Synthesis and antidiabetic evaluation of benzimidazole-tethered 1,2,3-triazoles

Article abstract of DOI:10.1002/ardp.202000090

Some novel benzimidazole-tethered 1,2,3-triazole derivatives (4a–r) were synthesized by a click reaction between 2-substituted 1-(prop-2-yn-1-yl)-1H-benzo[d]imidazole and in situ azide. The structures of the synthesized compounds were confirmed by spectroscopic studies (one- and two-dimensional nuclear magnetic resonance, Fourier transform infrared, and high-resolution mass spectra). The synthesized compounds were evaluated for their antidiabetic activity. Compounds 4a–r exhibited a good-to-moderate α-amylase and α-glucosidase inhibitory activity, with IC₅₀ values ranging from 0.0410 to 0.0916 μmol/ml and 0.0146 to 0.0732 μmol/ml, respectively. Compounds 4e, 4g, and 4n were found to be most active. Furthermore, the binding conformation of the most active compounds was ascertained by docking studies.

Full text of DOI:10.1002/ardp.202000090

Received: 27 March 2020
Revised: 12 May 2020
Accepted: 2 June 2020
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DOI: 10.1002/ardp.202000090
F U L L P A P E R
Synthesis and antidiabetic evaluation of
benzimidazole‐tethered 1,2,3‐triazoles
Laxmi Deswal1 | Vikas Verma1
| Devinder Kumar1 | Chander P. Kaushik1
|
Ashwani Kumar2
| Yogesh Deswal1 | Suman Punia1
1Department of Chemistry, Guru Jambheshwar
University of Science and Technology,
Hisar, India
Abstract
Some novel benzimidazole‐tethered 1,2,3‐triazole derivatives (4a–r) were synthe-
sized by a click reaction between 2‐substituted 1‐(prop‐2‐yn‐1‐yl)‐1H‐benzo[d]imi-
dazole and in situ azide. The structures of the synthesized compounds were
confirmed by spectroscopic studies (one‐ and two‐dimensional nuclear magnetic
resonance, Fourier transform infrared, and high‐resolution mass spectra). The syn-
thesized compounds were evaluated for their antidiabetic activity. Compounds 4a–r
exhibited a good‐to‐moderate α‐amylase and α‐glucosidase inhibitory activity, with
IC50 values ranging from 0.0410 to 0.0916 µmol/ml and 0.0146 to 0.0732 µmol/ml,
respectively. Compounds 4e, 4g, and 4n were found to be most active. Furthermore,
the binding conformation of the most active compounds was ascertained by docking
studies.
2Department of Pharmaceutical Sciences,
Guru Jambheshwar University of Science and
Technology, Hisar, India
Correspondence
Vikas Verma, Department of Chemistry,
Guru Jambheshwar University of Science
and Technology, Hisar 125001, India.
Email: vikas_chem_pu@yahoo.com
K E Y W O R D S
1,2,3‐triazole, antidiabetic, benzimidazole, docking, α‐amylase, α‐glucosidase
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1
INTRODUCTION
this class such as acarbose, voglibose, and miglitol are often reported
to cause side effects such as abdominal pain, diarrhea, and so forth.[7]
The International Diabetes Federation in 2019 reported that about
463 million people were suffering from diabetes, and this number is
supposed to increase at an alarming rate. It is estimated that 578
million people will have diabetes in 2030 and 700 million in 2045.[1]
Type‐II diabetes, which is the more common form of the disease
(accounts for 95% of the total diabetic population) is characterized
by a gradual increase in insulin resistance, followed by the dysfunc-
tioning of pancreatic beta cells, thus leading to hyperglycemia.[2]
Diabetes can also lead to other serious health issues such as vision
loss, nerve damage, kidney failure, and so forth. One of the important
tactics for treating this disease is to control and regulate the blood
glucose level by inhibiting carbohydrate hydrolytic enzymes.[3]
α‐Glucosidase is an important hydrolytic enzyme that plays a key
role in digestion of carbohydrates and it converts unabsorbed
disaccharides and oligosaccharides into monosaccharides.[4,5]
α‐Glucosidase inhibitors specifically inhibit α‐glucosidase in the brush
border of the enterocytes of the jejunum in the small intestine, thus
retarding the carbohydrate digestion.[6] Clinically accepted drugs of
Therefore, designing better drugs for the treatment of diabetes has
been a challenging area for medicinal chemists.
