Synthesis, antitumor, and DNA binding behavior of novel 4-(2-Hydroxyquinolin-3-yl)-6-Phenyl-5, 6 dihydropyrimidin derivatives in aqueous medium

Article abstract of DOI:10.1080/15257770.2010.496275

This article deals with the synthesis of 4-(2-hydroxyquinolin-3-yl)-6- phenyl-5,6-dihydropyrimidin derivatives (2a-f), on condensation with various aromatic aldehydes and ketones in aqueous ethanolic NaOH solution yielding the corresponding chalcones (3). These chalcones were further reacted with thiourea/urea in the presence of a base, which led to the formation of the titled derivatives (2a-f). The newly synthesized heterocyles were characterized by elemental analysis, FTIR, 1HNMR, and electronic and mass spectral data. The compounds (2a and 2b) were evulated for in vitro cyctotoxicity against human breast adenocarcinoma cell (MCF-7). In MTT cytotoxicity studies, both quinolinde derivatives were found most effective. The binding interaction behavior of the compound (2a) and (2d) with calf thymus-DNA (CT-DNA) was studied by electronic spectra, viscosity measurements, and thermal denaturation studies. On binding to CT-DNA, the absorption spectrum underwent bathochromic and hypochromic shifts. The binding constant (Kb) observed 4.3 × 10⁵ M^-1 for (2a), and 3.8 × 10⁵ M^-1 for (2d) suggested that compound (2a) binds more strongly with base pairs than (2d).

Full text of DOI:10.1080/15257770.2010.496275

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Synthesis, Antitumor, and DNA
Binding Behavior of Novel 4-(2-
Hydroxyquinolin-3-yl)-6-Phenyl-5, 6
Dihydropyrimidin Derivatives in Aqueous
Medium
Devappa S. Lamani a , K. R. Venugopala Reddy a , H. S. Bhojya Naik a
, K. S. R. Pai b , Ravishankar Kumar b , Fasiulla c , H. R. Prakash Naik
a & L. R. Naik d
a Department of Studies in Industrial Chemistry, Jnana Sahyadri
Kuvempu University, Shankaraghatta, Shimoga District, Karnataka,
India
b Department of Pharmacology, Manipal College of Pharmaceutical
Sciences, Manipal University, Manipal, Karnataka, India
c Department of Chemistry, Manipal Institute of Technology, Manipal,
Udupi District, Karnataka, India
d Department of Physics, Karnataka University, Dharwad, India
Version of record first published: 19 Jul 2010.
To cite this article: Devappa S. Lamani , K. R. Venugopala Reddy , H. S. Bhojya Naik , K. S. R. Pai ,
Ravishankar Kumar , Fasiulla , H. R. Prakash Naik & L. R. Naik (2010): Synthesis, Antitumor, and DNA
Binding Behavior of Novel 4-(2-Hydroxyquinolin-3-yl)-6-Phenyl-5, 6 Dihydropyrimidin Derivatives in
Aqueous Medium, Nucleosides, Nucleotides and Nucleic Acids, 29:8, 591-605
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Nucleosides, Nucleotides and Nucleic Acids, 29:591–605, 2010
Copyright ꢀC Taylor and Francis Group, LLC
ISSN: 1525-7770 print / 1532-2335 online
DOI: 10.1080/15257770.2010.496275
SYNTHESIS, ANTITUMOR, AND DNA BINDING BEHAVIOR OF NOVEL
4-(2-HYDROXYQUINOLIN-3-YL)-6-PHENYL-5, 6 DIHYDROPYRIMIDIN
DERIVATIVES IN AQUEOUS MEDIUM
Devappa S. Lamani,1 K. R. Venugopala Reddy,1 H. S. Bhojya Naik,1
K. S. R. Pai,2 Ravishankar Kumar,2 Fasiulla,3 H. R. Prakash Naik,1
and L. R. Naik4
1Department of Studies in Industrial Chemistry, Jnana Sahyadri Kuvempu University,
Shankaraghatta, Shimoga District, Karnataka, India
2Department of Pharmacology, Manipal College of Pharmaceutical Sciences,
Manipal University, Manipal, Karnataka, India
3Department of Chemistry, Manipal Institute of Technology, Manipal, Udupi District,
Karnataka, India
4Department of Physics, Karnataka University, Dharwad, India
This article deals with the synthesis of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5,6-
2
dihydropyrimidin derivatives (2a–f), on condensation with various aromatic aldehydes and
ketones in aqueous ethanolic NaOH solution yielding the corresponding chalcones (3). These
chalcones were further reacted with thiourea/urea in the presence of a base, which led to the
formation of the titled derivatives (2a–f). The newly synthesized heterocyles were characterized
1
by elemental analysis, FTIR, HNMR, and electronic and mass spectral data. The compounds
(2a and 2b) were evulated for in vitro cyctotoxicity against human breast adenocarcinoma cell
(MCF-7). In MTT cytotoxicity studies, both quinolinde derivatives were found most effective. The
binding interaction behavior of the compound (2a) and (2d) with calf thymus-DNA (CT-DNA)
was studied by electronic spectra, viscosity measurements, and thermal denaturation studies. On
binding to CT-DNA, the absorption spectrum underwent bathochromic and hypochromic shifts. The
binding constant (Kb) observed 4.3 × 105 M−1 for (2a), and 3.8 × 105 M−1 for (2d) suggested
that compound (2a) binds more strongly with base pairs than (2d).
Keywords Quinoline; Claisen Schmidt condensation; antitumor activity; MTT assay,
DNA binding; viscosity measurements; aqueous medium
Received 23 September 2008; accepted 21 May 2010.
D. S. L. thanks Indian Institute of Science Bangalore, Karnataka University Dharwad for providing
spectral data. K. R. V. R. would like to thank associate professor Dr. Sreedhra Pai, Department of
Pharmacology, Manipal College of Pharmaceutical Sciences, Manipal University for the in vitro antitumor
activity; project students, Shilashree, Sindura, Vinaya, and Preyadhrshini Sathish; and Kuvempu University
for awarding the research fellowship.
Address correspondence to K. R. Venugopala Reddy, Department of Studies in Industrial Chemistry,
Jnana Sahyadri Kuvempu University, Shankaraghatta–577 451, Shimoga District, Karnataka, India. E-mail:
venurashmi30@rediffmail.com
591

