Acetylcholinesterase inhibition activity of some quinolinyl substituted triazolothiadiazole derivatives

Article abstract of DOI:10.1134/S1068162015020089

A series of aralkanoic acids was converted into aralkanoic acid hydrazides through their esters formation. The aralkanoic acid hydrazides upon treatment with carbon disulfide and methanolic potassium hydroxide yielded potassium dithiocarbazinate salts, which on refluxing with aqueous hydrazine hydrate yielded 5-aralkyl-4-amino-3-mercapto-1,2,4-triazoles. The target compounds, 3-aralkyl-6-(substitutedquinolinyl) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazoles, were synthesized by condensing various quinolinyl substituted carboxylic acids with 5-aralkyl-4-amino-3-mercapto-1,2,4-triazoles in phosphorus oxychloride. The structures of the newly synthesized triazolothiadiazoles were characterized by IR, ¹H NMR, ¹³C NMR, and elemental analysis studies. The structure of one of the 5-aralkyl-4-amino-3-mercapto-1,2,4-triazoles was unambiguously deduced by single crystal X-ray diffraction analysis. All the synthesized compounds were screened for their acetylcholinesterase inhibition activities. Four of the triazolothiadiazoles exhibited excellent acetylcholinesterase inhibition activities as compared to the reference inhibitor.

Full text of DOI:10.1134/S1068162015020089

ISSN 1068ꢀ1620, Russian Journal of Bioorganic Chemistry, 2015, Vol. 41, No. 2, pp. 170–177. © Pleiades Publishing, Ltd., 2015.
Acetylcholinesterase Inhibition Activity of Some Quinolinyl
Substituted Triazolothiadiazole Derivatives1
Muhammad Rafiqa, Muhammad Saleemb, Muhammad Hanifc, Qamar Abbasa,
, 2
Ki Hwan Leeb, and SungꢀYum Seoa
a Department of Biology, Kongju National University, Kongju, 314ꢀ701 Korea
b Department of Chemistry, Kongju National University, Kongju, 314ꢀ701 Korea
c Department of Chemistry, QuaidꢀiꢀAzam University, Islamabad, 45320 Pakistan
Received May 12, 2014; in final form, August 28, 2014
Abstract—A series of aralkanoic acids was converted into aralkanoic acid hydrazides through their esters forꢀ
mation. The aralkanoic acid hydrazides upon treatment with carbon disulfide and methanolic potassium
hydroxide yielded potassium dithiocarbazinate salts, which on refluxing with aqueous hydrazine hydrate
yielded 5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles. The target compounds, 3ꢀaralkylꢀ6ꢀ(substitutꢀ
edquinolinyl) [1,2,4]triazolo[3,4ꢀb][1,3,4]thiadiazoles, were synthesized by condensing various quinolinyl
substituted carboxylic acids with 5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles in phosphorus oxychloride.
The structures of the newly synthesized triazolothiadiazoles were characterized by IR, 1H NMR, 13C NMR,
and elemental analysis studies. The structure of one of the 5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles
was unambiguously deduced by single crystal Xꢀray diffraction analysis. All the synthesized compounds were
screened for their acetylcholinesterase inhibition activities. Four of the triazolothiadiazoles exhibited excelꢀ
lent acetylcholinesterase inhibition activities as compared to the reference inhibitor.
Keywords: quinolinyl substituted triazolothiadiazole, Xꢀray diffraction analysis, acetylcholinesterase inhibition
assay
DOI: 10.1134/S1068162015020089
2 1
INTRODUCTION
The role of cholinergic system has been an intenꢀ
sive issue of interest in Alzheimer disease, which is a
neurodegenerative disorder causing deterioration of
memory and other cognitive functions [5, 6]. In Alzheꢀ
imer’s disease, a cholinergic deficiency in the brain has
been reported [7]. Therefore, the synthesis and study
of inhibitors of acetylcholinesterase may aid to the
development of therapeutically useful compounds to
treat such neurological disorders. Acetylcholinestꢀ
erase inhibitors donepezil hydrochloride, galantamine
hydrobromide, and rivastigmine tartrate are the curꢀ
rent approved drugs for the treatment of Alzheimer’s
patients [8]. However, acetylcholinesterase inhibitors
present some limitations, such as their short halfꢀlives
and excessive side effects caused by activation of
peripheral cholinergic systems, as well as hepatotoxicꢀ
ity, which is the most frequent and important side
effects of these drug therapies [9–11]. For this reason,
alternative and complementary therapies need to be
developed. The present work includes the synthesis of
new series of biheterocycles containing triazole and
thiadiazole ring coupled with substituted quinolinyl
carboxylic acids that show good acetylcholinesterase
inhibition activity.
Acetylcholinesterase is a serine hydrolase (AChE,
acetylcholine hydrolase, EC 3.1.1.7) that plays an
essential role in the cholinergic synapses. Hydrolysis
of the neurotransmitter acetylcholine in the nervous
system by acetylcholinesterase is known to be one of
the most efficient enzyme catalytic reactions [1]. The
basis of this high efficiency has been sought by means
of ligandꢀbinding studies using various substrates and
has led to the suggestion that the active center is comꢀ
posed of a cationic esteratic subsite containing the
active serine, an anionic site, which accommodates
the choline moiety of acetylcholine, and a peripheral
anionic site (PAS) [2]. The primary physiologic that
the active center is composed of a cationic esteratic
subsite containing the active serine, an anionic site,
which accommodates the choline moiety of acetylꢀ
choline, and a peripheral anionic site (PAS) [2]. The
primary physiologic role of the acetylcholinesterase
peripheral site is to accelerate the hydrolysis of acetylꢀ
choline at low substrate concentrations [3, 4].
1
The article is published in the original.
Corresponding author: eꢀmail: dnalove@kongju.ac.kr.
2
170

ACETYLCHOLINESTERASE INHIBITION ACTIVITY
171
RESULTS AND DISCUSSION
analysis. Condensation of the 5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀ
mercaptoꢀ1,2,4ꢀtriazoles (Va ) with quinolinyl subꢀ
stituted carboxylic acids in phosphorus oxychloride
resulted in the synthesis of target compounds,
3ꢀaralkylꢀ6ꢀ(substitutedquinolinyl)[1,2,4]triazolo[3,4ꢀ
–h
Synthetic pathway adopted for the synthesis of
5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles (Va
and 3ꢀaralkylꢀ6ꢀ(substitutedquinolinyl)[1,2,4]triazoꢀ
lo[3,4ꢀ ][1,3,4]thiadiazoles (VIa ) is illustrated in
Scheme 1. Briefly, substituted aralkanoic esters (IIa
were synthesized by the reaction of corresponding
substituted aralkanoic acid (Ia ) in the presence of
catalytic amount of sulfuric acid. The synthesized esꢀ
ters (IIa ) were converted into corresponding
aralkanoic acid hydrazides (IIIa ) by refluxing with
–h)
b
–v
b][1,3,4]thiadiazoles (VIa–v). Formation of triazoꢀ
–h)
lothiadiazoles (VIa ) was deduced from the IR specꢀ
–v
tral data by the disappearance of signals typical to NH
absorption for the primary and secondary NH group
–h
1
attached to triazole moiety. In the H NMR spectra,
the disappearance of singlet peaks for NH and NH2
protons of 5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriaꢀ
–h
–h
hydrazine hydrate (80%) in methanol; the reaction of
hydrazides with carbondisulfide in methanolic potasꢀ
sium hydroxide at low temperature yielded an interꢀ
zoles (Va
Condensation of quinolinyl carboxylic acids with
5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles (Va
–h) confirms the ring closure of thiadiazole.