Benzimidazole can be considered as a privileged structure in med-
icinal chemistry, and its derivatives exhibit a variety of biological activities
including antidiabetic activity.[8–16] Among different triazoles, 1,2,3‐
triazoles particularly have a broad range of applications in the field of
pharmaceutical and medicinal chemistry, and they are a part of many
molecules reported as antidiabetic.[17–24] It has been reported that ben-
zimidazole compounds, that is, 2‐aryl benzimidazoles (A) and 2‐(2‐
chlorophenyl)‐7‐methyl‐1H‐benzo[d]imidazole (B), act as α‐glucosidase
inhibitors,[25–27] but there is still a need to discover some new analogs for
improved inhibitory activity. Various studies have reported that by
combining a benzimidazole ring with an electron‐withdrawing group such
as triazole, the biological activity gets enhanced.[28] Compound C, a
hybrid of 2,4,5‐triarylimidazole–triazole, and compound D, a hybrid of
xanthone–triazole, showed a good antidiabetic activity (Figure 1).[29,30]
Encouraged by the above facts, the synthesis of benzimidazole‐
tethered 1,2,3‐triazoles (4a–r), with the objective of discovering new
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(a)
(b)
(c)
(d)
FIGURE 1 The design strategy for the benzimidazole‐tethered 1,2,3‐triazoles
benzimidazole–triazole hybrid as a potent antidiabetic agent, is re-
two bands, that is, 1,600–1,620 cm−1 (amide I) and 1,680–1,695 cm−1
(amide II) in IR. In 1H spectra, the characteristic NH peak was ob-
served at δ 10.29–12.29, and another singlet was observed due to a
triazole proton at δ 8.11–8.41. Two singlets for the methylene group
attached to N16 of triazole ring and C13 of the triazole ring appeared
in the region of δ 5.25–5.49 and 5.46–5.55, respectively. In 13C NMR
spectra of the compounds, signal due to C13 of the triazole moiety
appeared at δ 142.94–143.48, whereas peak owing to C17 of triazole
ring resonated at δ 125.66–125.56. The carbonyl carbon appeared at
δ 164.00–165.53. The peaks of 1H and 13C spectrum were confirmed
by correlating them with different types of correlation spectroscopy,
ported in this study.
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2
RESULTS AND DISCUSSION
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2.1
Chemistry
A library of 18 compounds was synthesized in the current study.
Compounds 1a,b (Scheme 1) and 2‐substituted 1‐(prop‐2‐yn‐1‐yl)‐
1H‐benzo[d]imidazoles 2a,b were formed by using substituted
benzimidazoles and propargyl bromide using the reported proce-
dure.[23] The N‐substituted 2‐bromoacetamides 3a–i were synthe-
sized by reacting aniline derivatives and bromoacetyl bromide in
tetrahydrofuran using triethylamine as a base.[31] Target com-
pounds, substituted {4‐[(1H‐benzoimidazol‐1‐yl)methyl]‐1H‐1,2,3‐
triazol‐1‐yl}‐acetamides (4a–r), were synthesized by Cu(I)‐
catalyzed Huisgen 1,3‐dipolar cycloaddition reaction of alkyne and
azide in which in situ azide was formed from N‐substituted 2‐
bromoacetamides[23] (Scheme 2).
