Synthesis, in?vitro evaluation and molecular docking studies of novel triazine-triazole derivatives as potential α-glucosidase inhibitors

Article abstract of DOI:10.1016/j.ejmech.2016.09.067

A novel series of triazine-triazole derivatives 7a–7m were synthesized, characterized by¹H NMR and evaluated for their α-glucosidase inhibitory activity. All the synthesized compounds displayed potent α-glucosidase inhibitory activity with IC₅₀range of 11.63?±?0.15 to 37.44?±?0.35?μM, when compared to the standard drug acarbose (IC₅₀?=?817.38?±?6.27?μM). Among the series, compound 7i (IC₅₀?=?11.63?±?0.15?μM) bearing 2,5-dichloro substitution at phenyl ring, represented the most potent α-glucosidase inhibitory activity. Molecular docking studies of the most active compounds with the homology modelled α-glucosidase were also performed to explore the possible inhibitory mechanism. Our studies shown that these triazine-triazole derivatives are a new class of α-glucosidase inhibitors.

Full text of DOI:10.1016/j.ejmech.2016.09.067

Accepted Manuscript
Synthesis, in vitro evaluation and molecular docking studies of novel triazine-triazole
derivatives as potential α-glucosidase inhibitors
Guangcheng Wang, Zhiyun Peng, Jing Wang, Xin Li, Juan Li
PII:
S0223-5234(16)30803-0
10.1016/j.ejmech.2016.09.067
EJMECH 8933
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 11 August 2016
Revised Date: 20 September 2016
Accepted Date: 21 September 2016
Please cite this article as: G. Wang, Z. Peng, J. Wang, X. Li, J. Li, Synthesis, in vitro evaluation and
molecular docking studies of novel triazine-triazole derivatives as potential α-glucosidase inhibitors,
European Journal of Medicinal Chemistry (2016), doi: 10.1016/j.ejmech.2016.09.067.
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ACCEPTED MANUSCRIPT
A novel series of triazine-triazole derivatives were synthesized and evaluated for
their α-glucosidase inhibitory activity. Among these compounds, 7i displayed the
most potent α-glucosidase inhibitory activity with IC50 values of 11.63±0.15 ꢀM as
compared to the standard drug acarbose (817.38±6.27 ꢀM).

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Synthesis, in vitro evaluation and molecular docking
studies of novel triazine-triazole derivatives as
potential α-glucosidase inhibitors
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Guangcheng Wang*, Zhiyun Peng, Jing Wang, Xin Li, Juan Li
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College of Chemistry and Chemical Engineering, Jishou University, Jishou 416000,
PR China
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*Corresponding Author
Tel.: +86 743 8563911
Fax: +86 743 8563911
E-mail address: wanggch123@163.com
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Abstract.
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A novel series of triazine-triazole derivatives 7a-7m were synthesized, characterized
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by H NMR and evaluated for their α-glucosidase inhibitory activity. All the
synthesized compounds displayed potent α-glucosidase inhibitory activity with IC50
range of 11.63±0.15 to 37.44±0.35 ꢀM, when compared to the standard drug acarbose
(IC50 = 817.38±6.27 ꢀM). Among the series, compound 7i (IC50 = 11.63±0.15 ꢀM)
bearing 2,5-dichloro substitution at phenyl ring, represented the most potent
α-glucosidase inhibitory activity. Molecular docking studies of the most active
compounds with the homology modeled α-glucosidase were also performed to
explore the possible inhibitory mechanism. Our studies shown that these
triazine-triazole derivatives are a new class of α-glucosidase inhibitors.
Keywords: 1,2,4-Triazine; 1,2,3-Triazole; Click chemistry; α-Glucosidase inhibitor;
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Molecular docking
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1. Introduction
Diabetes mellitus is a chronic metabolic disease which is characterized by high blood
sugar levels over a prolonged period [1]. Uncontrolled hyperglycemia can lead to
serious damage to many vital organs in the body, including kidney damage, heart
disease, and nerve damage [2, 3]. The goal of treatment of diabetes mellitus is
reduction of blood glucose levels and controlling subsequent complications.
