Synthesis, characterization, molecular docking, and biological activities of coumarin–1,2,3-triazole-acetamide hybrid derivatives

Article abstract of DOI:10.1002/ardp.202000109

Coumarins and their derivatives are receiving increasing attention due to numerous biochemical and pharmacological applications. In this study, a series of novel coumarin–1,2,3-triazole-acetamide hybrids was tested against some metabolic enzymes including α-glycosidase (α-Gly), α-amylase (α-Amy), acetylcholinesterase (AChE), butyrylcholinesterase (BChE), human carbonic anhydrase I (hCA I), and hCA II. The new coumarin–1,2,3-triazole-acetamide hybrids showed K_i values in the range of 483.50–1,243.04 nM against hCA I, 508.55–1,284.36 nM against hCA II, 24.85–132.85 nM against AChE, 27.17–1,104.36 nM against BChE,?590.42–1,104.36 nM against α-Gly,?and 55.38–128.63 nM against α-Amy. The novel coumarin–1,2,3-triazole-acetamide hybrids had effective inhibition profiles against all tested metabolic enzymes. Also, due to the enzyme inhibitory effects of the new hybrids, they are potential drug candidates to treat diseases such as epilepsy, glaucoma, type-2 diabetes mellitus (T2DM), Alzheimer's disease (AD), and leukemia. Additionally, these inhibition effects were compared with standard enzyme inhibitors like acetazolamide (for hCA I and II), tacrine (for AChE and BChE), and acarbose (for α-Gly and α-Amy). Also, those coumarin–1,2,3-triazole-acetamide hybrids with the best inhibition score were docked into the active site of the indicated metabolic enzymes.

Full text of DOI:10.1002/ardp.202000109

Received: 4 April 2020
Revised: 7 June 2020
Accepted: 9 June 2020
|
|
DOI: 10.1002/ardp.202000109
F U L L P A P E R
Synthesis, characterization, molecular docking, and biological
activities of coumarin–1,2,3‐triazole‐acetamide hybrid
derivatives
1
2
3
Nima Sepehri | Maryam Mohammadi‐Khanaposhtani | Nafise Asemanipoor |
4
3
3
3
Samanesadat Hosseini | Mahmood Biglar | Bagher Larijani | Mohammad Mahdavi |
5
6
7
7
Haleh Hamedifar | Parham Taslimi
| Nastaran Sadeghian | Ilhami Gulcin
1
Nano Alvand Company, Avicenna Tech Park,
Tehran University of Medical Sciences,
Abstract
Tehran, Iran
Coumarins and their derivatives are receiving increasing attention due to numerous
biochemical and pharmacological applications. In this study, a series of novel
coumarin–1,2,3‐triazole‐acetamide hybrids was tested against some metabolic enzymes
including α‐glycosidase (α‐Gly), α‐amylase (α‐Amy), acetylcholinesterase (AChE),
butyrylcholinesterase (BChE), human carbonic anhydrase I (hCA I), and hCA II.
2
Cellular and Molecular Biology Research
Center, Health Research Institute, Babol
University of Medical Sciences, Babol, Iran
3
Endocrinology and Metabolism Research
Center, Endocrinology and Metabolism Clinical
Sciences Institute, Tehran University of
Medical Sciences, Tehran, Iran
The new coumarin–1,2,3‐triazole‐acetamide hybrids showed
range of 483.50–1,243.04 nM against hCA I, 508.55–1,284.36 nM against hCA II,
4.85–132.85 nM against AChE, 27.17–1,104.36 nM against BChE, 590.42–1,104.36 nM
K
i
values in the
4
Department of Pharmaceutical Chemistry,
School of Pharmacy, Shahid Beheshti
University of Medical Sciences, Tehran, Iran
2
5
CinnaGen Medical Biotechnology Research
against α‐Gly, and 55.38–128.63 nM against α‐Amy. The novel coumarin–1,2,3‐triazole‐
acetamide hybrids had effective inhibition profiles against all tested metabolic enzymes.
Also, due to the enzyme inhibitory effects of the new hybrids, they are potential drug
candidates to treat diseases such as epilepsy, glaucoma, type‐2 diabetes mellitus (T2DM),
Alzheimer’s disease (AD), and leukemia. Additionally, these inhibition effects were com-
pared with standard enzyme inhibitors like acetazolamide (for hCA I and II), tacrine (for
AChE and BChE), and acarbose (for α‐Gly and α‐Amy). Also, those coumarin–1,2,3‐
triazole‐acetamide hybrids with the best inhibition score were docked into the active site
of the indicated metabolic enzymes.
Center, Alborz University of Medical Sciences,
Karaj, Iran
6
Department of Biotechnology, Faculty of
Science, Bartin University, Bartin, Turkey
7
Department of Chemistry, Faculty of Science,
Ataturk University, Erzurum, Turkey
Correspondence
Parham Taslimi, Department of Biotechnology,
Faculty of Science, Bartin University, 74100
Bartin, Turkey.
Email: parham_taslimi_un@yahoo.com and
ptaslimi@bartin.edu.tr
K E Y W O R D S
acetylcholinesterase, carbonic anhydrase, coumarin, enzyme inhibition, α‐glycosidase
1
| INTRODUCTION
CAs IV, IX, XII, XIV, and XV are membrane‐bound isoforms, CAs VA
and VB are mitochondrial forms, and CA VI is a secreted iso-
[
9–11]
Carbonic anhydrases (CAs; E.C.4.2.1.1) catalyze the reversible hy-
form.
Also, the human carbonic anhydrases (hCAs) IX and XII are
[
1–3]
dration of carbon dioxide (CO
2
) to bicarbonate anion and proton.
overexpressed in cancer cells, as they are transmembrane‐bound,
tumor‐associated enzymes, mainly in hypoxic tumors, with a low
expression in normal cells. An anticancer agent, for exhibition of
potent cytotoxicity without adverse effects, should selectively inhibit
tumor‐associated hCAs IX and XII over cytosolic CAs like hCA I and II
They belong to the lyase family and are ubiquitous metalloenzymes
that are present in most living organisms and are encoded by six
[
4–6]
distinct unrelated gene families: the α‐, β‐, γ‐, δ‐, and ζ‐CAs.
In
mammals, 16 α‐CA isozymes or CA‐related proteins have been de-
[
12,13]
scribed with different catalytic activities, subcellular localization, and
isoenzymes.
Especially, CA I and II isoforms have been found in
[
7,8]
tissue distribution.
The CAs I, II, III, VII, and XIII are cytosolic; the
the respiration and regulation of acid/base homeostasis. These
Arch Pharm. 2020;e2000109.
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[32,33]
2
complex processes involve the transport of CO /bicarbonate be-
overcome postprandial hyperglycemia.