–h)
mediate, potassium dithiocarbazinate (IVa
on reflux with aqueous hydrazine gave 5ꢀaralkylꢀ4ꢀ
aminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles (Va ). Structures
of
Va
The tautomeric structureof one of the triazoles (Vf
–h), which
was further confirmed by increased number of signals
1
in the aromatic region in both H NMR and
–h
5ꢀaralkylꢀ4ꢀaminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles 13C NMR spectra of 3ꢀaralkylꢀ6ꢀ(substitutedquinoliꢀ
(
–h
) were confirmed by IR and NMR spectroscopy. nyl)ꢀ[1,2,4]triazolo[3,4ꢀ
b][1,3,4]thiadiazoles (VIa–v).
Experimental values of elemental analysis were in
)
was determined using single crystal Xꢀray diffraction good aggrement with the calculated ones.
S
H
H
OH
N
O
N
n
i
ii
iii
n
n
N
S K
n
NH2
H
O
O
O
O
X
X
X
X
(Ia
–h)
(IIa–h
)
(
IIIa
–h)
(IVa–h
)
iv
I
⎯
Va, X = 4ꢀBr
b, X = 3ꢀBr
c, X = 4ꢀOCH3
d, X = 3ꢀOCH3
e, X = 2ꢀOCH3
f, X = 2ꢀF
n
= 1
= 1
= 1
= 1
= 1
= 1
= 2
= 1
n
n
n
n
n
n
n
NH2
X
N
N
N
n
v
S
N
n
N
S
NH
X
N
g, X = 3ꢀF
h, X = 4ꢀOCH3
Y
(
VIa
= 1 l, X = 3ꢀOCH3
= 1 m, X = 2ꢀOCH3 Y = 3ꢀQuinolinyl
= 1 n, X = 2ꢀF Y = 4ꢀQuinolinyl
= 1 o, X = 4ꢀOCH3 Y = 4ꢀQuinolinyl
= 1 p, X = 4ꢀBr
= 1 q, X = 2ꢀF
= 1 r, X = 4ꢀOCH3 Y = 5ꢀQuinolinyl
–
v
)
(Va–h)
VIa, X = 4ꢀBr
b, X = 3ꢀBr
Y = 2ꢀQuinolinyl
Y = 2ꢀQuinolinyl
n
Y = 3ꢀQuinolinyl
n
= 1
= 1
= 1
= 1
= 1
= 1
= 1
= 1
n
n
n
n
n
n
n
n
n
n
n
n
n
n
c, X = 4ꢀOCH3 Y = 2ꢀQuinolinyl
d, X = 3ꢀOCH3 Y = 2ꢀQuinolinyl
e, X = 2ꢀOCH3 Y = 2ꢀQuinolinyl
f, X = 2ꢀF
g, X = 3ꢀF
Y = 4ꢀQuinolinyl
Y = 5ꢀQuinolinyl
Y = 2ꢀQuinolinyl
Y = 2ꢀQuinolinyl
h, X = 4ꢀOCH3 Y = 3ꢀQuinolinyl
= 2 s, X = 4ꢀBr
= 1 t, X = 2ꢀF
= 1 u, X = 4ꢀOCH3 Y = 6ꢀQuinolinyl
Y = 5ꢀQuinolinyl
Y = 6ꢀQuinolinyl
i, X = 4ꢀBr
j, X = 2ꢀF
k, X = 3ꢀF
Y = 3ꢀQuinolinyl
Y = 3ꢀQuinolinyl
Y = 3ꢀQuinolinyl
n
n
n
n
n
n
= 1
= 1
= 1
= 1 v, X = 4ꢀBr Y = 6ꢀQuinolinyl
Scheme 1. Synthesis of 3ꢀaralkylꢀ6ꢀ(substitutedquinolinyl)[1,2,4]triazolo[3,4ꢀ
VIa–v). Reagents and conditions: ( ) H SO (conc.), methanol, reflux, 8–10 h; (ii) Hydrazine
hydrate (80%), methanol, reflux, 10–12 h; (iii) CS ,KOH, methanol, stirring, 0
b][1,3,4]thiadiazoles
(
i
2
4
°C, 1 h; (iv) Hydrazine
2
hydrate (80%), methanol, reflux, 10–12 h; (v) POCl ,substituted quinolinyl carboxylic acid, reflux, 4–6 h.
3
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 41
No. 2
2015

172
MUHAMMAD RAFIQ et al.
cause the labile hydrogen can be attached to the
N1
N2
nitrogen or sulfer atom, as shown in the thiol–thione
tautomeric forms below. In order to investigate the
major contributing structure, we have selected comꢀ
pound (Vf) for single crystal XRDꢀanalysis. The results
indicated that triazole derivative predominantly existed
in thione conformation as shown in (Figs. 1, 2).
C7 C8
C6
F1
C9
C5
N3
N4
C1
C4
C2
S1
C3
R
N
R
R
N
R
S
SH
Fig. 1. The molecular structure of one of the triazole molꢀ
ecules (Vf) with the atomꢀnumbering scheme. Displaceꢀ
ment ellipsoids are drawn at the 50% probability level.
N
N
N
N
H
(a
)
(b)
Scheme 2.
Crystal data. C9H9FN4S, Mw = 224.26, triclinic,
space group ꢀ1, = 6.54460(10), = 6.74380(10),
= 11.0531(2) = 100.2400(10)°,
= 96.1280(10) = 2,
3 at 100 K,
(000) = 232.
P
a
b
β
c
Å,
α
= 96.6600(10)°,
γ
°
,
V
= 472.758(13)
Å
Z
Dx = 1.575 mg/m3,
µ
= 0.326 mm–1,
F
Acetylcholinesterase Inhibition Activity
All the synthesized triazolothiadiazoles (VIa–v)
were evaluated for acetylcholinesterase inhibition acꢀ
tivity (table). Neostigmine methyl sulfate with IC50
value of 2.42 0.01 M was used as a reference inhibꢀ
itor. Almost all compounds exhibit moderate to excelꢀ
lent activities as compared to the reference inhibitor.
Compound (VIi) with IC50 value of 0.89 0.03
µ
Fig. 2. Packing diagram of compound (Vf) showing interꢀ
molecular hydrogen bonding.
µ
M
showed maximum acetylcholinesterase inhibition acꢀ
tivity among the whole synthesized series. It has 4ꢀ
bromobenzyl group as substituent X and 3ꢀquinolinyl
Crystal Structure Determination of Compound (Vf
)
group as substitent Y. Compounds (VIk) and (VIs
with IC50 values of 1.13 0.08 and 2.08 0.03
showed excellent inhibition exceeding that of the stanꢀ
ꢀ1,2,4ꢀtriazꢀ dard inhibitor. Compound (VIn) with IC50 values of
2.89 0.06 M showed comparable activity to the refꢀ
)
M
For the disubstituted 1,2,4ꢀtriazoles, two tautoꢀ
µ
meric forms (i.e., 4,5ꢀdisubstitutedꢀ4Hꢀ1,2,4ꢀtriazꢀ
oleꢀ3ꢀthiol (a) and 3,4ꢀdisubstitutedꢀ1
H
µ
oleꢀ5(4H)ꢀthione (b)) are theoretically possible beꢀ
erence inhibitor. These compounds possesed 3ꢀfluoꢀ
robenzyl, 2ꢀfluorobenzyl, and 4ꢀbromobenzyl groups
as substituent X and 3ꢀquinolinyl, 5ꢀquinolinyl, and
4ꢀquinolinyl groups as substituent Y, respectively.