α-Glucosidase is a membrane-bound enzyme at the epithelium of the small intestine
and play an important role in carbohydrate digestion [4]. Inhibition of α-glucosidase
can significantly decrease the postprandial hyperglycemia [5]. Thus, α-glucosidase
has been recognized as a therapeutic target for the treatment type-2 diabetes mellitus
and several α-glucosidase inhibitors have been used in clinic [6]. Furthermore,
α-glucosidase may also be used as therapeutic target for other carbohydrate mediated
diseases including cancer [7], HIV [8, 9] and hepatitis [10].
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1,2,4-Triazine is an important heterocyclic system, which is found in many
biologically active natural products such as fervenulin, toxoflavin, and reurhycin [11].
1,2,4-Triazine derivatives have been reported to exhibit a variety of biological
activities such as
antimalarial [12], anticonvulsant [13],
antifungal [14],
anti-inflammatory [15], anticancer [16], anti-HIV [17] and neuroprotective [18]
activities. Furthermore, recent studies have shown that some 1,2,4-triazine derivatives
have been identified to exhibit α-glucosidase inhibitory activity (Figure 1) [19, 20].
Such as, Rahim et al have reported that a novel series of triazinoindole derivatives as
inhibitors of α-glucosidase [19]. In our previous work, we have found that a new
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series of 1,2,4-triazine derivatives bearing carbazole or coumarin moieties show
potent α-glucosidase inhibitory activity [20, 21].
1,2,3-Triazoles is an important class of heterocyclic compounds, which have attracted
increasing attention in medicinal chemistry and drug discovery over the past decade
[22-25]. Previous studies revealed that 1,2,3-triazole derivatives possess a wide
variety of biological activities including antibacterial [26], antimalarial [27],
anticancer [28], anti-HIV [29] and antitubercular activities [30]. In particular, several
drugs containing 1,2,3-triazole group such as tazobactam, cephalosporin and
cefatrizine have been clinically used for the treatment of bacterial infections [31, 32].
On the other hand, recent studies have shown that numbers of compounds containing
the 1,2,3-triazole nucleus act as α-glucosidase inhibitors (Figure 1) [33, 34].
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Figure 1. Chemical structures of some α-glucosidase inhibitors containing
1,2,4-triazine or 1,2,3-triazole rings.
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In continuation of our interest in search of new α-glucosidase inhibitors [20], herein
we reported the synthesis of a novel series of triazine-triazole derivatives. The
synthesized compounds were tested for their in vitro α-glucosidase inhibitory activity.
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Furthermore, the structure-activity relationship (SAR) and molecular docking studies
of these compounds were also performed.
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2. Chemistry
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The triazine-triazole derivatives 7a-7m were synthesized as showed in Scheme 1.
Different substituted aniline 1a-1m reacted with 2-chloroacetyl chloride to obtain
compounds 2a-2m in excellent yields. Reaction of 2a-2m with NaN3 in DMF at 30 oC
for 24 h give the intermediates 3a-3m. Condensation of benzil 4 with
thiosemicarbazide in acetic acid at 120 oC for
3
h
provided the
5,6-diphenyl-1,2,4-triazine-3-thiol 5 [35], which was then reacted with propargyl
bromide in the presence of triethylamine as base to obtain compound 6. Finally,
triazine-triazole derivatives 7a-7m were achieved by reaction of compound 6 with
different intermediate 3 using sodium ascorbate and CuSO4·5H2O in DMF at room
temperature. The structures of all the new synthesized compounds 7a-7m were
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characterized by H NMR spectra. For instance, the H NMR spectrum of 7b (R =
4-Me) shown a singlet at δ 2.28 ppm due to methyl protons of the phenyl ring. Two
singlet signals at δ 4.69 and 5.01 ppm were corresponded to the methylene protons of
–S–CH2– and –CH2–CO–, respectively. The fourteen aromatic protons were appeared
as multiplet in the region of δ 7.06-7.51 ppm. The proton of –NH–CO– were appeared
at δ 8.03 as a singlet signal. A singlet of C5-H of triazole ring were observed at δ 7.89
ppm. All these data are in agreement with the structure of compound 7b.
