Synthesis, Characterization and biological applications of pyrazole-benzothiazolamine conjugates

Article abstract of DOI:10.14233/ajchem.2017.20395

Present paper describes the synthesis of some new pyrazole-benzothiazolamine conjugates by molecular conjunction by adopting appropriate synthetic routes. Purification of intermediates and final compounds has been done by recrystallization and chromatographic techniques. Characterization of all the newly synthesized pyrazole-benzothiazolamine derivatives was done by physical and spectral data. The experimental procedure for the preparation of seventeen pyrazole-benzthiazolamine conjugates has been incorporated. Data of synthesized compounds like IR, 1H NMR and Mass spectra were neatly presented. All these compounds were evaluated for their activity against Gram-positive and Gram-negative bacteria and various fungal strains. Anticancer activity has been carried out for the synthesized compounds using HeLa, DU145 and A549 cell lines using MTT assay and we found that the pyrazole-benzothiazolamine conjugates were possess antimicrobial, antifungal and anticancer activities.

Full text of DOI:10.14233/ajchem.2017.20395

Asian Journal of Chemistry; Vol. 29, No. 5 (2017), 1029-1034
ASIAN JOURNAL OF CHEMISTRY
https://doi.org/10.14233/ajchem.2017.20395
Synthesis, Characterization and Biological Applications
of Pyrazole-Benzothiazolamine Conjugates
*
RAMAYANAM S.K. SHARMA , RAMACHANDRULA KRISHNA KUMAR,
D. RAVI KUMAR, SHRIKANTH H. HAVELE and S.V. MURALI MOHAN RAO
Department of Chemistry, Krishna University, Dr. M.R.A.R. P.G. Centre, Nuzvid-521 201, India
*Corresponding author: E-mail: sharmarsk178@gmail.com
Received: 4 November 2016;
Accepted: 13 January 2017;
Published online: 10 March 2017;
AJC-18293
Present paper describes the synthesis of some new pyrazole-benzothiazolamine conjugates by molecular conjunction by adopting appropriate
synthetic routes. Purification of intermediates and final compounds has been done by recrystallization and chromatographic techniques.
Characterization of all the newly synthesized pyrazole-benzothiazolamine derivatives was done by physical and spectral data. The
experimental procedure for the preparation of seventeen pyrazole-benzthiazolamine conjugates has been incorporated. Data of synthesized
1
compounds like IR, H NMR and Mass spectra were neatly presented. All these compounds were evaluated for their activity against
Gram-positive and Gram-negative bacteria and various fungal strains.Anticancer activity has been carried out for the synthesized compounds
using HeLa, DU145 and A549 cell lines using MTT assay and we found that the pyrazole-benzothiazolamine conjugates were possess
antimicrobial, antifungal and anticancer activities.
Keywords: Pyrazole-benzothiazolamine conjugates, Characterization, Evaluation.
sulphate, trimethoxy acetophenone, ammonium chloride,
potassium bromate from Sigma Aldrich (all were LR grade)
were used as received.
INTRODUCTION
Several benzothiazole derivatives synthesis and their SAR
studies was carried by Wang et al. [1]. X-ray crystallographic
studies [2] of benzthiazoles were reported earlier. Literature
survey reveals the information that the benzothiazole moiety
is responsible for the anticancer activity [3,4], antimicrobial
activity [5-9], antiglutamate activity [10], antileishmanial
activity [11], DNA binding ability and anticancer activity [12],
antidiabetic activity [13] and such compounds were found to be
potent inhibitors of HIV-1 protease [14]. In view of importance
of the biological activity of bezothiazolamines, an attempt
is made to synthesize, characterize and to evaluate the biolo-
gical activity of derivatives of pyrazole-benzothiazolamine
conjugates.