Compound (VIv) with 4ꢀbromo group as substitent X
and 6ꢀquinolinyl group as substitent Y with IC50 values
Results of acetylcholinesterase inhibition assay of triazoꢀ
lothiadiazole derivatives (VIa
–v)
Compounds IC50,
µ
M
Compounds IC50, µM
(
(
(
(
(
VIa
VIb
VIc
VId
)
)
)
)
80.31 2.8
41.00 0.32
71.30 3.0
25 0.81
38.29 1.71
34.22 0.82
43 0.65
37.40 1.76
0.89 0.03
29.62 1.8
1.13 0.08
2.42 0.01 Neostigꢀ
mine methyl
sulfate
(
VIl
)
)
)
)
)
)
)
)
63.42 0.71
75 3.10
of 96.46 4.12
erase inhibition activity. The rest of the compounds
showed moderate activities with IC50 values in the
range of 25 0.81 to 86.67 3.33 M.
µ
M showed minimum acetylcholinestꢀ
(VIm
(
(
(
(
VIn
VIo
VIp
VIq
2.89 0.06
52.91 1.6
33.54 0.78
67.40 2.3
27.23 0.04
2.08 0.03
45.86 0.76
86.67 3.33
96.46 4.12
2.42 0.01
µ
VIe
)
)
(
VIf
EXPERIMENTAL
(VIg
)
)
)
(
(
(
VIr
VIs
VIt)
(
VIh
All the common solvents and chemicals were of anꢀ
alytical grade or dry distilled. The qualitative analysis
of the synthesized compounds was ascertained by thin
layer chromatography and the Rf values were deterꢀ
mined by employing precoated silica gel aluminium
plates, Kieslgel 60 F254 from Merck (Germany), usꢀ
ing chloroform–methanol, 9 : 1, as an eluent; TLC
(
VIi
VIj
VIk
(
(
)
(VIu
)
)
)
(VIv
Neostigꢀ
mine methyl
sulfate
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 41
No. 2
2015

ACETYLCHOLINESTERASE INHIBITION ACTIVITY
173
was visualized under UV lamp (VLꢀ4. LC, France) at ring. Carbon disulfide (0.125 mol) was added slowly
254 and 365 nm. Melting points were determined on a dropwise and the reaction mixture was stirred for 1 h at
Stuart melting point apparatus (SMP3) and are uncorꢀ
0
°
C
. The solid product of potassium dithiocarbaziꢀ
rected. The IR spectra (
υ
, cm–1) were recorded on a
nates (IVa ) formed was filtered, washed with chilled
diethyl ether, and dried. It waws directly used for the
next step without further purification. The potassium
–h
Bruker Optics Alpha FTꢀIR spectrophotometer.
NMR spectra were recorded in CDCl3 on a Bruker
Avance 300 MHz spectrometer with TMS as an interꢀ
dithiocarbazinates (IVa–h) were taken in 100 mL
round bottom flask with distilled water (20 mL),
hydrazine hydrate (0.250 mol), and refluxed overꢀ
night. The reaction mixture turned green with evoluꢀ
tion of hydrogen sulphide and finally it became homoꢀ
geneous. It was then poured in ice and neutralized
with concentrated hydrochloric acid. The white preꢀ
cipitate was filtered, washed with cold water, and crysꢀ
tallized from aquous methanol to afford 5ꢀaralkylꢀ4ꢀ
nal standard and reported in , ppm. The multipliciꢀ
δ
ties were expressed as s = singlet, d = doublet, t = tripꢀ
let. Single crystal data was obtained by using Bruker
SMART APEX II Xꢀray Diffractometer CCD
equipped with MO Xꢀray tube and graphite monoꢀ
chromator at the ambient temperature. Elemental
analysis was performed on Leco CHNSꢀ932 Elemenꢀ
tal Analyzer, Leco Corporation (USA).
aminoꢀ3ꢀmercaptoꢀ1,2,4ꢀtriazoles (Va –h).
General Procedure for the Synthesis of Substituted
Aromatic Esters (IIa
–h)
General Procedure for the Synthesis of 3ꢀAralkylꢀ6ꢀ
(Substituted Quinolinyl)[1,2,4]triazolo[3,4ꢀ
Aralkanoic acid (Ia 0.2 mol) was dissolved in
–h) (
b][1,3,4]thiadiazoles (VIa–v)
methanol (50 mL) in a round bottom flask equipped
with a reflux condenser and a calcium chloride drying
tube. Concentrated sulfuric acid (0.002 mol) was addꢀ
ed and the reaction mixture subjected to reflux for 8–
10 hours while monitored by thin layer chromatograꢀ
phy using chloroform–methanol, 9 : 1, solvent system.
After completion of the reaction, the excess alcohol
was removed under reduced pressure and resulting oil
was poured into water. The oily layer was separated and
the aqueous portion extracted with diethyl ether
A mixture of 4ꢀaminoꢀ5ꢀaralkyl [1,2,4]triazoleꢀ3ꢀ
thione (1 mmol) and substituted quinolinyl carboxylic
acids (1.1 mmol) in POCl3 (8–10 mL) was refluxed for
4–6 hours. The reaction mixture was slowly poured in
crushed ice with stirring and neutralized with
sodium hydrogen carbonate. Solid material was filꢀ
tered, washed with cold water, and dried to furnish
3ꢀaralkylꢀ6ꢀ(substitutedquinolinyl)[1,2,4]triazolo[3,4ꢀ
b
][1,3,4]thiadiazoles (VIa
3ꢀ(4ꢀBromobenzyl)ꢀ6ꢀ(quinolinꢀ2ꢀyl)ꢀ[1,2,4]triaꢀ
zolo[3,4ꢀ ][1,3,4]thiadiazole (VIa). Light brown solꢀ
id; yield 56%; mp 140–142
–v) as colored powder [12].
(3
×
50 mL). The combined organic layer was washed
with a dilute solution of sodium hydrogen carbonate
(100 mL) to remove unreacted sulfuric acid. The orꢀ
ganic layer was dried over anhydrous sodium sulphate.
The solvent was removed on a rotary evaporator after
b
°
C; Rf 0.71 (chloroform–
methanol, 9 : 1); IR: 3061 (sp2 CH), 2927, 2829 (sp3
CH), 1590 (C=N), 1502, 1478, 1471 (C=C), 1056
(C–S); 1H NMR (300 MHz): 8.36 (d, 1H,
filtration to afford corresponding esters (IIa
–h) as oily
J
= 8.4 Hz,
liquids.
ArꢀH), 8.19 (d, 2H, J = 8.4 Hz, ArꢀH), 7.96–7.90 (m,
1H, ArꢀH), 7.88–7.81 (m, 1H, ArꢀH), 7.74–7.66 (m,
General Procedure for the Synthesis of Aralkanoic Acid
Hydrazides (IIIa
1H, ArꢀH), 7.54–7.47 (m, 2H, ArꢀH), 7.38 (d, 2H,
13
–h)
J
= 8.4 Hz, ArꢀH), 4.49 (s, 2H, CH2); C NMR
The respective substituted aromatic esters (IIa–h)
(75 MHz): 163.21, 159.12, 147.14, 146.32, 143.21,
140.12, 143.21, 133.95, 131.95, 130.93, 129.75,
129.34, 128.82, 127.88, 121.37, 117.34, 30.82; Anal.
calcd. for C19H12BrN5S: C, 54.04; H, 2.86; N, 16.58;
S, 7.59; Found: C, 53.94; H, 2.80; N, 16.51; S, 7.45.