1a
The structure of synthesized compounds was confirmed by
various spectroscopic techniques, that is, one‐dimensional nuclear
magnetic resonance (NMR), two‐dimensional (2D) NMR, Fourier
transform infrared, and high‐resolution mass spectra (HRMS). The
formation of compounds 4a–r was ascertained by the presence of
1b
SCHEME 1 The synthesis of compounds 1a,b. Reagents and
conditions: (i) Acetic acid, reflux, 4–5 hr, (ii) (a) CS2, KOH, ethanol;
(b) CH3I, ethanol

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SCHEME 2 The synthesis of compounds 4a–r. Reagents and conditions: (i) K2CO3, propargyl bromide, DMF, 10–15°C, stirring 4–5 hr;
(ii) NaN3, CuSO4.5H2O, sodium ascorbate, DMF/H2O (8:2), 50°C, stirring 5–6 hr
like correlation spectroscopy (COSY), total correlation spectroscopy
(TOCSY), heteronuclear single‐quantum coherence (HSQC), and
heteronuclear multiple bond correlation (HMBC). The mass spectra
of the compounds showed signals due to [M+] and [M++1] ions, which
were in good agreement with their calculated values.
In the 1H spectra of compound 4j (Figure 2), one singlet at δ
8.13 ppm was assigned as a triazole proton, that is, C17–H. One
doublet of doublets of one proton at 7.61 was due to C6–H. The four
triplets at δ 7.54, 7.56, 7.32, and 7.08 ppm were attributed to C3–H,
The 13C NMR spectrum of compound 4j (Figure 2) indicated the
presence of 16 signals. Distortionless enhancement by polarization
transfer 13C NMR clearly indicated the presence of two secondary
carbons and eight tertiary carbons, thereby suggesting that the re-
maining six are quaternary carbons. To establish the assignment of
each carbon HSQC and HMBC, NMR experiments were carried out.
HSQC revealed the assignment of carbon signals at δ 129.34
(C22/C26), 125.57 (C17), 124.26 (C24), 122.02 (C1), 122.06 (C2),
119.74 (C23/C25), 118.01 (C6), 110.29 (C3), 52.68 (C18), 39.21 (C12),
→
→
C
22–H/C26–H, C23–H/C25–H, and C24–H, respectively. Two protons
and 14.93 (C11), because the key correlation was as follows: δ 8.13
→
→
→
118.01, 7.54
(C1–H and C2–H) gave a multiplet at δ 7.18 ppm due to C1–H and
C2–H, respectively. The above assignment was supported by 2D
NMR, for example, COSY and TOCSY. The 1H–1H correlation be-
tween C1–H/C2–H and C6–H, C1–H/C2–H and C3–H, C22–H/C26–H
and C23–H/C25–H, and C23–H/C25–H with both C22–H/C26–H and
125.57, 7.61
119.74, 7.18
110.29, 7.56
129.34, 7.32
→
124.26.
→
122.06 and 122.02, and 7.08
The HMBC experiment established the assignment of quaternary
carbon signals through a two‐bond correlation at δ 164.55 (C19),
152.68 (C8), 142.28 (C5), 143.45 (C13), 138.82 (C21), and 136.57 (C4).
The above assignment was confirmed through HMBC; the HMBC
C24–H was established through COSY spectrum. The TOCSY ex-
→
periment suggested the correlation of C3–H with C1–H/C2–H and
C6–H and C22–H/C26–H with C23–H/C25–H and C24–H.
correlation of signals is as follows: δ 5.46
152.68, 143.45,
→
→
119.74;
136.57, and 125.57; 5.29
125.57 and 164.51; 7.08

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FIGURE 2 The structure and 1H and 13C nuclear magnetic resonance signals of compound 4j
→
→
→
138.82 and 129.34; 7.56 119.74 and
7.18
118.01; 7.32
and compound 4e was most active for α‐glucosidase inhibition, with
→
→
143.45.
124.26; 7.61
122.02 and 122.06; 8.13
IC50 value 0.0146 µmol/ml.
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Molecular docking
2.2
Pharmacology
2.3
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2.2.1
Inhibition of α‐glucosidase and α‐amylase
To ascertain the binding conformation and interactions of the active
compounds with the active site residues, docking simulations were
carried out in the binding pocket of α‐glucosidase. The most stable
binding conformation of compounds 4e, 4g, and 4n along with the
interacting residues of the binding site is depicted in Figure 4.
