3268,
161 (NH and NH ), 1661 (br, C¼O); H NMR (DMSO-
1
2
d ): δ/ppm = 2.01 (s, 3H, CH ), 4.98 (s, 2H, NH ), 6.93–
6
3
2
7.52 (m, 9H, Ar–H), 8.80 (s, 1H, NH), 9.32 (s, 1H, NH),
9.89 (s, 2H, NH ), 10.24 (s, 1H, NH). Anal. Calcd. for
2
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

Month 2018
Acetanilide-based Heterocycles
2
-(4-Acetamidophenylimino)-5-(4-methoxybenzylidene)-
N-(4-Acetamidophenyl)-2-(3-cyano-4,6-dimethyl-2-oxo-5-(4-
thiazolidin-4-one (20a).
Orange crystals, yield 80%, mp
tolylazo)pyridin-1(2H)-yl)acetamide (22b). Orange powder,
ꢀ
1
ꢀ1
2
94–295°C; IR ( ν /cm ): 3218, 3103 (NH), 1661 (br,
yield 67%, mp 285–286°C; IR ( ν /cm ): 3275, 3171
1
1
C¼O); H NMR (DMSO-d ): δ/ppm = 2.06 (s, 3H, CH ),
(NH), 2224 (CꢁN), 1664 (C¼O); H NMR (DMSO-d ):
6
3
6
3
.82 (s, 3H, OCH ), 7.14 (d, J = 8.00 Hz, 2H, Ar–H),
δ/ppm = 2.03 (s, 3H, CH ), 2.33 (s, 3H, CH ), 2.53 (s,
3
3
3
7.43–7.53 (m, 4H, Ar–H), 7.62 (d, J = 8.00 Hz, 2H, Ar–
3H, CH ), 2.66 (s, 3H, CH ), 5.01 (s, 2H, CH ), 7.32 (d,
3 3 2
H), 7.72 (s, 1H, CH¼C), 9.98 (s, 1H, NH), 12.30 (s, 1H,
J = 8.80 Hz, 2H, Ar–H), 7.41–7.47 (m, 4H, Ar–H), 7.67
(d, J = 8.80 Hz, 2H, Ar–H), 9.82 (s, 1H, NH), 10.39 (s,
1H, NH). Anal. Calcd. for C H N O (456.19): C,
NH). Anal. Calcd. for C H N O S (367.10): C, 62.11;
1
9 17 3 3
H, 4.66; N, 11.44%. Found: C, 62.18; H, 4.61; N, 11.49%.
25 24 6 3
2
-(4-Acetamidophenylimino)-5-(2,5-dimethoxybenzylidene)-
6
1
5.78; H, 5.30; N, 18.41%. Found: C, 65.66; H, 5.34; N,
thiazolidin-4-one (20b). Orange crystals, yield 75%, mp
8.32%.
ꢀ
1
>
300°C; IR ( ν /cm ): 3217, 3107 (NH), 1662 (br,
N-(4-Acetamidophenyl)-2-(3-cyano-4,6-dimethyl-2-oxo-5-(4-
1
C¼O); H NMR (DMSO-d ): δ/ppm = 2.05 (s, 3H,
methoxyphenyl-azo)pyridin-1(2H)-yl)acetamide
Orange powder, yield 75%, mp 289–290°C; IR (ν/cm ):
(22c).
6
ꢀ
1
CH ), 3.78 (s, 3H, OCH ), 3.84 (s, 3H, OCH ), 6.84 (s,
3
3
3
1
1
4
H, Ar–H), 6.98–7.06 (dd, 2H, Ar–H), 7.58–7.65 (m,
H, Ar–H), 7.74 (s, 1H, CH¼C), 9.99 (s, 1H, NH), 12.26
3295 (NH), 2223 (CꢁN), 1665 (C¼O); H NMR
(DMSO-d ): δ/ppm = 2.02 (s, 3H, CH ), 2.46 (s, 3H,
6 3
(s, 1H, NH). Anal. Calcd. for C H N O S (397.11): C,
CH
2H, CH
2
3
), 2.70 (s, 3H, CH
3 3
), 3.83 (s, 3H, OCH ), 5.09 (s,
2
0 19 3 4
60.44; H, 4.82; N, 10.57%. Found: C, 60.33; H, 4.85; N,
), 7.11 (d, J = 8.80 Hz, 2H, Ar–H), 7.51–7.53
1
0.49%.
(m, 4H, Ar–H), 7.84 (d, J = 8.80 Hz, 2H, Ar–H), 9.89 (s,
1H, NH), 10.47 (s, 1H, NH). Anal. Calcd. for
C H N O (472.19): C, 63.55; H, 5.12; N, 17.79%.
(
5Z)-2-(4-Acetamidophenylimino)-5-(4-nitrobenzylidene)-
thiazolidin-4-one (20c). Orange crystals, yield 70%, mp
>
25 24 6 4
ꢀ
1
300°C; IR ( ν /cm ): 3214, 3162 (NH), 1676 (br,
Found: C, 63.40; H, 5.06; N, 17.87%.