For this purpose, miglitol,
tween metabolizing tissues and excretion sites (lungs and kidneys),
acarbose, and voglibose as α‐glycosidase and α‐amylase inhibitors are
used in diabetic patients for reducing postprandial hyperglycemia. α‐
Amylase is an endogenous hydrolytic enzyme and secreted by the
pancreas. It cleaves α‐(1,4)‐glycoside bonds of starch into shorter,
linear, and branched dextrin chains, which are further hydrolyzed to
glucose units by α‐glycosidase. However, a long‐term usage of these
which facilitate CO
2
elimination in capillaries and pulmonary micro-
+
vasculature, elimination of H ions in the renal tubules and collecting
ducts, and reabsorption of bicarbonate in the brush border and thick
ascending limb of Henle's loop in kidneys. So, CA isoforms, which are
involved in these processes, are important therapeutic targets with
their inhibition potential to treat a range of disorders including
antiglycemic agents exhibits some undesired side effects, including
[34]
[
14–16]
glaucoma, edema, obesity, cancer, osteoporosis, and epilepsy.
stomach distress, bloating, nausea, and diarrhea.
Hence, exploring
Alzheimer's disease (AD) is the most common form of dementia
in elderly people, which generally occurs due to the progressive
dysfunction of cholinergic neurons as a result of the β‐amyloid
accumulation in some selected portions of the hippocampus and
the cortex, which results in a decrease of learning and mem-
natural derivatives and safe inhibitors of both amylolytic enzymes has
crucial importance for postprandial hyperglycemia.
Coumarins are fused heterocyclic ring systems consisting of
benzene and 2‐pyrone rings. They belong to the neoflavonoids family
of plant secondary metabolites. They have the ability to make non-
covalent interactions, namely hydrogen bonds, electrostatic, hydro-
phobic, metal coordination, van der Waals force, etc., with different
[
17–19]
ory.
Until date, there are several molecular factors and me-
chanisms have been claimed that the role of acetylcholine (ACh) in
amyloid β‐protein (Aβ‐protein), N‐methyl‐D‐aspartate receptor, tau
neurofibrillary tangles, and oxidative stress, dyshomeostasis of
[
35]
active sites of the enzymes.
Thus, they display a wide range of
pharmacological activities like antimicrobial, antituberculosis, anti‐
[
19,20]
biometals are recorded.
Also, several hypotheses based on
inflammatory, antihyperlipidemic, antidepressant, antiviral, antic-
[
36]
these factors have been proposed to explain the mechanism of AD
development. The role of ACh deficit is well known to be associated
with the AD. Recently, acetylcholinesterase inhibitors (AChEIs)
have been determined as the most prescribed drugs to treat
ancer, and antioxidant activities.
Furthermore, coumarin scaffold
is widely used in design of new and potent CAs inhibitors, antic-
[
37–39]
holinesterase agents, and α‐glycosidase inhibitors.
In contrast,
1,2,3‐triazole ring, like coumarin, in addition to various biological
properties, is a popular scaffold for the design of new bioactive hy-
brid compounds with inhibitory activities against CAs, cholinesterase,
[
21–23]
AD.
The mammals’s brain contains two major cholinesterases
(
ChEs) forms including AChE and BChE. Both forms differ geneti-
[
40–43]
cally, structurally, and in their kinetics. In the human brain, BChE is
found in glial cells and neurons as well as in neurotic plaques and
tangles in patients with AD. Whereas AChE activity decreases
progressively in the brain of AD patients, BChE activity shows
and α‐glycosidase.
In the present study, we synthesized
a
novel series of
coumarin–1,2,3‐triazole‐acetamide hybrids 8a–m and evaluated
them against CA isoenzymes (hCA I and II), both cholinergic enzymes
(AChE and BChE), and both digestive enzymes (α‐glycosidase and α‐
amylase). Furthermore, the enzyme inhibition profiles of these new
compounds were compared with acetazolamide (AZA), tacrine, and
acarbose as standard inhibitors for target enzymes.
[
24,25]
some increase.
As the AChE accounts for approximately 80%
of ACh hydrolysis in a normal brain, it has received much attention
[
26,27]
as compared with the BChE.
Several lines of proof indicate
that as the disease progresses, the activity of AChE progressively
decreases and that of BChE significantly increases in the temporal
cortex and hippocampus. Consequently, a simultaneous inhibition
of both cholinergic enzymes (AChE and BChE) should provide
benefits in the treatment of AD. So far, AChE inhibition is one of
the most common and successful methods to treat AD. For this
reason, drug therapies have been developed focusing on AChE in-
hibitors, which increase cholinergic transmission through the in-
hibition of AChE responsible for ACh overthrow. Thus, AChE
inhibitors have become more promising agents in AD treatment. In
this perspective, coumarin derivatives were designed for the
management of central nervous system disorders, which include
2 | RESULTS AND DISCUSSION
2.1 | Chemistry
The synthetic route for the preparation of candidate coumarin–1,2,3‐
triazole‐acetamide hybrids 8a–m is depicted in Scheme 1.
7‐Hydroxycoumarin 1 reacted with propargyl bromide 2 in the pre-
2 3
sence of KI and K CO in acetone at reflux to give 7‐(prop‐2‐yn‐1‐
yloxy)‐2H‐chromen‐2‐one 3. In contrast, the reaction between amine
derivatives 4a–n and chloroacetyl chloride 5 in dimethylformamide
(DMF) at room temperature led to produce N‐phenyl (or benzyl)‐2‐
chloroacetamides 6a–n. Then, the latter compounds and sodium
[
27–29]
AChE and BChE inhibitions.
Diabetes is a major global health problem and associated with a
high‐carbohydrate diet, and starch is one of the main sources of car-
bohydrates. Controlling the blood glucose level is one of the most
effective methods to reduce diabetes and prevent diabetes‐related
azide reacted in the presence of NEt
at room temperature to give azide derivatives 7a–n. Finally, 7‐(prop‐
2‐yn‐1‐yloxy)‐2H‐chromen‐2‐one 3, CuSO ·5H O, and sodium as-
3 2
in the mixture of H O/t‐BuOH
[30,31]
complications.
α‐Glycosidase is an essential target to manage the
4
2
postprandial glucose level in diabetic patients. It is well known that the
inhibition of digestive enzymes like α‐glycosidase and α‐amylase by
delaying glucose absorption is one of the effective controlling ways to
corbate were added to the freshly prepared derivative 7a–n and the
reaction was continued for 22 hr at room temperature to give the
corresponding products 8a–m in good yields.

SEPEHRI ET AL.
