Design and synthesis of new donepezil analogs derived from arylpiperazine scaffold as acetylcholinesterase inhibitors

Article abstract of DOI:10.1080/10426507.2020.1830773

Newly synthesized 4-substituted phenyl-2-(4-substituted phenylpiperazine-1-yl)thiazole derivatives (4a–v) were evaluated in terms of their acetylcholinesterase (AChE) inhibition activities. Twenty-two compounds were tested against AChE at six different concentrations that varied between 10^?4 and 10^?9 M. The concentrations that inhibited AChE were calculated between 1.15 and 3.45 μM in seven compounds (4a, 4b, 4h, 4l, 4m, 4q, 4r). Compounds 4m, 4b, and 4l represented 1.15, 1.31, 1.34 μM (IC₅₀) inhibitions, respectively. Although the inhibition values are lower than that of donepezil, they are considerable. Modeling studies of these analogs revealed similar positioning with donepezil, in which Ar–Ar interactions with Tyr337 and Trp 286 exist.

Full text of DOI:10.1080/10426507.2020.1830773

Phosphorus, Sulfur, and Silicon and the Related Elements
ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/gpss20
Design and synthesis of new donepezil analogs
derived from arylpiperazine scaffold as
acetylcholinesterase inhibitors
Zafer Sahin , Sevde Nur Biltekin , Emre Fatih Bülbül , Leyla Yurttas , Barkin
Berk & Şeref Demirayak
To cite this article: Zafer Sahin , Sevde Nur Biltekin , Emre Fatih Bülbül , Leyla Yurttas , Barkin
Berk & Şeref Demirayak (2020): Design and synthesis of new donepezil analogs derived from
arylpiperazine scaffold as acetylcholinesterase inhibitors, Phosphorus, Sulfur, and Silicon and the
Related Elements, DOI: 10.1080/10426507.2020.1830773
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PHOSPHORUS, SULFUR, AND SILICON AND THE RELATED ELEMENTS
https://doi.org/10.1080/10426507.2020.1830773
Design and synthesis of new donepezil analogs derived from
arylpiperazine scaffold as acetylcholinesterase inhibitors
b
€ € d
Zafer Sahina,b , Sevde Nur Biltekinc , Emre Fatih Bulbul , Leyla Yurttas
, Barkin Berka , and S¸eref
Demirayaka
b
aDepartment of Pharmaceutical Chemistry, School of Pharmacy, Istanbul Medipol University, Istanbul, Turkey; Department of
c
Pharmaceutical Chemistry, Faculty of Pharmacy, Anadolu University, Eskis¸ehir, Turkey; Department of Pharmaceutical Microbiology, School
d
of Pharmacy, Istanbul Medipol University, Istanbul, Turkey; Department of Medicinal Chemistry, Institute of Pharmacy, Martin Luther
University Halle-Wittenberg, Halle (Saale), Germany
ABSTRACT
ARTICLE HISTORY
Received 17 June 2020
Accepted 27 September 2020
Newly synthesized 4-substituted phenyl-2-(4-substituted phenylpiperazine-1-yl)thiazole derivatives
(4a–v) were evaluated in terms of their acetylcholinesterase (AChE) inhibition activities. Twenty-
two compounds were tested against AChE at six different concentrations that varied between
10ꢀ4 and 10ꢀ9 M. The concentrations that inhibited AChE were calculated between 1.15 and
3.45 mM in seven compounds (4a, 4b, 4h, 4l, 4m, 4q, 4r). Compounds 4m, 4b, and 4l represented
1.15, 1.31, 1.34 mM (IC50) inhibitions, respectively. Although the inhibition values are lower than
that of donepezil, they are considerable. Modeling studies of these analogs revealed similar posi-
tioning with donepezil, in which Ar–Ar interactions with Tyr337 and Trp 286 exist.
KEYWORDS
Thiazole; piperazine;
anticholinesterase activity;
acetylcholinesterase inhib-
ition; molecular modeling
GRAPHICAL ABSTRACT
Introduction
abilities, regulate behavior, and slow down the progress of AD
by modulating the levels of certain neurotransmitters in the
brain such as acetylcholinesterase (AChE) and glutamate.[6]
Inhibition of AChE is the most promising strategy for AD
treatment since accumulation of b-amyloid plaques is con-
trolled by AChE, as well. Similarly, it was shown that there is
an inverse relationship between b amyloid plaque deposition
and ACh level in the brain. In this manner, AChE inhibition
elevates the level of acetylcholine in the brain, which in turn
reduces b amyloid plaque aggregation. Thus, AChE is an
important target to control the disease.[7] Clinically existing
drugs against AD include rivastigmine, galantamine, ensacu-
lin, and donepezil. All of these drugs are critical AChE inhibi-
tors, yet the significance of donepezil is distinguished from
According to the World Health Organization, by 2019
approximately 47 million people, worldwide, were affected by
dementia.[1] It is estimated that by the end of 2030, 75 million
people will be affected. In 2018, the United States of America
invested more than $270 billion for patients suffering from
dementia.[2] Alzheimer’s disease (AD), as the most common
form of dementia, is an irreversible neurodegenerative brain
disease.[3] In the last 20 years, multiple drugs have been clinic-
ally tested and various strategies including memory improve-
ment, blocking of Ab (amyloid b) plaque accumulation, gene
therapy, neuronal function repairment, and increasing the
immune response by vaccines have been proposed. Despite
many efforts, only four drugs have been approved for clinical
use.[4,5] The aim of the current strategies is to preserve mental others by being a very potent inhibitor with 0.07 lM
CONTACT Zafer Sahin
zsahin@medipol.edu.tr
Department of Pharmaceutical Chemistry, School of Pharmacy, Istanbul Medipol University, Istanbul, Turkey.
Supplemental data for this article is available online at https://doi.org/10.1080/10426507.2020.1830773.
ß 2020 Taylor & Francis Group, LLC

2
Z. SAHIN ET AL.
Figure 1. Design strategy of the titled compounds.
experimental IC50 value. Although donepezil is highly potent
and relatively safe, its extrapyramidal and cardiovascular side
effects encourage scientists to discover new molecules.[8]
Donepezil analogs have wide structure–activity relationships.
Cheung et al. have investigated the binding profile of donepe-
zil and the AChE through enlightening the enzyme–ligand
complex crystal structure.[9]
Structure–activity relationships of donepezil propose an aro-
matic core system, which is attached to a basic aliphatic ring
along with a carbon chain or a small ring bridge (Figure 1).[10–12]
Piperazine–thiazole analogs bearing phenyl and benzoyl groups
on both sides are evidenced inhibitors of AChE (Figure 1). On
the other hand, the thiazole ring with different modification is
reported as an acetylcholinesterase inhibitor in the litera-
Results and discussion
Chemistry
All the target compounds were synthesized successfully with
yields ranging between 64% and 85% and their structures were
determined using NMR and IR spectroscopy and High
Resolution Mass Spectrometry. No characteristic peaks were
detected in IR since no free H, carbonyl, or triple bond existed.
Aromatic and aliphatic C–H stretched out between 2800 and
3150cmꢀ1. Molecular weights were determined using HRMS.
NMR results revealed the piperazine CH2 groups as two sepa-
rated singlet peaks, around 3.1 and 3.6 ppm. Methoxy groups
were observed around 3.8 ppm in compounds 4d, 4e, 4l–v.
Compounds containing methyl groups (4b, 4c, 4m, 4n) yielded
ture.[13–18] Piperazine is a significant ring contributing to broad CH3 peaks around 2.3 ppm. The single hydrogen of thiazole has
pharmacological activities since it is an ionizable molecule. been detected around 7.6 ppm, which is critical to exhibit that
Besides, piperazine adds solubility to a molecule, and promotes one position was not substituted. The number of aromatic C–H
its ADME properties.[19,20]
peaks corresponded to the anticipated hydrogen count.
