Synthesis of 4-(2-substituted hydrazinyl)benzenesulfonamides and their carbonic anhydrase inhibitory effects

Article abstract of DOI:10.3109/14756366.2015.1047359

In this study, 4-(2-substituted hydrazinyl)benzenesulfonamides were synthesized by microwave irradiation and their chemical structures were confirmed by ¹H NMR, ¹³CNMR, and HRMS. Ketones used were: Acetophenone (S1), 4-methylacetophe

Full text of DOI:10.3109/14756366.2015.1047359

http://informahealthcare.com/enz
ISSN: 1475-6366 (print), 1475-6374 (electronic)
J Enzyme Inhib Med Chem, Early Online: 1–6
2015 Informa UK Ltd. DOI: 10.3109/14756366.2015.1047359
!
RESEARCH ARTICLE
Synthesis of 4-(2-substituted hydrazinyl)benzenesulfonamides and their
carbonic anhydrase inhibitory effects
Halise Inci Gul1, Kaan Kucukoglu1, Cem Yamali1, Sinan Bilginer1, Hafize Yuca2, Iknur Ozturk2, Parham Taslimi3,
Ilhami Gulcin3,4, and Claudiu T. Supuran5
2
1Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Ataturk University, Erzurum, Turkey, Faculty of Pharmacy, Ataturk University,
3
4
Erzurum, Turkey, Department of Chemistry, Faculty of Science, Ataturk University, Erzurum, Turkey, Department of Zoology, College of Science,
5
King Saud University, Riyadh, Saudi Arabia, and Polo Scientifico, Laboratorio di Chimica Bioinorganica, Universita degli Studi di Firenze,
Sesto Fiorentino (Firenze), Italy
Abstract
Keywords
In this study, 4-(2-substituted hydrazinyl)benzenesulfonamides were synthesized by microwave
irradiation and their chemical structures were confirmed by 1H NMR, 13CNMR, and HRMS.
Ketones used were: Acetophenone (S1), 4-methylacetophenone (S2), 4-chloroacetophenone
(S3), 4-fluoroacetophenone (S4), 4-bromoacetophenone (S5), 4-methoxyacetophenone (S6),
4-nitroacetophenone (S7), 2-acetylthiophene (S8), 2-acetylfuran (S9), 1-indanone (S10),
2-indanone (S11). The compounds S9, S10 and S11 were reported for the first time, while
S1–S8 was synthesized by different method than literature reported using microwave
irradiation method instead of conventional heating in this study. The inhibitory effects of 4-(2-
substituted hydrazinyl)benzenesulfonamide derivatives (S1–S11) against hCA I and II were
studied. Cytosolic hCA I and II isoenzymes were potently inhibited by new synthesized
sulphonamide derivatives with Kis in the range of 1.79 0.22–2.73 0.08 nM against hCA I and
in the range of 1.72 0.58–11.64 5.21 nM against hCA II, respectively.
2-Acethylfuran, 2-acethylthiophene,
acetophenones, carbonic anhydrase,
enzyme inhibition, indanone, sulfonamide
History
Received 30 March 2015
Revised 17 April 2015
Accepted 29 April 2015
Published online 5 June 2015
Introduction
Carbonic anhydrases (CAs, EC 4.2.1.1) are zinc (Zn2+)-contain-
ing metalloenzymes that catalyze the reversible hydration of car-
to the imidazole groups of three histidine residues and to the
substrate H2O/–OH that reacts with CO214–17. There are five
cytosolic forms (CA I, II, III, VII and XIII), five membrane
associated isozymes (CA IV, IX, XII, XIV and XV), two mito-
chondrial forms (CA VA and VB), and a secreted CA isoenzyme
(CA VI). There are three additional non-catalytic CA isoforms
bon dioxide (CO2) to bicarbonate (HCOꢀ3 ) and a proton (H+)1–3
:
CO2 þ H2O , HCOꢀ3 þ Hþ
(CA VIII, X and XI) whose functions remain unclear18–20
.
An understanding of the impact of this crucial equilibrium to
human health continues to develop; e.g. the CA-mediated
regulation of pH in the hypoxic tumor microenvironment is
proposed for using as a therapeutic target4–7. There are six main
classes of these enzymes: a-, b-, g-, d-, z- and Z-CAs8,9. The a-,
b-, d- and Z-CAs contain a Zn2+ ion at the active site, the g-CAs
are probably Fe2+ enzymes, while the metal ion is usually
replaced by Cd2+ in the z-CAs10. Humans encode 12 catalytically
active a-CA isozymes, which differ by molecular features,
oligomeric arrangement, cellular localization, distribution in
It was reported that the sulfonamide compounds (R–SO2NH2)
coordinate to the active site Zn2+ and to block the reaction
catalysis. The CAs inhibition has been of therapeutic interest for
several decades. The clinical usage of carbonic anhydrase
inhibitor (CAIs) has been established as antiglaucoma agents,
diuretics and antiepileptic. CAIs were also used in the treatment
of mountain sickness, osteoporosis, gastric and duodenal ulcers,
and neurological disorders21–23
.