(0.02 mol) were dissolved in methanol (100 mL) in a
round bottom flask fitted with a reflux condenser and
a calcium chloride drying tube. Hydrazine hydrate
(80%, 0.04 mol) was added slowly, and the reaction
was monitored by thin layer chromatography. After
completion of the reaction, the mixture was concenꢀ
trated under reduced pressure. The resulting crude
solid was filtered, washed with water, and crystallized
from aqueous ethanol to get aralkanoic acid
3ꢀ(3ꢀBromobenzyl)ꢀ6ꢀ(quinolinꢀ2ꢀyl)ꢀ[1,2,4]triꢀ
azolo[3,4ꢀ
solid; yield 56%; mp 196–198
b
][1,3,4]thiadiazole (VIb). Magenta color
°
C; Rf 0.58 (chloroꢀ
form–methanol, 9 : 1); IR: 3035 (sp2 CH), 2939, 2899
(sp3 CH), 1588 (C=N), 1563, 1502, 1485 (C=C),
1032 (C–S); 1H NMR (300 MHz): 8.63–8.59 (m, 1H,
ArꢀH), 8.44–8.40 (m, 1H, ArꢀH), 8.39–8.36 (m, 1H,
ArꢀH), 8.34–8.29 (m, 1H, ArꢀH), 8.23 (d, 1H,
hydrazides (IIIa–h).
General Procedure for the Synthesis of 5ꢀAralkylꢀ4ꢀ
Aminoꢀ3ꢀMercaptoꢀ1,2,4ꢀTriazoles (Va
–h)
J
= 8.7 Hz, ArꢀH), 7.97–7.92 (m, 1H, ArꢀH), 7.89–
7.81 (m, 1H, ArꢀH), 7.52–7.43 (m, 1H, ArꢀH), 4.03
13
The aralkanoic acid hydrazides (IIIa ) (0.01 mol)
were treated with a solution of potassium hydroxide
(0.125 mol) in methanol (50 mL) at
–h
(s, 2H, CH2); C NMR (75MHz): 170.18, 155.71,
0°C
under stirꢀ 147.65, 147.09, 145.20, 137.76, 133.32, 131.02,
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 41
No. 2
2015

174
MUHAMMAD RAFIQ et al.
130.56, 129.73, 129.19, 128.94, 127.93, 127.50, CH), 1593 (C=N), 1502, 1478, 1473 (C=C), 996 (C–
S); 1H NMR (300 MHz): 8.34 (d, 1H,
H), 8.22–8.14 (m, 2H, ArꢀH), 7.91 (d, 1H,
125.33, 124.83, 122.99, 117.41, 30.08.
J
= 8.7 Hz, Arꢀ
= 8.1 Hz,
J
3ꢀ(4ꢀMethoxybenzyl)ꢀ6ꢀ(quinolinꢀ2ꢀyl)ꢀ[1,2,4]
ArꢀH), 7.87–7.78 (m, 1H, ArꢀH), 7.73–7.64 (m, 1H,
ArꢀH), 7.47–7.40 (m, 1H, ArꢀH), 7.34–7.24 (m, 1H,
ArꢀH), 7.19–7.10 (m, 2H, ArꢀH), 4.58 (s, 2H, CH2);
13C NMR (75 MHz): 169.44, 162.51, 159.23, 154.61,
150.60, 146.00, 137.58, 135.31, 133.82, 131.03,
130.96, 129.30, 128.74, 127.86, 124.42, 121.95,
117.41, 115.74, 24.25; Anal. calcd. for C19H12FN5S:
C, 63.15; H, 3.35; N, 19.38; S, 8.87; Found: C, 63.08;
H, 3.25; N, 19.35; S, 8.77.
triazolo[3,4ꢀ
yield 65%; mp 197–199
methanol, 9 : 1); IR: 3028 (sp2 CH), 2910, 2832 (sp3
CH), 1578 (C=N), 1512, 1498, 1473 (C=C), 1039
(C–S); 1H NMR (300 MHz): 8.23–8.15 (m, 2H, Arꢀ
b
][1,3,4]thiadiazole (VIc). Brown solid;
°
C; Rf 0.67 (chloroform–
H), 7.92 (d, 1H,
1H, ArꢀH), 7.72–7.64 (m, 1H, ArꢀH), 7.43 (d, 2H,
= 8.7 Hz, ArꢀH), 7.34 (m, 1H, ArꢀH), 6.90 (d, 2H,
= 8.7 Hz, ArꢀH), 4.47 (s, 2H, CH2), 3.79 (s, 3H,
J = 8.1 Hz, ArꢀH), 7.88–7.78 (m,
J
J
OCH3); 13C NMR (75 MHz): 169. 35, 158.80, 154.49,
147.61, 147.44, 146.31, 137.55, 130.87, 130.11,
129.73, 129.30, 128.74, 127.86, 127.01, 117.39,
114.16, 55.30, 30.54; Anal. calcd. for C20H15N5OS:
C, 64.33; H, 4.05; N, 18.75; S, 8.59; Found: C, 64.23;
H, 4.01; N, 18.65; S, 8.49.
3ꢀ(3ꢀFluorobenzyl)ꢀ6ꢀ(quinolinꢀ2ꢀyl)ꢀ[1,2,4]triaꢀ
zolo[3,4ꢀ
yield 67%; mp 199–201
b
][1,3,4]thiadiazole (VIg). Off white solid;
°C; Rf 0.62 (chloroform–
methanol, 9 : 1); IR: 3022 (sp2 CH), 2900, 2819 (sp3
CH), 1588 (C=N), 1543, 1535, 1501, 1492 (C=C),
995 (C–S); 1H NMR (300 MHz): 8.63–8.59 (m, 1H,
ArꢀH), 8.44–8.40 (m, 1H, ArꢀH), 8.39–8.36 (m, 1H,
ArꢀH), 8.34–8.23 (m, 2H, ArꢀH), 8.23 (d, 1H,
3ꢀ(3ꢀMethoxybenzyl)ꢀ6ꢀ(quinolinꢀ2ꢀyl)ꢀ[1,2,4]
triazolo[3,4ꢀ
yield 76%; mp 214–215
b
][1,3,4]thiadiazole (VId). Brown solid;
J
= 8.7 Hz, ArꢀH), 7.39–7.32 (m, 1H, ArꢀH), 7.12–
°C; Rf 0.66 (chloroform–
13
methanol, 9 : 1); IR: 3032 (sp2 CH), 2923, 2852 (sp3
CH), 1584 (C=N), 1553, 1539, 1500, 1482 (C=C),
1070 (C–S); 1H NMR (300 MHz): 8.63–8.59 (m, 1H,
ArꢀH), 8.44–8.40 (m, 1H, ArꢀH), 8.39–8.36 (m, 1H,
ArꢀH), 8.34–8.29 (m, 1H, ArꢀH), 8.23 (d, 1H,
7.06 (m, 3H, ArꢀH), 4.25 (s, 2H, CH2); C NMR
(75MHz): 170.18, 158.71, 152.65, 150.62, 147.09,
144.20, 139.76, 137.46, 134.02, 130.56, 131.73,
129.19, 128.94, 127.93, 127.50, 124.83, 123.07,
117.41, 25.60; Anal. calcd. for C19H12FN5S: C, 63.15;
H, 3.35; N, 19.38; S, 8.87; Found: C, 63.09; H, 3.34;
N, 19.32; S, 8.81.