The instruments such as infrared spectra are recorded on
Perkin Elmer model 283B and Nicolet 740 FT-IR instruments
and values are given in cm-1. Proton nuclear magnetic reso-
nance spectra are recorded on varian Gemini-200, varian
Unity-200 and advance-300 MHz Bruker UX-NMR instru-
ment. The samples are made in chloroform-d (1:1) or/and
DMSO-d6 using tetramethyl silane (Me4Si) as the internal
standard. ESI mass spectra were recorded on Micro mass
Quattro LC using ESI+ software with capillary voltage 3.98
kV and ESI mode positive ion trap detector. High-resolution
mass spectra (HRMS) were recorded on a QSTAR XL Hybrid
MS-MS mass spectrometer. Elemental analysis is carried out
onVARFIO EL, se elementor.Analytical thin-layer chromato-
graphy (TLC) is performed on pre coated silica-gel-60 F254
(0.5 mm) glass plates.Visualization of the sports on TLC plates
is achieved either to iodine vapour or UV light. Employing
TLC techniques using appropriate solvent system for deve-
lopment monitored all the reasons. Moisture sensitive reactions
are carried out by standard syringe-septum techniques. Dry
ether, dry toluene are made by distilling them from sodium
benzophenone ketyl and dry methanol is prepared by using
potassium hydroxide. All extracts are extracted with ethyl
EXPERIMENTAL
Sodium ethoxide, hydrazine hydrochloride, lithium
aluminium hydride, tetrahydrofuran, dimethyl sulphoxide,
2-amino-6-methoxy benzothiazole, 2-amino-6-fluoro benzo-
thiazole, 2-amino-6-chloro benzothiazole, 2-amino-6-carbo-
ntrifluoro benzothiazole, 2-amino-benzothiazole, ethyl acetate,
2-iodo benzoic acid, acetone, conc. sulphuric acid, hexane,
monomethoxy acetophenone, dimethoxy acetophenone,
acetophenone, ethanol, acetophenone, anhydrous sodium

1030 Sharma et al.
Asian J. Chem.
acetoacetate and water and concentrated at reduced Pressure
on Buchi-R-3000 rotary evaporated below 50 °C. Yields of
materials judged homogenous by TLC and NMR spectroscopy.
Melting Points were recorded on Melter Fp-51 instrument and
were uncorrected were used for the characterization of synthe-
sized compounds.
was prepared using above method by the addition of [3-(3,4,5-
trimethoxyphenyl)-1H-pyrazol-5-yl]methanol 10c (2.64 g, 10
mmol) IBX (3.36 g 1.2 mmol). 3-(Benzo[d][1,3]-dioxol-5-
yl)-1H-pyrazole-5-carbaldehyde 11d was prepared using
above method by the addition of (3-(benzo[d][1,3]dioxol-5-
yl)-1H-pyrazol-5-yl)methanol 10d (2.18 g, 10 mmol) IBX
(3.36 g, 1.2 mmol). The obtained carbaldehydes were as such
taken in the next step for the synthesis of pyrazole-benzo-
thiazole conjugates 12a-12q.
Preparation of ethyl 2,4-dioxo-4-(substituted phenyl)
butanoates 8(a-d): Diethyl oxalate (1 mol) was added to
freshly prepared sodium ethanolate at 0 °C. After 10 min
substituted acetophenones 7(a-d) (1 mol) were added slowly
in small portions maintaining the temperature 0 °C. After
completion of addition the stirring was continued at room
temperature for 4 h, the reaction mixture was neutralized by
diluted H2SO4 and extracted with ethyl acetate to offer solid
products 8(a-d) (yield 85-90 %) which were taken as such for
the next step without purification.
Preparation of pyrazole and benzothiazole amine
conjugates (12a-12q): To 3-subtituted phenyl-1H-pyrazole-5-
carbaldehydes 11(a-d) (1 mmol) obtained in the above step
was added substituted 2-amino benzothiazoles (1.0 mmol) in
ethanol and heated to reflux for 6 h. The completion of reaction
was confirmed by TLC in ethyl acetate and hexane solvent
system. The product so obtained was filtered and washed twice
with ethanol. Futher these solid products were recrystallized
in ethanol to get pure conjugates of pyrazole and benzothiazole
amine (12a-12q) in good yields (Scheme-I).
Preparation of ethyl 3-substituted phenyl-1H-pyrazole-
5-carboxylates 9(a-d): To each ethyl 2, 4-dioxo-4-(substituted
phenyl)butanoates 8(a-d) (1 mol) was added hydrazine
dihydrochloride (NH2-NH2·2HCl) (1.5 mol) in ethanol and
heated to reflux for 3 h. The solvent was removed under vacuum
then added water to the residue and the compound was
extracted with ethyl acetate (50 mL × 4). The organic layer
was dried on anhydrous Na2SO4 and evaporated the solvent to
obtain crude product that was further purified by column
chromatography using ethyl acetate and hexane. The pure
compounds 9(a-d) were eluted at 30-40 % of ethyl acetate
with good yields.
(E)-6-Fluoro-N-[(3-(3,4,5-trimethoxyphenyl)-1H-
pyrazol-5-yl)methylene]benzo[d]thiazol-2-amine (12a):
Yield: 71 %, brown coloured solid. m.p.: 143-145 °C; IR (Neat,
cm-1): 3234, 3000, 2931, 2831, 1526, 1476, 1259, 1197, 1065,
1025, 980, 897, 797, 753, 669; 3243.56 ν(N-H) , 1259.57-
1
ν(C-N) aryl 1065.47 ν(C-F): H NMR (DMSO, 300 Hz): δ
values: 3.70-3.87 (M, 9H)) –OCH3; 6.85-7.02 (M, 2H) ArH;
7.06-7.23 (M, 1H) ArH; 7.53-7.79 (M, 3H) ArH; 8.19 (S, 1H)
ArH ppm; Mass (ESI) (m/z): 365 [M+], 367 [M+2] peak.