The aim of the study was to investigate the CA I and II
inhibiting properties of the compounds to be synthesized. For
this aim, ketones were changed as acetophenones derivatives,
which carry a substituent at 4-position of phenyl ring that has
electron donating or attracting property, 2-acethylthiophene,
2-acethylfuran, 1-indanone and 2-indanone.
organs and tissues, expression levels, and kinetic properties11–13
.
These CAs comprise CA I, II, III, IV, VA, VB, VI, VII, IX, XII,
XIII and XIV, all of which contain a zinc ion (Zn2+) coordinated
Materials and methods
Addresses for correspondence: Prof. Dr Halise Inci Gul, Department of
Pharmaceutical Chemistry, Faculty of Pharmacy, Ataturk University,
Erzurum, Turkey. Tel: +90 442 231 5219. Fax: +90 442 231 5201.
E-mail: incigul1967@yahoo.com
Prof. Dr Ilhami Gulcin, Department of Chemistry, Faculty of Science,
Ataturk University, Erzurum, Turkey. Tel: +90 442 231 4375. Fax: +90
442 231 4109. E-mail: igulcin@atauni.edu.tr
1H NMR (400 MHz) and 13C NMR (100 MHz) spectra were
obtained using a Varian Mercury Plus spectrometer (Varian Inc.,
Palo Alto, CA). Chemical shifts (d) are reported in ppm. Mass
spectra were undertaken on an HPLC-TOF Waters Micromass
LCT Premier XE (Waters Corporation, Milford, MA) mass

2
H. I. Gul et al.
J Enzyme Inhib Med Chem, Early Online: 1–6
spectrometer using an electrospray ion source (ESI). Melting 2-indanone (S11)], 4-hydrazinobenzenesulfonamide hydrochlor-
points were determined using an Electrothermal 9100 (IA9100, ide (9 mmol) and sodium acetate (9 mmol) were dissolved in
Bibby Scientific Limited, Staffordshire, UK) instrument and are ethanol (20 mL). Reaction mixture was heated for 60 min at 78 ꢁC,
uncorrected. Reactions were carried out in a CEM Discover 150 W in a microwave oven. Reactions were followed by thin
Microwave Synthesis System, 908010 (Matthews, NC).
layer chromatography (TLC) using methanol:chloroform (4.5:0.5)
solvent system. Ethanol was removed under vacuum. A solid
separated out was filtered, dried and crystallized from ethanol or
H2O:N,N-dimethylformamide (1:4) to afford S1–S11. The yield
(%) of compounds were as follows: S1 (61), S2 (63), S3 (40), S4
(86), S5 (40), S6 (70), S7 (87), S8 (50), S9 (47), S10 (38), S11
(92). Chemical structures of known compounds were confirmed
General procedure for the synthesis of compounds
S1–S11
An appropriate ketone (9 mmol) [Acetophenone (S1),
4-methylacetophenone
4-fluoroacetophenone
4-methoxyacetophenone (S6),
(S2),
(S4),
4-chloroacetophenone
4-bromoacetophenone
4-nitroacetophenone
(S3),
(S5),
(S7),
1
by H NMR (data were not presented), and HRMS (Table 1).
1
Structures of new compounds, S9–S11, were confirmed by H
NMR, 13C NMR and HRMS.
2-acetylthiophene (S8), 2-acetylfuran (S9), 1-indanone (S10),
Table 1. The physical data of the compounds S1–S11.
Melting
point (ꢁC)
Crystallization
solvent
Melting
point (ꢁC)
Crystallization
solvent
HRMS [MH]+
Calc. Mass
HRMS [MH]+
Found Mass
Compounds
Ketone
O
S1
257–259
234–235
Ethanol
247–248y
224–226y
DMF:H2Oy
Ethanoly
290.0963
290.0958
CH
3
O
S2
Ethanol
304.1120
304.1111
CH
3
H C
3
O
S3
S4
S5
S6
237–238
210–212
241–243
215–217
DMF:H2O
Ethanol
Ethanol
Ethanol
221–222y
210–212z
210–212y
230–232z
DMF:H2Oy
Ethanolz
Ethanoly
Ethanolz
324.0574
308.0869
368.0068
320.1069
324.0562
308.0866
368.0058
320.1056
CH
3
3
3
Cl
F
O
O
CH
CH
Br
O
CH
3
H CO
3
O
S7
266–268
Ethanol
254–255y
DMF:H2Oy
335.0814
335.0813
CH
3
O N
2
O
S8
217–218
249–251
247–249
Ethanol
Ethanol
230–232z
Ethanolz
296.0527
280.0756
302.0963
296.0518
280.0753
302.0945
S
CH
3
O
S9
–
–
–
–
O
CH
3
O
S10
DMF:H2O
S11
214–216
DMF:H2O
–
–
302.0970
302.0962
O
ySynthesis of some pyrazolyl benzenesulfonamide derivatives as dual anti-inflammatory antimicrobial agents23
.
zSynthesis and biological evaluation of some 4-functionalized-pyrazoles as antimicrobial agents46
.