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O
N3
(d)
+
Br (a)
n N
H
N
+
R
O
O
O
HO
O
O
O
O
O
N
N
O
7
a–n
1
2
3
8
a–n
HN
R
n
(
c)
O
O
8a: n = 0 R = H
8b: n = 0 R = 4-CH3
8h: n = 0 R = 3-Br
8i: n = 0 R = 4-Br
NH2+
(b)
Cl
n
Cl
n N
H
R
+ NaN3
Cl
4
a–n
R
8c: n = 0 R = 4-OCH3 8j: n = 0 R = 2-CH -3-Cl
5
3
6
a–n
8
8
8
8
d: n = 0 R = 4-F
8
8
8
k: n = 0 R = 2-CH -3-NO2
l: n = 1 R = H
m: n = 1 R = 4-F
3
e: n = 0 R = 2,4-diF
f : n = 0 R = 2-Cl
g: n = 0 R = 2,3-diCl
SCHEME 1 (a) KI, K
2 3 3 3 2
CO , acetone, reflux, 3hr; (b) dimethylformamide, room temperature, 30min; (c) NaN , NEt , H O, t‐BuOH;
(
d) CuSO ·5H O, sodium ascorbate, room temperature, 22hr
4
2
[
52]
2
.2 | Enzyme inhibition results
established clinical drugs with diverse therapeutic activities.
It
was reported that coumarins are a new class of non‐zinc‐
mediated CAIs and suggested that the coumarin hydrolysis pro-
duct is a cis‐2‐hydroxy‐cinnamic acid derivative, but not the
coumarin moiety bound within the enzyme active site. The same
research group has also reported that 7‐hydroxycoumarin and
derivatives had selective inhibition effects over CA iso-
The enzyme inhibitory effects of all the newly synthesized com-
pounds 8a–m were evaluated against hCA I, hCA II (cytosolic), as well
[
44,45]
as AChE, BChE, and α‐glycosidase using the former assays.
The
inhibitory activities were compared with AZA cytosolic hCA I and
hCA II isoenzymes, tacrine for AChE and BChE, and acarbose for
α‐glycosidase and α‐amylase enzymes (Table 1). The following
structure–activity relationship and results were deduced as follows:
[
53–55]
forms.
Also, novel 6‐ and 7‐substituted coumarins have
been reported to be selective tumor‐associated CA IX and XII
[
56,57]
inhibitors.
Recently, it was reported that 7,8‐disubstituted
1
. The target coumarin derivatives (8a–m) were screened for their
potential against the two cytosolic hCA isoforms, namely hCA I
and hCA II, using AZA as a standard and clinical drug. The com-
pounds 8a–m have shown a significant inhibition at the nano-
molar level against the cytosolic isoform hCA I, which exists in
coumarin derivatives showed inhibition effects in the low nano-
molar range of 0.078 to >100 μM against cytosolic hCA I
[
58]
isoenzyme.
Also, CA II showed a primary Na transportation mechanism in
the eyes. As a result of this mechanism, dominant CA II isoform is
responsible for regulating the intraocular pressure (IOP) that
[45]
erythrocytes at the high level.
It is well known that lower K
i
[
59,60]
values indicate a strong inhibitor affinity of inhibitors to en-
leads to a secondary chronic open‐angle glaucoma.
The high
[
46]
zymes.
centration of <1 µM except 8a and 8c. The inhibition types and K
constants of novel coumarin derivatives (8a–m) were obtained
All the target compounds exhibited inhibitory con-
IOP is harmful for the optic nerve in the eye. Thus, CA II inhibition
reduces a high IOP normally accompanying glaucoma. The in-
i
hibitors of CA II should be closely examined for glaucoma treat-
[61]
[47]
[48,49]
from Lineweaver–Burk graphs,
The novel coumarin derivatives 8a–m showed nanomolar inhibi-
tion levels against hCA I, with K values between 483.50 ± 28.74
and 1,243.04 ± 222.54 nM (Table 1). Also, compound 8g had the
best inhibition effect toward hCA I (K : 483.50 ± 28.74 nM).
The standard compound AZA exhibits value of
,320.59 ± 99.17 nM for cytosolic hCA I isoenzyme (Ki‐AZA/Ki‐8g
.73). AZA has well‐tolerated pharmacokinetic effects and is
as described previously.
ment.
Another pharmaceutical application of CA II inhibitors is
their usage in the treatment of bone resorption generally found
[
62]
i
during postmenopausal osteoporosis.
In this context, sulfa-
mates as well‐known CA II inhibitors are directly bound to the
[
63]
i
metal ions in the active site cavity of hCA II isoform.
Regarding
a
K
i
the inhibition effects against hCA II, the tested novel
1
2
:
coumarin–1,2,3‐triazole‐acetamide hybrids 8a–m demonstrated a
similar inhibition potency with
K
i
values ranging from
known to decrease headache, nausea, tiredness, dizziness, and
glaucoma; however, it displays some undesired side effects in-
cluding dizziness, blurred vision, dry mouth, drowsiness, diarrhea,
508.55 ± 48.11 to 1,284.36 ± 266.94 nM. However, AZA that is
[
64]
used to treat glaucoma and some other diseases
i
had a K value
of 22.17 ± 0.65 nM against hCA II. As seen in the Table 1, similar
to CA I inhibition, compound 8g had the greatest selectivity
against hCA II isoenzyme. It is a promising coumarin–1,2,3‐
triazole‐acetamide hybrid 8g in all novel synthesized coumarin
derivatives. In another words, it has a maximum inhibition profile
and can be used for the treatment of glaucoma after advanced
examinations (Ki‐AZA/Ki‐8g: 2.95; Table 1). It is well known that the
detailed understanding of the CA inhibition mechanism with the
[
50]
loss of appetite, vomiting, nausea, and increased urination.
All
undesirable side effects are due to its nonspecific CA inhibition.
The results clearly displayed that the novel coumarin derivatives
8a–m had an effective inhibition profile than that of AZA against
cytosolic hCA I isoform (Table 1). It is known that coumarin de-
rivatives are well‐studied CAIs and their CA inhibition effects are
[51]
well established.
Coumarins are present in a number of well‐

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coumarins might be useful for the design of new pharmacological
all novel coumarin–1,2,3‐triazole‐acetamide hybrids 8a–m had ef-
fective α‐glycosidase inhibition effects than that of acarbose
[
55]
applications.
It was reported that the hCA II isozyme was in-
hibited by some novel synthesized coumarin derivatives, with
(K
Also, highly effective
(K : 590.42 ± 59.04 nM). Similarly, novel coumarin–1,2,3‐triazole‐
acetamide hybrids 8a–m showed good IC50 values (IC50
55.38–128.63 nM) against α‐amylase in comparison to standard in-
hibitor acarbose (IC50 = 154.93). K values of novel coumarin–1,2,3‐
i
: 1,260.08 ± 123.54 nM) as a standard α‐glycosidase inhibitor.
[
55]
inhibition ranges of 32 nM–243 μM.
These results are compa-
i
Ks
were calculated for compound 8h
tible with our study. Also, 7,8‐disubstituted coumarin derivatives
i
showed inhibition effects in the low nanomolar range of 0.058 to
=
[
58]
>100 μM against cytosolic hCA II isoenzyme.