592
D. S. Lamani et al.
INTRODUCTION
Quinoline derivatives are useful heterocyclic aromatic compounds widely
used in medicinal chemistry. Additionally, quinoline derivatives have been
extensively studied as potential antitumor agents, as they are capable of
binding to DNA.[1] Many 4-hydroxy-1,2-dihydro-2-quinolinones have a wide
spectrum of pharmacological applications such as anti-herpes simplex virus
(anti-HSV),[2] anti-convulsion,[3] and anti-inflammatory[4] activities. Further,
the utility of quinoline derivatives in the preparation of some dyes and pig-
ments has been reported.[5] In addition, quinolines are valuable synthons
used for the preparation of nano and mesostructures with enhanced elec-
tronic and photonic properties.[6] Due to wide range of applicability in
the fields of medicine, bioorganics, industry, and synthetic organic chem-
istry, there has been an increasing interest in the development of efficient
methodologies for the synthesis of quinoline derivatives.
On other hand, chalcone are important starting materials for the syn-
theses of different classes of heterocyclic compounds such as pyrazolines,
thiophenes, and pyrimidines. The synthesis of novel pyrimido-quinoline
derivatives by a convenient method remains an important synthetic task
because of their great importance in pharmacological field.[7–10]
There is evidence that anticancer activity was due to the intercalation
between the drug and the base pairs of DNA and interference with nor-
mal functioning of the enzyme topoisomerase II involved in the breaking
and releasing of DNA strands.[11] In recent years, various fused systems such
as thiophene,[12] furan and pyridine analogues of ellipticine,[13] and ben-
zothiazoloquinoline[14] have been studied for their intercalative properties.
Recently, Cao et al. studied DNA affinity properties of Safranine T, which fea-
tures a planar phenazine ring and have shown that the electrostatic binding
plays an important role in the intercalation of safranine T.[12] The results
of these various binding studies have been useful in designing new and
promising anticancer agents for clinical use.[15]
The recent interest in green chemistry has posed a new challenge for
organic synthesis to find new reaction conditions that reduce the emission of
volatile organic solvents and the use of hazardous toxic chemicals. Organic
reactions in aqueous media have attracted increasing interest currently be-
cause of environmental issues and the understanding of biochemical pro-
cesses. Water as reaction solvent offers many practical and economic advan-
tages including low cost, safe handling, and environmental compatibility. Re-
cently, many organic reactions in aqueous media have been described in the
literature.[16] Hence in continuation of our research program, [17–20,31,32,34]
we described a highly efficient, cost-effective, and environmentally method
for the synthesis of new series of novel quinoline derivatives (2a–f). The
newly synthesized selected compounds were used for in vitro antitumor and
DNA binding studies.