(E)-N-[(3-(3,4-Dimethoxyphenyl)-1H-pyrazol-5-
yl)methylene]-6-methoxybenzo[d]thiazol-2-amine (12b):
Yield: 79 %, brown coloured solid. m.p.: 148-152 °C; 1H NMR
(DMSO, 300 Hz): δ values: 3.79 (s, 3H) –OCH3, δ = 3.82 (s,
6H) –OCH3 δ = 6.787 (s, 3H) ArH, δ = 7.07-7-34 (m, 2H) Ar-
H δ = 7.28 (s, 2H) Ar-H δ = 7.48-7-63 (m, 1H) Ar-H ppm;
Mass (ESI) (m/z): 395 [M+] peak.
Preparation of (3-substituted phenyl-1H-pyrazol-5-
yl)methanols 10(a-d): To the ethyl 3-substituted phenyl-1H-
pyrazole-5-carboxylates 9(a-d), obtained in the above step was
added LiAlH4 (0.5 mol) in dry THF at 0 °C and stirred for 1h
at room temperature. Added saturated NH4Cl solution drop
wise to quench the unreacted LiAlH4 and removed the THF
under vacuum then extracted with ethyl acetate (100 mL × 4).
The organic layer was dried on anhydrous Na2SO4 and evapo-
rated ethyl acetate to obtain colour less solid products of (3-
substitutedphenyl-1H-pyrazol-5-yl)methanols 10(a-d) (yield
70-80 %). The alcohols produced in this step were pure and
no further purification was required. These compounds were
taken as such for the next step.
(E)-N-[(3-(3,4-Dimethoxyphenyl)-1H-pyrazol-5-yl)-
methylene]-6-fluorobenzo[d]thiazol-2-amine (12c): Yield:
75 %, brown coloured solid; m.p.: 163-165 °C; IR (Neat, cm-1):
3261, 3016, 2937, 2837, 1608, 1545, 1460, 1414, 1331, 1258,
1205, 1159, 1028, 979, 797, 635, 549, 433; 3261 ν(N-H), 1258
ν(C-N) aryl, 1028 ν(C-F), 3016 aromatic C-H stretching; 1H
NMR (DMSO, 300 Hz): δ = 3.65-3.95 (m, 6H) –OCH3, δ =
6.62-7.03 (m, 2H) ArH, δ = 7.18-7.30 (m, 2H) Ar-H δ = 7.48
(s, 5H) Ar-H ppm; Mass (ESI) (m/z): 383 [M+H] peak
(E)-4-Chloro-N-((3-(3,4-dimethoxy phenyl)-1H-pyrazol-
5-yl)methylene)benzo[d]thiazol-2 amine (12d): Yield: 73 %,
light yellow coloured solid; m.p.: 158-160 °C; IR (Neat, cm-1):
3208, 2991, 2830, 1591, 1533, 1434, 1262, 1195, 1107, 1027,
981, 858, 763, 710, 670; 3261 ν(N-H), 1258 ν(C-N) aryl, 3016
Preparation of 3-subtitutedphenyl-1H-pyrazole-5-
carbaldehydes 11(a-d): To the (3-substituted phenyl-1H-
pyrazol-5-yl)methanols 10(a-d) produced in the above step
was added IBX (1.2 mol) in DMSO and stirred for 1 h at room
temperature. Ice cold water was added to the reaction mixture
and extracted with ethyl acetate (50 mL × 4). The organic
layer was dried on anhydrous Na2SO4 and evaporated the ethyl
acetate to obtain corresponding 3-subtituted phenyl-1H-
pyrazole-5-carbaldehydes 11(a-d) in good yields (80-85 %).
3-(4-Methoxyphenyl)-1H-pyrazole-5-carbaldehyde 11a
was prepared using above method by the addition of [3-(4-
methoxyphenyl)-1H-pyrazol-5-yl]methanol 10a (2.04 g, 10
mmol) IBX (3.36 g, 1.2 mmol. 3-(3,4-dimethoxyphenyl)-1H-
pyrazole-5-carbaldehyde 11b was prepared using above method
by the addition of [3-(3,4-dimethoxyphenyl)-1H-pyrazol-5-
yl]methanol 10b (2.34 g, 10 mmol) IBX (3.36 g, 1.2 mmol).