Fluorine atom of the compound 4e seemed to be very im-
portant for binding with the enzyme. This atom formed one hy-
drogen bond with His348 (green dots) and made two halogen
bonds with Asp349 (light blue dots). Amide group NH created a
hydrogen bond with oxygen atom of Glu276. π Orbitals of fluor-
ophenyl ring had π–cation and π–anion interactions (yellow dots)
with Arg439 and Asp214, respectively. π Orbitals of benzimidazole
and triazole rings interacted with π electrons of Phe157 through
interesting T‐shaped π–π stacked interactions (pink dots). Similarly,
compound 4g displayed almost the same type of interactions, ex-
cept that of fluorine atom of 4e, as this is not present in 4g. In
compound 4n, m‐fluorine atom showed hydrogen bonding with
The inhibitory potential against α‐glucosidase and α‐amylase en-
zymes was tested using acarbose as a standard drug, and the IC50
values for both enzymes are listed in Table 1. All compounds showed
inhibitory activity in the range of IC50 = 0.0410–0.0916 µmol/ml for
α‐amylase and IC50 = 0.0146–0.0732 µmol/ml for α‐glucosidase, and
compounds 4e, 4g, 4h, 4n, and 4p were found to be good inhibitors.
In the case of α‐amylase inhibition, F and OCH3 group at R2
substitution were more potent. Furthermore, F group at the meta
position of the phenylacetamide ring was more active than F at para
position, except in 4h, and in α‐glucosidase inhibition, the same trend
was observed (Figure 3). The compounds with nonpolar substituents
were less active as compared with
a polar substituent. The
structure–activity relationship of all the compounds (4a–r) showed
that substitution at R1 exhibited variable results. Compound 4n was
most active for α‐amylase inhibition, with IC50 value 0.0410 µmol/ml,

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TABLE 1 IC50 values of compounds 4a–r
went deep into the active site, whereas acarbose could not pene-
trate the active site deeply due to its larger size. A cartoon diagram
of the protein along with docked acarbose and 4e, 4g, as well as 4n
is shown in Figure 5.
α‐Glucosidase
α‐Amylase inhibition inhibition
IC50
value
IC50
IC50 value value
(µmol/ml) (µg/ml)
IC50 value
(µmol/ml)
S. No. Compounds (µg/ml)
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2.4
Druglikeness of target compounds
1
4a
31.74
20.1
0.0916
0.0528
0.0559
0.0689
0.0534
0.0829
0.0448
0.0512
0.0795
0.0843
0.0542
0.0586
0.0641
0.0410
0.0834
0.0431
0.0513
0.0733
0.0237
20.95
19.94
15.55
19.53
5.304
26.72
5.8
0.0605
0.0524
0.0397
0.0503
0.0146
0.0681
0.0154
0.0268
0.0732
0.0602
0.0452
0.0393
0.0311
0.0162
0.0649
0.0200
0.0251
0.0447
0.0064
2
4b
4c
Druglikeness refers to the similarity of the properties and structural
features of the compounds with the known drug molecules. Some
important properties to determine drug‐likeness were calculated by
Molinspiration property calculator (https://www.molinspiration.com).
These properties give information about the bioavailability of organic
compounds in the human body. Various properties calculated are
listed in Table 2. It is interesting to note from the table that there is
zero violation (Nviolation) of Lipinski's rule of five, so it can be in-
ferred that all the target compounds follow this rule, that is, (a) the
molecular weight is <500, (b) the calculated partition coefficient
(miLogP) < 5, (c) <5 hydrogen bond donors (OH and NH moieties),
and (d) <10 hydrogen bond acceptors (especially N and O). Thus, they
are drug‐like compounds that can be used orally.[32] Veber et al.[33]
have suggested that the number of rotatable bonds should be less
than 10 and the polar surface area should be less than 140 Å. All
these synthesized molecules passed these criteria for good oral
bioavailability. There is zero violation, which confirms the good drug‐
likeness of the synthesized molecules.