Novel Quinoline Derivatives
RESULTS AND DISCUSSION
593
The chemistry of title derivatives (2a–f) performed following steps as
per the (Scheme 1). The structural elucidation of the newly synthesized
compounds was established on the basis of their elemental analysis, infrared
(IR), 1H NMR, and mass spectral data.
The IR spectra of the compound (2a–f) showed the absence of aldehyde
carbonyl stretching frequency, instead it gave a band at 1657–1720 cm−1 and
two broad peaks observed in the region of 3401 cm−1 3268 cm−1 due to –OH-
and –NH- groups, respectively. The generation of new sharp absorption
band at 1290–1293 cm−1, which attributed to tautomeric form of (C=S)
O
O
R
H
H 3
C
+
N
O H
1
2
H 2
O
60-65 0 C
NaOH/Ethanol
N
O H
R
O
3
N H 2
O
N
H 2
N
S
N H 2
E than ol/ dil K O H
E than ol/d ilK O H
H 2
N
O H
N
O H
N
S
N
O
R
R
N H
N H
2 a -c
R = H , C H 3, O C H 3
2 d -f
N
O H
N
O H
N
O H
R
N
S H
N
R
N
SCHEME 1 Synthesis of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5,6-dihydropyrimidin derivatives.

594
D. S. Lamani et al.
stretching frequency. [17,18] Further structure (2a) was confirmed by 1H-NMR
spectra show a broad peak δ: 11.72–11.70 due to (Ar-OH-) second position
of quinoline (2a) and resonate peak δ: 11.32–11.23 ppm, corresponds to
the cyclized (-NH-pyrimidene). The signal exhibits multiplets at δ: 7.18–8.00
pp-m for aromatic protons,[19,20] and its mass spectra having molecular ion
peak at m/z = 333[M + H]. The obtained elemental analysis values are
in good agreement with theoretical data. Hence, we synthesized six more
title compounds, which exhibited similar spectral data, were summarized in
experimental section.
IN VITRO ANTITUMOR ACTIVITY
In vitro cytotoxicity of synthesized compounds was assessed by standard
MTT bioassay in MCF-7 cancer cells at 24 hours of test compounds exposure.
The IC50 (concentration required to inhibit 50% of cell growth) values for
derivatives (2a and 2d) were 50.1 and 76.36 µg/ml at 24 hours, respectively.
The percentage of DMSO (0.1%) used in the experiment did not affect the
growth of the cells. In the MTT time course study the compounds showed
dose-dependent and time dependent activities. The IC50 values for deriva-
tives (2a) and (2d) were 43.35, 59.56 µg/ml and 23.08, 26.82 µg/ml at 48
and 78 hours, respectively. In this study, MTT assays revealed substantial
cytotoxicity in MCF-7 cell with increasing exposure to test compounds, the
IC50 values of compounds at 48 and 72 hours were significantly reduced as
compared with 24 hours values. The reduction in the cytotoxicity activity of
test compound with increased exposure could be related to degradation of
the same in aqueous media. The resulting protocol shows that the in vitro an-
titumor activity of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5,6-dihydropyrimidin
derivatives exhibited significant cytotoxicity against breast adenocarcinoma
cells (MCF-7). It is clear from these data the compound (2a) has the high-
est levels of cytotoxicity activity than that of the compound (2d).[21] Many
anticancer drugs are effective against MCF-7 by causing apoptosis through
the expression of caspase-3, generacting reactive oxygen species (ROS) and
damaging DNA.[22] (See Tables 1–3.)
TABLE 1 In vitro cytotoxic activity of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5,6-dihydropyrimidine against
MCF-7 cells (human breast adenocarcinoma cells) by MTT assay at 24 hours of exposure
% Cytotoxicity
Treatment
20 µg/ml
40 µg/ml
60 µg/ml
80 µg/ml
100 µg/ml
IC50
2a
2d
38.69 0.57a 44.71 0.13a 55.76 0.37a 56.94 1.49a
63.15 1.29a
60.50 1.15a
50.10
76.36
45.61 1.84a 31.38 1.38a 33.57 1.2a 53.57 2.71a
aAll the values are mean SEM of three samples.