DOI: 10.3109/14756366.2015.1047359
Synthesis of 4-(2-substituted hydrazinyl)benzenesulfonamides
3
4-{2-[1-(Furan-2-yl)ethylidene]hydrazino}
benzenesulfonamide (S9)
visualized by SDS-PAGE (BIO-RAD Mini-PROTEANÕ system
casting stand, Shanghai, China). After this process, a single band
was observed for each isoenzyme33. This protein imaging method
was previously described34–36. In this application, the imaging
method was performed out in 10 and 3% acrylamide for the
1
Melting point 249–251 ꢁC. H NMR (DMSO-d6): d 9.68 (s, 1H,
NH), 7.64 (d, 1H, J ¼ 0.7 Hz, furyl), 7.62 (d, 2H, J ¼ 8.8 Hz,
phenyl), 7.22 (d, 2H, J ¼ 8.8 Hz, phenyl), 7.07 (s, 2H, SO2NH2),
6.76 (d, 1H, J ¼ 3.3 Hz, furyl), 6.55 (d, 1H, J ¼ 3.7 Hz, furyl), 2.17
(s, 3H, CH3); 13C NMR (DMSO-d6): d 153.2, 148.9, 144.1, 136.8,
134.1, 127.9, 112.6, 112.5, 109.5, 13.6; HRMS (ESI+) Calc. for
C12H14N3O3S [MH]+ 280.0756; found 280.0753.
running and the stacking gel, respectively, with 0.1% SDS37,38
.
Both CA isoenzymes activities were determined according to
the method given by Verpoorte et al.39 and described previ-
ously40. Briefly, the absorbance changing at 348 nm for
p-nitrophenylacetate (NPA) to p-nitrophenolate (NP) was rec-
orded over a 3-min period at the room temperature (25 ꢁC) using a
spectrophotometer (Shimadzu, UV-VIS Spectrophotometer,
UVmini-1240, Kyoto, Japan)41. The protein quantity was spec-
trophotometrically measured at 595 nm during the purification
steps according to the Bradford method42. As used in previous
studies, bovine serum albumin (BSA) was used as the standard
protein43. For determining the inhibition effect of each sulfona-
mide derivative, an activity (%)-[Sulfonamide] graph was drawn.
To determine Ki values, three different sulfonamide derivatives
concentrations were tested. In these experiments, NPA was used
as the substrate at five different concentrations and the
Lineweaver–Burk curves were drawn44 as previously described45.
4-{2-(2,3-Dihydro-1H-inden-1-ylidene)hydrazino}
benzenesulfonamide (S10)
1
Melting point 247–249 ꢁC. H NMR (DMSO-d6): d 9.58 (s, 1H,
NH), 7.66–7.63 (m, 1H, phenyl of inden-1-on), 7.62 (d, 2H,
J ¼ 8.8 Hz, phenyl), 7.31–7.27 (m, 3H), 7.25 (d, 2H, J ¼ 8.8 Hz,
phenyl), 7.06 (s, 2H, SO2NH2), 3.05 (d, 2H, J ¼ 6.6 Hz), 2.80
(d, 2H, J ¼ 6.6 Hz); 13C NMR (DMSO-d6): d 154.0, 149.3, 147.9,
139. 2, 133.5, 129.9, 127.9, 127.6, 126.3, 121.2, 112.2, 28.8, 27.5;
HRMS (ESI+) Calc. for C15H16N3O2S [MH]+ 302.0963; found
302.094.
4-{2-(1,3-Dihydro-2H-inden-2-ylidene)hydrazino}
benzenesulfonamide (S11)
Results and discussion
1
Melting point 214–216 ꢁC. H NMR (DMSO-d6/CD3OD): d 9.4
The compounds designed were synthesized successfully
(Schemes 1 and 2) and their chemical structures were confirmed
by spectral data. The physical data of the compounds were shown
in Table 1. The compounds S9–S11 are reported for the first time
by a microwave irradiation. The synthetic procedure for the
known compounds S1–S8 was different than literature proced-
ure46,47. Compounds S1–S8 were synthesized by the microwave
irradiation method in this study, while they were synthesized by a
conventional heating method in literature46,47. Microwave irradi-
ation method decreased the reaction time 2 or 3 times comparing
to conventional heating. The yields of the reactions were 40–87%
for S1–S8 in microwave irradiation, while they were synthesized
(s, 1H, NH), 7.59 (d, 2H, J ¼ 8.8 Hz, phenyl), 7.21 (bs, 2H), 7.19
(d, 2H, d, 2H, J ¼ 5.5 Hz), 7.13 (d, 2H, J ¼ 8.8 Hz, phenyl), 7.04
(s, 2H, SO2NH2), 3.84 (s, 2H), 3.77 (s, 2H); 13C NMR (CD3OD):
d 153.9, 149.7, 139.7, 139.1, 132.3, 127.6, 126.9, 126.8, 124.9,
124.5, 111.6, 38.3, 34.3; HRMS (ESI+) Calc. for C15H16N3O2S
[MH]+ 302.0970; found 302.0962.
