Synthesis of novel benzenesulfonamide bearing 1,2,3-triazole linked hydroxy-trifluoromethylpyrazolines and hydrazones as selective carbonic anhydrase isoforms IX and XII inhibitors

Article abstract of DOI:10.1016/j.bioorg.2019.01.002

A series of twenty four hydroxy-trifluoromethylpyrazoline-carbonyl-1,2,3-triazoles and four hydrazones bearing benzenesulfonamide moieties was obtained by condensation of carboxyhydrazides with substituted 1,3-diketones. All the newly synthesized compound

Full text of DOI:10.1016/j.bioorg.2019.01.002

Bioorganic Chemistry 85 (2019) 198–208
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Synthesis of novel benzenesulfonamide bearing 1,2,3-triazole linked
hydroxy-trifluoromethylpyrazolines and hydrazones as selective carbonic
anhydrase isoforms IX and XII inhibitors
Vikas Sharma , Rajiv Kumar , Silvia Bua , Claudiu T. Supuranb,⁎, Pawan K. Sharmaa,⁎
a
a
b
a
Department of Chemistry, Kurukshetra University, Kurukshetra 136119, India
b
Università degli Studi di Firenze, Laboratorio di Chimica Bioinorganica, Rm 188, and Neurofarba Department, Sezione di Scienze Farmaceutiche, Via U. Schiff 6, I-50019
Sesto Fiorentino, Firenze, Italy
A R T I C L E I N F O
A B S T R A C T
Keywords:
A series of twenty four hydroxy-trifluoromethylpyrazoline-carbonyl-1,2,3-triazoles and four hydrazones
bearing benzenesulfonamide moieties was obtained by condensation of carboxyhydrazides with substituted
Pyrazolines
Hydrazones
1
,3-diketones. All the newly synthesized compounds were investigated as inhibitors of physiologically and
1
,2,3-Triazole
pharmacologically relevant human (h) carbonic anhydrsae (CA, EC 4.2.1.1) cytosolic isoforms hCA I and II,
as well as transmembrane tumor-assosciated isoforms hCA IX and XII. These compounds exhibited excellent
CA inhibitory potency against the four CA isoenzymes as compared to clinically used reference drug acet-
azolamide (AAZ). Some compounds bearing bulkier group at C-5′ position of 1,2,3-triazoles ring were
weaker inhibitors of hCA I. Inhibition assay against hCA II indicates, that several derivatives
exhibited upto 27-fold more effective inhibitory activity compared to AAZ. Five of the assayed compounds
Benzenesulfonamide
Carbonic anhydrase isoforms I, II, IX, XII
displayed low nanomolar potency (K ≤ 10 nM) against hCA IX, whereas five compounds were found to be
i
endowed with excellent inhibitory potencies (K ≤ 5 nM) against hCA XII. The biological activity
i
profile presented herein will be useful for designing new leads and provide candidates for preclinical
investigations.
1
. Introduction
vertebrates, with at least sixteen different hCAs isoforms encoded in
their genome [11–13]. Out of the fifteen human isoforms (CA XV is not
Carbonic anhydrases (CA, EC 4.2.1.1) are metalloenzymes using
encoded in the primate genome), hCA I and II, the most abundant
isozymes, are primarily involved in important physiologic processes
such as respiration and regulation of the acid/base homeostasy, being
drug targets in retinal pathologies, cerebral edema, glaucoma and
epilepsy [14]. On the other hand, hCA IX and XII are multidomain
proteins which contribute for creating the pH regulating system
during tumor proliferation, producing an extracellular acidosis, thus
helping the growth of tumor cells in hypoxic and acidic micro-
environment [15–17]. Therefore, specifically targeting the trans-
membrane tumor-associated isozymes hCA IX and XII over the off-
target hCA I and II is a promising strategy for designing effective and
safer agents in the cancer therapy, with one such compound (SLC-
0111) in Phase II clinical studies [18].
the zinc ion as cofactor in the active site and generates hydro-
gencarbonate and a proton (or a hydronium cation) by catalyzing the
reversible reaction between CO
2
and water with rates approaching
8
−1 −1
diffusion-controlled limits (kcat/K
M
∼ 10 M
s
) [1–3]. CAs are
ubiquitous enzymes found in all organisms over the tree of life and are
engaged in physiological functions including pH regulation, biosyn-
thetic processes (ureagenesis, lipogenesis, gluconeogenesis), CO
2
homeostasis, respiration, production of biological fluids, lung elec-
trolyte secretion, calcification, chemosensing [4], and in metabolic/
signaling pathways (e.g., sexual development in fungi) in many pa-
thogens [5–7]. Indeed seven (α, β, γ, δ, ζ, η and θ-CAs) genetic families
encoding CAs have involved in organisms all over the tree of life,
+
assuring the homeostasis of CO
2
, H and bicarbonate, which differ in
The tissue dependent expression of CA isozymes regulates variety of
biological functions that makes them targets for developing the CA
their central metal active ion [8–10]. α-CAs is the prominent class in
Abbreviations: CA, carbonic anhydrase; hCA, human carbonic anhydrase; CAIs, carbonic anhydrase inhibitors; AAZ, acetazolamide; K , inhibition constant; nM,
i
nanomolar
⁎
Corresponding authors.
E-mail addresses: claudiu.supuran@unifi.it (C.T. Supuran), pksharma@kuk.ac.in (P.K. Sharma).
https://doi.org/10.1016/j.bioorg.2019.01.002
Received 4 November 2018; Received in revised form 30 December 2018; Accepted 1 January 2019
Available online 03 January 2019
0045-2068/ © 2019 Published by Elsevier Inc.

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
Fig. 1. Chemical structures of the clinically used sulfonamide CA inhibitors and derivatives incorporating the pyrazoles, pyrazolines and 1,2,3-triazole ring 1–8,
together with the newly designed sulfonamides 9–13.
inhibitor (CAI) based drugs. Primary sulfonamide based CAI drugs such
as acetazolamide (AAZ), methazolamide (MZA), ethoxzolamide (EZA),
celecoxib (CLX), etc. (Fig. 1) are clinically used for decades to treat
several diseases such as glaucoma, edema and also for the management
of osteoporosis, idiopathic intracranial hypertension, etc. [19,20].
Sulfonamide inhibits CA activity by swapping water molecule from the
zinc coordination sphere. The binding of sulfonamide to metal active
site is irreversible and the removal of water molecule from the active
site of metal inhibits all the reactions responsible for the activity of CA,
which is crucial for catalysis [21,22].
2. Results and discussion
2.1. Chemistry
The synthetic pathway employed for the synthesis of targeted sul-
fonamides is depicted in Scheme 1. Carboxylic acid hydrazides of 1,2,3-
triazole bearing benzenesulfonamide 18, required for the synthesis of
target molecules, were prepared by the diazotization of commercially
available sulfanilamide (14) which upon subsequent reaction with so-
dium azide at 0 °C yielded 4-azidobenzenesulfonamide (15) [39].
Compound 15 was subsequently treated with differently substituted β-
ketoesters 16, to afford 1,5-diaryl-1,2,3-triazole carboxylates 17, which
on further reaction with hydrazinehydrate yielded carboxylic acid hy-
drazides of 1,2,3-triazole 18 [35]. Variously substituted 1,3-diketones
19 were prepared by the reaction of corresponding acetophenones with
ethyl trifluoroacetate by using sodium ethoxide as a base in dry benzene
[40]. First of all, it was attempted to synthesize single product by
conventional method of refluxing carboxylic acid hydrazides of 1,2,3-
triazole bearing benzenesulfonamide 18 and 1,3-diketones 19 in
ethanol containing a few drops of conc. HCl for 5–6 hrs but it resulted
into a mixture of products. NMR of the isolated solid product confirmed
the presence of a mixture of hydroxypyrazolines and pyrazoles. For
getting the single desired product, different reaction conditions were
unsuccessfully attempted leading to a mixture of products. Finally, re-
action in DMF with catalytic amount of conc. HCl, stirring for 5 hrs at
Many contributions in the literature survey revealed that the
pyrazolines and 1,2,3-triazoles [23–25] represent an integral biolo-
gical architecture in medicinal chemistry associated with a diverse
array of activities such as anti-inflammatory [26], antimicrobial [27],
antitumor [28–30], antifungal [31], antiproliferative [32], anti-tu-
bercular [33] and CAI activities [23–25]. In a previous study from our
group, some pyrazolylpyrazolines bearing benzenesulfonamide 1
have been reported to show selective inhibition of hCA IX and XII
[
34]. Recently, some 4-functionalized 1,5-diaryl-1,2,3-triazoles con-
taining benzenesulfonamide 2–8 have also been reported from our
group acting as strong inhibitors of human carbonic anhydrase I, II,
IV and IX [35–37]. These results indicate that pyrazoline and 1,2,3-
triazole scaffolds are promising entities for exploring molecules of
interest as CAIs.
To study the combined effect of these moieties on biological
activity and continuing our interest in the field of designing benzene-
sulfonamide bearing heterocyclic compounds as CAIs, [34–38]
we report herein synthesis and biological evaluation of hydroxy-tri-
50 °C followed by addition of few drops of conc. H
2
SO at 90 °C fol-
4
lowed by further stirring for 8–9 hrs, afforded a solid product on
workup with water, which was purified by recrystillization. However
under the same reaction conditions, carboxylic acid hydrazides of 1,2,3-
triazole bearing benzenesulfonamide 18 on reacting with thienyl sub-
stituted 1,3-diketones 19 afforded unexpected hydrazone derivatives
13. Attempts to further cyclize the hydrazones 13 under variety of
different conditions did not yield the desired results. Postulated
fluoromethylpyrazoline-carbonyl-1,2,3-triazoles
9–12
and
tri-
fluoromethylhydrazone-carbonyl-1,2,3-triazoles 13 bearing benzene-
sulfonamide as carbonic anhydrase hCA I, II, IX and XII inhibitors
(
Fig. 1).