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Scheme 1. Reagents and conditions: (a) Et3N, CH2Cl2, room temperature, 24 h; (b)
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NaN3, DMF, 30 C, 20 h; (c) AcOH, reflux, 3 h; (d) Et3N, MeOH, room temperature,
24 h; (e) sodium ascorbate, CuSO4·5H2O, DMF, r.t, 2 h.
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3. Results and discussion
3.1. α-Glucosidase inhibition assay
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The newly synthesized triazine-triazole derivatives 7a-7m were tested for their
α-glucosidase inhibitory activity by in vitro enzyme assay. The commercially
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available α-glucosidase inhibitors acarbose was used as a positive control for this assay
The results were shown in Table 1. All the tested compounds (7a-7m) displayed
potent α-glucosidase inhibitory activity with IC50 values of 27.71±0.31, 18.22±0.28,
20.17±0.25, 37.44±0.35, 25.69±0.29, 15.97±0.17, 19.13±0.21, 21.00±0.18,
11.63±0.15, 16.94±0.19, 15.24±0.20, 17.02±0.25 and 14.45±0.17 ꢀM, respectively,
when compared to the standard drug acarbose (IC50 = 817.38±6.27 ꢀM, The value of
IC50 is similar to previous literature report [36, 37]). Among all the tested molecules,
compound 7i (11.63±0.15 ꢀM) bearing 2,5-dichloro substitution at phenyl ring,
represented the most potent α-glucosidase inhibitory activity. It was found to be
seventy folds more active than the standard drug acarbose (817.38±6.27 ꢀM).
.
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Based on our results, the structure-activity relationship (SAR) of this class of
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compounds can be summarized. Introduction of electron-donating groups such as
methyl (7b, 7c and 7e), ethoxyl (7j and 7k) and methoxyl (7l) into the phenyl ring,
results in a slight increase the inhibitory activity, except the phenyl ring have ortho-
methyl group (7d). Furthermore, compounds 7f (4-Cl), 7g (2,4-Cl2), 7h (4-F), 7i
(2,5-Cl2) and 7m (3-CF3) with electron-withdrawing group also displayed potent
inhibitory activity, with IC50 values of 15.97±0.17, 19.13±0.21, 21.00±0.18,
11.63±0.15 and 14.45±0.17 ꢀM, respectively. Among them, compound 7i (2,5-Cl2,
IC50 = 11.63±0.15 ꢀM) was found to be the most active compound of the series. The
activity of compound 7g (2,4-Cl2, IC50 = 19.13±0.21 ꢀM ) was lower than compound
7i (2,5-Cl2, IC50 = 11.63±0.15 ꢀM), which indicates that the position of substituents
on the phenyl ring influences inhibitory activity. Additionally, 7m (IC50 = 14.45±0.17
ꢀM) with strong electron-withdrawing 3-CF3 substitution on the phenyl ring, was
found to be the second most active compound. In summary, these results indicated the
difference of biological activity among this class of compounds due to the pattern of
substitution in the phenyl ring. The binding interactions of the most active analogs
were confirmed through molecular docking studies.
Table 1. α-Glucosidase inhibitory activity of triazine-triazole derivatives 7a-7m.
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Compound
R
IC50 (ꢀM)a
27.71±0.31
18.22±0.28
20.17±0.25
37.44±0.35
H
7a
7b
7c
7d
4-Me
3-Me
2-Me
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2,4-Me2
4-Cl
25.69±0.29
15.97±0.17
19.13±0.21
21.00±0.18
11.63±0.15
16.94±0.19
15.24±0.20
17.02±0.25
14.45±0.17
817.38±6.27
7e
7f
2,4-Cl2
4-F
7g
7h
2,5-Cl2
4-OEt
2-OEt
4-OMe
3-CF3
7i
7j
7k
7l
7m
Acarbose
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a Acarbose is standard for α-glucosidase inhibition activity
3.2. Homology model
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The crystallographic structure of Saccharomyces cerevisiae α-glucosidase enzyme has
not been published yet, a number of homology models of α-glucosidase have been
reported in the literature [36, 38]. In order to expose the binding mode between the
compounds and Saccharomyces cerevisiae α-glucosidase at the molecular level, the
3D structure of α-glucosidase was built by means of modeller 9.15 homology
modeling software (http://salilab.org/modeller/). The sequence in FASTA format of
α-glucosidase was retrieved from UniProt (access code P53341). The crystallographic
structure of Saccharomyces cerevisiae isomaltase (PDB ID: 3AJ7) shows high
sequence similarity (72.4%) with α-glucosidase, which was selected as the template
for homology modeling.