Acetylcholinesterase, which is initially retrieved from According to 13C NMR results, thiazole’s carbon at the second
Pacific electric ray and mice, has been recently obtained position was monitored as the highest peak in the spectrum
from humans. The donepezil AChE complex has been exten- (167–170 ppm). Methoxy and methyl groups were observed at
55ppm and 21–23ppm, respectively. Spectroscopic results indi-
cated that structure of the compounds were correct and valid.
sively anaylzed by X-ray crystallographic techniques.
Additionally, AChE has a narrow cavity consisting of two
parts, namely catalytic site and peripheral anionic site (PAS).
Cation–p interacting tryptophan, hydrogen bonding glycine,
and serine (have an important function in acetylation with
hydroxyl group) are critical amino acids. Furthermore, done-
pezil is known to stretch out to PAS by aromatic interactions
with Trp286 and Arg296, which lead to the identification of
the benzyl group of donepezil and propylcoumarin group of
ensaculin.[9,21] In this current study, 22 compounds with
Enzyme inhibition
Enzyme inhibition tests were carried out the Ellman method
along with minor modifications to the previous studies. The
tests were performed in concentrations ranging between
10ꢀ4 and 10ꢀ9 M (see Table 1). Seven out of 22 yielded up
to 70% inhibition at 10ꢀ4 M, whereas most of the com-
disubstituted (phenyl, piperazine) thiazole structures were pounds exhibited more than 50% inhibition. IC50 values
synthesized and tested against human AChE. were detected using six concentrations. Accordingly, IC50

PHOSPHORUS, SULFUR, AND SILICON AND THE RELATED ELEMENTS
3
Table 1. The inhibition percents of compounds 4a–4v against AChE at 10ꢀ4 M
Table 2. IC50 (mM) of the active compounds and donepezil against AChE.
concentrations.
Concentrations
AChE % inhibition
10ꢀ4 M
AChE % inhibition
10ꢀ4 M
Compound
IC50 (mM)
Compound
Compound
4a
4b
4h
4l
4m
4q
4r
1.80 0.02
1.31 0.18
2.90 0.14
1.34 0.10
1.15 0.14
2.19 0.14
3.45 0.10
0.07 0.002
4a
4b
4c
4d
4e
4f
4g
4h
79.28 0.73
79.60 0.65
57.09 0.63
63.14 0.36
66.86 0.43
61.70 0.13
20.91 0.37
74.03 0.53
50.83 0.35
27.37 0.13
15.35 0.88
%98.02 0.35
4l
82.79 0.55
82.47 0.43
65.96 0.22
59.17 0.54
46.26 0.59
76.21 0.32
70.31 0.35
58.47 0.18
35.11 0.95
43.70 0.15
24.02 0.36
4m
4n
4o
4p
4q
4r
4s
4t
4u
4v
Donepezil
Bold value represents the most active compound in the series and modeling
studies is presented with this molecule.
4i
4j
4k
Donepezil
The inhibition values higher than 70% are represented in bold.
According to our results, however, the absence of benzoyl
did not contribute to the affinity. The activity decreased,
even though they were still comparable with the previous
research. Remarkably, none of the 4-substituted R0 group
values of the compounds were calculated as follows; 4m:
1.15 mM, 4b: 1.31 mM, 4l: 1.34 mM, 4a: 1.80 mM, 4q: 2.19 mM, was found to be active, except for 4r, which included 4-F
4h: 2.90 mM, 4r: 3.45 mM (Table 2). Although these mole- group. However, F was also not a bulky substituent. The R0
groups of other active compounds were H, methyl and
fluorine at third positions. The posing of donepezil and
tested compounds was comparable. Interactions with Trp
86, Tyr 337, and Trp 286 amino acids from both sides of
4m were evidently demonstrated. According to the bio-
logical findings, a carbonyl group was required for the posi-
tioning through Arg2960Phe295 aminoacids’ NH groups in
order to achieve improved results, which was supported by
the comparison of previous study and donepezil. To the best
of our knowledge, this characteristic of donepezil analogs
has not been extensively discussed before (see Figure 3).
cules were not very active compared to donepezil (0.07 mM),
chemical structure consistencies and close IC50 results of
abovementioned seven compounds were critically significant.
Compounds 4l–v had 2-methoxy groups on the phenyl
attached-piperazine. Meanwhile, phenyl group attached-thia-
zole beared H, 3-methyl, 3-F and 4-F substituents in 4l, 4m,
4q, and 4r, respectively. 4a and 4b held H and 3-methyl
substituents, which were similar to 4l and 4m.
Molecular modeling
Compound 4m exhibited ꢀ72.4944 glide model score and
ꢀ9.7421 docking score. Trp86 and Tyr 337 are important
amino acids at the catalytic site, forming p–p and p–cation
interactions. On the other hand, Trp 286 and Tyr 341 are
important amino acids for the peripheric anionic site as
well.[22] Poses in Figure 2 demonstrate interactions with
these amino acids. Donepezil’s indanone interacts with
Trp286, whereas the benzyl group on the other side interacts
with Trp 86 at the catalytic site. Tyr337 was shown to form
a p–cation interaction with piperidine. Docking poses
revealed the pose consistency of our designed compounds
with donepezil, in which both of them interacted and
docked as shown in Figure 2. Phenyl–thiazole moiety of the
tested compounds matched with the indenonyl methyl
group of donepezil. Supportively, benzyl piperidine part of
donepezil represented similar posing with phenyl piperazine
of the tested compounds, namely 4a–l. Remarkably, it was
not discussed before that Phe295 or Arg296 linked amino
acids NH groups are also key feature to form hydrogen
bonding with carbonyl groups on ligands. This interaction is
likely to provide enhanced binding and enzymatic results.
Our modeling studies strongly supported the current infor-
mation about donepezil analog scaffolds. It was shown that,
in trisubstituted thiazoles (Figure 1), benzoyl group inter-
acted with Arg296 and increased the binding ability through
this interaction. Inversely, in this study, we aimed to evalu-
ate whether the absence of a benzoyl group would increase
Methods
Chemistry
All chemical and biological substances used in this study
were purchased from Sigma Aldrich Chemical Corp. (St.
Louis, Missouri, USA) and Merck KGaA (Merck,
Darmstadt, Germany). Thin Layer Chromatography (TLC)
was performed with a Silica Gel 60 F254 TLC plate (Merck
KGaA, Darmstadt, Germany) in order to determine the
purities of the intermediate compounds. Melting points were
detected using Stuart Melting Point Apparatus SMP30
(Staffordshire, UK). Leco TruSpec Micro CHN/CHNS elem-
ental analyzer (Saint Joseph, Michigan, USA) was operated
to investigate the C, H, and N fractions of the crystallized
compounds. Spectroscopic data were obtained with the fol-
lowing instruments: FT-IR, Perkin Elmer Spectrum Two FT-
IR Spectrometer (Perkin Elmer, Inc., Waltham,
Massachusetts, USA); 1H NMR (300 MHz) and 13C NMR
(75 MHz)
Bruker
300 MHz
UltraShield
NMR
Spectrophotometer (Bruker Corp., Billerica, Massachusetts,
USA); in DMSO-d6 using TMS as a standard. Finally,
Shimadzu 8040 LC/MS/MS ITTOF system (Shimadzu,
Tokyo, Japan) mass spectrometer was used to distinguish
the mass spectra of compounds by the electron spray
method (ESI). The Supplemental Materials contain sample
1H and 13C NMR and high resolution mass spectra for the
the binding affinity due to the narrowness of the cavity. products 4 (Figures S1–S66).

4
Z. SAHIN ET AL.
Figure 2. 2D and 3D poses of donepezil (A) and 4m (B).
