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Journal Name
New Journal of Chemistry
DOI: 10.1039/C5NJ02237A
on a Flash EA 1112 series CHNSꢀO analyzer. The completion of the
reaction was checked by thin layer chromatography (TLC) with
readymade aluminium sheets (Merck F254). The names of the
structures were mentioned as per chemdraw.
AFS
PZA
Streptomycin
ꢀꢀ
ꢀꢀ
ꢀꢀ
ꢀꢀ
ꢀꢀ
ꢀꢀ
<10
ꢀꢀ
ꢀꢀ
ꢀꢀ
3.12
6.25
ABS; antibacterial standard Ciprofloxacin; AFS; antifungal standard
Fluconazole; PZA; antiꢀtb standard Pyrazinamide; ꢀꢀ: not detected
inhibition; control; dimethylsulfoxide
A mixture of 1,3ꢀdisubstituted pyrazoleꢀ4ꢀcarbaldehyde
(1aꢀe) (0.01mol), acetyl derivative (2, 3 and 4) (0.91 g, 0.01mol) in
ethanol (10 ml) were stirred in the presence of 10% sodium
hydroxide solution (2 ml) at an ambient temperature for 5 h. The
resultant yellow color reaction mass was filtered and washed with 5
ml of ethanol to get chalcone intermediate (5aꢀe), (7aꢀe) and (9aꢀe)
respectively in reasonably good yields (80ꢀ95%). The compound
(5aꢀe), (7aꢀe) and (9aꢀe) (0.005 mol) was taken in ethanol (10 ml)
and added excess amount of hydrazine hydrate (1.3 g, 0.005 mol).
The reaction mixture was heated at reflux temperature for 2 h and
reaction was monitored by TLC [Hexane : Ethylacetate (4:1)]. The
reaction mass was cooled to room temperature and stirred for 0.5 h.
The solid product was filtered and washed with ethanol to get (6aꢀe),
(8aꢀe) and (10aꢀe) respectively.
The in vitro cytotoxicity study was carried out using HeLa
cells at Stellixir Biotech Pvt. Ltd, Bangalore. The five compounds
6a, 6d, 8b, 10c and 10e which had highest to moderate activity for
antimicrobial and antituberculosis were tested for cytotoxicity with
HeLa cells represented in Figure 1. The IC50 value of synthetic
compound found to be moderately effective for the HeLa cells. The
control cells which are not treated with any compound have shown
100% viability. One of the synthetic compound 8b has shown
highest cytotoxicity (12.83 µg/mL) among tested compounds. The
compounds having cytotoxicity below 50 µg/mL IC50 value are
usually considered as toxic compounds.
1′ꢀ(4ꢀchlorophenyl)ꢀ5ꢀ(2,3ꢀdihydrobenzofuranꢀ5ꢀyl)ꢀ3′ꢀ
phenylꢀ3,4ꢀdihydroꢀ2H,1′Hꢀ[3,4′]bipyrazolyl (6a). IR (KBr vmax
cmꢀ1): 3322 (NꢀH str), 3064 (ArꢀH str), 2919 (CꢀH aliphatic str),
1607 (C=N str), 1497 (C=C str), 824 (CꢀCl str); 1H NMR (400 MHz,
DMSOꢀd6, ppm): δ 2.90ꢀ2.96 (t, 1H, HA, J = 12.8 Hz), 3.19 (t, 2H, ꢀ
CH2), 3.42ꢀ3.49 (t, 1H, HB, J = 13.6 Hz), 4.56 (t, 2H, ꢀCH2), 4.89ꢀ
4.92 (t, 1H, HX, J = 10.4 Hz), 6.77 (m, 1H, ArꢀH), 7.33ꢀ7.36 (m, 2H,
ArꢀH), 7.43ꢀ7.56 (m, 6H, ArꢀH), 7.77ꢀ7.95 (m, 4H, ArꢀH), 8.62 (s,
1H, ꢀNH); 13C NMR (100 MHz, DMSOꢀd6, ppm): δ 160.5, 151.2,
138.8, 133.2, 130.7, 130.1, 129.9, 129.7, 129.1, 128.6, 128.4, 128.2,
128.0, 126.3, 122.9, 121.0, 120.2, 109.2, 71.7, 55.5, 29.3; MS: m/z =
441.2 (M+), ANAL. Calcd. for C26H21ClN4O; calcd: C, 70.82; H,
4.80; N, 12.71; found: C, 70.83; H, 4.80; N, 12.72.