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The quality of homology model was validated by the Ramachandran plot using the
PROCHECK (http://services.mbi.ucla.edu/PROCHECK/). The Ramachandran plot
(Figure 2) of the modelled α-glucosidase enzyme provided further evidence of the
model strength (92.3% of residues in most favored regions, 6.8% of residues in
additional allowed region, 0.8% of residues in generously allowed region and only
0.2% of residues in disallowed regions). The good results obtained from the
Psi/Phi Ramachandran plot suggested that the homology model could be used for the
next phase of docking studies.
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Figure 2. Ramachandran plot of the modelled α-glucosidase enzyme.
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3.3. Molecular docking
Molecular docking simulations was carried out to investigate the binding mode of
these compound with Saccharomyces cerevisiae α-glucosidase. The theoretical
binding mode between 7a and Saccharomyces cerevisiae α-glucosidase was shown in
Figure 3A. Compound 7a adopted a compact conformation in the pocket of the
α-glucosidase. The mono-phenyl ring of 7a bind at the bottom of the α-glucosidase
pocket and made a high density of van der Waals contacts, whereas the
diphenyltriazinyl group of 7a was positioned near the entrance of the pocket and
made only a few contacts. Detailed analysis showed that the mono-phenyl group of 7a
formed arene-cation interactions with the residue Arg-439. In addition, the
mono-phenyl group and the diphenyltriazinyl group of 7a formed CH-π interactions
with the residues Tyr-71 and Phe-177, respectively. It was shown that Glu-276 (bond
length: 2.0 Å) and Asp-349 (bond length: 2.9 Å) formed two hydrogen bonds with 7a,
which was the main interactions between 7a and α-glucosidase.
On the other hand, molecular docking studies of the standard drug acarbose with
α-glucosidase was also performed. The result was shown in Figure 3B. Acarbose
adopted a U-shaped conformation in the pocket of the α-glucosidase. The the
pyranose and cyclohexenyl rings of acarbose bind at the bottom of the α-glucosidase
pocket and made a high density of van der Waals contacts, whereas the two pyranose
rings in the middle of acarbose were positioned near the entrance of the pocket and
made only a few contacts. It was shown that Asp-68 (bond length: 3.1 Å), Gln-181
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(bond length: 2.0 Å) and Asp-349 (bond length: 1.6 Å) formed three hydrogen bonds
with acarbose, which was the main interactions between acarbose and α-glucosidase.
Results of molecular docking on α-glucosidase showed that compound 7a has similar
binding affinity as compared to standard drug.
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Figure 3. Compound 7a (A) and acarbose (B) were docked to the binding pocket of
the Saccharomyces cerevisiae α-glucosidase.
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To increase the activity of 7a, electron-withdrawing group (2,5-Cl2) was introduced to
the phenyl ring of 7a to obtain 7i. Compound 7i was docked to the binding pocket of
the Saccharomyces cerevisiae α-glucosidase, and the theoretical binding mode
between 7i and Saccharomyces cerevisiae α-glucosidase was shown in Figure 4.
Compound 7i adopted a compact conformation in the pocket of the α-glucosidase.
The 2,5-dichlorophenyl ring of 7i bind at the bottom of the α-glucosidase pocket and
made a high density of van der Waals contacts, whereas the diphenyltriazinyl group of
7i was positioned near the entrance of the pocket and made only a few contacts.
