M. Mushtaque et al. / Journal of Molecular Liquids 241 (2017) 912–921
919
base pair of DNA as shown in Fig. 8 and made several polar and non-
polar interactions.
The oxygen atoms of compound (6) form three hydrogen bonds
with nitrogen(N2) DG10 and DG16(0.174) A, and also with oxygen(O4)
of DC11(0.356) of chain A of DNA. The hydrophobic interactions are also
shown in Fig. 8. These typical docking observations were in good agree-
ment with DNA binding experimental results. The phenomena can be
explained by considering the non-covalent interactions such as hydro-
gen bonds, van der Waals forces, electrostatic and hydrophobic bonds.
The docking energy (ΔGbinding) produced by AutoDock is sum of various
factors as:
ΔGbinding ¼ ΔGvdw þ ΔGeleþΔGh bondþΔGdesolv þ ΔGtor
Fig. 9. Percentage cells viability of (2Z, 5Z)-5-[(4-chlorophenyl)methylidene]-3-
Interestingly, it can be seen that, the sum of Vdw + Hb +
dissolvation energy is quite high as −8.38 kcal/mol. Electrostatic energy
is −0.0 kcal/mol. Torsional free energy is 1.19 kcal/mol. Finally docking
energy is −7.19 kcal/mol. It can be concluded that DNA binding is in
good agreement with docking studies.
cyclopropyl-2-(phenylimino)-1,3-thiazolidin-4-one against MCF-7 cells.
1HNMR was recorded on Bruker Spectrospin DPX 300 MHz Bruker
Spectrospin using CDCl3 as a solvent and trimethylsilane (TMS) as an in-
ternal standard. Splitting patterns are designated as follows; s, singlet;
d, doublet; m, multiplet. Chemical shift values are given in ppm. The
ESI-MS was recorded on micrOTOF-Q II 10330 Electronspray ionization
mass spectrometer (Bruker). X-ray data were collected on Bruker
SMART Apex CCD diffractometer (SAI, Universidade da Coruña).
4.3. Cell line profile
4.3.1. Cytotoxicity studies
3-(4, 5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide
(MTT) is reduced by the succinate dehydrogenase system of mitochon-
drial cells to produce water insoluble purple formazan crystals
[52–53]and measured spectrophotometrically. Since the amount of
formazan produced is directly proportional to the number of active
cells in the culture, MTT has long been used to assess cells viability in
cell proliferation and cytotoxicity [54–55]. In the present study, lead
compound (6) which was found active by molecular simulation and
bound to the base pairs of DNA, and was screened against MCF-7 cell
lines to ensure its cytotoxic effect. The lead compound (6) displayed
non-cytotoxic effect. A sub-confluent population of MCF-7 cells was
treated with increasing concentration of compound (6) and the number
of viable cells was measured after 48 h. The concentration range of com-
pound (6) was 0-320 μM. The cell viability (%) obtained with continuous
exposure for 48 h are depicted in Fig. 9.
6.1. General procedure for the synthesis of 1-isopropyl phenylthiourea
Phenylisothiocynate (1 mmol) was dissolved in toluene and then
cyclopropyl amine (1 mmol) was added slowly at room temperature.
After 5 min, the white precipitation appeared. Stirring was continued
for 1 h. The precipitate was collected by filtration, washed with toluene,
and dried to afford product (95.0%) as white powder.
6.1.1. 3-cyclopropyl-1-phenylthiourea (3)
Yield 95 %. m.p: 95–99 °C; I.R λmax (cm−1): 3265.14 (\\NH),
3010.03 (C\\H, Ar\\H), 1240.83 (C_S); 1H NMR (CDCl3) δ (ppm):
7.976 (bs, 1H, J = 7.5 Hz,\\NH), 7.684 (m, 5H, Ar\\H), 6.458(bs, 1H,
\\NH), 2.858 (bh, 1H, CH-cyclopropyl), 0.922–0.831 (m, 2H, CH2 of
cyclopropyl), 0.725 (bs, 2H, CH2 of cyclopropyl).
At 0 μM concentration, cell viability was found to be 100% but with
the increase of concentrations 10 to 320 μM, the cell viability was
found to be almost same. Hence, it can be concluded that compound
(6) is non-toxic against MCF-7 cell line.
5. Conclusion
6.2. General procedure for the synthesis of thiazolidinone
From our studies, it can be concluded that the synthesized com-
pound (6) adopted (2Z, 5Z)-configuration. Quantum chemical calcula-
tion suggested that compound (6) adopted (2Z,5Z)-configuration out
of four possible configurations. Rests of the theoretical calculations are
in good agreement with experimental results. Moreover, the docking
energy of the docked molecule with DNA (PDBID: 1BNA) was found to
be −7.19 kcal/mol. The DNA-binding constant was found to be
3.3 × 106 L mol−1. In vitro study against MCF-7 cell line suggested
that compound (6) was non-cytotoxic at the range of 10–320 μM
concentration.
(1.0 mmol) 3-cyclopropyl-1-phenylthiourea was dissolved in abso-
lute ethanol. (2.17 mmol) anhydrous sodium acetate and (1.25 mmol)
chloroacetic acid were added in a sequence. The suspension was
refluxed for 12 h. The completion of the reaction was monitored by
TLC plate. After the completion of the reaction, solvent was evaporated
on rotator vacuum. In a reaction mixture, water was added and the
aqueous layer was back-extracted with ethyl acetate. The organic
layer was washed with saturated sodium bicarbonate and brine. The
combined organic extracts were dried over Na2SO4, and concentrated
in vacuum, after the concentration light yellow product was obtained
which was crystallization in chloroform to yellow powder product
(80%).
6. Experimental protocol
All the required chemicals were purchased from Merck and Aldrich
Chemical Company (USA). Precoated aluminium sheets (Silica gel 60
F254, Merck Germany) were used for thin-layer chromatography
(TLC) and spots were visualized under UV light. Ct-DNA (as sodium
salt) was obtained from SRL Pvt. Ltd., Mumbai, India. The concentrations
of DNA were determined spectrometrically with an extinction coeffi-
cient of 6600 M−1 cm−1 at 258 nm. FT-IR spectra were recorded on
Perkin Elmer model 1600 FT-IR RX1 spectrophotometer as KBr discs.
6.2.1. (2Z)-3-Cyclopropyl-2-(phenylimino)-1,3-thiazolidin-4-one (4)
Yield 78%; m. P: 68–70 °C; I.R λmax (cm−1): 1724 (C_O), 1628
(C_N), 1592 (C_C), 1248 (Ter. amine); 1H NMR (CDCl3) δ (ppm);
7.385–7.319 (m, 2H, Ar\\H), 7.160 (t, 1H, J = 7.5 Hz, Ar\\H), 6.959–
6.931 (m, 2H, Ar\\H), 3.731(s, 2H, CH2), 2.822–2.749 (m, 1H, CH of
cyclopropyl), 1.140–1.094(m, 2H, CH2 of cyclopropyl), 1.091–1.051(m,
2H, CH2 of cyclopropyl).