Biochemistry
For determination of the inhibition effects of sulfonamides,
both CA isoenzyme (hCA I and II) were purified by Sepharose-
4B-L-tyrosine-sulfanilamide affinity chromatography in a single
purification step24–26. The column material including Sepharose-
4B-L-tyrosine-sulfanilamide was prepared according to a previ-
ous method27–29. Thus, homogenate solution acidity was adjusted
to 8.7 with a pH-meter using solid Tris. Subsequently, the
supernatant was transferred to the previously prepared Sepharose-
4B-L-tyrosine-sulphanilamide affinity column30–32. The proteins
flow in the column eluates was spectrophotometrically deter-
mined at 280 nm. For determination of both isoenzymes purity,
sodium dodecyl sulphate-polyacrylamide gel electrophoresis
(SDS-PAGE) for both isoenzymes was performed after a purifi-
cation step. The presence and purity of both isoenzymes were
with the yield of 82–89% in conventional heating in literature46,47
.
Up to now, the sulfonamide group (R-SO2NH2) is the most
important and largely used Zn2+-binding function for the design
of CAIs; accordingly, the majority of the clinically used CAIs are
sulfonamides48. Since the first evidence of their CA inhibitory
properties, these molecules were largely investigated by means of
kinetic, pharmacological and physiological studies14,48–50
.
The first evidence that sulfonamides could act as potent CAIs
came from a study, reported by Mann and Keilin51. Subsequently,
many aromatic sulfonamides were synthesized and investigated
for their CA inhibitory action52. Among these, benzenesulfona-
mides constitute the most common and best-characterized class.
NH2
Scheme 1. Synthesis of the compounds
S1–S9.
NH2
O
S
O
O
S
O
O
CH COONa
3
+
R
CH3
CH3
S2
EtOH
60 min,78°C
150 Watt
NH
HN
N
NH2 . HCl
R
CH3
Cl
F
Br
OCH3
NO2
S
O
R =
S1
S3
S4
S5
S6
S7
S8
S9

4
H. I. Gul et al.
J Enzyme Inhib Med Chem, Early Online: 1–6
NH
S
2
O
O
O
O
O
NH
S
NH
2
O
S
N
N
O
NH
NH
2
i
i
O
S11
S10
HN
NH . HCl
2
i= Sodium acetate, ethanol, 60 min., 78° C, 150 Watt
Scheme 2. Synthesis of the compounds S10 and S11.
Table 2. Human carbonic anhydrase isoenzymes (hCA I and II) inhibition
values with 4-(2-substituted hydrazinyl)benzenesulfonamide derivatives
(S1–S11) using esterase assay.
in obtaining novel classes of drugs devoid of various undesired
side-effects28. We report here the first study on the inhibitory
effects of 4-(2-substituted hydrazinyl)benzenesulfonamide deriva-
tives (S1–S11) against hCA I and II using esterase activity.
Low cytosolic isoenzyme hCA I is ubiquitously expressed in
the body, and can be found in high concentrations in the blood and
gastrointestinal tract57. Specifically, hCA I is found in many
tissues, however, but it was demonstrated that this isozyme is
involved in retinal and cerebral edema, and its inhibition may be a
valuable tool for fighting these conditions. Also, it was reported
that if Ki value of a tested compound was less than 50 mM
(Ki450 mM), it was considered low inhibition and this inhibitor
was accepted to be inactive against hCA I40,58. The results
obtained from this study clearly indicate that 4-(2-substituted
hydrazinyl)benzenesulfonamide derivatives (S1–S11) have effect-
ive inhibition profile against slow cytosolic isoform hCA I, and
Ki (nM)
Compounds
hCA I
hCA II
S1
S2
S3
S4
S5
S6
S7
S8
2.23 0.14
2.42 0.51
2.15 0.42
2.67 0.30
1.97 0.23
2.73 0.48
1.90 0.43
2.73 0.08
2.52 0.38
2.06 0.66
1.79 0.22
5.41 1.28
8.02 3.04
6.93 2.51
7.91 3.30
9.12 4.42
11.64 5.21
6.34 2.64
1.97 0.40
10.69 4.73
9.43 1.94
6.73 2.18
1.72 0.58
S9
S10
S11
AZA*
6.82 1.59 cytosolic dominant rapid isozymes hCA II with low nanomolar
range. These compounds bind to hCA I in the nanomolar range. Ki
*Acetazolamide (AZA) was used as a standard inhibitor for all hCA.
values are in the range of 1.79 0.22–2.73 0.08 nM for hCA I
isoenzyme. On the other hand, acetazolamide (AZA) being a
broad-specificity CA inhibitor owing to its widespread inhibition
of CAs, showed Ki value of 5.41 1.28 nM against hCA I. 4-{2-
(1,3-dihydro-2H-inden-2-ylidene)hydrazino}benzenesulfonamide
(S11), possessing 1,3-dihydro-2H-inden-2-one was the best hCA I
inhibitor (Ki: 1.79 0.22 nM). The inhibition effects of all 4-(2-
substituted hydrazinyl)benzenesulfonamide derivatives (S1–S11)
are higher than that of acetazolamide (AZA; Ki: 5.41 1.28 nM).
