280 Letters in Drug Design & Discovery, 2012, Vol. 9, No. 3
Murty et al.
5-(3-Chlorophenyl)-4H-1,2,4-Triazol-3-yl 3-[4-(2-pyrimidi-
nyl)piperazino]propyl sulfide (8j)
6XN-CH2); 2.37 (s, 3H, Ar-CH3); 2.02-1.93 (m, 2H,
CH2CH2CH2); 1.26 (t, J= 6.8 Hz, 3H, CH3CH2). IR (KBr,
cm-1): 3406 (N-H), 2930, 2823 (-C-H), 1615 (C=C), 1450,
1327, 1267 (C-N), 830, 752 (=C-H ben). ESI-MS (m/z): 346
(M+1)+.
White solid. mp 115-117 °C. 1H NMR (200 MHz, CDCl3)
ꢂ: 8.24 (d, J= 4.7 Hz, 2H, 2XAr-H); 8.06-8.00 (m, 1H, Ar-
H); 7.97-7.89 (m, 1H, Ar-H); 7.42-7.30 (m, 2H, 2XAr-H);
6.45 (t, J= 4.7 Hz, 1H, Ar-H); 3.88-3.75 (m, 4H, 2XN-CH2);
3.25 (t, J= 7.0 Hz, 2H, S-CH2); 2.59-2.34 (m, 6H, 3XN-
CH2); 1.98 (quintet, J= 7.0 Hz, 2H, CH2CH2CH2). IR (KBr,
cm-1): 3448 (N-H), 2924, 2846 (-C-H), 1587 (C=C), 1485,
1255 (C-N), 795 (=C-H ben). ESI-MS (m/z): 417 (M+1)+.
3-[4-(3-Chlorophenyl)piperazino]propyl
enyl)-4H-1,2,4-triazol-3-yl] sulfide (8e)
[5-(4-chloroph-
White solid. mp 72-74 °C. 1H NMR (200 MHz, CDCl3) ꢂ:
7.96 (d, J= 8.8 Hz, 2H, 2XAr-H); 7.33 (d, J= 8.1 Hz, 2H,
2XAr-H); 7.18-7.04 (m, 1H, Ar-H); 6.94-6.62 (m, 3H,
3XAr-H); 3.44-3.04 (m, 6H, S-CH2 & 2XN-CH2); 2.77-2.45
(m, 6H, 3XN-CH2); 2.00 (quintet, J= 7.3 Hz, 2H,
CH2CH2CH2). IR (KBr, cm-1): 3077 (C-N), 2942, 2825 (-C-
H), 1594 (C=C), 1486, 1327, 1239 (C-N), 839, 755 (=C-H
ben). ESI-MS (m/z): 448 (M+).
Cytotoxicity Studies
Cell Lines and Cell Culture
The cell lines U937 (human leukemic monocytic
lymphoma), THP-1 human acute monocytic leukemia),
Colo205 (human colorectal cancer), MCF7 (human breast
adenocarcinoma) and HL-60 (human myeloid leukemia)
were obtained from the National Centre for Cellular Sciences
(NCCS), Pune, India. Cells were cultured either in RPMI -
1640 (U937, THP-1, Colo205 and HL-60) or MEM (MCF7)
media, supplemented with 10% heat- inactivated fetal bovine
serum (FBS), 1 mM NaHCO3 , 2 mM -glutamine, 100
units/ml penicillin and 100 ꢀg/ml streptomycin. All cell lines
were maintained in culture at 37° C in an atmosphere of 5%
CO2.
5-(4-Chlorophenyl)-4H-1,2,4-triazol-3-yl 3-[4-(2-pyrimidi-
nyl)piperazino]propyl sulfide (8f)
White solid. mp 128-129 °C. 1H NMR (200 MHz, CDCl3)
ꢂ: 8.24 (d, J= 5.1 Hz, 2H, 2XAr-H); 7.99 (d, J= 8.9 Hz, 2H,
2XAr-H); 7.40 (d, J= 8.9 Hz, 2H, 2XAr-H); 6.47 (t, J= 5.1
Hz, 1H, Ar-H); 3.85-3.74 (m, 4H, 2XN-CH2); 3.23 (t, J= 7.4
Hz, 2H, S-CH2); 2.57-2.30 (m, 6H, 3XN-CH2); 1.97 (quintet,
J= 7.4 Hz, 2H, CH2CH2CH2). IR (KBr, cm-1): 3171 (N-H),
2930, 2852 (-C-H), 1586, 1493 (C=C), 1448, 1260 (C-N),
752 (=C-H ben). ESI-MS (m/z): 417 (M+1)+.