199

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
Ar
2
-
2
-
1
6
17
18 C H 4-H CC H 4-ClC H4 4-BrC H4 2-Naphthyl
Thienyl
6
5
3
6
4
6
6
Pyridyl
1
9
-
-
-
19a
19b
19c
9c
19d
9d
19e
9e
19f
9f
19g
13a
13b
13c
13d
CH3
C H
16a 17a 18a 9a
16b 17b 18b 10a
16c 17c 18c 11a
16d 17d 18d 12a
9b
10b
11b
12b
10c
11c
12c
10d
11d
12d
10e
11e
12e
10f
11f
12f
6
5
R
4
4
-H CC H
3
6
4
-FC H
6
4
Scheme 1. Synthesis of target compounds 9–13. Reaction conditions: (i) HCl, NaNO
2
, H
2
O, 0 °C; (ii) NaN
3
, 0 °C; (iii) Piperidine, DMSO, 70 °C; (iv) NH
2
NH
2
·H O,
2
+
EtOH, Reflux; (v) DMF, H
.
structures of the newly prepared sulfonamides were in full agreement
formation of structure 9a with position of CF
3
group at C-5 of 5-hy-
1
13
19
19
with their spectral data (IR, H NMR, C NMR, F NMR and HRMS). In
IR spectra of 9–13 the absorption frequencies due to OeH and NH
−1
droxypyrazoline [41–45]. Finally, the F NMR spectrum gave further
2
evidence to structure 9a as it displayed a signal at δ −76.8 ppm, which
−
1
groups appeared in the region 3656–3533 cm
and 3394–3194 cm
is typical for a C
5
–CF
3
of 5-hydroxypyrazoline rather than a C
3
–CF
3
respectively while absorption band corresponding to C]O and SO
2
which should be at around δ −67 ppm [42–45]. Further hydrazones 13
−
1
-1
13
appeared in the range of 1690–1643 cm
and 1358–1142 cm re-
exhibited a peak at δ 185 ppm in their C NMR spectra for carbonyl
spectively. On the other hand hydrazones 13 showed extra peak for C]
group which was at around δ 160 ppm in case of pyrazolines.
−
1
1
O absorption at 1720–1705 cm
.
H NMR spectra in DMSO-d of
6
compounds 9–13 revealed singlet at δ 8.58–8.34 ppm for OeH, while
free SO NH group resonated around δ 7.63–7.54 ppm as singlet in-
tegrating for two protons. Two characteristic doublets at
.15–3.65 ppm and δ 3.80–3.46 ppm for CH confirm the formation of
hydroxypyrazolines 9–12. At the same time, singlet at δ 4.78–4.72 ppm
2.2. CA inhibition studies
2
2
δ
The CA inhibitory profile of all the newly synthesized 28 com-
pounds 9–13 was evaluated in vitro for their ability to inhibit physio-
logically relevant hCA isoforms I and II (cytosolic) as well as hCA IX and
XII (transmembrane, tumor-associated isoforms) by means of the
4
2
for the CH group and another singlet at δ 11.90–11.81 ppm for NH
2
confirm the formation of hydrazones 13.
stopped-flow CO hydration assay [46] and their activities were com-
2
As there is a possibility of formation of two regioisomers 9a and 9a′
pared to the standard reference drug acetazolamide (AAZ). The fol-
lowing structure activity relationship can be compiled from the in-
hibition data (Table 1).
1
9
13
(
Fig. 2.), F and C NMR spectroscopy had been used to identify the
13
position of CF
3
group. In the C NMR spectra, the observed signal from
2
C-5 was a quartet at δ 91.84 ppm ( JCF = 33.7 Hz) and the C-3 atom
gave a singlet at δ 153.3 ppm. This supports 9a for if the structure was
1. The slow cytosolic isoforms hCA I was inhibited by the compounds
2
9
a′, the signal for the sp -hybridised C-3 atom would be a quartet with
9–13 with K ’s ranging between 41.4 and 947.7 nM. Compounds
i
a significant downfield shift around δ 141 ppm rather than the value
9a–9f, 10a–10c, 10f and 12a showed excellent inhibition activity
(K ≤ 100 nM) as compared to standard drug AAZ (K = 250 nM)
against hCA I. Among these, compounds 9d, 9e, 10a and 10f
emerged as the most efficient hCAI inhibitiors with K values of 41.4,
3
observed at δ 91.84 ppm, i.e. the CF
3
group is attached to an sp -hy-
i
i
bridised carbon rather than the C]N carbon atom and also appearance
1
of quartet for CF
3
around δ 123 ppm ( JCF = 284.0 Hz) confirmed the
i
200

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
Fig. 2. Proposed structure for product 9a and 9a′.
Table 1
carbonyl-1,2,3-triazoles 13 showed that the hydrazones 13 were
better inhibitors of hCA II.
Inhibition data of human CA isoforms hCA I, II, IX and XII with sulfonamides
9
–13 reported here and the standard sulfonamide inhibitor acetazolamide
3. The inhibition potential against tumor associated hCA IX isoform of
all newly synthesized compounds 9–13 investigated here, was in the
(
AAZ) by a stopped flow CO hydrase assay [46].
2
(nM)a
range of K = 0.7–206.9 nM. Furthermore twelve compounds 9a,
Compounds
R
Ar
K
i
i
9
c, 9d, 10a, 10d, 10f, 11f, 12a, 12c, 12d, 13a, 13b showed better
inhibition with K ≤ 24.5 nM for hCA IX as compared to reference
drug AAZ (K
hCA I
hCA II
hCA IX
hCA XII
i
i
= 25 nM). Indeed, two compounds 12f and 13d were
9
9
9
9
9
9
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
a
CH
CH
CH
CH
CH
CH
3
3
3
3
3
3
C
6
4-H
H
5
58.0
0.42
0.43
0.93
0.46
1.7
16.3
90.5
20.8
24.0
79.9
26.5
22.8
28.2
161.2
24.5
28.3
9.9
8.7
found to be respectively 35 and 17-fold more effective than the
standard drug AAZ.
b
3
CC
6
H
4
68.8
21.7
55.9
8.9
c
4-ClC
6
6
H
4
H
72.9
d
4-BrC
4
41.4
4
. Tumor associated isoform hCA XII was inhibited by all the synthe-
e
2-Naphthyl
2-Pyridyl
55.5
64.2
88.1
42.1
8.7
sized 28 novel compounds 9–13 with K values 0.87–98.0 nM.
i
f
86.4
23.5
0.57
6.3
Amongest all the tested compounds, two analogues 10f
(K = 0.87 nM), and 13b (K = 1.2 nM) displayed inhibitory poten-
0a
0b
0c
0d
0e
0f
1a
1b
1c
1d
1e
1f
2a
2b
2c
2d
2e
2f
3a
3b
3c
3d
C
C
C
C
C
C
6
6
6
6
6
6
4-H
4-H
4-H
4-H
4-H
4-H
H
H
H
H
H
H
5
5
5
5
5
5
C
6
4-H
H
5
3
53.1
CC
6
H
4
82.6
i
i
4-ClC
6
6
H
4
70.2
22.8
7.9
9.0
tial approximately 5-fold superior than the AAZ (K = 5.7 nM). It
i
4-BrC
H
4
265.5
172.4
41.5
7.6
can be concluded from the table that the trend of CA inhibitory
2-Naphthyl
27.8
1.9
10.2
0.87
73.9
52.1
98.0
60.4
4.2
potential against hCA XII was found to be 10f < 13b < 13d <
2-Pyridyl
1
1e = 12a < 12c with K ≤ 5.1 nM from the most effective to the
i
3
CC
CC
CC
CC
CC
CC
6
6
6
6
6
6
H
H
H
H
H
H
4
4
4
4
4
4
C
6
4-H
H
5
3
151.5
501.8
709.1
774.0
947.7
555.9
84.8
3.6
31.2
206.9
48.9
33.2
134.3
6.0
3
3
3
3
3
CC
6
H
4
24.5
53.3
30.6
48.3
38.6
0.90
78.2
7.2
least effective compound.
4-ClC
6
6
H
4
5
6
. In general hydrazones 13a-13d showed excellent inhibiting poten-
tial against hCA I, II, IX and XII except, compound 13c in which tolyl
substituent is present at C-5′ position of 1,2,3-triazole ring. It can be
concluded that bulkier group at C-5′ position of 1,2,3-triazole ring in
general reduces the CAI activity in the newly synthesized com-
pounds.
4-BrC
H
4
2-Naphthyl
2-Pyridyl
31.8
4.2
4-FC
4-FC
4-FC
4-FC
4-FC
4-FC
6
6
6
6
6
6
H
H
H
H
H
H
4
4
4
4
4
4
C
6
H
5
3
6.1
4-H
4-ClC
4-BrC
CC
6
H
4
240.4
518.0
541.9
661.2
692.1
69.8
27.0
14.4
23.6
25.6
0.7
6.0
6
6
H
4
5.1
H
4
8.4
56.9
9.0
. The selectivity ratio for inhibiting the tumor-associated isoforms
hCA IX and XII over the off-targeted cytosolic isoforms hCA I and II
has been presented in Table 2. It is evident that the compounds
didn’t show a consistent behavior in their property against all the
four tested isoforms (hCA I, II, IX and XII). It can be observed that
some of the investigated derivatives showed promising levels of
selective inhibition of the transmembrane over the cytosolic iso-
forms. It was also observed that the compounds which showed ex-
cellent inhibition of isoform hCA IX and XII were also shown to be
highly selective. Compound 12b and 12e showed maximum in-
hibition selectivity ratio for hCA IX and XII over hCA II and 12c
showed over hCA I. All other compounds of this series that is 12c-
2-Naphthyl
2-Pyridyl
2-Thienyl
2-Thienyl
2-Thienyl
2-Thienyl
–
94.3
2.5
24.1
7.7
CH
3
H
0.47
2.4
12.5
11.6
26.8
1.4
C
6
5
3
69.0
1.2
4-H
CC
6
4
H
4
91.1
9.5
34.7
2.4
4-FC
6
H
65.2
6.1
AAZ
–
250
12
25
5.7
AAZ = acetazolamide, reference compound, a standard sulfonamide CAI, is also
provided for comparison.
a
Mean from 3 different assays, by a stopped flow technique (errors were in
the range of ± 5–10% of the reported values).
5
5.5, 53.1 and 41.5 nM respectively, with more than 4-fold in-
1
2f were also shown to be excellently selective for hCA IX over hCA
creased activity than AAZ. Some of the compounds 11b–11f
and 12c–12f were weak inhibitors of off-targeted hCA I.
I having selectivity ratio more than 10. Thus, the biological activity
of these sulfonamides constitute an interesting precursor for ex-
ploring in more detail the hydroxy-trifluoromethylpyrazoline-car-
bonyl-1,2,3-triazoles 9–12 and trifluoromethylhydrazone-carbonyl-
Trifluoromethylhydrazone-carbonyl-1,2,3-triazole
13
emanate
better inhibition potential than hydroxy-trifloromethylpyrazoline-
carbonyl-1,2,3-triazole 9–12 against hCA I. In terms of SAR, in-
hibition potency against hCA I of tested compounds in general de-
creases as the substituent size increases at C-5′ of 1,2,3-triazole ring.
. The dominant cytosolic isoform hCA II was effectively inhibited by
most of the compounds 9a–9e, 10a, 10b, 10d, 10f, 11a, 12a, 12c,
1
,2,3-triazoles 13 incorporating benzenesulfonamide, for designing
isoform-selective metalloenzyme inhibitors.
