Design and synthesis of novel quinazolinone-pyrazole derivatives as potential α-glucosidase inhibitors: Structure-activity relationship, molecular modeling and kinetic study

Article abstract of DOI:10.1016/j.bioorg.2021.105127

In this study, a new series of quinazolinone-pyrazole hybrids were designed, synthesized and screened for their α-glucosidase inhibitory activity. The results of the in vitro screening indicated that all the molecular hybrids exhibited more inhibitory activity (IC₅₀ values ranging from 60.5 ± 0.3 μM-186.6 ± 20 μM) in comparison to standard acarbose (IC₅₀ = 750.0 ± 10.0 μM). Limited structure–activity relationship suggested that the variation in the inhibitory activities of the compounds affected by different substitutions on phenyl rings of diphenyl pyrazole moiety. The enzyme kinetic studies of the most potent compound 9i revealed that it inhibited α-glucosidase in a competitive mode with a Ki of 56 μM. Molecular docking study was performed to predict the putative binding interaction. As expected, all pharmacophoric moieties used in the initial structure design playing a pivotal role in the interaction with the binding site of the enzyme. In addition, by performing molecular dynamic investigation and MM-GBSA calculation, we investigated the difference in structural perturbation and dynamic behavior that is observed over α-glycosidase in complex with the most active compound and acarbose relative to unbound α-glycosidase enzyme.

Full text of DOI:10.1016/j.bioorg.2021.105127

Bioorganic Chemistry 114 (2021) 105127
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Design and synthesis of novel quinazolinone-pyrazole derivatives as
potential α-glucosidase inhibitors: Structure-activity relationship,
molecular modeling and kinetic study
Fateme Azimia, Homa Azizianb, Mohammad Najafic, Farshid Hassanzadeha,
Hojjat Sadeghi-aliabadia, Jahan B. Ghasemid, Mohammad Ali Faramarzie,
,
,*
Somayeh Mojtabavie, Bagher Larijanif, Lotfollah Saghaeia *, Mohammad Mahdavif
a Department of Medicinal Chemistry, Faculty of Pharmacy and Pharmaceutical Science, Isfahan University of Medical Science, Hezar Jerib, 817416-73461 Isfahan, Iran
b Department of Medicinal Chemistry, School of Pharmacy-International Campus, Iran University of Medical Science, Tehran, Iran
c Department of Chemistry, Isfahan University of Technology, Isfahan 84156-83111, Iran
d School of Chemistry, University College of Science, University of Tehran, P.O. Box 14155-6455, Tehran, Iran
e Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Tehran University of Medical Sciences, P.O. Box 14155-6451, Tehran 1417614411, Iran
f Endocrinology and Metabolism Research Center, Endocrinology and Metabolism Research Institute, Tehran University of Medical Sciences, Tehran, Iran
A R T I C L E I N F O
A B S T R A C T
Keywords:
In this study, a new series of quinazolinone-pyrazole hybrids were designed, synthesized and screened for their
Quinazolinone
Pyrazole
α
-glucosidase inhibitory activity. The results of the in vitro screening indicated that all the molecular hybrids
exhibited more inhibitory activity (IC50 values ranging from 60.5 ± 0.3 µM-186.6 ± 20 μM) in comparison to
α
-Glucosidase inhibitors
standard acarbose (IC50 = 750.0 ± 10.0 µM). Limited structure–activity relationship suggested that the variation
in the inhibitory activities of the compounds affected by different substitutions on phenyl rings of diphenyl
pyrazole moiety. The enzyme kinetic studies of the most potent compound 9i revealed that it inhibited
Enzyme inhibition
Molecular dynamic simulation
α
-glucosidase in a competitive mode with a Ki of 56 μM. Molecular docking study was performed to predict the
putative binding interaction. As expected, all pharmacophoric moieties used in the initial structure design
playing a pivotal role in the interaction with the binding site of the enzyme. In addition, by performing molecular
dynamic investigation and MM-GBSA calculation, we investigated the difference in structural perturbation and
dynamic behavior that is observed over
α-glycosidase in complex with the most active compound and acarbose
relative to unbound -glycosidase enzyme.
α
1. Introduction
metabolism, it is considered as a therapeutic target for other
carbohydrate-mediated diseases, including cancer [5], hepatitis [6] and
viral infection [7,8]. Currently, carbohydrate mimics acarbose (Gluco-
bay), miglitol (Glyset), and voglibose (Volix, Basen) are clinically
α
-Glucosidase is a membrane-bound enzyme at the epithelium of the
small intestine which possesses key functions in carbohydrate digestion
[1]. It catalyzes the hydrolysis of the -1–4-glycosidic bond from the
α
approved drugs that act as inhibitors of both α-amylase and α-glucosi-
non-reducing end of carbohydrate substrates and releases absorbable
glucose, which is mainly responsible for increasing blood glucose levels
[2]. The control of the blood sugar level is a critical strategy in the
management of diabetes mellitus, especially type II diabetes and
reducing chronic complications associated with the disease. Thus, in-
dase enzymes [9,10]. However, continuous administration of these
drugs is associated with undesirable gastrointestinal adverse reactions
such as diarrhea, bloating, abdominal pain, etc. [11,12]. Based on the
biological importance of α-glucosidase and the inefficiencies of existing
drugs, the design and development of new α-glucosidase inhibitors are
hibition of the
peutic approach for the treatment of type-2 diabetes mellitus [3,4].
Furthermore, since -glucosidase plays a pivotal role in carbohydrate
α
-glucosidase enzyme is considered as a useful thera-
still attractive and challenging [13–18].
Pyrazole is an attractive pharmacophore in medicinal and pharma-
ceutical chemistry. Pyrazole and its derivatives exhibit a broad spectrum
α
* Corresponding authors.
E-mail addresses: saghaie@pharm.mui.ac.ir (L. Saghaei), momahdavi@tums.ac.ir (M. Mahdavi).
https://doi.org/10.1016/j.bioorg.2021.105127
Received 29 April 2021; Received in revised form 21 June 2021; Accepted 23 June 2021
Available online 29 June 2021
0045-2068/© 2021 Published by Elsevier Inc.

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
of pharmacological activities including anti-Alzheimer [19,20], anti-
tumor [21,22], antihypertensive [23], anti-inflammatory [24,25],
analgesic [26], antimicrobial [27,28], neuroprotective [29], antiviral
[30], and antibacterial [31]. In particular, some of the pyrazole de-
rivatives like Celecoxib, Viagra and Fipronil have been approved for
clinical use [32]. Moreover, pyrazole scaffolds can be traced in some
compounds that have been reported as potent antidiabetic and hypo-
glycemic agents (Fig. 1). Kenneth et al. prepared a series of pyrazole and
pyrazolone derivatives as potent antihyperglycemic agents [33] and
Shuangjie et al. reported the pyrazole-based derivatives as a potent
glucagon receptor antagonist [34,35]. Apart from this, recent studies
molecular hybridization for the design of new bioactive compounds,
herein novel quinazolinone-pyrazole hybrids were designed and syn-
thesized as potent α-glucosidase inhibitors. In a bid to stitch effective
pharmacophores together to generate new molecules, para-chlor-
ophenyl appendage on 2-position of quinazolinone moiety was main-
tained. Next, the pyrazole moiety insert at position 3 of quinazolinone.
Amide bond was included in the design of the new hybrid as part of the
initial design because of its unique features, including the polarity,
amide proton exchange rate, and protein binding (Fig. 2) [61–65].
2. Result and discussion
have shown that several entities of pyrazole act as potent
α-glucosidase
inhibitors. Structures of some potent pyrazole core containing
α-gluco-
2.1. Chemistry
sidase inhibitors are shown in Fig. 2(A–C) [36–38]. It is pertinent to
mention that Teneligliptin, an oral DPP-4 inhibitor containing pyrazole,
was approved for the treatment of type-II diabetes [39]. Also, report
based on metabolic stability and pharmacological efficiency of pyrazole
derivatives as antidiabetic agents added to our interest toward the
synthesis of novel compounds containing pyrazole moiety and evalua-
tion of their bioactivities [40,41].
The synthetic strategy for the intermediates and target compounds
9a-p has been depicted in Scheme 1. Quinazolinone 2 was obtained from
the condensation of anthranilamide 1 and 4-choloro benzaldehyde in
the presence of NaHSO3 in DMAc. It was then alkylated with ethyl
bromoacetate in acetone and under basic conditions to afforded ester 3,
which was further mixed with hydrazine hydrate in methanol to form
acetohydrazides 4.
On the other hand, quinazolinone and its derivatives have appealed
to the medicinal chemists due to its presence in a wide range of bioactive
compounds. Versatile biological properties including anti-virus [42],
anti-inflammatory [43,44], anti-bacterial [45], anti-tubercular [46],
anti-malaria[47], anti-cancer [48,49], anti-fungal [50], anti-allergic
[51,52], anti-HIV [53] and anti-convulsant [54,55] have been success-
fully documented in the literature. Rudolph et al. introduced quinazo-
linone derivatives as orally available ghrelin receptor antagonists for the
treatment of diabetes and obesity [56]. Also, recent studies have
The procedure for the synthesis of 1,3-disubstituted pyrazole-4-
carbaldehydes 8a-p was carried out via two-step methods. Initially,
condensation of different substituted acetophenones 5 with phenyl-
hydrazine or 4-methyl phenylhydrazine in the presence of sulfuric acid
in absolute ethanol afforded hydrazone intermediates 7a-p. Next, the
synthesized hydrazone intermediates 8a-p undergoes Vilsmeier–Hack
reaction with DMF/POCl3 to form pyrazole-4-carbaldehydes derivative
8a-p. Finally, compounds 4 and variously substituted aldehyde 8a-p
were refluxed overnight in ethanol in the presence of a catalytic
amount of anhydrous acetic acid to get the target quinazolinone-
pyrazole derivatives 9a-p.
confirmed the
rivatives [57,58]. Our group has synthesized several quinazolinone-
1,2,3-triazole hybrids as new -glucosidase inhibitor. The results
revealed that all of them presented a superior inhibitory profile than
standard acarbose (IC50 = 181.0–474.5 M comparing with acarbose
(IC50 = 750.0 M) (Fig. 2D) [59]. Peng et al. synthesized several 2-aryl
quinazolinone derivatives that were subsequently assayed for -gluco-
sidase inhibitory activity. Among them, 2-(4-chlorophenyl)-quinazolin-
α-glucosidase inhibitory activity of quinazolinone de-
α
μ
2.2. In vitro
α-glucosidase inhibitory activity
μ
α
All the synthesized new hybrids 9a-p were screened for their in vitro
α
-glucosidase inhibition against yeast α-glucosidase in comparison to
4(3H)-one was identified as potent inhibitors of α-glucosidase because of
acarbose as the standard drug (Table 1). To obtain the best insight into
the effects of the different substitutions on phenyl rings attached to
pyrazole core, a variety of compounds 9 were synthesized. As can be
its strong interaction with the enzyme (Fig. 2E) [60].
In light of the above, and also inspired by the importance of
Fig. 1. Pyrazole derivatives as antidiabetic and hypoglycemic agents.
2