Novel Quinoline Derivatives
595
TABLE 2 In vitro cytotoxic activity of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5,6-dihydropyrimidine against
MCF-7 cells (human breast adenocarcinoma cells) by MTT assay at 48 hours of exposure
% Cytotoxicity
Treatment
20 µg/ml
40 µg/ml
60 µg/ml
80 µg/ml
100 µg/ml
IC50
2a
2d
20.32 0.65a 28.95 0.44a 84.33 0.49a 89.26 0.30a
35.50 0.21a 31.66 1.24a 49.77 0.49a 57.77 2.37a
90.11 0.55a
60.19 0.27a
43.35
59.56
aAll the values are mean SEM of three samples.
DNA BINDING STUDIES (ABSORPTION SPECTRAL STUDIES)
The application of electronic absorption spectroscopy in calf-thymus
DNA (CT-DNA) binding studies is one of the most important techniques.[23]
The binding of the molecules to DNA has been well characterized by the
large hypochromism. After intercalation the π∗ orbital of compounds could
couple with π orbital of base pairs, thus decreasing the π∗—π transition
of energy, and resulting bathochromism. Hence, the decrease the absorp-
tion intensity and significant red shift due to stacking interaction between
drug and CT-DNA.[24,25] The DNA binding studies were characterized by
absorbance maximum at 329 nm for (2a) and 259 nm for (2d). The addi-
tion of increasing higher concentration of DNA led to hypochromic and
bathochromic (red shift) changes in its visible absorption spectra as a result
of formation of more stable complexes (Figures 1 and 2). The interaction of
(2a) and (2d) with CT-DNA resulted in the decrease of absorption intensity
accompanied by a shift toward higher wavelengths (∼3 and 5 nm). Around
9–12% reduction intensity of absorption was observed at 329 and 259 nm
peak maximum in the presence of an excess of calf thymus DNA.[26] The
lowest observation value observed in spectral changes (including red shift,
hypochromicity) were used to evaluate intrinsic binding constant (Kb), it
observed value 4.3 × 105 M−1 for (2a) and 3.8 × 105 M−1 for (2d) from the
spectral result suggested that compound (2a) bind more strongly with base
pairs than that (2d).[27,28]
TABLE 3 In vitro cytotoxic activity of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5,6-dihydropyrimidine against
MCF-7 cells (human breast adenocarcinoma cells) by MTT assay at 72 hours of exposure>
% Cytotoxicity
Treatment 20 µg/ml
40 µg/ml
60 µg/ml
80 µg/ml
100 µg/ml
IC50
2a
2d
40.09 0.80a
56..6 0.14a 72.61 0.27a 77.27 2.33a
81.23 0.59a
79.27 0.62a
23.08
26.82
43.80 1.62a 54.05 2.32a 61.41 1.91a 66.53 0.42a
aAll the values are mean SEM of three samples.

596
D. S. Lamani et al.
FIGURE 1 Ultraviolet-absorption spectra in Tris-HCl buffer on addition CT-DNA (2a) [DNA] = 0.5 µm,
= 10 µm, drug, 20 µm; 30 µm; 40 µm; 50 µm; Arrow indicates the absorbance changing on the increase
of DNA concentration.
Viscosity Measurements
To further clarify the interaction modes of (2a) and (2d) with DNA were
investigated by viscosity measurements. An increase in viscosity of native
DNA is regarded as a diagnostic feature of an intercalation process.[29,30] We
have measured the viscosity changes in short, rod-like DNA fragments. The
relative length increase (L/L0) of the complex formed between (2a), (2d),
and DNA is shown in Figure 3. It is evident that binding of (2a) and (2d)
increased the viscosity of DNA corresponding to an increase in the contour
length of the DNA fragments. In order to elucidate the binding mode of
the present compound, the viscosity measurements were carried out on
CT-DNA by varying the concentration of added compound. The effects of
the compounds on the viscosity of rod-like DNA were shown (Figure 3). The
presence of compound had an obvious effect on relative viscosity of CT-DNA
with an increase in concentration of the added compounds.[31]
Thermal Denaturation
Other strong evidence for the intercalative binding of (2a) and (2d)
into the double helix DNA was obtained from DNA melting studies. The