General synthesis of 2-((4-substitutedphenylpiperazine-1-
carbonothioyl)thio)acetic acid derivatives (1a–b)
General synthesis of N-substitutedphenyl-N0-piperazine-
thioamide derivatives (2a–b)
The piperazine derivative (600 mmol) was dissolved in pyri-
dine (150 ml) and 20% NaOH solution containing 600 mmol
sodium hydroxide. Carbon disulfide (1.2 mol) was added to
the mixture dropwise at 0–2 ꢁC. After stirring for 30 min at
0–2 ꢁC, equal moles of chloroacetic acid (600 mmol) were
added and boiled for 1 h. Subsequent to reaction completion,
pyridine was evaporated and water was added to the mix-
ture. Due to sodium salt formation, it was neutralized with
acetic acid and the precipitated product was removed by fil-
tration. This method is adapted from the previous synthesis
Mercaptoacetic acid group was removed from 1a–b by
hydrolyzing in excess ammonia solution. 1a–b was dissolved
in ammonia solution and refluxed for an hour. The reaction
mixture was cooled down to room temperature and the pre-
cipitate was filtered off. Solid material was crystallized
from ethanol.
a-Bromoacetophenone derivatives (3a–k)
The acetophenone derivative (600 mmol) was dissolved in
400 ml acetic acid. Catalytic amount of hydrobromic acid
was added to a flask. Equal mole of bromine (600 mmol)
ꢀ
studies of Spalinska et al., which were carried out by dithio-
carbamate derivatives (see Figure 4).[23]

PHOSPHORUS, SULFUR, AND SILICON AND THE RELATED ELEMENTS
5
Figure 2. Continued.
was diluted in acetic acid and added dropwise. Following (C–H), 1329–1598 (C ¼ C, C ¼ N). 1H NMR (300 MHz,
DMSO-d6) d (ppm): 3.31 (4H, t, J: 5.08 Hz piperazine C2,6),
3.63 (4H, t, J: 5.07 Hz piperazine C3,5), 6.85 (1H, t, J:
7.23 Hz, pip-phenyl-C4), 7.02 (2H, d, J: 7.95 Hz pip-phenyl
the completion of the reaction, water was added to the mix-
ture and the obtained intermediate was filtrated.[24]
C2,6), 7.24–7.32 (4H, m, thiazole C5, pip-phenyl-C3,5, th-phe-
General synthesis of 4-(substituted phenyl)-2-(4-(substi-
tuted phenyl) piperazine-1-yl)thiazole derivatives (4a–v)
Equimolar quantities of piperazinethiourea (2a–b)
nyl C4), 7.40 (2H, t, J: 7.44 Hz, th-phenyl C3,5), 7.88 (2H, d,
J: 7.14 Hz, th-phenyl C2,6). 13C NMR (75 MHz, DMSO-d6) d
(ppm): 40.28 (piperazine CH2), 48.51 (piperazine CH2),
(50 mmol) and bromoacetophenones (3a–k) (50 mmol) were 103.34, 116.71, 120.27, 126.20, 128.06, 128.52, 129.6, 129.90,
135.01, 150.98 (thiazole C4), 170.82 (thiazole C2). For
C19H19N3S calcd % C, 70.99; H, 5.66; N, 13.07. Found; C,
71.01%; H, 5.63%; N, 13.04%. HRMS (m/z) M þ H calcd
322.1372; found: 322.1358.
refluxed in ethanol (60 ml) for 5 h. The reaction mixture was
cooled down. Water and NaHCO3 solution were added to
the mixture until the neutralization completed. The crystal-
lization process was accomplished using ethanol.[25]
4-Phenyl-2-(4-phenylpiperazine-1-yl)thiazole (4a). Yield
4-(3-Methylphenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com-
70%, mp; 129–130 ꢁC. FT-IR ῡ
(cmꢀ1): 2820–3125 pound 4b). Yield 80%, mp; 121–122.5 ꢁC. FT-IR ῡ max
max

6
Z. SAHIN ET AL.
Figure 3. Superposing of 4m (C: White) and donepezil (C: Black) at active site.
Figure 4. Synthetic method for compounds 4a–v.
(cmꢀ1): 2826–3129 (C–H), 1324–1598 (C ¼ C, C ¼ N). 1H 119.96, 123.38, 126.75, 128.69, 128.87, 129.49, 135.05,
NMR (300 MHz, DMSO-d6) d (ppm): 2.34 (3H, s, CH3), 138.03, 151.26 (thiazole C4), 170.79 (thiazole C2). For
3.30 (4H, m, piperazine C2,6), 3.61 (4H, t, J: 5.23 Hz, pipera- C20H21N3S calcd %; C, 71.61; H, 6.31; N, 12.43. Found; %
C, 71.64; H, 6.33; N, 12.44. HRMS (m/z) M þ H: calcd
zine C3,5), 6.83 (1H, t, J: 7.32 Hz, pip-phenyl-C4), 7.01 (2H,
336.1529; found: 336.1527.
d, J: 7.77 Hz, pip-phenyl C2,6), 7.10 (1H, d, J: 7.90 Hz, th-
phenyl C4), 7.21–7.30 (4H, m, pip-phenyl-C3,5, th-phenyl C5,
thiazole C5), 7.67 (2H, t, J: 8.03 Hz, th-phenyl C2,6). 13C 4-(4-Methylphenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com-
NMR (75 MHz, DMSO-d6) d (ppm): 21.59 (CH3), 40.28 pound 4c). Yield 81%, mp; 181–183 ꢁC. FT-IR ῡmax (cmꢀ1):
(piperazine CH2), 48.50 (piperazine CH2), 103.12, 116.55, 2813–3122 (C–H), 1326–1599 (C ¼ C, C ¼ N). 1H NMR

PHOSPHORUS, SULFUR, AND SILICON AND THE RELATED ELEMENTS
7
(300 MHz, DMSO-d6) d (ppm): 2.31 (3H, s, CH3), 3.27 (4H, 12.38. Found; C, 67.25; H, 5.37; N, 12.40. HRMS (m/z)
m, piperazine C2,6), 3.60 (4H, t, J: 5.04 Hz piperazine C3,5), M þ H: calcd 340.1278; found: 340.1275.
6.83 (1H, t, J: 7.20 Hz, pip-phenyl-C4), 7.00 (2H, d, J:
7.85 Hz, pip-phenyl C2,6), 7.18–7.27 (5H, m, pip-phenyl-C3,5
,
4-(4-Fluorophenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com-
pound 4g). Yield 80%, mp; 142–144 ꢁC. IR ῡmax (cmꢀ1):
2833–3090 (C–H), 1308–1609 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.29 (4H, m, piperazine
C2,6), 3.60 (4H, t, J: 5.11 Hz piperazine C3,5), 6.82 (1H, t,
pip-phenyl-C4), 7.01 (2H, d, J: 7.74 Hz, pip-phenyl C2,6),
7.17–7.28 (5H, m, th-phenyl C3,5, pip-phenyl C3,5, thiazole
C5), 7.90 (2H, td, J: 6.02 Hz, J: 2.22 Hz, pip-phenyl C3,5). 13C
NMR (75 MHz, DMSO-d6) d (ppm): 40.29 (piperazine CH2),
48.40 (piperazine CH2), 103.00, 116.55, 119.97, 128.22,
th-phenyl C3,5, thiazole C5), 7.76 (2H, d, J: 8.13 Hz, th-phe-
nyl C2,6–H). 13C NMR (75 MHz, DMSO-d6) d (ppm): 21.27
(CH3), 40.29 (piperazine CH2), 48.42 (piperazine CH2),
102.38, 116.55, 119.96, 126.12, 129.12, 129.48, 129.54,
129.86, 132.47, 137.27, 151.18 (thiazole C4), 170.79 (thiazole
C2). For C20H21N3S calcd % C; 71.61; H, 6.31; N, 12.43.
Found; C, 71.63; H, 6.30; N, 12.45. HRMS (m/z) M þ H:
calcd 336.1529; found: 336.1534.