2
3
. Conclusions
1
2d, 12f and 13a–13d with K in the range of sub-nanomolar to
i
In summary, this study reports the synthesis and unambiguous
single digit nanomolar 0.42–9.5 nM. Inhibition constant decreases
by expanding the substituent on aryl group at C-5′ of 1,2,3-triazole
ring. Overall comparison of the hydroxy-trifloromethylpyrazoline-
carbonyl-1,2,3-triazoles 9–12 with trifluoromethylhydrazone-
characterization of twenty four novel hydroxy-trifluoromethylpyrazo-
line-carbonyl-1,2,3-triazoles and four hydrazones. In light of the CA
inhibition profile, it can be concluded that synthesized sulfonamide
compounds investigated as inhibitors of four hCA isoforms, the
201

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
NMR, 13C NMR and F NMR spectra were recorded on Bruker Avance
19
Table 2
Selectivity ratios for inhibiting the tumor-associated isoforms hCA IX and XII
over cytosolic isoforms hCA I and II, with AAZ and compounds 9–13.
III 400 MHz, 100 MHz and 282.4 MHz respectively, using deuterated
dimethyl sulfoxide (DMSO-d ) as solvent, and tetramethylsilane (TMS)
6
*
as internal standard at room temperature. Chemical shifts are expressed
Compounds
R
Ar
Selectivity ratio
at δ (ppm) values relative to Me Si (TMS). The apparent resonance
4
I/IX
II/IX
I/XII
II/XII
multiplicity is described as: s (singlet), d (doublet), t (triplet), q (qua-
tret), dd (doublet of doublets), td (triplet of doublets) and m (multiplet)
for NMR assignments and strong (s), medium (m) for IR assignments.
The coupling constants in NMR are expressed in hertz (Hz). High re-
solution mass spectra were obtained from Xevo G2-S QTof UPLC/MS
spectrometer.
9
9
9
9
9
9
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
a
CH
CH
CH
CH
CH
CH
3
3
3
3
3
3
C
6
4-H
H
5
3.558
0.760
3.505
1.725
0.695
3.260
2.329
2.929
0.435
10.837
6.092
4.192
4.856
2.425
14.501
23.313
7.057
92.650
13.902
8.904
35.972
22.962
25.828
988.714
5.584
5.948
3.399
46.571
10.000
0.026
0.005
0.045
0.019
0.021
0.887
0.025
0.223
0.141
0.322
0.982
0.192
0.115
0.118
1.090
0.922
0.360
6.433
0.148
2.896
0.500
0.356
3.684
3.571
0.038
0.207
0.354
4.357
0.480
6.667
0.048
0.020
0.017
0.052
0.026
0.267
0.014
0.724
2.533
1.039
2.725
2.184
0.049
0.470
0.544
0.507
11.500
1.214
0.214
13.033
1.412
0.148
10.478
0.104
0.061
2.000
0.274
2.542
2.105
b
3
CC
6
H
4
3.171
c
4-ClC
6
6
H
4
H
1.304
d
4-BrC
4
4.652
e
2-Naphthyl
0.864
f
2-Pyridyl
0.981
0a
0b
0c
0d
0e
0f
1a
1b
1c
1d
1e
1f
2a
2b
2c
2d
2e
2f
3a
3b
3c
3d
C
C
C
C
C
C
6
6
6
6
6
6
H
H
H
H
H
H
5
5
5
5
5
5
CC
CC
CC
CC
CC
CC
C
6
4-H
H
5
1.261
3
CC
6
H
4
9.494
4.2. General procedure for the synthesis of 4-(4-(5-hydroxy-3-aryl-5-
(trifluoromethyl)-4,5-dihydro-1H-pyrazole-1-carbonyl)-5-alkyl/aryl-1H-
4-ClC
6
6
H
4
7.800
4-BrC
H
4
34.934
16.902
47.701
2.050
1
,2,3-triazol-1-yl)benzenesulfonamide (9–12) and 4-(5-alkyl/aryl-4-(2-
2-Naphthyl
2-Pyridyl
(
1,1,1-trifluoro-4-oxo-4-(thiophen-2-yl)butan-2-ylidene)hydrazine-1-
carbonyl)-1H-1,2,3-triazol-1-yl)benzenesulfonamide (13)
4-H
4-H
4-H
4-H
4-H
4-H
3
6
6
6
6
6
6
4
4
4
4
4
4
H
H
H
H
H
H
4
4
4
4
4
4
C
6
4-H
H
5
3
3
3
3
3
3
CC
6
H
4
9.631
4-ClC
6
6
H
4
7.236
To a solution of Carboxylic acid hydrazides of 1,2,3-triazole bearing
4-BrC
H
4
12.815
225.643
17.481
20.190
40.067
101.569
9.524
benzenesulfonamide 18 (1.1 mmol) and substituted 1,3-diketones 19
2-Naphthyl
2-Pyridyl
(
1.1 mmol) in DMF (15 ml), added conc. HCl (1 ml) and then stirred at
4-FC
4-FC
4-FC
4-FC
4-FC
4-FC
6
6
6
6
6
6
H
H
H
H
H
H
C
6
H
5
50 °C for 5 hrs. Thereafter, few drops of conc. H
2
SO
4
were added and
4-H
4-ClC
4-BrC
3
CC
6
H
4
the reaction mixture was further stirred at 90 °C in silicon oil bath for
6
6
H
4
9
–12 hrs. The reaction was monitored through TLC and after comple-
H
4
tion, the reaction mixture was allowed to cool and poured into ice cold
water to obtain a solid which was filtered and dried to afford crude
solid. Crude product thus obtained was recrystallized in appropriate
solvent. It is pertinent to mention here that under the same reaction
conditions thienyl substituted 1,3-diketones 19 on reacting with
Carboxylic acid hydrazides of 1,2,3-triazole bearing benzenesulfona-
mide 18 afforded corresponding hydrazone-carbonyl-1,2,3-triazoles 13.
2-Naphthyl
2-Pyridyl
2-Thienyl
2-Thienyl
2-Thienyl
2-Thienyl
–
73.467
28.718
9.065
CH
3
C
6
4-H
H
5
57.500
2.625
3
CC
6
H
4
4-FC
6
H
4
27.167
43.860
AAZ
–
AAZ = acetazolamide, reference compound, a standard sulfonamide CAI, is also
provided for comparison.
4.2.1. 4-(4-(5-Hydroxy-3-phenyl-5-(trifluoromethyl)-4,5-dihydro-1H-
pyrazole-1-carbonyl)-5-methyl-1H-1,2,3-triazol-1-yl)benzenesulfonamide
*
The K ratios are indicative of isozyme selectivity: a weak selective in-
i
hibitor is characterized by a low value ratio.
(
9a)
Recrystallized from ethanol; Yield 68%; Colour: Pale Yellow; mp:
10–212 °C; silica gel F-254 TLC R 0.63 (CHCl :CH OH, 90:10, v/v); IR
cytosolic CA I and II, as well as the transmembrane, tumor-associated
CA IX and XII, interesting inhibitory activities were observed against
almost all these isoforms. The most active inhibitors in both the series
2
f
3
3
−1
(
KBr) (ν, cm ): 3618 (s, OeH), 3356, 3263 (m, NeH stretch), 1666 (s,
1
C]O stretch), 1342, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
were found to be the compounds 9d and 10f, which had K value of
i
d
6
) δ (ppm): 8.39 (s, 1H, OH), 8.09 (d, J = 8.4 Hz, 2H, Ar), 7.97 (d,
H
around 41.5 nM for hCA I (involved in edema), and compound 9a, 9b,
J = 8.4 Hz, 2H, Ar), 7.77 (dd, J = 7.6 Hz, J = 2.0 Hz, 2H, Ar), 7.63 (s,
9
d and 13a with K value of around 0.45 nM for hCA II (an anti-
i
2
H, SO NH ), 7.51–7.46 (m, 3H, Ar), 4.02 (d, JHA-HB = 19.2 Hz, 1H,
2
2
glaucoma drug target). Compound 12f showed the most effective in-
pyrazoline), 3.67 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 2.44 (s, 3H,
hibitory action for the tumor associated isoform hCA IX (an isoform
13
CH
3
); C NMR (100 MHz, DMSO-d
6
) δ (ppm): 159.83, 152.68, 145.00,
C
involved in tumors) (K = 0.7 nM) being a better inhibitor compared to
i
1
1
40.03, 137.71, 136.21, 130.73, 129.92, 128.78, 127.08, 126.69,
the reference drug AAZ (K
i
= 25 nM). Compound 10f K
i
= 0.87 nM was
1
2
25.79, 123.20 (q,
JCF = 284.0 Hz, C
19
5
-CF
3
), 91.84 (q,
J
CF = 33.7 Hz,
–CF );
SH , calcd
found to be the most effective inhibitor against hCA XII. Our findings
identify the advantage of using compounds with flexible smaller sub-
stituents at the C-5 of 1,2,3-triazole ring may provide an avenue to
overcome CA isoform specificity, as they may unveil both nanomolar
affinity and preferential binding for the tumor associated membrane
bound isoforms.
C-5), 44.20, 9.79; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.05 (C
5
3
+
+
HRMS (ESI-MS) m/z 495.1057 (M + H) , C20
H
17
3
F N
6 4
O
4
95.1062.
4
.2.2. 4-(4-(5-Hydroxy-3-(p-tolyl)-5-(trifluoromethyl)-4,5-dihydro-1H-
pyrazole-1-carbonyl)-5-methyl-1H-1,2,3-triazol-1-yl)benzenesulfonamide
(
9b)
Recrystallized from ethanol; Yield 66%; Colour: White; mp:
4
. Experimental protocols
1
85–187 °C; silica gel F-254 TLC R
f
0.64 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−1
4.1. General
(KBr) (ν, cm ): 3626 (s, OeH), 3364, 3286 (m, NeH stretch), 1643 (s,
1
C]O stretch), 1327, 1149 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
All the chemicals were purchased from Alfa Aesar, Himedia and
d
6
) δ
J = 8.8 Hz, 2H, Ar), 7.66 (d, J = 8.0 Hz, 2H, Ar), 7.62 (s, 2H, SO
7.28 (d, J = 8.0 Hz, 2H, Ar), 3.97 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline),
3.64 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 2.43 (s, 3H, CH ), 2.35 (s,
H
(ppm): 8.34 (s, 1H, OH), 8.09 (d, J = 8.8 Hz, 2H, Ar), 7.96 (d,
used as received without further purification. Melting points were de-
termined in open glass capillaries in an electrical melting point appa-
ratus and are uncorrected. Analytical thin-layer chromatography (TLC)
was carried out on Merck silica gel F-254 aluminium plates using a
mixture of chloroform and methanol as eluent while UV lamp was used
to visualize the spots. Compounds were named following IUPAC rules
as applied by ChemDraw Professional 17.0. IR spectra were measured
2
NH ),
2
3
1
3
3H, CH
3
); C NMR (100 MHz, DMSO-d
6
) δ (ppm): 159.75, 152.61,
C
144.98, 140.66, 140.07, 137.70, 136.15, 129.34, 127.18, 127.07,
1
126.65, 125.77, 123.21 (q,
J
CF = 284.0 Hz,
C
5
-CF ), 91.62 (q,
3
2
19
J
CF = 33.1 Hz, C-5), 44.24, 20.96, 9.78;
F
NMR (DMSO-d
6
,
1
on ABB MB 3000 DTGS IR instrument using the KBr pellet technique. H
282.4 MHz) –δ 76.06 (C
5
–CF
3
); HRMS (ESI-MS) m/z 509.1212
202

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
(
M + H)+, C21
H
19
F
3
N
6
O
4
SH+, calcd 509.1219.