(3,4,5-Trimethoxyphenyl)-1H-pyrazole-5-carbaldehyde 11c
1
-aromatic C-H stretching, 797 -C-Cl stretching; H NMR
(DMSO, 300 Hz): δ = 3.65-3.87 (m, 6H) –OCH3, δ = 7.03 (s,
1H) ArH, δ = 7.08 (s, 1H) Ar-H δ = 7.10 (s, 1H) Ar-H δ =
7.30-7.42 (m, 3H) Ar-H δ = 7.69-7.84 (m, 1H) Ar-H ppm;
Mass (ESI) (m/z): 399 [M+] peaks.
(E)-N-[(3-(3,4-Dimethoxyphenyl)-1H-pyrazol-5-yl)-
methylene]-6-methylbenzo[d]thiazol-2-amine (12e): Yield:
78 %, light yellow coloured solid, m.p.: 147-151 °C; IR (Neat,
cm-1): 3237, 2931, 2833, 1608, 1524, 1459, 14333, 1259, 1238,

Vol. 29, No. 5 (2017)
Synthesis and Biological Applications of Pyrazole-Benzothiazolamine Conjugates 1031
R1, R2, R3, R4 = Alkyl groups
(i) NaOEt/EtOH/Diethyl oxalate, 4 h, room temperature; (ii) NH2NH2·2HCl/EtOH, 3 h, Reflux;
(iii) LiAlH4/THF, 1 h, room temperature; (iv) IBX/DMSO, 1 h, room temperature; (v) 2-Aminobenzothiazoles/EtOH, 4 h, Reflux
Scheme-I
1190, 1142, 1024, 979, 868, 802, 761, 660; 1258 ν(N-H), 1258
ν(C-N) aryl, 3016 -aromatic C-H stretching; 1H NMR (DMSO,
300 Hz): δ = 3.69-3.77 (m, 6H) –OCH3, δ = 6.90-7.11 (m,
2H) ArH, δ = 7.13-7.25 (m, 1H) Ar-H δ = 7.33 (s, 3H) Ar-H δ
= 7.53-7.63 (m, 1H) Ar-H, δ = 9.60-9.71 (s, 1H) N-H ppm;
Mass (ESI) (m/z): 379 [M+] peaks.
cream coloured solid; m.p.: 166-168 °C; IR (Neat, cm-1): 3443,
3187, 2834, 2704, 1614, 1539, 1412, 1373, 1253, 1178, 1111,
1072, 966, 833, 762, 660, 533; 3187 ν(N-H), 1253 ν(C-N)
aryl, 3016 -aromatic C-H stretching, 762-C-Cl stretching; 1H
NMR (DMSO, 300 Hz): δ = 3.74-3.85 (m, 3H) –OCH3, δ =
6.75-7.06 (m, 4H) ArH, δ = 7.22-7.32 (d, 1H) ArH, δ = 7.45-
7.56 (m, 1H)Ar-H, δ = 7.65-7.75 (m, 2H)Ar-H, δ = 9.74-9.95
(m, 1H) N-H ppm; Mass (ESI) (m/z): 369 [M+] peak.
(E)-N-[(3-(Benzo[d][1,3]dioxol-5-yl)-1H-pyrazol-5-
yl)methylene]-6-methoxybenzo[d]thiazol-2-amine (12j):
Yield: 78 %, light red coloured solid. m.p.: 152-155 °C; IR
(Neat, cm-1): 3188, 3002, 1606, 1548, 1465, 1335, 1288, 1246,
1221, 1166, 1115, 1036, 978, 934, 865, 796, 655; 3188 ν(N-H),
1246 ν(C-N) aryl, 3002 aromatic ν(C-H). 1H NMR (DMSO,
300 Hz): δ = 6.03 (s, 6H) –OCH2 -O, δ = 6.65-7.0 (m, 3H)
ArH, δ = 6.99-7.04 (m, 2H)ArH, δ = 7.12-7.36 (m, 2H)Ar-H,
δ = 7.82 (s, 1H) Ar-H ppm; Mass (ESI) (m/z): 391 [M+23]
peak.
(E)-6-Fluoro-N-[(3-(4-methoxyphenyl)-1H-pyrazol-5-
yl)methylene]-benzo[d]thiazol-2-amine (12f): Yield: 79 %,
cream coloured solid, m.p.: 163-165 °C; IR (Neat, cm-1): 3012,
2835, 2705, 1611, 1544, 1457, 1287, 1250, 1196, 1114, 1071,
1030, 965, 835, 791, 663; 3261 ν(N-H), 1258 ν(C-N) aryl,
3016 -aromatic ν(C-H), 1071-C-F stretching; 1H NMR (DMSO,
300 Hz): δ = 3.72-3.76 (m, 3H) –OCH3, δ = 6.84-6.77 (m,
3H) ArH, δ = 7.10-7.44 (m, 2H) Ar-H, δ = 7.57-7.78 (m, 4H)
Ar-H, δ = 9.70 (s, 1H) N-H ppm; Mass (ESI) (m/z): 352 [M+]
peaks.