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
Fig. 2. Design strategy of novel quinazolinone-pyrazole derivatives as novel
α
-glucosidase inhibitors based on molecular hybridization of pharmacophoric units of
potent reported α-glucosidase inhibitors A–D.
seen in Table 1, synthesized compounds can be considered in two cat-
respectively, showed the lowest inhibitory activity in this series of
compounds. Similarly, it seems that the presence of electron-donating
–
egories of 9a-i (series A, Table 1, R2 H) and 9j-m (series B, Table 1,
–
–
R2 CH ) and then a limited structure–activity relationship has been
groups were again unfavoured for α-glucosidase inhibitory activity. In
–
3
also established based on variations in substitution pattern at the R1
this series, NO2 substituted derivative 9n showed the second highest
activity, with an IC50 value of 105.7 ± 0.8 µM. It is also interesting to
note that, among the synthesized compounds, derivatives with unsub-
stituted at R1 position have shown less inhibitory activity, as in com-
pounds 9a (IC50 = 186.6 ± 2.0 µM) and 9j (IC50 = 148.7 ± 1.6 µM).
From these results, it could be concluded that substitution at R1
position as well as their electron property plays an important role in in
position. All the new synthetic derivatives displayed excellent -gluco-
α
sidase inhibitory activity having IC50 values in the range 60.5 ±
0.3–186.6 ± 2.0 µM and 91.7 ± 0.7–167.2 ± 1.9 µM, for 8a-i and 8j-m,
respectively, when compared to standard acarbose (750 ± 10 µM). It is
worth mentioning that all compounds 9 demonstrated inhibitory ac-
tivities around 12–4 folds more than standard. The best activity was
obtained by compound 9i having bromo group at the R1 position and no
substitution at R2 position.
vitro
the methyl group at the R2 position was affected by the nature of sub-
stitutions at the R1 position. In this regard, -glucosidase inhibitory
α-glucosidase inhibitory activity. On the other hand, the efficacy of
In series A, replacing bromine with other halogens (F and Cl) led to a
decrease in the inhibitory activity as observed in the compounds 9g
(IC50 = 79.5 ± 0.5 µM) and 9 h (IC50 = 107.1 ± 0.8 µM), respectively. It
α
activity of compounds 9k, 9m, and 9n substituted with methyl, hydroxyl
and nitro at the R1 position, respectively, indicated that the introduction
of methyl substituent at the R2 position improved inhibitory activity
(inhibitory activity: 9k > 9b, 9m > 9d, 9n > 9e). In a different manner,
the presence of methyl group at R2 position caused bromo derivative 9p
showed two-fold lower activity than its corresponding analog 8l in series
A. Similarly, in chloro substituted derivative 9h, the absence of the
methyl group led to better activity (inhibitory activity: 9h > 9o).
Interestingly, the presence of the methyl group in methoxy derivative
had a negligible effect on the inhibitory efficacity (9c and 9l).
should be noted that trifluoromethyl substituted compound 9f (IC50
=
143.6 ± 1.6
μ
M) showed lower activity than other halogenated com-
-glucosidase inhibitory activity of
pounds. Therefore, it seems that the
α
halogenated analogous 9f-i was not only affected by the size of halogen
groups but also by their electronic property of them. In this series of
compounds, it was observed that the introduction of the electron-
withdrawing NO2 group (compound 9e) led to good activity with IC50
= 113.2 ± 1.0 µM. Finally, compounds 9b-d depicted versatile inhibitory
activity affected by the electron-donating capability of the substitution.
The order of activity for compounds 9b-d, was found to be 9b > 9c > 9d
2.3. Kinetic study
(IC50s = 138.3, 151.7, and 176.2 μM, respectively), suggesting that an
electron-donating group was less favorable to the activity.
To evaluate the mechanism of inhibition of the new synthesized
hybrid, the enzyme kinetic studies of the most active compound 9i were
performed. The inhibition mode was determined by the Line-
weaver–Burk plots and the Ki value was calculated from the second plots
of the kinetic parameter (Km) versus different concentrations of inhib-
itor 9i (Fig. 3A–B). As can be seen in Fig. 3A, the Lineweaver–Burk plots
revealed that by increasing the concentration of compound 9i, the Vmax
was not affected, while the Km increased [66–69]. It indicated that
compound 9i competes with the substrate for binding at the active site of
Further investigation comes back to the series B of compounds (9j-m)
which possessed a methyl group at the R2 position. In the case of halo-
substituted derivatives 9o (IC50 = 124.6 ± 1.1 μM) and 9p (IC50 =
133.6 ± 1.4 μM), comparing the corresponding inhibitory activity with
their counterparts in the first category revealed that the introduction of
methyl group did not induce higher activity. Compound 9k, the most
active member of the series B and the third most active compound tested
(IC50 = 91.7 ± 0.7 µM), having a methyl group at both R1 and R2 po-
sition. Replacement of methyl with hydroxyl or methoxy group
dramatically decreased the inhibitory activity. So that compounds 9l
the α-glucosidase enzyme. Moreover, the inhibitory constant (Ki) of this
compound was calculated as 56 µМ.
and 9m, with IC50 values of 155.2 ± 1.8 and 167.2 ± 1.9
μM,
Similarly, many recently developed organic-nitrogen compounds
3

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
Scheme 1. Synthesis of target compounds 10a-p. Reagents and conditions: (a) NaHSO3, DMAC, 180 ◦C, 12 h; (b) Ethyl bromoacetate, K2CO3, acetone, rt, overnight;
(c) N2H4⋅H2O, EtOH, reflux, 4 h; (d) H2SO4, EtOH, reflux, 8–12 h; (e) DMF, POCl3, 60 ◦C, 5–8 h; (f) AcOH, EtOH, reflux, overnight.
competitively inhibited the
α
-glucosidase [70,71]. Interestingly, to the
the GOLD 5.3 tool. Since the x-ray crystallographic structure S. cerevisiae
best of our knowledge, none of the reported pyrazole-based inhibitor
competitively inhibited the enzyme activity.
α-glucosidase is still not available, a homology model of this enzyme was
built and validated according to our reported method [73]. Compounds
9i, 9g, 9k, 9m, and 9a were selected for docking studies. The superposed
structure of these compounds and details of their interactions in the
active site of a-glucosidase is shown in Fig. 4.
2.4.
α-Amylase inhibition assay
From the docking calculation study, it was observed that compounds
Inhibition of
α
-amylase is also a relevant strategy to control post-
well nested into the active pocket of α-glucosidase and several favorable
prandial hyperglycemia through delayed carbohydrate digestion [72].
In this regard, the most potent compounds (9i, 9g, and 9k) were also
interactions support the high potency against the enzyme. The predicted
binding mode of studied compounds indicated that the following in-
teractions were mutual for them. (1) pyrazole core made T-shaped
evaluated for their inhibitory potential against
α
-amylase. Results
-amylase (at 300
α-amylase inhibitor
revealed that these compound had no activity against
M) when compared with acarbose as a standard
α
π
-stacking interaction with Phe177, whereas the phenyl groups attached
μ
to the pyrazole moiety established hydrophobic interaction with resi-
dues Arg439, Tyr71, Ala278 and Phe157. (2) -alkyl and T-shaped
(IC50 = 108 ± 0.71 μM).
π
π-π
Non-halogenated derivative in recent developed bis-azo-containing
Schiff base exhibited the highest inhibitory potential against both en-
interactions formed between quinazolinone moiety and Arg312 and
Phe157, respectively. (3) Furthermore, 4-chlorophenyl group directing
toward the hydrophobic pocket comprising Phe300 and Val303. This
pharmacophore showed two interactions with Phe300 and Val 303
through chloro and one ᴨ-alkyl interaction with Val303 through phenyl
ring, which could further fitted ligands tightly into the active site of the
enzyme. (4) The amide linker forms two hydrogen bonds with active site
residues, which played a significant role in stabilizing the enzyme-
zymes (
α-glucoside and α-amylase) [70].
2.5. Docking
To acquire insight into the binding interactions of the newly syn-
thesized quinazolinone-pyrazole derivatives in the active site of
α
-glucosidase, in silico docking studies were carried out by employing
4

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
positioned in a hydrophobic pocket comprising the side chains of Asp68,
His348, Tyr71 and Arg439 that allows the formation of highly stable
complexes. As can be seen in the Fig. 5B and C, the comparison of
interaction modes of compound 9g and 9i revealed that diphenyl pyr-
azole moiety in compound 9g, unlike compound 9i established fewer
hydrophobic interactions with the active site residues. On the other
hand, the oxygen carbonyl of amide moiety of the compound 9g formed
additional hydrogen bond interaction with Arg439, which could be a
key factor in stabilizing this compound in the active site of the enzyme.
Compound 9k, the most active compound in series B, formed two
additional hydrogen bonds compared to compound 9i. Arg439 estab-
lished a hydrogen bond with the oxygen atom of amide carbonyl and
Tyr313 formed a hydrogen bond with carbonyl group of quinazolinone
moiety. Also, this compound is involved in hydrophobic interactions
with several residues such as His348, His245, His279, Tyr71 and Ala278
through methyl-substituents at R1 and R2 (Fig. 4D). For further inves-
tigation, the interaction mode of least active compounds in the series A
and B (9a and 9m) were also analyzed in detail. Similar to compound 9g,
an additional hydrogen bond is established between the oxygen atom of
the amide carbonyl of compound 9a and the side chains of Arg439
(Fig. 4E). Compound 9m showed additional hydrophobic interactions
with His245, Leu218, Ala278 and Phe157 through methyl-substituted
phenyl group while hydroxyl substitution was making a hydrogen
bond with the oxygen atom of Asp68 (Fig. 4F). Other interactions are the
same in both compound 9a and 9m.
Table 1
In vitro α-glucosidase inhibitory activity of compounds 9a-p.
Compound
R1
R2
IC50(µM)a
9a
H
H
186.6 ± 2.0
138.3 ± 1.5
151.7 ± 1.8
176.2 ± 1.9
113.2 ± 1.0
143.6 ± 1.6
79.5 ± 0.5
9b
CH3
OCH3
OH
NO2
CF3
F
H
9c
H
9d
H
9e
H
9f
H
9g
H
9 h
9i
Cl
H
107.1 ± 0.8
60.5 ± 0.3
Br
H
9j
H
CH3
CH3
CH3
CH3
CH3
CH3
CH3
–
148.7 ± 1.6
91.7 ± 0.7
9k
CH3
OCH3
OH
NO2
Cl
9l
155.2 ± 1.8
167.2 ± 1.9
105.7 ± 0.8
124.6 ± 1.1
133.6 ± 1.4
750.0 ± 10.0
9m
9n
Further studies on gold fitness docking scores of compounds 9i
(72.09), 9g (74.46), 9k (969.67), 9a (77.27) and 9m (64.48) revealed
that they bound more easily to the active site than the standard drug
acarbose (35.71). However, there was no good correlation between the
docking scores and biological activities of compounds, indicating that
some other factors rather than their binding mode to the active site were
important.
9o
9p
Br
Acarbose
–
a
Values are the mean ± SD. All experiments were performed at least three
times.
inhibitor complex. Asp408 made H-acceptor interaction with NH unit of
amide moiety while Tyr313 formed H-acceptor interaction with the
carbonyl of the same moiety. Similar types of interactions were also
observed in newly developed N-alkyl–deoxynojirimycin derivatives
[71].
2.6. Molecular dynamic (MD) simulation
MD simulation was performed in order to understand the effect of the
best biologic active compound over the enzyme active site [74]. For this
purpose, the structural perturbations incurred by the most potent
compound (9i) has been investigated through the study the RMSD,
RMSF and their effect on the active site environment in comparison to
Therefore, as expected, all pharmacophores used in the design of new
hybrid have participated in binding with the catalytic residue that play a
significant role in enhancing inhibition activity against the
dase enzyme.
α-glucosi-
acarbose as α-glycosidase standard inhibitor and the apo-enzyme. The
The most potent compound 9i made further interactions in addition
to those described above and hydrophobic interactions have a greater
contribution for binding of it (Fig. 4B). Bromo substituted phenyl is
appropriate pose for MD simulation procedure of the compound 9i and
acarbose were achieved by induced fit docking method.
The stability of protein–ligand complex trajectories assessed by
Fig. 3. Kinetic study of
α-glucosidase inhibition by compound 9i. (A) The Lineweaver–Burk plot in the absence and presence of different concentrations of compound
9i ( M); (B) the secondary plot between 1/Vmax and various concentrations of compound 9i.
μ
5