AZA, 5-Acetamido-1,3,4-thiadiazole-2-sulfonamide) is con-
sidered to be a good CA inhibitor and is approved for the
treatment of a range of conditions including glaucoma, epilepsy
and altitude sickness5.
CA II is involved in several diseases including glaucoma,
epilepsy, edema and probably altitude sickness. Against the
physiologically dominant isoform hCA II, 4-(2-substituted
hydrazinyl)benzenesulfonamide derivatives (S1–S11) demon-
strated Kis of 1.72 0.58–11.64 5.21 nM (Table 2). As with
CA I, the compound of S11 (4-{2-(1,3-dihydro-2H-inden-2-
ylidene)hydrazino}benzenesulfonamide) was the best hCA II
inhibitor (Ki: 1.72 0.58 nM). However, the average Ki value of
Up to now, a lot of studies have been reported on the interaction of
benzenesulfonamides with CA isoenzymes. Especially, benzene-
sulfonamides include halogens, acetamido and alkoxycarbonyl
moieties had good inhibitory properties and interesting physico-
chemical features were observed for compounds possessing
carboxy-, ureido-, hydrazido-, thioureido- and methylamino-
moieties48. It was reported that the interaction of the benzene-
sulfonamide moiety with the CA active site is rather similar, with
the sulfonamide moiety involved in the canonical coordination of
the Zn2+ catalytic ion and the phenyl ring. Also, the substituents
of the phenyl ring can establish different types of interactions,
which involve the hydrophobic, or the hydrophilic region of the
active site48.
Benzenesulfonamide derivatives have been largely investigated
both in the search of more active CAIs and to develop compounds
with a different biological activity53. So far, two different types of
such derivatives have been characterized: those where the two
sulfonamide moieties are present on the same phenyl ring10 and
those where the two sulfonamide moieties are present on two
different phenyl rings, separated by urea, carboxyamido, guan-
idine moieties, etc., as spacers53–55. These sulfonamide deriva-
tives have been largely clinically used for the treatment of
glaucoma56 and several neurological disorders53.
4-(2-substituted
hydrazinyl)benzenesulfonamide derivatives
(S1–S11) was found to be 2.28 nM for hCA I. Conversely, the
average Ki value of these compounds for hCA II was found to be
7.32 nM. These results showed that 4-(2-substituted hydrazinyl)-
benzenesulfonamide derivatives (S1–S11) have higher inhibition
affinity toward hCA I than that of hCA II isoenzyme. Also, AZA,
which may interact with the distinct hydrophobic and hydrophilic
halves of the CA II active site, showed Ki value of 6.82 1.59 nM.
Two physiologically relevant CA isoforms (hCA I and II) were
included in our study. 4-(2-Substituted hydrazinyl)benzenesulfo-
namide derivatives (S1–S11) were tested for their inhibition
properties against hCA I and II isoenzymes, showing generally an
efficient inhibition. CA I and II inhibiting effects of the 4-
Conclusion
(2-substituted
hydrazinyl)benzenesulfonamide
derivatives
(S1–S11) are presented in Table 2. It was well known that The 4-(2-substituted hydrazinyl)benzenesulfonamide derivatives
developing isoenzyme-specific CAIs should be highly beneficial (S1–S11) were evaluated for their hCA I and II isoenzymes

DOI: 10.3109/14756366.2015.1047359
Synthesis of 4-(2-substituted hydrazinyl)benzenesulfonamides
5
¨
17. Goksu S, Naderi A, Akbaba Y, et al. Carbonic anhydrase inhibitory
properties of novel benzylsulfamides using molecular modeling and
experimental studies. Bioorg Chem 2014;56:75–82.
inhibition properties. These compounds were found to be
sufficiently active. hCA I and II isoenzymes were potently
inhibited by new synthesized sulphonamide derivatives with Kis
in the range of 1.79 0.22–2.73 0.08 nM against hCA I and in
the range of 1.72 0.58–11.64 5.21 nM against hCA II,
respectively.
18. Sethi KK, Verma SM, Tanc¸ M, et al. Carbonic anhydrase inhibitors:
synthesis and inhibition of the cytosolic mammalian carbonic
anhydrase isoforms I, II and VII with benzene sulfonamides
incorporating 4,5,6,7-tetrachlorophthalimide moiety. Bioorg Med
Chem 2013;21:5168–74.