2
. The cholinesterase inhibition effects of novel coumarin–1,2,3‐
triazole‐acetamide hybrids 8a–m were evaluated according to the
spectrophotometric Ellman’s method using tacrine as a reference
i
triazole‐acetamide hybrids 8a–m could not be calculated because
starch as a substrate is a polysaccharide and its concentration can-
[68]
[
65]
[66,67]
compound,
from the electric eel (E.C. 3.1.1.7) and BChE from equine serum
E.C. 3.1.1.8). Both cholinergic enzymes inhibition results are re-
ported in Table 1. Novel coumarin–1,2,3‐triazole‐acetamide hybrids
a–m had K values ranging from 24.85 ± 3.75 to 132.85 ± 16.54 nM
for AChE (Table 1). However, novel coumarin–1,2,3‐triazole‐
acetamide hybrids 8a–m had K values ranging from 27.17 ± 4.05 to
,104.36 ± 294.52 nM for BChE, which is found mainly in the liver
and blood plasma. However, tacrine (TAC) had K values of
73.22 ± 17.35 and 228.18 ± 31.22 nM against indicated AChE and
as described previously.
AChE was obtained
not be calculated.
(
2.3 | Docking study
8
i
To identify the interaction modes of the most potent compounds in
the active site of the studied enzymes of hCA I, hCA II, α‐glycosidase,
α‐amylase, AChE, and BChE, molecular modeling was performed by
AutoDock Tools (1.5.6). Compound 8g as the most potent compound
against hCA I and hCA II was evaluated in the active site of these
isoenzymes. As we explained in our previous studies, AZA as a
standard inhibitor against CA can interact with Zn301 and His200 in
the active site of hCA I and Zn265, Asn67, and Thr199 of the hCA II
i
1
i
1
BChE, respectively. It could be seen from the table that all novel
target coumarin–1,2,3‐triazole‐acetamide hybrids 8a–m demon-
strated marked inhibitory effects against both cholinesterase with
[
17]
K
i
values in the sub‐nanomolar range; however, compound 8d
showed perfect inhibition effect against AChE (K
4.85 ± 3.75 nM; Ki‐TAC/Ki‐8d: 6.97; Table 1). In contrast, in a recent
study, it was found that novel tacrine–coumarin hybrids had in-
hibition effects with K values of 133–258.000 nM against AChE
and K values of 99–22.000 nM against BChE.
active site.
As can be seen in the Table 1, the activity of compound
a
i
:
8g was about threefold better than AZA against hCA I and hCA II.
The docking result of compound 8g demonstrated that this com-
pound also fitted in the active site of hCA isoenzymes and interacted
with important residues. As can be seen in Figure 1a, compound 8g
formed interactions with Zn301, His200, Thr199, Gln92, Ala121,
Pro202, Leu198, Ala135, and Leu131 in the active site of hCA I
isoenzyme. This compound showed important interactions with
2
i
[28]
i
Our results have
shown potent results, as represented in the present study. It was
reported that some novel disubstituted coumarins demonstrated a
[
69]
medium inhibition potency against hCA I with K
i
values in the range
Zn265 and Gln92 as key residues of hCA II active site.
Compound
[
54]
of 1.3–9.3 μM.
. Finally, for the α‐glycosidase inhibition, coumarin–1,2,3‐triazole‐
acetamide hybrids 8a–m showed K values between 590.42 ± 59.04
and 1,104.36 ± 294.52 nM (Table 1). The results demonstrated that
8g also interacted with amino acids His94, Ala65, Val143, Leu198,
Val121, Leu141, and Trp209 in the active site of hCA II.
3
i
An antidiabetic study on synthesized compounds 8a–m revealed
that the activity of compound 8h against α‐glycosidase and
FIGURE 1 Interaction modes of compound 8g in the active sites of (a) human carbonic anhydrase (hCA I) and (b) hCA II isoenzymes

6
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(
a)
8h
(b)
8d
FIGURE 2 (a) The interaction mode of compound 8h in the active site of α‐glycosidase; (b) the interaction mode of compound 8d in the
active site of α‐amylase
[
72]
compound 8d against α‐amylase was better than others (Table 1).
Compound 8h interacted with important residues Arg312, Asn241,
Glu304, Ser308, and Pro309 in the active site of α‐glycosidase
catalytic triad (CT), and peripheral anionic site (PAS).
The docking
study of compound 8d as most potent cholinesterase inhibitor among
the synthesized compounds demonstrated that this compound in-
teracted with AS residues Trp84, Phe300, and Phe301, and PAS re-
sidues Trp279 and Tyr334 in the active site of AChE (Figure 3a).
Furthermore, compound 8d formed interactions with active site re-
sidues Phe288 (acyl pocket), Ser286, and Arg289. The interaction
mode of this compound in the active site of BChE is also shown in
Figure 3b. Compound 8d formed interactions with CT residues
Ser198 and His438, oxyanion hole residues Gly116, Gly117, and
[70]
(Figure 2a).
This compound also formed a hydrophobic interaction
with Val305. As can be seen in Figure 2b, compound 8d interacted
with important residues His201, Ile235, and Tyr151 in the active site
[
71]
of α‐amylase.
This compound also interacted with α‐amylase
active site residues Lys200, Ser199, Ala307, Gly308, and Glu240.
As discussed extensively in the literature, important components
of active site of AChE and BChE include the anionic site (AS),
FIGURE 3 Interaction modes of compound 8d in the active sites of (a) acetylcholinesterase and (b) butyrylcholinesterase enzymes

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Ala199, and acyl pocket residue Val288. This compound also formed
4.1.3 | General procedure for the synthesis of azide
derivatives 7a–n
an interaction with Gly115 via a 4‐fluoro substituent.
For a click reaction, N‐phenyl (or benzyl)‐2‐chloroacetamides 6a–n
were converted to azide derivatives 7a–n in situ by a reaction be-
tween compounds 6a–n (1.1 mmol), sodium azide (0.9 mmol), and
3
| CONCLUSION
In conclusion, we have reported the synthesis of novel
coumarin–1,2,3‐triazole‐acetamide hybrids 8a–m, which are de-
signed as target inhibitors against hCA I and hCA II isoenzymes,
AChE and BChE, α‐glycosidase and α‐amylase enzymes. All of the
compounds have showed a significant sub‐nanomolar inhibition
against all metabolic enzymes, especially compound 8e for cytosolic
hCA I and II isoenzymes, 8d for both cholinergic enzymes and
α‐amylase enzyme, and 8h for α‐glycosidase enzyme. Different moi-
eties of the compounds are of crucial importance for the inhibition
for indicated enzyme receptors. Novel coumarin–1,2,3‐triazole‐
acetamide hybrids 8a–m could modulate different pathways involved
in disease progression. The results may be useful for designing and
synthesizing new metabolic enzyme inhibitors, which are potential
lead candidates for the design of novel drugs to treat some diseases
including epilepsy, glaucoma, leukemia, AD, and T2DM in the future.
NEt
3 2
(1.3 mmol) in the mixture of H O and t‐BuOH (10 ml, 1:1) at
room temperature for 1 hr.