(E)-N-[(3-(3,4-Dimethoxyphenyl)-1H-pyrazol-5-yl)-
methylene]-6-(trifluoromethyl)benzo[d]thiazol-2-amine
(12g): Yield: 75 %, white coloured solid, m.p.: 155-157 °C;
IR (Neat, cm-1): 3257, 2937, 2834, 1529, 1473, 1325, 1259,
1161, 1107, 1079, 979, 830, 714, 646; 3261 ν(N-H), 1258
ν(C-N) aryl, 3016 -aromatic C-H stretching, 1079 -C-F
stretching; 1H NMR (DMSO, 300 Hz): δ = 3.68-3.84 (m, 3H)
–OCH3, δ = 6.88-7.02 (m, 1H) ArH δ = 7.22 (s, 1H) ArH, δ =
7.34 (s, 2H) Ar-H, δ = 7.59-7.79 (m, 2H) Ar-H, δ = 7.63-7.78
(m, 3H) Ar-H, δ = 8.27 (s, 1H) Ar-H, δ = 9.90 (s, 1H) N-H
ppm; Mass (ESI) (m/z): 433 [M+] peaks.
(E)-N-[(3-(Benzo[d][1,3]dioxol-5-yl)-1H-pyrazol-5-
yl)methylene]-6-fluorobenzo[d]thiazol-2-amine (12k):
Yield: 72 %, cream coloured solid; m.p.: 153-158 °C; IR (Neat,
cm-1): 3234, 3000, 2931, 2831, 1526, 1476, 1259, 1197, 1065,
1025, 980, 897, 797, 753, 669; 3234 ν(N-H), 1259 ν(C-N)
1
aryl, 3000 aromatic ν(C-H), 1025-C-F stretching; H NMR
(DMSO, 300 Hz): δ = 5.87-5.99 (m, 2H) –O-CH2-O, δ = 7.16-
7.30 (m, 3H) Ar-H, δ = 7.82 (s, 4H) Ar-H, δ = 6.97-7.20 (s,
1H) Ar-H ppm; Mass (ESI) (m/z): 367 [M+] peaks.
(E)-6-methoxy-N-[(3-(4-methoxyphenyl)-1H-pyrazol-
5-yl)methylene]benzo[d]thiazol-2-amine (12h): Yield: 76 %,
brown coloured solid; m.p.: 169-171 °C; IR (Neat, cm-1): 3212,
3021, 2958, 2832, 2707, 1609, 1546, 1467, 1250, 1175, 1058,
964, 831, 793, 659, 534; 3261 ν(N-H), 1258 ν(C-N) aryl, 3016
-aromatic C-H stretching, 1H NMR (DMSO, 300 Hz): δ = 3.72-
3.86 (m, 6H) –OCH3, δ = 6.75-7.04 (m, 4H) Ar, δ = 7.10-7.37
(m, 1H)ArH, δ = 7.42-7.64 (m, 1H)Ar-H, δ = 7.72 (s, 2H)Ar-
H ppm; Mass (ESI) (m/z): 365 [M+] peaks.
(E)-6-Methoxy-N-[(3-(3, 4, 5-trimethoxyphenyl)-1H-
pyrazol-5-yl)methylene]benzo[d]thiazol-2-amine (12l):
Yield: 72 %, brown coloured solid; m.p.: 145-148 °C; IR (Neat,
cm-1): 3342, 3204, 2932, 2829, 1600, 1548, 1468, 1422, 1365,
1282, 1226, 1125, 1055, 1003, 903, 832, 764, 661; 3204
ν(N-H), 1282 ν(C-N) aryl, 3204 aromatic C-H stretching; 1H
NMR (DMSO, 300 Hz): δ = 3.73-3.88 (m, 12H) –OCH3, δ =
6.79-6.84 (m, 1H) ArH, δ = 6.99-7.04 (m, 2H) ArH, δ = 7.22-
7.39 (m, 1H) Ar-H, δ = 7.46-7.60 (m, 1H) Ar-H, δ = 8.02 (s,
1H)Ar-H, δ = 9.31-9.36 (m, 1H) N-H ppm; Mass (ESI) (m/z):
425 [M+] peaks.