Novel Quinoline Derivatives
597
FIGURE 2 Ultraviolet-absorption spectra in Tris-HCl buffer on addition CT-DNA (2d) [DNA] = 0.5
µm, = 10 µm, drug, 20 µm; 30 µm; 40 µm; 50 µm; Arrow indicates the absorbance changing on the
increase of DNA concentration.
intercalation of small molecules into the double helix is known to increase
the DNA melting temperature (Tm), at which the double helix denatures
into single stranded DNA, owing to the increased stability of the helix in
the presence of an intercalator.[32] The molar extinction coefficient of DNA
bases at 260 nm in the double helical form is much less than the single
stranded form; hence, melting of the helix leads to an increase in the ab-
sorbance at 260 nm. The DNA melting studies were carried out with CT-DNA
in the absence and presence of (2a) and (2d). Tm (melting temperature)
for CT-DNA was 60 5◦C in the absence of compounds, but in the presence
of (2a) and (2d) the Tm of CT-DNA increased by 7–10◦C. These variations
in DNA melting temperature strongly supported the intercalation of com-
pounds into the double helix DNA (Figure 4).[33]
EXPERIMENTAL SECTION
Materials and Methods
All organic solvents used for the synthesis were of analytical grade.
The thin layer chromatography (TLC) was performed on Baker-Flex sil-
ica gel 1B-F (1.55) plates using ethyl acetate and petroleum ether (1:8).

598
D. S. Lamani et al.
compound (2a)/[DNA]
compound (2d)/[DNA]
1.18
1.16
1.14
1.12
1.10
1.08
1.06
1.04
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
Compound (2a), (2d)/[DNA]
FIGURE 3 Effect of increasing amount of the (2a) and (2d) on the relative viscosities of CT-DNA, at
25◦C.
Melting point was determined on a Mel-Temp apparatus and were un-
corrected. IR spectra were recorded in the matrix of KBr with Perkin-
1
Elmer 1430 spectrometer. H NMR spectra were recorded on Jeol spec-
trometer (400 MHz), and chemical shifts (δ) given in ppm relative to
the TMS in CDCl3 solvent. Mass spectra were recorded by electron ion-
ization (EI) on a finnigan MAT 312 spectrometer. C, H, and N analysis
were performed at Cochin University, Sophisticated Test & Instrumentation
Center, Kochi, Kerala, India. Ammonium hexaflurophosphate (NH4PF6)
purchased from Qualigens (India). Tris-HCl buffer (5 mM Tris-HCl, 50
mM NaCl, pH = 7.2, Tris = Tris(hydroxymethyl) amino methane) so-
lution was prepared using deionized double distilled water. CT-DNA was
purchased from Bangalore Gene, Bangalore, India. Ultraviolet absorption
spectra were determined in a Perkin-Elmer (Germany) model 554, UV-
Vis recording spectrophotometer using quartz cuvettes of 10 mm path
-length.
MTT Assay in Cultured Human Cancer Cells
Cell line: Human breast adenocarcinoma cells (MCF-7) were received
from NCCS Pune, India. MEM (minimum essential medium) media was sup-
plemented with 10% fetal bovine serum (FBS), 1% l glutamine, and 50 ug/ml