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
Fig. 4. (a) 9i, 9g, 9k, 9m and 9a superimposed in the active site pocket of modeled
α-glucosidase. The predicted binding modes of (B) 9i, (C) compound 9g, (D)
compound 9k (E) compound 9m and (F) compound 9a, in the active site pocket. Residues that may be involved in the interactions of compound binding are drawn
with stick model and shown in different colors. (The colored symbols are as following, dark gray: carbon, light gray: hydrogen, red: oxygen, blue: nitrogen, green:
chlorine. The possible hydrogen-bond interactions are indicated with dashed green lines). (For interpretation of the references to color in this figure legend, the
reader is referred to the web version of this article.)
analyzing the root mean square deviation (RMSD) of the enzymes’s
can be evaluated through investigating the RMSF value in which eval-
uate the fluctuation of the protein backbone structure from its average
position throughout the whole simulation time [75]. The residue RMSF
backbone over the 20 ns MD simulation. The RMSD value of the un-
bounded
α-glycosidase enzyme depicts higher RMSD value than the
other two bounded-state enzyme complexes. The RMSD value signifi-
cantly increased during the first 5 ns up to 2.7 Å and steadily fluctuated
to the next 10 ns and become more stable for the last 5 ns of simulation
time with the value of 2.6 Å. Moreover, based on the RMSD value of
values of the α-glycosidase unbounded enzyme and the enzyme in the
bounded state showed that, the unbounded enzyme (blue color line) had
higher RMSF fluctuations compared to the -glycosidase bound-states
α
(green and red colored line) (Fig. 6-a). According to the result, the
mentioned lower RMSF value occurs upon ligand binding to the enzyme,
in which residues movement decrease as a result of non-bonding inter-
action between the ligand and the enzyme active site residues. More-
over, Fig. 6a depicted that there are four structural segments which
revealed different RMSF pattern upon ligand binding including; B
domain loop and the active site lid (both in pink dash box), A domain
and B domain sides of the active site entrance (orange and blue dash
boxes, respectively).
α
-glycosidase complexed with acarbose and compound 9i, the bounded
state enzymes were stable during the simulation time with the lower
RMSD value of 1.8 Å and 2 Å, respectively (Fig. 5). The mentioned
outcome may cause as a result of more structural rigidity due to the
active site bounding state and proposed the importance of active site
domain on the structural flexibility of α-glycosidase enzyme.
The RMSD value for the 5 ns at the terminal section of MD simulation
was used to investigate the structural specificity of
complexes.
α-glycosidase-ligand
Based on Fig. 6a and b, the RMSF value of the B domain loop residues
Moreover, the detailed residue-based flexibility of protein structure
would have decreased in α-glycosidase/acarbose and compound 9i
bound-state rather than unbounded α-glycosidase enzyme. Furthermore,
both acarbose and compound 9i bonding state show the same RMSF
value and the ligand-binding pattern except the region over the active
site lid. The flexibility of the active site lid was not only the lowest in
unbounded enzyme, but the mentioned segment flexibility revealed
higher in α-glycosidase complexed with acarbose rather than compound
9i. Furthermore, Fig. 6b depicts the interaction of acarbose and com-
pound 9i with several residues located both on A and B domain sides of
the active site mouth. Also, acarbose showed more interaction with the
A side domain of the active site entrance (orange dash line in α-glyco-
sidase/acarbose), while compound 9i provided a higher number of in-
teractions with the residues of the B domain side (blue dash line in
α-glycosidase/compound 9i). So, it can be proposed that the lower RMSF
value of the active site lid has occurred as a result of higher ligand in-
teractions with the B domain side of the active site mouth. In addition,
Fig. 6c and d depict the close-up representation of the active site mouth
in association with the corresponding residues of A and B domains at
both sides of the active site entrance.
Fig. 5. RMSD of the α-glycosidase backbone in complexed with acarbose (in
red), compound 9i (in green) and the unbound enzyme (in blue) for over 20 ns
MD simulation time. (For interpretation of the references to color in this figure
legend, the reader is referred to the web version of this article.)
6

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
Fig. 6. RMSF plot of the
α
-glycosidase backbone in complexed with compound 9i (in yellow) and acarbose (in green) and the unbounded enzyme (in red) for over 20
ns MD simulation time (a). Ligand binding location of α-glycosidase in bound and unbound-state for over 20 ns MD simulation time (α-helical and ß-strand regions are
highlighted in red and blue backgrounds, respectively) (b). 3D representation of
and orange, respectively. The the flexible regions correspond to the B domain loop and the active site lid are colored in pink (c). Close-up representation of
-glycosidase active site (d). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
α-glycosidase structure. Enzyme domain of A, B and C are colored in yellow, blue,
α
Overall, based on the result of the RMSF plot, acarbose and com-
pound 9i have almost the same interaction pattern through the whole
b represent four important structural moieties in stabilizing compound
9i into the active site of α-glycosidase. The first one is the phenyl pyr-
α
-glycosidase active site structure. The only dissimilarity comes from the
azole and the quinazoline rings, which oriented toward the front side of
the active site and interacted with Phe177 and Phe157 at the B side of
active site lid flexibility in which we can propose that compound 9i
along with more interaction over the B domain side of the active site
entrance, has a dominant effect in the rigidity of active site lid rather
than acarbose which has more interaction with the A domain side of the
the active site mouth through Van Der Waals and T-shape π-π hydro-
phobic interactions for about 51% and 60% of the simulation time,
respectively. The next part is related to the bromo phenyl and chloro
phenyl moieties which pointed toward the back part of the active site
and mainly interacted with Arg212 and Arg312 residues through
α
-glycosidase active site entrance.
Fig. 7a and b represent the detailed orientation and interactions that
occurred more than 30% of the simulation time during the equilibrated
π-cation interactions for almost the whole amount of simulation time
phase over the
α
-glycosidase complexed with compound 9i. The inter-
(95% and 91% of MD time, respectively). Along with these interactions,
the acetohydrazide group interacted with Asp408 located at the back
part of the active site (previously known as back wall helix) through H-
action inspection depicts compound 9i stabilized into the enzyme do-
mains by interacting with residues Phe300, Glu276, Arg212, Arg312,
Pro309, and Leu218 from the A domain side and Phe157 and Phe177
from the B domain side of the active site mouth. In the case of acarbose,
it disposed vertically and formed non-binding interactions with residues
Phe311, Asn241, Arg439, Asp68, His245, Asp349, Asp214, which
belong to the domain A of the enzyme (Fig. 7c). Furthermore, Fig. 7a and
–
bond interaction with the C O group for about 61% of the simulation
–
time. Besides these stabilizing interactions, the pyrazole ring provided
steady interaction with Glu276 which is one of the active site catalytic
residue for significantly 85% of simulation time.
Based on the MD study, it can reveal that compound 9i not only like
7

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
Fig. 7. The detailed orientation and ligand atom interactions that occurred more than 30.0% of the simulation time during the equilibrated phase over α-glycosidase
complexed with compound 9i (a, b) and acarbose (c, d). Domain A, domain B and the flap region covered the mouth of the active site colored in yellow, blue and
pink, respectively. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
as acarbose stabilized through hydrophobic and H-bond interaction with
A domain side of the active side mouth (Fig. 7d) but also provided
persistent and strong interactions with the B domain side of the active
side mouth, the back side of the active site and the catalytic residue
during the MD simulation time which may propose the contribution to
compound. The analysis of the structure–activity relationship indicated
that substitution at R1 position as well as their electron property plays an
important role in in vitro α-glucosidase inhibitory activity. On the other
hand, the efficacy of the methyl group at the R2 position was affected by
the nature of substitutions at the R1 position. Analysis of the kinetics of
the higher
α-glycosidase inhibition activity of the mentioned compound.
enzyme inhibition indicated that compound 9i inhibited α-glucosidase
Finally, the binding free energies calculated by performing the MM-
GBSA method, is which known as one of the rigorous and efficient
methods to estimate relative binding affinities have been performed to
predict the strength of compound’s affinity over the enzyme active site
[76,77]. In this way, a number of 102 snapshots were extracted at time
interval of 50 ps from the last 5 ns of equilibrated MD trajectories which
can provide scope for predicting the biological activity of compound 9i.
The calculated binding free energies (ΔGbind) and the individual energy
in a competitive inhibition manner (Ki = 56 µM). Further, the in silico
studies were performed to get insights into the molecular interaction of
compounds with the active site of the enzyme. Docking study of com-
pounds 9i, 9g, 9k, 9m and 9a revealed that they behave almost similar to
exhibit the inhibitory potential and well fitted in the active site of
α
-glucosidase through both hydrophobic and hydrogen interactions. As
expected, all pharmacophoric moiety used in the initial structure design
(biphenyl pyrazole, 2-(4-chlorophenyl)-quinazolin-4(3H)-one and
amide moiety) playing a pivotal role in the interaction with the binding
components reveal that the binding free energies of
α-glycosidase
complexed with compound 9i are obviously higher than acarbose
(ꢀ 93.14 ± 7.80 vs. ꢀ 62.49 ± 2.90 kcal.molꢀ 1, respectively) during the
equilibrated phase of MD simulation time which may attribute to the
more stabilizing effect of compound 9i over the A and B domain of the
enzyme.
site of the α-glucosidase enzyme. Hence, these compounds could be
promising hits for further research towards the treatment of hypergly-
cemia and associated disorders. MD simulations showed compound 9i
oriented inside the active site and stabilized through four important
structural moieties; the phenyl pyrazole and the quinazoline rings which
oriented toward the front side of the active site, the bromo phenyl and
the chloro phenyl moiety which pointed toward the back part of the
active site, the acetohydrazide group interacted at the back part of the
active site and the pyrazole ring provided steady interaction with
Glu276 which is one of the active site catalytic residue. These groups
constructed a new nucleus which provided a significant role for inhi-
bition activity of the mentioned compound.
3. Conclusion
In conclusion, a novel series of quinazolinone-pyrazole derivatives
were designed, synthesized and evaluated for α-glucosidase inhibitory
activities. The results of the in vitro screening indicated that newly
synthesized hybrids demonstrated inhibitory activities around 12–4
folds more than acarbose as standard drug. Among them, compound 9i
with the IC50 values of 60.5 ± 0.3 µM was found to be the most potent
8