(E)-4-Chloro-N-[(3-(4-methoxyphenyl)-1H-pyrazol-5-
yl)methylene]benzo[d]thiazol-2-amine (12i): Yield: 78 %,

1032 Sharma et al.
Asian J. Chem.
(E)-6-Fluoro-N-[(3-(3,4,5-trimethoxyphenyl)-1H-
fungal strain like Candida albicans MTCC3017 were tested for
the synthesized compounds against the standards chloramphi-
nicol and nistatin. 0.1 mL of standard inoculum (1-2 × 107
cfu/mL) of each organism was added to freshly prepared sterile
Mueller Hilton Agar plates (HiMedia Laboratories) and the
compounds dissolved in DMSO was added on the sterile disc,
after 18-24 h of incubation the plates were observed for zone
of inhibition. The MIC values were determined in each case as
the minimum concentration of the compound which inhibited
the growth of organism.
pyrazol-5-yl)methylene]-benzo[d]thiazol-2-amine (12m):
Yield: 73 %, brown coloured solid; m.p.: 167-168 °C; IR (Neat,
cm-1): 3171, 2970, 1608, 1554, 1461, 1423, 1340, 1290, 1120,
1010, 905, 842, 764, 659, 566; 3171 ν(N-H), 1290 ν(C-N)
1
aryl, 3000 aromatic ν(C-H), 1010-C-F stretching; H NMR
(DMSO, 300 Hz): δ = 3.77-3.94 (m, 9H) –OCH3, δ = 6.79-
6.85 (m, 1H) ArH, δ = 6.96-7.08 (m, 3H) ArH, δ = 7.32-7.39
(m, 1H) Ar-H, δ = 7.59 (s, 2H) Ar-H, δ = 9.09-9.17 (m, 1H)
N-H ppm; Mass (ESI) (m/z): 413 [M+] peaks.
(E)-6-Methoxy-N-[(3-(3,4,5-trimethoxyphenyl)-1H-
pyrazol-5-yl)methylene]-benzo[d]thiazol-2-amine (12n):
Yield: 74 %, brown coloured solid; m.p.: 158-161 °C; IR (Neat,
cm-1): 3208, 3004, 2831, 1594, 1538, 1469, 1423, 1325, 1284,
1236, 1130, 1082, 1046, 1013, 896, 831, 796, 715, 670; 3208
ν(N-H), 1284 ν(C-N) aryl, 3004 aromatic ν(C-H); 1H NMR
(DMSO, 300 Hz): δ = 3.71-3.97 (m, 12H) –OCH3, δ = 6.46 (s,
1H) ArH, δ = 6.82 (s, 1H) ArH, δ = 6.93-6.97 (s, 1H) Ar-H, δ
= 7.05-7.16 (s, 2H) Ar-H, δ = 7.25-7.44 (s, 1H)Ar-H, δ = 7.66
(s, 1H) Ar-H, δ = 8.34 (s, 1H) Ar-H ppm; Mass (ESI) (m/z):
463 [M+ k] peaks.
Antifungal activity: Selectively 12b was chosen to
check the antifungal activity against selected Candida strains
(Issattchenkia orientalis MTCC 3020, Candida albicans
MTCC 1637, Candida albicans MTCC 3019, Candida
albicans MTCC 3958, Candida albicans MTCC 227, Candida
albicans MTCC 7315, Candida albicans MTCC 854, Candida
albicans MTCC 183, Candida albicans MTCC 3018 and
Candida parapsilosis MTCC 1744) and showed good activity
against Candida albicans MTCC3017.
Anticancer activity: Anticancer activity has been carried
out for the synthesized compounds using HeLa, DU145 and
A549 cell lines by MTT Assay.
(E)-6-Chloro-N-[(3-(3,4,5-trimethoxyphenyl)-1H-
pyrazol-5-yl)methylene]-benzo[d]thiazol-2-amine (12o):
Yield: 72 %, brown coloured solid, m.p.: 144-145 °C; IR (Neat,
cm-1): 3180, 2932, 2828, 1595, 1539, 1466, 1421, 1280, 1235,
1203, 1129, 1045, 1000, 890, 811, 781, 652; 3180 ν(N-H),
RESULTS AND DISCUSSION
Sodium ethoxide acts as a strong base which abstracts
proton from methylene carbon of acetophenone to result
carbanion. The active carbanion abstracts carbonyl carbon of
diethyl oxalate by Claisen-Schmidt condensation. Further
removal of ethanol produces 8(a-d) compounds. Hydrazine
dihydro chloride reacts with 2,4-dioxo-4-substituted pentanoic
acid ester via Knorr condensation to form compound 9(a-d).