HB = 19.6 Hz, 1H, pyrazoline), 3.68 (d, JHA-HB = 19.6 Hz, 1H, pyrazo-
1
3
line), 2.45 (s, 3H, CH
3
); C NMR (100 MHz, DMSO-d
6
) δ
C
(ppm):
4
.2.3. 4-(4-(3-(4-Chlorophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
159.89, 153.48, 149.54, 148.92, 145.02, 139.79, 137.67, 137.09,
1
dihydro-1H-pyrazole-1-carbonyl)-5-methyl-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (9c)
136.48, 127.06, 125.80, 125.23, 123.12 (q,
J
CF = 284.0 Hz, C
5
-CF ),
3
2
19
120.81, 91.99 (q,
J
CF = 33.3 Hz, C-5), 44.07, 9.82; F NMR (DMSO-
Recrystallized from ethanol; Yield 72%; Colour: Brown; mp:
d
6
, 282.4 MHz) –δ 76.16 (C
5
–CF
3
); HRMS (ESI-MS) m/z 496.1006
+
+
2
05–207 °C; silica gel F-254 TLC R
f
3
0.64 (CHCl :CH
3
OH, 90:10, v/v); IR
(M + H) , C19
H
16
F
3
N O
7 4
SH , calcd 496.1015.
−
1
(
KBr) (ν, cm ): 3636 (s, OeH), 3340, 3248 (m, NeH stretch), 1690 (s,
1
C]O stretch), 1342, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
4.2.7. 4-(4-(5-Hydroxy-3-phenyl-5-(trifluoromethyl)-4,5-dihydro-1H-
pyrazole-1-carbonyl)-5-phenyl-1H-1,2,3-triazol-1-yl)benzenesulfonamide
(10a)
d
6
) δ
H
(ppm): 8.41 (s, 1H, OH), 8.08 (d, J = 8.8 Hz, 2H, Ar), 7.96 (d,
NH ),
.54 (d, J = 8.4 Hz, 2H, Ar), 4.03 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline),
.67 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 2.44 (s, 3H, CH ); C NMR
J = 8.8 Hz, 2H, Ar), 7.78 (d, J = 8.4 Hz, 2H, Ar), 7.62 (s, 2H, SO
2
2
7
3
Recrystallized from ethanol; Yield 72%; Colour: Yellow; mp:
1
3
3
208–210 °C; silica gel F-254 TLC R
f
0.64 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−1
(
100 MHz, DMSO-d
6
) δ
C
(ppm): 160.38, 152.30, 145.58, 140.47,
(KBr) (ν, cm ): 3622 (s, OeH), 3394, 3271 (m, NeH stretch), 1659 (s,
1
1
38.24, 136.95, 135.88, 129.43, 129.02, 127.63, 126.33, 123.69 (q,
C]O stretch), 1327, 1173 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
1
1
2
J
CF = 284.0 Hz, C
5
-CF
3
), 92.60 (q, JCF = 33.9 Hz, C-5), 44.66, 10.33;
d
6
) δ
J = 8.4 Hz, 2H, Ar), 7.57 (s, 2H, SO
7.30–7.26 (m, 5H, Ar), 3.75 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 3.50
(d, JHA-HB = 19.2 Hz, 1H, pyrazoline); C NMR (100 MHz, DMSO-d
(ppm): 159.46, 152.26, 144.84, 140.44, 137.90, 137.53, 130.61,
129.57, 129.37, 129.15, 128.45, 128.39, 126.82, 126.66, 126.30,
H
(ppm): 8.40 (s, 1H, OH), 7.94 (d, J = 8.4 Hz, 2H, Ar), 7.70 (d,
9
F NMR (DMSO-d
6
, 282.4 MHz) –δ 75.99 (C
5
–CF
3
); HRMS (ESI-MS) m/
2
NH ), 7.45–7.34 (m, 5H, Ar),
2
+
+
+
z 529.0668 (M + H) , 531.0638 (M + H + 2) , C20
H16ClF
3
N
6
O
4
SH
,
1
3
calcd 529.0672
6
)
δ
C
4
.2.4. 4-(4-(3-(4-Bromophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
1
2
dihydro-1H-pyrazole-1-carbonyl)-5-methyl-1H-1,2,3-triazol-1-yl)
125.58, 123.08 (q,
J
CF = 284.0 Hz, C
5
-CF
3
), 91.43 (q,
J
CF = 33.4 Hz,
–CF ); HRMS
SH , calcd 557.1219.
1
9
benzenesulfonamide (9d)
C-5), 44.00; F NMR (DMSO-d , 282.4 MHz) –δ 76.28 (C
6
5
3
+
+
Recrystallized from ethanol; Yield 64%; Colour: White; mp:
(ESI-MS) m/z 557.1212 (M + H) , C25
H
19
F
3
N
6 4
O
2
16–218 °C; silica gel F-254 TLC R
f
3
0.65 (CHCl :CH OH, 90:10, v/v); IR
3
−
1
(
KBr) (ν, cm ): 3644 (s, OeH), 3333, 3240 (m, NeH stretch), 1690 (s,
4.2.8. 4-(4-(5-Hydroxy-3-(p-tolyl)-5-(trifluoromethyl)-4,5-dihydro-1H-
pyrazole-1-carbonyl)-5-phenyl-1H-1,2,3-triazol-1-yl)benzenesulfonamide
(10b)
1
C]O stretch), 1350, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
d
6
) δ
H
(ppm): 8.42 (s, 1H, OH), 8.09 (dd, J = 8.8 Hz, J = 2.0 Hz, 2H,
Ar), 7.96 (dd, J = 8.8 Hz, J = 2.0 Hz, 2H, Ar), 7.73–7.67 (m, 4H, Ar),
Recrystallized from ethanol; Yield 67%; Colour: Yellow; mp:
7
.62 (s, 2H, SO NH ), 4.03 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 3.67
2
2
190–192 °C; silica gel F-254 TLC R
f
0.65 (CHCl
3
:CH OH, 90:10, v/v); IR
3
1
3
−1
(
d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 2.44 (s, 3H, CH ); C NMR
100 MHz, DMSO-d ) δ (ppm): 160.38, 152.40, 145.59, 140.46,
38.24, 136.97, 132.35, 129.77, 129.20, 127.64, 126.33, 124.73,
3
(KBr) (ν, cm ): 3656 (s, OeH), 3356, 3256 (m, NeH stretch), 1674 (s,
1
(
6
C
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
1
1
4
d
6
) δ (ppm): 8.35 (s, 1H, OH), 7.93 (d, J = 8.8 Hz, 2H, Ar), 7.68 (d,
H
1
2
23.83 (q,
J
CF = 284.0 Hz, C
5
-CF
3
), 92.60 (q,
J
CF = 33.4 Hz, C-5),
J = 8.8 Hz, 2H, Ar), 7.57 (s, 2H, SO
2
NH ), 7.34 (d, J = 8.0 Hz, 2H, Ar),
2
1
9
4.61, 10.34; F NMR (DMSO-d
6
, 282.4 MHz) –δ 75.98 (C
5
–CF
3
);
+
7.28 (s, 5H, Ar), 7.16 (d, J = 8.0 Hz, 2H, Ar), 3.71 (d, JHA-HB = 19.2 Hz,
+
HRMS (ESI-MS) m/z 573.0159 (M + H) , 575.0141 (M + H + 2)
,
1H, pyrazoline), 3.46 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 2.30 (s,
+
13
C
20
H
16BrF
3
N
6
O
4
SH , calcd 573.0167.
3H, CH
3
); C NMR (100 MHz, DMSO-d
6
) δ (ppm): 159.39, 152.21,
C
1
44.83, 140.51, 137.90, 137.48, 129.57, 129.14, 128.96, 128.44,
1
4
.2.5. 4-(4-(5-Hydroxy-3-(naphthalen-2-yl)-5-(trifluoromethyl)-4,5-
126.82, 126.68, 126.64, 126.29, 123.10, (q,
J
CF = 283.4 Hz, C
5
-CF ),
3
6
2
19
dihydro-1H-pyrazole-1-carbonyl)-5-methyl-1H-1,2,3-triazol-1-yl)
91.32 (q,
J
CF = 33.78 Hz, C-5), 44.07, 20.93; F NMR (DMSO-d
,
benzenesulfonamide (9e)
282.4 MHz) –δ 76.26 (C
5
–CF
3
); HRMS (ESI-MS) m/z 571.1365
Recrystallized from ethanol; Yield 66%; Colour: Pale Yellow; mp:
(M + H)+, C26
H
21
F
3
N
6
O
4
SH , calcd 571.1375.
+
2
37–239 °C; silica gel F-254 TLC R
f
3
0.64 (CHCl :CH OH, 90:10, v/v); IR
3
−
1
(
KBr) (ν, cm ): 3618 (s, OeH), 3364, 3271 (m, NeH stretch), 1659 (s,
4.2.9. 4-(4-(3-(4-Chlorophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
dihydro-1H-pyrazole-1-carbonyl)-5-phenyl-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (10c)
1
C]O stretch), 1335, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
d
6
8
) δ
H
(ppm): 8.45 (s, 1H, OH), 8.31 (s, 1H, Ar), 8.11–8.09 (m, 2H, Ar),
NH2, Ar), 4.15 (d, JHA-
.03–7.91 (m, 6H, Ar), 7.63–7.60 (m, 4H, SO
2
Recrystallized from ethanol; Yield 69%; Colour: Yellow; mp:
HB = 19.2 Hz, 1H, pyrazoline), 3.80 (d, JHA-HB = 19.2 Hz, 1H, pyrazo-
212–214 °C; silica gel F-254 TLC R
f
0.64 (CHCl
3
:CH OH, 90:10, v/v); IR
3
1
3
−1
line), 2.48 (s, 3H, CH
3
); C NMR (100 MHz, DMSO-d
6
) δ
C
(ppm):
(KBr) (ν, cm ): 3618 (s, OeH), 3356, 3263 (m, NeH stretch), 1682 (s,
1
1
60.38, 153.28, 145.57, 140.58, 138.29, 136.96, 134.25, 133.08,
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
1
29.03, 128.95, 128.60, 128.09, 127.65, 127.44, 126.39, 123.33 (q,
d
6
) δ (ppm): 8.43 (s, 1H, OH), 7.93 (dd, J = 8.8 Hz, J = 2.0 Hz, 2H,
H
1
2
J
CF = 284.0 Hz, C
5
-CF
3
), 123.20, 92.57 (q,
J
CF = 34.4 Hz, C-5),
Ar), 7.69 (dd, J = 8.8 Hz, J = 2.0 Hz, 2H, Ar), 7.57 (s, 2H, SO
2
NH ),
2
1
9
4
4.79, 10.39; F NMR (DMSO-d
6
, 282.4 MHz) –δ 75.97 (C
5
–CF
3
);
7.47–7.41 (m, 4H, Ar), 7.28 (s, 5H, Ar), 3.75 (d, JHA-HB = 19.2 Hz, 1H,
+
+
13
HRMS (ESI-MS) m/z 545.1212 (M + H) , C24
H
F
19 3
6
N O
4
SH , calcd
pyrazoline), 3.47 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline); C NMR
5
45.1219.
