Synthesis and molecular docking studies of some 4-phthalimidobenzenesulfonamide derivatives as acetylcholinesterase and butyrylcholinesterase inhibitors

Article abstract of DOI:10.1080/14756366.2016.1226298

A series of 4-phthalimidobenzenesulfonamide derivatives were designed, synthesized and evaluated for the inhibitory activities against acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). Structures of the title compounds were confirmed by spectral and elemental analyses. The cholinesterase (ChE) inhibitory activity studies were carried out using Ellman’s colorimetric method. The biological activity results revealed that all of the title compounds (except for compound 8) displayed high selectivity against AChE. Among the tested compounds, compound 7 was found to be the most potent against AChE (IC₅₀= 1.35 ± 0.08 μM), while compound 3 exhibited the highest inhibition against BuChE (IC₅₀= 13.41 ± 0.62 μM). Molecular docking studies of the most active compound 7 in AChE showed that this compound can interact with both the catalytic active site (CAS) and the peripheral anionic site (PAS) of AChE.

Full text of DOI:10.1080/14756366.2016.1226298

Journal of Enzyme Inhibition and Medicinal Chemistry
ISSN: 1475-6366 (Print) 1475-6374 (Online) Journal homepage: http://www.tandfonline.com/loi/ienz20
Synthesis and molecular docking studies of some
4-phthalimidobenzenesulfonamide derivatives as
acetylcholinesterase and butyrylcholinesterase
inhibitors
Zeynep Soyer, Sirin Uysal, Sulunay Parlar, Ayse Hande Tarikogullari Dogan &
Vildan Alptuzun
To cite this article: Zeynep Soyer, Sirin Uysal, Sulunay Parlar, Ayse Hande Tarikogullari
Dogan & Vildan Alptuzun (2016): Synthesis and molecular docking studies of
some 4-phthalimidobenzenesulfonamide derivatives as acetylcholinesterase and
butyrylcholinesterase inhibitors, Journal of Enzyme Inhibition and Medicinal Chemistry, DOI:
10.1080/14756366.2016.1226298
To link to this article: http://dx.doi.org/10.1080/14756366.2016.1226298
Published online: 21 Oct 2016.
Submit your article to this journal
Article views: 2
View related articles
View Crossmark data
Full Terms & Conditions of access and use can be found at
http://www.tandfonline.com/action/journalInformation?journalCode=ienz20
Download by: [Cornell University Library]
Date: 23 October 2016, At: 23:49

JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY, 2016
http://dx.doi.org/10.1080/14756366.2016.1226298
ORIGINAL ARTICLE
Synthesis and molecular docking studies of some
4-phthalimidobenzenesulfonamide derivatives as acetylcholinesterase
and butyrylcholinesterase inhibitors
Zeynep Soyer, Sirin Uysal, Sulunay Parlar, Ayse Hande Tarikogullari Dogan and Vildan Alptuzun
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Ege University, Bornova, Izmir, Turkey
ABSTRACT
ARTICLE HISTORY
Received 1 July 2016
Revised 16 August 2016
Accepted 16 August 2016
Published online 7 Septem-
ber 2016
A series of 4-phthalimidobenzenesulfonamide derivatives were designed, synthesized and evaluated for
the inhibitory activities against acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). Structures of
the title compounds were confirmed by spectral and elemental analyses. The cholinesterase (ChE) inhibi-
tory activity studies were carried out using Ellman’s colorimetric method. The biological activity results
revealed that all of the title compounds (except for compound 8) displayed high selectivity against
AChE. Among the tested compounds, compound 7 was found to be the most potent against
KEYWORDS
AChE (IC50¼ 1.35 0.08 lM), while compound
3 exhibited the highest inhibition against BuChE
Acetylcholinesterase
inhibitor;butyrylcholinestera-
se inhibitor; molecular
docking; phthalimide;
sulfonamide
(IC50¼ 13.41 0.62 lM). Molecular docking studies of the most active compound 7 in AChE showed that
this compound can interact with both the catalytic active site (CAS) and the peripheral anionic site (PAS)
of AChE.
Introduction
inhibitors were designed based on this pharmacophore23–29
.
Meanwhile, sulfonamide derivatives are another important class of
pharmacophores in medicinal chemistry effective in a number of
different therapeutic areas. They act as antibacterials, diuretics, car-
bonic anhydrase inhibitors, anticonvulsants, anti-inflammatory,
Alzheimer’s disease (AD), characterized by memory loss and other
cognitive impairments, is currently one of the most difficult pro-
gressive neurodegenerative disorders to treat1–4. In the past deca-
des, various pathogenesis hypothesis of AD have been proposed,
such as cholinergic hypothesis, amyloid cascade hypothesis, oxida-
tive stress hypothesis and tau protein hypothesis. Among them,
cholinergic hypothesis was a widely accepted theory, which sug-
gests that the low level of acetylcholine in specific regions of the
brain is the major cause leading to learning and memory dysfunc-
tions. Based on the cholinergic hypothesis, one possible approach
to treat AD is to restore the level of acetylcholine by using revers-
ible inhibitors to inhibit cholinesterases that include acetylcholin-
esterase (AChE) and butyrlcholinesterase (BuChE)3–10. Currently,
four AChE inhibitors have been approved by European and US
agency: tacrine, donepezil, galantamine and rivastigmine
(Figure 1). These agents are important for the palliative treatment
of AD, but their clinically efficacy is limited, mainly due to their
poor selectivity, bioavailability and adverse side effects on periph-
eral nervous system and liver. Thus, the search for new ChE inhibi-
anticancers, antihypertensives and AChE inhibitors30–34
.
In this study, we designed a series of 4-phthalimidobenzenesul-
fonamide derivatives as potent cholinesterase inhibitors, in which
two pharmacophores (phthalimide and sulfonamide) were com-
bined. Accordingly, the synthesis, AChE and BuChE inhibitory activ-
ities and molecular docking studies of designed compounds are
reported.