Novel Quinoline Derivatives
599
[DNA]
(2a)/[DNA]
(2d)/[DNA]
1.25
1.20
1.15
1.10
1.05
1.00
0
20
40
60
80
100
Temperature 0C
FIGURE 4 Melting cures of CT-DNA in the presence and absences of (2a) and (2d).
gentamicin sulphate in a CO2 incubator (NuAire, dPlymouth, USA) in a hu-
midified atmosphere of 5% CO2 and 95% air. MTT (3-(4,5-dimethylthiazol-
2-yl)-2,5-diphenyltetrazolium bromide) reagent was received from Sigma
Aldrich (USA). Tissue culture flasks, 96-well microculture plates were from
Tarson and Nunc (USA). Gentamycin was obtained from KMC Hospital,
Manipal, India.
In Vitro Cytotoxic Activities (MTT Assay)
In vitro cytotoxicity was determined using a standard MTT assay[34,35]
with protocol appropriate for the individual test system. In brief, exponen-
tially growing cells were plated in 96-well plated (1 × 104 cells/well in 100 µl
of medium) and incubated for 24-hour attachment. Test compounds were
prepared prior to the experiment by dissolving 0.1% DMSO and diluted with
medium. The cells were then exposed to different concentrations of test com-
pounds (20–100 µg/ml) in the volume of 100 µl/well. Cells in the control
wells received the same volume of medium containing 0.1% DMSO After 24
hours, the medium was removed and cell cultures were incubated with 100 µl
MTT reagent (1mg/ml) for 4 hours at 37◦C. The formation of the viable cells
was solubilized by the addition of 100 µl DMSO. The suspension was placed
on micro-vibrator for 5 minutes and absorbance was recorded at 540 nm

600
D. S. Lamani et al.
by the ELISA reader (BIOTEK-ELx800). The experiment was performed in
triplicate. The percentage of growth inhibition was calculated with respect to
vehicle control using the formula,% Inhibition = [{(Control absorbance −
Blank absorbance) − (Test absorbance − Blank absorbance)}/(Control
absorbance − Blank absorbance)] × 100. For MTT time course study, MCF-7
cells (1 × 104 cells/well) seeded in 96-well plates were exposed to different
concentrations of test compounds (20–100 µg/ml). The percentage cytotox-
icity and IC50 values were determined at 48 and 72 hours of test compound
incubation. Data obtained were expressed as mean SEM.
UV-Visible Absorption Studies
The concentration of CT-DNA per nucleotide [C(p)] was measured us-
ing its known extinction coefficient at 260 nm (6600 M−1cm−1).[36] The
absorbance at 260 nm (A260) and at 280 nm (A280) for CT-DNA was mea-
sured to check purity. The ratio A260/A280 was found to be 1.8–1.9, indicating
that CT-DNA was satisfactorily free from protein. Buffer [5mM tris (hydrox-
ymethyl) aminomethane, pH 7.2, 50mM NaCl] was used for the absorption,
viscosity, and thermal denaturation studies.
Absorption titration experiments were carried out by varying DNA con-
centration (0–100 µM) and maintaining the compound concentration con-
stant (30 µM). Absorption spectra were recorded after each successive ad-
dition of DNA and equilibration (approximately 10 minutes). For both the
compound (2a) and (2d) observed data were then fit into (1) in order to
obtain the intrinsic binding constant Kb.
[DNA]/(εa − ε f ) = [DNA]/(εb − ε f ) + 1/Kb(εb − ε f ),
(1)
where εa,ε, and εb are the apparent, free, and bound compound extinction
coefficients at 329 nm for (2a) and 256 nm for (2d). A plot of [DNA]/
(εa−ε) versus [DNA] gave a slope of 1/(εb−ε) and an intercept y equal to
1/Kb(εb-ε), where Kb is the ratio of the slope to the intercept y.[37]
Viscosity Measurements
Viscosity measurements were carried out using a semimicro dilution
capillary viscometer at room temperature. Each experiment was performed
three times and an average flow time was calculated. Data were presented as
(η/ηo) versus binding ratio, where η is the viscosity of DNA in the presence
of complex and ηo is the viscosity of DNA alone.[38]
Thermal Denaturation
Melting studies were carried out by monitoring the absorption of CT-
DNA (50 µM) at 260 nm at various temperatures in the presence (5–10 µM)
and the absence of each complex. The melting temperature (Tm), at which