Test Concentrations
3-(4-Benzylpiperazino)propyl [5-(4-fluorophenyl)-4H-1,2,
4-triazol-3-yl] sulfide (8g)
Initially, stock solutions of each test substances were
prepared in 100% Dimethyl Sulfoxide (DMSO, Sigma
Chemical Co., St. Louis, MO) with a final concentration of 8
mg/ml. Exactly 150 ꢀl of stock was diluted to 1 ml in culture
medium to obtain experimental stock concentration of 1200
ꢀg/ml. This solution was further serially diluted with media
to generate a dilution series of 20 ꢀg/ml to 600 ꢀg/ml.
Exactly 100 ꢀl of each diluent was added to 100 ꢀl of cell
suspension (total assay volume of 200 ꢀl) and incubated for
24 h at 37 °C in 5% CO2.
1
Brown gummy liquid. H NMR (200 MHz, CDCl3) ꢂ:
8.08-7.94 (m, 2H, 2XAr-H); 7.33-7.16 (m, 5H, 5XAr-H);
7.11-6.97 (m, 2H, 2XAr-H ); 3.54 (s, 2H, Ph-CH2); 3.12 (t,
J= 6.0 Hz, 2H, S-CH2); 2.72-2.43 (m, 10H, 5XN-CH2); 1.97
(quintet, J= 6.0 Hz, 2H, CH2CH2CH2). IR (KBr, cm-1): 3405
(N-H), 2932, 2830 (-C-H), 1606 (C=C), 1455, 1326, 1226
(C-N), 847, 752 (=C-H ben). ESI-MS (m/z): 412 (M+1)+.
5-(3-Methylphenyl)-4H-1,2,4-Triazol-3-yl [3-(4-phenylpip-
erazino)propyl] sulfide (8h)
Cytotoxicity
White solid. mp 75-77 °C. 1H NMR (200 MHz, CDCl3) ꢂ:
7.88-7.74 (m, 2H, 2XAr-H); 7.37-7.10 (m, 4H, 4XAr-H);
6.89-6.70 (m, 3H, 3XAr-H); 3.27-3.05 (m, 6H, S-CH2 &
2XN-CH2); 2.62-2.45 (m, 6H, 3XN-CH2); 2.40 (s, 3H, Ar-
CH3); 1.95 (quintet, J= 7.0 Hz, 2H, CH2CH2CH2). IR (KBr,
cm-1): 3421 (N-H), 2921, 2852 (-C-H), 1599 (C=C), 1456,
1328, 1234 (C-N), 741, 685 (=C-H ben). ESI-MS (m/z): 394
(M+1)+.
Cytotoxicty was measured using the MTT [3-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide]
assay, according to the method of Mossman [27]. Briefly, the
cells (2 x 104) were seeded in each well containing 0.1 ml of
medium in 96 well plates. After overnight incubation at 37
°C in 5% CO2, the cells were treated with 100 ꢀl of different
test concentrations of test compounds (2 to 60 ꢀg) at
identical conditions with five replicates each. The final test
concentrations were equivalent to 10 to 300 ꢀg/ml or 10 to
300 ppm. The cell viability was assessed after 24 h, by
adding 10 ꢀl of MTT (5 mg/ml) per well. The plates were
incubated at 37 °C for additional three hours. The medium
was discarded and the formazan blue, which formed in the
cells, was dissolved with 100 ꢀl of DMSO. The rate of color
formation was measured at 570 nm in a spectrophotometer
(Spectra MAX Plus; Molecular Devices; supported by
SOFTmax PRO-5.4). The percent inhibition of cell viability
was determined with reference to the control values (without
test compound). The data were subjected to linear regression
analysis and the regression lines were plotted for the best
straight-line fit. The IC50 (inhibition of cell viability)
5-(3-Methylphenyl)-4H-1,2,4-Triazol-3-yl 3-[4-(2-pyridyl)
piperazino]propyl sulfide (8i)
White solid. mp 109-101 °C. 1H NMR (200 MHz, CDCl3)
ꢂ: 8.14 (dd, J= 3.8, 1.5 Hz, 1H, Ar-H); 7.94-7.74 (m, 2H,
2XAr-H); 7.42 (t, J= 7.6 Hz, 1H, Ar-H); 7.22 (t, J= 7.6 Hz,
1H, Ar-H); 7.12 (d, J= 6.8 Hz, 1H, Ar-H); 6.61-6.54 (m, 2H,
2XAr-H); 3.665-3.57 (m, 4H, 2XN-CH2); 3.17 (t, J= 6.8 Hz,
2H, S-CH2); 2.62-2.52 (m, 6H, 3XN-CH2); 2.34 (s, 3H, Ar-
CH3); 1.97 (quintet, J= 6.0 Hz, 2H, CH2CH2CH2). IR (KBr,
cm-1): 3067 (N-H), 2926, 2828 (-C-H), 1595 (C=C), 1483,
1247 (C-N), 772 (=C-H ben). ESI-MS (m/z): 395 (M+1)+.