Materials and methods
Chemistry
All chemicals, reagents and solvents were high-grade commercial
products and used without further purification. Reactions were
checked by thin-layer chromatography (TLC) on precoated silica
gel aluminum plates (Kieselgel 60, F254, E. Merck, Germany); spots
were visualized by UV at 254 nm. Melting points were determined
using a Stuart SMP30 (Staffordshire, ST15 OSA, United Kingdom)
melting point apparatus and are not corrected. IR spectra of the
compounds were recorded on a Perkin Elmer 100 Fourier trans-
form FT-IR (ATR) spectrophotometer (Perkin Elmer Inc., MA). 1H
NMR spectra were recorded on a Varian AS 400 Mercury Plus NMR
spectrometer (Varian, Palo Alto, CA) at 400 MHz using DMSO-d6
and Aceton-d6 as solvent. Chemical shifts were given in ppm (d)
with TMS as an internal standard. J values were given in Hertz.
tors is still of great interest3,4,11–19
.
The crystal structure of AChE in complex with inhibitors
revealed that there are two binding sites, a peripheral anionic site
(PAS) and catalytic active site (CAS). According to the structure
characteristics of AChE, several new compounds were synthesized
as anti-AD agents3,11,12,14,20–22
.
Phthalimide derivatives are important compounds due to
their various bioactivities such as anticancer, anti-inflammatory,
anticonvulsant and AChE inhibitory activity. Literature survey
revealed that phthalimide structure had been proved to Abbreviations for 1H NMR data quoted are as follows: s (singlet);
interact with the active site of AChE and several novel AChE d, (doublet); t, (triplet); q, (quartet); m, (multiplet); bs, (broad
_
CONTACT Zeynep Soyer
zeynep.soyer@ege.edu.tr
Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Ege University, 35100 Bornova, Izmir, Turkey
ß 2016 Informa UK Limited, trading as Taylor & Francis Group

2
Z. SOYER ET AL.
4-(1,3-Dioxoisoindolin-2-yl)-N-(2-methoxyphenyl)
O
1
benzenesulfonamide (2) Yield 49%. mp 205 ꢀC. H NMR (DMSO-d6):
O
NH2
d 9.57 (1H, bs, NH), 7.98–7.94 (2H, m, phthalimide-H), 7.92–7.89
(2H, m, phthalimide-H), 7.80 (2H, d, J ¼ 9.2 Hz, benzene-H), 7.61
(2H, d, J ¼ 7.6 Hz, benzene-H), 7.24 (1H, d, J ¼ 6.8 Hz, anilide-H),
7.13 (1H, t, J ¼ 7.8 Hz, anilide-H), 6.90 (2H, d, J ¼ 7.6 Hz, anilide-H),
3.48 (3H, s, OCH3) ppm. IR (tmaks cmꢁ1) (FT/ATR): 3465, 3298, 3100,
3011, 2963, 2837, 1786, 1707, 1337, 1166. Anal. calcd. for
C21H6N2O5S: C, 61.76; H, 3.95; N, 6.86; S, 7.85. Found C,
61.51; H, 4.15; N, 7.21; S, 7.86. MS (APCI) m/z (%): 288 (100), 409
(M þ Hþ, 75).
O
N
N
Donepezil
Tacrine
OH
O
O
O
N
CH3
O
H3C
CH3
4-(1,3-Dioxoisoindolin-2-yl)-N-(2-isopropylphenyl)
1
benzenesulfonamide (3) Yield 66%. mp 186 ꢀC. H NMR (DMSO-d6):
H3C
N
N
d 9.76 (1H, bs, NH), 7.98–7.93 (2H, m, phthalimide-H), 7.92–7.89
(2H, m, phthalimide-H), 7.79 (2H, d, J ¼ 7.2 Hz, benzene-H), 7.65
(2H, d, J ¼ 6.4 Hz, benzene-H), 7.26 (1H, d, J ¼ 8.4 Hz, anilide-H),
7.19 (1H, t, J ¼ 7.8 Hz, anilide-H), 7.07 (1H, t, J ¼ 7.2 Hz, anilide-H),
6.94 (1H, d, J ¼ 8.0 Hz, anilide-H), 3.15–3.10 (1H, m, CH), 0.92 (6H,
d, J ¼ 6.8 Hz, 2xCH3) ppm. IR (tmaks cmꢁ1) (FT/ATR): 3474, 3249,
3201, 2978, 2966, 2929, 2868, 2821, 2749, 1787, 1721, 1335, 1162.
Anal. calcd. for C23H20N2O4S: C, 65.70; H, 4.79; N, 6.66; S, 7.63.
Found C,65.53; H, 4.99; N, 7.07; S,7.84. MS (APCI) m/z (%): 421
(M þ Hþ, 100).
CH3
Galantamine
Rivastigmine
Figure 1. Chemical structures of FDA approved AChE inhibitors.
singlet). Mass spectra (APCI-MS) were measured on a Thermo
MSQ Plus LC/MS (Thermoscientific Inc., San Jose, CA). Elemental
analyses (C, H, N and S) were performed by Leco TruSpec Micro
(Leco, St. Joseph, MI). The analytical results for the elements were
within 0.4% of the theoretical values.
4-(1,3-Dioxoisoindolin-2-yl)-N-(p-tolyl)benzenesulfonamide (4) Yield
52%. mp 212 ꢀC; 224–226 ꢀC38. H NMR (DMSO-d6): d 10.22 (1H, bs,
1
General procedure for the synthesis of N-phenylphthalimide (1a)
Phthalic anhydride (3.38 mmol) and aniline (4.06 mmol) were
heated at 160 ꢀC in the sand bath, until completely melted. After
cooling, the residue was crystallized from water35. Yield is 45%.