J
= 8.7 Hz, ArꢀH), 7.97–7.92 (m, 1H, ArꢀH), 7.89–
7.81 (m, 1H, ArꢀH), 7.74–7.65 (m, 2H, ArꢀH), 7.52–
7.43 (m, 1H, ArꢀH), 4.25 (s, 2H, CH2), 3.21 (s, 3H,
3ꢀ(4ꢀMethoxyphenethyl)ꢀ6ꢀ(quinolinꢀ3ꢀyl)ꢀ[1,2,4]
OCH3); 13C NMR (75MHz): 170.18, 155.71, 152.65,
147.09, 145.20, 137.76, 133.32, 131.02, 130.56,
129.73, 129.19, 128.94, 127.93, 127.50, 124.83,
123.07, 122.95, 117.41, 55.56, 25.60; Anal. calcd. for
C20H15N5OS: C, 64.33; H, 4.05; N, 18.75; S, 8.59;
Found: C, 64.24; H, 4.01; N, 18.65; S, 8.51.
triazolo[3,4ꢀ
yield 58%; mp 221–224
b
][1,3,4]thiadiazole (VIh). Brown solid;
°
C; Rf 0.63 (chloroform–
methanol, 9 : 1); IR: 3012 (sp2 CH), 2929, 2856 (sp3
CH), 1579 (C=N), 1541, 1512, 1479 (C=C), 1013
(C–S); 1H NMR (300 MHz): 8.55–8.49 (m, 1H, Arꢀ
H), 8.24–8.17 (m, 1H, ArꢀH), 8.01–7.95 (m, 1H, Arꢀ
H), 7.94–7.85 (m, 1H, ArꢀH), 7.75–7.67 (m, 1H, Arꢀ
3ꢀ(2ꢀMethoxybenzyl)ꢀ6ꢀ(quinolinꢀ2ꢀyl)ꢀ[1,2,4]
H), 7.40–7.35 (m, 1H, ArꢀH), 7.40 (d, 2H,
ArꢀH), 6.92 (d, 2H, = 8.4 Hz, ArꢀH), 3.89 (s, 3H,
OCH3), 3.01 (t, 2H, = 8.4 Hz, CH2), 2.90 (t, 2H,
= 8.4 Hz, CH2); 13C NMR (75 MHz): 163.46,
J = 8.4 Hz,
triazolo[3,4ꢀ
b
][1,3,4]thiadiazole (VIe). Light yellow
J
solid; yield 65%; mp 203–205
°C; Rf 0.61 (chloroꢀ
form–methanol, 9 : 1); IR: 3006 (sp2 CH), 2969, 2827
J
(sp3 CH), 1589 (C=N), 1502, 1494, 1486 (C=C),
J
1
157.61, 159.38, 154.46, 147.03, 146.31, 135.28, 132.11,
128.81, 128.61, 128.37, 126.88, 123.16, 122.29, 120.60,
117.58, 55.55, 30.38, 25.66; Anal. calcd. for
C21H17N5OS: C, 65.10; H, 4.42; N, 18.08; S, 8.28;
Found: C, 65.00; H, 4.35; N, 18.01; S, 8.21.
1067 (C–S): H NMR (300 MHz): 8.34 (d, 1H,
J
= 8.7 Hz, ArꢀH), 8.22–8.14 (m, 2H, ArꢀH), 7.91–
7.28 (m, 1H, ArꢀH), 7.87–7.78 (m, 1H, ArꢀH), 7.73–
7.64 (m, 1H, ArꢀH), 7.47–7.40 (m, 1H, ArꢀH), 7.34–
7.24 (m, 1H, ArꢀH), 7.19–7.10 (m, 2H, ArꢀH), 4.58
(s, 2H, CH2), 3.01 (s, 3H, OCH3); 13C NMR
(75 MHz): 169.44, 159.23, 154.61, 150.60, 146.77,
146.00, 137.58, 135.74, 133.56, 131.03, 129.30,
128.74, 127.86, 124.42, 121.95, 120.56, 117.41,
115.74, 55.25, 24.25; Anal. calcd. for C20H15N5OS: C,
64.33; H, 4.05; N, 18.75; S, 8.59; Found: C, 64.28; H,
4.01; N, 18.63; S, 8.49.
3ꢀ(4ꢀBromobenzyl)ꢀ6ꢀ(quinolinꢀ3ꢀyl)ꢀ[1,2,4]triaꢀ
zolo[3,4ꢀ
66%; mp 170–173
b
][1,3,4]thiadiazole (VIi). Red solid; yield
°C; Rf 0.68 (chloroform–methaꢀ
nol, 9 : 1); IR: 3056 (sp2 CH), 2929, 2871 (sp3 CH),
1597 (C=N), 1541, 1488, 1477 (C=C), 1059 (C–S);
1H NMR (300 MHz): 8.60–8.39 (m, 1H, ArꢀH),
8.34–8.27 (m, 1H, ArꢀH), 8.11–7.99 (m, 1H, ArꢀH),
3ꢀ(2ꢀFluorobenzyl)ꢀ6ꢀ(quinolinꢀ2ꢀyl)ꢀ[1,2,4]triꢀ 7.94–7.84 (m, 1H, ArꢀH), 7.75–7.67 (m, 1H, ArꢀH),
azolo[3,4ꢀ ][1,3,4]thiadiazole (VIf). Brown solid; 7.40–7.35 (m, 1H, ArꢀH), 7.4 (d, 2H, = 8.4 Hz,
yield 61%; mp 150–152 C; Rf 0.58 (chloroform– ArꢀH), 6.92 (d, 2H, = 8.4 Hz, ArꢀH), 4.56 (s, 2H,
CH2); C NMR (75 MHz): 163.46, 159.38, 156.92,
b
J
°
J
13
methanol, 9 : 1); IR: 3053 (sp2 CH), 2927, 2840 (sp3
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 41
No. 2
2015

ACETYLCHOLINESTERASE INHIBITION ACTIVITY
175
147.55, 148.03, 139.28, 135.11, 131.76, 129.81, 128.61, solid; yield 66%; mp 203–205
°C; Rf 0.61 (chloroꢀ
form–methanol, 9 : 1); IR: 3006 (sp2 CH), 2969, 2851
(sp3 CH), 1589 (C=N), 1533, 1494, 1489, (C=C),
1011 (C–S): 1H NMR (300 MHz): 8.34–8.29 (m, 1H,
ArꢀH), 8.20–8.11 (m, 2H, ArꢀH), 7.91–7.84 (m, 1H,
ArꢀH), 7.87–7.78 (m, 1H, ArꢀH), 7.73–7.64 (m, 2H,
ArꢀH), 7.47–7.40 (m, 1H, ArꢀH), 7.34–7.10 (m, 2H,
ArꢀH), 4.58 (s, 2H, CH2), 3.01 (s, 3H, OCH3);
128.37, 126.88, 123.16, 122.29, 120.60, 117.58, 25.66;
Anal. calcd. for C19H12BrN5S: C, 54.04; H, 2.86; N,
16.58; S, 7.59; Found: C, 54.01; H, 2.80; N, 16.50; S,
7.40.
3ꢀ(2ꢀFluorobenzyl)ꢀ6ꢀ(quinolinꢀ3ꢀyl)ꢀ[1,2,4]triaꢀ
zolo[3,4ꢀ
61%; mp 150–152
b
][1,3,4]thiadiazole (VIj). Yellow solid; yield
°C; Rf 0.72 (chloroform–methaꢀ
13C NMR (75 MHz): 169.44, 159.23, 154.61, 152.60,
146.00, 145.36, 137.58, 135.36, 133.27, 131.03,
129.30, 128.74, 127.86, 124.42, 121.95, 120.98,
117.41, 115.74, 55.25, 24.25; Anal. calcd. for
C20H15N5OS: C, 64.33; H, 4.05; N, 18.75; S, 8.59;
Found: C, 64.28; H, 3.95; N, 18.66; S, 8.49.
nol, 9 : 1); IR: 3064 (sp2 CH), 2983, 2852 (sp3 CH),
1583 (C=N), 1542, 1543, 1492 (C=C), 1009 (C–S);
1H NMR (300 MHz): 8.55–8.49 (m, 1H, ArꢀH),
8.24–8.17 (m, 1H, ArꢀH), 8.01–7.95 (m, 1H, ArꢀH),
7.94–7.85 (m, 1H, ArꢀH), 7.75–7.67 (m, 1H, ArꢀH),
7.40–7.35 (m, 1H, ArꢀH), 7.34–7.25 (m, 2H, ArꢀH),
7.00–6.90 (m, 2H, ArꢀH), 4.56 (s, 2H, CH2);
13C NMR (75 MHz): 163.46, 157.38, 156.35, 155.46,
147.03,145.35, 135.28, 132.11, 130.52, 128.81,
128.61, 128.37, 126.88, 123.16, 122.29, 120.60,
116.58, 55.55, 25.66; Anal. calcd. for C19H12FN5S: C,
63.15; H, 3.35; N, 19.38; S, 8.87; Found: C, 63.08; H,
3.28; N, 19.31; S, 8.76.