19. Oztu¨rk Sarıkaya SB, Gu¨lc¸in I, Supuran CT. Carbonic anhydrase
inhibitors. Inhibition of human erythrocyte isozymes I and II with a
series of phenolic acids. Chem Biol Drug Des 2010;75:515–20.
¨
_
Declaration of interest
The authors report no conflict of interest and are responsible for the
contents and writing of the paper.
_
20. Topal M, Gu¨lc¸in I. Rosmarinic acid: a potent carbonic anhydrase
isoenzymes inhibitor. Turk J Chem 2014;38:894–902.
¨
21. Innocenti A, Oztu¨rk Sarıkaya SB, et al. Carbonic anhydrase
The authors Gul and Gulcin thank the Ataturk University Research
Fund (Project number 2012/75 and 2012/74) and Research Chairs
Program at King Saud University for their financial supports. Also,
Gulcin would like to extend his sincere appreciation to the Research
Chairs Program at King Saud University for funding this research.
inhibitors. Inhibition of mammalian isoforms I-XIV with a series
of natural product polyphenols and phenolic acids. Bioorg Med
Chem 2010;18:2159–64.
_
¨
¨
22. C¸ etinkaya Y, Goc¸er H, Gulc¸in I, Menzek A. Synthesis and carbonic
anhydrase isoenzymes inhibitory effects of brominated diphenyl-
methanone and its derivatives. Arch Pharm 2014;347:354–9.
23. Akıncıoglu A, Topal M, Gu¨lc¸in I, Goksu S. Novel sulfamides and
sulfonamides incorporating tetralin scaffold as carbonic anhydrase
and acetylcholine esterase inhibitors. Arch Pharm 2014;347:68–76.
24. Gu¨lc¸in I, Beydemir S¸, Bu¨yu¨kokuroglu ME. In vitro and in vivo
effects of dantrolene on carbonic anhydrase enzyme activities. Biol
Pharm Bull 2004;27:613–16.
25. Beydemir S¸ , Gu¨lc¸in I. Effect of melatonin on carbonic anhydrase
from human erythrocyte in vitro and from rat erythrocyte in vivo.
J Enzyme Inhib Med Chem 2004;19:193–7.
References
_
˘
¨
1. ArasHisar S¸ , Hisar O, Beydemir S¸ , et al. Effect of vitamin E on
carbonic anhydrase enzyme activity in rainbow trout (Oncorhynchus
mykiss) erythrocytes in vitro and in vivo. Acta Vet Hung 2004;52:
413–22.
_
˘
_
2. Hisar O, Beydemir S¸ , Gu¨lc¸in I, et al. Effect of low molecular weight
_
plasma inhibitors of rainbow trout (Oncorhyncytes mykiss) on
human erythrocytes carbonic anhydrase-II isozyme activity in vitro
and rat erythrocytes in vivo. J Enzyme Inhib Med Chem 2005;20:
35–9.
_
26. C¸ oban TA, Beydemir S¸ , Gu¨lc¸in I, Ekinci D. Morphine inhibits
erythrocyte carbonic anhydrase in vitro and in vivo. Biol Pharm Bull
2007;30:2257–61.
27. Aksu K, Nar M, Tanc¸ M, et al. The synthesis of sulfamide analogues
of dopamine related compounds and their carbonic anhydrase
inhibitory properties. Bioorg Med Chem 2013;21:2925–31.
28. Atasaver A, Ozdemir H, Gu¨lc¸in I, Ku¨frevioglu OI. One-step
purification of lactoperoxidase from bovine milk by affinity
chromatography. Food Chem 2013;136:864–70.
29. Akbaba Y, Bastem E, Topal F, et al. Synthesis and carbonic
anhydrase inhibitory effects of novel sulfamides derived from
1-aminoindanes and anilines. Arch Pharm 2014;347:950–7.
_
3. Hisar O, Beydemir S¸, Gu¨lc¸in I, et al. The effect of melatonin
hormone on carbonic anhydrase enzyme activity in rainbow trout
(Oncorhynchus mykiss) erythrocytes in vitro and in vivo. Turk J Vet
Anim Sci 2005;29:841–5.
4. Ward C, Langdon SP, Mullen P, et al. New strategies for targeting
the hypoxic tumour microenvironment in breast cancer. Cancer
Treat Rev 2013;39:171–9.
5. Tanpure RP, Ren B, Peat TS, et al. Carbonic anhydrase inhibitors
with dual-tail moieties to match the hydrophobic and hydrophilic
halves of the carbonic anhydrase active site. J Med Chem 2015;58:
1494–501.
6. Neri D, Supuran CT. Interfering with pH regulation in tumours as a
therapeutic strategy. Nat Rev Drug Discov 2011;10:767–77.