Mp 204–205 ꢀC.
NH), 7.98–7.94 (2H, m, phthalimide-H), 7.92–7.89 (2H,
m, phthalimide-H), 7.86 (2H, d, J ¼ 8.0 Hz, benzene-H), 7.62 (2H, d,
J ¼ 8.4 Hz, benzene-H), 7.03 (2H, d, J ¼ 8.8 Hz, anilide-H), 6.99 (2H,
d, J ¼ 8.4 Hz, anilide-H), 2.17 (3H, s, CH3) ppm. IR (tmaks cmꢁ1
)
(FT/ATR): 3282, 1786, 1713, 1594, 1509, 1340, 1161. Anal. calc. for
C21H16N2O4S. 0.3 C2H6O: C, 63.86; H, 4.42; N, 6.98; S, 7.89. Found C,
64.17; H, 4.64; N, 7.28; S, 7.72. MS (APCI) m/z (%): 379 (100), 393
(M þ Hþ, 11).
General procedure for the synthesis of 4-phthalimidobenzenesul-
fonyl chloride (1b)
To a solution of chlorosulfonic acid (1.35 mmol) and phosphorus
pentachloride (0.67 mmol) that had been stirred for 10 min, com-
pound 1a (0.67 mmol) was added in small portions. After the mix-
ture was stirred and heated at 50 ꢀC for 30 min, it was poured into
ice water and extracted with chloroform. The organic phase was
separated, dried over anhydrous sodium sulfate and evaporated at
reduced pressure to furnish compound 1b36. Yield is 86%. Mp
256 ꢀC.
4-(1,3-Dioxoisoindolin-2-yl)-N-(4-methoxyphenyl)
benzenesulfonamide (5) Yield 48%. mp 161 ꢀC; 168–170 ꢀC38. 1H
NMR (DMSO-d6): d 10.04 (1H, bs, NH), 7.97–7.94 (2H, m, phthali-
mide-H), 7.92–7.89 (2H, m, phthalimide-H), 7.82 (2H, d, J ¼ 8.4 Hz,
benzene-H), 7.63 (2H, d, J ¼ 9.2 Hz, benzene-H), 7.01 (2H, d,
J ¼ 8.8 Hz, anilide-H), 6.81 (2H, d, J ¼ 8.8 Hz, anilide-H), 3.65 (3H, s,
OCH3) ppm. IR (tmaks cmꢁ 1) (FT/ATR): 3243, 1790, 1713, 1592,
1508, 1334, 1172. Anal. calcd. for C21H16N2O5S. 0.01 C2H6O: C,
61.74; H, 3.96; N, 6.85; S, 7.84. Found C, 62.15; H, 4.39; N, 7.10; S,
7.70. MS (APCI) m/z (%): 288 (100), 409 (M þ Hþ, 38).
General procedure for the synthesis of final compounds (1–11)
4-Phthalimidobenzenesulfonyl chloride (1.1 mmol) and appropriate
amines (2.2 mmol) were refluxed in acetone (25 mL). After comple- 4-(1,3-Dioxoisoindolin-2-yl)-N-(4-chlorophenyl)benzenesulfonamide
(6) Yield 59%. mp 212 ꢀC; 212–214 ꢀC38. 1H NMR (DMSO-d6): d
tion of reaction (monitored by TLC), reaction mixture was poured
into ice water, then obtained precipitate was filtered and recrystal-
10.56 (1H, bs, NH), 7.98–7.94 (2H, m, phthalimide-H), 7.91–7.89 (2H,
m, phthalimide), 7.89 (2H, d, J ¼ 8.4 Hz, benzene-H), 7.66 (2H, d,
J ¼ 8.8 Hz, anilide-H), 7.30 (2H, d, J ¼ 8.4 Hz, benzene-H), 7.13 (2H,
d, J ¼ 8.6 Hz, anilide-H) ppm. IR (tmaks cmꢁ 1) (FT/ATR): 3252, 1784,
1714, 1592, 1334, 1164, 712. Anal. calcd. for C20H13ClN2O4S. 0.9
C3H6O: C, 58.57; H, 3.95; N, 6.02; S, 6.88. Found C, 58.17; H, 3.55; N,
5.56; S, 6.88. MS (APCI) m/z (%): 222 (100), 413 (M þ Hþ, 13), 415
(M þ H þ 2þ, 4).
lized from ethanol37.
4-(1,3-Dioxoisoindolin-2-yl)-N-(o-tolyl)benzenesulfonamide (1) Yield
66%. mp 235 ꢀC; 391–393 ꢀC38. 1H NMR (DMSO-d6): d 9.68 (1H, bs,
NH), 7.99–7.92 (2H, m, phthalimide-H), 7.92–7.90 (2H, m, phthali-
mide-H), 7.80 (2H, d, J ¼ 8.8 Hz, benzene-H), 7.65 (2H, d, J ¼ 8.8 Hz,
benzene-H), 7.14–7.11 (1H, m, anilide-H), 7.10 (2H, t, J ¼ 4.4 Hz, ani-
lide-H), 7.01–6.98 (1H, m, anilide-H), 2.04 (3H, s, CH3) ppm. IR (tmaks
cmꢁ1) (FT/ATR): 3485, 3312, 3085, 2797, 1785, 1717, 1592, 1501,
4-(1,3-Dioxoisoindolin-2-yl)-N,N-diethylbenzenesulfonamide
(7)
1327, 1161. Anal. calcd. for C21H16N2O4S: C, 64.27; H, 4.11; N, 7.14; Yield 68%. mp 181 ꢀC; 178–180 ꢀC38. 1H NMR (DMSO-d6):
d
S, 8.17. Found C, 64.26; H, 4.28; N, 7.56; S, 8.04. MS (APCI) m/z (%): 7.99–7.97 (2H, m, phthalimide-H), 7.92–7.90 (2H, m, phthalimide-H),
393 (M þ Hþ, 100).
