Triazoloquinazolines as a new class of potent α-glucosidase inhibitors: In vitro evaluation and docking study

Article abstract of DOI:10.1371/journal.pone.0220379

Previously, we synthesized triazoloquinazolines 1-14 and characterized their structure. In this study, we aimed to evaluate the in vitro activity of the targets 1-14 as α-glucosidase inhibitors using α-glucosidase enzyme from Saccharomyces cerevisiae type 1. Among the tested compounds, triazoloquinazolines 14, 8, 4, 5, and 3 showed the highest inhibitory activity (IC₅₀ = 12.70 ± 1.87, 28.54 ± 1.22, 45.65 ± 4.28, 72.28 ± 4.67, and 83.87 ± 5.12 μM, respectively) in relation to that of acarbose (IC₅₀ = 143.54 ± 2.08 μM) as a reference drug. Triazoloquinazolines were identified herein as a new class of potent α-glucosidase inhibitors. Molecular docking results envisaged the plausible binding interaction between the target triazoloquinazolines and α-glucosidase enzyme and indicated considerable interaction with the active site residues.

Full text of DOI:10.1371/journal.pone.0220379

RESEARCH ARTICLE
Triazoloquinazolines as a new class of potent
α-glucosidase inhibitors: in vitro evaluation
and docking study
Hatem A. Abuelizz1, El Hassane Anouar2, Rohaya Ahmad3, Nor Izzati Iwana Nor Azman3,
1
Mohamed Marzouk4, Rashad Al-Salahi
*
ID
1 Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, Riyadh, Saudi
Arabia, 2 Department of Chemistry, College of Sciences and Humanities, Prince Sattam bin Abdulaziz
University, Al Kharj, Saudi Arabia, 3 Faculty of Applied Sciences, Universiti Teknologi MARA, shah Alam,
Selangor Darul Ehsan, Malaysia, 4 Chemistry of Natural Products Group, Centre of Excellence for Advanced
Sciences, National Research Centre, Dokki, Cairo, Egypt
a1111111111
a1111111111
a1111111111
a1111111111
a1111111111
* ralsalahi@ksu.edu.sa
Abstract
Previously, we synthesized triazoloquinazolines 1–14 and characterized their structure. In
this study, we aimed to evaluate the in vitro activity of the targets 1–14 as α-glucosidase
inhibitors using α-glucosidase enzyme from Saccharomyces cerevisiae type 1. Among the
tested compounds, triazoloquinazolines 14, 8, 4, 5, and 3 showed the highest inhibitory
activity (IC50 = 12.70 ± 1.87, 28.54 ± 1.22, 45.65 ± 4.28, 72.28 ± 4.67, and 83.87 ± 5.12 μM,
respectively) in relation to that of acarbose (IC50 = 143.54 ± 2.08 μM) as a reference drug.
Triazoloquinazolines were identified herein as a new class of potent α-glucosidase inhibi-
tors. Molecular docking results envisaged the plausible binding interaction between the tar-
get triazoloquinazolines and α-glucosidase enzyme and indicated considerable interaction
with the active site residues.
OPEN ACCESS
Citation: Abuelizz HA, Anouar EH, Ahmad R,
Azman NIIN, Marzouk M, Al-Salahi R (2019)
Triazoloquinazolines as a new class of potent α-
glucosidase inhibitors: in vitro evaluation and
docking study. PLoS ONE 14(8): e0220379.
https://doi.org/10.1371/journal.pone.0220379
Editor: Israel Silman, Weizmann Institute of
Science, ISRAEL
Received: February 7, 2019
Accepted: July 15, 2019
Published: August 14, 2019
Introduction
Copyright: © 2019 Abuelizz et al. This is an open
access article distributed under the terms of the
Creative Commons Attribution License, which
permits unrestricted use, distribution, and
reproduction in any medium, provided the original
author and source are credited.
Diabetes mellitus (DM) is a life-threatening, chronic metabolic disorder of multiple etiologies,
characterized by hyperglycemia accompanied with disturbances in protein, fat, and carbohy-
drate metabolism [1–4]. In 2011, approximately 366 million people were diagnosed with dia-
betes, according to the International Diabetes Federation (IDF) studies. This number is
predicted to reach 522 million by 2030. Diabetes severely damages several vital organs, leading
to heart attack, stroke, and neuropathy complications [5,6]. Diabetes has become pandemic in
human society owing to its prevalence, and high rate of associated mortality and morbidity
[7]. Thus, treating hyperglycemia and controlling the subsequent complications are the main
aims of diabetes therapy.
Data Availability Statement: All relevant data are
within the paper and its Supporting Information
files.
Funding: The authors extend their appreciation to
the Deanship of Scientific Research at King Saud
University for funding this study through research
group no. RG-1435–068.
