In vitro and in silico evaluation of new thiazole compounds as monoamine oxidase inhibitors

Article abstract of DOI:10.1016/j.bioorg.2018.12.019

New twenty compounds bearing thiazole ring (3a-3t) were designed and synthesized as monoamine oxidase (MAO) inhibitors. The fluorometric enzyme inhibition assay was used to determine the biological effects of synthesized compounds. Most of them showed rem

Full text of DOI:10.1016/j.bioorg.2018.12.019

BioorganicChemistry85(2019)97–108
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
In vitro and in silico evaluation of new thiazole compounds as monoamine
oxidase inhibitors
⁎
Begüm Nurpelin Sağlıka,b, , Betül Kaya Çavuşoğlua, Derya Osmaniyea,b, Serkan Leventa,b
,
Ulviye Acar Çevika,b, Sinem Ilgınc, Yusuf Özkaya,b, Zafer Asım Kaplancıklıa, Yusuf Öztürkd
a Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Anadolu University, Eskişehir, Turkey
b Doping and Narcotic Compounds Analysis Laboratory, Faculty of Pharmacy, Anadolu University, Eskişehir, Turkey
c Department of Pharmaceutical Toxicology, Faculty of Pharmacy, Anadolu University, Eskişehir, Turkey
d Department of Pharmacology, Faculty of Pharmacy, Anadolu University, Eskişehir, Turkey
A R T I C L E I N F O
A B S T R A C T
Keywords:
New twenty compounds bearing thiazole ring (3a-3t) were designed and synthesized as monoamine oxidase
(MAO) inhibitors. The fluorometric enzyme inhibition assay was used to determine the biological effects of
synthesized compounds. Most of them showed remarkable inhibitory activity against both MAO-A and MAO-B.
By comparing their IC50 values, it can be seen that active derivatives displayed generally selectivity on MAO-B
enzyme. Compounds 3j and 3t, which bear dihydroxy moiety at the 3rd and 4th position of phenyl ring, were the
most active derivatives in the series against both isoenzymes. Compounds 3j and 3t showed significant inhibition
Thiazole derivatives
MAO inhibitors
Enzyme inhibitor activity
Molecular docking
profile on MAO-A with the IC50 values of 0.134
0.004 µM and 0.123
0.005 µM, respectively, while they
0.001 µM and 0.025 0.001 µM, re-
performed selectivity against MAO-B with the IC50 values of 0.027
spectively. Also, docking studies about these compounds were carried out to evaluate their binding modes on the
active regions of MAO-A and MAO-B.
1. Introduction
a person has got. However, some disorders in the nervous system as
depression, Parkinson’s disease (PD) and Alzheimer’s disease (AD) may
Monoamine Oxidases (MAOs) are flavin adenine dinucleotide (FAD)
based enzymes being responsible for the catalysis of oxidative deami-
nation of biological amines in both peripheral tissues and central ner-
vous system (CNS). Two different types of MAOs are known as MAO-A
and MAO-B. Both of them are placed to the mitochondrial membrane in
neural, glial and some other tissues as integral proteins [1]. MAO-A
consists of 527 amino acids, while MAO-B contains 520 amino acids.
Their amino acid sequences have 70% similarity. MAO-A crystallizes as
a monomer, whereas MAO-B exists as dimers. Specificities of MAO-A
and MAO-B cause minor differences in the active sites [2–4]. The vo-
lume of the MAO-A active site is ∼400 Å while the MAO-B active has
∼700 Å volume [4–6]. Although both of two isoenzymes have sub-
strate-binding cavity, their substrate selectivity is quite different. In-
hibition of norepinephrine and serotonin was carried out by MAO-A,
whereas inhibition of phenyl ethylamine and benzyl amine is carried
out by MAO-B. Both enzymes are able to oxidize dopamine and tyr-
amine. However, dopamine and tyramine are efficiently oxidized by
MAO-A and MAO-B, respectively [7–10].
affect the life of human. These disorders have been studied extensively
and numerous drugs have been improved up to day [1,5,11–13]. MAO
inhibitors are one of the promising choices in the treatment of these
disorders. The increasing functional and structural information about
MAO enzyme help to create better selective and reversible inhibitors.
Current studies are targeting MAO-B for the treatment of PD and AD via
generating the selective MAO-B inhibitors, while selective MAO-A in-
hibitors are important for the treatment of depression [14–16].
In the literature there are many thiazole compounds found to be
effective against MAO enzymes. Among them the compounds bearing
arylidene-hydrazinyl-thiazole substructure were reported to inhibit
MAO enzymes, significantly [17–19] (Fig. 1). The SARs of previously
reported hydrazinyl-thiazole compounds have clearly suggested the
required structural features for a compound to possess enzyme in-
hibitory activity [20–27]. Prompted from the SARs data of these studies
we recently performed a study including the MAO inhibitory activity of
some benzylidene-hydrazinyl-thiazoles. [28]. Results of such study di-
rected us to design and synthesize similar compounds for enhancing the
MAO inhibitory activity. Hence, in the present study new benzylidene-
A healthy nervous system is one of the most valuable properties that
⁎ Corresponding author at: Anadolu University, Faculty of Pharmacy, Department of Pharmaceutical Chemistry, 26470 Eskişehir, Turkey.
E-mail address: bnsaglik@anadolu.edu.tr (B.N. Sağlık).
https://doi.org/10.1016/j.bioorg.2018.12.019
Received 24 October 2018; Received in revised form 11 December 2018; Accepted 12 December 2018
Availableonline13December2018
0045-2068/©2018ElsevierInc.Allrightsreserved.