7.94 (2H, d, J ¼ 8.4 Hz, benzene-H), 7.69 (2H, d, J ¼ 8.4 Hz,

JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY
3
Table 1. AChE and BuChE inhibitory activities and log P values of
the title compounds.
benzene-H), 3.19 (4H, q, J ¼ 7.2 Hz, 2xCH2), 1.06 (6H, t, J ¼ 7.0 Hz,
2xCH3) ppm. IR (tmaks cmꢁ 1) (FT/ATR): 2976, 1782, 1711, 1593,
1349, 1290, 1180. Anal. calcd. for C18H18N2O4S: C, 60.32; H, 5.06; N,
7.82; S, 8.95. Found C, 60.40; H, 5.19; N, 7.53; S, 8.76. MS (APCI)
m/z (%): 359 (M þ Hþ, 100).
ꢂ
IC50 SEM (lM)
Compound No
AChE
BuChE
log P†
1
2
3
4
5
6
7
8
6.84 0.17
6.79 0.41
8.19 0.13
8.61 0.18
8.52 0.18
8.21 0.14
1.35 0.08
>100
74.36 2.34
44.24 13.38
13.41 0.62
85.01 13.67
87.16 13.18
78.15 5.39
>100
>100
>100
75.18 23.82
56.67 3.26
3.60
3.30
4.41
3.60
3.30
3.94
2.52
2.27
2.66
3.05
1.51
2-(4-(Pyrrolidin-1-ylsulfonyl)phenyl)isoindoline-1,3-dione (8) Yield
1
45%. mp 175 ꢀC. H NMR (Acetone-d6): d 8.02–8.00 (2H, m, phthali-
mide-H), 7.98 (2H, d, J ¼ 8.4 Hz, benzene-H), 7.96–7.95 (2H, m,
phthalimide-H), 7.82 (2H, d, J ¼ 8.4 Hz, benzene-H), 3.30–3.26 (4H,
m, pyrolidine-H), 1.81–1.77 (4H, m, pyrolidine-H) ppm. IR (tmaks
cmꢁ 1) (FT/ATR): 2979, 2879, 1792, 1716, 1592, 1341, 1251, 1161.
Anal. calcd. for C18H16N2O4S . H2O: C, 57.74; H, 4.85; N, 7.48; S,
8.56. Found C, 58.11; H, 4.60; N, 7.08; S, 8.91. MS (APCI) m/z (%):
357 (M þ Hþ, 100).
9
10.39 0.61
8.05 0.54
7.95 0.22
10
11
ꢂ
Data are means standard error of the main of triplicate inde-
pendent experiments.
†Log P values calculated using MOE 2011.10.
2-(4-(Piperidin-1-ylsulfonyl)phenyl)isoindoline-1,3-dione (9) Yield
85%. mp 176 ꢀC; 165–167 ꢀC38. 1H NMR (Acetone-d6): d 8.01–7.95
(4H, m, phthalimide-H), 7.93 (2H, d, J ¼ 8.4 Hz, benzene-H), 7.83
(2H, d, J ¼ 8.4 Hz, benzene-H), 3.06–3.03 (4H, m, piperidine-H),
1.68–1.62 (4H, m, piperidine-H), 1.50–1.47 (2H, m, piperidine-H)
ppm. IR (tmaks cmꢁ 1) (FT/ATR): 3281, 2973, 2933, 1781, 1712, 1672,
1575, 1354, 1237, 1177. Anal. calcd. for C19H18N2O4S. 0.25H2O: C,
60.87; H, 4.97; N, 7.47; S, 8.55. Found C, 60.53; H, 4.68; N, 7.37; S,
8.60. MS (APCI) m/z (%): 371 (M þ Hþ, 100).
method of Ellman et al., with galantamine as the reference com-
pound41–45. Prior to use, all solutions were adjusted to 20 ꢀC.
Enzyme solution (100 lL) and inhibitor solution (100 lL) were
added into a cuvette containing the phosphate buffer (3.0 mL,
0.1 M; pH 8.0). After 5-min incubation, required aliquots of the
DTNB solution (100 lL) and of the AChI/BChI (20 lL) were added.
After rapid and immediate mixing, the absorption was measured
at 412 nm by UV spectroscopy. As a reference, an identical solu-
tion of the enzyme without the inhibitor is processed following
the same protocol. The blank reading contained 3.0 mL buffer,
200 lL water, 100 lL DTNB and 20 lL substrate. The enzyme activ-
ity was determined in the presence of at least five different con-
centrations of an inhibitor. Each concentration was assayed in
triplicate. The samples were investigated immediately after prepar-
ation. The AChE/BuChE inhibitory activities of the title compounds
are summarized in Table 1.