According to a report by the Pharmaceutical Research and Manufacturers of America
(PhRMA), 182 new antidiabetic agents (30 for type 1, 100 for type 2, and 52 for related condi-
tion of DM) have been developed by American biopharmaceutical companies, and these drugs
are still either in clinical trials or under review by the Food and Drug Administration [7].
Competing interests: The authors have declared
that no competing interests exist.
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
1 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
Currently, a wide range of oral antidiabetic drugs are being utilized due to the absence of effi-
cient and affordable interventions [7]. In most cases, the prescribed antidiabetic drugs are
responsible for various side effects such as liver problems, diarrhea, lactic acidosis, and high
rate of secondary failure.
α-Glucosidase is present in the brush border membrane of the intestine. It catalyzes the
hydrolysis of α-(1!4)-glycosidic linkage of sugar (disaccharides and starch), releasing free
monosaccharides (α-D-glucose) during the final step of carbohydrate digestion. Thus, α-glu-
cosidase inhibitors (AGIs) can suppress postprandial hyperglycemia and decrease carbohy-
drate digestion rate; therefore, reduce the glucose level in the blood stream [1,8–13].
α-Glucosidase inhibitors are a unique class of anti-diabetic drugs, described as an attractive
therapeutic target for type-II diabetes and may also be used for other diseases in which carbo-
hydrates or its biosynthesis are potential targets including cancer, hyperlipoproteinemia, HIV,
and obesity [14,15]. In contrast to other oral antidiabetic drugs, AGIs exert their effect locally
in the intestine, rather than modulating some biochemical processes in the body. Accordingly,
extensive research has been carried out, elucidating the possible role of AGIs in the treatment
of prediabetic conditions, such as impaired glucose tolerance IGT and impaired fasting glucose
IFG [16].
α-Glucosidase inhibitors (acarbose, miglitol, and voglibose) are characterized by similar
structure to that of the sugar moieties (disaccharides and oligosaccharides); thus, they bind to
α-glucosidase via the carbohydrate site [17–19]. During the last two decades, the AGIs have
been introduced as antidiabetic drugs and recommended as the first line therapy by the IDF
and American Association of Clinical Endocrinologists (AACE) [20]. These drugs are well tol-
erated; however, their localized gastrointestinal adverse effects such as diarrhea, bloating, and
flatulence restrict the long-term acquiescence for treatment [8,9].
Recently, nitrogen-containing heterocyclic compounds without glycosyl, such as triazole,
quinazoline, imidazole, thiazole, and pyrazole, have been documented as potent in vitro AGIs.
Moreover, some natural and synthesized flavonoids (such as luteolin, naringenin, anthocya-
nins, and baicalein), coumarin, chromones and their derivatives (Fig 1 d-f) have been targeted
as potent AGIs [8,9,21–26]. Triazole and quinazoline derivatives have gained considerable
attention owing to their important pharmacological activities (Fig 1). For instance, carbazole-
linked triazole compounds have shown potent α-glucosidase inhibition activity in relation to
that of the standard acarbose (Fig 1C) [22]. Furthermore, substituted quinazolines have been
reported to possess antihyperglycemic and α-glucosidase inhibition effects (Fig 1A and 1B). In
particular, the modified 4-Cl/Br-2-phenyl-quinazolines reversibly inhibited α-glucosidase
enzyme activity in a non-competitive manner (Fig 1B), resulting in excellent inhibitory activity
comparable to that of acarbose [24].
In our previous study, we have shown that benzoquinazoline derivatives have the potential
to act as a new class of AGIs. Their IC50 value was 69.20 1.76, 59.60 0.52, 49.40 0.50,
50.20 0.37, and 88.20 0.89 μM, whereas that of acarbose was 143.54 μM [8]. In addition,
our 2-phenoxy-pyridotriazolopyrimidine was found to possess potent α-glucosidase inhibiti-
ory (104.07 4.89 μM) in relation to acarbose (143.54 μM) (Fig 1G) [9]. Particularly, our study
suggested that different types of interactions between benzoquinazolines and α-glucosidase,
such as H-bonding, free binding energy, closest amino acids, and hydrophobic effects, are key
factors involved in increasing the inhibitory effects [8]. Further, structure–activity relationship
(SAR) study on benzoquinazolines suggested that the type of functional groups attached at
position 2 of benzoquinazoline was found to increase α-glucosidase inhibitory activity [8] and
that the activity largely depends on electron donating/withdrawing substituents (Fig 1H).
The undesired effects of available AGIs have encouraged researchers to discover safer and
new generation of α-glucosidase inhibitor agents. Therefore, it is necessary to clarify how the
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
2 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
Fig 1. Synthetic routes for compounds 1–14.
https://doi.org/10.1371/journal.pone.0220379.g001
α-glucosidase inhibitory activity can be affected by combining the quinazoline and triazole
moieties in one molecule. However, to the best of our knowledge, there are no studies on the
relationship between triazoloquinazolines and α-glucosidase inhibition activity. To clarify and
gain deeper insight into the SAR of triazoloquinazolines with α–glucosidase, in the present
study, we evaluated the in vitro α-glucosidase inhibitory activity of the target triazoloquinazo-
lines (1–14).