(100 MHz, DMSO-d
6
) δ
C
(ppm): 159.41, 151.29, 144.86, 140.31,
1
37.87, 137.70, 135.21, 129.60, 129.16, 128.49, 128.27, 126.83,
1
4
.2.6. 4-(4-(5-Hydroxy-3-(pyridin-2-yl)-5-(trifluoromethyl)-4,5-dihydro-
H-pyrazole-1-carbonyl)-5-methyl-1H-1,2,3-triazol-1-yl)
126.28, 125.61, 123.02 (q,
J
CF = 283.2 Hz,
C
5
-CF ), 91.61 (q,
3
2
19
1
J
CF = 34.0 Hz, C-5), 44.89; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.23
+
benzenesulfonamide (9f)
(C
5
–CF
3
); HRMS (ESI-MS) m/z 591.0826 (M + H)
,
593.0801
+
+
Recrystallized from ethanol; Yield 62%; Colour: Off White; mp:
(M + H + 2) , C25
H18ClF
3
N
6
O SH , calcd 591.0829.
4
2
04–206 °C; silica gel F-254 TLC R
f
3
0.63 (CHCl :CH OH, 90:10, v/v); IR
3
−
1
(
KBr) (ν, cm ): 3533 (s, OeH), 3348, 3256 (m, NeH stretch), 1674 (s,
4.2.10. 4-(4-(3-(4-Bromophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
dihydro-1H-pyrazole-1-carbonyl)-5-phenyl-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (10d)
1
C]O stretch), 1342, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
d
6
2
) δ
H, Ar), 7.98–7.95 (m, 3H, Ar), 7.91 (td, J = 7.6 Hz, J = 1.6 Hz, 1H,
NH ), 7.53–7.50 (m, 1H, Ar), 3.97 (d, JHA-
H
(ppm): 8.69–8.67 (m, 1H, Ar) 8.48 (s, 1H, OH), 8.10–8.07 (m,
Recrystallized from ethanol/THF; Yield 68%; Colour: Pale Yellow;
Ar), 7.62 (s, 2H, SO
2
2
mp: 215–217 °C; silica gel F-254 TLC R
f
0.63 (CHCl
3
:CH OH, 90:10, v/
3
203

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
−
1
1
2
v); IR (KBr) (ν, cm ): 3634 (s, OeH), 3356, 3263 (m, NeH stretch),
J
CF = 284.0 Hz, C
5
-CF
3
), 123.15, 92.04 (q,
J
CF = 34.4 Hz, C-5),
–CF );
SH , calcd
1
19
1
682 (s, C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz,
44.59, 21.22; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.30 (C
5
3
+
+
DMSO-d
d, J = 8.4 Hz, 2H, Ar), 7.58–7.55 (m, 4H, SO
J = 8.4 Hz, 2H, Ar), 7.28 (s, 5H, Ar), 3.75 (d, JHA-HB = 19.2 Hz, 1H,
pyrazoline), 3.48 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline); C NMR
(ppm): 159.41, 151.40, 144.86, 140.31,
6
) δ
H
(ppm): 8.41 (s, 1H, OH), 7.94 (d, J = 8.4 Hz, 2H, Ar), 7.68
NH2, Ar,), 7.38 (d,
HRMS (ESI-MS) m/z 571.1369 (M + H) , C26
H
21
F
3
N
6 4
O
(
2
571.1375.
1
3
4.2.14. 4-(4-(5-Hydroxy-3-(p-tolyl)-5-(trifluoromethyl)-4,5-dihydro-1H-
pyrazole-1-carbonyl)-5-(p-tolyl)-1H-1,2,3-triazol-1-yl)benzenesulfonamide
(11b)
(
100 MHz, DMSO-d
6
) δ
C
1
37.86, 137.70, 131.40, 129.60, 129.16, 128.61, 128.48, 126.82,
1
1
26.29, 125.60, 124.10, 123.01 (q, JCF = 285.0 Hz, C
5
-CF
3
), 91.61 (q,
Recrystallized from ethanol; Yield 67%; Colour: Pale Yellow; mp:
2
19
J
CF = 33.7 Hz, C-5), 43.84; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.22
208–210 °C; silica gel F-254 TLC R
f
0.65 (CHCl
3
:CH OH, 90:10, v/v); IR
3
+
−1
(
C
5
–CF
3
); HRMS (ESI-MS) m/z 635.0317 (M + H)
,
637.0300
(KBr) (ν, cm ): 3552 (s, OeH), 3364, 3263 (m, NeH stretch), 1674 (s,
+
+
1
(
M + H + 2) , C25
H
18BrF
3
N
6
O
4
SH , calcd 635.0324.
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
d
6
) δ
Ar), 7.68 (d, J = 8.4 Hz, 2H, Ar), 7.56 (s, 2H, SO
J = 8.4 Hz, 2H, Ar), 7.19–7.15 (m, 4H, Ar), 7.10 (d, J = 8.0 Hz, 2H,
Ar), 3.77 (d, HA-HB = 19.2 Hz, 1H, pyrazoline), 3.49 (d,
HB = 19.2 Hz, 1H, pyrazoline), 2.31 (s, 3H, CH ), 2.17 (s, 3H, CH C
H
(ppm): 8.34 (s, 1H, OH), 7.94 (dd, J = 8.4 Hz, J = 2.0 Hz, 2H,
4
.2.11. 4-(4-(5-Hydroxy-3-(naphthalen-2-yl)-5-(trifluoromethyl)-4,5-
2
NH ), 7.36 (d,
2
dihydro-1H-pyrazole-1-carbonyl)-5-phenyl-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (10e)
J
J
HA-
1
3
Recrystallized from ethanol; Yield 71%; Colour: Pale Yellow; mp:
3
3
);
2
29–231 °C; silica gel F-254 TLC R
f
3
0.65 (CHCl :CH
3
OH, 90:10, v/v); IR
NMR (100 MHz, DMSO-d
6
) δ (ppm): 159.54, 152.32, 144.80, 140.53,
C
−
1
(
KBr) (ν, cm ): 3610 (s, OeH), 3317, 3256 (m, NeH stretch), 1659 (s,
140.40, 139.30, 138.00, 137.45, 129.07, 129.04, 128.99, 126.86,
1
1
C]O stretch), 1327, 1173 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
126.82, 126.61, 126.27, 123.11 (q, JCF = 283.7 Hz, C
5
-CF ), 91.40 (q,
3
2
19
d
7
7
6
) δ
.84 (d, J = 8.8 Hz, 1H, Ar), 7.72 (dd, J = 6.8 Hz, J = 1.6 Hz, 2H, Ar),
.59–7.53 (m, 5H, SO NH2, Ar), 7.34 (dd, J = 8.0 Hz, J = 1.6 Hz, 2H,
H
(ppm): 8.44 (s, 1H, OH), 8.05 (s, 1H, Ar), 7.96–7.90 (m, 4H, Ar),
J
CF = 34.0 Hz, C-5), 122.58, 44.09, 20.93, 20.67; F NMR (DMSO-d
6
,
282.4 MHz) –δ 76.29 (C
5
–CF
3
); HRMS (ESI-MS) m/z 585.1523
(M + H)+, C27
H
F
23 3
N
6
O
SH , calcd 585.1532.
+
2
4
Ar) 7.29–7.22 (m, 3H, Ar), 3.87 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline),
1
3
3
.64 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline); C NMR (100 MHz, DMSO-
4.2.15. 4-(4-(3-(4-Chlorophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
dihydro-1H-pyrazole-1-carbonyl)-5-(p-tolyl)-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (11c)
d
1
1
6
) δ (ppm): 159.43, 152.25, 144.85, 140.40, 137.94, 137.70, 133.58,
C
32.26, 129.56. 129.20, 128.46, 128.40, 127.86, 127.73, 127.58,
1
27.43, 126.96, 126.83, 126.34, 125.66, 124.48 (q,
J
CF = 284.0 Hz,
Recrystallized from chloroform/THF; Yield 73%; Colour: White; mp:
2
19
C
5 3
-CF ), 122.98, 122.89, 91.54 (q,
J
CF = 34.1 Hz, C-5), 44.04;
F
206–208 °C; silica gel F-254 TLC R
f
0.63 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−1
NMR (DMSO-d
6
, 282.4 MHz) –δ 76.19 (C
5
–CF
3
); HRMS (ESI-MS) m/z
(KBr) (ν, cm ): 3642 (s, OeH), 3371, 3271 (m, NeH stretch), 1674 (s,
+
+
1
6
07.1368 (M + H) , C29
H
21
F
3
N
6
O
4
SH , calcd 607.1375.
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
d
6
) δ (ppm): 8.41 (s, 1H, OH), 7.94 (d, J = 8.4 Hz, 2H, Ar), 7.67 (d,
H
4
.2.12. 4-(4-(5-Hydroxy-3-(pyridin-2-yl)-5-(trifluoromethyl)-4,5-dihydro-
H-pyrazole-1-carbonyl)-5-phenyl-1H-1,2,3-triazol-1-yl)
J = 8.4 Hz, 2H, Ar), 7.56 (s, 2H, SO
2
NH ), 7.48–7.42 (m, 4H, Ar), 7.15
2
1
(d, J = 8.0 Hz, 2H, Ar), 7.09 (d, J = 8.0 Hz, 2H, Ar), 3.80 (d, JHA-
benzenesulfonamide (10f)
HB = 19.2 Hz, 1H, pyrazoline), 3.50 (d, JHA-HB = 19.2 Hz, 1H, pyrazo-
1
3
Recrystallized from ethanol; Yield 62%; Colour: White; mp:
line), 2.16 (s, 3H, CH
3
); C NMR (100 MHz, DMSO-d
6
) δ
C
(ppm):
2
10–212 °C; silica gel F-254 TLC R
f
3
0.64 (CHCl :CH
3
OH, 90:10, v/v); IR
159.52, 151.38, 144.82, 140.21, 139.32, 137.96, 135.21, 129.09,
−
1
1
(
KBr) (ν, cm ): 3618 (s, OeH), 3348, 3214 (m, NeH stretch), 1682 (s,
129.07, 128.49, 128.41, 126.85, 126.25, 123.04 (q,
J
CF = 283.7 Hz,
1
2
19
C]O stretch), 1335, 1173 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
C
5
-CF
3
), 122.61, 91.67 (q, JCF = 34.0 Hz, C-5), 43.91, 20.64; F NMR
d
6
) δ
H
(ppm): 8.56–8.54 (m, 1H, Ar) 8.49 (s, 1H, OH), 7.94 (dd,
(DMSO-d
6
,
282.4 MHz) –δ 76.25 (C
5
–CF ); HRMS (ESI-MS) m/z
+ +
3
+
J = 8.4 Hz, J = 1.6 Hz, 2H, Ar), 7.78 (td, J = 8.0 Hz, J = 1.6 Hz, 1H,
605.1075 (M + H) , 607.1047 (M + H + 2) , C26
H20ClF
N
3 6
4
O SH ,
Ar) 7.70 (dd, J = 8.4 Hz, J = 1.6 Hz, 2H, Ar), 7.57 (s, 2H, SO
2
NH ),
2
calcd 605.0985.