_
7. C¸ oban TA, Beydemir S¸, Gu¨lc¸in I, Ekinci D. The inhibitory effect of
ethanol on carbonic anhydrase isoenzymes: in vivo and in vitro
studies. J Enzyme Inhib Med Chem 2008;23:266–70.
8. Boztas¸ M, C¸ etinkaya Y, Topal M, et al. Synthesis and carbonic
anhydrase isoenzymes I, II, IX, and XII inhibitory effects of
dimethoxy-bromophenol derivatives incorporating cyclopropane
moieties. J Med Chem 2015;58:640–50.
9. Yıldırım A, Atmaca U, Keskin A, et al. N-Acylsulfonamides
strongly inhibit human carbonic anhydrase isoenzymes I and II.
Bioorg Med Chem 2015;23:2598–605.
10. Alterio V, De Simone G, Monti SM, et al. Carbonic anhydrase
inhibitors: inhibition of human, bacterial, and archaeal isozymes
with benzene-1,3-disulfonamides-solution and crystallographic stu-
dies. Bioorg Med Chem Lett 2007;17:4201–7.
¨
_
¨ _
˘
_
30. S¸ entu¨rk M, Gu¨lc¸in I, Beydemir S¸ , et al. In vitro inhibition of human
carbonic anhydrase I and II isozymes with natural phenolic
compounds. Chem Biol Drug Des 2011;77:494–9.
¨
31. Oztu¨rk Sarıkaya SB, Topal F, S¸ entu¨rk M, et al. In vitro inhibition of
a-carbonic anhydrase isozymes by some phenolic compounds.
Bioorg Med Chem Lett 2011;21:4259–62.
32. Nar M, C¸ etinkaya Y, Gu¨lc¸in I, Menzek A. (3,4-
Dihydroxyphenyl)(2,3,4-trihydroxyphenyl)methanone and its
derivatives as carbonic anhydrase isoenzymes inhibitors. J Enzyme
Inhib Med Chem 2013;28:402–6.
33. Laemmli DK. Cleavage of structural proteins during the assembly of
_
the head of bacteriophage T4. Nature 1970;227:680–5.
_
¨ _
˘
34. Gu¨lc¸in I, Ku¨frevioglu OI, Oktay M. Purification and characteriza-
tion of polyphenol oxidase from nettle (Urtica dioica L.) and inhib-
ition effects of some chemicals on the enzyme activity. J Enzyme
Inhib Med Chem 2005;20:297–302.
˘
35. S¸ is¸ecioglu M, C¸ ankaya M, Gu¨lc¸in I, Ozdemir M. Interactions of
melatonin and serotonin to lactoperoxidase enzyme. J Enzyme Inhib
_
¨
_
11. S¸ entu¨rk M, Gu¨lc¸in I, Das¸tan A, et al. Carbonic anhydrase inhibitors.
Inhibition of human erythrocyte isozymes I and II with a series of
antioxidant phenols. Bioorg Med Chem 2009;17:3207–11.
Med Chem 2010;25:779–83.
_
36. S¸ is¸ecioglu M, Gu¨lc¸in I, C¸ ankaya M, et al. Purification and
_
12. Arabaci B, Gu¨lc¸in I, Alwasel S. Capsaicin: a potent inhibitor of
˘
characterization of peroxidase from Turkish black radish
(Raphanus sativus L.). J Med Plants Res 2010;4:1187–96.
carbonic anhydrase isoenzymes. Molecules 2014;19:10103–14.
13. Gu¨ney M, Cos¸kun A, Topal F, et al. Oxidation of cyanobenzocy-
cloheptatrienes: synthesis, photooxygenation reaction and carbonic
anhydrase isoenzymes inhibition properties of some new benzo-
tropone derivatives. Bioorg Med Chem 2014;22:3537–43.
14. Supuran CT. Carbonic anhydrases: novel therapeutic applications for
inhibitors and activators. Nat Rev Drug Discov 2008;7:168–81.
_
15. Coban TA, Beydemir S, Gu¨cin I, et al. Sildenafil is a strong activator
_
37. Gu¨lc¸in I, Beydemir S¸, C¸ oban TA, Ekinci D. The inhibitory effect
of dantrolene sodium and propofol on 6-phosphogluconate
dehydrogenase from rat erythrocyte. Fresen Environ Bull 2008;17:
1283–7.
_
¨ _
˘
38. S¸ entu¨rk M, Gu¨lc¸in I, C¸ iftci M, Ku¨frevioglu OI. Dantrolene inhibits
human erythrocyte glutathione reductase. Biol Pharm Bull 2008;31:
2036–9.
39. Verpoorte JA, Mehta S, Edsall JT. Esterase activities of human
carbonic anhydrases B and C. J Biol Chem 1967;242:4221–9.
40. Akıncıoglu A, Akbaba Y, Goc¸er H, et al. Novel sulfamides as
potential carbonic anhydrase isoenzymes inhibitors. Bioorg Med
Chem 2013;21:379–85.
of mammalian carbonic anhydrase isoforms I-XIV. Bioorg Med
Chem 2009;17:5791–5.