Materials and methods
α-Glucosidase inhibitory assay
Chemicals and reagents. Sodium phosphate monobasic, sodium chloride, 0.2g of potas-
sium chloride and 0.2g potassium dihydrogen phosphate All chemical and reagent used were
purchased from Sigma-Aldrich (St Louis, Missouri).
Preparation of reagents. For phosphate-buffered saline (PBS), NaH2PO4 (1.4 g), NaCl (8
g), KCl (0.2 g), and KH2PO4 (0.2 g) were dissolved in deionized H2O (1 L) at 20˚C. Using
NaOH (1M) and HCl (1M), the pH of the solution was adjusted to 6.5 at 20˚C. p-Nitrophenyl-
α-D-glucopyranoside (2 mM) (p-NPG-substrate) was prepared by dissolving p-NPG (6 mg) in
buffer solution (10 mL). α-Glucosidase enzyme Type 1, lyophilized powder (Sigma-Aldrich)
from Saccharomyces cerevisiae (a mixture of MAL12 and MAL32) was employed. The enzyme
solution was prepared by dissolving 1 mg of in 1000 μL of cold PBS (pH 6.5). Subsequently,
50 μL of α-glucosidase enzyme solution was mixed with cold buffered saline (12 mL) to a
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
3 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
concentration of 0.125 unit/mL. For sample screening, a solution of 200 μg/mL concentration
was prepared in 100% DMSO, and then subjected to two-fold serial dilution in 5% DMSO in a
96-well microplate to obtain concentrations of 100, 50, 25, 12.5, 6.25, 3.13, and 1.56 μg/mL.
The samples were screened using Epoch 2 Microplate Spectrophotometer from BioTek. To
each well of the 96-well microplate, 10 μL of DMSO (5%) was added. The stock solution
(10 μL) was added to the first well and mixed. Then, 10 μL of sample was withdrawn from the
first well and added to the second well, and the process was repeated for two-fold serial dilu-
tion. Subsequently, 20 μL of α-glucosidase enzyme, 40 μL of PBS (pH 6.5), and 20 μL of deion-
ized H2O were added to each well of the 96-well microtiter plate with constant stirring. The
plate was pre-incubated at 37˚C for 10 min, and then 10 μL of p-NPG (2 mM) solution was
added to the mixture; its absorbance at 0 min was measured at a wavelength of 405 nm. The
reaction mixture was incubated at 37˚C for 30 min, and subsequently, its absorbance was mea-
sured. Dimethyl sulfoxide (5%) was used as the negative control and acarbose (200 μg/mL)
(Sigma-Aldrich) as the positive control. The experiment was performed in triplicate. Accord-
ing to a previously described methodology [27], the α-glucosidase inhibitory activity was
determined and the inhibition (%) of this activity was calculated using the following equation:
ðA30min À A0minÞcontrol À ðA30min À A0minÞexp
%inhibition ¼
� 100
ðA30min À A0minÞcontrol
Molecular docking. Autodock package was used to assess triazoloquinazolines as AGIs
[28]. The original docked ligands were downloaded from the RCSB data bank website with
PDB code 3W37 [28], and X-ray analysis was performed to coordinate the target α-glucosi-
dase. Molecular docking was performed according to Abuelizz et al [8, 9, 29].
Results and discussion
The target triazoloquinazolines 1–14 (Fig 2) were characterized in our previous study [30–34].
Cyclocondensation reaction of dialkyl-N-cyanoimido(dithio)carbonates with 2-hydrazinobne-
zoic acid afforded the parents 1 and 3, whereas the parent 2 was obtained by oxidation of 1
with H2O2 (Fig 3). Reaction of compounds 1–3 with appropriate alkyl(heteroalkyl)halides in
basic medium furnished the N-alkylated products 4–14.
In the present study, we assayed their enzymatic inhibitory activity against α-glucosidase.
The IC50 value of the target molecules are listed in Table 1 and S1 Fig
Generally, the structural diversity of triazoloquinazolines based on substituent groups at
positions 2 and 4 largely contribute to their diverse inhibitory activities.