4-(3-Methoxyphenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com- 129.48, 131.70, 131.74, 151.24, 160.47, 163.71 (thiazole C4),
pound 4d). Yield 84%, mp; 99–101 ꢁC. FT-IR ῡmax (cmꢀ1):
170.93 (thiazole C2). For C19H18FN3S calcd; % C, 67.23; H,
5.35; N, 12.38. Found; C, 67.23; H, 5.36; N, 12.41. HRMS
(m/z) M þ H: calcd 340.1278; found: 340.1285.
2820–3122 (C–H), 1326–1599 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.28 (4H, t, J: 5.16 Hz,
piperazine C2,6), 3.60 (4H, t, J: 5.03 Hz, piperazine C3,5),
3.80 (3H, s, OCH3), 6.80–6.88 (2H, m, thiazole C5, th-phe-
nyl-C4), 7.00 (2H, d, J: 7.99 Hz, pip-phenyl C2,6), 7.22–7.33
(4H, m, pip-phenyl-C3,5, th-phenyl C3,5), 7.44 (2H, d, J:
4.24 Hz, th-phenyl C2,6). 13C NMR (75 MHz, DMSO-d6) d
(ppm): 48.40 (piperazine CH2), 55.53 (piperazine CH2),
55.57 (OCH3), 103.67, 111.73, 113.57, 116.55, 118.65, 119.96,
129.48, 130.04, 136.50, 150.97, 151.24, 159.95 (thiazole C4),
170.73 (thiazole C2). For C20H21N3OS calcd; % C, 68.35; H,
6.02; N, 11.96. Found; C, 68.33; H, 6.00; N, 11.97. HRMS
(m/z) M þ H: calcd 352.1478; found: 352.1469.
4-(3-Chlorophenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com-
pound 4h). Yield 85%, mp; 116–117.6 ꢁC. IR ῡmax (cmꢀ1):
2833–3109 (C–H), 1308–1609 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.29 (4H, m, piperazine C2,6),
3.61 (4H, t, J: 5.00 Hz piperazine C3,5), 6.83 (1H, t, J: 7.30 Hz,
pip-phenyl-C4), 7.00 (2H, d, J: 8.15 Hz, pip-phenyl C2,6), 7.25
(2H, t, J: 7.95 Hz, pip-phenyl C3,5), 7.33 (1H, dt, J: 8.06 Hz, J:
1.10 Hz, th-phenyl C4), 7.39–7.46 (2H, m, th-phenyl C5, thia-
zole C5), 7.83 (1H, dt, J: 7.71 Hz, J:1.38 Hz, th-phenyl C2), 7.93
(1H, t, J: 1.82 Hz, th-phenyl C6). 13C NMR (75 MHz, DMSO-
d6) d (ppm): 40.29 (piperazine CH2), 48.34 (piperazine CH2),
104.82, 116.55, 114.79, 116.56, 119.97, 124.69, 129.49, 130.90,
133.89, 137.11, 149.47, 152.22 (thiazole C4), 170.92 (thiazole
C2). For C19H18ClN3S calcd; % C, 64.12; H, 5.10; N, 11.81.
Found; C, 64.15; H, 5.10; N, 11.83. HRMS (m/z) M þ H: calcd
356.0983; found: 356.0981.
4-(4-Methoxyphenyl)-2-(4-phenylpiperazine-1-yl)thiazole
(compound 4e). Yield 78%, mp; 178–179 ꢁC. IR ῡmax (cmꢀ1):
2830–3086 (C–H), 1336–1609 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.29 (4H, m, piperazine
C2,6), 3.58 (4H, m, piperazine C3,5), 3.77 (1H, s, OCH3),
6.83 (1H, t, J: 7.25 Hz, pip-phenyl-C4), 6.97 (4H, m, th-phe-
nyl C3,5, pip-phenyl C2,6), 7.12 (1H, s, thiazole C5), 7.24
(2H, t, J: 7.86 Hz, th-phenyl C2,6), 7.81 (2H, d, J: 8.73 Hz, th-
phenyl C3,5). 13C NMR (75 MHz, DMSO-d6) d (ppm): 40.01
(piperazine CH2), 48.36 (piperazine CH2), 55.60 (OCH3),
100.28, 114.43, 115.68, 116.54, 119.95, 127.52, 129.48, 151.25
(thiazole C4), 170.00 (thiazole C2). For C20H21N3OS calcd; %
C, 68.35; H, 6.02; N, 11.96. Found; C, 68.36; H, 6.03; N,
11.98. HRMS (m/z) M þ H: calcd 352.1478; found: 352.1467.
4-(4-Chlorophenyl)-2-(4-phenylpiperazine-1-yl)
thiazole
(compound 4i). Yield 84%, mp; 157–159 ꢁC. IR ῡmax (cmꢀ1):
2830–3090 (C–H), 1336–1609 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.29 (4H, m, piperazine
C
2,6), 3.61 (4H, t, J: 4.93 Hz, piperazine C3,5), 6.83 (1H, t, J:
7.25 Hz, pip-phenyl-C4), 7.00 (2H, t, J: 8.01 Hz, pip-phenyl
2,6), 7.25 (2H, t, J: 8.07 Hz, pip-phenyl C3,5), 7.37 (1H, s,
thiazole C5), 7.44 (2H, td, J: 8.65 Hz, J:1.97 Hz, th-phenyl
3,5), 7.89 (2H, td, J: 8.63 Hz, J:1.96 Hz, th-phenyl C2,6). 13C
C
C
NMR (75 MHz, DMSO-d6) d (ppm): 40.29 (piperazine CH2),
48.34 (piperazine CH2), 104.05, 116.56, 119.97, 127.88,
129.00, 129.49, 132.41, 133.94, 149.83, 151.23 (thiazole C4),
170.95 (thiazole C2). For C19H18ClN3S calcd; % C, 64.12; H,
5.10; N, 11.81. Found; C, 64.13; H, 5.11; N, 11.84. HRMS
(m/z) M þ H: calcd 356.0983; found: 356.0974.
4-(3-Fluorophenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com-
pound 4f). Yield 75%, mp; 84–86 ꢁC. IR ῡ max (cmꢀ1):
2840–3129 (C–H), 1336–1599 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.30 (4H, m, piperazine
C
2,6), 3.62 (4H, t, J: 4.78 Hz, piperazine C3,5), 6.83 (1H, t, J:
7.23 Hz, pip-phenyl-C4), 7.00 (2H, d, J: 8.03 Hz, pip-phenyl
2,6), 7.11 (1H, td, J:8.60 Hz, J: 2.46 Hz, th-phenyl C4), 7.25
C
(2H, t, J: 7.80 Hz, pip-phenyl C3,5), 7.39–7.47 (2H, m, thia- 4-(3-Nitrophenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com-
zole C5), 7.65 (1H, d, J: 11.01 Hz, th-phenyl C5), 7.73 (1H, pound 4j). Yield 76%, mp; 142–145 ꢁC. IR ῡmax (cmꢀ1):
d, J: 7.84 Hz, th-phenyl C6). 13C NMR (75 MHz, DMSO-d6) 2774–3109 (C–H), 1306–1537 (C ¼ C, C ¼ N). 1H NMR
d (ppm): 40.28 (piperazine CH2), 48.34 (piperazine CH2), (300 MHz, DMSO-d6) d (ppm): 3.32 (4H, m, piperazine
104.74, 114.51, 114.79, 116.56, 119.97, 122.19, 129.49, C2,6), 3.65 (4H, t, J: 5.04 Hz piperazine C3,5), 6.86 (1H, t, J:
130.92, 131.04, 137.54, 151.23, 164.57 (thiazole C4), 170.87 7.25 Hz, pip-phenyl-C4), 7.03 (2H, d, J: 8.00 Hz, pip-phenyl
(thiazole C2). For C19H18FN3S calcd; % C, 67.23; H, 5.35; N,
C2,6), 7.26 (2H, t, J: 8.00 Hz, pip-phenyl C3,5), 7.63 (1H, s,

8
Z. SAHIN ET AL.
thiazole C5), 7.68 (1H, t, J: 8.00 Hz, th-phenyl C5), 8.14 (1H, 69.01; H, 6.34; N, 11.50. Found; C, 69.02; H, 6.32; N, 11.48.