7
.53 (d, J = 8.0 Hz, 1H, Ar), 7.44–7.41 (m, 1H, Ar), 7.31–7.24 (m, 5H,
Ar), 3.65 (d,
J
HA-HB = 19.6 Hz, 1H, pyrazoline), 3.47 (d,
J
HA-
4.2.16. 4-(4-(3-(4-Bromophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
dihydro-1H-pyrazole-1-carbonyl)-5-(p-tolyl)-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (11d)
1
3
HB = 19.6 Hz, 1H, pyrazoline);
C NMR (100 MHz, DMSO-d
6
) δ
C
(
ppm): 159.49, 152.97, 149.23, 148.31, 144.87. 140.19, 137.89,
1
37.84, 136.58, 129.55, 129.18, 128.45, 126.81, 126.32, 125.62,
Recrystallized from ethanol/THF; Yield 67%; Colour: Pale Yellow;
1
1
25.13, 123.0 (q,
J
CF = 284.3 Hz,
C
5
-CF
3
), 120.93, 91.55 (q,
mp: 207–209 °C; silica gel F-254 TLC R
f
0.64 (CHCl
3
:CH OH, 90:10, v/
3
2
19
−1
J
CF = 33.9 Hz, C-5), 43.84; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.41
v); IR (KBr) (ν, cm ): 3564 (s, OeH), 3348, 3263 (m, NeH stretch),
1
(
C
5
24
–CF
3
F
);
HRMS
(ESI-MS)
m/z
558.1162
(M + H)+
,
1674 (s, C]O stretch), 1342, 1165 (s, SO
2
stretch); H NMR (400 MHz,
+
C
H
18
3
N
7
O
4
SH , calcd 558.1171.
DMSO-d
6
) δ (ppm): 8.41 (s, 1H, OH), 7.93 (d, J = 8.8 Hz, 2H, Ar), 7.67
H
(
d, J = 8.8 Hz, 2H, Ar), 7.59–7.56 (m, 4H, SO
2
NH , Ar), 7.39 (d,
2
4
.2.13. 4-(4-(5-Hydroxy-3-phenyl-5-(trifluoromethyl)-4,5-dihydro-1H-
pyrazole-1-carbonyl)-5-(p-tolyl)-1H-1,2,3-triazol-1-yl)benzenesulfonamide
11a)
Recrystallized from ethanol; Yield 65%; Colour: White; mp:
J = 8.8 Hz, 2H, Ar), 7.15 (d, J = 8.4 Hz, 2H, Ar), 7.09 (d, J = 8.4 Hz,
2H, Ar), 3.79 (d, JHA-HB = 19.6 Hz, 1H, pyrazoline), 3.49 (d, JHA-
1
3
(
HB = 19.6 Hz, 1H, pyrazoline), 2.16 (s, 3H, CH ); C NMR (100 MHz,
3
DMSO-d
6
) δ (ppm): 160.09, 152.06, 145.38, 140.77, 139.89, 138.52,
C
2
20–222 °C; silica gel F-254 TLC R
f
3
0.64 (CHCl :CH
3
OH, 90:10, v/v); IR
138.23, 131.99, 129.65, 129.32, 129.15, 127.41, 126.82, 124.64,
−
1
1
2
(
KBr) (ν, cm ): 3600 (s, OeH), 3333, 3256 (m, NeH stretch), 1690 (s,
123.58 (q,
JCF = 284.6 Hz, C
5
-CF ), 123.16, 92.25 (q, JCF = 33.9 Hz,
3
1
19
C]O stretch), 1350, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
C-5), 44.42, 21.20;
F
NMR (DMSO-d
6
,
282.4 MHz) –δ 76.25
+
d
6
) δ
J = 8.4 Hz, 2H, Ar), 7.58 (s, 2H, SO
d, J = 8.0 Hz, 2H, Ar), 7.09 (d, J = 8.0 Hz, 2H, Ar), 3.80 (d, JHA-
HB = 19.2 Hz, 1H, pyrazoline), 3.52 (d, JHA-HB = 19.2 Hz, 1H, pyrazo-
H
(ppm): 8.40 (s, 1H, OH), 7.94 (d, J = 8.4 Hz, 2H, Ar), 7.68 (d,
(C
5
–CF
3
); HRMS (ESI-MS) m/z 649.0472 (M + H)
,
651.4555
+
+
2
NH ), 7.47–7.36 (m, 5H, Ar), 7.16
2
(M + H + 2) ,C26
H
20BrF
3
N
6
O
4
SH , calcd 649.0498.
(
4.2.17. 4-(4-(5-Hydroxy-3-(naphthalen-2-yl)-5-(trifluoromethyl)-4,5-
dihydro-1H-pyrazole-1-carbonyl)-5-(p-tolyl)-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (11e)
1
3
line), 2.16 (s, 3H, CH
3
); C NMR (100 MHz, DMSO-d
60.14, 152.91, 145.37, 140.89, 139.86, 138.55, 138.07, 131.18,
30.07, 129.64, 128.98, 127.42, 127.20, 126.85, 125.34 (q,
6
) δ
C
(ppm):
1
1
Recrystallized from CHCl /THF; Yield 68%; Colour: White; mp:
3
204

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
1
3
2
30–232 °C; silica gel F-254 TLC R
f
0.65 (CHCl
3
:CH
3
OH, 90:10, v/v); IR
(d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 2.31 (s, 3H, CH
3
); C NMR
−
1
1
(
KBr) (ν, cm ): 3644 (s, OeH), 3364, 3263 (m, NeH stretch), 1666 (s,
(100 MHz, DMSO-d
6
) δ
C
(ppm): 162.47 (d,
J
CF = 246.7 Hz) 159.25,
1
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
152.34, 144.86, 140.57, 140.52, 127.77, 136.80, 131.70 (d,
3
1
2
d
7
5
6
) δ
.86 (d, J = 8.4 Hz, 1H, Ar), 7.71 (d, J = 8.4 Hz, 2H, Ar), 7.58–7.54 (m,
H, SO NH2, Ar), 7.20 (d, J = 8.0 Hz, 2H, Ar), 7.08 (d, J = 8.0 Hz, 2H,
HA-HB = 19.2 Hz, 1H, pyrazoline), 3.64 (d,
); C NMR (100 MHz,
(ppm): 159.56, 152.37, 144.81, 140.30, 139.28, 138.03,
H
(ppm): 8.45 (s, 1H, OH), 8.07 (s, 1H, Ar), 7.96–7.91 (m, 4H, Ar),
J
J
J
CF = 8.8 Hz) 129.00, 126.85, 126.65, 126.29, 123.11, (q,
4
CF = 284.0 Hz,
C
5
-CF
3
), 122.17 (d,
J
CF = 3.3 Hz), 115.61 (d,
2
19
2
CF = 21.9 Hz), 91.36 (q, JCF = 33.7 Hz, C-5), 44.04, 20.94; F NMR
Ar), 3.92 (d,
J
J
HA-
(DMSO-d
6
, 282.4 MHz) –δ 76.27 (C
5
–CF
3
), 110.92 (F); HRMS (ESI-MS)
1
3
+
+
HB = 19.2 Hz, 1H, pyrazoline), 2.09 (s, 3H, CH
3
m/z 589.1276 (M + H) , C26
H
20
F
4
N
6
O
4
SH , calcd 589.1281.
DMSO-d
6
) δ
C
1
1
1
37.64, 133.59, 132.29, 129.09, 128.38, 127.89, 127.72, 127.60,
4.2.21. 4-(4-(3-(4-Chlorophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
dihydro-1H-pyrazole-1-carbonyl)-5-(4-fluorophenyl)-1H-1,2,3-triazol-1-
yl)benzenesulfonamide (12c)
1
27.42, 127.09, 126.86, 126.76, 123.14 (q,
J
CF = 284.7 Hz, C
5
-CF ),
3
2
19
22.83, 122.65, 91.60 (q,
J
CF = 34.0 Hz, C-5),44.06, 20.60; F NMR
(
DMSO-d
6
,
282.4 MHz) –δ 76.22 (C
5
–CF
3
); HRMS (ESI-MS) m/z
Recrystallized from ethanol; Yield 68%; Colour: White; mp:
+
+
6
21.1526 (M + H) , C30
H
23
F
3
N
6
O
4
SH , calcd 621.1532.