Compounds 1–14 were investigated for their inhibitory potential against α-glucosidase,
and they demonstrated significant activity with the IC50 values ranging between 12.70 1.87
and 180.34 1.28 μM (Table 1 and S1 Fig). Triazoloquinazolines 3–5, 8, 12, and 14 were the
most potent α-glucosidase inhibitors with the IC50 values of 83.87 5.12, 45.65 4.28,
72.28 4.67, 28.54 1.22, 97.53 0,94, and 12.70 1.87 μM, respectively, in relation to that of
acarbose (IC50 = 143.54 μM). Triazoloquinazolines 1 and 2 showed good inhibitory effect
(155.86 1.36 and 180.43 1.28 μM), which was comparable to that of the standard acarbose,
whereas compounds 6, 7, 9–11, and 13 showed less than 50% inhibition, and were not
screened for their IC50 values. Triazoloquinazoline 14, 8, and 4 were more active, and approxi-
mately 12, 5, and 3 times more potent than acarbose, respectively. It is interesting to note that
all the target compounds have the same triazoloquinazoline scaffold and differ only in the sub-
stituents at positions 2 and/or 4 with the ‘parent’ compounds 1, 2 and 3 bearing a 2-SMe,
2-SO2Me or 2-OPh substituent and a free amine position 4. Therefore, the difference in their
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
4 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
Fig 2. Structure of the target triazoloquinazolines (1–14).
https://doi.org/10.1371/journal.pone.0220379.g002
inhibitory potential might be attributed to the structural diversity of the different substituents
designed at position 2 and/or 4-NH along with the presence of 8-Me or not, exploring a key
idea for a tentative SAR. This in vitro enzyme inhibitory study rationalized the preliminary
SAR of the parents 1–3 and indicated that the chemical modifications in such target com-
pounds produced eleven derivatives (4–14) with different enzymatic inhibitory activities.
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
5 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
Fig 3. Synthetic routes for compounds 1–14.
https://doi.org/10.1371/journal.pone.0220379.g003
The SAR study suggested that the inhibitory activity mainly relies on the presence of ali-
phatic or aromatic groups at the position R2. However, in case of aromatic group, it relies on
the type and position of its substituents. This activities changing depends on the presence of
an electron withdrawing or donating groups which might be responsible for its binding with
enzyme. Moreover, the presence of bulky moiety as aromatic group substituents affects on the
compound affinity toward the enzyme. In the present study, the substitution pattern of the
triazoloquinazoline scaffold at position 4 in parent compounds 1–3 induced potent activity.
Thus, the compounds 4 (Ethyl) and 5 (Allyl) were more potent, with a two- and three-fold
enhanced inhibitory activity ((IC50 = 72.28 4.67 and 45.65 4.28 μM) compared with that of
acarbose (IC50 = 143.54 2.08 μM). This might be because the transformed targets 4 and 5
attained adequate conformation to fit the active site of α-glucosidase. Similarly, the transfor-
mation of parent 3 into 14 drastically increased its inhibitory activity, suggesting that the pres-
ence of heteroalkyl group could enhance the affinity toward α-glucosidase. Moreover, this
indicated that the relative changes in triazoloquinazoline bulky structure lead to significant dif-
ference in its ability to inhibit α-glucosidase as shown by compounds 12 (N-Ethyl pyridine)
and 14 (3-CN-benzyl). Notably, a sharp decrease in the inhibitory activity was observed in all
compounds bearing a 4-aromatic substituent (6, 9, 10, 11, 13). Although, there was an increase
in the lipophilicity character, the methyl group in 6, 10, 11, 13 does not display any interaction
role with the active site of the glucosidase enzyme.
As mentioned in the introductory part regarding the benzoquinazolines (potent glucosidase
inhibitors), we observed that such derivatives bearing electron withdrawing group substituted
Table 1. α-Glucosidase inhibitory activity of triazoloquinazolines 1–14.
Sample
IC50 (μM)
155.86 1.36
1
2
180.34 1.28
3
83.87 5.12
4
45.65 4.28
5
72.28 4.67
6
16.24% inhibition at 0.28 μM
19.15% inhibition at 0.34 μM
28.54 1.22
7
8
9
10
16.89% inhibition at 0.24 μM
14.83% inhibition at 0.25 μM
19.52% inhibition at 0.23 μM
97.53 0.94
11
12
13
27.93% inhibition at 0.21 μM
12.70 1.87
14
Acarbose
143.54 2.08
https://doi.org/10.1371/journal.pone.0220379.t001
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
6 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
on benzyl group showed higher inhibitory effects. Recognizing the importance of the quinazo-
line moiety in several benzoquinazoline and triazoloquinazoline pharmaceutical compounds,
we investigated the modifications of our targets that might contribute to improve α-glucosi-
dase inhibitory activity. Moreover, our SAR study on benzoquinazolines validated that the
increasing of the number of H-bonds formed, the binding energy of the stable complex formed
between the docked benzoquinazolines, and the amino acids in the active site of the enzyme
and the number of aromatic rings positively enhanced the inhibitory effect [8].
In view of the aforementioned facts, one of our aims was to incorporate such electron with-
drawing substitutions in the target triazoloquinazolines 6, 9, 10, 11, and 13. Contrarily, the
inhibitory activity was abolished when parents 1 and 3 were transformed into 6, 9, 10, 11, and
13. This suggests that the insertion of aromatic substitutions with electron withdrawing groups
might either behave as a deactivating group that restricts the affinity between inhibitors and
enzyme or did not adequately fit the conformation of the enzyme’s active site.