4.1.4 | General procedure for the synthesis of
coumarin–1,2,3‐triazole‐acetamide hybrids 8a–m
Finally, 7‐(prop‐2‐yn‐1‐yloxy)‐2H‐chromen‐2‐one 3 (1 mmol), so-
4 2
dium ascorbate, and CuSO ·5H O (7 mol %) were added to azide
derivatives 7a–n, and the obtained mixture was stirred at room
temperature for 22 hr. Then, the reaction mixture was poured
into crushed ice in water and formed precipitates were filtered
off, washed with water, and purified by recrystallization (in ethyl
acetate) to give pure coumarin–1,2,3‐triazole‐acetamide hy-
brids 8a–m.
The original spectra of the investigated compounds, together
with their InChI keys and some biological activity data, are provided
as Supporting Information Data,
4
4
4
| EXPERIMENTAL
.1 | Chemistry
2‐(4‐{[2‐(Oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1H‐1,2,3‐triazol‐1‐yl)‐
N‐phenylacetamide 8a
.1.1 | Synthesis of 7‐(prop‐2‐yn‐1‐yloxy)‐2H‐
chromen‐2‐one 3
White powder, yield: 79%; infrared (IR; KBr disc), ν: 3,398, 1,702,
–
1
1
1,672, 1,540, 1,120 cm
00 MHz, dimethyl sulfoxide [DMSO]‐d
(s, 2H, CH ), 6.31 (d, J = 7.2 Hz, 1H, He), 7.03–7.11 (m, 2H, Hb, H4),
;
H nuclear magnetic resonance (NMR;
4
6
): 5.31 (s, 2H, CH ), 5.38
2
A suspension of 7‐hydroxycoumarin 3 (1 mmol) and K
2
CO
3
(2 mmol)
2
in acetone (10 ml) was stirred at room temperature for 30 min. Then,
7.20 (1H, m, Ha), 7.34 (t, J = 8.0 Hz, 2H, H3, H5), 7.58 (d, J = 8.4 Hz,
2H, H2, H6), 7.64 (d, J = 8.8 Hz, 1H, Hc), 8.00 (d, J = 7.2 Hz, 1H, Hd),
the latter mixture was added to a suspension of propargyl bromide 2
1
3
(
1.2 mmol) and KI in acetone (5 ml) dropwise, and the obtained
8.33 (s, 1H, triazole), 10.51 (s, 1H, NH); C NMR (100 MHz, DMSO‐
): 52.7, 62.0, 102.0, 113.0, 113.1, 113.4, 119.6, 124.2, 127.1, 129.4,
130.0, 136.6, 142.2, 144.7, 155.7, 160.7, 161.6, 164.6. Anal. calcd. for
: C, 63.83; H, 4.29; N, 14.89. Found: C, 63.93; H, 4.19;
mixture was stirred at reflux for 3 hr. Then, the solvent (acetone) was
evaporated under reduced pressure, the residue was dissolved in
ethyl acetate (30 ml), ethyl acetate was washed with water
d
6
20 16 4 4
C H N O
(4 × 10 ml) and brine (2 × 10 ml), and dried over Na
2
SO
4
. The solvent
N, 14.85.
(ethyl acetate) was evaporated under vacuum, and the obtained re-
sidue was recrystallized from ethyl acetate to obtain pure 7‐(prop‐2‐
yn‐1‐yloxy)‐2H‐chromen‐2‐one 3.
2‐(4‐{[2‐(Oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1H‐1,2,3‐triazol‐1‐yl)‐
N‐(p‐tolyl)acetamide 8b
White powder, yield: 75%; IR (KBr disc), ν: 3,298, 1,712, 1,679,
–
1
1
1
,547, 1,125 cm
;
H NMR (400 MHz, DMSO‐d
6
): 2.55 (s, 3H,
4
.1.2 | General procedure for the synthesis of
CH
3
), 5.31 (s, 2H, CH ), 5.36 (s, 2H, CH ), 6.31 (d, J = 9.2 Hz, 1H,
2
2
N‐phenyl (or benzyl)‐2‐chloroacetamides 6a–n
He), 7.03–7.05 (dd, J = 8.4, 2.4 Hz, 1H, Hb), 7.13 (d, J = 9.2 Hz, 2H,
H3, H5), 7.20 (1H, d, J = 2.4 Hz, Ha), 7.47 (2H, d, J = 8.0 Hz, H2,
H6), 7.65 (1H, d, J = 8.8 Hz, Hc), 8.01 (d, J = 9.6 Hz, 1H, Hd), 8.32
A solution of amine derivatives 4a–n (1 mmol) and chloroacetyl
chloride 5 (1.1 mmol) in DMF (15 ml) was stirred at room tempera-
ture for 30 min. Then, the mixture was diluted with cold water,
poured into ice, and precipitates were filtered off. The obtained re-
sidue was washed with cold water to obtain pure N‐phenyl (or
benzyl)‐2‐chloroacetamide 6a–n.
1
3
(s, 1H, triazole), 10.37 (s, 1H, NH); C NMR (100 MHz, DMSO‐
): 20.9, 52.6, 55.6, 102.0, 113.0, 113.1, 113.3, 119.6, 127.1,
129.7, 133.2, 136.3, 142.1, 144.7, 155.7, 160.7, 161.6, 164.3.
Anal. calcd. for C21 : C, 64.61; H, 4.65; N, 14.35. Found:
C, 64.56; H, 14.30; N, 14.34.
d
6
18 4 4
H N O

8
of 11
SEPEHRI ET AL.
|
N‐(4‐Methoxyphenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐
H‐1,2,3‐triazol‐1‐yl)acetamide 8c
White powder, yield: 71%; IR (KBr disc) ν: 3,127, 3,085, 2,980, 2,950,
134.6, 142.2, 144.7, 155.7, 160.7, 161.6, 165.3. Anal. calcd. for
1
20 4 4
C H15ClN O : C, 58.47; H, 3.68; N, 13.64. Found: C, 58.43; H, 3.61;
N, 13.54.