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
1132 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.92 (3H, d,
J = 6.5 Hz, CH3), 1.12–1.25 (2H, m, piperidine protons), 1.49–1.58
(1H, m, piperidine proton), 1.67 (2H, d, J = 12.7 Hz, piperidine
protons), 2.72 (2H, d, J = 11.4 Hz, piperidine protons), 3.77 (2H, d,
J = 12.6 Hz, piperidine protons), 6.95 (2H, d, J = 8.9 Hz, aromatic
protons), 7.26 (1H, s, aromatic proton), 7.30 (1H, d, J = 7.2 Hz,
aromatic proton), 7.40 (2H, t, J = 7.7 Hz, aromatic protons), 7.47
(2H, d, J = 8.8 Hz, aromatic protons), 7.85 (2H, d, J = 7.2 Hz, aromatic
protons), 7.91 (1H, s, CH]N), 11.89 (1H, s, NH). 13C NMR (75 MHz,
DMSO-d6, ppm) δ 22.23 (CH3), 30.70, 33.71, 48.40, 103.52, 115.36,
124.21, 125.95, 127.92, 129.03, 135.26, 142.37, 152.16, 168.84.
HRMS (m/z): [M+H]+ calcd for C22H24N4S: 377.1794; found
377.1782.
Fig. 1. General chemical structure of 2-[2-(Arylidene)hydrazinyl]-4-sub-
stitutedphenylthiazoles as MAO inhibitors. Ar: phenyl, pyridin-2-yl, pyridin-3-
yl, pyridin-4-yl, naphthalen-1-yl, naphthalen-2-yl R: H, CH3 Ŕ: 4-F, 4-CN were
reported in previous studies [17–27]; Ar: 4-(Morpholin-4-yl)/(pyrrolidin-1-yl)
phenyl R:H, Ŕ: H, 4-CH3, 4-CN, 4-Br, 4-CI, 4-F, 2,4-DiCI, 2,4-DiF were reported
in our recent study [28]; Ar: 4-(3- or 4-methylpiperidin-1-yl)phenyl R: H, Ŕ: H,
4-CH3, 4-Br, 4-CI, 4-F, 4-NO2, 4-CN, 4-OH, 2,4-DiF, 3,4-DiOH were synthesized
in this study.
2.1.3.2. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
methylphenyl)thiazole (3b). FTIR (ATR, cm−1): ʋmax 3296 (NeH), 3091
(aromatic CeH), 2918 (aliphatic CeH), 1600–1516 (C]N and C]C),
1224 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.92 (3H, d,
J = 6.5 Hz, CH3), 1.12–1.27 (2H, m, piperidine protons), 1.51–1.62
(1H, m, piperidine proton), 1.69 (2H, d, J = 12.6 Hz, piperidine
protons), 2.31 (3H, s, CH3), 2.73–2.93 (2H, m, piperidine protons),
3.76 (2H, d, J = 12.6 Hz, piperidine protons), 6.98–7.05 (2H, m,
aromatic protons), 7.18–7.21 (3H, m, aromatic protons), 7.48 (2H, d,
J = 8.7 Hz, aromatic protons), 7.73 (2H, d, J = 8.1 Hz, aromatic
protons), 7.91 (1H, s, CH]N), 11.87 (1H, s, NH). 13C NMR (75 MHz,
DMSO-d6, ppm) δ 21.26 (CH3), 22.18 (CH3), 30.54, 33.61, 47.48, 48.72,
102.65, 114.21, 115.63, 125.91, 127.92, 129.60, 132.59, 137.15,
142.18, 150.95, 154.12, 168.72. HRMS (m/z): [M+H]+ calcd for
hydrazinyl-thiazoles were synthesized and tested for their MAO in-
hibition potency both in in vitro and in silico assays.
2. Materials and methods
2.1. Chemistry
All chemicals were purchased from Sigma-Aldrich Chemical Co
(Sigma-Aldrich Corp., St. Louis, MO, USA) and Merck Chemicals (Merck
KGaA, Darmstadt, Germany). All melting points (m.p.) were de-
termined by MP90 digital melting point apparatus (Mettler Toledo,
Ohio, USA) and were uncorrected. All reactions were monitored by
thin-layer chromatography (TLC) using Silica Gel 60 F254 TLC plates
(Merck KGaA, Darmstadt, Germany). Spectroscopic data were recorded
with the following instruments: 1H NMR, Bruker DPX 300 NMR spec-
trometer; 13C NMR Bruker DPX 75 NMR spectrometer (Bruker
Bioscience, Billerica, MA, USA) in DMSO-d6, using TMS as internal
standard; The IR spectra were recorded by an IRAffinity-1S Fourier
transform IR (FTIR) spectrometer (Shimadzu, Tokyo, Japan). M+1
peaks were determined by Shimadzu LC/MS IT-TOF system (Shimadzu,
Tokyo, Japan).
C23H26N4S: 391.1951; found 391.1940.
2.1.3.3. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
bromophenyl)thiazole (3c). FTIR (ATR, cm−1): ʋmax 3387 (NeH), 3074
(aromatic CeH), 2949 (aliphatic CeH), 1602–1440 (C]N and C]C),
1049 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.97 (3H, d,
J = 6.5 Hz, CH3), 1.45–1.49 (1H, m, piperidine proton), 1.71–1.91
(4H, m, piperidine protons), 3.26 (2H, br s, piperidine protons), 3.67
(2H, d, J = 11.9 Hz, piperidine protons), 7.41–7.53 (3H, m, aromatic
protons), 7.60 (2H, d, J = 8.6 Hz, aromatic protons), 7.69–7.71 (2H, m,
aromatic protons), 7.80 (2H, d, J = 8.5 Hz, aromatic protons), 8.02
(1H, s, CH]N), 12.22 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ
21.70 (CH3), 28.86, 32.41, 47.72, 48.12, 105.05, 121.01, 128.02,
129.19, 129.87, 132.01, 134.25, 137.36, 142.67, 150.97, 154.94,
168.73. HRMS (m/z): [M + H]+ calcd for C22H23BrN4S: 455.0900;
found 455.0886.
2.1.1. General procedure for synthesis of 4-(3/4-methylpiperidin-1-yl)
benzaldehydes (1a, 1b)
To a solution of 3- or 4-methylpiperidine (3.4 g, 0.034 mol) in DMF
(10 mL) was added 4-fluorobenzaldehyde (4.3 g, 0.034 mol) and K2CO3
(4.7 g, 0.034 mol) then the reaction mixture stirred under reflux for
24 h. After cooling to the ambient temperature, the reaction mixture
was poured into ice-water. The obtained solid was filtered off, washed
with water and crystallized from ethanol to afford compounds 1a, 1b.
2.1.3.4. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
chlorophenyl)thiazole (3d). FTIR (ATR, cm−1): ʋmax 3298 (NeH), 3087
(aromatic CeH), 2918 (aliphatic CeH), 1602–1462 (C]N and C]C),
1226 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.92 (3H, d,
J = 6.5 Hz, CH3), 1.12–1.26 (2H, m, piperidine protons), 1.49–1.61
(1H, m, piperidine proton), 1.68 (2H, d, J = 12.4 Hz, piperidine
protons), 2.73 (2H, t, J = 10.5 Hz, piperidine protons), 3.78 (2H, d,
J = 12.7 Hz, piperidine protons), 6.96 (2H, d, J = 8.9 Hz, aromatic
protons), 7.34 (1H, s, aromatic proton), 7.44–7.48 (4H, m, aromatic
protons), 7.86 (2H, d, J = 8.6 Hz, aromatic protons), 7.91 (1H, s,
CH=N), 11.89 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ
22.23 (CH3), 30.69, 33.69, 48.41, 104.37, 115.38, 127.66, 127.96,
129.05, 132.27, 134.12, 142.55, 168.97. HRMS (m/z): [M+H]+ calcd
for C22H23ClN4S: 411.1405; found 411.1388.
2.1.2. General procedure for synthesis of 2-(4-(3- or 4-methylpiperidin-1-
yl)benzylidene)hydrazine-1-carbothioamides (2a, 2b)
A mixture of compounds 1a or 1b (5 g, 25 mmol), and thiosemi-
carbazide (2.3 g, 25 mmol) were refluxed in ethanol for 4 h. When the
reaction was completed as indicated by TLC, the reaction mixture was
cooled to attain room temperature. The precipitated product was fil-
tered and recrystallized from ethanol to give 2a, 2b.
2.1.3. General procedure for synthesis of 2-[2-(4-(3- or 4-methylpiperidin-
1-yl)benzylidene) hydrazinyl]-4-(substituted or disubstituted phenyl)
thiazole derivatives (3a-3 t)
The target compounds (3a-3 t) were synthesized via the reaction of
corresponding thiosemicarbazone (2a, 2b) (0.55 g, 2 mmol) and ap-
propriate phenacyl bromide derivatives (2 mmol) in ethanol (40 mL)
under reflux for 3 h. The mixture was cooled, precipitated product was
collected by filtration, dried and recrystallized from ethanol.
2.1.3.5. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
fluorophenyl)thiazole (3e). FTIR (ATR, cm−1): ʋmax 3298 (NeH), 3111
(aromatic CeH), 2918 (aliphatic CeH), 1600–1516 (C]N and C]C),
1222 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.92 (3H, d,
J = 6.5 Hz, CH3), 1.12–1.25 (2H, m, piperidine protons), 1.49–1.59
(1H, m, piperidine proton), 1.67 (2H, d, J = 12.6 Hz, piperidine
2.1.3.1. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-
phenylthiazole (3a). FTIR (ATR, cm−1): ʋmax 3292 (NeH), 3072
(aromatic CeH), 2924 (aliphatic CeH), 1600–1442 (C]N and C]C),
98