HRMS (m/z) M þ H: calcd 366.1635; found: 366.1638.
d, J: 8.08 Hz, th-phenyl C6), 8.31 (1H, dt, J: 8.16 Hz, J:
1.60 Hz, th-phenyl C4), 8.65 (1H, t, J: 2.0 Hz, th-phenyl C2).
13C NMR (75 MHz, DMSO-d6) d (ppm): 40.28 (piperazine 4-(4-Methylphenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)th-
iazole (compound 4n). Yield 68%, mp; 201–204 ꢁC. IR ῡmax
CH2), 48.30 (piperazine CH2), 106.00, 116.76, 120.38,
120.44, 122.56, 129.54, 130.62, 132.28, 136.59, 148.64,
148.74, 150.86 (thiazole C4), 171.09 (thiazole C2). For
C19H18N4O2S calcd; % C, 62.28; H, 4.95; N, 15.29. Found;
C, 62.31; H, 4.94; N, 15.26. HRMS (m/z) M þ H: calcd
367.1223; found: 367.1219.
(cmꢀ1): 2650–3100 (C–H), 1307–1609 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 2.31 (3H, s, CH3),
3.28 (2H, brs, piperazine C2), 3.32 (2H, brs, piperazine
C6), 3.62 (2H, brs, piperazine C3), 3.73 (2H, brs, pipera-
zine C5), 3.86 (3H, s, OCH3), 6.92–7.26 (7H, m, Ar), 7.76
(2H, t, J: 6.63 Hz, th-phenyl C2,6). 13C NMR (75 MHz,
DMSO-d6) d (ppm): 21.33 (CH3), 40.28 (piperazine CH2),
47.99 (piperazine CH2), 56.26 (OCH3), 102.75, 119.95,
121.48, 125.50, 129.25, 131.93, 137.53, 138.15, 148.46,
150.39, 152.32, 168.64 (thiazole C4), 170.49 (thiazole C2).
For C21H23N3OS calcd; % C, 69.01; H, 6.34; N, 11.50.
Found; C, 69.00; H, 6.35; N, 11.49. HRMS (m/z) M þ H:
calcd 366.1635; found: 366.1634.
4-(4-Nitrophenyl)-2-(4-phenylpiperazine-1-yl)thiazole (com-
pound 4k). Verim 77%, ed; 158–161 ꢁC. IR ῡmax (cmꢀ1):
2777–3135 (C–H), 1315–1599 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.29 (4H, m, piperazine
C2,6), 3.63 (4H, t, J: 5.04 Hz, piperazine C3,5), 6.83 (1H, t, J:
7.25 Hz, pip-phenyl-C4), 7.00 (2H, d, J: 8.15 Hz, pip-phenyl
C2,6), 7.25 (2H, t, J: 7.95 Hz, pip-phenyl C3,5), 7.68 (1H, s,
thiazole C5), 8.12 (2H, d, J: 8.96 Hz, th-phenyl C2,6), 8.24
(2H, d, J: 9.00 Hz, th-phenyl C3,5). 13C NMR (75 MHz,
DMSO-d6) d (ppm): 40.29 (piperazine CH2), 48.32 (pipera-
zine CH2), 108.02, 116.56, 120.00, 124.48, 126.97, 129.49,
141.05, 146.71, 148.99, 151.19 (thiazole C4), 171.10 (thiazole
C2). For C19H18N4O2S calcd; % C, 62.28; H, 4.95; N, 15.29.
Found; C, 62.30; H, 4.92; N, 15.28. HRMS (m/z) M þ H:
calcd 367.1223; found: 367.1233.
4-(3-Methoxyphenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)
thiazole (compound 4o). Yield 69%, mp; 89–91 ꢁC. IR ῡmax
(cmꢀ1): 2835–3100 (C–H), 1314–1589 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 3.25 (4H, brs, pipera-
zine C2,6), 3.75 (4H, brs, piperazine C3,5), 3.80 (3H, s,
OCH3), 3.84 (3H, s, OCH3), 6.86–6.96 (2H, m, pip-phenyl
C4,6), 7.07 (3H, m, Ar), 7.31 (1H, t, J: 7.85 Hz, th-phenyl
C2), 7.35 (1H, s, thiazole C5), 7.45 (2H, t, J: 8.73 Hz, th-phe-
nyl C5,6). 13C NMR (75 MHz, DMSO-d6) d (ppm): 40.28
(piperazine CH2), 48.36 (piperazine CH2), 55.58 (OCH3),
56.07 (OCH3), 103.78, 111.77, 112.67, 113.66, 118.66, 119.37,
121.40, 124.76, 130.05, 136.28, 150.65, 152.39, 159.95 (thia-
zole C4), 170.60 (thiazole C2). For C21H23N3O2S calcd; % C,
66.12; H, 6.08; N, 11.01. Found; C, 66.10; H, 6.38; N, 11.03.
HRMS (m/z) M þ H: calcd 382.1584; found: 382.1578.
4-Phenyl-2-(4-(2-methoxyphenyl)piperazine-1-yl)thiazole (com-
pound 4l). Verim 70%, ed; 156–158 ꢁC. IR ῡmax (cmꢀ1):
2780–3073 (C–H), 1363–1576 (C ¼ C, C ¼ N). 1H NMR
(300 MHz, DMSO-d6) d (ppm): 3.42 (4H, brs, piperazine
C2,6), 3.79 (4H, brs, piperazine C3,5), 3.88 (3H, s, OCH3)
7.01 (1H, t, J: 7.36 Hz, pip-phenyl-C4), 7.13 (1H, d, J:
8.21 Hz pip-phenyl C6), 7.23 (1H, t, J: 7.36 Hz, pip-phenyl
C5), 7.28–7.45 (6H, m, pip-phenyl C2,3, thiazole C5, th-phe-
nyl C3,4,5), 7.87 (2H, d, J: 7.08 Hz, th-phenyl C2,6). 13C NMR
(75 MHz, DMSO-d6) d (ppm): 40.28 (piperazine CH2), 48.44
(piperazine CH2), 56.44 (OCH3), 103.86, 113.16, 120.17,
126.30, 128.74, 129.03, 133.89, 134.53, 136.16, 150.30,
152.26, 168.66 (thiazole C4), 170.45 (thiazole C2). For
C20H21N3OS calcd; % C, 68.35; H, 6.02; N, 11.96. Found; C,
68.31; H, 6.02; N, 11.98. HRMS (m/z) M þ H: calcd
352.1478; found: 352.1479.
4-(4-Methoxyphenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)
thiazole (compound 4p). Yield 70%, mp; 217–219 ꢁC. IR ῡmax
(cmꢀ1): 2770–3090 (C–H), 1359–1573 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 3.34 (4H, brs, pipera-
zine C2,6), 3.73 (4H, brs, piperazine C3,5), 3.78 (3H, s,
OCH3), 3.87 (3H, s, OCH3), 6.93–7.25 (7H, m, Ar), 7.79
(2H, d, J: 9.05 Hz, th-phenyl C2,6). 13CNMR (75 MHz,
DMSO-d6) d (ppm): 40.28 (piperazine CH2), 48.01 (pipera-
zine CH2), 55.66 (OCH3), 56.26 (OCH3), 101.60, 114.40,
119.97, 121.49, 126.14, 126.43, 127.27, 127.68, 129.83,
149.93, 152.30, 159.48 (thiazole C4) 170.45 (thiazole C2). For
C21H23N3O2S calcd; % C, 66.12; H, 6.08; N, 11.01. Found;
C, 66.10; H, 6.08; N, 11.00. HRMS (m/z) M þ H: calcd
382.1584; found: 382.1567.