–
1 1
2
,834, 1,727, 1,626, 1,404, 1,298, 1,251, 1,135, 1,047 cm ; H NMR
400 MHz, DMSO‐d ): 3.72 (s, 3H, CH ), 5.32 (s, 2H, CH ), 5.36 (s, 2H,
), 6.31 (d, J = 9.2 Hz, 1H, He), 6.91 (d, J = 9.2 Hz, 2H, H3, H5),
(
6
3
2
N‐(2,3‐Dichlorophenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐
1H‐1,2,3‐triazol‐1‐yl)acetamide 8g
CH
2
7
2
8
.03–7.06 (m, 1H, Hb), 7.20 (d, J = 2 Hz, 1H, Ha), 7.49 (d, J = 8.8 Hz,
White powder, yield: 70%; IR (KBr disc) ν: 3,480, 3,100, 3,075, 2,938,
–
1 1
H, H2, H6), 7.56 (d, J = 8.4 Hz, 1H, Hc), 8.01 (d, J = 9.6 Hz, 1H, Hd),
2,876, 1,739, 1,642, 1,614, 1,285, 1,230, 1,124, 1,050, 1,032 cm ; H
NMR (400 MHz, DMSO‐d ): 5.31 (s, 2H, CH ), 5.51 (s, 2H, CH ), 6.30
1
3
.32 (s, 1H, triazole), 10.37 (s, 1H, NH); C NMR (100 MHz, DMSO‐
6
2
2
d
6
): 52.6, 55.6, 62.0, 102.0, 113.0, 113.1, 113.4, 114.4, 121.2, 127.1,
(d, J = 9.2 Hz, 1H, He), 7.03 (d, J = 8.0 Hz, 1H, Hb), 7.18 (s, 1H, Ha),
7.36 (t, J = 8.0 Hz, 1H, H5), 7.48 (d, J = 8.0 Hz, 1H, H4), 7.62
(d, J = 8.4 Hz, 1H, Hc), 7.73 (d, J = 8.0 Hz, 1H, H6), 8.00 (d, J = 9.6 Hz,
1
30.0, 131.9, 142.1, 144.8, 155.7, 155.9, 160.7, 161.6, 164.0. Anal.
calcd. for C21
18 4 5
H N O : C, 62.07; H, 4.46; N, 13.79. Found: C, 62.05;
1
3
H, 4.40; N, 13.75.
1H, Hd), 8.34 (s, 1H, triazole), 10.28 (s, 1H, NH); C NMR (100 MHz,
DMSO‐d ): 52.4, 62.0, 101.9, 113.1, 113.3, 124.9, 125.4, 127.2,
127.5, 128.6, 129.9, 132.4, 136.5, 142.3, 144.7, 155.7, 160.7, 161.6,
165.5. Anal. calcd. for C20 14Cl : C, 53.95; H, 3.17; N, 12.58.
Found: C, 53.85; H, 3.11; N, 12.54.
6
N‐(4‐Fluorophenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1H‐
,2,3‐triazol‐1‐yl)acetamide 8d
White powder, yield: 75%; IR (KBr disc), ν: 3,398, 1,707, 1,677, 1,535,
1
H
2 4 4
N O
–
1 1
1
,110 cm
s, 2H, CH
.16–7.21 (m, 3H, Ha, Ph), 7.60 (m, 2H, Ph), 7.65 (d, J = 8.8 Hz, 1H,
Hc), 8.00 (d, J = 9.6 Hz, 1H, Hd), 8.32 (s, 1H, triazole), 10.57 (s, 1H,
;
H NMR (400 MHz, DMSO‐d
6 2
): 5.31 (s, 2H, CH ), 5.36
(
2
), 6.31 (d, J = 9.6 Hz, 1H, He), 7.03–7.06 (m, 1H, Hb),
N‐(3‐Bromophenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1H‐
1,2,3‐triazol‐1‐yl)acetamide 8h
7
White powder, yield: 78%; IR (KBr disc) ν: 3,300, 3,112, 3,087, 2,935,
−1 1
1
3
NH); C NMR (100 MHz, DMSO‐d
6
): 52.6, 62.0, 102.0, 113.0, 113.1,
13.4, 115.9, 116.1, 121.4, 121.5, 127.1, 130.0, 135.2, 142.2, 144.8,
55.7, 157.5, 159.9, 160.7, 161.6, 164.5. Anal. calcd. for
15FN : C, 60.91; H, 3.83; N, 14.21. Found: C, 60.89; H, 3.80;
N, 14.19.
2,879, 1,729, 1,664, 1,282, 1,242, 1,123, 1,087, 1,016 cm ; H NMR
1
1
(400 MHz, DMSO‐d ): 5.31 (s, 2H, CH ), 5.39 (s, 2H, CH ), 6.31
6
2
2
(d, J = 9.6 Hz, 1H, He), 7.03–7.05 (dd, J = 8, 2.4 Hz, 1H, Hb), 7.20
(d, J = 2.0 Hz, 1H, Ha), 7.27–7.33 (m, 2H, H5, H6), 7.48 (d, J = 7.2 Hz,
1H, H4), 7.65 (d, J = 8.6 Hz, 1H, Hc), 7.92 (s, 1H, H2), 8.01
C
20
H
4 4
O
(d, J = 9.5 Hz, 1H, Hd), 8.32 (s, 1H, triazole), 10.69 (s, 1H, NH);
1
3
N‐(2,4‐Difluorophenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐
H‐1,2,3‐triazol‐1‐yl)acetamide 8e
White powder, yield: 79%; IR (KBr disc) ν: 3,410, 3,242, 3,087, 2,938,
C NMR (100 MHz, DMSO‐d
113.4, 118.4, 122.0, 122.1, 126.9, 127.1, 130.0, 131.4, 140.4, 142.2,
144.7, 155.7, 160.7, 161.6, 165.1. Anal. calcd. for C20 15BrN : C,
52.76; H, 3.32; N, 12.31. Found: C, 52.66; H, 3.30; N, 12.34.
6
): 52.6, 62.0, 102.0, 113.0, 113.1,
1
H
4 4
O
−
1 1
2
,877, 1,745, 1,654, 1,289, 1,244, 1,121, 1,099, 1,025 cm ; H NMR
400 MHz, DMSO‐d ): 5.31 (s, 2H, CH ), 5.36 (s, 2H, CH ), 6.30
d, J = 9.2 Hz, 1H, He), 7.03–7.10 (m, 2H, Ha, H2), 7.19 (m, 1H, Hb),
.33–7.39 (m, 1H, H5), 7.63 (d, J = 8.8 Hz, 1H, Hc), 7.83–7.89 (m, 1H,
H6), 8.00 (1H, d, J = 9.5 Hz, Hd), 8.33 (s, 1H, triazole), 10.37 (s, 1H,
(
(
6
2
2
N‐(4‐Bromophenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1H‐
1,2,3‐triazol‐1‐yl)acetamide 8i
7
White powder, yield: 79%; IR (KBr disc) ν: 3,410, 3,149, 3,037, 2,945,
−1 1
13
NH); C NMR (100 MHz, DMSO‐d
6
): 52.3, 60.23, 62.0, 101.9, 104.5,
2,886, 1,715, 1,648, 1,289, 1,274, 1,120, 1,087, 1,020 cm ; H NMR
1
1
1
04.7, 105.0, 111.6, 111.7, 111.8, 113.0, 113.3, 122.3, 122.4, 122.5,
(400 MHz, DMSO‐d ): 5.31 (s, 2H, CH ), 5.3 (s, 2H, CH ), 6.31
6
2
2
25.6, 125.7, 129.9, 142.2, 144.7, 153.0, 153.1, 155.4, 155.5, 155.7,
57.8, 157.9, 159.9, 160.2, 160.4, 160.7, 161.6, 165.3. Anal. calcd. for
(d, J = 9.6 Hz, 1H, He), 7.03–7.05 (m, 1H, Hb), 7.20 (d, J = 2 Hz, 1H,
Ha), 7.53–7.55 (m, 4H, H2, H3, H5, H6), 7.65 (d, J = 8.8 Hz, 1H, Hc),
8.01 (d, J = 9.6 Hz, 1H, Hd), 8.32 (s, 1H, triazole), 10.64 (s, 1H, NH);
20 14 2 4 4
C H F N O : C, 58.26; H, 3.42; N, 13.59. Found: C, 58.23; H, 3.31;
1
3
N, 13.54.