Detailed analysis showed that one of the phenyl group and the 2,5-dichlorophenyl
group of 7i formed arene-cation interactions with the residues Arg-312 and Arg-439,
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respectively. In addition, the dichlorophenyl group of 7i formed a π-π stacking with
the residue Tyr-71. It was shown that Glu-276 (bond length: 2.0 Å) and His-279 (bond
length: 2.4 Å) formed two hydrogen bonds with 7i, which was the main interactions
between 7i and α-glucosidase. In summary, the above molecular simulations give us
rational explanation of the interactions between 7i and α-glucosidase, which provided
valuable information for further development of α-glucosidase inhibitors.
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Figure 4. Compound 7i was docked into the binding pocket of the Saccharomyces
cerevisiae α-glucosidase.
4. Conclusion
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In conclusion, we designed and synthesized a novel series of triazine-triazole
derivatives 7a-7m. All the synthesized compounds were tested for their α-glucosidase
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inhibitory activity. Among them, compound 7i (IC50 = 11.63±0.15 ꢀM) having
2,5-dichloro substitution at phenyl ring was found to be the most active compound,
with seventy folds more active than the standard drug acarbose (IC50
=
817.38±6.27 ꢀM). Molecular docking studies showed that these triazine-triazole
derivatives were binding to the active site of α-glucosidase enzyme with the
hydrophobic interactions, arene-cation interactions, π-π interactions and hydrogen
bonds interactions. Hence, this study identified a new structural type of α-glucosidase
inhibitors, which could be used as lead molecules for further research and
development of potent α-glucosidase inhibitors.
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5. Experimental section.
5.1. Chemistry.
All starting materials and reagents were purchased from commercial suppliers. TLC
was performed on 0.20 mm Silica Gel 60 F254 plates (Qingdao Ocean Chemical
Factory, Shandong, China). Nuclear magnetic resonance spectra (NMR) were
recorded on a Bruker spectrometer (400 MHz) with TMS as an external reference and
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reported in parts per million. The Supplemental Materials contain sample H NMR
spectrum for 7a-7m (Figures S1–S13).
5.1.1. General procedures for the synthesis of triazine-triazole derivatives (7a-7m)
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A mixture of 6 (303 mg, 1.0 mmol), 3 (1.0 mmol), CuSO4·5H2O (0.025 g; 0.1 mmol)
and sodium ascorbate (0.10 g, 0.5 mmol) in DMF (20 mL) was stirred at room
temperature for 4 h. After the completion of the reaction, the mixture was poured into
100 mL of ice-cold water and the precipitate was filtered. The crude product was
purified by chromatography to give the title product 7a-7m.
5.1.1.1.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-phenyla
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cetamide (7a)
Yellow solid, yield 33.8 %, m.p. 197-199 C, 1H NMR (400 MHz, CDCl3) δ: 4.69 (s,
2H, SCH2), 5.12 (s, 2H, CH2CO), 7.11 (t, 1H, J = 7.2 Hz, ArH), 7.27 (d, 2H, J = 8.4
Hz, ArH), 7.32 (d, 2H, J = 7.6 Hz, ArH), 7.36 (d, 2H, J = 7.2 Hz, ArH), 7.40-7.43 (m,
4H, ArH), 7.47(d, 2H, J = 8.0 Hz, ArH), 7.51 (d, 2H, J = 7.6 Hz, ArH), 7.88 (s, 1H,
CH-triazole), 8.12 (s, 1H, NH).
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5.1.1.2.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(p-tolyl)
acetamide (7b)
Yellow solid, yield 63.1 %, m.p. 173-174 C, 1H NMR (400 MHz, CDCl3) δ: 2.28 (s,
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3H, CH3), 4.69 (s, 2H, SCH2), 5.01 (s, 2H, CH2CO), 7.06 (d, 2H, J = 8.4 Hz, ArH),
7.28-7.38 (m, 6H, ArH), 7.40-7.43 (m, 2H, ArH), 7.47(d, 2H, J = 8.4 Hz, ArH), 7.51
(d, 2H, J = 8.4 Hz, ArH), 7.89 (s, 1H, CH-triazole), 8.03 (s, 1H, NH).