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
protons), 2.72 (2H, td, J1 = 2.2 Hz, J2 = 12.3 Hz, piperidine protons),
3.77 (2H, d, J = 12.7 Hz, piperidine protons), 6.95 (2H, d, J = 8.9 Hz,
aromatic protons), 7.20–7.25 (3H, m, aromatic protons), 7.46 (2H, d,
J = 8.9 Hz, aromatic protons), 7.85–7.91 (3H, m, aromatic protons,
CH]N), 11.88 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 22.23
(CH3), 30.70, 33.70, 48.39, 103.28, 115.36, 115.86 (d, J = 21 Hz),
124.18, 127.90 (d, J = 7.5 Hz), 131.92, 142.47, 152.17, 162.00 (d,
J = 242.3 Hz), 168.94. HRMS (m/z): [M + H]+ calcd for C22H23FN4S:
395.1700; found 395.1692.
protons), 7.14–7.20 (2H, m, aromatic protons), 7.30–7.38 (1H, m,
aromatic protons), 7.47 (2H, d, J = 8.8 Hz, aromatic protons), 7.92
(1H, s, CH]N), 7.98–8.06 (1H, m, aromatic protons), 11.91 (1H, s,
NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 22.22 (CH3), 30.68, 33.68,
48.41, 104.98, 107.59, 107.78, 112.14, 112.42, 115.38, 127.98,
130.83, 142.63, 152.14, 158.24, 161.74, 163.15, 168.31. HRMS (m/
z): [M+H]+ calcd for C22H22F2N4S: 413.1606; found 413.1604.
2.1.3.10. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-
(3,4-hydroxyphenyl)thiazole (3j). FTIR (ATR, cm−1): ʋmax 3263 (NeH),
3059 (aromatic CeH), 2974 (aliphatic CeH), 1620–1487 (C]N and
C]C), 1265–1082 (CeN and CeO). 1H NMR (300 Mhz, DMSO-d6, ppm)
δ 0.93 (3H, d, J = 5.6 Hz, CH3), 1.53–1.76 (5H, m, piperidine protons),
3.15 (1H, br s, piperidine proton), 3.50–3.60 (3H, m, piperidine
protons), 6.41–6.50 (1H, m, aromatic proton), 6.74 (2H, d,
J = 8.2 Hz, aromatic protons), 6.95–6.96 (1H, m, aromatic proton),
7.23–7.73 (5H, m, aromatic protons), 8.02 (1H, s, CH]N), 8.99 (2H, br
s, OH), 12.22 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 21.77
(CH3), 29.19, 32.32, 47.72, 106.73, 115.63, 116.39, 117.51, 119.53,
120.72, 124.73, 127.94, 128.12, 129.12, 142.93, 145.63, 145.98,
152.43, 169.49. HRMS (m/z): [M+H]+ calcd for C22H24N4O2S:
409.1693; found 409.1689.
2.1.3.6. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
nitrophenyl)thiazole (3f). FTIR (ATR, cm−1): ʋmax 3309 (NeH), 3034
(aromatic CeH), 2922 (aliphatic CeH), 1597–1566 (C]N and C]C),
1514–1336 (NO2), 1224 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ
0.92 (3H, d, J = 6.5 Hz, CH3), 1.16–1.27 (2H, m, piperidine protons),
1.52–1.58 (1H, m, piperidine proton), 1.70 (2H, d, J = 12.3 Hz,
piperidine protons), 2.79 (2H, br s, piperidine protons), 3.77 (2H, d,
J = 12.6 Hz, piperidine protons), 7.01 (2H, d, J = 6.2 Hz, aromatic
protons), 7.50 (2H, d, J = 8.5 Hz, aromatic protons), 7.67 (1H, s,
aromatic proton), 7.94 (1H, s, CH]N), 8.10 (2H, d, J = 9 Hz,
aromatic protons), 8.27 (2H, d, J = 9 Hz, aromatic protons), 12.04
(1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 22.16 (CH3), 30.49,
33.56, 48.82, 108.57, 115.62, 124.57, 126.76, 128.05, 141.23, 142.76,
146.61, 148.95, 169.22. HRMS (m/z): [M+H]+ calcd for C22H23N5O2S:
422.1645; found 422.1636.
2.1.3.11. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-
phenylthiazole (3k). FTIR (ATR, cm−1): ʋmax 3267 (NeH), 3091
(aromatic CeH), 2945 (aliphatic CeH), 1600–1442 (C]N and C]C),
1232 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.91 (3H, d,
J = 6.5 Hz, CH3), 1.00–1.13 (1H, m, piperidine proton), 1.46–1.69
(4H, m, piperidine protons), 2.40 (1H, t, J = 11.5 Hz, piperidine
proton), 2.69 (1H, t, J = 10.2 Hz, piperidine proton), 3.71 (2H, t,
J = 11.2 Hz, piperidine protons), 6.96 (2H, d, J = 8.6 Hz, aromatic
protons), 7.27–7.31 (2H, m, aromatic protons), 7.40 (2H, t, J = 7.7 Hz,
aromatic protons), 7.47 (2H, d, J = 8.8 Hz, aromatic protons), 7.85
(2H, d, J = 8.5 Hz, aromatic protons), 7.91 (1H, s, CH]N), 11.89 (1H,
s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 19.68 (CH3), 24.88, 30.52,
32.93, 48.53, 56.02, 103.54, 115.35, 125.95, 127.95, 129.03, 135.26,
142.37, 168.83. HRMS (m/z): [M + H]+ calcd for C22H24N4S:
377.1794; found 377.1781.
2.1.3.7. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
cyanophenyl)thiazole (3 g). FTIR (ATR, cm−1): ʋmax 3248 (NeH), 3105
(aromatic CeH), 2968 (aliphatic CeH), 2223 (C^N), 1602–1514 (C]
N and C]C), 1226 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.91
(3H, d, J = 6.5 Hz, CH3), 1.11–1.25 (2H, m, piperidine protons),
1.49–1.58 (1H, m, piperidine proton), 1.67 (2H, d, J = 12.7 Hz,
piperidine protons), 2.72 (2H, td, J1 = 2.1 Hz, J2 = 12.4 Hz,
piperidine protons), 3.77 (2H, d, J = 12.7 Hz, piperidine protons),
6.95 (2H, d, J = 8.9 Hz, aromatic protons), 7.47 (2H, d, J = 8.8 Hz,
aromatic protons), 7.58 (1H, s, aromatic proton), 7.86 (2H, d,
J = 8.5 Hz, aromatic protons), 7.93 (1H, s, CH]N), 8.02 (2H, d,
J = 8.5 Hz, aromatic protons), 11.96 (1H, s, NH). 13C NMR (75 MHz,
DMSO-d6, ppm) δ 22.23 (CH3), 30.70, 33.70, 48.35, 107.47, 109.94,
115.32, 119.48, 124.01, 126.54, 128.02, 133.13, 139.36, 142.84,
149.27, 152.24, 169.15. HRMS (m/z): [M+H]+ calcd for C23H23N5S:
402.174; found 402.1735.
2.1.3.12. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
methylphenyl)thiazole (3l). FTIR (ATR, cm−1): ʋmax 3238 (NeH), 3047
(aromatic CeH), 2987 (aliphatic CeH), 1600–1442 (C]N and C]C),
1234 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.91 (3H, d,
J = 6.5 Hz, CH3), 1.03–1.15 (1H, m, piperidine proton), 1.54–1.76
(4H, m, piperidine protons), 2.26–2.42 (4H, m, piperidine protons,
CH3), 2.71–2.73 (1H, m, piperidine proton), 2.56–2.73 (2H, m,
piperidine protons), 6.96–7.06 (3H, m, aromatic protons), 7.19 (3H,
t, J = 8.4 Hz, aromatic protons), 7.40–7.51 (2H, m, aromatic protons),
7.73 (1H, d, J = 8.1 Hz, aromatic proton), 7.91 (1H, s, CH]N), 11.89
(1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 19.61 (CH3), 21.28
(CH3), 24.76, 30.48, 32.73, 47.41, 48.76, 102.68, 114.08, 115.66,
125.91, 127.95, 128.17, 129.23, 129.60, 132.05, 137.17, 142.12,
150.06, 154.58, 168.91. HRMS (m/z): [M+H]+ calcd for C23H26N4S:
391.1951; found 391.1946.
2.1.3.8. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
hydroxyphenyl)thiazole (3 h). FTIR (ATR, cm−1): ʋmax 3329 (NeH),
3068 (aromatic CeH), 2918 (aliphatic CeH), 1600–1514 (C]N and
C]C), 1226–1047 (CeN and CeO). 1H NMR (300 Mhz, DMSO-d6, ppm)
δ 0.91 (3H, d, J = 6.5 Hz, CH3), 1.14–1.25 (2H, m, piperidine protons),
1.57–1.70 (3H, m, piperidine protons), 2.83–2.92 (2H, m, piperidine
protons), 3.60–3.74 (2H, m, piperidine protons), 6.78 (2H, d,
J = 8.7 Hz, aromatic protons), 6.96–7.15 (3H, m, aromatic protons),
7.46–7.53 (2H, m, aromatic protons), 7.64 (2H, d, J = 8.7 Hz, aromatic
protons), 7.93 (1H, s, CH]N), 9.57 (1H, s, OH), 11.89 (1H, s, NH). 13
C
NMR (75 MHz, DMSO-d6, ppm) δ 22.08 (CH3), 30.76, 33.31, 47.56,
100.62, 114.30, 115.73, 116.81, 127.36, 127.90, 129.32, 129.62,
131.76, 133.34, 157.70, 164.31. HRMS (m/z): [M+H]+ calcd for
2.1.3.13. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
bromophenyl)thiazole (3m). FTIR (ATR, cm−1): ʋmax 3338 (NeH), 3076
(aromatic CeH), 2974 (aliphatic CeH), 1600–1458 (C]N and C]C),
1234 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.93 (3H, d,
J = 6.5 Hz, CH3), 1.03–1.15 (1H, m, piperidine proton), 1.54–1.79
(4H, m, piperidine protons), 2.79–2.97 (1H, m, piperidine proton),
3.63–3.95 (3H, m, piperidine protons), 7.03–7.43 (6H, m, aromatic
protons), 7.59 (2H, d, J = 8.6 Hz, aromatic protons), 7.80 (1H, d,
J = 8.5 Hz, aromatic proton), 7.97 (1H, s, CH]N), 12.13 (1H, s, NH).
13C NMR (75 MHz, DMSO-d6, ppm) δ 19.45 (CH3), 30.28, 34.62, 46.72,
48.37, 103.72, 114.81, 115.37, 125.82, 127.31, 128.01, 130.26,
C
22H24N4OS: 393.1744; found 393.1738.
2.1.3.9. 2-[2-(4-(4-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(2,4-
difluorophenyl)thiazole (3i). FTIR (ATR, cm−1): ʋmax 3290 (NeH), 3130
(aromatic CeH), 2951 (aliphatic CeH), 1600–1487 (C]N and C]C),
1136 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.92 (3H, d,
J = 6.5 Hz, CH3), 1.12–1.25 (2H, m, piperidine protons), 1.48–1.60
(1H, m, piperidine proton), 1.68 (2H, d, J = 12.6 Hz, piperidine
protons), 2.73 (2H, t, J = 11.9 Hz, piperidine protons), 3.77 (2H, d,
J = 12.7 Hz, piperidine protons), 6.96 (2H, d, J = 8.8 Hz, aromatic
99