2-(4-((2-Methylpiperidin-1-yl)sulfonyl)phenyl)isoindoline-1,3-dione
(10) Yield 18%. mp 182 ꢀC; 182–184 ꢀC39. 1H NMR (DMSO-d6): d
7.99–7.97 (2H, m, phthalimide-H), 7.94 (2H, d, J ¼ 8.8 Hz, benzene-
H), 7.92–7.90 (2H, m, phthalimide-H), 7.70 (2H, d, J ¼ 8.0 Hz,
benzene-H), 4.18–4.12 (1H, m, piperidine-H), 3.64–3.61 (1H, m,
piperidine-H), 3.03–2.96 (1H, m, piperidine-H), 1.56–1.43 (5H, m,
piperidine-H), 1.26–1.17 (1H, m, piperidine-H), 1.02 (3H, d,
J ¼ 6.4 Hz, CH3) ppm. IR (tmaks cmꢁ 1) (FT/ATR): 2933, 2872, 1788,
1719, 1592, 1335, 1263, 1163. Anal. calcd. for C20H20N2O4S. H2O: C,
59.69; H, 5.51; N, 6.96; S, 7.97. Found C, 59.27; H, 5.16; N, 6.88; S,
8.31. MS (APCI) m/z (%): 385 (M þ Hþ, 100).
Molecular docking study
The crystal structures of donepezil in complex with AChE (PDB
code 1EVE resolved at 2.5 Å) were taken from the Protein Data
Bank. Heteroatoms and water molecules in the PDB file were
removed and hydrogen atoms were added to the protein by using
MOE 2014.09.146. Prior to the docking calculations, an energy mini-
mization using the AMBER99 force field was performed on the
enzyme. Compound 7 was built and protonated using the proton-
ate 3D protocol and energy minimized using the MMFF94 force
field via MOE 2014.09.1. Docking of the ligand was carried out
using the GOLD 5.2.1 program with default settings47,48. A sphere
of 22 Å around the carbonyl group of Glu199 was defined as the
binding site for the ligand docking and 250 confirmations was
allowed. The Chemscore and Goldscore standard precision (sp)
were calculated and analyzed. The putative binding mode was car-
ried out through visual inspection (Figure 2).
2-(4-(Morpholinosulfonyl)phenyl)isoindoline-1,3-dione (11) Yield
11%. mp 222 ꢀC; 212 ꢀC40. 1H NMR (DMSO-d6): d 8.01–7.98 (2H, m,
phthalimide-H), 7.95–7.92 (2H, m, phthalimide-H), 7.90 (2H, d,
J ¼ 8.0 Hz, benzene-H), 7.77 (2H, d, J ¼ 8.0 Hz, benzene-H), 3.63 (4H,
t, J ¼ 4.4 Hz, morpholine-H), 2.91 (4H, t, J ¼ 4.6 Hz, morpholine-H)
ppm. IR (tmaks cmꢁ 1) (FT/ATR): 2952, 2895, 2866, 2828, 1776, 1715,
1682, 1590, 1349, 1260, 1162. Anal. calcd. for C18H16N2O5S. 0.3
C2H6O: C, 57.84; H, 4.65; N, 7.25; S, 8.30. Found C, 57.40; H, 4.73; N,
7.33; S,7.91. MS (APCI) m/z (%): 373 (M þ Hþ, 100).
Biological activity
AChE (E.C.3.1.1.7., Type VI-S, from electric eel) and BuChE
(E.C.3.1.1.8., from equine serum) were purchased from Sigma-Aldrich
(Steinheim, Germany). 5,50-Dithiobis(2-nitrobenzoic acid) (DTNB,
Ellman’s reagent) acetylthiocholine iodide (AChI) and butyrylthio-
choline iodide (BChI) used as substrates were obtained from Fluka.
Buffer compounds (potassium dihydrogen phosphate, potassium
hydroxide) and sodium hydrogen carbonate were purchased from
Merck (Darmstadt, Germany). Spectrophotometric measurements
were performed on a Shimadzu 160-A UV-Vis spectrophotometer.
Results and discussion
Chemistry
As shown in Scheme 1, the synthesis of the title compounds was
realized in three steps according to the procedure in the litera-
ture35–37. Initially, phthalic anhydride and aniline were reacted to
yield N-phenylphthalimide. Then, N-phenylphthalimide was treated
with chlorosulfonic acid to give the 4-phthalimidobenzenesulfonyl
Acetylcholinesterase/butyrylcholinesterase activity assay
The inhibitory effects of the synthesized compounds on AChE and
BuChE were evaluated using a slightly modified colorimetric chloride. Finally, 4-phthalimidobenzenesulfonyl chloride was

4
Z. SOYER ET AL.
promptly converted to final sulfonamide derivatives by SN2 data are not available. The AChE and BuChE inhibitory activities of
nucleophilic reaction with appropriate amine. the all compounds have not been described in the literature, and
The synthesis of the compounds 1, 4–7, 9–11 were reported were reported for the first time in this study.
previously36,38–40,49,50. Compound 8 is listed compound with regis-
The structures of the title compounds were confirmed by spec-
try number CASRN 898471–20-0, whereas corresponding scientific tral and elemental analyses.
With regard to IR data, diagnostic vibrational bands were pro-
vided by sulfonamide and phthalimide moieties of the final com-
pounds. SO2-stretching bands of sulfonamide chromophore were
observed between1327–1354 and 1161–1180 cmꢁ1, in addition,
NH-stretching bands for compounds 1–6 were detected in the
range of 3243–3485 cmꢁ 1. Two characteristic absorption bands
were appeared at around 1782–1792 and 1707–1721 cmꢁ 1 in
spectra indicating the presence of phthalimide carbonyl groups51.
1H NMR spectra of the compounds were consistent with expected
resonance signals in terms of chemical shifts and integrations. In
1H NMR, the NH proton of secondary sulfonamide group was seen
as a broad singlet between at d 9.57–10.56 ppm (for compounds
1–6). The protons of phthalimide ring were observed as multiplets
in the range of d 8.02–7.89 ppm. Aromatic protons of phenyl ring
linked to phthalimide structure were detected at d 7.61–7.98 ppm
as two doublets with 2H integration according to the AA’BB’ pat-
tern. On the other hand, resonance signals of all the aromatic and
aliphatic protons were observed in the expected regions with
expected multiplicities confirming the proposed structure52.
The structures of the title compounds were further verified by
APCI spectra where the m/z values of molecular ion peaks were in
complete agreement with the calculated molecular weight for
each compound.
The purity levels of the compounds were determined by elem-
ental analyzes (C, H, N, S) and results were within 0.4% of the cal-
culated values.