C NMR (100 MHz, DMSO‐d
13.4, 115.8, 121.6, 127.1, 130.0, 132.2, 138.2, 142.2, 144.8, 155.7,
160.7, 161.6, 164.8, 179.9. Anal. calcd. for C20 15BrN : C, 52.76;
H, 3.32; N, 12.31. Found: C, 52.67; H, 3.31; N, 12.36.
6
): 52.6, 62.0, 102.0, 113.0, 113.1,
1
N‐(2‐Chlorophenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1H‐
,2,3‐triazol‐1‐yl)acetamide 8f
White powder, yield: 77% IR (KBr disc) ν: 3,400, 3,142, 3,067, 2,930,
H
4 4
O
1
−
1 1
2,876, 1,725, 1,644, 1,282, 1,244, 1,127, 1,097, 1,026 cm ; H NMR
N‐(3‐Chloro‐2‐methylphenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]‐
methyl}‐1H‐1,2,3‐triazol‐1‐yl)acetamide 8j
(
(
(
400 MHz, DMSO‐d ): 5.31 (s, 2H, CH ), 5.50 (s, 2H, CH ), 6.31
6
2
2
d, J = 9.6 Hz, 1H, He), 7.03–7.05 (dd, J = 2.4 Hz, 1H, Hb), 7.20
d, J = 2.0 Hz, 1H, Ha), 7.23 (t, J = 7.6 Hz, 1H, H4), 7.34 (t, J = 7.2 Hz,
White powder, yield: 72%; IR (KBr disc) ν: 3,412, 3,132, 3,060, 2,920,
−
1 1
2,856, 1,745, 1,648, 1,262, 1,125 cm
;
H NMR (400 MHz, DMSO‐
), 5.43 (s, 2H, CH ), 6.31
1H, H5), 7.52 (d, J = 7.4 Hz, 1H, H3), 7.64 (d, J = 8.8 Hz, 1H, Hc), 7.75
d
6
): 2.26 (s, 3H, CH ), 5.30 (s, 2H, CH
3
2
2
(
d, J = 7.6 Hz, 1H, H6), 8.00 (d, J = 7.2 Hz, 1H, Hd), 8.34 (s, 1H, tria-
(d, J = 9.6 Hz, 1H, He), 7.03–7.05 (m, 1H, Hb), 7.20 (d, 1H, J = 2.0 Hz,
Ha), 7.23 (d, J = 8.0 Hz, 1H, H6), 7.32 (d, J = 8.0 Hz, 1H, H5), 7.37
(d, J = 9.5 Hz, 1H, H4), 7.65 (d, J = 8.8 Hz, 1H, Hc), 8.01 (d, J = 9.5 Hz,
1
3
zole), 10.12 (s, 1H, NH); C NMR (100 MHz, DMSO‐d
6
): 52.4, 62.0,
1
01.9, 113.0, 113.1, 113.3, 126.3, 126.7, 127.2, 128.0, 129.9, 130.1,

SEPEHRI ET AL.
9 of 11
|
1
3
[75]
1
H, Hd), 8.32 (s, 1H, triazole), 10.10 (s, 1H, NH); C NMR (100 MHz,
the spectrophotometric method of Verpoorte et al.,
as described
[76]
DMSO‐d
6
): 15.5, 52.3, 62.0, 102.0, 113.0, 113.1, 113.4, 124.8, 126.9,
in our previous studies in detail.
was used as a substrate for the enzymatic reaction.
zyme unit is adopted as the amount of CA which had absorbance
p‐Nitrophenyl acetate (p‐NPA)
[
77]
1
1
27.1, 127.4, 130.0, 134.3, 137.4, 142.2, 144.7, 155.7, 160.7, 161.6,
65.1. Anal. calcd. for C21 17ClN : C, 59.37; H, 4.03; N, 13.19%.
One CA en-
H
4 4
O
[
78]
Found: C, 59.27; H, 4.01; N, 13.16.
difference at 348 nm over 3 min at 25°C.
For determination of inhibition kinetics of novel coumarin–1,2,3‐
triazole‐acetamide hybrids, activity (%) and [coumarin] graphs were
drawn. From these graphs, half‐maximal inhibitory concentrations
N‐(2‐Methyl‐3‐nitrophenyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]‐
methyl}‐1H‐1,2,3‐triazol‐1‐yl)acetamide 8k
Yellow powder, yield: 70%; IR (KBr disc) ν: 3,146, 3,088, 2,942, 2,913,
(IC50s) for novel coumarin–1,2,3‐triazole‐acetamide hybrids were
–
1
1
[79]
2
,868, 1,723, 1,614, 1,395, 1,282, 1,263, 1,170, 1,122, 1,050 cm . H
NMR (400 MHz, DMSO‐d ): 2.50 (s, 3H, CH ), 5.31 (s, 2H, CH ), 5.48
), 6.31 (d, J = 7.2 Hz, 1H, He), 7.02–7.04 (m, 1H, Hb), 7.19
d, J = 1.6 Hz, 1H, Ha), 7.44 (t, J = 8.0 Hz, 1H, H5), 7.64 (d, J = 8.8 Hz,
H, Hc), 7.71 (d, J = 7.6 Hz, 1H, H6), 7.75 (d, J = 8.0 Hz, 1H, H4), 8.00
determined.
i
Also, for K values, three different concentrations of
6
3
2
the novel coumarin–1,2,3‐triazole‐acetamide hybrids were used.
Then, Lineweaver–Burk graphs were drawn according to these
(s, 2H, CH
2
(
measurements. K values of novel coumarin–1,2,3‐triazole‐acetamide
i
[40]
1
hybrids 8a–m were determined from Lineweaver–Burk graphs,
[80]
as
1
3
(d, J = 9.6 Hz, 1H, Hd), 8.34 (s, 1H, triazole), 10.28 (s, 1H, NH);
C
previously described.
NMR (100 MHz, DMSO‐d
6
): 14.2, 52.3, 62.0, 101.9, 113.0, 113.1,
For measuring the quantity of protein during the purification
[
81]
113.3, 121.6, 126.9, 127.1, 129.9, 130.2, 137.7, 142.3, 144.7, 151.3,
155.7, 160.7, 161.6, 165.4. Anal. calcd. for C21 : C, 57.93; H,
3.94; N, 16.09. Found: C, 57.95; H, 3.95; N, 16.05.
process, Bradford’s technique was used,
while bovine serum al-
[
82]
H
17 5
N O
6
bumin was used as the standard protein.