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
131.99, 137.92, 142.73, 151.34, 154.82, 168.19. HRMS (m/z): [M
piperidine protons), 2.84–3.11 (2H, m, piperidine protons), 3.60–3.68
(2H, m, piperidine protons), 6.61–6.84 (2H, m, aromatic protons), 7.04
(1H, s, aromatic proton), 7.33–7.44 (2H, m, aromatic protons), 7.65
(4H, d, J = 8.6 Hz, aromatic protons), 8.01 (1H, s, CH]N), 9.59 (1H, s,
OH), 12.18 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 19.25
(CH3), 23.22, 30.08, 34.04, 48.71, 101.07, 115.75, 126.08, 127.44,
128.06, 129.91, 132.72, 134.64, 142.72, 157.63, 168.43. HRMS (m/z):
[M+H]+ calcd for C22H24N4OS: 393.1744; found 393.1737.
+H]+ calcd for C22H23BrN4S: 455.0900; found 455.0885.
2.1.3.14. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
chlorophenyl)thiazole (3n). FTIR (ATR, cm−1): ʋmax 3381 (NeH), 3078
(aromatic CeH), 2974 (aliphatic CeH), 1602–1438 (C]N and C]C),
1234 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.92 (3H, d,
J = 6.5 Hz, CH3), 1.05–1.13 (1H, m, piperidine proton), 1.59–1.75
(4H, m, piperidine protons), 2.80 (1H, br s, piperidine proton),
3.65–3.76 (3H, m, piperidine protons), 7.03–7.27 (4H, m, aromatic
protons), 7.36 (1H, s, aromatic proton), 7.44–7.55 (3H, m, aromatic
protons), 7.86 (1H, d, J = 8.6 Hz, aromatic proton), 7.95 (1H, s, CH]
N), 12.00 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 19.53
(CH3), 24.63, 30.38, 33.46, 48.73, 104.41, 115.99, 127.67, 128.01,
129.07, 129.96, 132.92, 134.92, 142.46, 168.37. HRMS (m/z): [M
+H]+ calcd for C22H23ClN4S: 411.1405; found 411.1395.
2.1.3.19. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-
(2,4-difluorophenyl)thiazole (3s). FTIR (ATR, cm−1): ʋmax 3273 (NeH),
3049 (aromatic CeH), 2924 (aliphatic CeH), 1595–1485 (C]N and
C]C), 1261 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.91 (3H, d,
J = 6.6 Hz, CH3), 1.04–1.12 (1H, m, piperidine proton), 1.50–1.79 (4H,
m, piperidine protons), 2.35–2.43 (1H, m, piperidine proton),
2.64–2.75 (1H, m, piperidine proton), 3.68–3.79 (2H, m, piperidine
protons), 6.94–7.01 (2H, m, aromatic protons), 7.14–7.20 (2H, m,
aromatic protons), 7.30–7.38 (1H, m, aromatic proton), 7.47 (1H, d,
J = 8.9 Hz, aromatic proton), 7.65 (1H, d, J = 8.3 Hz, aromatic
proton), 7.92 (1H, s, CH]N), 7.98–8.06 (1H, m, aromatic proton),
11.90 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 19.68 (CH3),
24.88, 30.53, 32.97, 48.44, 55.91, 104.98, 107.68 (d, J = 13.3 Hz),
112.25 (d, J = 21.7 Hz), 114.43, 115.27, 128.00, 130.83, 142.67,
152.21, 158.40, 168.30. HRMS (m/z): [M+H]+ calcd for
2.1.3.15. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
fluorophenyl)thiazole (3o). FTIR (ATR, cm−1): ʋmax 3338 (NeH), 3070
(aromatic CeH), 2945 (aliphatic CeH), 1602–1438 (C]N and C]C),
1232 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.92 (3H, d,
J = 6.5 Hz, CH3), 1.00–1.14 (1H, m, piperidine proton), 1.54–1.77
(4H, m, piperidine protons), 2.70 (1H, br s, piperidine proton),
3.56–3.73 (3H, m, piperidine protons), 6.96–7.07 (3H, m, aromatic
protons), 7.20–7.32 (3H, m, aromatic protons), 7.43–7.50 (2H, m,
aromatic protons), 7.86–7.92 (2H, m, aromatic proton, CH]N), 11.91
(1H, s, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 19.59 (CH3), 24.75,
30.45, 32.77, 56.23, 103.33, 114.06, 115.38, 115.68 (d, J = 3.2 Hz),
115.99, 127.86, 127.97, 128.92, 130.26 (d, J = 8 Hz), 131.77, 132.13,
142.34, 161.88 (d, J = 243.7 Hz), 163.34, 166.52, 168.93. HRMS (m/
z): [M+H]+ calcd for C22H23FN4S: 395.1700; found 395.1687.
C22H22F2N4S: 413.1606; found 413.1600.
2.1.3.20. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-
(3,4-hydroxyphenyl)thiazole (3t). FTIR (ATR, cm−1): ʋmax 3238 (NeH),
3122 (aromatic CeH), 2953 (aliphatic CeH), 1595–1485 (C]N and
C]C), 1263 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.91 (3H, d,
J = 6.3 Hz, CH3), 1.11–1.20 (1H, m, piperidine proton), 1.74–2.18 (4H,
m, piperidine protons), 2.78–3.08 (2H, m, piperidine protons),
3.51–3.55 (2H, m, piperidine protons), 6.42–6.51 (1H, m, aromatic
proton), 6.95 (1H, s, aromatic proton), 7.10–7.43 (3H, m, aromatic
protons), 7.59–7.72 (3H, m, aromatic proton), 8.02 (1H, s, CH]N),
9.02 (2H, br s, OH), 12.47 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6,
ppm) δ 19.27 (CH3), 29.85, 33.18, 48.19, 106.18, 115.63, 116.41,
119.50, 121.18, 124.86, 127.86, 128.16, 129.46, 142.72, 145.62,
145.97, 152.38, 169.87. HRMS (m/z): [M + H]+ calcd for
2.1.3.16. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
nitrophenyl)thiazole (3p). FTIR (ATR, cm−1): ʋmax 3346 (NeH), 3080
(aromatic CeH), 2904 (aliphatic CeH), 1597–1446 (C]N and C]C),
1508–1338 (NO2), 1107 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ
0.93 (3H, d, J = 6.5 Hz, CH3), 1.12–1.21 (1H, m, piperidine proton),
1.69–1.82 (4H, m, piperidine protons), 2.78–2.99 (1H, m, piperidine
proton), 3.63 (1H, t, J = 12.8 Hz, piperidine proton), 4.49 (2H, br s,
piperidine protons), 7.11–7.14 (1H, m, aromatic proton), 7.43–7.51
(2H, m, aromatic protons), 7.63–7.73 (2H, m, aromatic protons),
7.85–8.12 (3H, m, CH]N, aromatic protons), 8.22–8.29 (2H, m,
aromatic protons), 12.30 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6,
ppm) δ 19.19 (CH3), 30.02, 31.25, 32.90, 48.27, 108.73, 113.27,
124.60, 126.81, 128.11, 129.50, 146.67, 169.02. HRMS (m/z): [M
+H]+ calcd for C22H23N5O2S: 422.1645; found 422.1637.
C22H24N4O2S: 409.1693; found 409.1687.
2.2. MAO inhibition assay
The fluorometric enzyme inhibition assay was carried out to in-
vestigate the inhibition profiles of the synthesized compounds as in
previously defined by us [29,30]. Ampliflu™ Red (10-Acetyl-3,7-dihy-
droxyphenoxazine) was used as fluorescence reagent in this assay. All
reagents and enzymes (Ampliflu™ Red, peroxidase from horseradish,
hMAO-A, hMAO-B, H2O2, tyramine hydrochloride, moclobemide and
selegiline) were supplied from Sigma-Aldrich (Steinheim, Germany).
In the enzymatic assay, three different daily prepared solutions were
used. (I) Inhibitor solutions: Synthesized compounds and reference
agents were prepared in 2% DMSO in 10−3–10−9 M concentrations
(10 mL for each concentration). (II) Enzyme solutions: Recombinant
hMAO-A (0.5 U/mL) and recombinant hMAO-B (0.64 U/mL) enzymes
were dissolved in the phosphate buffer and final volumes were adjusted
to 10 mL. (III) Working solution: Horseradish peroxidase (200 U/mL,
100 μL), Ampliflu™ Red (20 mM, 200 μL) and tyramine (100 mM,
200 μL) were dissolved in the phosphate buffer and final volume was
adjusted to 10 mL.
2.1.3.17. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
cyanophenyl)thiazole (3q). FTIR (ATR, cm−1): ʋmax 3298 (NeH), 3080
(aromatic CeH), 2958 (aliphatic CeH), 2484 (C^N), 1602–1512 (C]
N and C]C), 1147 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.93
(3H, d, J = 6.5 Hz, CH3), 1.18–1.24 (1H, m, piperidine proton),
1.78–1.97 (4H, m, piperidine protons), 2.96–3.22 (2H, m, piperidine
protons), 3.57–3.65 (2H, m, piperidine protons), 7.37–7.55 (3H, m,
aromatic protons), 7.65–7.73 (3H, m, aromatic protons), 7.87 (2H, d,
J = 8.6 Hz, aromatic protons), 8.03 (2H, d, J = 8.3 Hz, CH]N,
aromatic proton), 12.30 (1H, s, NH). 13C NMR (75 MHz, DMSO-d6,
ppm) δ 19.14 (CH3), 23.82, 29.93, 48.72, 108.08, 110.06, 119.45,
126.58, 128.09, 133.18, 139.16, 141.17, 149.27, 168.90. HRMS (m/z):
[M+H]+ calcd for C23H23N5S: 402.1747; found 402.1738.
The solutions of inhibitor (20 μL/well) and hMAO-A (100 μL/well)
or hMAO-B (100 μL/well) were added to the flat black bottom 96-well
micro test plate, and incubated at 37 °C for 30 min. After this incubation
period, the reaction was started by adding a working solution (100 μL/
well). The mixture was incubated at 37 °C for 30 min and the fluores-
cence (Ex/Em = 535/587 nm) was measured at 5 min intervals. Control
2.1.3.18. 2-[2-(4-(3-Methylpiperidin-1-yl)benzylidene)hydrazinyl]-4-(4-
hydroxyphenyl)thiazole (3r). FTIR (ATR, cm−1): ʋmax 3267 (NeH),
3126 (aromatic CeH), 2951 (aliphatic CeH), 1571–1512 (C]N and
C]C), 1141 (CeN). 1H NMR (300 Mhz, DMSO-d6, ppm) δ 0.93 (3H, d,
J = 6.5 Hz, CH3), 1.16 (1H, br s, piperidine proton), 1.78–1.81 (4H, m,
100