3ꢀ(2ꢀFluorobenzyl)ꢀ6ꢀ(quinolinꢀ4ꢀyl)ꢀ[1,2,4]triaꢀ
zolo[3,4ꢀ
71%; mp 142–144
nol, 9 : 1); IR: 3033 (sp2 CH), 2922, 2843 (sp3 CH),
1581 (C=N), 1542, 1492 (C=C), 1019 (C–S); H
NMR (300 MHz): 8.84–8.67 (m, 1H, ArꢀH), 8.52 (d,
b][1,3,4]thiadiazole (VIn). White solid; yield
°C; Rf 0.61 (chloroform–methaꢀ
1
1H, J = 8.4 Hz, ArꢀH), 7.91–7.83 (m, 1H, ArꢀH),
7.75–7.68 (m, 1H, ArꢀH), 7.67–7.64 (m, 2H, ArꢀH),
3ꢀ(3ꢀFluorobenzyl)ꢀ6ꢀ(quinolinꢀ3ꢀyl)ꢀ[1,2,4]triaꢀ
7.37–7.27 (m, 1H, ArꢀH), 7.20–7.16 (m, 2H, ArꢀH),
13
zolo[3,4ꢀ
yield 58%; mp 206–208
b][1,3,4]thiadiazole (VIk). Brown powder;
°
C; Rf 0.59 (chloroform– 6.90–6.84 (m, 1H, ArꢀH), 4.61 (s, 2H, CH2): C
methanol, 9 : 1); IR: 3045 (sp2 CH), 2953, 2896 (sp3
CH), 1587 (C=N), 1543, 1519, 1496 (C=C), 1034
(C–S); 1H NMR (300 MHz): 8.63–8.59 (m, 1H, Arꢀ
H), 8.44–8.40 (m, 1H, ArꢀH), 8.39–8.36 (m, 1H, Arꢀ
H), 8.34–8.29 (m, 1H, ArꢀH), 8.24–8.17 (m, 1H,
NMR (75 MHz): 163.79, 162.58, 159.30, 154.93,
149.68, 149.15, 146.39, 133.65, 131.31, 131.26,
130.63, 130.51, 129.48, 129.00, 124.88, 123.62,
121.86, 115.87, 24.69; Anal. calcd. for C19H12FN5S:
C, 63.15; H, 3.35; N, 19.38; S, 8.87; Found: C, 63.09;
H, 3.29; N, 19.25; S, 8.71.
J
= 8.7 Hz, ArꢀH), 7.97–7.92 (m, 1H, ArꢀH), 7.39–
7.32 (m, 1H, ArꢀH), 7.12–7.06 (m, 3H, ArꢀH), 4.25
13
(s, 2H, CH2); C NMR (75MHz): 170.18, 155.71,
154.51, 153.65, 147.09, 145.20, 137.76, 133.32,
131.02, 130.56, 129.73, 129.19, 128.94, 127.93,
127.50, 124.83, 123.07, 117.41, 25.60; Anal. calcd. for
C19H12FN5S: C, 63.15; H, 3.35; N, 19.38; S, 8.87;
Found: C, 63.01; H, 3.18; N, 19.14; S, 8.64.
yield 65%; mp 150–152
°
C; Rf 0.72 (chloroform–
methanol, 9 : 1); IR: 3064 (sp2 CH), 2983, 2843 (sp3
CH), 1583 (C=N), 1542, 1543, 1492 (C=C), 1022
(C–S); 1H NMR (300 MHz): 8.64–8.57 (m, 1H, Arꢀ
H), 8.45 (d, 1H,
= 8.4 Hz, ArꢀH), 7.93–7.84 (m, 1H, ArꢀH), 7.76–
7.64 (m, 2H, ArꢀH), 7.40 (d, 2H, = 8.4 Hz, ArꢀH),
6.92 (d, 2H, = 8.4 Hz, ArꢀH), 4.50 (s, 2H, CH2),
J = 8.1 Hz, ArꢀH), 8.25 (d, 1H,
J
3ꢀ(3ꢀMethoxybenzyl)ꢀ6ꢀ(quinolinꢀ3ꢀyl)ꢀ[1,2,4]triꢀ
J
azolo[3,4ꢀ
solid; yield 59%; mp 230–232
b
][1,3,4]thiadiazole (VIl). Saddle brown
J
°C; Rf 0.62 (chloroꢀ
13
form–methanol, 9 : 1); IR: 3027 (sp2 CH), 2945, 2871
(sp3 CH), 1580 (C=N), 1533, 1509, 1480 (C=C),
1021 (C–S); 1H NMR (300 MHz): 8.63–8.59 (m, 1H,
ArꢀH), 8.44–8.40 (m, 1H, ArꢀH), 8.39–8.36 (m, 1H,
ArꢀH), 8.34–8.29 (m, 1H, ArꢀH), 8.24–8.17 (d, 1H,
3.81 (s, 3H, OCH3); C NMR (75MHz): 163.47,
158.91, 157.11, 152.97, 149.70, 149.08, 147.81,
133.80, 130.71, 130.49, 128.97, 126.69, 124.89,
123.67, 121.85, 114.21, 55.34, 30.93; Anal. calcd. for
C20H15N5OS: C, 64.33; H, 4.05; N, 18.75; S, 8.59;
Found: C, 64.25; H, 3.95; N, 18.69; S, 8.49.
J
= 8.7 Hz, ArꢀH), 7.97–7.92 (m, 1H, ArꢀH), 7.89–
7.81 (m, 1H, ArꢀH), 7.74–7.65 (m, 2H, ArꢀH), 7.52–
7.43 (m, 1H, ArꢀH), 4.25 (s, 2H, CH2), 3.10 (s, 3H,
3ꢀ(4ꢀBromobenzyl)ꢀ6ꢀ(quinolinꢀ4ꢀyl)ꢀ[1,2,4]triꢀ
azolo[3,4ꢀ
67%; mp 176–178
nol, 9 : 1); IR: 3054 (sp2 CH 8.40 (d 2869 (sp3 CH),
1577 (C=N), 1551, 1521, 1502, 1499 (C=C), 1034
(C–S); 1H NMR (300 MHz): 8.77–8.64 (m, 1H, ArꢀH),
b
][1,3,4]thiadiazole (VIp). Red solid; yield
OCH3); 13C NMR (75MHz): 170.18, 156.99, 155.71,
151.65, 147.09, 145.20, 137.76, 133.32, 131.02,
130.56, 129.73, 129.19, 128.94, 127.93, 127.50,
124.83, 123.07, 117.41, 55.25, 25.60; Anal. calcd. for
C20H15N5OS: C, 64.33; H, 4.05; N, 18.75; S, 8.59;
Found: C, 64.23; H, 4.01; N, 18.65; S, 8.49.