4-(3-Methylphenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)th-
iazole (compound 4m). Yield 67%, mp; 187–190 ꢁC. IR ῡmax
(cmꢀ1): 2780–3083 (C–H), 1366–1589 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 2.35 (3H, s, CH3),
3.27 (2H, brs, piperazine C2), 3.35 (2H, brs, piperazine C6),
3.65 (2H, brs, piperazine C3), 3.69 (2H, brs, piperazine C5),
3.86 (3H, s, OCH3), 6.97 (1H, m, pip-phenyl-C4), 7.05–7.35
(6H, m, Ar), 7.67 (2H, t, J: 6.91 Hz, th-phenyl C2,6). 13C
NMR (75 MHz, DMSO-d6) d (ppm): 21.55 (CH3), 40.28
(piperazine CH2), 47.92 (piperazine CH2), 56.26 (OCH3),
103.52, 112.84, 112.97, 119.79, 126.23, 128.74, 129.36,
133.87, 134.53, 138.09, 148.60, 150.46, 152.31, 168.68 (thia-
4-(3-Fluorophenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)th-
iazole (compound 4q). Yield 71%, mp; 181–183 ꢁC. IR ῡmax
(cmꢀ1): 2800–3100 (C–H), 1366–1609 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 3.21 (4H, m, pipera-
zine C2,6), 3.61 (4H, m, piperazine C3,5), 3.90 (3H, s,
OCH3), 6.70–7.23 (5H, m, Ar), 7.45–7.76 (2H, m, Ar),
7.67–7.74 (3H, m, Ar). 13C NMR (75 MHz, DMSO-d6) d
zole C4), 170.46 (thiazole C2). For C21H23N3OS calcd; % C, (ppm): 40.27 (piperazine CH2), 47.68 (piperazine CH2),

PHOSPHORUS, SULFUR, AND SILICON AND THE RELATED ELEMENTS
9
56.34 (OCH3), 105.15, 112.62, 112.89, 113.04, 114.61, 115.70, phenyl C5), 8.14 (1H, d, J: 8.12 Hz, th-phenyl C6), 8.31 (1H, d, J:
119.91, 121.52, 125.94, 130.94, 136.07, 149.50, 152.27, 160.74 8.12 Hz, th-phenyl C4), 8.65 (1H, s, th-phenyl C2). 13C NMR
(thiazole C4), 170.53 (thiazole C2). For C20H20FN3OS calcd; (75MHz, DMSO-d6) d (ppm): 40.28 (piperazine CH2), 48.71
% C, 65.02; H, 5.46; N, 11.37. Found; C, 65.01; H, 5.48; N, (piperazine CH2), 55.91 (OCH3), 105.87, 112.46, 118.90, 120.44,
11.40. HRMS (m/z) M þ H: calcd 370.1384; found: 370.1370. 121.32, 122.55, 123.69, 130.62, 132.28, 136.64, 148.65, 152.48
(thiazole C4), 171.19 (thiazole C2). For C20H20N4O3S calcd; %
C, 60.59; H, 5.08; N, 14.13. Found; C, 60.55; H, 5.07; N, 14.16.
HRMS (m/z) M þ H: calcd 397.1329; found: 397.1332.
4-(4-Fluorophenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl) thia-
zole (compound 4r). Yield 70%, mp; 177–178.5 ꢁC. IR ῡmax
(cmꢀ1): 2774–3170 (C–H), 1350–1576 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 3.37 (4H, brs, pipera-
zine C2,6), 3.76 (4H, brs, piperazine C3,5), 3.88 (3H, s,
OCH3), 7.00 (1H, t, J: 7.70 Hz, pip-phenyl C4), 7.12 (1H, d,
J: 8.34 Hz, pip-phenyl C6), 7.18–7.30 (4H, m, Ar), 7.33 (1H,
s, thiazole C5), 7.91 (2H, d, J: 8.66 Hz, th-phenyl C3,5). 13C
NMR (75 MHz, DMSO-d6) d (ppm): 40.27 (piperazine CH2),
48.67 (piperazine CH2), 56.33 (OCH3), 103.46, 112.89,
115.71, 121.52, 126.56, 128.21, 130.56, 136.80, 149.59,
152.28, 160.55, 163.79 (thiazole C4), 170.60 (thiazole C2).
For C20H20FN3OS calcd; % C, 65.02; H, 5.46; N, 11.37.
Found; C, 65.00; H, 5.45; N, 11.36. HRMS (m/z) M þ H:
calcd 370.1384; found: 370.1374.
4-(4-Nitrophenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)th-
iazole (compound 4v). Yield 65%, mp; 147–149 ꢁC. IR ῡmax
(cmꢀ1): 2700–3145 (C–H), 1337–1596 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 3.10 (4H, t, J: 4.87 Hz,
piperazine C2,6), 3.62 (4H, t, piperazine C3,5), 3.80 (3H, s,
OCH3), 6.88–7.00 (4H, m, Ar), 7.66 (1H, s, thiazole C5),
8.12 (2H, d, J: 9.20 Hz, th-phenyl C2,6), 8.25 (2H, d, J:
9.20 Hz, th-phenyl C3,5). 13C NMR (75 MHz, DMSO-d6) d
(ppm): 40.28 (piperazine CH2), 48.77 (piperazine CH2),
55.87 (OCH3), 107.90, 112.42, 118.82, 123.51, 124.45, 126.95,
141.08, 141.12 146.68, 148.98, 152.49 (thiazole C4), 171.15
(thiazole C2). For C20H20N4O3S calcd; % C, 60.59; H, 5.08;
N, 14.13. Found; C, 60.57; H, 5.08; N, 14.14. HRMS (m/z)
M þ H: calcd 397.1329; found: 397.1335.
4-(3-Chlorophenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)th-
iazole (compound 4s). Yield 64%, mp; 88–90 ꢁC. IR ῡ max
(cmꢀ1): 2856–3099 (C–H), 1360–1592 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 3.10 (4H, t, J: 4.81 Hz,
piperazine C2,6), 3.59 (4H, t, J: 4.81 Hz, piperazine C3,5), 3.80
(3H, s, OCH3), 6.86–7.00 (4H, m, Ar, thiazole C5), 7.32–7.45
(3H, m, Ar), 7.84 (1H, dt, J: 7.70Hz, J: 1.29Hz, th-phenyl C6),
7.92 (1H, s, th-phenyl C3). 13C NMR (75MHz, DMSO-d6) d
(ppm): 40.28 (piperazine CH2), 48.79 (piperazine CH2), 55.86
(OCH3), 104.69, 112.41, 118.79, 121.30, 123.44, 124.67 125.81,
127.71, 130.88, 133.88, 137.14 141.22, 149.45, 152.50 (thiazole
C4), 170.99 (thiazole C2). For C20H20ClN3OS calcd; % C, 62.25;
H, 5.22; N, 10.89. Found; C, 62.22; H, 5.25; N, 10.91. HRMS (m/
z) M þ H: calcd 386.1088; found: 386.1079.
Enzyme assay
AChE inhibition experiments were performed in accordance
with our previous studies,[26] which followed Ellman method
with minor modifications.[27] During the experiments, the
following instructions provided by the “EliteTM
Acetylcholinesterase Assay Kit (Green Fluorescence)”
(MyBiosource, Catalog number: 433569) were followed:
acetylcholinesterase enzyme solutions were prepared in dou-
ble-distilled H2O with 0.1% bovine serum albumin (BSA) at
a concentration of 10 mU/mL. The acetylthiocholine reaction
mixture (1:2, 5:500 [v/v] acetylthiocholine stock solution:
EliteTM Green stock solution: assay buffer, 50 lL) and the
compound solution (50 lL) prepared in 1% DMSO at a con-
centration ranged between10ꢀ4 and 10ꢀ9 M were added to
each well and incubated at 25 ꢁC for 20 min. For the blank
control group, only the solvent was added instead of the
sample solution and the same protocol was applied. After
the incubation period, fluorescence was measured at 490 nm
(excitation) and 520 nm (emission) using a fluorescence
plate reader. All experiments were performed in triplicate.