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
experiments were carried out simultaneously by replacing the inhibitor
solution with 2% DMSO (20 μL). To check the probable inhibitory effect
of inhibitors on horseradish peroxidase, a parallel reading was per-
formed by replacing enzyme solutions with %3 H2O2 solution (20 mM
100 μL/well). In addition, the possible capacity of the inhibitors to
modify the fluorescence generated in the reaction mixture due to non-
enzymatic inhibition was determined by mixing inhibitor and working
solutions.
The specific fluorescence emission (used to obtain the final results)
was calculated after subtraction of the background activity, which was
determined from vials containing all components except the hMAO
isoforms, which were replaced by phosphate buffer (100 μL/well).
Blank, control and all concentrations of inhibitors were analyzed in
quadruplicate and inhibition percent was calculated by using following
equation:
Scheme 1. Synthesis of the compounds (3a-3t). Reactants, reagents, and con-
ditions; (i) K2CO3, DMF, reflux, 24 h; (ii) EtOH, reflux, 4 h; (iii) appropriate
phenacyl bromides, EtOH, reflux, 3 h.
(FCt2 FCt1) (FIt2 FIt1)
%Inhibition =
× 100
FCt2 FCt1
[35] interface and then were submitted to the Protein Preparation Wi-
zard protocol of the Schrödinger Suite 2016 Update 2 [36]. The ligands
were prepared by the LigPrep 3.8 [37] to assign the protonation states at
where FCt2: Fluorescence of a control well measured at t2 time, FCt1:
Fluorescence of a control well measured at t1 time, FIt2: Fluorescence of
an inhibitor well measured at t2 time, FIt1: Fluorescence of an inhibitor
well measured at t1 time. The IC50 values were calculated from a dose-
response curve obtained by plotting the percentage inhibition versus
the log concentration with the use of Microsoft Office Excel 2013. The
pH 7.4
1.0 and the atom types, correctly. Bond orders were assigned
and hydrogen atoms were added to the structures. The grid generation
was formed using Glide 7.1 [38]. The grid box with dimensions of
20 Å × 20 Å × 20 Å was centered in the vicinity of the flavin (FAD) N5
atom on the catalytic site of the protein to cover all binding sites and
neighboring residues [39–41]. Flexible docking runs were performed
with single precision docking mode (SP).
results were displayed as mean
standard deviation (SD).
2.3. Enzyme kinetic studies
3. Results and discussion
Enzyme kinetic studies were performed for compounds 3j and 3 t in
order to determine the inhibition type on MAO enzymes. The same
materials were used in the MAO inhibition assay. The most active
compounds 3j and 3t were tested at three different concentrations
(IC50/2, IC50 and 2 × IC50). The solutions of inhibitor (20 μL/well) and
enzyme were added to the flat black bottom 96-well micro test plate,
and incubated at 37 °C for 30 min. After incubation period, the working
solution, including various concentrations (20, 10, 5, 2.5, 1.25, and
0.625 μM) of tyramine (100 μL/well) was added. The increase of the
fluorescence (Ex/Em = 535/587 nm) was recorded for 30 min. A par-
allel experiment was carried out without inhibitor. All processes were
assayed in quadruplicate. The results were analyzed as Lineweaver-
Burk plots using Microsoft Office Excel 2013. The Km / Vmax (slope)
values of the Lineweaver-Burk plots were replotted versus the inhibitor
concentration, and the Ki values were determined from the x-axis in-
tercept as -Ki.
3.1. Chemistry
The title compounds (3a-3t) were obtained according the multi-step
strategy outlined in Scheme 1. Reaction of 3 or 4-substituted piperidine
with 4-fluorobenzaldehyde resulted in the formation of 4-substituted
benzaldehyde derivatives (1a, 1b) which were then treated with thio-
semicarbazide to afford the corresponding thiosemicarbazones (2a,
2b). Subsequently, the intramolecular cyclization of compounds (2a,
2b) with appropriate phenacyl bromide derivatives generated the target
thiazole compounds (3a-3 t) in good yields. Some properties of the
compounds were presented in Table 1.
The structural elucidation of the final compounds was performed by
FT-IR 1H NMR, 13C NMR, and HRMS. In the IR spectra, characteristic
stretching bands noted at between 3387 and 3238 cm−1
,
1620–1438 cm−1, 1263–1049 cm−1 belong to NeH, C]N and C]C,
CeN bonds, respectively.
2.4. Cytotoxicity test
In the 1H NMR spectra, methyl protons linked to piperidine were
observed at about 0.92–0.97 ppm as doublet peak. Azomethine (eCH]
N) and NeH protons were recorded as singlet peaks at the region of
7.91–8.03 ppm and 11.87–12.47 ppm, respectively. Aromatic protons
were assigned in range 6.41–8.29 ppm. In the 13C NMR spectra, peaks
at about 19.14–22.23 ppm were recorded for methyl carbons attached
to piperidine. Aromatic carbons were generally detected at
100.62–169.87 ppm range. Molecular ion peaks were designated in
agreement with molecular weights of the compounds.
MTT test was performed to determine the cytotoxic profile of the
most active compounds 3j and 3t using the NIH/3T3 mouse embryonic
fibroblast cell line (ATCC® CRL-1658™, London, UK). The method was
executed in the same way as we have reported in our previous work
[31].
2.5. Theoretical determination of ADME properties
ADME parameters of all synthesized compounds were assessed with
the help of QikProp 4.8 software [32].
3.2. MAO inhibition assay
2.6. Molecular docking studies
The fluorometric method was applied in two steps in order to in-
vestigate the hMAO-A and hMAO-B isoenzymes inhibitory activity of all
synthesized compounds (3a-3t). Final compounds were prepared and
tested at the initial concentrations (10−3 and 10−4 M) in the first step.
According to the first step results, the compounds showing > 50% in-
hibition were selected for the second stage. In this second step,
10−5–10−9 M concentrations of the selected compounds were prepared
and used. Considering Table 2, only compounds 3a, 3b, 3k, and 3l
In order to determine the binding modes of compounds 3j and 3 t on
hMAO-A and hMAO-B enzyme active site, docking studies were per-
formed. X-ray crystal structures of hMAO-A (PDB ID: 2Z5X) [33] and
hMAO-B (PDB ID: 2V5Z) [34] were retrieved from the Protein Data
Bank (www.pdb.org).
The structures of ligands were built using the Schrödinger Maestro
101

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
Table 1
Some properties of the synthesized compounds (3a-3t).
Compound
R1
R2
R3
R4
R5
Yield (%)
M.p. (˚C)
Molecular formula
Molecular weight
3a
3b
3c
3d
3e
3f
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
H
H
H
H
H
H
H
H
H
H
F
H
H
H
H
H
H
H
H
H
OH
H
H
H
H
H
H
H
H
H
OH
H
75
70
68
70
69
71
70
70
76
67
72
75
67
65
68
60
65
69
71
73
187
230
209
182
165
205
217
170
175
233
150
120
157
157
146
183
216
235
155
209
C22H24N4S
C23H26N4S
376,52
390,55
455,42
410,96
394,51
421,52
401,53
392,52
412,50
408,52
376,52
390,55
455,42
410,96
394,51
421,52
401,53
392,52
412,50
408,52
H
CH3
Br
H
C22H23
BrN4S
ClN4S
FN4S
N5O2S
N5S
H
Cl
C22 23
C22 23
C22 23
C23 23
C22 24
C22 22
C22 24
C22 24
C23 26
C22 23
C22 23
C22 23
C22 23
C23 23
C22 24
C22 22
C22 24
H
H
F
H
H
NO2
CN
OH
F
H
3g
3h
3i
H
H
H
H
N4OS
F2N4S
N4O2S
N4S
H
H
3j
H
H
H
H
H
H
H
H
H
H
F
OH
H
H
3k
3l
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
CH3
H
H
CH3
Br
H
N4S
3m
3n
3o
3p
3q
3r
3s
3t
H
H
BrN4S
ClN4S
FN4S
N5O2S
N5S
H
Cl
H
H
F
H
H
NO2
CN
OH
F
H
H
H
H
H
N4OS
F2N4S
N4O2S
H
H
H
H
OH
H
substitution and para methyl substitution of phenyl group caused a
dramatic activity loss. On the other hand, all other compounds, bearing
halogen, hydroxyl, nitro or cyano substituents, displayed moderate to
good inhibition against both isoenzymes. According to biological ac-
tivity results, compounds 3j and 3t were found as the most active de-
rivatives in the series against both MAO isoenzymes. When these
compounds are structurally examined, it can be seen that their sub-
stitution pattern is different from those of compounds reported both in
our previous [28] and present studies. Both 3j and 3t carry hydroxyl
groups at meta and para positions of phenyl ring. Thus, it can be sug-
gested that 3,4-dihydroxy substitution has an important influence on
both hMAO-A and hMAO-B enzyme inhibition.
Table 2
% Inhibition of compounds 3a-3t, moclobemide and selegiline at the initial
concentrations against MAO-A and MAO-B.
Compound
MAO-A IC50 (µM)
MAO-B IC50 (µM)
*SI
Selectivity
3c
0.849
0.452
0.187
0.594
0.380
0.280
0.194
0.134
0.765
0.442
0.202
0.524
0.338
0.277
0.209
0.123
6.054
–
0.023
0.011
0.012
0.018
0.014
0.011
0.022
0.004
0.028
0.017
0.011
0.020
0.013
0.010
0.016
0.005
0.173
0.665
0.152
0.117
0.184
0.137
0.109
0.184
0.027
0.698
0.149
0.114
0.173
0.136
0.104
0.196
0.025
–
0.018
0.005
0.003
0.007
0.004
0.003
0.017
0.001
0.024
0.004
0.003
0.006
0.005
0.003
0.022
0.001
1,28
2,97
1,60
3,23
2,77
2,57
1,05
4,96
1,10
2,97
1,77
3,03
2,49
2,66
1,07
4,92
–
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-B
MAO-A
MAO-B
3d
3e
3f
3g
3h
3i
3j
3m
The N1 of hydrazine moiety is another important region of the
compounds for an intrinsic enzyme inhibition. According to literature
N1 of the hydrazine group has steric and electronic influences on en-
zyme-inhibitor interaction [20,21]. From this knowledge in the present
study we incorporated 4-(3- or 4-methylpiperidin-1-yl)benzylidene
fragment to N1 of hydrazine. Although 3- or 4-methyl substitutions of
piperidine do not cause different inhibition potencies each other, they
have a great contribution on the enzyme inhibition potency of the
compounds compared with the compounds that we reported previously
[28].
3n
3o
3p
3q
3r
3s
3t
Moclobemide
Selegiline
0.039
0.001
–
* The selectivity index (SI) was calculated as IC50 (MAO-A) / IC50 (MAO-B).
could not pass the second step of hMAO-A and hMAO-B enzymes in-
hibition assay. The IC50 values of compounds against hMAO-A and
hMAO-B enzymes are given in Table 2. Compounds 3j and 3t showed
3.3. Enzyme kinetic studies
IC50 values of 0.134
0.004 µM and 0.123
0.005 µM, respectively,
0.173 μM
on hMAO-A. In hMAO-B inhibition study, compounds 3j and 3t were
the most active derivatives with IC50 values of 0.027 0.001 µM and
0.025 0.001 µM. Selegiline was used as a reference drug in hMAO-B
enzyme activity and showed an IC50 of 0.039
MAO inhibitors are divided into two types as reversible or irrever-
sible inhibitors. The first produced MAO inhibitors inhibit based on
mechanism and they bind to proteins covalently. These irreversible
inhibitors have serious side effects like hallucination, schizophrenia and
hypertension. They perform covalent binding to the proteins to yield
reactive products. In addition to these kinds of MAO inhibitors also had
noticeable hepatotoxic side effects, by inactivating the P450, which
cannot be tolerated. Therefore, for newly developed drug candidates,
reversible inhibition types as uncompetitive, competitive, non-compe-
titive, or mixed type are required in order to avoid such side effects.
The enzyme kinetic studies of compounds 3j and 3t on both hMAO-
A and hMAO-B enzymes were carried out in order to determine the
inhibition type and thus predict whether or not they have the side ef-
fects mentioned above. Kinetic studies were applied with a similar
procedure to the MAO inhibition assay. The type of inhibition was
specified with the help of Lineweaver-Burk graphics [42]. Enzyme ki-
netics were examined by recording substrate velocity curves in the
absence and presence of the most effective compounds 3j and 3t, which
whereas standard drug moclobemide had an IC50 of 6.054
0.001 µM.
Selectivity indexes (SI), calculated as IC50(MAO-A)/IC50(MAO-B),
are presented in Table 2. SI < 1 means selectivity towards MAO-A,
while SI > 1 shows selectivity towards MAO-B. The data in Table 2
shows that synthesized compounds possess selectivity towards MAO-B.
However, very close SIs to 1 suggests that enzyme selectivity is not
evident.
In terms of SARs the phenyl group at C4 of the thiazole is the
structural property that should be evaluated first. Because, significant
changes in enzyme inhibition have been observed by the alterations of
substituent type and position on phenyl group. It has been reported that
ortho and/or para substitution of phenyl group are responsible for en-
zyme inhibition [20,21]. Interestingly, in the present study non-
102