3
21.89
26.75
19.44
32.54
16.88
18.64
31.53
31.89
22.39
24.82
26.94
16.24
32.75
17.59
20.33
31.41
15.31
4
4d
4e
5
6
4f
7
4g
8
4h
4i
9.76
9
29.03
22.8
10
11
12
13
14
15
16
17
18
19
4j
4k
4l
18.67
16.64
13.09
6.44
4m
4n
4o
4p
4q
4r
25.48
8.18
9.93
19.14
4.12
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3
CONCLUSION
Acarbose
Note: The high activity value is marked in bold.
Benzimidazole–1,2,3‐triazoles, designed as a potential antidiabetic agent,
were synthesized by employing a click reaction in DMF/H2O in a good
yield. The structure of the newly synthesized benzimidazole–1,2,3‐
triazoles was elucidated utilizing different spectroscopic techniques and
they were found to exhibit a good antidiabetic activity. Compounds 4e,
4g, 4n, and 4p showed good inhibitory activity against α‐amylase, and 4e,
4g, and 4n were found to be more potent against α‐glucosidase. Fur-
thermore, all the compounds displayed good absorption, distribution,
His111 and Gln181, and formed a halogen bond with Asp68. The
docking scores of these conformations were −10.4, −9.2, and
−8.2 kcal/mol for compounds 4e, 4g, and 4n, respectively. The ex-
perimental activity also followed the same order, that is,
4e > 4g > 4n. Thus, these computational study results were in a
good correlation with the experimental outcomes. Compounds
0.1
0.09
0.08
0.07
0.06
0.05
0.04
0.03
0.02
0.01
0
Compounds
FIGURE 3 α‐Amylase and α‐glucosidase
inhibitory activity of benzimidazole derivatives
α-amylase inhibition
α-glucosidase inhibition

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FIGURE 4 Binding interactions of compounds (a) 4e, (b) 4g, and (c) 4n in the active site of α‐glucosidase
metabolism, and excretion properties and zero violation of rules for
drug‐likeness. Docking studies showed that compound 4e went deeper
into the active site than acarbose, thereby effectively binding with the
enzyme. In addition, it also revealed the role of hydrogen bonding, ha-
logen bonding, and T‐shaped π–π interactions with the active site re-
sidues, and the docking scores were well correlated with the
experimental results. The correlation of docking score with in vitro
results further supported the experimental results. The findings of the
above investigations would certainly be helpful to the researchers
working in the field of diabetes in designing better therapeutics, and
these molecules may act as a good lead.
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4
EXPERIMENTAL
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4.1
Chemistry
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4.1.1
General
In the present study, chemicals were procured from Alfa Aesar/Sigma
Aldrich and used as such without further purification. To detect the
melting point of synthesized compounds, open capillary method was
used. Standard literature procedures were used for drying the solvents.
To perform thin‐layer chromatography (TLC), precoated Merck silica gel
(SIL G/UV254; ALUGRAM) plates were used in ethyl acetate/hexane
mixture solvent system, followed by observing spots under ultraviolet
light (254 nm). The 1H NMR (400 MHz) and 13C NMR (100 MHz) spectra
were taken in dimethyl sulfoxide (DMSO)‐d6 by Bruker Avance III. The
chemical shifts of DMSO for 1H (δ 2.50 ppm) and 13C (δ 39.50 ppm) were
used as a reference. The IR spectra of synthesized compounds were
recorded on Shimadzu IR Affinity‐I using KBr powder as a standard in
the region of 4,000–400 cm−1. The HRMS were recorded by mass
spectrometer Esquire 3000 with electrospray ionization (ESI) resources.