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
4. Experimental
pure product [79].
All the chemicals and solvents were purchased from Merck (Ger-
many) and Sigma Aldrich (USA). Melting points (mp) of all derivatives
were measured on a Kofler hot stage apparatus and uncorrected. Re-
actions were monitored by thin-layer chromatography (TLC) using
precoated silica-gel plates (Merck, TLC silica gel 60 F254), and the plots
were visualized through UV light at wavelengths of 254 and 365 nm. 1H
and 13C NMR spectra was recorded on Avance Bruker AV-500 MHz
spectrometer in DMSO‑d6 and tetramethylsilane was used as internal
standard. Infrared spectra (IR) were recorded on Shimadzu 8400 S FT-IR
using KBr disc. Elemental analysis of title compounds 9a-p was per-
formed on an Elemental Analyzer system GmbH VarioEL CHN mode.
4.5. General procedure for the synthesis of quinazolinone-pyrazole
hybrids 9a-p
Equimolar quantities (1 mmol) of appropriate pyrazole-4-
carbaldehyde derivative 8a-p and 2-(2-(4-chlorophenyl)-4-oxoquinazo-
lin-3(4H)-yl)acetohydrazide 4 were dissolved in warm ethanol with a
catalytic amount of glacial acetic acid and the reaction was refluxed
for>16 h. After completion (TLC), the solvent was evaporated in vac-
uum to half its volume and cooled to room temperature. After cooling,
the solid obtained was separated by filtration, washed with water, dried,
and recrystallized with ethanol to get the target compound 9a-p. The
target compounds were obtained as a mixture of Z and E isomers. The
percentage of isomers was calculated by NMR [80].
4.1. General procedure for the synthesis of 2-(4-chlorophenyl)quinazolin-
4(3H)-one 2
4.5.1. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((1,3-
diphenyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9a)
Anthranilamide 1 (20 mmol) was treated with 4-choloro benzalde-
hyde (20 mmol) in DMAC in the presence of NaHSO3 (40 mmol) and the
mixture was heated at 180 ◦C for 15 h. After completion of the reaction
(checked by TLC), the mixture was cooled to room temperature and
poured into ice–water (100 mL). The formed precipitate was collected
by filtration and washed successively with water. The product was
recrystallized from ethyl acetate/petroleum ether to provide 2-(4-
chlorophenyl)quinazolin-4(3H)-one 2 as pale yellow solid [60].
White powder; yield: 83%, mp 248–249 C. IR (KBr, cmꢀ 1): 3450
◦
1
–
–
–
–
–
–
(N H), 3053 (C H), 1668 (C O), 1649 (C O), 1639 (C N). H NMR
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 58%:42%) δ 11.83
(s, 1H, NH), 11.61 (s, 1H, NH), 9.11 (s, 1H, H5-pyr), 8.99 (s, 1H, H5-pyr),
[8.53–8.38 (m, 2H), 8.30–8.24 (m, 2H), 8.06–7.94 (m, 4H), 7.82–7.67
–
(m, 3H), 7.60–7.41 (m, 7H), 7.40–7.32 (m, 1H)] (19H, Ar H and
CH N), 5.67 (s, 2H, CH ), 5.25 (s, 2H, CH ). 13C NMR (101 MHz,
–
–
2
2
DMSO, mixture of two isomers) δ 167.8, 165.9, 165.7, 163.2, 157.5,
151.8, 151.2, 140.7, 138.9, 137.0, 135.9, 135.8, 135.7, 135.6, 134.6,
132.2, 131.9, 129.7, 129.6, 129.5, 129.5, 128.7, 128.6, 128.6, 128.5,
128.5, 128.4, 128.4, 128.3, 127.6, 127.5, 127.1, 126.9, 123.5, 123.3,
118.7, 118.6, 116.7, 116.5, 114.3, 114.3, 64.6, 63.5. Anal. Calcd for
4.2. General procedure for the synthesis of ethyl 2-(2-(4-chlorophenyl)-4-
oxoquinazolin-3(4H)-yl)acetate 3
The mixture of 2-(4-chlorophenyl)quinazolin-4(3H)-one 2 (15
mmol) and Anhydrous K2CO3 (30 mmol) in acetone (15 mL) stirred for
10 min at room temperature. Then, ethyl bromoacetate (15 mmol) was
added to the stirring solution and the reaction was continued at room
temperature for 15 h. Upon completion, checked by TLC, the mixture
was poured into 100 mL of water. The precipitated products 3 were
filtered off, washed with water, and recrystallized from ethanol–water
[78].
C
32H23ClN6O2: C, 68.75; H, 4.15; N, 15.03. Found: C, 68.02; H, 4.56; N,
15.66.
4.5.2. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((1-phenyl-3-
p-tolyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9b)
White powder; yield: 85%, mp 259–261 C. IR (KBr, cmꢀ 11): 3405
◦
–
–
–
–
–
–
(N H), 3120 (C H), 1689 (C O), 1655 (C O), 1644 (C N). H NMR
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 58%:42%) δ 11.82
(s, 1H, NH), 11.60 (s, 1H, NH), 9.09 (s, 1H, H5-pyr), 8.96 (s, 1H, H5-pyr),
[8.52–8.40 (m, 2H), 8.31–8.20 (m, 2H), 8.05–7.95 (m, 4H), 7.76–7.59
4.3. General procedure for the synthesis of 2-(2-(4-chlorophenyl)-4-
oxoquinazolin-3(4H)-yl)acetohydrazide 4
–
(m, 3H), 7.58–7.46 (m, 4H), 7.42–7.30 (m, 3H)] (18H, Ar H and
–
–
5 mL hydrazine hydrate was added to a solution of compound 3 (10
mmol) in ethanol (15 mL). The reaction mixture was heated under reflux
for 6 h. Reaction progress was monitored using TLC. After completion of
the reaction, the solvent volume was reduced to half the original volume
in a rotary evaporator, under reduced pressure. Upon cooling, the
separated solid was filtered, washed with water and dried. The solid
obtained was recrystallized with ethanol to get crystals of pure product 4
[78].
CH N), 5.70 (s, 2H, CH ), 5.25 (s, 2H, CH ), 2.40 (s, 3H, CH ), 2.33 (s,
3H, CH3). 13C NMR (101 MHz, DMSO, mix2ture of two isomer3s) δ 167.8,
166.0, 163.1, 157.5, 151.8, 151.2, 151.2, 140.8, 138.9, 138.1, 138.0,
137.1, 135.9, 135.6, 134.6, 129.7, 129.6, 129.5, 129.5, 129.3, 129.3,
129.1, 129.0, 128.6, 128.2, 128.1, 127.6, 127.5, 127.1, 126.8, 123.5,
123.3, 118.7, 118.6, 116.6, 116.4, 114.3, 114.3, 64.6, 63.5, 20.8, 20.8.
Anal. Calcd for C33H25ClN6O2: C, 69.17; H, 4.40; N, 14.67. Found: C,
69.56; H, 3.88; N, 14.98.
2
4.4. General procedure for the synthesis of 1,3-disubstituted pyrazole-4-
carbaldehydes 8a-p
4.5.3. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-(4-
methoxyphenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9c)
White powder; yield: 79%, mp 241–243 C. IR (KBr, cmꢀ 1): 3450
◦
1
–
–
–
–
–
–
A mixture of substituted acetophenone 6 (10 mmol) and 4-
substituted phenylhydrazine 7 (12 mmol) in ethanol (15 mL) was
heated under reflux in the presence of a catalytic amount of anhydrous
acetic acid (0.5 mL) at 70 ◦C for 8–12 h. After completion of the reaction,
the mixture was cooled at room temperature and the precipitate was
allowed to settle down, filtered, washed with ethanol and dried in vacuo
to afford hydrazone intermediate 7a-p. In the next step, hydrazone in-
termediate 7a-p (10 mmol) was added to a cold solution of dime-
thylformamide (10 mL) and phosphoryl chloride (30 mmol) and the
resulting mixture was heated at 60 ◦C for 5–8 h. After the completion of
the reaction (checked by TLC), the mixture was cooled and poured into a
slurry of ice/saturated solution of sodium bicarbonate. The solid product
obtained was filtered, washed with and crystallized in ethanol to afford
(N H), 3115 (C H), 1678 (C O), 1649 (C O), 1639 (C N). H NMR
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 58%:42%) δ 11.82
(s, 1H, NH), 11.60 (s, 1H, NH), 9.07 (s, 1H, H5-pyr), 8.96 (s, 1H, H5-pyr),
[8.51–8.42 (m, 2H), 8.34–8.18 (m, 2H), 8.03–7.98 (m, 4H), 7.74–7.67
(m, 3H), 7.57–7.50 (m, 4H), 7.43–7.27 (m, 1H), 7.12–7.06 (m, 2H)]
–
–
(18H, Ar H and CH N), 5.70 (s, 2H, CH ), 5.25 (s, 2H, CH ), 3.84 (s,
–
3H, OCH3), 3.76 (s, 3H, OCH3). 13C NMR (101 MHz, DMSO, 2mixture of
two isomers) δ 167.8, 166.0, 165.7, 163.1, 159.5, 159.4, 157.5, 151.6,
151.2, 151.0, 140.9, 138.9, 137.2, 135.9, 135.8, 135.7, 135.6, 134.6,
129.7, 129.7, 129.7, 129.6, 129.5, 129.5, 128.6, 128.2, 127.6, 127.5,
127.1, 126.8, 124.5, 124.2, 123.5, 123.3, 118.6, 118.5, 116.5, 116.2,
114.3, 114.3, 114.1, 113.9, 63.5, 55.2, 55.1. Anal. Calcd for
2
C33H25ClN6O3: C, 67.29; H, 4.28; N, 14.27. Found: C, 67.62; H, 3.88; N,
9