Biological activity
Inhibitory activities of the synthesized compounds against AChE
and BuChE were evaluated by modified Ellman method, using gal-
antamine as the reference compound41–45. The AChE inhibitory
activity was determined by using electric eel acetylcholinesterase
and the BuChE inhibitory activity was tested by using equine
serum butyrylcholinesterase. The IC50 values for AChE and BuChE
inhibitions are summarized in Table 1.
Figure 2. Proposed binding mode for compound 7 inside AChE (1EVE pdb code).
The active compound is showed as green stick in AChE. The most involved resi-
dues are named and represented as brown sticks for AChE. Hydrogen bond inter-
actions are represented as blue dashed lines.
O
O
O
O
O
S
O
S
a
b
c
N
N
Cl
N
A
O
O
O
O
O
O
O
A= 2-methylaniline(1)
(2)
2-methoxyaniline
2-isopropylaniline(3)
4-methylaniline(4)
(5)
4-methoxyaniline
4-chloroaniline(6)
(7)
diethylamine
pyrolidine(8)
(9)
piperidine
2-methylpiperidine(10)
morpholine(11)
a
b
c
) Aniline, 30 min. ) ClSO3H, PCl5, 30 min. ) Amines, acetone, 2h
Scheme 1. Synthesis of the final compounds 1–11.

JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY
5
According to the biological activity results, target compounds
exhibited higher inhibitory activity against AChE than BuChE. In
addition, all compounds displayed high selectivity against AChE.
Among the tested compounds, compounds 1, 2 and 3 bearing
the substituent at ortho position of N-phenyl ring on sulfona-
mide group showed slightly better AChE inhibitory activity com-
pared to the para-substituted derivatives. Compound 7 with
diethyl substituent on nitrogen atom of the sulfonamide is the
most active compound with IC50 value of 1.35 lM against AChE.
The conversion of sulfonanilide structure of the compounds 1–6
and cyclic sulfonamide structure of the compounds 8–11 to
N,N-dietil sulfonamide produced the most active compound 7
against AChE. This finding let us to consider that the character-
ization of amide nitrogen is important for AChE inhibitory
activity.
Acknowledgements
The authors would like to thank the Pharmaceutical Sciences
Research Center (FABAL) at Ege University Faculty of Pharmacy for
spectral analyses of the compounds.
Disclosure statement
The authors declare no conflicts of interests.
Funding
This study was supported by research grants from Ege University
(Project Number: 12/Ecz/016).
Regarding BuChE activity results, generally, it is found that the
tested compounds have moderate to weak inhibition potency.
Compound 3 bearing isopropyl substituent at ortho position of
N-phenyl ring on sulfonamide exhibited the highest inhibitory
activity in the series. However, compound 7, the most active com-
pound against AChE, did not exhibit any inhibition against BuChE.
This situation possibly results from the differences of the amino
acids in the active site of the both enzymes.
References
1. Fargo K, Bleiler L. Alzheimer’s association report: 2014
Alzheimer’s disease facts and figures. Alzheimers Dement
2014;10:47–92.
2. Mucke L. Neuroscience: Alzheimer's disease. Nature
2009;461:895–7.
3. Khan I, Ibrar A, Zaib S, et al. Active compounds from a
diverse library of triazolothiadiazole and triazolothiadiazine
scaffolds: Synthesis, crystal structure determination, cytotox-
icity, cholinesterase inhibitory activity, and binding mode
analysis. Bioorg Med Chem 2014;22:6163–73.
4. Khan I, Bakht SM, Ibrar A, et al. Exploration of a library of tri-
azolothiadiazole and triazolothiadiazine compounds as a
highly potent and selective family of cholinesterase and
monoamine oxidase inhibitors: design, synthesis, X-ray dif-
fraction analysis and molecular docking studies. RSC Adv
2015;5:21249–67.
Based on these activity results, 4-phthalimidobenzenesulfona-
mide derivatives could be described as selective AChE inhibitors.
As a potential compound for treatment of AD, log p was
thought as an important physicochemical parameter to evaluate
or predict the ability to cross blood–brain barrier (BBB). It was
reported that log p with the optimum central nervous system
(CNS) penetration was around 2 0.753. The lipophilicities of the
synthesized compounds were calculated using MOE 2011.10
(Molecular Operating Environment Chemical Computing Group).
As shown in Table 1, log p values of synthesized compounds
ranged from 1.51–4.41, which indicated that all the compounds
are sufficiently lipophilic to pass the BBB.
€
€
5. Goc¸er H, Akincioglu A, Goksu S, et al. Carbonic anhydrase
and acetylcholinesterase inhibitory effects of carbamates
and sulfamoylcarbamates.
J Enzyme Inhib Med Chem
2015;30:316–20.
Docking studies
6. Goedert M, Spillantini MG. A century of Alzheimer's disease.
Science 2006;314:777–81.
7. Bolognesi ML, Andrisano V, Bartolini M, et al. Heterocyclic
inhibitors of AChE acylation and peripheral sites. Il Farmaco
2005;60:465–73.
8. Bartus RT, Dean RL, Beer B, Lippa AS. The cholinergic
hypothesis of geriatric memory dysfunction. Science
1982;217:408–17.
To explore the possible binding mode of the phthalimide deriv-
atives with AChE (PDB code 1EVE), docking studies were per-
formed using Gold 5.2.1. for the most active compound 7 in
the series. As shown in Figure 2, compound 7 exhibited two
binding modes with AChE. In the bottom of the gorge, the
phthalimide moiety interact with Trp84 via the p–p stacking
interaction and the oxygen atom of sulfonamide group create a
hydrogen bond with hydroxyl group of Tyr121. Summing up, it
can be proposed that compound 7 can interact both with CAS
and PAS of AChE.
9. Blennow K, Leon MJ, Zetterberg H. Alzheimer's disease.
Lancet 2006;368:387–403.
ꢀ
10. Konrath EL, Passos CS, Klein-Junior LC, Henriques AT.
Alkaloids as a source of potential anticholinesterase inhibi-
tors for the treatment of Alzheimer's disease. J Pharm
Pharmacol 2013;65:1701–25.