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
0.12
0.10
0.08
0.06
0.04
0.02
0.00
-0.02
IC50 / 2 (0.062 µM)
IC50 (0.123 µM)
2 x IC50 (0.246 µM)
Control
-2
-1.5
-1
-0.5
0
0.5
1
1.5
2
1/[S] µM-1
(A)
0.08
0.06
0.04
0.02
0
y = 0.2164x + 0.0116
R² = 0.9989
-0.1
-0.05
0
0.05
0.1
0.15
0.2
0.25
0.3
-0.02
Compound 3t (µM)
(B)
Fig. 2. (A) Lineweaver–Burk plots for the inhibition of hMAO-A by compound 3 t. [S], substrate concentration (μM); V, reaction velocity (nmol/min/mg protein).
Inhibitor concentrations are shown at the left. Km values from 2 × IC50 to Control; 5.111, 3.192, 2.208 and 0.923 (μM). Vmax value of the competitive inhibition;
79.950
2.341 (nmol/min/mg protein). (B) Secondary plot for calculation of steady-state inhibition constant (Ki) of compound 3t. Ki was calculated as 0.054 µM.
were prepared at 2xIC50, IC50 and IC50/2 concentrations. The substrate
(tyramine) was used at different concentrations ranging from 0.625 µM
to 20 µM, and thus the initial velocity measurements were obtained.
Then, Dixon plots were formed by using the data gained from Line-
weaver-Burk graphics in order to calculate Ki values of compounds 3j
and 3t. The graphical analysis of steady-state inhibition data are pre-
sented for 3t in Figs. 2 and 3 and given for 3j in Supporting
Information Figs. S1 and S2.
3.4. Cytotoxicity test
As a general medicinal chemistry approach a drug candidate should
also not be toxic in addition to its essential biological activity. Therefore,
the most active derivatives 3j and 3t were screened for their cytotoxicity
profiles. Thus, MTT assay was applied by using healthy NIH/3T3 mouse
embryonic fibroblast cell line (ATCC CRL1658), which is recommended for
cytotoxicity screening by ISO (10993-5, 2009) [43]. It can be seen from
Table 3, compounds 3j and 3t had IC50 values of > 1000 µM and 600 µM,
respectively, against NIH/3T3 cells. These IC50 values are very higher than
those obtained in MAO-B enzyme inhibition assay (0.027 µM and
0.025 µM, respectively). As a result, compounds 3j and 3t have been
proven to be non-toxic in their effective concentration against hMAO-B.
In the Lineweaver-Burk graphics of compounds 3j and 3 t, inter-
section of plots on y-axis and different slopes and intercepts on x-axis
describe the competitive inhibition type. Hence, it can be stated that
these compounds are reversible and competitive inhibitors against
hMAO-A and hMAO-B enzymes, so have similar inhibition features with
the substrate. The Ki values for compounds 3j and 3t were calculated as
0.071 µM and 0.052 µM, respectively, for the inhibition of hMAO-A,
whereas their Ki values on hMAO-B enzyme kinetics were found as
0.012 µM and 0.010 µM, respectively.
3.5. Calculated ADME parameters
Having favorable ADME (absorption, distribution, metabolism, ex-
cretion) properties is one of the most challenging obstacles for drug
103

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
0.12
0.10
0.08
0.06
0.04
0.02
0.00
-0.02
IC50 / 2 (0.013 µM)
IC50 (0.025 µM)
2 x IC50 (0.050 µM)
Control
-2
-1.5
-1
-0.5
0
0.5
1
1.5
2
1/[S] µM-1
(A)
0.08
0.06
0.04
0.02
0
y = 1.2526x + 0.0109
R² = 0.9944
-0.02 -0.01
0
0.01
0.02
0.03
0.04
0.05
0.06
Compound 3t (µM)
-0.02
(B)
Fig. 3. (A) Lineweaver–Burk plots for the inhibition of hMAO-B by compound 3t. [S], substrate concentration (μM); V, reaction velocity (nmol/min/mg protein).
Inhibitor concentrations are shown at the left. Km values from 2 × IC50 to Control; 7.796, 4.267, 3.042 and 0.935 (μM). Vmax value of the competitive inhibition;
104.832
2.023 (nmol/min/mg protein). (B) Secondary plot for calculation of steady-state inhibition constant (Ki) of compound 3t. Ki was calculated as 0.010 µM.
ADME predictions of all synthesized compounds (3a-3t) were car-
ried out by QikProp 4.8 software [32]. QikProp also provides acceptable
Table 3
The IC50 value of the compounds 3j and 3t against NIH/3T3 cell line.
ranges for comparing the predicted properties of compounds with those
of 95% of known drugs. In addition to ADME properties, drug-likeness
properties were also estimated by QikProp. The drug-likeness of a
compounds was assessed according to Lipinski’s Rule of Five [45],
which considers molecular weight (< 500 Da), number of hydrogen
bond acceptors (≤10) and donors (≤5), and octanol/water partition
coefficient (≤5) and Jorgensen's rule of three [46], which regards logS
(> -5.7), PCaco (> 22 nm/s), primary metabolites (PM) (< 7). These
rules are essential for the optimization of biologically active com-
pounds, and thus, they must not be violated.
Compound
Cell Line
IC50(µM)
3j
3t
NIH/3T3
NIH/3T3
> 1000
600
development. ADME expresses the properties of absorption, distribu-
tion, metabolism and excretion of a molecule within an organism. All
these features are important for any drug. The early optimization could
be provided by ADME calculation. Early resolution of problems that
cause ADME failure prevents later time loss. Identification and elim-
ination of adverse compounds make the research process more effective
and efficient [44]. For this reason, to predict the pharmacokinetic
properties of new drug candidates as early as possible in the drug de-
velopment process is very important.
The predicted ADME properties are presented in Table 4 and this
table contains the following parameters: Molecular weight (MW),
number of rotatable bonds (RB), dipole moment (DM), molecular vo-
lume (MV), number of hydrogen donors (DHB), number of hydrogen
acceptors (AHB), polar surface area (PSA), octanol/water partition
coefficient (log P), aqueous solubility (log S), apparent Caco-2 cell
104

