D. He et al. / Journal of Molecular Structure 1076 (2014) 730–736
731
À1
against the normal human cell [2]. Thus we link o-nitrophenylace-
Yield 85%, m.p.: 120–121 °C. IR (KBr)
1780 (AC@O), 1526 (ANO ), 1348 (ANO
(CAN); H NMR (400 MHz, CDCl ) d: 8.14 (d, J = 7.6 Hz, 1H), 7.63
m
(cm ): 3433 (ANAC@O),
tic acid as the carrier with hydroxamic acid. These derivatives are
easier reduced and hydrolyzed in tumor cells than normal cells
because of the low pH value, the highly active reductase and
hydrolase [5], which can lead to the rapid reduction of nitro and
hydrolysis of ester. Then o-nitrophenylacetic acid may cyclize to
form a stable six-membered ring and release benzohydroxamic
acid with antitumor activity. So the conjugation of o-nitrophenyl-
acetic acid and hydroxamic acid which is easier synthesized
increases the selectivity to tumor cell. Meanwhile, the conjugation
not undergoing nitro reduction may also occur hydrolysis with
associated hydrolase, and then release hydroxamic acid.
2
2
), 1077 (NAO), 863
1
3
(d, J = 6.4 Hz, 1H), 7.62 (d, J = 6.4 Hz, 1H), 7.50 (t, J = 7.6 Hz, 2H),
7.47 (d, J = 6.0 Hz, 1H), 7.44 (d, J = 8.4 Hz, 1H), 7.34 (d, J = 8.0 Hz,
1H), 7.30 (t, J = 7.2 Hz, 2H), 7.26 (d, J = 6.4 Hz, 1H), 7.23 (d,
J = 8.8 Hz, 1H), 4.18 (s, 2H, ACH ); C NMR (100 MHz, CDCl ) d:
2 3
167.6, 165.6, 148.2, 140.0, 137.4, 137.4, 133.9, 133.5, 131.3,
1
3
131.3, 130.3, 129.3, 129.1, 129.1, 128.8, 128.4, 128.1, 128.1,
+
126.9, 125.4, 37.3. HRMS (ESI): (M + NH
428.8427), error = 2.3 ppm.
4
) 428.1008 (calculated
Here, with reference to the prospect of the potent biological
effect and the high selectivity to tumor cell resulting in the low
toxicity, 4-chloro-N-(2-(2-nitrophenyl)acetoxy)-N-phenylbenza-
mide was synthesized, and its single crystal X-ray diffraction anal-
ysis, spectroscopic and electrochemical measurements, properties,
density functional theory calculations, and in vitro biological activ-
ity assay were carried out.
Determination of the crystal structure
A colorless crystal of the title compound with dimensions of
0
.37 mm  0.35 mm  0.25 mm was selected for X-ray diffraction
analysis. The data collection was performed at 293 K on Super-
Nova, Dual, Cu at zero, Eos diffractometer with Mo K
a radiation
(k = 0.71073 Å) [8,9]. The structure was solved by direct methods
with SHELX.97 [10] and refined by SHELXL.97 [11]. All non-hydro-
gen atoms were refined with anisotropic thermal parameters. The
hydrogen atoms were placed in the calculated positions.
Experiment
Materials and measurements
Electrochemistry
Nitrobenzene, 4-chlorobenzoyl chloride, and 2-nitrophenylace-
tic acid were purchased from j&k (China). The melting point deter-
mination was performed on electrothermal PIF YRT-3 apparatus
without thermometer rectification. Infrared spectra were recorded
with a Perkin-Elmer Spectrum 2000 FTIR spectrometer in KBr pel-
Cyclic voltammetry (CV) was performed with a CHI 600 electro-
chemical workstation (Shanghai Shenhua Instrument Co., Ltd.
Shanghai, China) in a conventional three-electrode electrochemical
cell using glassy carbon as the working electrode, a saturated cal-
omel electrode as reference electrode, and a Pt slice electrode as
auxiliary electrode. The target compound solution containing
1
13
lets. H NMR and C NMR (d ppm) spectra were recorded (CDCl
3
solution) on a Varian Mercury (400 MHz) using TMS as the internal
standard. Mass spectra were recorded on a VGZAB-HS (70 eV)
spectrometer with ESI source as ionization.
À3
0.10 mol/L KCl at the concentration of 6.0 Â 10 mol/L was pre-
pared with pH 6.5 phosphate buffer. The cyclic voltammetry inves-
tigation was performed by sweeping the potential between À1.9 V
and +1 V (vs. Ag/AgCl) for one cycle at the scan rate of 100 mV/s to
record the reduction potential. The oxygen of all solutions was
removed by purging high-purity nitrogen.
Preparation of N-(4-chlorophenyl)-N-(2-(2-
nitrophenoxy)acetoxy)benzamide
Acylation reaction of 4-chlorobenzoyl chloride and the reduc-
tive matter of nitrobenzene gave compound (1) [6,7]. Compound
In vitro cytotoxicity assay
(2) (10 mmol) was slowly dropwise poured into the solution of
compound (1) (10 mmol) in 30 mL dichloromethane. The mixture
was stirred for 1.5 h and monitored by thin layer chromatography
The proliferation inhibition assay of the title compound was
tested in MCF-7, HCT-116, PC-3, A549, NCI-H460 cell lines, which
(
TLC). After filtration, the solution was concentrated under reduced
2
were maintained at 37 °C in 5% CO in dulbecco’s modification of
pressure, and then recrystallization from methanol gave the white
powder (Scheme 1). The colorless crystal of title compound suit-
able for X-ray structure analysis was obtained from dichlorometh-
ane–ethanol (v/v, 1:3) with slow evaporation at room temperature.
eagle’s medium dulbecco (DMEM), supplemented with 10% fetal
calf serum. Exponentially growing cells, representing an asynchro-
nous population, were seeded in 96-well plates, at the density of
5000–10,000 cells per well while the marginal wells should be
O
Cl
H
N
NO2
OH
Cl
a
b
O
Cl
O
HO
N
O
O
NO
N
2
c
2
Cl
O
NO
Cl
2
1
3
Scheme 1. Synthetic route of 4-chloro-N-(2-(2-nitrophenyl)acetoxy)-N-phenylbenzamide. Reagents and conditions: (a) NH2NH2ÁH2O, Raney nickel, CH3CH2OH/
ClCH2CH2Cl (3:2), 0 °C; (b) tetrahydrofuran, NaHCO3, 0 °C; (c) CH2Cl2, NaHCO3, rt.