5.1.1.3.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(m-tolyl
)acetamide (7c)
Yellow solid, yield 51.8 %, m.p. 156-158 C, 1H NMR (400 MHz, CDCl3) δ: 2.29 (s,
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3H, CH3), 4.69 (s, 2H, SCH2), 5.11 (s, 2H, CH2CO), 6.92 (d, 1H, J = 7.2 Hz, ArH),
7.15 (t, 1H, J = 8.0 Hz, ArH), 7.21 (d, 1H, J = 8.0 Hz, ArH), 7.26-7.30 (m, 1H, ArH),
7.32 (d, 2H, J = 7.6 Hz, ArH), 7.35 (d, 2H, J = 7.6 Hz, ArH), 7.39-7.42 (m, 2H, ArH),
7.47(d, 2H, J = 8.4 Hz, ArH), 7.51 (d, 2H, J = 8.4 Hz, ArH), 7.88 (s, 1H, CH-triazole),
8.13 (s, 1H, NH).
5.1.1.4.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(o-tolyl)
acetamide (7d)
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Yellow solid, yield 57.2 %, m.p. 110-112 C, H NMR (400 MHz, CDCl3) δ: 2.06 (s,
3H, CH3), 4.70 (s, 2H, SCH2), 5.15 (s, 2H, CH2CO), 7.06 (t, 1H, J = 7.2 Hz, ArH),
7.11 (d, 1H, J = 7.2 Hz, ArH), 7.17 (t, 1H, J = 8.0 Hz, ArH), 7.30-7.38 (m, 4H),
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7.40-7.44 (m, 2H, ArH), 7.47 (d, 2H, J = 8.4 Hz, ArH), 7.51 (d, 2H, J = 8.4 Hz, ArH),
7.77 (d, 1H, J = 8.0 Hz, ArH), 7.81 (s, 1H, NH), 7.89 (s, 1H, CH-triazole).
5.1.1.5.
N-(2,4-Dimethylphenyl)-2-(4-(((5,6-diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3
-triazol-1-yl)acetamide (7e)
o
Yellow solid, yield 59.7 %, m.p. 171-173 C, 1H NMR (400 MHz, CDCl3) δ: 2.01 (s,
3H, CH3), 2.25 (s, 3H, CH3), 4.70 (s, 2H, SCH2), 5.14 (s, 2H, CH2CO), 6.93 (s, 1H,
ArH), 6.95 (d, 1H, J = 8.0 Hz, ArH), 7.30-7.38 (m, 4H), 7.40-7.43 (m, 2H, ArH), 7.47
(d, 2H, J = 8.0 Hz, ArH), 7.51 (d, 2H, J = 8.4 Hz, ArH), 7.57 (d, 1H, J = 8.0 Hz, ArH),
7.68 (s, 1H, NH), 7.89 (s, 1H, CH-triazole).
5.1.1.6.
N-(4-Chlorophenyl)-2-(4-(((5,6-diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-tria
zol-1-yl)acetamide (7f)
o
Yellow solid, yield 42.7 %, m.p. 184-186 C, 1H NMR (400 MHz, CDCl3) δ: 4.67 (s,
2H, SCH2), 5.12 (s, 2H, CH2CO), 7.19 (d, 2H, J = 8.0 Hz, ArH), 7.29-7.35 (m, 3H,
ArH), 7.37-7.42 (m, 4H, ArH), 7.45 (d, 2H, J = 8.4 Hz, ArH), 7.50 (d, 2H, J = 7.2 Hz,
ArH), 7.88 (s, 1H, CH-triazole), 8.62 (s, 1H, NH).
5.1.1.7.