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
Table 4
Calculated ADME parameters of compounds 3a-3t.
Comp.
MW
RB
DM
MV
DHB
AHB
PSA
logP
logS
PCaco
logBB
PMDCK
PM
%HOA
VRF
VRT
3a
3b
3c
3d
3e
3f
376.518
390.545
455.414
410.963
394.509
421.516
401.528
392.518
412.499
408.517
376.518
390.545
455.414
410.963
394.509
421.516
401.528
392.518
412.499
408.517
4
4
4
4
4
5
5
5
4
6
4
4
4
4
4
5
5
5
4
6
2.451
1.975
4.494
4.810
4.868
12.450
8.783
2.720
4.004
4.467
2.453
1.977
4.591
4.908
4.871
12.555
8.885
2.730
4.122
4.467
1276.917
1335.853
1329.949
1321.037
1293.032
1359.315
1343.614
1300.086
1302.694
1321.534
1276.949
1335.885
1322.774
1313.861
1293.064
1352.140
1336.439
1300.118
1295.519
1321.566
1
1
1
1
1
1
1
2
1
3
1
1
1
1
1
1
1
2
1
3
5
40.192
40.192
40.194
40.194
40.196
88.858
65.987
62.874
38.754
84.384
40.192
40.192
38.829
38.829
40.196
87.493
64.622
62.874
37.389
84.384
5.606
5.912
6.179
6.102
5.842
4.905
4.842
4.842
6.004
4.125
5.606
5.912
6.132
6.054
5.842
4.858
4.796
4.842
5.957
4.126
−7.471
−8.040
−8.334
−8.217
−7.839
−7.716
−8.418
−7.158
−8.068
−6.805
−7.471
−8.039
−8.224
−8.108
−7.838
−7.608
−8.310
−7.158
−7.959
−6.804
4525.096
4525.091
4524.991
4524.925
4524.816
465.090
0.051
4998.277
4998.272
10000.000
10000.000
9037.418
427.377
1
2
1
1
1
2
1
2
1
3
1
2
1
1
1
2
1
2
1
3
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
1
1
1
1
1
0
0
0
1
0
1
1
1
1
1
0
0
0
1
0
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
5
0.042
5
0.232
5
0.219
5
0.163
6
−1.235
−0.864
−0.638
0.257
3g
3h
3i
6.5
5.75
5
935.800
909.948
1363.085
4699.197
490.613
1366.369
10000.000
452.783
3j
6.5
5
−1.241
0.051
3k
3l
4525.096
4525.091
4524.991
4524.925
4524.816
465.090
4998.277
4998.272
10000.000
10000.000
9037.418
427.377
5
0.042
3m
3n
3o
3p
3q
3r
3s
3t
5
0.235
5
0.223
5
0.163
6
−1.221
−0.853
−0.637
0.260
6.5
5.75
5
935.800
909.948
1363.085
4699.197
490.613
1366.369
10000.000
452.783
6.5
−1.241
MW: Molecular weight RB: Number of rotatable bonds DM: Computed dipole moment MV: Total solvent-accessible volume DHB: Estimated number of hydrogen
bond donors AHB: Estimated number of hydrogen bond acceptors PSA: Van der Waals surface area of polar nitrogen and oxygen atoms and carbonyl carbon atoms
logP: Predicted octanol/water partition coefficient logS: Predicted aqueous solubility PCaco: Predicted apparent Caco-2 cell permeability logBB: Predicted brain/
blood partition coefficient PMDCK: Predicted apparent MDCK cell permeability PM: Number of likely metabolic reactions %HOA: Predicted human oral absorption
percent VRF: Number of violations of Lipinski’s rule of five. The rules are: MW < 500, logP < 5, DHB ≤ 5, AHB ≤ 10, Positive PSA value. VRT: Number of
violations of Jorgensen’s rule of three. The three rules are: logS > -5.7, PCaco > 22 nm/s, PM < 7.
permeability (PCaco), brain/blood partition coefficient (logBB), ap-
parent MDCK cell permeability (PMDCK), number of likely primer
metabolic reactions (PM), and percent of human oral absorption (%
HOA), the violations of rules of three (VRT) and five (VRF). According
to Lipinski's rule of five and Jorgensen's rule of three all compounds
(3a-3t) are in accordance with the rule by causing no more than one
violation.
Table 5
Docking scores of the compounds 3a-3t against MAO-A and MAO-B enzymes.
Compound
MAO-A Docking Score
MAO-B Docking Score
(kcal/mol)
(kcal/mol)
3a
3b
3c
3d
3e
3f
−9.749
−8.937
−7.692
−9.108
−9.732
−9.873
−7.641
−8.101
−9.866
−10.222
−8.821
−7.706
−7.849
−7.796
−8.67
−8.911
−9.247
−8.932
−9.143
−9.736
−8.375
−9.191
−9.454
−9.099
−10.354
−8.672
−7.619
−8.948
−9.015
−8.924
−8.076
−9.269
−9.417
−9.333
−10.079
Medicines targeted specifically to central nervous system should
cross the blood-brain barrier (BBB). Thus, the ability to cross the blood
brain barrier of these drugs of a great importance and should be eval-
uated in the early stage of drug development. For this purpose, ap-
parent MDCK cell permeability (PMDCK) and brain/blood partition
coefficient (logBB) were calculated. The MDCK cells are considered as a
good mimic for the blood brain barrier. According to software predic-
tions, the PMDCK values of < 25 and > 500 nm/s are recommended as
poor and great for non-active transport of compound. So as to pass BBB,
LogBB is another important parameter, which has recommended values
of −3–1.2. According to Table 4, results of PMDCK and logBB are
among these reference values and it can be said that all synthesized
compounds are capable of crossing BBB.
3g
3h
3i
3j
3k
3l
3m
3n
3o
3p
3q
3r
3s
3t
−9.034
−9.423
−9.217
−9.116
−10.217
As a consequence of ADME and BBB Permeability estimates, it can
be suggested that the final compounds may have a good pharmacoki-
netic profile.
docking poses of compounds are given in Figs. 4, 5 and in Supporting
information (Figs. S3-S12).
3.6. Molecular docking studies
The docking poses related to hMAO-A enzyme of compounds 3j and
3t are presented in Fig. 4A, 5A, 5B and in Supporting information
Figs. S3-S7. Both compounds showed similar interactions. In com-
pound 3j the thiazole ring was in an interaction with phenyl of Tyr407
by doing π-π interaction, while for compound 3t, this interaction was
observed with phenyl of Tyr444. The N1 nitrogen of the hydrazine
moiety emerged in terms of polar interaction. The N1 nitrogen formed a
hydrogen bond with hydroxyl of Tyr444 in both compounds. The last
interaction belonging to these compounds was a hydrogen bond, which
was formed by hydroxyl moiety at the 3rd position of phenyl ring. In
the compound 3j, this hydroxyl group was in interaction with carbonyl
of Arg51, while, hydroxyl moiety of compound 3t created this inter-
action with amino of Lys305. These two compounds carry the hydroxyl
Compounds 3j and 3t were found as the most active derivatives in
the series as mentioned in MAO enzyme activity part. The common
feature of these compounds is bearing dihydroxy moiety at 3rd and 4th
positions of the phenyl ring. Therefore, the docking studies were con-
ducted in order to determine in silico binding modes of these com-
pounds and to be able to evaluate the contribution of the dihydroxy
moiety to both MAO-A and MAO-B enzyme activities. The X-ray crystal
structures of hMAO-A (PDB ID: 2Z5X) [33] and hMAO-B (PDB ID:
2V5Z) [34] were obtained from Protein Data Bank (www.pdb.org). The
docking scores were calculated for all compounds. The obtained data
(Table 5) confirmed the biological activity results. The most active
compounds 3j and 3t also displayed the highest docking scores. The
105

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
Fig. 4. A: The three-dimensional interacting modes of compound 3t in the active region of hMAO-A. The inhibitor and the important residues in the active site of the
enzyme are presented by tube model. The FAD molecule is coloured red with ball and stick model. B: The three-dimensional interacting modes of compound 3t in the
active region of hMAO-B. The inhibitor and the important residues in the active site of the enzyme are presented by tube model. The FAD molecule is coloured green
with ball and stick model. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
group at the 3rd position of the phenyl ring, unlike the remainder of the
series. It can be suggested that owing to this additional interaction this
position has a significant effect on the biological activity. The presence
of a group capable of hydrogen bonding such as hydroxyl moiety at this
position has a positive contribution on enzyme inhibition. All these
findings explain the greater potency of compounds 3j and 3t on hMAO-
A enzyme compared to the other compounds. Also, it can be seen from
Fig. 5A, B and Supporting information Figures S5, S6 that van der
Waals and electrostatic interactions provide stronger binding to enzyme
active site. Both compounds have favorable van der Waals interactions
with Tyr60, Phe99, Phe103, Pro104, Trp119, Leu164, Leu167, Phe168,
Leu171, Cys172, Ile198, Ser200, Gln206, Leu328, Tyr398 and Tyr435,
displayed with pink and red colours as described in the user guide of
Glide [38]. Especially, it is thought that piperidine and methyl moiety at
its 4th position enhance van der Waals interaction with enzyme active
region. These compounds also show promising electrostatic contribu-
tions with Phe103, Leu167, Leu171, Ile198, Tyr326 and Leu328.
Docking studies of compounds 3j and 3t against hMAO-B enzyme
were also performed considering hMAO-A docking assays. The docking
poses of hMAO-B are given in Fig. 4B, Fig. 5C, D and in Supporting
information Figs. S8-S12. It can be understood from the docking poses
that these compounds showed five identical interactions. The phenyl
ring next to the piperidine was in an interaction with phenyl of Tyr326
by doing π-π interaction. The other π-π interaction was observed be-
tween thiazole ring and phenyl of Tyr398. In both compounds, dihy-
droxy substitution at 3rd and 4th positions of phenyl ring is very es-
sential for polar interactions by doing three hydrogen bonds to bind
enzyme active site. The hydroxy substituent at the 3rd position had two
hydrogen bonds by interacting with amino of Ser59 acting as a hy-
drogen acceptor and carbonyl of Tyr60 acting as a hydrogen donor. The
hydroxy moiety at the 4th position of phenyl formed another hydrogen
bond with carbonyl of Gly434. This last hydrogen bond was observed in
hMAO-B docking runs, unlike hMAO-A docking studies. Besides, van
der Waals and electrostatic interactions of compounds 3j and 3t are
presented in Fig. 5C, D and in Supporting information Figs. S10, S11.
It can be seen that piperidine and methyl groups support binding to
residues of enzyme active region by van der Waals interactions. Both
compounds formed van der Waals interactions with Gly58, Ser59,
Tyr60, Phe103, Trp119, Leu164, Leu167, Phe168, Leu171, Cys172,
Ile198, Ile199, Gln206, Ile316, Tyr326, Phe343, Tyr398 and Tyr435,
which were shown as pink and red colours [38]. Electrostatic interac-
tions between docked compounds and Gly58, Ser59, Tyr60, Phe168,
Gln206, Gly434 and Tyr435 were also observed.
All these findings explain more potent activity of compounds 3j and
106