°C; Rf 0.64 (chloroform–methaꢀ
8.40 (d, 1H, J = 8.1 Hz, ArꢀH), 8.19 (d, 1H, J = 8.4 Hz,
ArꢀH), 8.17–8.09 (m, 1H, ArꢀH), 7.93–7.84 (m, 2H,
ArꢀH), 7.76–7.64 (m, 2H, ArꢀH), 6.97–6.84 (d, 2H,
3ꢀ(2ꢀMethoxybenzyl)ꢀ6ꢀ(quinolinꢀ3ꢀyl)ꢀ[1,2,4]
triazolo[3,4ꢀb][1,3,4]thiadiazole (VIm). Light yellow
13
J
= 8.4 Hz, ArꢀH), 4.50 (s, 2H, CH2); C NMR
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 41
No. 2
2015

176
MUHAMMAD RAFIQ et al.
(75 MHz): 164.46, 159.33, 157.12, 147.63, 146.48, CH), 1587 (C=N),1573, 1477, (C=C), 1069 (C–S);
1H NMR (300 MHz): 9.08–9.03 (m, 1H, ArꢀH),
8.35–8.21 (m, 1H, ArꢀH), 7.56–7.47 (m, 1H, ArꢀH),
7.48–7.40 (m, 1H, ArꢀH), 7.35–7.30 (m, 1H, ArꢀH),
7.30–7.24 (m, 1H, ArꢀH), 7.24–7.15 (m, 4H, ArꢀH),
4.58 (s, 2H, CH2); 13C NMR (75 MHz): 165.77,
162.51, 159.24, 152.73, 149.73, 146.28, 136.78,
131.19, 131.08, 129.36, 129.25, 127.59, 126.63,
124.42, 122.64, 122.05, 120.89, 115.50, 24.38; Anal.
calcd. for C19H12FN5S: C, 63.15; H, 3.35; N, 19.38; S,
8.87; Found: C, 63.09; H, 3.25; N, 19.31; S, 8.7.
140.11, 137.62, 133.95, 131.95, 130.93, 129.75,
129.34, 128.82, 127.88, 121.37, 117.34, 30.82; Anal.
calcd. for C19H12BrN5S: C, 54.04; H, 2.86; N, 16.58;
S, 7.59; Found: C, 54.01; H, 2.71; N, 16.49; S, 7.50.
3ꢀ(2ꢀFluorobenzyl)ꢀ6ꢀ(quinolinꢀ5ꢀyl)ꢀ[1,2,4]triazoꢀ
lo[3,4ꢀ
yield 65%; mp 188–190
b
][1,3,4]thiadiazole (VIq). Light brown solid;
°C; Rf 0.64 (chloroform–
methanol, 9 : 1); IR: 3043 (sp2 CH), 2938, 2871 (sp3
CH), 1587 (C=N), 1541, 1501, 1487 (C=C), 1042
(C–S); 1H NMR ( 300 MHz): 8.70–8.63 (m, 1H, ArꢀH),
8.52–8.46 (m, 1H, ArꢀH), 8.38–8.32 (m, 1H, ArꢀH),
7.96–7.91 (m, 1H, ArꢀH), 7.88–7.81 (m, 2H, ArꢀH),
7.60–7.54 (m, 1H, ArꢀH), 7.20–7.13 (m, 1H, ArꢀH),
7.01–6.94 (m, 2H, ArꢀH), 4.61 (s, 2H, CH2);
3ꢀ(4ꢀMethoxybenzyl)ꢀ6ꢀ(quinolinꢀ6ꢀyl)ꢀ[1,2,4]
triazolo[3,4ꢀ
yield 61%; mp 170–172
b
][1,3,4]thiadiazole (VIu). Yellow solid;
°C; Rf 0.69 (chloroform–
methanol, 9 : 1); IR: 3031 (sp2 CH), 2912, 2843 (sp3
CH), 1593 (C=N), 1520, 1486, (C=C), 1052 (C–S);
1H NMR (300 MHz): 9.00–8.94 (m, 1H, ArꢀH),
7.56–7.47 (m, 1H, ArꢀH), 7.48–7.40 (m, 1H, ArꢀH),
13C NMR (75 MHz): 165.06, 162.58, 159.31, 156.37,
148.29, 146.21, 133.69, 131.32, 129.97, 129.36,
128.65, 126.23, 125.33, 124.47, 123.07, 122.02,
121.81, 115.85, 24.65; Anal. calcd. for C19H12FN5S:
C, 63.15; H, 3.35; N, 19.38; S, 8.87; Found: C, 63.07;
H, 3.28; N, 19.27; S, 8.74.
7.36 (d, 2H,
7.20–7.10 (m, 2H, ArꢀH), 6.82 (d, 2H,
J
= 8.7 Hz, ArꢀH), 7.35–7.26 (m, 1H, ArꢀH),
= 8.7 Hz, ArꢀH),
J
13
4.58 (s, 2H, CH2), 3.82 (s, 3H, OCH3); C NMR
(75 MHz): 165.77, 162.51, 159.24, 152.73, 149.73,
147.48, 138.78, 134.19, 129.36, 129.25, 127.59,
126.63, 124.42, 122.37, 122.05, 115.79, 55.35, 24.38;
Anal. calcd. for C20H15N5OS: C, 64.33; H, 4.05; N,
18.75; S, 8.59; Found: C, 64.29; H, 3.95; N, 18.65; S,
8.51.
3ꢀ(4ꢀMethoxybenzyl)ꢀ6ꢀ(quinolinꢀ5ꢀyl)ꢀ[1,2,4]
triazolo[3,4ꢀ
yield 63%; mp 147–149
b
][1,3,4]thiadiazole (VIr). Brown solid;
°C; Rf 0.59 (chloroform–
methanol, 9 : 1); IR: 3030 (sp2 CH), 2969, 2836 (sp3
CH), 1583 (C=N), 1511, 1501, 1499 (C=C), 1066
(C–S); 1H NMR (300 MHz): 9.09–9.03 (m, 1H, ArꢀH),
8.84–8.77 (m, 1H, ArꢀH), 8.39–8.32 (m, 1H, ArꢀH),
7.94–7.88 (m, 1H, ArꢀH), 7.87–7.79 (m, 1H, ArꢀH),
3ꢀ(4ꢀBromobenzyl)ꢀ6ꢀ(quinolinꢀ6ꢀyl)ꢀ[1,2,4]triaꢀ
zolo[3,4ꢀ
yield 62%; mp 176–177
b
][1,3,4]thiadiazole (VIv). Light green solid;
°C; Rf 0.64 (chloroform–
7.55–7.47 (m, 1H, ArꢀH), 7.39 (d, 2H,
J = 8.7 Hz,
methanol, 9 : 1); IR: 3059 (sp2 CH), 2900, 2831 (sp3
CH), 1580 (C=N), 1561, 1509, 1497 (C=C), 1041
(C–S); 1H NMR (300 MHz): 8.83–8.77 (m, 1H, ArꢀH),
7.65–7.57 (m, 1H, ArꢀH), 7.48–7.40 (m, 1H, ArꢀH),
ArꢀH), 6.92 (d, 2H, = 8.7 Hz, ArꢀH), 4.21 (s, 2H,
J
13
CH2), 3.74 (s, 3H, OCH3); C NMR (75 MHz):
164.68, 158.88, 156.55, 155.40, 148.27, 147.58,
134.43, 130.33, 129.87, 126.34, 125.36, 125.32,
124.12, 122.99, 119.87, 114.14, 55.35, 30.98; Anal.
calcd. for C20H15N5OS: C, 64.33; H, 4.05; N, 18.75; S,
8.59; Found: C, 64.28; H, 4.01; N, 18.56; S, 8.49.