Molecular docking results
The potential antidiabetic activity of the target triazoloquinazolines was explored by evaluating
their ability to inhibit α-glucosidase. The results revealed that compound 14 is the best inhibi-
tor of α-glucosidase. In light to our previous studies using similar parameters [8], molecular
docking study was carried out to elucidate the binding modes between the docked triazoloqui-
nazolines (4, 8, 12 and 14) and α-glucosidase enzyme. The binding energy, number of hydro-
gen bonds, and the number of closest residues surrounding the docked triazoloquinazolines 4,
8, 12 and 14 into the active site of α-glucosidase were determined (Table 2 and Figs 4 and 5).
All the stable complexes formed between the docked triazoloquinazolines and α-glucosidase
displayed negative binding energy. This indicates that the inhibition of α-glucosidase by the
tested compounds is favorable from thermodynamic point of view.
From the results in Table 2, it is obvious that the higher activity of 14 compared with that of
4, 8 and 12 is mainly refer to the number of hydrogen bonds formed in the 14-α-glucosidase
complex. Indeed, three hydrogen bonds were formed between compound 14 and the amino
acids in the active site of α-glucosidase, whilst only one hydrogen bond is formed between the
docked compounds (4, 8 and 12) and the amino acids into the active site of α-glucosidase. Tria-
zoloquinazoline 14 is substituted by phenoxy group at position 2 and propyl isoindole group at
position 4. Docking study results showed that compound 14 forms hydrogen bond between the
´
˚
oxygen atom of phenoxy group of 14 and Arg A 552 of distance 3.04 A. The second and third
hydrogen bonds are established between the carbonyl group of isoindole of compound 14 and
´
˚
Ala A 234 and Asn A 237 of distances 2.98 and 3.02 A, respectively. However, in case of the
docked 4, 8 and 12, only one conventional hydrogen bond was formed between the oxo group
´
˚
of quinazoline and the Arg A 552 of 4, 8 and 12 of distances 2.78, 2.86 and 2.94 A, respectively.
Compound 8 showed higher inhibition efficiency compared with 4, which may refer to the sta-
bility of 8-α-glucosidase (Table 2). Both compounds exhibit almost similar intermolecular inter-
action with the active amino acids of α-glucosidase (Figs 4 and 5). In 8-α- glucosidase complex,
Table 2. Docking binding energy and the inhibition activity of the docked triazoloquinazolines into the active site of α-glucosidase.
Synthesized derivatives
Free binding energy (kcal/mol)
Number of HBs
Number of closest residues
IC50 (μM)
45.56
4
-7.28
-7.74
1
1
7
8
12
7
28.54
-11.76
-10.48
-10.56
1
10
11
10
97.53
14
3
12.70
Acarbose
10
143.54
https://doi.org/10.1371/journal.pone.0220379.t002
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
7 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
Fig 4. 3D of the closest interactions between the active site residues of α-glucosidase and the highly active compounds 14, 4, 8, and
acarbose.
https://doi.org/10.1371/journal.pone.0220379.g004
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
8 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
9 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
Fig 5. 2D of the closest interactions between the active site residues of α-glucosidase and the moderately active compounds 14, 4, 8, and acarbose.
https://doi.org/10.1371/journal.pone.0220379.g005
the allyl group at position 4 of 8 forms two intermolecular interactions with ASP 232 and ALA
234 amino acids. The hydrogen atom of -CH2- of the allyl group of 8 forms a carbon hydrogen
´
˚
bond with the oxygen atom of the carbonyl group of ASP 232 of 2.15 A, which might explain
the stability of complex formed with 8 compared with the one formed with 4 (Figs 4 and 5).
ALA 234 of α-glucosidase forms an alkyl interaction with the hydrogen atom (= CH2) of allyl
group at position 4 of 8, whilst the ethylate position 4 in 4 showed no interactions with amino
acids of alpha-glucosidase. Both compounds exhibit Pi-Pi T-Shaped interactions with quinazo-
lin-4(1H)-one and TRP329 and TRP432 amino acids (Fig 5).