1
C¼O); H NMR (DMSO-d ): δ/ppm = 2.05 (s, 3H,
6
N-(4-Acetamidophenyl)-2-(3-cyano-4,6-dimethyl-2-oxo-5-(4-
chlorophenyl-azo)pyridin-1(2H)-yl)acetamide (22d). Brown
CH ), 7.63–7.75 (m, 4H, Ar–H), 7.77 (s, 1H, CH¼C),
3
ꢀ
1
7
2
.83 (d, J = 8.80 Hz, 2H, Ar–H), 8.34 (d, J = 8.40 Hz,
H, Ar–H), 10.21 (s, 1H, NH), 12.40 (s, 1H, NH). Anal.
powder, yield 70%, mp 268–270°C; IR (ν/cm ): 3315
+
(NH), 2224 (CꢁN), 1661 (C¼O); MS m/z (%): 476 (M ,
Calcd. for C H N O S (382.07): C, 56.54; H, 3.69; N,
34.60), 416 (100), 415 (88.67), 374 (16.73), 353 (16.93),
1
8 14 4 4
1
4.65%. Found: C, 56.71; H, 3.63; N, 14.56%.
329 (17.43), 327 (47.46). Anal. Calcd. for C H ClN O
6 3
(476.14): C, 60.44; H, 4.44; N, 17.62%. Found: C, 60.62;
H, 4.37; N, 17.74%.
Synthesis of 2-amino-1-(4-acetamidophenyl)-3-cyano-5-oxo-
24
21
2
-(4-Acetamidophenylimino)-5-(4-bromobenzylidene)-
thiazolidin-4-one (20d).
>
Brown powder, yield 65%, mp
ꢀ
1
300°C; IR ( ν /cm ): 3218, 3106 (NH), 1662 (br,
+
+
C¼O); MS m/z (%): 417 (M , Br-81, 100), 415 (M , Br-
4,5-dihydro-1H-pyrrole (27).
A mixture of 3 (1.13 g,
79, 95.38), 375 (15.99). Anal. Calcd. for C H BrN O S
5 mmol) and malononitrile (5 mmol) in EtOH (30 mL)
containing 0.5 mL triethylamine was heated under reflux
for 3 h. After cooling, the precipitate that formed was
picked up by filtration and then recrystallized from the
dioxane.
1
8
14
3 2
(415.00): C, 51.93; H, 3.39; N, 10.09%. Found: C, 51.77;
H, 3.31; N, 10.21%.
General procedure for the synthesis of N-(4-
acetamidophenyl)-2-(3-cyano-4,6-dimethyl-2-oxo-5-(substituted
ꢀ
1
phenylazo)pyridinyl)acetamides 22a–d.
A suspension of
chloroacetamide derivative 3 (5 mmol, 1.13 g), 5-arylazo-
-cyano-4,6-dimethyl-pyridin-2-one (5 mmol), and 0.70 g
Gray powder, yield 78%, mp >300°C; IR ( ν /cm ):
3328, 3225, 3185 (NH and NH
), 2175 (CꢁN), 1728,
): δ/ppm = 2.07 (s, 3H,
), 6.78 (s, 2H, NH ), 7.17 (d,
J = 8.70 Hz, 2H, Ar–H), 7.69 (d, J = 8.70 Hz, 2H, Ar–
H), 10.13 (s, 1H, NH). Anal. Calcd. for C13
2
1
3
1647 (C¼O); H NMR (DMSO-d
CH ), 3.36 (s, 2H, CH
6
of anhydrous potassium carbonate in 20-mL
dimethylformamide was heated on water bath for 4 h.
The reaction mixture was cooled and then poured onto
ice water. The precipitate that formed was filtered off and
recrystallized from dioxane to give N-substituted pyridine
derivatives 22a–d.
3
2
2
H N O
12 4 2
(256.10): C, 60.93; H, 4.72; N, 21.86%. Found: C, 60.81;
H, 4.75; N, 21.78%.
In vitro anticancer activity. The evaluation of in vitro
N-(4-Acetamidophenyl)-2-(3-cyano-4,6-dimethyl-2-oxo-5-
cytotoxicity effects of the synthesized aminoacetanilide
scaffolds was carried out against mammary gland breast
cancer (MCF-7) cell line. This cell line was obtained
from ATCC via Holding company for biological products
and vaccines (VACSERA), Cairo, Egypt. Cytotoxicity de-
terminations are based the transformation of the yellow
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bro-
mide (MTT) to a purple formazan derivative by mitochon-
drial succinate dehydrogenase in practical cells. The
phenylazopyridin-1(2H)-yl)acetamide (22a).
Red crystals,
ꢀ
1
yield 55%, mp 278–280°C; IR ( ν /cm ): 3303 (NH),
1
2
224 (CꢁN), 1666 (C¼O); H NMR (DMSO-d6):
δ/ppm = 2.02 (s, 3H, CH ), 2.55 (s, 3H, CH ), 2.64 (s,
3
3
3
9
H, CH ), 5.05 (s, 2H, CH ), 7.26–7.64 (m, 9H, Ar–H),
3 2
.90 (s, 1H, NH), 10.41 (s, 1H, NH). Anal. Calcd. for
C H N O (442.18): C, 65.15; H, 5.01; N, 18.99%.
2
4 22 6 3
Found: C, 65.22; H, 5.06; N, 19.06%.
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

E. Abdel-Latif, E. M. Keshk, A.-G. M. Khalil, A. Saeed, and H. M. Metwally
Vol 000
method of this MTT test was performed as previously
described in detail [27,30].
[16] Counihan, J. L.; Duckering, M.; Dalvie, E.; Ku, W.; Bateman,
L. A.; Fisher, K. J.; Nomura, D. K. ACS Chem Biol 2017, 12, 635.
[17] Mortlock, A. A.; Foote, K. M.; Heron, N. M.; Jung, F. H.;
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Journal of Heterocyclic Chemistry
DOI 10.1002/jhet