Pyrazole moiety is formed in this step. Ester undergoes
reduction with lithium aluminium hydride to form alcohol
10(a-d). Then alcohol undergoes selective oxidation with IBX
to form aldehyde 11(a-d). In the final step, 11(a-d) reacts with
2-amino benzothiazoles to form pyrazole-benzothiazole
conjugates 12(a-d).
1
1235 ν(C-N) aryl, 3016 aromatic ν(C-H), 781 ν(C-Cl); H
NMR (DMSO, 300 Hz): δ = 3.52-3.81 (m, 9H) –OCH3, δ =
6.65 (s, 1H) ArH, δ = 6.77-6.88 (m, 2H) ArH, δ = 7.08-7.18
(m, 2H) Ar-H, δ = 7.41 (s, 2H) Ar-H, δ = 8.82 (brs, 1H) N-H
ppm; Mass (ESI) (m/z): 365 [M+], 367 [M+2] peak.
(E)-N-[(3-(Benzo[d] [1,3]dioxol-5-yl)-1H-pyrazol-5-
yl)methylene]-6-chlorobenzo[d]thiazol-2-amine (12p):
Yield: 71 %, brown coloured solid; m.p.: 155-157 °C; IR (Neat,
cm-1): 3180, 3001, 1598, 1541, 1460, 1335, 1288, 1245, 1220,
1167, 1113, 1039, 978, 935, 865, 798, 619, 553; 3180 ν(N-H),
1
1245 ν(C-N) aryl, 3001 aromatic ν(C-H), 798 ν(C-Cl); H
NMR (DMSO, 300 Hz): δ = 5.90-6.14 (m, 2H) –O-CH2 –O-,
δ = 6.68-7.04 (m, 3H) ArH, δ = 7.11-7.43 (m, 2H) ArH, δ =
7.88 (s, 3H) Ar-H ppm; Mass (ESI) (m/z): 383 [M+] peaks.
(E)-N-[(3-(Benzo[d][1,3]dioxol-5-yl)-1H-pyrazol-5-yl)-
methylene]-6-(trifluoro methyl)benzo[d] thiazol-2-amine
(12q): Yield: 75 %, m.p.: 159-162 °C; IR (Neat, cm-1): 3223,
3021, 2902, 1610, 1572, 1539, 1462, 1324, 1285, 1244, 1163,
1112, 1080, 1040, 35, 863, 830, 801, 717, 645; 3223 ν(N-H),
Biological activity study reveals that few of the synthe-
sized compounds have shown broad spectrum of antimicrobial
activity against Gram-positive and Gram-negative organisms.
The compound 12h was found to be most active against Gram-
positive, Gram-negative bacteria and fungus (Candida
albicans), it showed excellent activity against Bacillus subtilis
MTCC121 (MIC 4 µg/disc), Staphylococcus MLS-16
MTCC2940 (MIC 8 µg/disc), Micrococcus luteus MTCC2470
(MIC 4 µg/disc), Staphylococcus aureus MTCC 96 (MIC 6
µg/disc) and Escherichia coli MTCC739 (MIC 16 µg/disc).
The compounds 12a, 12i, 12j, 12k and 12l also showed
moderate to good inhibitory activity but less as compared to
12h. Overall rest of the synthesized compounds showed
average antimicrobial activity when compared to standard
compounds and MIC values ranged from (6-80 µg/disc). As
per the results observed and thus obtained from the anti-
microbial activity, Gram-positive organisms are more affected
than Gram-negative organisms by the lead compounds. The
antimicrobial activity and inhibitory zone of synthesized
compounds against tested strains were incorporated in the
Tables 1 and 2, respectively.
1
1244 ν(C-N) aryl, 3021 aromatic ν(C-H), 1080 ν(C-F); H
NMR (DMSO, 300 Hz): δ = 5.90-6.0 (m, 2H) –O-CH2-O-, δ
= 6.78-6.99 (m, 2H) ArH, δ = 7.24-7.34 (m, 2H) ArH, δ =
7.54-7.84 (m, 3H) Ar-H, δ = 8.08-8.22 (m, 1H) Ar-H ppm;
Mass (ESI) (m/z): 417 [M+] peaks.