215–217 °C; silica gel F-254 TLC R
f
0.66 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−
1
(
KBr) (ν, cm ): 3605 (s, OeH), 3356, 3263 (m, NeH stretch), 1674 (s,
1
4
.2.18. 4-(4-(5-Hydroxy-3-(pyridin-2-yl)-5-(trifluoromethyl)-4,5-dihydro-
H-pyrazole-1-carbonyl)-5-(p-tolyl)-1H-1,2,3-triazol-1-yl)
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
1
d
6
) δ (ppm): 8.43 (s, 1H, OH), 7.95 (d, J = 8.8 Hz, 2H, Ar), 7.70 (d,
H
benzenesulfonamide (11f)
J = 8.8 Hz, 2H, Ar), 7.58 (s, 2H, SO
2
NH ), 7.50 (d, J = 8.8 Hz, 2H, Ar),
2
Recrystallized from ethanol; Yield 59%; Colour: Off White; mp:
7.44 (d, J = 8.8 Hz, 2H, Ar), 7.38–7.34 (m, 2H, Ar), 7.15 (t, J = 8.8 Hz,
2
09–211 °C; silica gel F-254 TLC R
f
0.65 (CHCl
3
:CH
3
OH, 90:10, v/v); IR
2H, Ar), 3.80 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 3.49 (d, JHA-
−
1
13
(
KBr) (ν, cm ): 3604 (s, OeH), 3350, 3286 (m, NeH stretch), 1690 (s,
HB = 19.2 Hz, 1H, pyrazoline);
C NMR (100 MHz, DMSO-d
6
) δ
C
1
1
C]O stretch), 1327, 1157 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
(ppm): 162.49 (d,
J
CF = 246.7 Hz) 159.29, 151.42, 144.89, 140.366,
3
d
6
) δ
H
(ppm): 8.58 (dd, J = 4.0 Hz, J = 0.8 Hz, 1H, Ar), 8.49 (s, 1H,
137.74, 136.99, 135.28, 131.72 (d,
J
CF = 8.8 Hz) 128.54, 128.44,
OH), 7.94 (d, J = 8.4 Hz, 2H, Ar), 7.79 (td, J = 8.0 Hz, J = 1.6 Hz, 1H,
128.25, 126.87, 126.29, 123.04 (q, 1JCF = 283.5 Hz, C
5
-CF ), 122.18
3
4
2
Ar) 7.69 (d, J = 8.4 Hz, 2H, Ar), 7.57 (s, 2H, SO
2
NH
2
), 7.52 (d,
(d,
J
CF = 3.2 Hz), 115.69 (d,
JCF = 21.9 Hz), 91.67 (q,
2
19
J = 8.0 Hz, 1H, Ar), 7.43 (dd, J = 6.8 Hz, J = 1.6 Hz, 1H, Ar), 7.16 (d,
J = 8.0 Hz, 2H, Ar), 7.06 (d, J = 8.0 Hz, 2H, Ar), 3.69 (d, JHA-
HB = 19.2 Hz, 1H, pyrazoline), 3.50 (d, JHA-HB = 19.2 Hz, 1H, pyrazo-
J
CF = 33.8 Hz, C-5), 43.89; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.22
+
(C
5
–CF
3
), 110.87 (F); HRMS (ESI-MS) m/z 609.0726 (M + H)
,
+
+
611.0701 (M + H + 2) , C25
H
17ClF
4
N
6
O SH , calcd 609.0735.
4
1
3
line), 2.13 (s, 3H, CH
3
); C NMR (100 MHz, DMSO-d
6
) δ
C
(ppm):
1
60.15, 153.62, 149.82, 149.00, 145.40, 140.64, 139.88, 138.50,
4.2.22. 4-(4-(3-(4-Bromophenyl)-5-hydroxy-5-(trifluoromethyl)-4,5-
dihydro-1H-pyrazole-1-carbonyl)-5-(4-fluorophenyl)-1H-1,2,3-triazol-1-
yl)benzenesulfonamide (12d)
1
38.44, 137.16, 129.63, 127.41, 126.87, 125.69, 123.60 (q,
1
2
J
CF = 284.0 Hz, C
5
-CF
3
), 123.19, 121.47, 92.19 (q, JCF = 33.7 Hz, C-
1
9
5
), 44.41, 21.18; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.43 (C
5
–CF
3
);
Recrystallized from ethanol; Yield 69%; Colour: Off White; mp:
+
+
HRMS (ESI-MS) m/z 572.1325 (M + H) , C25
H
20
F
3
7
N O
4
SH , calcd
207–209 °C; silica gel F-254 TLC R
f
0.63 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−1
5
72.1328.
(KBr) (ν, cm ): 3610 (s, OeH), 3368, 3256 (m, NeH stretch), 1682 (s,
1
C]O stretch), 1350, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
4
.2.19. 4-(5-(4-Fluorophenyl)-4-(5-hydroxy-3-phenyl-5-(trifluoromethyl)-4,5-
d
6
) δ
Ar), 7.68 (dd, J = 6.8 Hz, J = 1.6 Hz, 2H, Ar), 7.59–7.57 (m, 2H,
SO NH , 2H, Ar), 7.42 (d, J = 8.8 Hz, 2H, Ar), 7.37–7.33 (m, 2H, Ar),
H
(ppm): 8.58 (s, 1H, OH), 7.95 (dd, J = 6.8 Hz, J = 1.6 Hz, 2H,
dihydro-1H- pyrazole-1-carbonyl)-1H-1,2,3-triazol-1-yl)benzenesulfonamide
(12a)
2
2
Recrystallized from ethanol; Yield 66%; Colour: Brown; mp:
7.15 (t, J = 8.8 Hz, 2H, Ar), 3.80 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline),
1
3
2
08–210 °C; silica gel F-254 TLC R
f
0.64 (CHCl
3
:CH
3
OH, 90:10, v/v); IR
3.49 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline); C NMR (100 MHz, DMSO-
−
1
1
(
KBr) (ν, cm ): 3618 (s, OeH), 3356, 3271 (m, NeH stretch), 1674 (s,
d
6
) δ
C
(ppm): 162.33 (d,
J
CF = 249.6 Hz), 159.26, 151.54, 144.87,
1
3
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
140.34, 137.73, 137.00, 131.73 (d,
J
CF = 8.8 Hz), 131.45, 129.64,
1
d
6
) δ
H
(ppm): 8.39 (s, 1H, OH), 7.94 (dd, J = 7.2 Hz, J = 1.8 Hz, 2H,
NH ),
.49–7.47 (m, 2H, Ar), 7.44–7.42 (m, 1H, Ar), 7.39–7.34 (m, 4H, Ar),
127.86, 126.29, 123.17 (q,
J
CF = 284.5 Hz, C
5
-CF
3
), 122.17 (d,
4
2
2
Ar), 7.70 (dd, J = 7.2 Hz, J = 1.8 Hz, 2H, Ar), 7.57 (s, 2H, SO
2
2
J
CF = 3.3 Hz), 115.66 (d, JCF = 21.9 Hz), 91.65 (q, JCF = 33.2 Hz, C-
1
9
7
7
5), 43.82; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.22 (C
5
–CF
3
), 110.88
+
.16–7.11 (m, 2H, Ar), 3.80 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 3.50
(F); HRMS (ESI-MS) m/z 653.0225 (M + H)
,
655.0208
1
3
+
+
(
d, JHA-HB = 19.2 Hz, 1H, pyrazoline); C NMR (100 MHz, DMSO-d
6
)
(M + H + 2) , C25
H
17BrF
4
N
6
O
4
SH , calcd 653.0229.
1
δ
C
1
(ppm): 162.46 (d, JCF = 246.4 Hz) 159.31, 152.38, 144.87, 140.47,
3
37.77, 136.85, 131.70 (d,
J
CF = 8.8 Hz) 130.67, 129.36, 128.42,
4.2.23. 4-(5-(4-Fluorophenyl)-4-(5-hydroxy-3-(naphthalen-2-yl)-5-
(trifluoromethyl)-4,5-dihydro-1H-pyrazole-1-carbonyl)-1H-1,2,3-triazol-1-
yl)benzenesulfonamide (12e)
1
1
26.85, 126.67, 126.31, 123.06 (q,
J
CF = 283.0 Hz, C
5
-CF ), 122.17
3
4
2
(
d,
J
CF = 3.2 Hz), 115.63 (d,
JCF = 21.9 Hz), 91.48 (q,
2
19
J
CF = 33.6 Hz, C-5), 43.98; F NMR (DMSO-d
6
, 282.4 MHz) –δ 76.28
Recrystallized from ethanol; Yield 67%; Colour: Brown; mp:
(
C
5
25
–CF
3
), 110.93 (F); HRMS (ESI-MS) m/z 575.1118 (M + H)+
,
233–235 °C; silica gel F-254 TLC R
f
0.65 (CHCl
3
:CH OH, 90:10, v/v); IR
3
+
−1
C
H
18
F
N
4 6
O
4
SH , calcd 575.1124.
(KBr) (ν, cm ): 3590 (s, OeH), 3387, 3294 (m, NeH stretch), 1666 (s,
1
C]O stretch), 1335, 1173 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
4
.2.20. 4-(5-(4-Fluorophenyl)-4-(5-hydroxy-3-(p-tolyl)-5-
d
6
) δ (ppm): 8.44 (s, 1H, OH), 8.08 (s, 1H, Ar), 7.97–7.85 (m, 5H, Ar),
H
(
trifluoromethyl)-4,5-dihydro-1H-pyrazole-1-carbonyl)-1H-1,2,3-triazol-1-
7.72 (d, J = 8.8 Hz, 2H, Ar), 7.58–7.56 (m, 2H, SO
2
NH , 3H, Ar),
2
yl)benzenesulfonamide (12b)
7.43–7.39 (m, 2H, Ar), 7.13 (t, J = 8.8 Hz, 2H, Ar), 3.92 (d, JHA-
Recrystallized from ethanol; Yield 64%; Colour: Yellow; mp:
HB = 19.2 Hz, 1H, pyrazoline), 3.61 (d, JHA-HB = 19.2 Hz, 1H, pyrazo-
1
3
2
10–212 °C; silica gel F-254 TLC R
f
0.64 (CHCl
3
:CH
3
OH, 90:10, v/v); IR
line);
C
NMR (100 MHz, DMSO-d
6
)
δ
C
(ppm): 163.03 (d,
−
1
1
(
KBr) (ν, cm ): 3618 (s, OeH), 3350, 3210 (m, NeH stretch), 1674 (s,
J
CF = 245.9 Hz), 159.85, 152.97, 145.44, 141.01, 138.38, 137.55,
1
3
C]O stretch), 1335, 1173 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
134.17, 132.85, 132.32 (d,
J
CF = 8.8 Hz), 128.99, 128.48, 128.35,
d
6
) δ
J = 8.4 Hz, 2H, Ar), 7.56 (s, 2H, SO
.19–7.12 (m, 4H, Ar), 3.75 (d, JHA-HB = 19.2 Hz, 1H, pyrazoline), 3.47
H
(ppm): 8.34 (s, 1H, OH), 7.94 (d, J = 8.4 Hz, 2H, Ar), 7.70 (d,
128.17, 128.03, 127.50, 127.42, 126.91, 123.42, 123.09 (q,
1
2
4
2
NH ), 7.39–7.34 (m, 4H, Ar),
2
J
J
CF = 282.5 Hz,
C
5
-CF
3
), 122.77 (d,
J
CF = 3.2 Hz), 116.19, (d,
2
19
7
CF = 22.2 Hz), 92.16 (q,
J
CF = 33.8 Hz, C-5), 44.60;
F NMR
205

V. Sharma et al.
Bioorganic Chemistry 85 (2019) 198–208
1
3
(
DMSO-d
6
, 282.4 MHz) –δ 76.18 (C
5
–CF
3
), 110.94 (F); HRMS (ESI-MS)
3H, CH
3
); C NMR (100 MHz, DMSO-d
6
) δ (ppm): 185.86, 145.55,
C
+
+
m/z 625.1275 (M + H) , C29
H
20
F
4
N
6
O
4
SH , calcd 625.1281.