The reference acarbose showed moderate activity (IC50 = 143.54 μM) compared with that
of the active compounds 4, 8, 12 and 14. The hydrogen bonds are the main binding interaction
modes between the acarbose and the amino acids at the active site (Figs 4 and 5), which main-
tain complex stability. Indeed, 12 hydrogen bonds are formed between the active amino acids of
α-glucosidase -glucosidase. 10 of out of the 12 hydrogen bonds are established between the
hydroxyl groups of acarbose and the active amino acids of α-glucosidase. The two other hydro-
gen bonds are formed between (i) ASP 568 and hydrogen atom of amine NH in acarbose of a
´
˚
distance of 1.74 A; (ii) ASN 237 and the lone pair of the oxygen atom of tetrahydro-2H-pyrane
´
˚
in acarbose of a distance of 2.90 A. The other non-hydrogen bonding interactions are of type
carbon hydrogen bond, Pi-Alkyl and Sulfur-X interactions (Fig 5). The lower activity of the ref-
erence drug compared with 4, 8, 12 and 14 may refer to the fact that the core structure of the
triazoloquinazoline and the additional phenoxy can participate in the formation of p-p interac-
tions, which are not established in acarbose-alpha-glucosidase complex. For instance, in com-
pound 14, it is found that (i) Pi-Pi stacked interaction is formed between the phenyl ring of
´
˚
PHE 601 amino acid of α-glucosidase and the phenoxyl group of 14 with a distance of 5.91 A;
(ii) Pi-Pi T-shaped interaction is formed between the isoindoline moiety of TRP 432 amino acid
´
˚
and the triazole of 14 with a distance of 5.80 A; and (iii) Pi-Donor hydrogen between the hydro-
gen atom of the amide of ALA 234 and Pi orbitals of isoindoline-1,3-dionyl group of 14 with a
´
˚
distance of 4.08 A. Moreover, p-anion interactions might also exist as a noncovalent intermolec-
ular force between the additional phenyl groups of some of these compounds and ionized resi-
dues of amino acid. Overall, the results suggest multiple binding modes of these AGIs.
Conclusions
The target triazoloquinazolines 1–14 were identified as a novel class of potential AGIs. Their
structural features and various substitutions at positions 2 and 4 in the triazole and quinazoline
rings apparently played crucial roles in the inhibitory activity. However, difference in the α-
glucosidase inhibitory activity is conferred by the type of substituents at both positions in the
triazoloquinazoline ring. The substitution pattern in the quinazoline moiety at position 4 in
parent compounds 1–3 induced potent activity, with compound 3 being more potent. Triazo-
loquinazolines 4, 8, and 12 and 14 showed the highest potential inhibitory activity against α-
glucosidase and indicated their potential as potent lead candidates. Furthermore, molecular
modeling study confirmed the importance of binding energy in the stability of complex
formed between the docked triazoloquinazolines and the amino acid residues in the active site
of the enzyme. It also demonstrated considerable interaction with the active site residues,
which is in agreement with the reported IC50 values. Thus, triazoloquinazolines might be the
best candidates for designing and discovering novel AGIs, after in vitro/in vivo efficacy, safety,
and clinical studies.
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
10 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
Supporting information
S1 Fig. α-Glucosidase inhibitory activity of triazoloquinazolines.
(TIF)
Acknowledgments
The authors extend their appreciation to the Deanship of Scientific Research at King Saud
University for funding this study through research group no. RG-1435–068.
Author Contributions
Conceptualization: Hatem A. Abuelizz, Mohamed Marzouk, Rashad Al-Salahi.
Data curation: El Hassane Anouar, Mohamed Marzouk, Rashad Al-Salahi.
Funding acquisition: Rashad Al-Salahi.
Investigation: Rohaya Ahmad, Nor Izzati Iwana Nor Azman.
Methodology: Rohaya Ahmad, Nor Izzati Iwana Nor Azman.
Software: Hatem A. Abuelizz, El Hassane Anouar.
Validation: Rohaya Ahmad, Rashad Al-Salahi.
Writing – original draft: Hatem A. Abuelizz, El Hassane Anouar, Mohamed Marzouk,
Rashad Al-Salahi.
Writing – review & editing: Hatem A. Abuelizz, Mohamed Marzouk, Rashad Al-Salahi.
References
1. Zhen J, Dai Y, Villani T, Giurleo D, Simon JE, Wu Q. Synthesis of novel flavonoid alkaloids as α-glucosi-
dase inhibitors. Bioorg Med Chem. 2017; 25(20):5355–5364. https://doi.org/10.1016/j.bmc.2017.07.
055 PMID: 28797772
2. IDF Diabetes Atlas, 6th ed. 2014 update. Brussels, Belgium: International Diabetes Federation, Interna-
tional Diabetes Federation 2014.
3. Tundis R, Loizzo M, Menichini F. Natural products as alpha-amylase and alpha-glucosidase inhibitors
and their hypoglycaemic potential in the treatment of diabetes: an update. Mini Rev Med Chem. 2010;
10(4):315–331. PMID: 20470247
4. Guariguata L, Whiting D, Hambleton I, Beagley J, Linnenkamp U, Shaw J. Global estimates of diabetes
prevalence for 2013 and projections for 2035. Diabetes Res Clin Pract. 2014; 103(2):137–149. https://
doi.org/10.1016/j.diabres.2013.11.002 PMID: 24630390
5. Hati S, Madurkar SM, Bathula C, Thulluri C, Agarwal R, Siddiqui FA, Dangi P, Adepally U, Singh A,
Singh S, Sen S. Design, synthesis and biological evaluation of small molecules as potent glucosidase
inhibitors. Eur J Med Chem. 2015; 100:188–196. https://doi.org/10.1016/j.ejmech.2015.04.059 PMID:
26087029
6. Wang QQ, Cheng N, Yi WB, Peng SM, Zou XQ. Synthesis, nitric oxide release, and α-glucosidase inhi-
bition of nitric oxide donating apigenin and chrysin derivatives. Bioorg Med Chem. 2014; 22(5):1515–
1521. https://doi.org/10.1016/j.bmc.2014.01.038 PMID: 24508143
7. Kalita H, Boruah DC, Deori M, Hazarika A, Sarma R, Kumari S, Kandimalla R, Kotoky J, Devi R. Antidia-
betic and antilipidemic effect of Musa balbisiana root extract: A potent agent for glucose homeostasis in
streptozotocin-induced diabetic rat. Front Pharmacol. 2016; 7:102. https://doi.org/10.3389/fphar.2016.