Biological activity study
Antimicrobial activity: The antimicrobial activities of
the synthesized compounds were tested using disc diffusion
method. Various Gram-positive bacteria like Bacillus subtilis
MTCC121, Staphylococcus aureus MTCC 96, Staphylococcus
MLS-16 MTCC2940, Micrococcus luteus MTCC2470, Gram-
negative bacteria like Escherichia coli MTCC739, Pseudomonas
aeruginosa MTCC2453, Klebsiella planticola MTCC530,

Vol. 29, No. 5 (2017)
Synthesis and Biological Applications of Pyrazole-Benzothiazolamine Conjugates 1033
TABLE-1
ANTIMICROBIAL ACTIVITY OF SYNTHESIZED COMPOUNDS (MIC, µg/mL)
Bacillus
subtilis
MTCC121
Staphylococcus Micrococcus Staphylococcus Escherichia Pseudomonas
aeruginosa
MTCC2453
Klebsiella
planticola
MTCC530
Candida
albicans
MTCC3017
luteus
MTCC2470
aureus
MTCC 96
coli
MTCC739
Compound
MLS-16
MTCC2940
12a
12b
12c
12d
12e
12f
15
80
41
80
80
80
78
4
12
80
10
80
80
75
80
9
16
80
80
80
80
80
80
5
10
80
80
80
80
75
80
8
25
80
80
80
80
80
75
15
22
78
40
40
80
80
80
80
80
40
80
80
12
80
80
40
26
40
80
25
40
40
60
40
80
80
24
80
80
80
80
80
80
40
25
80
40
24
80
80
80
80
80
24
20
34
80
80
80
80
40
25
80
25
24
80
80
80
80
80
12g
12h
12i
10
40
8
15
80
15
10
80
80
35
60
80
25
80
12
15
80
80
35
80
80
10
80
5
12j
12k
12l
20
76
62
40
80
80
40
80
80
35
75
80
12m
12n
12o
12p
12q
a = Gram-positive bacteria; b = Gram-negative bacteria; c = Fungus.
TABLE-2
INHIBITORY ZONE (DIAMETER, mm) OF SYNTHESIZED COMPOUNDS AGAINST TESTED STRAINS
Bacillus
subtilis
MTCC121
Staphylococcus Micrococcus Staphylococcus Escherichia Pseudomonas
aeruginosa
MTCC2453
Klebsiella
planticola
MTCC530
Candida
albicans
MTCC3017
luteus
MTCC2470
aureus
MTCC 96
coli
MTCC739
Compound
MLS-16
MTCC2940
12a
12h
12i
20
30
15
5
15
25
20
3
20
30
12
4
16
25
15
3
10
20
10
3
5
10
6
10
5
5
5
12
3
10
3
12j
12k
12l
3
15
10
15
15
20
15
30
5
6
10
5
5
10
10
5
10
a = Gram-positive bacteria; b = Gram-negative bacteria; c = Fungus; Standard (20 µg/disc) was used as positive reference. Synthesized compounds
(20 µg/disc).
Antifungal activity: Among the tested Candida strains,
Candida albicans MTCC 1637 and Candida albicans MTCC
3020 showed the best activity as compared to others. Overall
a very good antifungal activity was observed against most
organisms used when compared to standard compounds and
MIC values ranged from (4.0-80 µg/disc). Antifungal activity
of synthesized compounds were incorporated in Table-3.
Anticancer activity: Few of the compounds have shown
moderate to best activity against cancer cell lines. Compounds
12b, 12n and 12p displayed moderate activity against A549
cell lines; 12a, 12i and 12o against HeLa cell lines; 12h and
12i against DU145. Best activity has been shown by 12b, 12c
and 12f against HeLa cell lines; 12m, 12n, 12o and 12q against
A549 cell lines (Table-4).
Conclusion
In conclusion, the synthesis and characterization of some
new pyrazole-benzothiazolamine conjugates are reported and
the majority of the synthesized compounds were biological
active against Gram-positive and Gram-negative bacteria and
various fungal strains. The compound 12h was found to be
most active against Gram-positive, Gram-negative bacteria and
TABLE-3
ANTIFUNGAL ACTIVITY OF SYNTHESIZED COMPOUNDS (MIC, µg/mL)
Organisms
12a
30
25
40
25
40
40
30
30
40
25
12h
30
40
45
50
40
60
40
40
50
30
12i
35
25
40
40
50
40
35
50
35
30
12j
75
80
80
65
60
80
80
80
70
65
12k
35
25
25
30
45
40
40
50
35
35
12l
35
25
40
35
35
45
40
40
35
30
Issattchenkia orientalis MTCC 3020
Candida albicans MTCC 1637
Candida albicans MTCC 3019
Candida albicans MTCC 3958
Candida albicans MTCC 227
Candida albicans MTCC 7315
Candida albicans MTCC 854
Candida albicans MTCC 183
Candida albicans MTCC 3018
Candida parapsilosis MTCC 1744

1034 Sharma et al.
Asian J. Chem.
TABLE-4
ANTICANCER ACTIVITY OF SYNTHESIZED
COMPOUNDS (MIC, µg/mL)
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