142.54, 140.30, 138.23, 136.46, 135.48, 130.54, 129.53, 129.44,
1
19
1
27.42, 126.91, 125.84 (q,
J
CF = 237.8 Hz), 37.28, 21.39; F NMR
4
.2.24. 4-(5-(4-Fluorophenyl)-4-(5-hydroxy-3-(pyridin-2-yl)-5-
(DMSO-d
6
, 282.4 MHz) –δ 68.64 (CF
3
); HRMS (ESI-MS) m/z 577.0932
+
+
(
trifluoromethyl)-4,5-dihydro-1H-pyrazole-1-carbonyl)-1H-1,2,3-triazol-1-
(M + H) , C24
H
F
19 3
N
6
O
4
S H , calcd 577.0939.
2
yl)benzenesulfonamide (12f)
Recrystallized from ethanol; Yield 60%; Colour: Brown; mp:
4.2.28. 4-(5-(4-fluorophenyl)-4-(2-(1,1,1-trifluoro-4-oxo-4-(thiophen-2-
yl)butan-2-ylidene)hydrazine-1-carbonyl)-1H-1,2,3-triazol-1-yl)
benzenesulfonamide (13d)
2
09–211 °C; silica gel F-254 TLC R
f
0.64 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−
1
(
KBr) (ν, cm ): 3618 (s, OeH), 3364, 3263 (m, NeH stretch), 1682 (s,
1
C]O stretch), 1327, 1165 (s, SO
2
stretch); H NMR (400 MHz, DMSO-
Recrystallized from ethanol; Yield 74%; Colour: Yellow; mp:
d
6
) δ
J = 8.4 Hz, 2H, Ar), 7.79 (t, J = 8.0 Hz, 1H, Ar) 7.70 (d, J = 8.4 Hz, 2H,
Ar), 7.57 (s, 3H, SO NH , 1H, Ar), 7.44 (t, J = 6.2 Hz, 1H, Ar),
.38–7.35 (m, 2H, Ar), 7.11 (t, J = 8.8 Hz, 2H, Ar), 3.70 (d, JHA-
H
(ppm): 8.57 (d, J = 4.8 Hz, 1H, Ar), 8.48 (s, 1H, OH), 7.94 (d,
138–140 °C; silica gel F-254 TLC R
f
0.70 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−
1
(KBr) (ν, cm ): 3364, 3271 (m, NeH stretch), 1705, 1643 (s, C]O
1
2
2
stretch), 1358, 1157 (s, SO
2
stretch); H NMR (400 MHz, DMSO-d
6
) δ
H
7
(ppm): 11.90 (s, 1H, Ar), 8.13 (m, 2H, Ar), 7.90 (d, J = 8.4 Hz, 2H, Ar),
HB = 19.6 Hz, 1H, pyrazoline), 3.50 (d, JHA-HB = 19.6 Hz, 1H, pyrazo-
7.59 (d, J = 8.4 Hz, 2H, Ar), 7.54 (s, 2H, SO
2
NH ), 7.44 (s, 2H, Ar),
2
1
3
13
line);
C
NMR (100 MHz, DMSO-d
6
)
δ
C
(ppm): 162.34 (d,
7.34–7.25 (m, 3H, Ar), 4.73 (s, 2H, CH
2
); C NMR (100 MHz, DMSO-
1
J
CF = 249.4 Hz), 159.35, 153.13, 149.30, 148.33, 144.90, 137.72,
d
6
) δ (ppm): 185.97, 145.62, 142.54, 139.64, 138.02, 136.49, 135.49,
C
3
1
1
1
1
37.17, 136.65, 131.76 (d,
J
CF = 8.7 Hz), 126.84, 126.32, 125.18,
129.45, 127.44, 126.93, 125.40 (q,
J
CF = 240.6 Hz), 116.20, 115.98,
1
4
19
23.34 (q, JCF = 283.0 Hz, C
5
-CF
3
), 122.20, 120.90 (d, JCF = 3.2 Hz),
96.73, 37.32; F NMR (DMSO-d
6
, 282.4 MHz) –δ 68.67 (CF
3
), 110.43
2
2
19
+
+
15.62, (d,
J
CF = 22.0 Hz), 91.28 (q,
J
CF = 33.2 Hz, C-5), 43.74;
F
(F); HRMS (ESI-MS) m/z 581.0785 (M + H) , C23
H
F
16 4
N
6
O
4
S
2
H
,
NMR (DMSO-d
6
, 282.4 MHz) –δ 76.40 (C
5
–CF
3
), 110.90 (F); HRMS
calcd 581.0689.
+
+
(
ESI-MS) m/z 576.1069 (M + H) , C24
H
17
F N
4 7
O
4
SH , calcd 576.1077.
5
. CA inhibition assay
4
.2.25. 4-(5-Methyl-4-(2-(1,1,1-trifluoro-4-oxo-4-(thiophen-2-yl)butan-2-
ylidene)hydrazine-1-carbonyl)-1H-1,2,3-triazol-1-yl)benzenesulfonamide
An SX.18MV-R Applied Photophysics (Oxford, UK) stopped-flow
(
13a)
Recrystallized from ethanol; Yield 76%; Colour: Off White; mp:
instrument has been used to assay the inhibition of various CA isozymes
[47]. Phenol Red (at a concentration of 0.2 mM) has been used as an
indicator, working at the absorbance maximum of 557 nm, with 10 mM
1
42–144 °C; silica gel F-254 TLC R
f
0.70 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−
1
(
KBr) (ν, cm ): 3333, 3217 (m, NeH stretch), 1720, 1682 (s, C]O
Hepes (pH 7.4) as a buffer, 0.1 M Na
2
SO
4
or NaClO (for maintaining
4
1
stretch), 1327, 1142 (s, SO
2
stretch); H NMR (400 MHz, DMSO-d
6
) δ
H
constant the ionic strength; these anions are not inhibitory in the used
concentration), following the CA-catalyzed CO hydration reaction for a
period of 5–10 s. Saturated CO solutions in water at 25 °C were used as
(
ppm): 11.81 (s, 1H, NH), 8.16 (dd, J = 4.8 Hz, J = 0.8 Hz, 1H, Ar),
2
8
.13 (dd, J = 6.0 Hz, J = 6.0 Hz, 1H, Ar), 8.07 (dd, J = 6.8 Hz,
2
J = 1.8 Hz, 2H, Ar), 7.89 (dd, J = 6.8 Hz, J = 1.8 Hz, 2H, Ar), 7.61 (s,
substrate. Stock solutions of inhibitors were prepared at a concentration
of 10 mM (in DMSO-water 1:1, v/v) and dilutions up to 0.01 nM done
with the assay buffer mentioned above. At least 7 different inhibitor
concentrations have been used for measuring the inhibition constant.
Inhibitor and enzyme solutions were pre-incubated together for 10 min
at room temperature prior to assay, in order to allow for the formation
of the E-I complex. Triplicate experiments were done for each inhibitor
concentration, and the values reported throughout the paper are the
mean of such results. The inhibition constants were obtained by non-
linear least-squares methods using the Cheng-Prusoff equation, as re-
ported earlier, and represent the mean from at least three different
determinations [47–49]. All CA isozymes used here were recombinant
proteins obtained as reported earlier by our group.
2
H, SO
2
NH
2
), 7.35–7.34(m, 1H, Ar), 4.78 (s, 2H, CH
2
), 2.59 (s, 3H,
1
3
CH
3
); C NMR (100 MHz, DMSO-d ) δ (ppm): 185.44, 145.25, 145.06,
6
C
1
1
2
42.02, 138.53, 137.30, 135.95, 134.96, 128.89, 127.09, 125.85,
1
19
25.78 (q,
J
CF = 240.2 Hz), 34.70, 9.42;
F
NMR (DMSO-d
6
,
,
+
82.4 MHz) –δ 68.57 (CF
3
); HRMS (ESI-MS) m/z 501.0616 (M + H)
+
C
18
H
15
F
N
3 6
O
4
S
2
H
, calcd 501.0626.
4
.2.26. 4-(5-Phenyl-4-(2-(1,1,1-trifluoro-4-oxo-4-(thiophen-2-yl)butan-2-
ylidene)hydrazine-1-carbonyl)-1H-1,2,3-triazol-1-yl)benzenesulfonamide
13b)
Recrystallized from ethanol; Yield 71%; Colour: Brown; mp:
(
1
49–151 °C; silica gel F-254 TLC R
f
0.69 (CHCl
3
:CH OH, 90:10, v/v); IR
3
−
1
(
KBr) (ν, cm ): 3310, 3194 (m, NeH stretch), 1705, 1659 (s, C]O
1
stretch), 1350, 1173 (s, SO
2
stretch); H NMR (400 MHz, DMSO-d
6
) δ
H
Disclosure statement
(
ppm): 11.90 (s, 1H, Ar), 8.13 (m, 2H, Ar), 7.90 (d, J = 8.8 Hz, 2H, Ar),
7
.60 (d, J = 8.8 Hz, 2H, Ar), 7.54 (s, 2H, SO
2
NH
2
), 7.47–7.38 (m, 5H,
The authors report no conflicts of interest. The authors alone are
responsible for the content and writing of this article.
1
3
Ar), 7.34 (t, J = 8.8.4 Hz, J = 4.4 Hz, 1H, Ar), 4.72 (s, 2H, CH
2
);
C
NMR (100 MHz, DMSO-d
6
) δ
C
(ppm): 183.71, 145.03, 141.98, 137.74,
1
1
37.60, 134.90, 133.49, 130.08, 129.94, 128.88, 128.35, 126.83,
Acknowledgement
1
19
26.34, 126.12 (q,
J
CF = 240.0 Hz), 124.76, 114.50, 36.71; F NMR
); HRMS (ESI-MS) m/z 563.0776
, calcd 563.0783.
(
(
DMSO-d
6
, 282.4 MHz) –δ 68.66 (CF
3
One of the authors (Vikas Sharma) is grateful to the Council of
Scientific and Industrial Research, New Delhi, India for the award of
Junior Research Fellowship and the other (Rajiv Kumar), is thankful to
University Grants Commission, New Delhi, India for the award of Senior
Research Fellowship. Authors are also thankful to Materials Research
Centre, MNIT Jaipur for providing HRMS facility.
+
+
M + H) , C23
H
17
F
3
N
6
O
4
2
S H
4
.2.27. 4-(5-(p-Tolyl)-4-(2-(1,1,1-trifluoro-4-oxo-4-(thiophen-2-yl)butan-
-ylidene)hydrazine-1-carbonyl)-1H-1,2,3-triazol-1-yl)benzenesulfonamide
2
(
13c)
Recrystallized from ethanol; Yield 77%; Colour: Pale Yellow; mp:
1
45–147 °C; silica gel F-254 TLC R
f
0.71 (CHCl
3
:CH
3
OH, 90:10, v/v); IR
References
−
1
(
KBr) (ν, cm ): 3343, 3256 (m, NeH stretch), 1713, 1643 (s, C]O
1
stretch), 1358, 1157 (s, SO
2
stretch); H NMR (400 MHz, DMSO-d
6
) δ
H
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