Novel Quinoline Derivatives
601
50% of double-stranded DNA becomes single-stranded and the curve width
(σT), the temperature range between 10% and 90% noticed absorption
increases occurred and calculated as reported.[39]
GENERAL SYNTHESIS
Synthesis of (2)-1-(2-hydroxyquinolin-3-yl)-
3-phenylprop-2-en-1-one (3)
To a mixture of ketone (2) (1.63 mL 0.014 mol) and the appropriate
aldehyde (1) (2.55g 0.014 mol) in oxygen-free ethanol (25 mL) was added a
solution of sodium hydroxide in distilled water (5 mL) with constant stirring
at 0◦C. The reaction mixture was stirred at 60
5◦C for and additional
2 hours on a magnetic stirrer at room temperature. The completion of
reaction was checked by TLC and poured in ice-cold water; the solid mass
that separated out was filtered, washed with water, and crystallized from
ethyl acetate to furnish the desired product as yellowish crystals. Yield 79%
180–182◦C
Synthesis of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5,6-
dihydropyrimidine-2(1H)-thione(2a)
A mixture of (1.55 g 0.005 mol) required chalcone (3), (0.42 g 0.0056
mol) thiourea and KOH (0.66 g) in 20 mL ethanol was heated under reflux
for 6–8 hours. The progress of the reaction was monitored by TLC. The reac-
tion mixture was poured onto crushed ice; the greenish yellow solid product
was separated by neutralizing diluted HCl and recrystalized from ethanol.
Yield 83%, m.p. 215–217◦C, IR (KBr): 1290 (C=S), 3401 (OH-quinoline),
3256, (-NH pyrimidine), 2988 (Ar-CH stretching), 1568 (C=N), 1415 (C=C
ring skeleton pyrimidine moiety), 1H NMR (400 MHz, CDCl3) 11.72 (s, 1H,
OH quinoline), 11.32 (s, 1H, NH pyrimidine), 7.18–7.53 (m, 5H, 4H, 5H,
6H, 7H, and 8H, Ar-H quinoline). 3.67 (d, 1H, -CH-), 4.23 (d, 2H, CH2),
7.85–8.00 (m, 5H, Ar-H, phenyle). Mass spectra m/z = 333[M++H]. Anal.
(%) for C19H15N3OS, Calcd. C, 68.45; H, 4.53; N, 12.60; S, 9.62; Found: C,
68.47; H, 4.44; N, 12.56; S, 9.68.
Synthesis of 4-(2-hydroxy-6-methylquinolin-3-yl)-6-phenyl-5,6-
dihydropyrimidine-2(1H)-thione(2b)
Yellowish solid with yield 78%, m.p. 250–252◦C, IR (KBr): 1290 (C=S),
3403 (OH- quinoline), 3255(-NH pyrimidine), 2982, (Ar-CH stretching),
1
1566 (C=N), 1417 (C=C ring skeleton pyrimidine moiety), H NMR (400
MHz, CDCl3) 11.72 (s, 1H, OH Quinoline), 11.32 (s, 1H, NH pyrimidine),
3.22, (s, 3H, CH3), 3.64 (d, 1H, -CH-), 4.26 (d, 2H, CH2), 7.15–7.52 (m, 4H,
4H, 5H, 7H, and 8H, Ar-H quinoline). 7.85–8.05 (m, 5H, Ar-H, phenyle).