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
Fig. 5. A: The van der Waals interaction of compound 3t with active region of hMAO-A. B: The electrostatic interaction of compound 3t with active region of hMAO-
A C: The van der Waals interaction of compound 3t with active region of hMAO-B. D: The electrostatic interaction of compound 3t with active region of hMAO-B. The
residues displaying favourable van der Waals interactions have been shown with red and pink colour. The residues in electrostatic interaction are coloured with blue,
red, and pink according to the distance from ligand by Per-Residue Interaction panel. (For interpretation of the references to colour in this figure legend, the reader is
referred to the web version of this article.)
3t against hMAO-B enzyme. Furthermore, by looking at the general
structure of the series, it can be suggested that both hMAO-A and
hMAO-B inhibitions are positively affected by the presence of a hydroxy
group, which is capable of a hydrogen bonding at the 3rd and 4th po-
sitions of phenyl ring.
Declaration of interest
The authors declare no conflicts of interest.
Appendix A. Supplementary material
4. Conclusion
Supplementary data to this article can be found online at https://
doi.org/10.1016/j.bioorg.2018.12.019.
In this study, our aim was to add new effective MAO compounds
bearing thiazole ring to the literature. For this purpose, we have de-
signed and synthesized new twenty thiazole derivatives and evaluated
their in vitro and in silico biological activity against MAO isoenzymes.
According to in vitro MAO enzyme inhibition assay results, compounds
3j and 3t were found as the most active derivatives in the series against
both MAO-A and MAO-B enzymes. So as to clarify the inhibition type
and in silico features of them, enzyme kinetic and docking studies were
performed for these compounds. These compounds were assessed as
reversible and competitive inhibitors of MAO-A and MAO-B enzymes.
Also, it was indicated that these compounds were stronger than the
other compounds in the series owing their different substituents.
Furthermore, in vitro cytotoxicity and in silico ADME calculations have
also shown that these compounds are promising MAO inhibitor candi-
dates. Consequently, medicinal chemists may improve new molecules,
which are more potent and safer MAO inhibitors, derived from com-
pounds 3j and 3t for the treatment of neurological diseases based on
these results.
References
[1] C. Binda, A. Mattevi, D.E. Edmondson, Int. Rev. Neurobiol. 100 (2011) 1–11.
[2] M.B. Youdim, D. Edmondson, K.F. Tipton, Nat. Rev. Neurosci. 7 (4) (2006)
295–309.
[3] R.M. Geha, I. Rebrin, K. Chen, J.C. Shin, J. Biol. Chem. 276 (13) (2001) 9877–9882.
[4] L.J. Legobe, A. Petzer, J.P. Petzer, Bioorg. Chem. 45 (2012) 1–11.
[5] J. Wang, K. Huo, L. Ma, L. Tang, D. Li, X. Huang, Y. Yuan, C. Li, W. Wang, W. Guan,
H. Chen, C. Jin, J. Wei, W. Zhang, Y. Yang, Q. Liu, Y. Zhou, C. Zhang, Z. Wu, W. Xu,
Y. Zhang, T. Liu, D. Yu, Y. Zhang, L. Chen, D. Zhu, X. Zhong, L. Kang, X. Gan, X. Yu,
Q. Ma, J. Yan, L. Zhou, Z. Liu, Y. Zhu, T. Zhou, F. He, X. Yang, Assoc. Mol. Syst. Biol.
536 (2011) 10–13.
[6] D.E. Edmondson, C. Binda, J. Wang, A.K. Upadhyay, A. Mattevi, Biochemistry 48
(2009) 4220–4230.
[7] N. Kaludercic, A. Carpi, R. Menabò, F. Di Lisa, N. Paolocci, Biochim. Biophys. Acta
(BBA)-Mol. Cell Res. 1813 (7) (2011) 1323–1332.
[8] F. Chimenti, D. Secci, A. Bolasco, P. Chimenti, A. Granese, S. Carradori, O. Befani,
P. Turini, S. Alcaro, F. Ortuso, Bioorg. Med. Chem. Lett. 16 (2006) 4135–4140.
[9] A.M. Helguera, A. Pérez-Garrido, A. Gaspar, J. Reis, F. Cagide, D. Vina,
M.N. Cordeiro, F. Borges, Eur. J. Med. Chem. 59 (2013) 75–90.
[10] D.S. Knudsen-Gerber, Consult. Pharm. 26 (2011) 48–51.
[11] W.J. Geldenhuys, M.O. Funk, C.J. Van der Schyf, R.T. Carroll, Bioorg. Med. Chem.
Lett. 22 (3) (2012) 1380–1383.
Acknowledgment
[12] R. Singh, S.N. Geetanjali, Chem. Biol. Lett. 1 (2014) 33–39.
[13] R.H. Haung, R. Faulkner, J. Biol. Chem. 256 (1981) 9211–9215.
[14] R. Pignatello, S. Mazzone, F. Castelli, P. Mazzone, G. Raciti, G. Mazzone, Pharmazie
49 (4) (1994) 272–276.
his study was financially supported by Anadolu University Scientific
Projects Fund, Project No: 1506S516.
107

B.N. Sağlık et al.
BioorganicChemistry85(2019)97–108
[15] D.E. Edmondson, C. Binda, A. Mattevı, Biochem. and Biophy. 464 (2007) 269–276.
[16] M. Ebadi, S.K. Srinivasan, M.D. Baxi, Prog. Neurobiol. 48 (1) (1996) 1–19.
[17] D. Secci, A. Bolasco, S. Carradori, M. D'Ascenzio, R. Nescatelli, M. Yáñez, Eur. J.
Med. Chem. 58 (2012) 405–417.
[28] N.Ö. Can, D. Osmaniye, S. Levent, B.N. Sağlık, B. Korkut, Ö. Atlı, Y. Özkay,
Z.A. Kaplancıklı, Eur. J. Med. Chem. 144 (2018) 68–81.
[29] Ö.D. Can, D. Osmaniye, Ü. Demir Özkay, B.N. Sağlık, S. Levent, S. Ilgın, M. Baysal,
Y. Özkay, Z.A. Kaplancıklı, Eur. J. Med. Chem. 131 (2017) 92–106.
[30] N.Ö. Can, D. Osmaniye, S. Levent, B.N. Sağlık, B. İnci, S. Ilgın, Y. Özkay,
Z.A. Kaplancıklı, Molecules 22 (8) (2017) 1381.
[18] G. Raciti, P. Mazzone, A. Raudino, G. Mazzone, A. Cambria, Bioorg. Med. Chem. 3
(1995) 1485–1491.
[19] S. Gritsch, S. Guccione, R. Hoffmann, A. Cambria, G. Raciti, T. Langer, J. Enzyme
Inhib. 16 (2001) 199–215.
[31] Ü. Demir Özkay, Ö.D. Can, B.N. Sağlık, U. Acar Çevik, S. Levent, Y. Özkay, S. Ilgın,
Ö. Atlı, Bioorg. Med. Chem. Lett. 26 (2016) 5387–5394.
[20] M. D'Ascenzio, S. Carradori, D. Secci, L. Mannina, A.P. Sobolev, C. De Monte,
R. Cirilli, M. Yáñez, S. Alcaro, F. Ortuso, Bioorg. Med. Chem. 22 (10) (2014)
2887–2895.
[32] QikProp, Version 4.8, Schr€odinger, LLC, New York, NY, 2016.
[33] S.Y. Son, J. Ma, Y. Kondou, M. Yoshimura, E. Yamashita, T. Tsukihara, Proc. Natl.
Acad. Sci. (PNAS) 105 (15) (2008) 5739–5744.
[21] M. D'Ascenzio, P. Chimenti, M.C. Gidaro, C. De Monte, D. De Vita, A. Granese,
L. Scipione, R. Di Santo, G. Costa, S. Alcaro, M. Yáñez, S. Carradori, J. Enzyme
Inhib. Med. Chem. 30 (6) (2015) 908–919.
[34] C. Binda, J. Wang, L. Pisani, C. Caccia, A. Carotti, P. Salvati, D.E. Edmondson,
A. Mattevi, J. Med. Chem. 50 (2007) 5848–5852.
[35] Maestro, version 10.6, Schrödinger, LLC, New York, NY (2016).
[36] Schrödinger, LLC, New York, NY (2016).
[22] A. Cambria, A. Raudino, A. Geronikaki, G. Buemi, G. Raciti, P. Mazzone,
S. Guccione, S. Ragusa, J. Enzyme Inhib. 14 (4) (1999) 307–321.
[23] F. Chimenti, D. Secci, A. Bolasco, P. Chimenti, A. Granese, S. Carradori, M. Yáñez,
F. Orallo, M.L. Sanna, B. Gallinella, R. Cirilli, J. Med. Chem. 53 (17) (2010)
6516–6520.
[37] LigPrep, version 3.8, Schrödinger, LLC, New York, NY (2016).
[38] Glide, version 7.1, Schrödinger, LLC, New York, NY (2016).
[39] M. Toprakçı, K. Yelekçi, Bioorg. Med. Chem. Lett. 15 (2005) 4438–4446.
[40] N. Gökhan-Kelekçi, Ö.Ö. Şimşek, A. Ercan, K. Yelekçi, Z.S. Şahin, Ş. Işık, G. Uçar,
A.A. Bilgin, Bioorg. Med. Chem. 17 (2009) 6761–6772.
[24] F. Chimenti, E. Maccioni, D. Secci, A. Bolasco, P. Chimenti, A. Granese, O. Befani,
P. Turini, S. Alcaro, F. Ortuso, M.C. Cardia, S. Distinto, J. Med. Chem. 50 (4) (2007)
707–712.
[41] B. Evranos-Aksöz, S. Yabanoğlu-Çiftçi, G. Uçar, K. Yelekçi, R. Ertan, Bioorg. Med.
Chem. Lett. 24 (2014) 3278–3284.
[25] F. Chimenti, E. Maccioni, D. Secci, A. Bolasco, P. Chimenti, A. Granese, S. Carradori,
S. Alcaro, F. Ortuso, M. Yáñez, F. Orallo, R. Cirilli, R. Ferretti, F. La Torre, J. Med.
Chem. 51 (16) (2008) 4874–4880.
[42] 1st ed., Elsevier, Burlington, MA, USA, 2011, pp. 47–58.
[43] International Organization for Standardization, Biological Evaluation of Medical
Devices-part 5: Tests for in Vitro Cytotoxicity ISO-10993-5, third ed., 2009.
[44] H. Van de Waterbeemd, E. Gifford, Nat. Rev. Drug Discov. 2 (3) (2003) 192–204.
[45] C.A. Lipinski, F. Lombardo, B.W. Dominy, P.J. Feeney, Adv. Drug Deliv. Rev. 46
(2001) 3–26.
[26] F. Chimenti, D. Secci, A. Bolasco, P. Chimenti, A. Granese, S. Carradori, E. Maccioni,
M.C. Cardia, M. Yáñez, F. Orallo, S. Alcaro, F. Ortuso, R. Cirilli, R. Ferretti,
S. Distinto, J. Kirchmair, T. Langer, Bioorg. Med. Chem. 18 (14) (2010) 5063–5070.
[27] F. Chimenti, A. Bolasco, D. Secci, P. Chimenti, A. Granese, S. Carradori, M. Yáñez,
F. Orallo, F. Ortuso, S. Alcaro, Bioorg. Med. Chem. 18 (15) (2010) 5715–5723.
[46] W.L. Jorgensen, E.M. Duffy, Adv. Drug Deliv. Rev. 54 (3) (2002) 355–366.
108