Conclusion
11. Shaikh S, Verma A, Siddiqui S, et al. Current acetylcholin-
In this study, eleven 4-phthalimidobenzenesulfonamide derivatives
were synthesized and evaluated for their in vitro AChE and BuChE
inhibition. Among the series, compound 7 and 3 showed the
highest activity against AChE and BuChE, respectively. Generally,
all the tested compounds displayed selectivity for AChE. Molecular
modeling study of the most active compound 7 against AChE
demonstrated binding interactions with both PAS and CAS of the
enzyme. According to the calculated log p values, all the com-
pounds might pass to the BBB. These sulfonamide derivatives
could be considered as new lead compounds to develop more
potent AChE and BuChE inhibitors.
esterase-inhibitors:
A neuroinformatics perspective. CNS
Neurol Disord Drug Targets 2014;13:391–401.
12. Toda N, Kaneko T, Kogen H. Development of an efficient
therapeutic agent for Alzheimer's disease: design and syn-
thesis of dual inhibitors of acetylcholinesterase and sero-
tonin transporter. Chem Pharm Bull 2010;58:273–87.
13. Farina R, Pisani L, Catto M, et al. Structure-based design and
optimization of multitarget-directed 2H-chromen-2-one
derivatives as potent inhibitors of monoamine oxidase B and
cholinesterases. J Med Chem 2015;58:5561–78.

6
Z. SOYER ET AL.
14. Bajda M, Więckowska A, Hebda M, et al. Structure-based 30. Shah SSA, Rivera G, Ashfaq M. Recent advances in medicinal
search for new inhibitors of cholinesterases. Int J Mol Sci
2013;14:5608–32.
chemistry of sulfonamides. Rational design as anti-tumoral,
anti-bacterial and anti-inflammatory agents. Mini Rev Med
15. Pohanka M. Inhibitors of acetylcholinesterase and butyryl-
Chem 2013;13:70–86.
€
cholinesterase meet immunity. Int
J
Mol Sci 2014;15:
€
ꢁ
31. Goc¸er H, Akıncıoglu A, Oztas¸kın N, et al. Synthesis, antioxi-
dant, and antiacetylcholinesterase activities of sulfonamide
derivatives of dopamine-related compounds. Arch Pharm
Chem Life Sci 2013;346:783–92.
9809–25.
16. Khan MA, Fazal-ur-Rehman S, Hameed A, et al.
Regioselective synthesis of novel 2,3,4,4a-tetrahydro-1H-car-
bazoles and their cholinesterase inhibitory activities. RSC
Adv 2015;5:59240–50.
32. Kang JE, Cho JK, Curtis-Long MJ, et al. Inhibitory evaluation
of sulfonamide chalcones on #946;-secretase and acylcholi-
nesterase. Molecules 2013;18:140–53.
33. Abbasi MA, Ahmad S, Rehman A, et al. Sulfonamide deriva-
tives of 2-amino-1-phenylethane as suitable cholinesterase
inhibitors. Trop J Pharm Res 2014;13:739–45.
_
ꢁ
ꢁ
€
17. Akincioglu A, Akincioglu H, Gulc¸in I, et al. Discovery of
potent carbonic anhydrase and acetylcholine esterase
inhibitors: novel sulfamoylcarbamates and sulfamides
derived from acetophenones. Bioorg Med Chem
2015;23:3592–602.
34. Mutahir S, Jonczyk J, Bajda M, et al. Novel biphenyl bis-
sulfonamides as acetyl and butyrylcholinesterase inhibitors:
synthesis, biological evaluation and molecular modeling
studies. Bioorg Chem 2016;64:13–20.
€
18. Yılmaz S, Akbaba Y, Ozgeris¸ B, et al. Synthesis and inhibitory
properties of some carbamates on carbonic anhydrase and
acetylcholin esterase. J Enzym Inhib Med Chem 2016;31:
1484–91.
35. Lida K, Kajiwara M. Synthesis of 13C and 15N multilabeled 5-
aminolevulinic acid. J Label Compd Radiopharm 2002;45:
139–43.
19. Scozzavafa A, Kalın P, Supuran CT, et al. The impact of
hydroquinone on acetylcholine esterase and certain human
carbonic anhydrase isoenzymes (hCA I, II, IX, and XII).
J Enzyme Inhib Med Chem 2015;30:941–6.
36. Lima LM, Castro P, Machado AL, et al. Synthesis and anti-
inflammatory activity of phthalimide derivatives, designed as
new thalidomide analogs. Bioorg Med Chem 2002;10:
3067–73.
37. Dewar MJS, King FE. Sulphanylamides of some aminopyra-
zoles and a note on the application of p-phthalimidobenze-
nesulphonyl chloride to the synthesis of sulfanilamides.
J Chem Soc 1945;29:114–16.
38. Timiri AK, Subasri S, Kesherwani M, et al. Synthesis and
molecular modeling studies of novel sulfonamide derivatives
as dengue virus 2 protease inhibitors. Bioorg Chem 2015;62:
74–82.
39. Heinrich DM, Flanagan JU, Jamieson SMF, et al. Synthesis
and structure activity relationships for 1-(4-(piperidin-1-ylsul-
fonyl)phenyl)pyrrolidin-2-ones as novel non-carboxylate
inhibitors of the aldo-ketoreductase enzyme AKR1C3. Eur J
Med Chem 2013;62:738–44.
40. Cremlyn RJ, Thandi K, Wilson R. Derivatives of cinnamide-4-
sulfonyl chloride and p-(phthalimido)benzenesulfonyl chlor-
ide. Ind J Chem Sect B 1984;23B:94–6.