1′ꢀ(4ꢀchlorophenyl)ꢀ5ꢀ(5ꢀmethylfuranꢀ2ꢀyl)ꢀ3′ꢀphenylꢀ
3,4ꢀdihydroꢀ2H,1′Hꢀ[3,4′]bipyrazolyl (8a). IR (KBr vmax cmꢀ1):
3316 (NꢀH str), 3117 (ArꢀH str), 2923 (CꢀH aliphatic str), 1594
(C=N str), 1499 (C=C str), 829 (CꢀClstr); 1H NMR (400 MHz,
DMSOꢀd6, ppm): δ 2.31 (s, 3H, ꢀCH3), 2.84ꢀ2.90 (t, 1H, HA, J =
13.0 Hz), 3.31 (m, 1H, HB), 4.89ꢀ4.95 (t, 1H, HX, J =10.6 Hz), 6.17
(s, 1H, ArꢀH), 6.50 (s, 1H, ArꢀH), 7.44ꢀ7.76 (m, 8H, ArꢀH), 7.94ꢀ
Figure 1: Cytotoxicity of 6a, 6d, 8b, 10c and 10e with HeLa cell line 7.95 (d, 2H, ArꢀH, J = 6.0 Hz), 8.59 (s, 1H, ꢀNH); 13C NMR (100
MHz, DMSOꢀd6, ppm): δ 152.9, 151.1, 147.3, 142.1, 138.7, 133.2,
130.7, 129.9, 129.1, 128.6, 128.4, 128.0, 123.7, 120.2, 110.9, 108.3,
This concludes that, compounds 6aꢀe, 8aꢀe and 10aꢀe derivatives
55.2, 13.9; MS: m/z =403.2 (M+), ANAL. Calcd. for C23H19ClN4O;
were synthesized, characterized and investigated for their inꢀvitro
calcd: C, 68.57; H, 4.75; N, 13.91; found: C, 68.60; H, 4.76; N,
antimicrobial by Resazurin reduction method and was used for
13.92.
determining the MIC in 96 well microplates.19 and antitubercular
activity by microplate alamar blue assay (MABA) method20 and
proved to be very good antimicrobial and antitubercular agents. The
results are consistent with specific substitution to utility of
tuberculosis chemotherapy and antimicrobial agents. The
compounds 10e, 10c showed best screening results among all the
synthesized compounds. This indicates, newly synthesized pyrazole
containing pyrazoline compounds might emerge as one of the
antituberculosis drugs.
5ꢀBiphenylꢀ4ꢀylꢀ1′ꢀ(4ꢀchlorophenyl)ꢀ3′ꢀphenylꢀ3,4ꢀ
dihydroꢀ2H,1′Hꢀ [3,4′]bipyrazolyl (10a). IR (KBr vmax cmꢀ1): 3312
(NꢀH str), 3073 (ArꢀH str), 2921 (CꢀH aliphatic str), 1593 (C=N str),
1497 (C=C str), 830 (CꢀCl str); 1H NMR (400 MHz, DMSOꢀd6,
ppm): δ 2.97ꢀ3.03 (dd, 1H, HA, J = 10.9 Hz), 3.49ꢀ3.56 (dd, 1H, HB,
J = 10.7 Hz), 4.97ꢀ5.03 (dt, 1H, Hx), 7.30ꢀ7.38 (m, 2H, ArꢀH), 7.45ꢀ
7.55 (m, 6H, ArꢀH), 7.64ꢀ7.65 (d, 1H, ArꢀH, J =3.3 Hz), 7.68 (s, 1H,
pyrazoleꢀ5H), 7.70ꢀ7.71 (dd, 5H, ArꢀH, J = 1.4 Hz), 7.80ꢀ7.82 (d,
2H, ArꢀH, J = 8.5 Hz), 7.89ꢀ7.91 (d, 2H, ArꢀH, J = 7.7 Hz), 8.61(s,
1H, ꢀNH); MS: m/z = 475.1 (M+), ANAL. Calcd. for C30H23ClN4;
calcd: C, 75.86; H, 4.88; N, 11.80; found: C, 75.90; H, 4.90; N,
11.90.
Experimental
All the chemicals were purchased from Sigma Aldrich and
SpectrochemꢀIndia. Melting points were determined by open
capillary method and are uncorrected. The IR spectrums (in KBr
pellet) were recorded on PerkinꢀElmer FTIRꢀ4000ꢀ400 cmꢀ1
spectrophotometer. NMR spectra were obtained on a Bruker
Acknowledgments
AMI thank, Director, National Institute of Technology Karnataka,
India for providing the research facilities. The authors extend their
appreciation to The Deanship of Scientific Research at King Saud
University for funding the work through research group project no.
RGPꢀVPPꢀ207.
1
Avanceꢀ400 spectrometer (400 MHz) for H NMR and 13C NMR
using tetramethylsilane (TMS) as internal standard. The mass
spectrum was recorded on LCꢀMS Applied biosystems MDS
SCIEXꢀAPI 4000 spectrometer. Elemental analysis was performed
This journal is © The Royal Society of Chemistry 2012
J. Name., 2012, 00, 1-3 | 3
Harikrishna