The inhibition rate (%) was calculated as the following equa-
tion I (%) ¼ 100 ꢀ (Abssample ꢀ Abscontrol) ꢂ 100. The inhib-
ition rate of the compounds and IC50 values were recorded
using GraphPad Prism software, version 7.02 (La Jolla,
California). The data were expressed as mean standard
deviation (SD).[28]
4-(4-Chlorophenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)th-
iazole (compound 4t). Yield 67%, mp; 104–107 ꢁC. IR ῡmax
(cmꢀ1): 2751–3079 (C–H), 1366–1564 (C ¼ C, C ¼ N). 1H
NMR (300 MHz, DMSO-d6) d (ppm): 3.21 (4H, brs, pipera-
zine C2,6), 3.66 (4H, brs, piperazine C3,5), 3.83 (3H, s,
OCH3), 6.91–7.10 (4H, m, Ar), 7.38 (1H, s, thiazole C5),
7.45 (2H, d, J: 8.93 Hz, th-phenyl C3,5), 7.89 (2H, d, J:
8.39 Hz, th-phenyl C2,6). 13C NMR (75 MHz, DMSO-d6) d
(ppm): 40.28 (piperazine CH2), 48.35 (piperazine CH2),
55.07 (OCH3), 104.14, 112.66, 119.36, 121.39, 124.70, 127.89,
129.00, 132.46, 133.85, 149.69, 152.39 (thiazole C4), 170.85
(thiazole C2). For C20H20ClN3OS calcd; % C, 62.25; H, 5.22;
N, 10.89. Found; C, 62.24; H, 5.23; N, 10.90. HRMS (m/z)
M þ H: calcd 386.1088; found: 386.1100.
4-(3-Nitrophenyl)-2-(4-(2-methoxyphenyl)piperazine-1-yl)th-
Molecular modeling
iazole (compound 4u). Yield 69%, mp; 164–166 ꢁC. IR ῡmax
1
(cmꢀ1): 2800–3110 (C–H), 1336–1615.9 (C¼ C, C ¼ N). H The modeling studies were performed by reported specifica-
NMR (300 MHz, DMSO-d6) d (ppm): 3.14 (4H, s, piperazine tions in the predecessor study.[26] The structures of the com-
C2,6), 3.64 (4H, s, piperazine C3,5), 3.81 (3H, s, OCH3), 6.96 pounds were obtained from the LigPrep module in the
€
(4H, m, Ar), 7.63 (1H, s, thiazole C5), 7.69 (1H, t, J: 8.12Hz, th- Maestro Suite (Scrodinger LCC, Oregon) and minimized

10
Z. SAHIN ET AL.
[3] Commins, S.; Kirby, B. The Complexities of Behavioural
Assessment in Neurodegenerative Disorders: Focus on
using OPLS 2005 forcefield. The X-ray structure of human
acetylcholinesterase (PDB ID: 4EY7) was used in the ana-
lysis.[9,26] PDB file was edited using the protein preparation
module of Maestro. GRID file was prepared using Maestro
Glide-Receptor Grid Generation Tool. Ligand docking was
performed with the Glide Module in the extra precision
(XP) mode. Poses of the most active compounds were sub-
jected to induced fit protocols with default parameter set-
tings and visualized in the Molecular Operating
Environment (MOE) program.
A
Alzheimer’s Disease. Pharmacol. Res. 2019, 147, 104363. DOI:
10.1016/j.phrs.2019.104363.
[4] Galimberti, D.; Ghezzi, L.; Scarpini, E. Immunotherapy Against
Amyloid Pathology in Alzheimer’s Disease. J. Neur. Sci. 2013,
333, 50–54. DOI: 10.1016/j.jns.2012.12.013.
[5] Mishra, P.; Kumar, A.; Panda, G. Anti-Cholinesterase Hybrids
as Multi-Target-Directed Ligands Against Alzheimer’s Disease
(1998–2018). Bioorg. Med. Chem. 2019, 27, 895–930. DOI: 10.
1016/j.bmc.2019.01.025.
[6] Anand, A.; Patience, A. A.; Sharma, N.; Khurana, N. The
Present and Future of Pharmacotherapy of Alzheimer’s Disease:
A
Comprehensive Review. Eur. J. Pharmacol. 2014, 815,
364–375. DOI: 10.1016/j.ejphar.2017.09.043.
Conclusion
[7] Anand, P.; Singh, B.; Singh, N. A Review on Coumarins as
Acetylcholinesterase Inhibitors for Alzheimer’s Disease. Bioorg.
Med. Chem. 2012, 20, 1175–1180. DOI: 10.1016/j.bmc.2011.12.042.
[8] Magnuson, T. M.; Keller, B. K.; Burke, W. J. Extrapyramidal
Donepezil is a well-known inhibitor of acetylcholinesterase
owing to its potent IC50 value as high as 0.07mM. This high
IC50 value is consistent with the previously conducted studies,
which manifests that the enzymatic results are accurate. In the
past, our group reported benzoyl attached-thiazole structures
with a similar scaffold, which exhibited IC50 value ranged
between 0.29 and 2.10 mM.[27,28] In addition to the considerable
binding and inhibition profile of 22 molecules reported in this
study, a carbonyl group positioned toward Arg 296 (likewise
the indenone ring on donepezil and benzoyl on predecessor
compounds) seemed to be an important SAR feature of done-
pezil analogs. Hence, hydrogen bond interaction with Arg296
or Phe295 linked amino acid’s NH group stands for a signifi-
cant characteristic. An aromatic site, linker and a basic center
(Figure 1) provided a considerable binding, even in the absence
of carbonyl. This scaffold can be developed through retaining a
carbonyl group between a linker and an aromatic site. The
linker might be modified by different five-membered hetero-
cycles or alkyl groups as well, whereas the basic center might
be modified by alternatives, which altogether contribute design-
ing new donepezil analogs. Additional molecules should be
developed in order to propose more potent inhibitors with an
activity similar to donepezil.
Side Effects in
a Patient Treated with Risperidone plus
Donepezil. Am. J. Psychiatry 1998, 155, 1458–1459. DOI: 10.
1176/ajp.155.10.1458a.
[9] Cheung, J.; Rudolph, M. J.; Burshteyn, F.; Cassidy, M. S.; Gary,
E. N.; Love, J.; Height, J. J. Structures of Human
Acetylcholinesterase in Complex with Pharmacologically
Important Ligands. J. Med. Chem. 2012, 55, 10282–10286. DOI:
10.1021/jm300871x.
[10] Sugimoto, H.; Iimura, Y.; Yamanishi, Y.; Yamatsu, K. Synthesis
and Structure–Activity Relationships of Acetylcholinesterase
Inhibitors:
1-Benzyl-4-[(5,6-Dimethoxy-1-Oxoindan-2-
yl)Methyl]Piperidine Hydrochloride and Related Compounds. J.
Med. Chem. 1995, 38, 4821–4829. DOI: 10.1021/jm00024a009.
[11] Sakya, S.; Karki, K. Donepezil, Rivastigmine and Galantamine:
Cholinesterase Inhibitors for Alzheimer’s Disease. In Modern
Drug Synth.; Li, J. J., Johnson, D. S., Eds.; John Wiley & Sons
Inc.: Hoboken, NJ, 2010; pp 249–274. DOI: 10.1002/
9780470768594.ch17.