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
14.82.
C, 66.18; H, 3.29; N, 14.
4.5.4. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-(4-
hydroxyphenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9d)
4.5.8. N′-((3-(4-chlorophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)-2-
(2-(4-chlorophenyl)-4-oxoquinazolin-3(4H)-yl)acetohydrazide (9 h)
White powder; yield: 81%, mp 256–258 C. IR (KBr, cmꢀ 1): 3477
White powder; yield: 79%, mp 263–265 C. IR (KBr, cmꢀ 1): 3456
◦
◦
1
1
–
–
–
–
–
–
–
–
–
–
–
(N H), 3035 (C H), 1681 (C O), 1651 (C O), 1642 (C N). H NMR
(N H), 3100 (C H), 1683 (C O), 1662 (C O), 1650 (C N). H NMR
–
–
–
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 58%:42%) δ 11.91
(s, 1H, NH), 11.68 (s, 1H, NH), 9.15 (s, 1H, H5-pyr), 9.03 (s, 1H, H5-pyr),
[(8.63–8.49 (m, 2H), 8.43–8.29 (m, 2H), 8.13–8.07 (m, 4H), 7.85–7.81
(m, 1H), 7.74–7.58 (m, 6H), 7.49–7.43 (m, 1H), 7.05–6.99 (m, 2H)]
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 57%:43%) δ 11.70
(br s, 1H, NH), 9.11 (s, 1H, H5-pyr), 8.98 (s, 1H, H5-pyr), [8.51–8.37 (m,
2H), 8.31–8.19 (m, 2H), 8.07–7.93 (m, 4H), 7.85–7.89 (m, 2H),
–
7.74–7.70 (m, 1H), 7.61–7.47 (m, 6H), 7.42–7.33 (m, 1H)] (18H, Ar
H
(18H, Ar H and CH N), 5.82 (s, 2H, CH ), 5.36 (s, 2H, CH ). 13C NMR
and CH N), 5.65 (s, 2H, CH ), 5.25 (s, 2H, CH ). 13C NMR (101 MHz,
–
–
–
–
–
(101 MHz, DMSO, mixture of two isomers2) δ 167.8, 166.0, 165.7, 163.1,
157.9, 157.8, 157.5, 152.1, 151.4, 151.2, 141.0, 139.0, 137.4, 135.9,
135.9, 135.7, 135.6, 134.6, 129.7, 129.7, 129.7, 129.6, 129.5, 129.5,
128.6, 128.5, 128.0, 127.6, 127.5, 126.8, 126.7, 123.5, 123.3, 122.8,
122.6, 118.6, 118.4, 116.3, 116.1, 115.5, 115.3, 114.3, 64.6, 63.5. Anal.
Calcd for C32H23ClN6O3: C, 66.84; H, 4.03; N, 14.62. Found: C, 66.23; H,
4.35; N, 14.24.
DMSO, mixture of two isom2ers) δ 167.9, 166.0, 165.8, 157.5, 151.2,
150.3, 149.8, 140.2, 138.8, 136.8, 135.9, 135.9, 135.6, 134.6, 133.3,
133.2, 131.1, 130.1, 130.0, 129.7, 129.6, 129.6, 129.5, 128.9, 128.6,
128.6, 128.5, 128.5, 127.7, 127.6, 127.5, 127.0, 123.5, 123.3, 118.7,
118.6, 116.8, 114.3, 64.7, 63.5. Anal. Calcd for C32H22Cl2N6O2: C,
64.76; H, 3.74; N, 14.16. Found: C, 64.36; H, 3.32; N, 14.56.
2
2
4.5.9. N′-((3-(4-bromophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)-2-
(2-(4-chlorophenyl)-4-oxoquinazolin-3(4H)-yl)acetohydrazide (9i)
4.5.5. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-(4-
nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9e)
White powder; yield: 83%, mp 271–273 C. IR (KBr, cmꢀ 1): 3446
◦
White powder; yield: 85%, mp 268–270 C. IR (KBr, cmꢀ 1): 3448
(N H), 3060 (C H), 1680 (C O), 1660 (C O), 1646 (C N). H NMR
1
◦
–
–
–
–
–
–
–
–
1
–
–
–
–
–
–
(N H), 3045 (C H), 1680(C O), 1654 (C O), 1640 (C N). H NMR
(400 MHz, DMSO‑d6+, mixture of two isomers E:Z = 62%:38%) δ 11.83
(s, 1H, NH), 11.62 (s, 1H, NH), 9.11 (s, 1H, H5-pyr), 9.00 (s, 1H, H5-pyr),
[8.50–8.41 (m, 2H), 8.32–8.19 (m, 2H), 8.05–7.94 (m, 4H), 7.81–7.66
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 57%:43%) δ 11.96
(s, 1H, NH), 11.70 (s, 1H, NH), 9.17 (s, 1H, H5-pyr), 9.06 (s, 1H, H5-pyr),
[8.50–8.48 (m, 1H), 8.42–8.22 (m, 5H), 8.17–8.10 (m, 2H), 8.05–7.96
(m, 4H), 7.75–7.70 (m, 1H), 7.62–7.49 (m, 4H), 7.44–7.38 (m, 1H)]
–
(m, 5H), 7.59–7.47 (m, 4H), 7.41–7.36 (m, 1H)] (18H, Ar H and
CH N), 5.64 (s, 2H, CH ), 5.26 (s, 2H, CH ). 13C NMR (101 MHz,
–
–
2
2
(18H, Ar H and CH N), 5.61 (s, 2H, CH ), 5.28 (s, 2H, CH ). 13C NMR
DMSO, mixture of two isomers) δ 167.8, 165.9, 165.7, 163.2, 157.5,
151.2, 150.4, 149.8, 140.4, 138.8, 136.8, 135.9, 135.8, 135.7, 135.6,
134.6, 131.6, 131.5, 131.4, 131.1, 130.4, 130.4, 129.7, 129.6, 129.6,
129.5, 129.0, 128.6, 127.9, 127.6, 127.5, 127.0, 123.5, 123.3, 122.0,
121.9, 118.8, 118.6, 116.8, 116.6, 114.3, 64.6, 63.4. Anal. Calcd for
–
–
–
(101 MHz, DMSO, mixture of two isomers2) δ 167.9, 165.9, 157.5, 151.2,
148.6, 147.0, 138.8, 138.7, 136.5, 135.9, 135.6, 134.6, 130.0, 129.7,
129.6, 129.5, 129.5, 129.4, 128.9, 128.6, 127.6, 127.5, 127.3, 123.7,
123.6, 123.3, 118.9, 118.8, 117.4, 114.3, 64.5, 63.4. Anal. Calcd for
2
C
32H22ClN7O4: C, 63.63; H, 3.67; N, 16.23. Found: C, 63.25; H, 3.12; N,
C32H22BrClN6O2: C, 60.25; H, 3.48; N, 13.17. Found: C, 59.98; H, 3.71;
16.45.
N, 13.37.
4.5.6. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((1-phenyl-3-
(4(trifluoromethyl)phenyl)-1H-pyrazol-4-yl)methylene)acetohydrazide (9f)
4.5.10. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-phenyl-
1-p-tolyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9j)
White powder; yield: 81%, mp 260–262 C. IR (KBr, cmꢀ 1): 3438
White powder; yield: 78%, mp 282–285 C. IR (KBr, cmꢀ 1): 3438
◦
◦
1
1
–
–
–
–
–
–
–
–
–
–
–
(N H), 3055 (C H), 1684 (C O), 1660 (C O), 1638 (C N). H NMR
(N H), 3095 (C H), 1681 (C O), 1673 (C O), 1649 (C N). H NMR
–
–
–
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 61%:39%) δ 11.84
(s, 1H, NH), 11.63 (s, 1H, NH), 9.15 (s, 1H, H5-pyr), 9.04 (s, 1H, H5-pyr),
[8.52––8.44 (m, 1H), 8.43–8.38 (m, 1H), 8.31–8.23 (m, 2H), 8.05–7.97
(m, 6H), 7.90–7.82 (m, 2H), 7.75–7.68 (m, 1H), 7.61–7.49 (m, 4H),
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 60%:40%) δ 11.93
(s, 1H, NH), 11.71 (s, 1H, NH), 9.17 (s, 1H, H5-pyr), 9.04 (s, 1H, H5-pyr),
[8.63–8.51 (m, 2H), 8.42–8.31 (m, 2H), 8.13–8.10 (m, 2H), 8.01–7.98
(m, 2H), 7.89–7.80 (m, 3H), 7.70–7.52 (m, 5H), 7.48–7.42 (m, 2H)]
–
–
–
–
–
(18H, Ar H and CH N), 5.78 (s, 2H, CH ), 5.36 (s, 2H, CH ), 2.48 (s,
2 2
7.43–7.37 (m, 1H)] (18H, Ar H and CH N), 5.57 (s, 2H, CH ), 5.27 (s,
–
2
2H, CH2). 13C NMR (101 MHz, DMSO, mixture of two isomers) δ 167.8,
165.9, 163.3, 157.5, 151.2, 149.5, 138.8, 136.6, 135.9, 135.7, 135.6,
134.6, 129.7, 129.6, 129.6, 129.2, 129.2, 129.1, 128.6, 128.2, 127.6,
127.5, 127.1, 125.4, 125.3, 125.3, 123.5, 123.3, 118.8, 118.7, 117.1,
114.3, 64.6, 63.3. Anal. Calcd for C33H22ClF3N6O2: C, 63.21; H, 3.54; N,
13.40. Found: C, 63.44; H, 3.21; N, 13.82.
3H, CH3), 2.46 (s, 3H, CH3). 13C NMR (101 MHz, DMSO, mixture of two
isomers) δ 167.8, 165.9, 165.7, 163.1, 157.5, 151.6, 151.2, 150.9,
140.7, 137.1, 136.7, 136.3, 135.9, 135.8, 135.7, 135.6, 134.6, 132.2,
131.9, 129.9, 129.9, 129.7, 129.6, 128.7, 128.6, 128.5, 128.5, 128.5,
128.4, 128.3, 128.0, 127.6, 127.5, 126.9, 123.5, 123.3, 118.7, 118.5,
116.5, 116.3, 114.3, 114.3, 64.6, 63.5, 20.4. Anal. Calcd for
C
33H25ClN6O2: C, 69.17; H, 4.40; N, 14.67. Found: C, 69.42; H, 4.59; N,
4.5.7. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-(4-
fluorophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9g)
14.26.
White powder; yield: 85%, mp 259–261 C. IR (KBr, cmꢀ 1): 3448
4.5.11. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((1,3-dip-
tolyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9k)
◦
1
–
–
–
–
–
–
(N H), 3047 (C H), 1670 (C O), 1650 (C O), 1641 (C N). H NMR
–
–
White powder; yield: 82%, mp 266–268 C. IR (KBr, cmꢀ 1): 3458
◦
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 59%:41%) δ 11.91
(s, 1H, NH), 11.70 (s, 1H, NH), 9.19 (s, 1H, H5-pyr), 9.07 (s, 1H, H5-pyr),
[8.58–8.49 (m, 2H), 8.40–8.27 (m, 2H), 8.10–8.06 (m, 4H), 7.95–7.88
(m, 2H), 7.82–7.78 (m, 1H), 7.68–7.55 (m, 4H), 7.51–7.38 (m, 3H)]
1
–
–
–
–
–
–
(N H), 3022 (C H), 1694 (C O), 1653 (C O), 1643 (C N). H NMR
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 58%:42%) δ 11.80
(s, 1H, NH), 11.59 (s, 1H, NH), 9.03 (s, 1H, H5-pyr), 8.90 (s, 1H, H5-pyr),
[8.51–8.42 (m, 3H), 8.32–8.20 (m, 2H), 8.03–7.99 (m, 2H), 7.87 (d, J =
8.4 Hz, 2H), 7.75–7.71 (m, 1H), 7.67–7.60 (m, 2H), 7.54–7.48 (m, 2H),
(18H, Ar H and CH N), 5.74 (s, 2H, CH ), 5.34 (s, 2H, CH ). 13C NMR
–
–
–
(101 MHz, DMSO, mixture of two isomers2) δ 167.8, 165.9, 165.7, 163.4,
163.2, 161.0, 157.5, 151.2, 150.7, 150.1, 140.5, 138.9, 136.9, 135.9,
135.8, 135.7, 135.6, 134.6, 130.6, 130.6, 130.5, 130.5, 129.7, 129.6,
129.6, 129.5, 128.7, 128.7, 128.6, 128.5, 127.6, 127.5, 126.9, 123.5,
123.3, 118.7, 118.6, 116.6, 116.5, 115.7, 115.5, 115.3, 114.3, 64.6,
63.4. Anal. Calcd for C32H22ClFN6O2: C, 66.61; H, 3.84; N, 14.56. Found:
2
– –
– –
–
7.38–7.30 (m, 4H)] (17H, Ar H and CH NCH N), 5.70 (s, 2H, CH ),
5.25 (s, 2H, CH2), 2.40 (s, 3H, CH3), 2.36 (s, 3H, CH3), 2.35 (s, 3H, CH23),
2.33 (s, 3H, CH3). 13C NMR (101 MHz, DMSO, mixture of two isomers) δ
167.8, 165.9, 165.7, 163.1, 157.5, 151.6, 151.2, 151.0, 140.9, 138.0,
137.9, 137.2, 136.7, 136.2, 135.9, 135.8, 135.7, 135.6, 134.6, 129.9,
10