7.39 (d, 2H,
ArꢀH), 7.20–7.13 (m, 2H, ArꢀH), 6.82 (d, 2H,
= 8.7 Hz, ArꢀH), 4.58 (s, 2H, CH2); 13C NMR
J = 8.7 Hz, ArꢀH), 7.35–7.26 (m, 1H,
J
(75 MHz): 165.77, 162.51, 159.24, 152.73, 149.73,
147.48, 138.78, 134.19, 131.08, 130.65, 129.36,
129.25, 127.94, 127.59, 126.63, 124.42, 122.64,
119.15, 24.38; Anal. calcd. for C19H12BrN5S: C, 54.04;
H, 2.86; N, 16.58; S, 7.59; Found: C, 53.91; H, 2.81;
N, 16.48; S, 7.56.
3ꢀ(4ꢀBromobenzyl)ꢀ6ꢀ(quinolinꢀ5ꢀyl)ꢀ[1,2,4]triaꢀ
zolo[3,4ꢀ
yield 66%; mp 154–157
b
][1,3,4]thiadiazole (VIs). Brown solid;
°C; Rf 0.59 (chloroform–methꢀ
anol, 9 : 1); IR: 3068 (sp2 CH), 2933, 2826 (sp3 CH),
1589 (C=N), 1557, 1511, 1496 (C=C), 1054 (C–S); 1H
NMR (300 MHz): 8.84–8.77 (m, 1H, ArꢀH), 8.39–8.32
(m, 1H, ArꢀH), 7.90–7.84 (m, 1H, ArꢀH), 7.89–7.73
(m, 1H, ArꢀH), 7.70–7.65 (m, 1H, ArꢀH), 7.50–7.43
XRDꢀAnalysis
(m, 1H, ArꢀH), 7.35 (d, 2H,
J = 8.5 Hz, ArꢀH), 6.82 (d,
Data collection. A colorless crystal of 0.22
×
0.20
×
13
2H, = 8.5 Hz, ArꢀH), 4.31 (s, 2H, CH2); C NMR
J
0.19 mm3 was mounted on a glass fiber in inert oil and
transferred to the cold gas stream of the spectrophoꢀ
tometer. A total of 8741 intensities leading to 2358 inꢀ
dependent reflections were collected from theta range
(75 MHz): 166.04, 156.22, 155.91, 149.74, 146.77,
136.80, 133.84, 131.95, 131.23, 129.18, 127.94,
127.48, 126.59, 125.33, 121.43, 119.12, 30.88; Anal.
calcd. for C19H12BrN5S: C, 54.04; H, 2.86; N, 16.58;
S, 7.59; Found: C, 53.94; H, 2.81; N, 16.51; S, 7.50.
of data collections 1.89° to 28.41
matic radiation.
°, using monochroꢀ
K
α
3ꢀ(2ꢀFluorobenzyl)ꢀ6ꢀ(quinolinꢀ6ꢀyl)ꢀ[1,2,4]triaꢀ
Structure solution and refinemen. The molecular
structure was solved by routine direct methods and reꢀ
fined on F2 using the program SHELXLꢀ97 (Sheldꢀ
zolo[3,4ꢀ
b
][1,3,4]thiadiazole (VIt). Light yellow solꢀ
id; yield 62%; mp 140–142
°
C; Rf 0.63 (chloroform–
methanol, 9 : 1); IR: 3011 (sp2 CH), 2969, 2891 (sp3 rick, 2008). The final wR2 was 0.1076 for all reflections
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 41
No. 2
2015

ACETYLCHOLINESTERASE INHIBITION ACTIVITY
177
and 136 parameters with a conventional
S = 0.923, max. Δρ = –0.883 e Å–3
R
1 of 0.0395; “Leades IndustryꢀUniversity Cooperation” Project,
supported by the Ministry of Education, Science &
Technology (MEST) and the Basic Science Research
Program through the National Research Foundation
.
Acetylcholine Esterase Inhibition Assay Protocol
of Korea (NRF) funded by the Ministry of Education
(NRF–2011–0015056).
The inhibitory activities of newly synthesized comꢀ
pounds were determined spectrophotometrically usꢀ
ing acetylthiocholine iodide as substrate from the
method of Ellman [13]. The assay solution consisted
REFERENCES
1. Quinn, D.M., Chem. Rev., 1987, vol. 87, pp. 955–979.
of 180
ing 0.1 M sodium chloride and 0.02 M magnesium
chloride, and 20 L of enzyme (AChE, acetylcholine
hydrolase, EC 3.1.1.7, acetyl cholinesterase from huꢀ
man erythrocytes) solution (0.03 U/mL); increasing
concentrations of test compounds (10 L) were added
to the assay solution and preꢀincubated for 30 min at
C. 5,5'ꢀDithioꢀbis(2ꢀnitrobenzoic acid) (0.3 mM,
20 L) and acetylthiocholine iodide (1.8 mM, 20 L)
were added to the reaction mixture and incubated at
37 C for 10 min, followed by the measurement of abꢀ
µ
L of 50 mM TrisꢀHCl buffer, pH 8.0, containꢀ
2. Colletier, J.P., Fournier, D., Greenblatt, H.M., Stogan, J.,
Sussman, J.L., Zaccai, G., Silman, I., and Weik, M.,
EMBO J., 2006, vol. 25, pp. 2746–2756.
µ
3. Mallender, W.D., Szeglets, T., and Rosenberry, T.L.,
J. Biol. Chem., 1999, vol. 274, pp. 8491–8499.
µ
4. Szeglets, T., Mallender, W.D., Thomas, P.J., and
Rosenberry, T.L., Biochemistry, 1999, vol. 38, pp. 122–
133.
4°
µ
µ
5. Abramov, A.Y., Canvari, L., and Duchen, M.R.,
°
J. Neurosci., 2003, vol. 23, pp. 5088–5095.
sorbance at 412 nm. For nonꢀenzymatic reaction, the
assays were carried out with a blank containing all
components except acetyl cholinesterase. The assay
measurements were carried out by using a micro plate
reader (OPTIMax, Tunable Micro plate Reader; waveꢀ
length range 340–850 nm; for 96ꢀwell plates). The reꢀ
action rates were compared and the percent inhibition
due to the presence of tested inhibitors was calculated.
Neostigmine methylsulfate was used as reference inꢀ
hibitor. Each concentration was analyzed in three inꢀ
dependent experiments run in triplicate. The IC50 valꢀ
ues were determined by the data analysis and graphing
software, Origin 8.6, 64ꢀbit.
6. Butterfield, D.A., Chem. Res. Toxicol., 1997, vol. 10,
pp. 495–506.
7. Greenblatt, H.M., Dvir, H., Silman, I., and Sussman, J.L.,
J. Mol. Neurosci., 2003, vol. 20, pp. 369–383.
8. Ellis, J.M., J. Am. Osteopth. Assoc., 2005, vol. 105,
pp. 145–158.
9. Farlow, M., Gracon, S.I., Hershey, L.A., Lewis, K.W.,
Sadowsky, C.H., and Ureno, J.D., J. Am. Med. Assoc.
1992, vol. 268, pp. 2523–2529.
,
10. Lahiri, D.K., Farlow, M.R., Grieg, N.H., and Samꢀ
bamurti, K., Drug. Dev. Res., 2002, vol. 56, pp. 267–
281.
11. Rogers, S.L., Farlow, M.R., Doody, R.S., Mohs, R.,
and Friedhoff, L.T., Neurology, 1998, vol. 50, pp. 136–
145.
ACKNOWLEDGMENTS
12. Eugene, L., Piatnitski, C., Hassan, M.E., and John, B.,
This work was supported by Business for Cooperaꢀ
tive R&D between Industry, Academy, and Research
Institute funded Korea Small and Medium Business
Administration in 2012 (grant no. C0036335) and by
Tetrahedron. Lett., 2008, vol. 49, pp. 6709–6711.
13. Ellman, G.L., Courtney, K.D., Andres, V., and Feathꢀ
erstone, R.M., Biochem. Pharmacol., 1961, vol. 7,
pp. 88–90.
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 41
No. 2
2015