N-(2,4-Dichlorophenyl)-2-(4-(((5,6-diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3
-triazol-1-yl)acetamide (7g)
o
Yellow solid, yield 42.3 %, m.p. 195-197 C, 1H NMR (400 MHz, CDCl3) δ: 4.71 (s,
2H, SCH2), 5.17 (s, 2H, CH2CO), 7.20 (dd, 1H, J = 8.8 Hz, 2.0 Hz, ArH), 7.30-7.32
(m, 2H, ArH), 7.34-7.38 (m, 3H, ArH), 7.40-7.43 (m, 2H, ArH), 7.46 (d, 2H, J = 8.4
Hz, ArH), 7.51 (d, 2H, J = 8.4 Hz, ArH), 7.90 (s, 1H, CH-triazole), 8.20 (d, 1H, J =
8.0 Hz, ArH), 8.22 (s, 1H, NH).
5.1.1.8.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(4-fluor
ophenyl)acetamide (7h)
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Yellow solid, yield 75.7 %, m.p. 200-201 C, 1H NMR (400 MHz, CDCl3) δ: 4.69 (s,
2H, SCH2), 5.11 (s, 2H, CH2CO), 6.96 (t, 2H, J = 8.8 Hz, ArH), 7.30-7.34 (m, 2H),
7.36-7.45 (m, 6H), 7.47 (d, 2H, J = 8.4 Hz, ArH), 7.51 (d, 2H, J = 8.4 Hz, ArH), 7.88
(s, 1H, CH-triazole), 8.16 (s, 1H, NH).
5.1.1.9.
N-(2,5-Dichlorophenyl)-2-(4-(((5,6-diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3
-triazol-1-yl)acetamide (7i)
o
Yellow solid, yield 46.0 %, m.p. 160-162 C, 1H NMR (400 MHz, CDCl3) δ: 4.71 (s,
2H, SCH2), 5.18 (s, 2H, CH2CO), 7.02 (dd, 1H, J = 8.8 Hz, 2.4 Hz, ArH), 7.21 (d, 1H,
J = 8.8 Hz, ArH), 7.30-7.38 (m, 4H), 7.40-7.43 (m, 2H, ArH), 7.46 (d, 2H, J = 8.4 Hz,
ArH), 7.51 (d, 2H, J = 8.4 Hz, ArH), 7.90 (s, 1H, CH-triazole), 8.28 (s, 1H, NH), 8.35
(d, 1H, J = 2.4 Hz, ArH).
5.1.1.10.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(4-etho
xyphenyl)acetamide (7j)
o
1
Yellow solid, yield 12.5 %, m.p. 175-176 C, H NMR (400 MHz, CDCl3) δ: 1.38 (t,
3H, J = 7.2 Hz, OCH2CH3), 3.94 (q, 2H, J = 7.2 Hz, OCH2CH3), 4.69 (s, 2H, SCH2),
5.10 (s, 2H, CH2CO), 6.77 (d, 2H, J = 8.8 Hz, ArH), 7.29-7.32 (m, 3H, ArH),
7.34-7.38 (m, 3H, ArH), 7.40-7.44 (m, 2H, ArH), 7.47 (d, 2H, J = 8.4 Hz, ArH), 7.51
(d, 2H, J = 8.4 Hz, ArH), 7.88 (s, 1H, CH-triazole), 7.95 (s, 1H, NH).
5.1.1.11.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(2-etho
xyphenyl)acetamide (7k)
o
1
Yellow solid, yield 28.1 %, m.p. 178-179 C, H NMR (400 MHz, CDCl3) δ: 1.38 (t,
3H, J = 7.2 Hz, OCH2CH3), 3.94 (q, 2H, J = 7.2 Hz, OCH2CH3), 4.69 (s, 2H, SCH2),
5.10 (s, 2H, CH2CO), 6.77 (d, 2H, J = 8.8 Hz, ArH), 7.29-7.32 (m, 3H, ArH),
7.34-7.38 (m, 3H, ArH), 7.40-7.44 (m, 2H, ArH), 7.47 (d, 2H, J = 8.0 Hz, ArH), 7.51
(d, 2H, J = 8.4 Hz, ArH), 7.88 (s, 1H, CH-triazole), 7.96 (s, 1H, NH).