˘
¨
16. Akbaba Y, Akıncıoglu A, Goc¸er H, et al. Carbonic anhydrase
inhibitory properties of novel sulfonamide derivatives of aminoin-
danes and aminotetralins. J Enzyme Inhib Med Chem 2014;29:
35–42.
˘
¨

6
H. I. Gul et al.
J Enzyme Inhib Med Chem, Early Online: 1–6
¨
41. Goc¸er H, Akıncıoglu A, Oztas¸kın N, et al. Synthesis, antioxidant 51. Mann T, Keilin D. Sulphanilamide as a specific carbonic anhydrase
˘
¨
and antiacetylcholinesterase activities of sulfonamide derivatives of
dopamine related compounds. Arc Pharm 2013;346:783–92.
42. Bradford MM. Rapid and sensitive method for the quantitation of
microgram quantities of protein utilizing the principle of protein-dye
binding. Anal Biochem 1976;72:248–54.
inhibitor. Nature 1940;146:164–5.
52. Supuran CT, Scozzafava A, Casini A. Development of sulfonamide
carbonic anhydrase inhibitors (CAIs). In: Supuran CT, Scozzafava
A, Conway J, eds. Carbonic anhydrase: its inhibitors and activators.
Boca Raton (FL): CRC Press; 2004:67–147.
_
¨
¨
43. Koksal E, Gulc¸in I. Purification and characterization of peroxidase 53. Alterio V, Di Fiore A, D’Ambrosio K, et al. X-ray crystallography of
from cauliflower (Brassica oleracea L.) buds. Protein Peptide Lett
2008;15:320–6.
44. Lineweaver H, Burk D. The determination of enzyme dissociation
constants. J Am Chem Soc 1934;56:658–66.
CA inhibitors and its importance in drug design. In: Supuran CT,
Winum JY, eds. Drug design of zinc-enzyme inhibitors: functional,
structural, and disease applications. Hoboken (NJ): Wiley; 2009:
73–138.
_
˘
¨
¨
¨
45. Koksal E, Aggul AG, Bursal E, Gulc¸in I. Purification and 54. Casini A, Abbate F, Scozzafava A, Supuran CT. Carbonic anhydrase
characterization of peroxidase from sweet gourd (Cucurbita
Moschata Lam. Poiret). Int J Food Propert 2012;15:1110–19.
46. Bekhit AA, Ashour HMA, Abdel-Rahman HM, et al. Synthesis of
inhibitors: X-ray crystallographic structure of the adduct of human
isozyme II with a bis-sulfonamide-two heads are better than one?
Bioorg Med Chem Lett 2003;13:2759–63.
some pyrazolyl benzenesulfonamide derivatives as dual anti-inflam- 55. Abbate F, Casini A, Owa T, et al. Carbonic anhydrase inhibitors:
matory antimicrobial agents. J Enzyme Inhib Med Chem 2009;24:
296–309.
47. Sharma PK, Chandak N, Kumar P, et al. Synthesis and biological
E7070, a sulfonamide anticancer agent, potently inhibits cytosolic
isozymes I and II, and transmembrane, tumor-associated isozyme
IX. Bioorg Med Chem Lett 2004;14:217–23.
evaluation of some 4-functionalized-pyrazoles as antimicrobial 56. Pastorekova S, Parkkila S, Pastorek J, Supuran CT.
agents. Eur J Med Chem 2011;46:1425–32.
Carbonic anhydrases: current state of the art, therapeutic applica-
tions and future prospects. J Enzyme Inhib Med Chem 2004;19:
199–229.
48. Alterio V, Di Fiore A, D’Ambrosio K, et al. Multiple binding modes
of inhibitors to carbonic anhydrases: how to design specific drugs
targeting 15 different isoforms? Chem Rev 2012;112:4421–68.
49. Clare BW, Supuran CT. A perspective on quantitative structure–
activity relationships and carbonic anhydrase inhibitors. Expert Opin
Drug Metab Toxicol 2006;2:113–37.
57. D’Ascenzio M, Carradori S, De Monte C, et al. Design, synthesis
and evaluation of N-substituted saccharin derivatives as selective
inhibitors of tumor-associated carbonic anhydrase XII. Bioorg Med
Chem 2014;22:1821–31.
50. Thiry A, Dogne JM, Supuran CT, Masereel B. Anticonvulsant 58. Vullo D, Scozzafava A, Pastorekova S, et al. Carbonic anhydrase
sulfonamides/sulfamates/sulfamides with carbonic anhydrase inhibi-
tory activity: drug design and mechanism of action. Curr Pharm Des
2008;14:661–71.
inhibitors: inhibition of the tumor-associated isozyme IX with
fluorine-containing sulfonamides. The first subnanomolar CA IX
inhibitor discovered. Bioorg Med Chem Lett 2004;14:2351–6.