00102 PMID: 27199747
8. Al-Salahi R, Ahmad R, Anouar E, Iwana Nor Azman NI, Marzouk M, Abuelizz HA. 3-Benzyl(phenethyl)-
2-thioxobenzo[g]quinazolines as a new class of potent α-glucosidase inhibitors: synthesis and molecu-
lar docking study. Future Med Chem. 2018; 10(16):1889–1905. https://doi.org/10.4155/fmc-2018-0141
PMID: 29882426
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
11 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
9. Abuelizz HA, Iwana NNI, Ahmad R, Anouar E, Marzouk M, Al-Salahi R, Synthesis, biological activity
and molecular docking of new tricyclic series as α-glucosidase inhibitor. BMC Chem. (2019) 13:52
https://doi.org/10.1186/s13065-019-0560-4 PMID: 31384800
10. Spasov AA, Babkov DA, Prokhorova TY, Sturova EA, Muleeva DR, Demidov MR, Osipov DV, Osyanin
VA, Klimochkin YN. Synthesis and biological evaluation of 2-acylbenzofuranes as novel α-glucosidase
inhibitors with hypoglycemic activity. Chem Biol Drug Des. 2017; 90(6):1184–1189. https://doi.org/10.
1111/cbdd.13038 PMID: 28585419
11. Iqbal S, Khan MA, Javaid K, Sadiq R, Fazal-Ur-Rehman S, Choudhary MI, Basha FZ. New carbazole
linked 1,2,3-triazoles as highly potent non-sugar α-glucosidase inhibitors. Bioorg Chem. 2017; 74:72–
81. https://doi.org/10.1016/j.bioorg.2017.07.006 PMID: 28756277
12. Zhang AJ, Rimando AM, Mizuno CS, Mathews ST. α-Glucosidase inhibitory effect of resveratrol and
piceatannol. J Nutr Biochem. 2017; 47:86–93. https://doi.org/10.1016/j.jnutbio.2017.05.008 PMID:
28570943
13. Wang G, Peng Y, Xie Z, Wang J, Chen M. Synthesis, α-glucosidase inhibition and molecular docking
studies of novel thiazolidine-2,4-dione or rhodanine derivatives. Med Chem Commun. 2017; 8:1477–
1484.
14. Adib M, Peytam F, Rahmanian-Jazi M, Mahernia S, Bijanzadeh HR, Jahani M et al. New 6-amino-pyr-
ido[2,3-d]pyrimidine-2,4-diones as novel agents to treat type 2 diabetes: A simple and efficient synthe-
sis, α-glucosidase inhibition, molecular modeling and kinetic study. Eur J Med Chem. 2018; 155:353–
363. https://doi.org/10.1016/j.ejmech.2018.05.046 PMID: 29902721
15. Ding SM, Lan T, Ye GJ, Huang JJ, Hu Y, Zhu YR et al. Novel oxazolxanthone derivatives as a new type
of α-glucosidase inhibitor: synthesis, activities, inhibitory modes and synergetic effect. Bioorg Med
Chem. 2018; 26(12):3370–3378 https://doi.org/10.1016/j.bmc.2018.05.008 PMID: 29776833
16. Chiasson JL, Robert GJ, Ramon G, Hanefeld M, Karasik A,Laakso M, et al. Acarbose treatment and
the risk of cardiovascular disease and hypertension in patients with impaired glucose tolerance: The
STOP-NIDDM trial. JAMA. 2003; 290(4):486–494. https://doi.org/10.1001/jama.290.4.486 PMID:
12876091
17. Salar U, Taha M, Khan KM, Ismail NH, Imran S, Perveen S et al. Syntheses of new 3-thiazolyl coumarin
derivatives, in vitro α-glucosidase inhibitory activity, and molecular modeling studies. Eur J Med Chem.
2016; 122:196–204. https://doi.org/10.1016/j.ejmech.2016.06.037 PMID: 27371923
18. Liu Z, Ma S. Recent advances in synthetic α-glucosidase inhibitors. Chem Med Chem. 2017; 12
(11):819–829. https://doi.org/10.1002/cmdc.201700216 PMID: 28498640
19. Imran S, Taha M, Ismail NH, Kashif SM, Rahim F, Jamil W et al. Synthesis of novel flavone hydrazones:
In-vitro evaluation of α-glucosidase inhibition, QSAR analysis and docking studies. Eur J Med Chem.