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5.1.1.12.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(4-meth
oxyphenyl)acetamide (7l)
Yellow solid, yield 41.4 %, m.p. 185-187 C, 1H NMR (400 MHz, CDCl3) δ: 3.75 (s,
o
3H, OCH3), 4.69 (s, 2H, SCH2), 5.10 (s, 2H, CH2CO), 6.78 (d, 2H, J = 8.8 Hz, ArH),
7.30-7.34 (m, 4H, ArH), 7.36 (d, 2H, J = 7.2 Hz, ArH), 7.40-7.44 (m, 2H, ArH), 7.47
(d, 2H, J = 7.6 Hz, ArH), 7.51 (d, 2H, J = 7.6 Hz, ArH), 7.89 (s, 1H, CH-triazole),
8.09 (s, 1H, NH).
5.1.1.13.
2-(4-(((5,6-Diphenyl-1,2,4-triazin-3-yl)thio)methyl)-1H-1,2,3-triazol-1-yl)-N-(3-(trifl
uoromethyl)phenyl)acetamide (7m)
Yellow solid, yield 36.5 %, m.p. 202-204 C, 1H NMR (400 MHz, CDCl3) δ: 4.69 (s,
o
2H, SCH2), 5.15 (s, 2H, CH2CO), 7.29-7.36 (m, 6H, ArH), 7.40-7.44 (m, 2H, ArH),
7.46 (d, 2H, J = 8.0 Hz, ArH), 7.50 (d, 2H, J = 8.0 Hz, ArH), 7.58 (d, 1H, J = 7.2 Hz,
ArH), 7.84 (d, 1H, J = 7.2 Hz, ArH), 7.90 (s, 1H, CH-triazole), 8.71 (s, 1H, NH).
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5.2. In vitro assay of α-glucosidase inhibitory activity
α-Glucosidase inhibitory activity was assayed by using 0.1 M phosphate buffer (pH
o
6.8) at 37 C. The enzyme (0.1 U/mL) in phosphate buffer saline was incubated with
o
various concentrations of test compounds at 37 C for 15 min. Then 1.25 mM
p-nitrophenyl α-D-glucopyranoside was added to the mixture as a substrate. After
further incubation at 37 oC for 30 min. The absorbance was measured
spectrophotometrically at 405 nm. The sample solution was replaced by DMSO as a
control. Acarbose was used as a positive control. All experiments were carried out in
triplicates. The% inhibition has been obtained using the formula: Inhibition (%) =
(1-∆Asample/∆Acontrol) * 100%. IC50 value is defined as a concentration of samples
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inhibiting 50% of α-glucosidase activity under the stated assay conditions.
5.3. Molecular docking
Molecular docking studies were performed to investigate the binding mode between
the compound 7a, 7i and acarbose with α-glucosidase using Autodock vina 1.1.2. The
3D structures of 7a, 7i and acarbose were obtained by ChemBioDraw Ultra 14.0 and
ChemBio3D Ultra 14.0 softwares. The AutoDockTools 1.5.6 package was employed
to generate the docking input files. The search grid of α-glucosidase was identified as
center_x: -19.676, center_y: -7.243, and center_z: -21.469 with dimensions size_x: 15,
size_y: 15, and size_z: 15. The value of exhaustiveness was set to 20. For Vina
docking, the default parameters were used if it was not mentioned. The best-scoring
poses as judged by the Vina docking score were chosen and visually analyzed using
PyMOL 1.7.6 software (http://www.pymol.org/).
Conflict of Interest
The authors confirm that this article content has no conflict of interest.
Acknowledgments
This work was supported by National Natural Science Foundation of China (Grant No.
81660574), Hunan Provincial Natural Science Foundation of China (2015JJ3099),
Scientific Research Fund of Hunan Provincial Education Department (15B194).
SUPPLEMENTARY DATA
Supplemental data for this article can be accessed on the publisher’s website at
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Highlights
►We designed and synthesized a new series of triazine-triazole derivatives.
► All the synthesized compounds displayed potent α-glucosidase inhibitory activity.
►Compound 7i was found to be the most active compound.
►The structure-activity relationship has been discussed.
►Molecular docking study was carried out to reveal the interaction between enzyme
and inhibitors.