FIGURE 5 α‐Glucosidase containing docked acarbose (blue) and
compounds 4e (cyan), 4g (magenta), and 4n (brown)

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TABLE 2 Molinspiration properties
Compounds
miLogP
2.14
2.77
2.05
1.99
2.28
3.42
2.19
2.3
TPSA
77.64
77.64
123.47
94.71
77.64
77.64
86.88
77.64
77.64
77.64
77.64
123.47
94.71
77.64
77.64
86.88
77.64
77.64
N atoms
26
MW
nON
7
nOHNH
Nviolation
nRotB
Volume
4a
4b
4c
4d
4e
4f
346.39
380.84
391.39
388.43
364.38
392.49
376.42
364.38
396.45
378.46
412.91
423.46
420.5
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
1
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
5
5
6
6
5
6
6
5
5
6
6
7
7
6
6
7
6
6
310.67
324.20
334
27
7
29
10
8
29
346.21
315.6
27
7
28
7
345.36
336.21
315.6
4g
4h
4i
28
8
27
7
3.3
30
7
354.66
328.8
4j
2.97
3.6
27
7
4k
4l
28
7
342.33
352.13
364.34
333.73
345.36
354.34
333.73
372.79
2.88
2.82
3.11
3.42
3.02
3.13
4.13
30
10
8
4m
4n
4o
4p
4q
4r
30
28
396.45
392.49
408.49
396.45
428.52
7
28
7
29
8
28
7
31
7
Abbreviations: MW, molecular weight; miLogP, calculated partition coefficient; nRotB, number of rotatable bonds; TPSA, topological polar surface area.
The original spectra are provided as Supporting Information, as
are the InChI codes of the investigated compounds, together with
some biological activity data.
The reaction mixture was refluxed for 10 hr. Then the mixture was
poured in a beaker and (10%) sodium hydroxide and some con-
centrated hydrochloric acid were added until the mixture became
acidic to precipitate 2‐mercaptobenzimidazole, which was filtered,
dried, and then recrystallized from ethanol and water.
|
4.1.2
Synthesis of 2‐methyl‐1H‐benzimidazole (1a)
2‐Mercaptobenzimidazole (2.0 g, 0.0133 mol) was dissolved in
absolute ethanol (15 ml) with methyl iodide (0.0133 mol) and sodium
hydroxide (0.53 g, 0.0133 mol) in a round flask (50 ml) with a reflux
condenser. The reaction mixture was refluxed for 5 hr and filtered
directly to get rid of the precipitated salt; the filtered sample was
cooled and recrystallized from ethanol and water.
A synthetic method reported in the literature[34] was followed to
synthesize 1a. o‐Phenylenediamine (12.5 g) was treated with 11.25 g
of 90% acetic acid in a 250‐ml round‐bottom flask. The reaction
mixture was heated at 100°C in a water bath for 2 hr. The reaction
was monitored by TLC. After the completion of the reaction, the
reaction mixture was cooled and a 10% sodium hydroxide solution
was slowly added through mixing by continuous rotation of the flask
until the reaction mixture was just alkaline to litmus and precipitate
was formed. The precipitate was filtered, washed with ice‐cold water,
and then purified.
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4.1.4
Synthesis of 2‐substituted 1‐(prop‐2‐yn‐1‐yl)‐
1H‐benzo[d]imidazoles (2a,b)
Potassium carbonate (4 mmol) was added to a solution of substituted
benzimidazole (2 mmol) in 15 ml dimethylformamide, and the mixture
was stirred for 10 min. Propargyl bromide (2.4 mmol) was added
dropwise and the reaction mixture was stirred at 10–15°C for
4–5 hr.[23] The workup of the reaction was done with ice‐cold water
and the compound was extracted with ethyl acetate. The ethyl
acetate layer was dried over anhydrous Na2SO4, filtered, and con-
centrated under vacuum to yield 2a,b. The synthesized compounds
were recrystallized with ethanol (spectroscopic data were in agree-
ment with published data).
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4.1.3
Synthesis of 2‐(methylthio)‐
1H‐benzimidazole (1b)
2‐(Methylthio)‐1H‐benzimidazole 1b was synthesized by a two‐step
procedure according to an earlier reported method.[35] In a 250‐ml
round‐bottom flask, o‐phenylenediamine (2.0 g) was dissolved in ab-
solute ethanol (50 ml) and then carbon disulfide (14 ml) was added.