2015; 105:156–170. https://doi.org/10.1016/j.ejmech.2015.10.017 PMID: 26491979
20. Kalra S. Recent advances in endocrinology. J Pak Med Assoc. 2014; 64(4):474–476. PMID: 24864650
21. Zhang Y, Gao H, Liu R, Chen L, Li X, Zhao L et al. Quinazoline-1-deoxynojirimycin hybrids as high
active dual inhibitors of EGFR and α-glucosidase. Bioorg Med Chem Lett. 2017; 27(18):4309–4313.
https://doi.org/10.1016/j.bmcl.2017.08.035 PMID: 28838691
22. Wang G, Peng Z, Wang J, Li X, Li J. Synthesis, in vitro evaluation and molecular docking studies of
novel triazine-triazole derivatives as potential α-glucosidase inhibitors. Eur J Med Chem. 2017;
125:423–429. https://doi.org/10.1016/j.ejmech.2016.09.067 PMID: 27689725
23. Wang G, Wang J, He D, Li X, Li J, Peng Z. Synthesis and biological evaluation of novel 1,2,4-triazine
derivatives bearing carbazole moiety as potent α-glucosidase inhibitors. Bioorg Med Chem Lett. 2016;
26(12):2806–2809. https://doi.org/10.1016/j.bmcl.2016.04.071 PMID: 27177827
24. Gurram V, Garlapati R, Thulluri C, Madala N, Kasani KS, Machiraju PK et al. Design, synthesis, and bio-
logical evaluation of quinazoline derivatives as α-glucosidase inhibitors. Med Chem Res. 2015; 24
(5):2227–2237.
25. Garlapati R, Pottabathini N, Gurram V, Kasani KS, Gundla R, Thulluri C et al. Development of α-glucosi-
dase inhibitors by room temperature C-C cross-couplings of quinazolinones. Org Biomol Chem. 2013;
11(29):4778–4791. https://doi.org/10.1039/c3ob40636a PMID: 23778751
26. Wei M, Chai WM, Wang R, Yang Q, Deng Z, Peng Y. Quinazolinone derivatives: Synthesis and com-
parison of inhibitory mechanisms on α-glucosidase. Bioorg Med Chem. 2017; 25(4):1303–1308. https://
doi.org/10.1016/j.bmc.2016.09.042 PMID: 28110817
27. Ahmad R, Hashim HM, Noor ZM, Ismail NH, Salim F, Lajis NH et al. Antioxidant and antidiabetic poten-
tial of Malaysian Uncaria. Res J Med Plant. 2011; 5(5):587–595.
28. Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK, Goodsell DS et al. AutoDock4 and AutoDock-
Tools4: automated docking with selective receptor flexibility. J Comput Chem. 2009; 30(16):2785–
2791. https://doi.org/10.1002/jcc.21256 PMID: 19399780
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
12 / 13

Triazoloquinazolines as a new class of potent α-glucosidase inhibitors
29. Abuelizz HA, El-Dib RE, Marzouk M, Anouar EH, A Maklad Y, Attia HN et al. Molecular docking and
anticonvulsant activity of newly synthesized quinazoline derivatives. Molecules. 2017; 22(7):pii:E1094.
30. Al-Salahi R, Tahir KE, Alswaidan I, Lolak N, Hamidaddin M, Marzouk M. Biological effects of a new set
1,2,4-triazolo[1,5-a]quinazolines on heart rate and blood pressure. Chem Cent J. 2014; 8(1):3. https://
doi.org/10.1186/1752-153X-8-3 PMID: 24428932
31. Al-Salahi R, Marzouk M, Awad G, Al-Omar M, Ezzeldin E. Antimicrobial activity of newly synthesized
methylsulfanyl-triazoloquinazoline derivatives. J Pharm Pharmacol. 2013; 64(11):1678–1687.
32. Al-Salahi R, Alswaidan I, Al-Omar M, Marzouk M. Synthesis and antimicrobial of new 2-Phenoxy-4H-
[1,2,4]triazolo[1,5-a]quinazoline derivatives. Life Sci J. 2013; 10(4):2018–2028.
33. Al-Salahi R, Geffken D. Synthesis of novel 2-methylsulfanyl-4H-[1,2,4]triaz olo[1,5-a]quinazolin-5-one
and derivatives. Synth Commun. 2011; 41(23):3512–3523.
34. Al-Salahi R, Geffken D. Synthesis and reactivity of [1, 2, 4]triazolo-annelated quinazolines. Molecules.
2010; 15(10):7016–7034.
PLOS ONE | https://doi.org/10.1371/journal.pone.0220379 August 14, 2019
13 / 13