41. Ozgun DO, Yamalı C, Gul HI, et al. Inhibitory effects of isatin
Mannich bases on carbonic anhydrases, acetylcholinesterase,
and butyrylcholinesterase.
2016;31:1498–501.
42. Sujayev A, Garibov E, Taslimi P, et al. Synthesis of
some tetrahydropyrimidine-5-carboxylates, determination
of their metal chelating effects and inhibition profiles
against acetylcholinesterase, butyrylcholinesterase and
_
€
20. Gulc¸in I, Scozzafava A, Supuran CT, et al. The effect of caffeic
acid phenethyl ester (CAPE) metabolic enzymes including
acetylcholinesterase, butyrylcholinesterase, glutathione-s-
transferase, lactoperoxidase and carbonic anhydrase
ısoenzymes I, II, IX and XII. J Enzym Inhib Med Chem
2015;31:1060–6.
€
€
€
21. Ozgeris¸ B, Goksu S, Kose LP, et al. Acetylcholinesterase and
carbonic anhydrase inhibitory properties of novel urea and
sulfamide derivatives incorporating dopaminergic 2-aminote-
tralin scaffolds. Bioorg Med Chem 2016;24:2318–29.
€
22. Goc¸er H, Topal F, Topal M, et al. Acetylcholinesterase and
carbonic anhydrase isoenzymes I and II inhibition profiles of
taxifolin. J Enzyme Inhib Med Chem 2016;31:441–7.
23. Sharma U, Kumar P, Kumar N, Singh B. Recent advances in
the chemistry of phthalimide analogues and their thera-
peutic potential. Mini Rev Med Chem 2010;10:678–704.
€ €
24. Alptuzun V, Kapkova P, Baumann K, et al. Synthesis and bio-
logical activity of pyridinium-type acetylcholinesterase inhibi-
tors. J Pharm Pharmacol 2003;55:1397–404.
25. Si W, Zhang T, Zhang L, et al. Design, synthesis and
bioactivity of novel phthalimide derivatives as acetyl-
cholinesterase inhibitors. Bioorg Med Chem Lett
2015;26:2380–2.
26. Ignasik M, Bajda M, Guzior N, et al. Design, synthesis and
evaluation of novel 2-(Aminoalkyl)isoindoline-1,3-dione
derivatives as dual-binding site acetylcholinesterase inhibi-
tors. Arch. Pharm. (Weinheim) 2012;345:509–16.
27. Zhao Q, Yang G, Mei X, et al. Novel acetylcholinesterase
inhibitors: Synthesis and structure–activity relationships of
phthalimide alkyloxyphenyl N,N-dimethylcarbamate deriva-
tives. Pestic Biochem Physiol 2009;95:131–4.
J Enzym Inhib Med Chem
carbonic anhydrase.
J
Enzym Inhib Med Chem
2016;31:1531–9.
_
€
43. Gulc¸in I, Scozzafava A, Supuran CT, et al. Rosmarinic acid
inhibits some metabolic enzymes including glutathione
S-transferase, lactoperoxidase, acetylcholinesterase, butyryl-
cholinesterase and carbonic anhydrase isoenzymes. J Enzym
Inhib Med Chem 2016;31:1698–702.
28. Alonso D, Dorronsoro I, Rubio L, et al. Donepezil-tacrine
hybrid related derivatives as new dual binding site inhibitors
of AChE. Bioorg Med Chem Lett 2005;13:6588–97.
44. Kapkova P, Stiefl N, Surig U, et al. Synthesis, biological activ-
ity, and docking studies of new acetylcholinesterase inhibi-
tors of the bispyridinium type. Arch Pharm Pharm Med
Chem 2003;336:523–40.
29. Aliabadi A, Foroumadi A, Mohammadi-Farani A, Mahvar
MG. Synthesis and evaluation of anti-acetylcholinester-
ase activity of 2-(2-(4-(2-Oxo-2-phenylethyl)piperazin-1-
yl)ethyl)isoindoline-1,3-dione derivatives with potential
45. Ellman GL, Courtney KD, Andres V, Feather-Stone RM.
Biochem Pharmacol 1961;7:88–95.
anti-Alzheimer
2013;16:1049–54.
effects.
Iran
J
Basic
Med
Sci

JOURNAL OF ENZYME INHIBITION AND MEDICINAL CHEMISTRY
7
46. Molecular Operating Environment (2014.09.1) Chemical 50. Arti S K, Pathak D. Synthesis and anticonvulsant screening of
Computing Group Inc. 1010 Sherbrooke Street West, Suite
91, Montreal H3A 2R7, Canada.
47. Jones G, Willett P, Glen RC, et al. Development and valid-
4-phthalimide-N-(40-substitutedphenylbenzenesulphona-
mide,4-succinimide-N-(40-substituted phenyl) benzenesulpho-
namide. Int J Pharm Tec Res 2011;3:2104–10.
ation of a genetic algorithm for flexible docking. J Mol Biol 51. Nakanishi K, Solomon PH. Infrared absorption spec-
1997;267:727–48.
48. Friesner RA, Banks JL, Murphy RB, et al. Glide: A new
troscopy. 2nd ed. San Fransisco: Holden-Day Inc;
1977.
approach for rapid, accurate docking and scoring. 1. Method 52. Hesse M, Meier H, Zeeh B. Spectroscopic methods in organic
and assessment of docking accuracy. J Med Chem 2004;47:
1739–49.
chemistry. In: Enders D, Noyori R, Trost BM, eds. Nuclear
Magnetic Resonance. New York: Georg Thieme Verlag
Stuttgart; 1997.
49. Cremlyn
RJ,
Swinbourne
FJ,
Nunes
RJ.
Phthalimidebenzenesulphonyl derivates. Ouimica Nova 53. Glave WR, Hansch C. Relationship between lipophilic charac-
1985;8:61–2. ter and anesthetic activity. J Pharm Sci 1972;61:589–91.