Sodium dodecyl sulfate‐
polyacrylamide gel electrophoresis was employed for visualizing the
[
83]
image of isoenzymes.
N‐Benzyl‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1H‐
,2,3‐triazol‐1‐yl)acetamide 8l
White powder, yield: 78%; IR (KBr disc), ν: 3,388, 1,732, 1,679, 1,556,
The inhibitory effect of novel coumarin–1,2,3‐triazole‐acetamide
1
hybrids on AChE and BChE activities was examined according to
[65]
[84]
spectrophotometric method of Ellman,
as described previously.
–
1 1
1,128 cm
;
H NMR (400 MHz, DMSO‐d
6
): 4.33 (d, J = 5.6 Hz, 2H,
α‐Glycosidase inhibition effect of novel coumarin–1,2,3‐triazole‐
CH
2
), 5.21 (s, 2H, CH ), 5.29 (s, 2H, CH ), 6.31 (d, J = 7.2 Hz, 1H, He),
2
2
acetamide hybrids was evaluated according to the method of Tao
[85]
[86]
7.02–7.05 (dd, J = 2.4, 8.8 Hz, 1H, Hb), 7.19 (m, 1H, Ha), 7.24–7.36
et al.
The absorbance of samples was recorded at 405 nm.
(
m, 5H, Ph), 7.65 (d, J = 8.8 Hz, 1H, Hc), 8.00 (d, J = 9.6 Hz, 1H, Hd),
Finally, the inhibitory effects of novel coumarin–1,2,3‐triazole‐
acetamide hybrids were determined on α‐amylase according to the
1
3
8
.27 (s, 1H, triazole), 8.90 (t, J = 5.5 Hz, 1H, NH); C NMR (100 MHz,
): 52.0, 62.0, 101.9, 113.0, 113.1, 113.4, 127.0, 127.4,
27.8, 129.8, 129.9, 130.0, 139.1, 142.1, 144.7, 155.7, 160.7, 161.6,
65.8. Anal. calcd. for C21 : C, 64.61; H, 4.65; N, 14.35.
[
87]
DMSO‐d
6
procedure given by Xiao et al.
by using starch as a substrate; the
[
81]
1
1
detailed information has been provided previously.
The absor-
[88]
H N
18 4
O
4
bances were spectrophotometrically measured at 580 nm.
Found: C, 64.60; H, 4.62; N, 14.30.
N‐(4‐Fluorobenzyl)‐2‐(4‐{[2‐(oxo‐2H‐chromen‐7‐yl)oxy]methyl}‐1
H‐1,2,3‐triazol‐1‐yl)acetamide 8m
4.3 | Docking study
White powder, yield: 73%; IR (KBr disc) ν: 38,764, 3,133, 3,093,
Docking studies of compound 8d as the most potent compound
against α‐amylase (pdb code: 4W93), AChE (pdb code: 1EVE), and
BChE (pdb code: 1P0I), compound 8g as the most potent compound
against hCA I (pdb code: 4WR7) and hCA II (pdb code: 5AML), and
compound 8h as the most potent compound against α‐glucosidase
–
1 1
1
,729, 1,617, 1,296, 1,241, 1,133, 1,047 cm
DMSO‐d ): 4.32 (d, J = 5.6 Hz, 2H, CH ), 5.20 (s, CH
CH ), 6.31 (d, J = 7.2 Hz, 1H, He), 7.03 (m, 1H, Hb), 7.15–7.19 (m, 3H,
Ha, Ph), 7.32 (m, 2H, Ph), 7.65 (d, J = 8.8 Hz, 1H, Hc), 8.01
d, J = 9.2 Hz, 1H, Hd), 8.27 (s, 1H, H‐triazole), 8.90 (t, J = 5.2 Hz, 1H,
;
H NMR (400 MHz,
6
2
2
, 2H), 5.29 (s, 2H,
2
[70]
(
(modeled enzyme) were performed by AutoDock Tools 1.5.6.
The
13
NH); C NMR (100 MHz, DMSO‐d
6
): 52.0, 62.0, 101.9, 113.0, 113.1,
13.4, 115.4, 115.6, 127.0, 129.8, 129.9, 130.0, 135.4, 135.5, 142.1,
44.8, 155.7, 160.5, 160.7, 161.6, 162.9, 165.9. Anal. calcd. for
17FN : C, 61.76; H, 4.20; N, 13.72. Found: C, 61.60; H, 4.22;
N, 13.61.
3D structures of the selected compounds were generated by Mar-
vinSketch 5.10.4, and then their pdbqt formats were provided by
AutoDock Tools. By using the same software, the pdbqt structures of
the target enzymes were also prepared and their AutoDock Tools
parameters were set as follows: (a) α‐amylase: box size:
1
1
C
21
H
4 4
O
60 × 60 × 60 Å; the center of box: x = −11.2985, y = 1.60, z = –24.868;
(b) AChE: box size: 40 × 40 × 40 Å; the center of box: x = 2.023,
4
.2 | Enzyme inhibition studies
y = 63.295, z = 67.062; (c) BChE: box size: 56 × 56 × 56 Å; the center
of box: x = 137.985, y = 122.725, z = 38.78; (d) hCA I: box size:
40 × 40 × 40 Å, the center of box: x = 1.8085, y = 72.66, z = 55.3515;
(e) hCA II: box size: 60 × 60 × 60 Å; the center of box: x = −3.8315,
y = 3.9065, z = 15.043; (f) α‐glycosidase: box size: 40 × 40 × 40 Å; the
In the present work, hCA I and II isoenzymes were purified by
Sepharose‐4B‐L‐tyrosine‐sulfanilamide affinity column chromato-
[
73,74]
graphy.
CA isoenzymes activity was determined according to

10 of 11
SEPEHRI ET AL.
|
center of box: x = 12.5825, y = −7.8955, z = 12.519. Each docked
system was carried out by 50 runs of the AutoDock search (by the
Lamarckian genetic algorithm). Finally, the best pose of each com-
pound was analyzed by Discovery Studio 2019 Client (Accelrys, Inc.,
San Diego, CA).
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CONFLICT OF INTERESTS
29] J. Song, Y. Wu, X. Ma, L. Feng, Z. Wang, G. Jiang, H. Tong, Carbohydr.
Polym. 2020, 230, 115586.
The authors declare that there are no conflicts of interests.
30] M. Boztas, P. Taslimi, M. A. Yavari, İ. Gülçin, E. Sahin, A. Menzek,
Bioorg. Chem. 2019, 89, 103017.
ORCID
31] U. Atmaca, R. Kaya, H. S. Kahraman, M. Çelik, İ. Gülçin, Bioorg. Chem.
Parham Taslimi
http://orcid.org/0000-0002-3171-0633
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