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4.1.5
Synthesis of N‐substituted
2H), 7.21–7.13 (m, 2H), 5.54 (s, 2H), 5.29 (s, 2H), and 2.67 (s, 3H,
CH3). 13C NMR (100 MHz, DMSO) δ = 164.79 (C═O), 142.96 (C13
triazole), 137.79, 129.31, 127.82, 125.60 (C17 triazole), 122.03,
121.80, 121.23, 118.63, 110.59, 63.74, 52.63, 38.62, and 14.24. IR
(KBr, ʋmax) = 3,251 (N–H str.), 3,135 (C–H str., triazole), 3,030 (C–H
str., aromatic), 1,695 (C═O str.), 1,622 (N–H bending), 1,556, 1,514,
and 1,458 (C═C str., aromatic) cm−1. HRMS (ESI): m/z calculated for
[M+H]+ C19H17N6ClO 381.1231; found 381.1229.
2‐bromoacetamides (3a–i)
The N‐substituted 2‐bromoacetamides 3a–i were synthesized by an
earlier reported method.[31] In a round‐bottom flask, substituted aniline
(1.0 mmol), bromoacetyl bromide (1.0 mmol) and triethylamine
(1.1 mmol) were stirred in tetrahydrofuran at 0–10°C for 3–4 hr. The
mixture was diluted with dichloromethane (50 ml), washed with satu-
rated ammonium chloride solution, dried over anhydrous sodium sul-
fate. The excess of solvent was removed under reduced pressure and
the obtained compound was crystallized from chloroform (spectro-
scopic data were in agreement with published data).
2‐{4‐[(2‐Methyl‐1H‐benzo[d]imidazol‐1‐yl)methyl]‐1H‐1,2,3‐triazol‐
1‐yl}‐N‐(2‐nitrophenyl)acetamide (4c)
Green color, yield: 58%, M.P.: 148–150°C, 1H NMR (400 MHz,
DMSO) δ = 10.73 (s, 1H, N–H), 8.16 (s, 1H, C–H triazole), 7.98
(d, J = 7.8 Hz, 1H), 7.82–7.64 (m, 3H), 7.58 (s, 1H), 7.41 (t, J = 7.0 Hz,
1H), 7.17 (d, J = 7.1 Hz, 2H), 5.55 (s, 2H), 5.37 (s, 2H), and 2.65 (s, 3H,
CH3). 13C NMR (100 MHz, DMSO) δ = 165.34 (C═O), 143.02 (C13
triazole), 142.88, 134.64, 130.74, 126.39, 126.02, 125.53 (C17 tria-
zole), 122.02, 121.78, 118.63, 110.63, 52.39, 38.64, and 14.23. IR
(KBr, ʋmax) = 3,286 (N–H str.), 3,145 (C–H str., triazole), 3,053 (C–H
str., aromatic), 1,691 (C═O str. amide), 1,608 (N–H bending), 1,553,
1,502, and 1,465 (C═C str., aromatic) cm−1. HRMS (ESI): m/z calcu-
lated for [M+H]+ C19H17N7O3 392.1471; found 392.1569.
|
4.1.6
Synthesis of substituted {4‐[(1H‐benzo‐
imidazol‐1‐yl)methyl]‐1H‐1,2,3‐triazol‐1‐yl}-
acetamides (4a–r)
In a round‐bottom flask, a mixture of 2‐substituted 1‐(prop‐2‐yn‐1‐
yl)‐1H‐benzo[d]imidazoles 2a,b (1.4 mmol), N‐substituted 2‐
bromoacetamides 3a–i (1.4 mmol), and sodium azide (2.8 mmol) was
taken in dimethylformamide/water (8:2), and then copper sulfate
pentahydrate (14 mol%) and sodium ascorbate (28 mol%) were ad-
ded, and the reaction mixture was stirred for 5–6 hr at 50°C.[36–38]
The reaction progress was monitored by TLC. After the completion of
reaction, workup was done by adding ammonia solution and product
was extracted with ethyl acetate (3 × 50 ml). The ethyl acetate layer
was washed with brine solution and dried using anhydrous sodium
sulfate. The solution was concentrated under reduced pressure and
residue was purified by recrystallization with CHCl3. The desired
products, 1,2,3‐triazole–benzimidazole hybrid, 4a–r, were isolated in
good yields.