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
129.8, 129.7, 129.6, 129.3, 129.2, 129.1, 128.7, 128.5, 128.5, 128.4,
128.2, 127.8, 127.6, 127.5, 126.8, 123.5, 123.3, 118.6, 118.4, 116.4,
116.2, 114.3, 114.3, 64.6, 63.5, 20.8, 20.8, 20.4. Anal. Calcd for
7.86–7.83 (m, 1H), 7.72–7.60 (m, 4H), 7.48–7.42 (m, 2H)] (17H,
–
– –
Ar HAr H and CH N), 5.77 (s, 2H, CH ), 5.36 (s, 2H, CH ), 2.48 (s,
–
2
2
3H, CH3), 2.46 (s, 3H, CH3). 13C NMR (101 MHz, DMSO, mixture of two
isomers) δ 167.9, 166.0, 157.5, 151.2, 149.5, 136.8, 136.6, 136.4,
135.9, 135.6, 134.6, 133.1, 131.2, 130.9, 130.1, 130.0, 129.9, 129.9,
129.7, 129.6, 128.6, 128.6, 128.5, 128.5, 127.6, 127.5, 123.5, 123.3,
118.6, 118.5, 116.6, 114.3, 63.5, 20.4. Anal. Calcd for C33H24Cl2N6O2:
C, 65.24; H, 3.98; N, 13.83. Found: C, 65.66; H, 4.23; N, 14.32.
C
34H27ClN6O2: C, 69.56; H, 4.64; N, 14.32. Found: C, 69.81; H, 4.21; N,
14.61.
4.5.12. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-(4-
methoxyphenyl)-1-p-tolyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9l)
White powder; yield: 78%, mp 275–277 C. IR (KBr, cmꢀ 1): 3462
◦
1
(N H), 3018(C H), 1682 (C O), 1651 (C O), 1648 (C N). H NMR
4.5.16. N′-((3-(4-bromophenyl)-1-p-tolyl-1H-pyrazol-4-yl)methylene)-2-
(2-(4-chlorophenyl)-4-oxoquinazolin-3(4H)-yl)acetohydrazide (9p)
–
–
–
–
–
–
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 60%:40%) δ 11.91
(s, 1H, NH), 11.69 (s, 1H, NH), 9.11 (s, 1H, H5-pyr), 8.98 (s, 1H, H5-pyr),
[8.60–8.52 (m, 2H), 8.44–8.24 (m, 2H), 8.11 (s, 2H), 7.96 (d, J = 8.0 Hz,
2H), 7.82–7.85 (m, 3H), 7.60 (d, J = 8.1 Hz, 2H), 7.49–7.35 (m, 2H),
White powder; yield: 81%, mp 281–283 C. IR (KBr, cmꢀ 1): 3477
◦
1
–
–
–
–
–
–
(N H), 3142 (C H),1683 (C O), 1645 (C O), 1639 (C N). H NMR
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 61%:39%) δ 11.83
(s, 1H, NH), 11.61 (s, 1H, NH), 9.05 (s, 1H, H5-pyr), 8.93 (s, 1H, H5-pyr),
[8.51–8.38 (m, 2H), 8.32–8.18 (m, 2H), 8.01 (d, J = 3.6 Hz, 2H),
7.89–7.83 (m, 2H), 7.78–7.66 (m, 5H), 7.53–7.50 (m, 2H), 7.36–7.31
–
–
–
7.25–7.10 (m, 2H)] (17H, Ar H and CH N), 5.79 (s, 2H, CH ), 5.34 (s,
2
2H, CH2), 3.93 (s, 3H, OCH3), 3.85 (s, 3H, OCH3), 2.45 (s, 3H, CH3). 13
C
NMR (101 MHz, DMSO, mixture of two isomers) δ 167.8, 166.0, 165.7,
163.1, 159.5, 159.4, 157.5, 151.4, 151.2, 150.8, 140.9, 137.3, 136.7,
136.1, 135.9, 135.8, 135.7, 135.6, 134.6, 129.9, 129.8, 129.7, 129.6,
129.6, 129.6, 128.6, 128.5, 127.9, 127.6, 127.5, 126.9, 124.6, 124.3,
123.5, 123.3, 118.6, 118.4, 116.2, 116.0, 114.3, 114.3, 114.1, 113.9,
64.6, 63.5, 55.2, 55.1, 20.4. Anal. Calcd for C34H27ClN6O3: C, 67.71; H,
4.51; N, 13.94. Found: C, 67.41; H, 4.83; N, 13.25.
–
–
–
(m, 2H)] (17H, Ar H and CH N), 5.64 (s, 2H, CH ), 5.26 (s, 2H, CH ),
2
2
2.36 (s, 3H, CH3), 2.35 (s, 3H, CH3). 13C NMR (101 MHz, DMSO, mixture
of two isomers) δ 167.8, 165.9, 165.7, 157.5, 151.2, 149.6, 140.5, 136.9,
136.6, 136.4, 135.9, 135.6, 134.6, 131.6, 131.4, 130.4, 130.3, 129.9,
129.9, 129.7, 129.6, 128.7, 128.6, 127.6, 127.5, 123.3, 121.8, 118.7,
118.5, 116.6, 114.3, 63.4, 20.4. Anal. Calcd for C33H24BrClN6O2: C,
60.80; H, 3.71; N, 12.89. Found: C, 60.52; H, 4.11; N, 12.52.
4.5.13. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-(4-
hydroxyphenyl)-1-p-tolyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9m)
4.6.
α
-Glucosidase inhibition assay
-glucosidase inhibition property of quinazolinone-pyrazole 9a-
White powder; yield: 75%, mp 265–267 C. IR (KBr, cmꢀ 1): 3435
◦
1
–
–
–
–
–
–
(N H), 2972 (C H), 1689 (C O), 1641 (C O), 1639 (C N). H NMR
–
–
The
α
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 57%:43%) δ 11.79
(s, 1H, NH), 11.55 (s, 1H, NH), 8.98 (s, 1H, H5-pyr), 8.85 (s, 1H, H5-pyr),
[8.52–8.38 (m, 2H), 8.33–8.17 (m, 2H), 8.02 (d, J = 3.4 Hz, 2H), 7.85
(d, J = 8.5 Hz, 2H), 7.75–7.71 (m, 1H), 7.63–7.47 (m, 4H), 7.35–7.30
p were determined according to the previously reported method [66].
The α-glucosidase enzyme ((EC3.2.1.20, Saccharomyces cerevisiae, 20
U/mg) and p-nitrophenyl glucopyranoside as substrate were purchased
from Sigma-Aldrich. Enzyme and desired concentrations of nitrophenol-
–
–
–
(m, 2H), 6.94–6.88 (m, 2H)] (17H, Ar H and CH N), 5.71 (s, 2H,
α-D-glycopyranoside (PNP) were stored in potassium phosphate buffer
CH2), 5.25 (s, 2H, CH2), 2.36 (s, 3H, CH3), 2.34 (s, 3H, CH3). 13C NMR
(101 MHz, DMSO, mixture of two isomers) δ 167.7, 166.0, 163.1, 157.9,
157.8, 157.5, 151.9, 151.2, 141.1, 137.5, 136.8, 136.1, 135.9, 135.7,
135.6, 134.6, 129.9, 129.8, 129.7, 129.7, 129.6, 129.6, 128.6, 128.5,
127.7, 127.6, 127.5, 126.6, 123.5, 123.3, 122.9, 118.5, 118.3, 116.0,
115.8, 115.5, 115.2, 114.3, 64.6, 63.5, 20.4. Anal. Calcd for
(pH 6.8). The standard and all tested compounds were dissolved in
DMSO (10 mM) and further diluted with potassium phosphate buffer for
subsequent use in the enzyme inhibition assay (10% final
concentration).
Firstly, a mixture containing phosphate buffer phosphate (135
μ
L, pH
L)
L) were added to the 96-well plate and pre-
L of 4 mM of p-nitrophenyl
6.8,50 mM), various concentrations of tested compounds 9a-p (20
μ
C
33H25ClN6O3: C, 67.29; H, 4.28; N, 14.27. Found: C, 66.81; H, 4.52; N,
and enzyme solution (20
μ
◦
14.73.
incubated at 37 C for 10 min. Then, 25
μ
glucopyranoside as substrate was added to each well and incubation was
continued at 37 ◦C for 20 min. Finally, the absorbance change was
measured at 405 nm using a spectrophotometer (Gen5, Power wave xs2,
BioTek, America). DMSO and acarbose were used as the control and
standard inhibitor, respectively. The percentage of inhibition for target
compounds, control, and the standard inhibitor was calculated by using
the following formula:
4.5.14. 2-(2-(4-Chlorophenyl)-4-oxoquinazolin-3(4H)-yl)-N′-((3-(4-
nitrophenyl)-1-p-tolyl-1H-pyrazol-4-yl)methylene)acetohydrazide (9n)
White powder; yield: 82%, mp 269–271 C. IR (KBr, cmꢀ 1): 3500
◦
1
–
–
–
–
–
–
(N H), 3003 (C H), 1685 (C O), 1651 (C O), 1640 (C N). H NMR
–
–
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 59%:41%) δ 12.06
(s, 1H, NH), 11.81 (s, 1H, NH), 9.24 (s, 1H, H5-pyr), 9.13 (s, 1H, H5-pyr),
[8.64–8.59 (m, 1H), 8.52–8.35 (m, 5H), 8.25–8.22 (m, 1H)8.13–8.10
(m, 3H), 8.02–7.99 (m, 2H), 7.73 (d, J = 8.7 Hz, 1H), 7.65–7.61 (m, 2H),
%Inhibition = [(Abs control Abs sam ple)/Abs control] × 100
–
–
–
7.50–7.45 (m, 2H)] (17H, Ar H and CH N), 5.73 (s, 2H, CH ), 5.39 (s,
IC50 values of tested compounds were obtained from the nonlinear
regression curve (logit method). All experiments were performed in
triplicate and the results are expressed as the mean ± SD.
2H, CH2), 2.49 (s, 3H, CH3), 2.48 (s, 3H, CH3). 13C NMR (2101 MHz,
DMSO, mixture of two isomers) δ 167.9, 166.1, 165.9, 163.3, 157.5,
151.2, 148.3, 147.0, 140.0, 138.9, 136.8, 136.6, 136.5, 135.9, 135.6,
134.6, 134.5, 130.0, 129.7, 129.7, 129.5, 129.4, 129.4, 128.6, 128.6,
127.6, 127.5, 127.4, 127.3, 123.8, 123.7, 123.6, 123.3, 118.8, 118.6,
117.2, 114.3, 64.2, 63.4, 20.4. Anal. Calcd for C33H24ClN7O4: C, 64.13;
H, 3.91; N, 15.86. Found: C, 64.48; H, 3.65; N, 16.22.
4.7. Kinetics of enzyme inhibition
The kinetic analysis was performed under the above-mentioned re-
action condition. The mode of inhibition of the most active compound 9i
was investigated by varying the concentration of p-nitrophenyl
α-D-
4.5.15. N′-((3-(4-chlorophenyl)-1-p-tolyl-1H-pyrazol-4-yl)methylene)-2-
(2-(4-chlorophenyl)-4-oxoquinazolin-3(4H)-yl)acetohydrazide (9o)
glucopyranoside as substrate in the absence and presence of compound
9i at different concentrations. The enzyme solution (1 U/mL, 20 L) was
incubated with different concentrations of 0, 20, 40, and 60 M of the
L) for 15 min at 30 ◦C. The change in the
μ
White powder; yield: 75%, mp 272–274 C. IR (KBr, cmꢀ 1): 3456
◦
μ
1
–
–
–
–
–
–
(N H), 3132 (C H), 1683 (C O), 1662 (C O), 1648 (C N). H NMR
–
–
most potent compound 9i (20
μ
(400 MHz, DMSO‑d6, mixture of two isomers E:Z = 58%:42%) δ 11.86
(br s, 1H, NH), 9.18 (s, 1H, H5-pyr), 9.03 (s, 1H, H5-pyr), [8.60–8.49 (m,
2H), 8.42–8.30 (m, 2H), 8.14–8.00 (m, 2H), 8.01–7.88 (m, 4H),
absorbance was measured at 405 nm for 20 min using a spectropho-
tometer (Gen5, Power wave xs2, BioTek, America) following addition
increasing concentrations of p-nitrophenyl glucopyranoside (2–10 mM)
11

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
as substrate. A Lineweaver–Burk plot was generated to identify the type
of inhibition and the Michaelis–Menten constant (Km) value was
determined from the plot between reciprocal of the substrate concen-
tration (1/[S]) and reciprocal of enzyme rate (1/V) over various in-
hibitor concentrations. The experimental inhibitor constant (Ki) value
was constructed by secondary plots of the inhibitor concentration [I]
versus Km.
interaction.
4.11. Prime MM-GBSA
The ligand-binding energies (ΔGBind) were calculated for the best
active compound and acarbose using Molecular mechanics/generalized
¨
born surface area (MM–GBSA) modules (Schrodinger LLC 2018) based
on the following equation:
4.8. In vitro α-amylase inhibition assay
[
]
ΔGBind = ECom plex ꢀ EReceptor + ELigand
The anti-α-amylase activity of the compounds 9i, 9g, and 9k was
where ΔGBind is the calculated relative free energy which includes both
ligand and receptor strain energy. EComplex is the MM-GBSA energy of the
minimized complex, and ELigand is the MM-GBSA energy of the ligand
after removing it from the complex and allowing it to relax. EReceptor is
the MM-GBSA energy of relaxed protein after separating it from the
ligand. The MM-GBSA calculation was performed by extracting the total
number of 102 snapshots from the last 5 ns trajectory (production phase)
with the snapshot interval of 50 ps for binding free energy calculation.
determined based on the colorimetric method described by Taha et al.
[17].
4.9. Molecular docking
Docking studies were performed using genetic algorithm-based
docking program (GOLD) implemented in the Discovery Studio 4.1
(Accelrys Software Inc.) to study the binding modes and important in-
teractions of some selected synthesized compounds. The 3D structure of
compounds has been drawn by using Chem Draw Ultra 12.0 software.
The Ligand structures were transferred into Discovery Studio (DS), typed
with CHARMm force field and partial charges were calculated by
Momany-Rone option. Subsequently, the resulting structures were
minimized with Smart Minimizer algorithm, which performs 1000 steps
of steepest descent with a RMS gradient tolerance of 3, followed by
Conjugate Gradient minimization. The 3D Modeled structure of the
Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Appendix A. Supplementary material
α
-glucosidase was prepared using the protein preparation protocol of
DS. In this step, complex typed with CHARMm force field, hydrogen
atoms were added, all water molecules were removed and pH adjusted
to neutral 7.4. A 9 Å radius sphere was defined as the binding region for
docking study. Other parameters were set by default protocol settings.
Selected synthesized compounds were docked into the active site of the
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.bioorg.2021.105127.
References
α
-glucosidase and 10 different conformations for each compound were
[1] A.J. Hirsh, S. Yao, J.D. Young, C.I. Cheeseman, Inhibition of glucose absorption in
the rat jejunum: a novel action of alpha-D-glucosidase inhibitors, Gastroenterology
113 (1) (1997) 205–211.
ranked by GOLD score. Finally, the pose with the best GOLD score was
selected to analyze the mode of binding.
[2] A.J. Scheen, Is there a role for α-glucosidase inhibitors in the prevention of type 2
diabetes mellitus? Drugs 63 (10) (2003) 933–951.
[3] F.A. van de Laar, Alpha-glucosidase inhibitors in the early treatment of type 2
diabetes, Vasc. Health Risk Manage. 4 (6) (2008) 1189.
4.10. Molecular dynamic simulation
[4] S.R. Joshi, E. Standl, N. Tong, P. Shah, S. Kalra, R. Rathod, Therapeutic potential of
Molecular dynamic (MD) simulation of this study was performed by
using the Desmond module implemented in Maestro interface (from
α
-glucosidase inhibitors in type 2 diabetes mellitus: an evidence-based review,
Expert Opin. Pharmacother. 16 (13) (2015) 1959–1981.
[5] R. Pili, J. Chang, R.A. Partis, R.A. Mueller, F.J. Chrest, A. Passaniti, The
¨
Schrodinger 2018-4 suite). In order to build the system for MD simula-
α
-glucosidase I inhibitor castanospermine alters endothelial cell glycosylation,
tion, the protein–ligand complexes were solvated with SPC explicit
water molecules and placed in the center of an orthorhombic box of
appropriate size in the Periodic Boundary Condition. Sufficient counter-
ions and a 0.15 M solution of NaCl were also utilized to neutralize the
system and to simulate the real cellular ionic concentrations, respec-
tively. The MD protocol involved minimization, pre-production, and
finally production MD simulation steps. In the minimization procedure,
the entire system was allowed to relax for 2500 steps by the steepest
descent approach. Then the temperature of the system was raised from
0 to 300 K with a small force constant on the enzyme in order to restrict
any drastic changes. MD simulations were performed via NPT (constant
number of atoms, constant pressure i.e. 1.01325 bar and constant tem-
perature i.e. 300 K) ensemble. The Nose-Hoover chain method was used
as the default thermostat with 1.0 ps interval and Martyna-Tobias-Klein
as the default barostat with 2.0 ps interval by applying isotropic
coupling style. Long-range electrostatic forces were calculated based on
Particle-mesh-based Ewald approach with the cut-off radius for
columbic forces set to 9.0 Å. Finally, the system subjected to produce MD
simulations for 20 ns for protein–ligand complex. During the simulation
every 1000 ps of the actual frame was stored. The dynamic behavior and
structural changes of the systems were analyzed by the calculation of the
root mean square deviation (RMSD) and RMSF. Subsequently, the
energy-minimized structure calculated from the equilibrated trajectory
system was evaluated for investigation of each ligand–protein complex
prevents angiogenesis, and inhibits tumor growth, Cancer Res. 55 (13) (1995)
2920–2926.
´
[6] N. Zitzmann, A.S. Mehta, S. Carrouee, T.D. Butters, F.M. Platt, J. McCauley, B.
S. Blumberg, R.A. Dwek, T.M. Block, Imino sugars inhibit the formation and
secretion of bovine viral diarrhea virus, a pestivirus model of hepatitis C virus:
implications for the development of broad spectrum anti-hepatitis virus agents,
Proc. Natl. Acad. Sci. 96 (21) (1999) 11878–11882.
[7] A. Mehta, N. Zitzmann, P.M. Rudd, T.M. Block, R.A. Dwek, α-Glucosidase inhibitors
as potential broad based anti-viral agents, FEBS Lett. 430 (1–2) (1998) 17–22.
[8] A.J. Rawlings, H. Lomas, A.W. Pilling, M.J.R. Lee, D.S. Alonzi, J.S. Rountree, S.
F. Jenkinson, G.W. Fleet, R.A. Dwek, J.H. Jones, Synthesis and Biological
Characterisation of Novel N-Alkyl-Deoxynojirimycin α-Glucosidase Inhibitors,
ChemBioChem 10 (6) (2009) 1101–1105.
[9] H. Bischoff, Pharmacology of alpha-glucosidase inhibition, Eur. J. Clin. Invest. 24
(1994) 3–10.
[10] M. Toeller, α-Glucosidase inhibitors in diabetes: efficacy in NIDDM subjects, Eur. J.
Clin. Invest. 24 (S3) (1994) 31–35.
[11] P. Hollander, Safety profile of acarbose, an α-glucosidase inhibitor, Drugs 44 (3)
(1992) 47–53.
¨
[12] M. Ozil, M. Emirik, S.Y. Etlik, S. Ülker, B. Kahveci, A simple and efficient synthesis
of novel inhibitors of alpha-glucosidase based on benzimidazole skeleton and
molecular docking studies, Bioorg. Chem. 68 (2016) 226–235.
[13] T. Luthra, V. Banothu, U. Adepally, K. Kumar, M. Swathi, S. Chakrabarti, S.
R. Maddi, S. Sen, Discovery of novel pyrido-pyrrolidine hybrid compounds as
alpha-glucosidase inhibitors and alternative agent for control of type 1 diabetes,
Eur. J. Med. Chem. 188 (2020), 112034.
[14] X.-T. Xu, X.-Y. Deng, J. Chen, Q.-M. Liang, K. Zhang, D.-L. Li, P.-P. Wu, X. Zheng,
R.-P. Zhou, Z.-Y. Jiang, Synthesis and biological evaluation of coumarin derivatives
as α-glucosidase inhibitors, Eur. J. Med. Chem. 189 (2020), 112013.
[15] M. Taha, S. Imran, M. Salahuddin, N. Iqbal, F. Rahim, N. Uddin, A. Shehzad, R.
K. Farooq, M. Alomari, K.M. Khan, Evaluation and docking of indole sulfonamide
12