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D. S. Lamani et al.
Mass spectra m/z = 347[M+H]. Anal. (%) for C20H17N3OS, Calcd. C, 69.14;
H, 4.93; N, 12.09; S, 9.23; Found: C, 69.17; H, 4.94; N, 12.02; S, 9.28.
Synthesis of 4-(2-hydroxy-6-methoxyquinolin-3-yl)-
6-phenyl-5,6-dihydropyrimidine-2(1H)-thione (2c)
Yellowish solid with yield 93%, m.p. 238–240◦C, IR (KBr): 1290 (C=S),
3400 (OH- quinoline), 3255(-NH pyrimidine), 2987, (Ar-CH stretching),
1
1566 (C=N), 1415 (C=C ring skeleton pyrimidine moiety), H NMR (400
MHz, CDCl3) 11.72 (s, 1H, OH quinoline), 11.33 (s, 1H, NH pyrimidine),
3.86 (s, 3H, -OCH3), 3.64 (d, 1H, -CH-), 4.25 (d, 2H, CH2), 7.15–7.53 (m, 4H,
4H, 5H, 7H, and 8H, Ar-H quinoline). 7.85–8.05 (m, 5H, Ar-H, phenyle).
Mass spectra m/z = 363 [M+H].Anal. (%) for C20H17N3OS, Calcd. C, 66.10;
H, 4.71; N, 11.56; S, 8.82; Found: C, 66.17; H, 4.74; N, 11.51; S, 8.88.
Synthesis of 4-(2-hydroxyquinolin-3-yl)-6-
phenyl-5,6-dihydropyrimidin-2(1H)-one (2d)
Brown solid with yield 93%, m.p. 225–226◦C, IR (KBr): 1690 (C=O),
3401 (OH- quinoline), 3258 (-NH pyrimidine), 2982, (Ar-CH stretching),
1
1569 (C=N), 1414 (C=C ring skeleton pyrimidine moiety), H NMR (400
MHz, CDCl3) 11.70 (s, 1H, OH quinoline), 11.33 (s, 1H, NH pyrimidine),
7.18–7.52 (m, 5H, 4H, 5H, 6H, 7H, and 8H, Ar-H quinoline). 3.63 (d, 1H,
-CH-), 4.26 (d, 2H, CH2), 7.87–8.02 (m, 5H, Ar-H, phenyle). Mass spectra
m/z = 318 [M+H]. Anal. (%) for C19H15N3O2, Calcd. C, 71.91; H, 4.76; N,
13.24; Found: C, 71.96; H, 4.69; N, 13.29.
Synthesis of 4-(2-hydroxy-6-methylquinolin-3-yl)-6-phenyl-5,6-
dihydropyrimidin-2(1H)-one(2e)
Dark brown solid with yield 93%, m.p. 246–248◦C, IR (KBr): 1690
(C=O), 3405 (OH- quinoline), 3253 (-NH pyrimidine), 2989 (Ar-CH stretch-
1
ing), 1568 (C=N), 1416 (C=C ring skeleton pyrimidine moiety), H NMR
(400 MHz, CDCl3) 11.72 (s, 1H, OH quinoline), 11.30 (s, 1H, NH pyrimi-
dine), 3.20 (s, 3H, CH3), 3.66 (d, 1H, -CH-), 4.24 (d, 2H, CH2), 7.18–7.52
(m, 5H, 4H, 5H, 7H, and 8H, Ar-H quinoline). 7.87–8.02 (m, 5H, Ar-H,
phenyle). Mass spectra m/z = 331[M+].Anal. (%) for C20H17N3O2, Calcd.
C, 72.49; H, 5.17; N, 12.68; Found: C, 72.37; H, 5.20; N, 12.66.0
Synthesis of 4-(2-hydroxy-6-methoxyquinolin-3-yl)-6-
phenyl-5,6-dihydropyrimidin-2(1H)-one (2f)
Dark brown solid with yield 93%, m.p. 240–242◦C, IR (KBr): 1690
(C=O), 3401 (OH- quinoline), 3253 (-NH pyrimidine), 2989, (Ar-CH
stretching), 1569 (C=N), 1416 (C=C ring skeleton pyrimidine moiety), 1H

Novel Quinoline Derivatives
603
NMR (400 MHz, CDCl3) 11.70 (s, 1H, OH quinoline), 11.30 (s, 1H, NH
pyrimidine), 3.82 (s, 3H, -OCH3), 3.66 (d, 1H, -CH-), 4.26 (d, 2H, CH2),
7.18–7.53 (m, 5H, 4H, 5H, 7H, and 8H, Ar-H quinoline). 7.87–8.02 (m, 5H,
Ar-H, phenyle). Mass spectra m/z = 347 [M+]. Anal. (%) for C20H17N3O3,
Calcd. C, 69.15; H, 4.93; N, 12.10; Found: C, 69.20; H, 4.91; N, 12.06.
CONCLUSIONS
The synthetic route adopted for synthesis of quinoline derivatives (2a–f)
was very simple and yielded good yield. The preliminary antitumor studies
of 4-(2-hydroxyquinolin-3-yl)-6-phenyl-5, 6-dihydropyrimidin derivatives ex-
hibited significant cytotoxicity against breast adenocarcinoma cells (MCF-7)
growth in vitro. The obtained protocol shows that compound (2a) exhibits
more prominent antitumor activity than that of compound (2d). In DNA
binding studies, indicate hypochromicity and bathochromic shifts of the
compounds (2a) and (2d) when it binds with base pairs of calf thymus-DNA.
The binding constant values of 4.3 × 105 M−1 for (2a) and 3.8 × 105 M−1 for
(2d) suggested that the compound (2a) binds more avidly to CT-DNA than
(2d). In addition, increasing viscosity of sonicated rod-like DNA fragments
and the melting temperature of CT-DNA, in the presence of compound
solutions supports the binding mode.
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