€
ꢁ
[12] Saglık, B. N.; Ilgın, S.; Ozkay, Y. Synthesis of New Donepezil
Analogues and Investigation of Their Effects on Cholinesterase
Enzymes. Eur. J. Med. Chem. 2016, 124, 1026–1040. DOI: 10.
1016/j.ejmech.2016.10.042.
[13] Andreani, A.; Burnelli, S.; Granaiola, M.; Guardigli, M.; Leoni,
A.; Locatelli, A.; Varoli, L. Chemiluminescent High-Throughput
Microassay Applied to Imidazo[2,1-b]Thiazole Derivatives as
Potential
Acetylcholinesterase
and
Butyrylcholinesterase
Inhibitors. Eur. J. Med. Chem. 2008, 43, 657–661. DOI: 10.
1016/j.ejmech.2007.04.018.
Consent for publication
[14] Channar, P. A.; Shah, M. S.; Saeed, A.; Khan, S. U.; Larik,
F. A.; Shabir, G.; Iqbal, J. Synthesis, Characterization and
Cholinesterase Inhibition Studies of New Arylidene
Aminothiazolylethanone Derivatives. Med. Chem. 2017, 13,
648–653. DOI: 10.2174/1573406413666170306113347.
[15] Mumtaz, A.; Shoaib, M.; Zaib, S.; Shah, M. S.; Bhatti, H. A.;
Saeed, A.; Hussain, I.; Iqbal, J. Synthesis, Molecular Modelling
and Biological Evaluation of Tetrasubstituted Thiazoles
Towards Cholinesterase Enzymes and Cytotoxicity Studies.
Bioorg. Chem. 2018, 78, 141–148. DOI: 10.1016/j.bioorg.2018.
02.024.
Not applicable.
Conflict of interest
The authors declare that no conflict of interest exists.
ORCID
Zafer Sahin
Sevde Nur Biltekin
http://orcid.org/0000-0002-5976-676X
http://orcid.org/0000-0003-1896-2729
€
[16] Yurtta¸s, L.; Kaplancıklı, Z. A.; Ozkay, Y. Design, Synthesis and
Evaluation of New Thiazole-Piperazines as Acetylcholinesterase
Inhibitors. J. Enzyme Inhib. Med. Chem. 2012, 28, 1040–1047.
DOI: 10.3109/14756366.2012.709242.
Leyla Yurttas
Barkin Berk
S¸eref Demirayak
http://orcid.org/0000-0002-0957-6044
http://orcid.org/0000-0001-6047-2796
http://orcid.org/0000-0002-0841-1299
[17] Rahim, F.; Javed, M. T.; Ullah, H.; Wadood, A.; Taha, M.;
Ashraf, M.; Khan, M. A.; Khan, F.; Mirza, S.; Khan, K. M.
Synthesis, Molecular Docking, Acetylcholinesterase and
Butyrylcholinesterase Inhibitory Potential of Thiazole Analogs
as New Inhibitors for Alzheimer Disease. Bioorg. Chem. 2015,
62, 106–116. DOI: 10.1016/j.bioorg.2015.08.002.
References
[1] Ulep, M. G.; Saraon, S. K.; McLea, S. Alzheimer Disease. J. Nurse
Pract. 2018, 14, 129–135. DOI: 10.1016/j.nurpra.2017.10.014.
[2] 2018 Alzheimer’s Disease Facts and Figures. Alzheimers
Dement. 2018, 14, 367–429. DOI: 10.1016/j.jalz.2018.02.001.
[18] Kurt, B. Z.; Gazioglu, I.; Sonmez, F.; Kucukislamoglu, M.
Synthesis, Antioxidant and Anticholinesterase Activities of

PHOSPHORUS, SULFUR, AND SILICON AND THE RELATED ELEMENTS
11
Novel Coumarylthiazole Derivatives. Bioorg. Chem. 2015, 59,
80–90. DOI: 10.1016/j.bioorg.2015.02.002.
Antibacterial Activity of Substituted Thiosemicarbazides and of
1,3,4-Thiadiazole or 1,2,4-Triazole Derivatives. Phosphorus
Sulfur Silicon Relat. Elem. 2006, 181, 609–625. DOI: 10.1080/
10426500500269851.
[19] Varadaraju, K. R.; Kumar, J. R.; Mallesha, L.; Muruli, A.;
Mohana, K. N. S.; Mukunda, C. K.; Sharanaiah, U. Virtual
Screening and Biological Evaluation of Piperazine Derivatives as
Human Acetylcholinesterase Inhibitors. Int. J. Alzheimers Dis.
2013, 2013, 653962–653913. DOI: 10.1155/2013/653962.
[24] Cowper, R. M.; Davidson, L. H. Phenacyl Bromide
[Acetophenone, a-bromo-]. Org. Synth. Coll. 1943, 2, 480. DOI:
10.15227/orgsyn.019.0024.
[20] Bikshapathi, R.; Sai Prathima, P.; Yashwanth, B.; Rajesh, P.; Rao,
J. V.; Jagadeesh Kumar, G.; Jagadeesh, N.; Rao, V. J. An Expeditious
Protocol for Synthesis of Baylis–Hillman Derived Piperazine
Derivatives and Evaluation of Their AChE Inhibition. Res. Chem.
Intermed. 2018, 44, 553–565. DOI: 10.1007/s11164-017-3119-9.
[21] Shafferman, A.; Barak, D.; Kaplan, D.; Ordentlich, A.; Kronman,
C.; Velan, B. Functional Requirements for the Optimal Catalytic
Configuration of the AChE Active Center. Chem. Biol. Interact.
2005, 157–158, 123–131. DOI: 10.1016/j.cbi.2005.10.021.
[25] Hantzsch, A. R.; Weber, J. H. Uerbindungen Des
Thiazols(Pyridins Der Thiophenreihe). Ber. Dtsch. Chem. Ges.
1887, 20, 3118–3132. DOI: 10.1002/cber.188702002200.
[26] Sahin, Z.; Ertas, M.; Bender, C.; Bu€lbu€l, E. F.; Berk, B.; Biltekin,
S. N.; Yurtta¸s, L.; Demirayak, S¸. Thiazole-Substituted
Benzoylpiperazine Derivatives as Acetylcholinesterase Inhibitors.
Drug Dev. Res. 2018, 79, 406–425. DOI: 10.1002/ddr.21481.
[27] Pohanka, M.; Hrabinova, M.; Kuca, K.; Simonato, J.-P.
Assessment of Acetylcholinesterase Activity Using Indoxylacetate
and Comparison with the Standard Ellman’s Method. Int. J. Mol.
Sci. 2011, 12, 2631–2640. DOI: 10.3390/ijms12042631.
ꢀ
~
[22] Camps, P.; Formosa, X.; Galdeano, C.; Gomez, T.; Munoz-
Torrero, D.; Scarpellini, M.; Viayna, E.; Badia, A.; Clos, M. V.;
Camins, A.; et al. Novel Donepezil-Based Inhibitors of Acetyl-
and Butyrylcholinesterase and Acetylcholinesterase-Induced
b-Amyloid Aggregation. J. Med. Chem. 2008, 51, 3588–3598.
DOI: 10.1021/jm8001313.
[28] Demirayak, S¸.; S¸ahin, Z.; Erta¸s, M.; Bu€lbu€l, E. F.; Bender, C.;
ꢁ
Biltekin, S. N.; Berk, B.; Saglık, B. N.; Levent, S.; Yurtta¸s, L.
Novel Thiazole-Piperazine Derivatives as Potential
Cholinesterase Inhibitors. J. Heterocyclic Chem. 2019, 56,
3370–3386. DOI: 10.1002/jhet.3734.
ꢀ
[23] Spalinska, K.; Foks, H.; Kedzia, A.; Wierzbowska, M.; Kwapisz,
ꢀ
E.; Gebska, A.; Zilkołwska-Klinkosz, M. Synthesis and