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
´
´
´
´
as a potent inhibitor of
α
-glucosidase enzyme in streptozotocin–induced diabetic
[40] M.E. Lopez-Viseras, B. Fernandez, S. Hilfiker, C.S. Gonzalez, J.L. Gonzalez, A.
´
albino wistar rats, Bioorg. Chem. 110 (2021), 104808.
J. Calahorro, E. Colacio, A. Rodríguez-Dieguez, In vivo potential antidiabetic
[16] M.T. Javid, F. Rahim, M. Taha, H.U. Rehman, M. Nawaz, S. Imran, I. Uddin,
activity of a novel zinc coordination compound based on 3-carboxy-pyrazole,
A. Mosaddik, K.M. Khan, Synthesis, in vitro
α-glucosidase inhibitory potential and
J. Inorg. Biochem. 131 (2014) 64–67.
´
´
´
molecular docking study of thiadiazole analogs, Bioorg. Chem. 78 (2018) 201–209.
[41] E. Hernandez-Vazquez, H. Ocampo-Montalban, L. Ceron-Romero, M. Cruz,
´
[17] M. Taha, S.A.A. Shah, M. Afifi, S. Imran, S. Sultan, F. Rahim, K.M. Khan, Synthesis,
J. Gomez-Zamudio, G. Hiriart-Valencia, R. Villalobos-Molina, A. Flores-Flores,
α
-glucosidase inhibition and molecular docking study of coumarin based
S. Estrada-Soto, Antidiabetic, antidyslipidemic and toxicity profile of ENV-2: A
potent pyrazole derivative against diabetes and related diseases, Eur. J. Pharmacol.
803 (2017) 159–166.
derivatives, Bioorg. Chem. 77 (2018) 586–592.
[18] M. Taha, S. Imran, F. Rahim, A. Wadood, K.M. Khan, Oxindole based oxadiazole
hybrid analogs: Novel
α
-glucosidase inhibitors, Bioorg. Chem. 76 (2018) 273–280.
[42] H. Li, R. Huang, D. Qiu, Z. Yang, X. Liu, J. Ma, Z. Ma, Synthesis and bioactivity of 4-
quinazoline oxime ethers, Prog. Nat. Sci. 8 (3) (1998) 359–365.
[43] N. Krishnarth, S.K. Verma, A. Chaudhar, Synthesis and anti-inflammatory activity
of some novel quinazolinone derivatives, FABAD J. Pharm. Sci. 45 (3) (2020)
205–210.
[19] Y.T. Han, K. Kim, G.-I. Choi, H. An, D. Son, H. Kim, H.-J. Ha, J.-H. Son, S.-J. Chung,
H.-J. Park, Pyrazole-5-carboxamides, novel inhibitors of receptor for advanced
glycation end products (RAGE), Eur. J. Med. Chem. 79 (2014) 128–142.
[20] Y. Zou, L. Xu, W. Chen, Y. Zhu, T. Chen, Y. Fu, L. Li, L. Ma, B. Xiong, X. Wang,
Discovery of pyrazole as C-terminus of selective BACE1 inhibitors, Eur. J. Med.
Chem. 68 (2013) 270–283.
[44] S. Farooq, A. Mazhar, N. Ullah, One-pot multicomponent synthesis of novel 3, 4-
dihydro-3-methyl-2 (1H)-quinazolinone derivatives and their biological evaluation
as potential antioxidants, enzyme inhibitors, antimicrobials, cytotoxic and anti-
inflammatory agents, Arabian J. Chem. 13 (12) (2020) 9145–9165.
[45] S. Malasala, J. Gour, M.N. Ahmad, S. Gatadi, M. Shukla, G. Kaul, A. Dasgupta,
Y. Madhavi, S. Chopra, S. Nanduri, Copper mediated one-pot synthesis of
quinazolinones and exploration of piperazine linked quinazoline derivatives as
anti-mycobacterial agents, RSC Adv. 10 (71) (2020) 43533–43538.
[46] A. Suresh, S. Srinivasarao, Y.M. Khetmalis, S. Nizalapur, M. Sankaranarayanan, K.
V.G.C. Sekhar, Inhibitors of pantothenate synthetase of Mycobacterium
tuberculosis–a medicinal chemist perspective, RSC Adv. 10 (61) (2020)
37098–37115.
[21] V. Aware, N. Gaikwad, S. Chavan, S. Manohar, J. Bose, S. Khanna, B.-R. Chandrika,
N. Dixit, K.S. Singh, A. Damre, Cyclopentyl-pyrimidine based analogues as novel
and potent IGF-1R inhibitor, Eur. J. Med. Chem. 92 (2015) 246–256.
[22] A. Kamal, A.B. Shaik, S. Polepalli, G.B. Kumar, V.S. Reddy, R. Mahesh,
S. Garimella, N. Jain, Synthesis of arylpyrazole linked benzimidazole conjugates as
potential microtubule disruptors, Bioorg. Med. Chem. 23 (5) (2015) 1082–1095.
[23] S. Kantevari, D. Addla, P.K. Bagul, B. Sridhar, S.K. Banerjee, Synthesis and
evaluation of novel 2-butyl-4-chloro-1-methylimidazole embedded chalcones and
pyrazoles as angiotensin converting enzyme (ACE) inhibitors, Bioorg. Med. Chem.
19 (16) (2011) 4772–4781.
[24] Y.-R. Li, C. Li, J.-C. Liu, M. Guo, T.-Y. Zhang, L.-P. Sun, C.-J. Zheng, H.-R. Piao,
Synthesis and biological evaluation of 1, 3-diaryl pyrazole derivatives as potential
antibacterial and anti-inflammatory agents, Bioorg. Med. Chem. Lett. 25 (22)
(2015) 5052–5057.
[47] T.S. Patel, J.D. Bhatt, R.B. Dixit, C.J. Chudasama, B.D. Patel, B.C. Dixit, Green
synthesis, biological evaluation, molecular docking studies and 3D-QSAR analysis
of novel phenylalanine linked quinazoline-4 (3H)-one-sulphonamide hybrid
entities distorting the malarial reductase activity in folate pathway, Bioorg. Med.
Chem. 27 (16) (2019) 3574–3586.
[25] T. Wang, D. Banerjee, T. Bohnert, J. Chao, I. Enyedy, J. Fontenot, K. Guertin,
H. Jones, E.Y. Lin, D. Marcotte, Discovery of novel pyrazole-containing benzamides
as potent RORγ inverse agonists, Bioorg. Med. Chem. Lett. 25 (15) (2015)
2985–2990.
[48] R. Venkatesh, S. Kasaboina, N. Jain, S. Janardhan, U.D. Holagunda, L. Nagarapu,
Design and synthesis of novel sulphamide tethered quinazolinone hybrids as
potential antitumor agents, J. Mol. Struct. 1181 (2019) 403–411.
[49] M.A.E.-A.M. El, M.S. Salem, S.A.M. Al-Mabrook, Synthesis and anticancer activity
of novel quinazolinone and benzamide derivatives, Res. Chem. Intermed. 44 (4)
(2018) 2545–2559.
[26] B.V. Kendre, M.G. Landge, S.R. Bhusare, Synthesis and biological evaluation of
some novel pyrazole, isoxazole, benzoxazepine, benzothiazepine and
benzodiazepine derivatives bearing an aryl sulfonate moiety as antimicrobial and
anti-inflammatory agents, Arabian J. Chem. 12 (8) (2019) 2091–2097.
[27] L.-G. Yu, T.-F. Ni, W. Gao, Y. He, Y.-Y. Wang, H.-W. Cui, C.-G. Yang, W.-W. Qiu,
The synthesis and antibacterial activity of pyrazole-fused tricyclic diterpene
derivatives, Eur. J. Med. Chem. 90 (2015) 10–20.
[50] S. Chen, M. Jiang, B. Chen, J. Salaenoi, S.-I. Niaz, J. He, L. Liu, Penicamide A, a
unique N, N′-ketal quinazolinone alkaloid from ascidian-derived fungus
Penicillium sp. 4829, Mar. Drugs 17 (9) (2019) 522.
[51] S. Zhou, G. Huang, Synthesis of anti-allergic drugs, RSC Adv. 10 (10) (2020)
5874–5885.
[28] R. Prasath, P. Bhavana, S. Sarveswari, S.W. Ng, E.R. Tiekink, Efficient ultrasound-
assisted synthesis, spectroscopic, crystallographic and biological investigations of
pyrazole-appended quinolinyl chalcones, J. Mol. Struct. 1081 (2015) 201–210.
[29] M. Khoobi, F. Ghanoni, H. Nadri, A. Moradi, M.P. Hamedani, F.H. Moghadam,
S. Emami, M. Vosooghi, R. Zadmard, A. Foroumadi, New tetracyclic tacrine
analogs containing pyrano [2, 3-c] pyrazole: efficient synthesis, biological
assessment and docking simulation study, Eur. J. Med. Chem. 89 (2015) 296–303.
[30] D. Manvar, S. Pelliccia, G. La Regina, V. Famiglini, A. Coluccia, A. Ruggieri,
S. Anticoli, J.-C. Lee, A. Basu, O. Cevik, New 1-phenyl-5-(1H-pyrrol-1-yl)-1H-pyr-
azole-3-carboxamides inhibit hepatitis C virus replication via suppression of
cyclooxygenase-2, Eur. J. Med. Chem. 90 (2015) 497–506.
[52] H. Jin, H.-G. Dan, G.-W. Rao, Research progress in quinazoline derivatives as multi-
target tyrosine kinase inhibitors, Heterocycl. Commun. 24 (1) (2018) 1–10.
[53] Z. Hajimahdi, R. Zabihollahi, M.R. Aghasadeghi, A. Zarghi, Design, Synthesis,
Docking Studies and Biological Activities Novel 2, 3-Diaryl-4-Quinazolinone
Derivatives as Anti-HIV-1 Agents, Curr. HIV Res. 17 (3) (2019) 214–222.
[54] M.K. Yadav, L. Tripathi, D. Goswami, Evaluation of Anticonvulsant Activity and
Toxicity Screening of Semicarbazones Derived from Quinazolinone Scaffold, Curr.
Bioact. Compd. 15 (5) (2019) 573–581.
[55] V.G. Ugale, S.B. Bari, S.C. Khadse, P.N. Reddy, C.G. Bonde, P.J. Chaudhari,
Exploring Quinazolinones as Anticonvulsants by Molecular Fragmentation
Approach: Structural Optimization, Synthesis and Pharmacological Evaluation
Studies, ChemistrySelect 5 (10) (2020) 2902–2912.
[31] P.N. Kalaria, S.P. Satasia, J.R. Avalani, D.K. Raval, Ultrasound-assisted one-pot
four-component synthesis of novel 2-amino-3-cyanopyridine derivatives bearing 5-
imidazopyrazole scaffold and their biological broadcast, Eur. J. Med. Chem. 83
(2014) 655–664.
[56] J. Rudolph, W.P. Esler, S. O’Connor, P.D. Coish, P.L. Wickens, M. Brands, D.
E. Bierer, B.T. Bloomquist, G. Bondar, L. Chen, Quinazolinone derivatives as orally
available ghrelin receptor antagonists for the treatment of diabetes and obesity,
J. Med. Chem. 50 (21) (2007) 5202–5216.
[32] M.F. Khan, M.M. Alam, G. Verma, W. Akhtar, M. Akhter, M. Shaquiquzzaman, The
therapeutic voyage of pyrazole and its analogs: a review, Eur. J. Med. Chem. 120
(2016) 170–201.
[57] V. Gurram, R. Garlapati, C. Thulluri, N. Madala, K.S. Kasani, P.K. Machiraju,
R. Doddapalla, U. Addepally, R. Gundla, B. Patro, Design, synthesis, and biological
[33] K. Kees, J. Fitzgerald Jr, K. Steiner, J. Mattes, B. Mihan, T. Tosi, D. Mondoro, M.
L. McCaleb, New Potent Antihyperglyceimc Agents in db/db Mice: Synthesis and
Structure-Activity Relationship Studies of (4-substitutedbenzyl)-(trifluoromethyl)
pyrazoles and-pyrazolones, J. Med. Chem 39 (1996) 3920–3928.
evaluation of quinazoline derivatives as α-glucosidase inhibitors, Med. Chem. Res.
24 (5) (2015) 2227–2237.
[58] K. Javaid, S.M. Saad, S. Rasheed, S.T. Moin, N. Syed, I. Fatima, U. Salar, K.M. Khan,
S. Perveen, M.I. Choudhary, 2-Arylquinazolin-4 (3H)-ones: A new class of
[34] S. Shu, X. Cai, J. Li, Y. Feng, A. Dai, J. Wang, D. Yang, M.-W. Wang, H. Liu, Design,
synthesis, structure–activity relationships, and docking studies of pyrazole-
containing derivatives as a novel series of potent glucagon receptor antagonists,
Bioorg. Med. Chem. 24 (12) (2016) 2852–2863.
α
-glucosidase inhibitors, Bioorg. Med. Chem. 23 (23) (2015) 7417–7421.
[59] M. Saeedi, M. Mohammadi-Khanaposhtani, P. Pourrabia, N. Razzaghi, R. Ghadimi,
S. Imanparast, M.A. Faramarzi, F. Bandarian, E.N. Esfahani, M. Safavi, Design and
synthesis of novel quinazolinone-1, 2, 3-triazole hybrids as new anti-diabetic
[35] S. Shu, A. Dai, J. Wang, B. Wang, Y. Feng, J. Li, X. Cai, D. Yang, D. Ma, M.-
W. Wang, A novel series of 4-methyl substituted pyrazole derivatives as potent
glucagon receptor antagonists: Design, synthesis and evaluation of biological
activities, Bioorg. Med. Chem. 26 (8) (2018) 1896–1908.
agents: in vitro α-glucosidase inhibition, kinetic, and docking study, Bioorg. Chem.
83 (2019) 161–169.
[60] M. Wei, W.-M. Chai, R. Wang, Q. Yang, Z. Deng, Y. Peng, Quinazolinone
[36] F. Chaudhry, S. Naureen, R. Huma, A. Shaukat, M. Al-Rashida, N. Asif, M. Ashraf,
derivatives: synthesis and comparison of inhibitory mechanisms on α-glucosidase,
M.A. Munawar, M.A. Khan, In search of new
α-glucosidase inhibitors:
Bioorg. Med. Chem. 25 (4) (2017) 1303–1308.
Imidazolylpyrazole derivatives, Bioorg. Chem. 71 (2017) 102–109.
[61] S. Moghimi, M. Toolabi, S. Salarinejad, L. Firoozpour, S.E.S. Ebrahimi, F. Safari,
S. Mojtabavi, M.A. Faramarzi, A. Foroumadi, Design and synthesis of novel
[37] F. Chaudhry, S. Naureen, S. Choudhry, R. Huma, M. Ashraf, M. Al-Rashida,
B. Jahan, M.H. Khan, F. Iqbal, M.A. Munawar, Evaluation of
α
-glucosidase
pyridazine N-aryl acetamides: In-vitro evaluation of α-glucosidase inhibition,
inhibiting potentials with docking calculations of synthesized arylidene-
pyrazolones, Bioorg. Chem. 77 (2018) 507–514.
docking, and kinetic studies, Bioorg. Chem. 102 (2020), 104071.
[62] E.S. Gosnell, S. Thikkurissy, Assessment and Management of Pain in the Pediatric
Patient, Pediatric Dentistry, Elsevier, 2019, pp. 97-115. e1.
[38] V. Pogaku, K. Gangarapu, S. Basavoju, K.K. Tatapudi, S.B. Katragadda, Design,
synthesis, molecular modelling, ADME prediction and anti-hyperglycemic
[63] M.U. Ahmad, M. Rafiq, B. Zahra, M. Islam, M. Ashraf, M. Al-Rashida, A. Khan,
J. Hussain, Z. Shafiq, A. Al-Harrasi, Synthesis of benzimidazole based hydrazones
evaluation of new pyrazole-triazolopyrimidine hybrids as potent
inhibitors, Bioorg. Chem. 93 (2019), 103307.
α-glucosidase
as non-sugar based α-glucosidase inhibitors: Structure activity relation and
[39] S.K. Sharma, A. Panneerselvam, K. Singh, G. Parmar, P. Gadge, O.C. Swami,
Teneligliptin in management of type 2 diabetes mellitus, Diabet. Metabolic
Syndrome Obesity: Targets Therapy 9 (2016) 251.
molecular docking, Drug Dev. Res. (2021).
[64] S. Khan, M. Tariq, M. Ashraf, S. Abdullah, M. Al-Rashida, M. Khalid, P. Taslimi,
M. Fatima, R. Zafar, Z. Shafiq, Probing 2-acetylbenzofuran hydrazones and their
metal complexes as α-glucosidase inhibitors, Bioorg. Chem. 102 (2020), 104082.
13

F. Azimi et al.
Bioorganic Chemistry 114 (2021) 105127
[65] S. Malik, A. Khan, M.M. Naseer, S.U. Khan, A. Ashraf, M. Ashraf, M. Rafiq,
K. Mahmood, M.N. Tahir, Z. Shafiq, Xanthenone-based hydrazones as potent
[72] F. Payan, Structural basis for the inhibition of mammalian and insect α-amylases by
plant protein inhibitors, Biochimica et Biophysica Acta (BBA) – Proteins Proteom.
1696 (2) (2004) 171–180.
α
-glucosidase inhibitors: synthesis, solid state self-assembly and in silico studies,
Bioorg. Chem. 84 (2019) 372–383.
[73] F. Azimi, J.B. Ghasemi, H. Azizian, M. Najafi, M.A. Faramarzi, L. Saghaei,
H. Sadeghi-Aliabadi, B. Larijani, F. Hassanzadeh, M. Mahdavi, Design and synthesis
[66] M. Adib, F. Peytam, M. Rahmanian-Jazi, S. Mahernia, H.R. Bijanzadeh, M. Jahani,
M. Mohammadi-Khanaposhtani, S. Imanparast, M.A. Faramarzi, M. Mahdavi, New
6-amino-pyrido [2, 3-d] pyrimidine-2, 4-diones as novel agents to treat type 2
of novel pyrazole-phenyl semicarbazone derivatives as potential α-glucosidase
inhibitor: Kinetics and molecular dynamics simulation study, Int. J. Biol.
Macromol. (2020).
diabetes: A simple and efficient synthesis, α-glucosidase inhibition, molecular
modeling and kinetic study, Eur. J. Med. Chem. 155 (2018) 353–363.
[67] M. Mohammadi-Khanaposhtani, S. Rezaei, R. Khalifeh, S. Imanparast, M.
A. Faramarzi, S. Bahadorikhalili, M. Safavi, F. Bandarian, E.N. Esfahani,
[74] S.A. Hollingsworth, R.O. Dror, Molecular dynamics simulation for all, Neuron 99
(6) (2018) 1129–1143.
[75] E. Fuglebakk, J. Echave, N. Reuter, Measuring and comparing structural
fluctuation patterns in large protein datasets, Bioinformatics 28 (19) (2012)
2431–2440.
M. Mahdavi, Design, synthesis, docking study, α-glucosidase inhibition, and
cytotoxic activities of acridine linked to thioacetamides as novel agents in
treatment of type 2 diabetes, Bioorg. Chem. 80 (2018) 288–295.
[76] T. Hou, J. Wang, Y. Li, W. Wang, Assessing the performance of the MM/PBSA and
MM/GBSA methods. 1. The accuracy of binding free energy calculations based on
molecular dynamics simulations, J. Chem. Inf. Model. 51 (1) (2011) 69–82.
[77] T. Hou, J. Wang, Y. Li, W. Wang, Assessing the performance of the MM/PBSA and
MM/GBSA methods: II. The accuracy of ranking poses generated from docking,
J. Comput. Chem. 32 (5) (2011) 866.
[68] Y. Wang, L. Ma, C. Pang, M. Huang, Z. Huang, L. Gu, Synergetic inhibition of
genistein and d-glucose on α-glucosidase, Bioorg. Med. Chem. Lett. 14 (11) (2004)
2947–2950.
[69] J.-M.G. Rodriguez, N.P. Hux, S.J. Philips, M.H. Towns, Michaelis-Menten graphs,
Lineweaver-Burk plots, and reaction schemes: investigating introductory
biochemistry students’ conceptions of representations in enzyme kinetics, J. Chem.
Educ. 96 (9) (2019) 1833–1845.
[78] E. Mentes¸e, N. Karaali, G. Akyüz, F. Yılmaz, S. Ülker, B. Kahveci, Synthesis and
evaluation of α-glucosidase and pancreatic lipase inhibition by quinazolinone-
[70] D. Shahzad, A. Saeed, F.A. Larik, P.A. Channar, Q. Abbas, M.F. Alajmi, M.I. Arshad,
coumarin hybrids, Chem. Heterocycl. Compd. 52 (12) (2016) 1017–1024.
[79] R. Pundeer, P. Ranjan, K. Pannu, O. Prakash, One-pot synthesis of some new
semicarbazone, thiosemicarbazone, and hydrazone derivatives of 1-phenyl-3-aryl-
pyrazole-4-carboxaldehyde from acetophenone phenylhydrazones using Vilsmeier-
Haack reagent, Synth. Commun. 39 (2) (2008) 316–324.
M.F. Erben, M. Hassan, H. Raza, Novel C-2 symmetric molecules as α-glucosidase
and α-amylase inhibitors: design, synthesis, kinetic evaluation, molecular docking
and pharmacokinetics, Molecules 24 (8) (2019) 1511.
[71] P. Lin, J.-C. Zeng, J.-G. Chen, X.-L. Nie, E. Yuan, X.-Q. Wang, D.-Y. Peng, Z.-P. Yin,
Synthesis, in vitro inhibitory activity, kinetic study and molecular docking of novel
[80] M. Mollazadeh, M. Mohammadi-Khanaposhtani, A. Zonouzi, H. Nadri, Z. Najafi,
B. Larijani, M. Mahdavi, New benzyl pyridinium derivatives bearing 2, 4-dioxo-
chroman moiety as potent agents for treatment of Alzheimer’s disease: Design,
synthesis, biological evaluation, and docking study, Bioorg. Chem. 87 (2019)
506–515.
N-alkyl–deoxynojirimycin derivatives as potential α-glucosidase inhibitors,
J. Enzyme Inhib. Med. Chem. 35 (1) (2020) 1879–1890.
14