N‐(2‐Acetylphenyl)‐2‐{4‐[(2‐methyl‐1H‐benzo[d]imidazol‐1‐yl)‐
methyl]‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4d)
White color, yield: 54%, M.P.: 124–126°C, 1H NMR (400 MHz,
DMSO) δ = 12.29 (s, 1H, N–H), 8.41 (s, 1H, C–H triazole), 7.89
(d, J = 8.0 Hz, 1H), 7.66 (t, J = 7.8 Hz, 2H), 7.54 (d, J = 7.5 Hz, 1H), 7.42
(d, J = 8.1 Hz, 1H), 7.32 (t, J = 7.6 Hz, 1H), 7.28–7.08 (m, 2H), 5.61
(s, 2H), 5.39 (s, 2H), 2.70 (s, 3H, CH3), and 2.15 (s, 3H, CH3). 13C NMR
(100 MHz, DMSO) δ = 202.77 (carbonyl carbon), 165.58, 143.32,
142.79 (C13 triazole), 138.32, 134.60, 132.02, 125.48 (C17 triazole),
124.26, 121.92, 121.71, 121.42, 118.53, 110.50, 53.26, 38.59, 29.03,
and 14.29. IR (KBr, ʋmax) = 3201 (N–H str.), 3,130 (C–H str., triazole),
3,076 (C–H str., aromatic), 1,720 (C═O str.), 1,641 (N–H bending),
1,608, 1,559, 1,531, and 1,465 (C–C str., aromatic) cm−1. HRMS (ESI):
m/z calculated for [M+H]+ C21H20N6O2 389.1726; found 389.1723.
2‐{4‐[(2‐Methyl‐1H‐benzo[d]imidazol‐1‐yl)methyl]‐1H‐1,2,3‐triazol‐
1‐yl}‐N‐phenylacetamide (4a)
Yellow color, yield: 60%, M.P.: 158–160°C, 1H NMR (400 MHz,
DMSO) δ = 10.43 (s, 1H, N–H), 8.18 (s, 1H, C–H triazole), 7.64
(d, J = 8.1 Hz, 1H), 7.54 (t, J = 7.7 Hz, 2H), 7.52 (d, J = 7.7 Hz, 1H), 7.32
(t, J = 7.9 Hz, 2H), 7.16 (dt, J = 16.3, 6.8 Hz, 2H), 7.08 (t, J = 7.4 Hz,
1H), 5.53 (s, 2H), 5.28 (s, 2H), and 2.67 (s, 3H, CH3). 13C NMR
(100 MHz, DMSO) δ = 164.57 (C═O), 152.19, 142.96 (C13 triazole),
142.73, 138.82, 135.39, 129.38, 125.58 (C17 triazole), 124.25,
121.98, 121.76, 119.66, 118.67, 110.52, 52.64, 38.57, and 14.22.
(KBr, ʋmax) = 3,305 (N–H str.), 3,140 (C–H str., triazole), 3,051 (C–H
str., aromatic), 1,697 (C═O str. amide), 1,616 (N–H bending), 1,558,
1,519, and 1,492 (C═C str., aromatic) cm−1. HRMS (ESI): m/z calcu-
lated for [M+H]+ C19H18N6O 347.1620; found 347.1617.
N‐(3‐Fluorophenyl)‐2‐{4‐[(2‐methyl‐1H‐benzo[d]imidazol‐1‐yl)‐
methyl]‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4e)
Brown color, yield: 74%, M.P.: 155–156°C, 1H NMR (400 MHz,
DMSO) δ = 10.67 (s, 1H, N–H), 8.19 (s, 1H, C–H triazole), 7.66
(d, J = 7.1 Hz, 1H), 7.53 (d, J = 9.6 Hz, 2H), 7.37 (dd, J = 15.0, 8.0 Hz,
1H), 7.28 (d, J = 8.2 Hz, 1H), 7.18 (dd, J = 16.4, 8.6 Hz, 2H), 6.92 (t,
J = 9.4 Hz, 1H), 5.54 (s, 2H), 5.31 (s, 2H), and 2.67 (s, 3H, CH3). 13C
NMR (100 MHz, DMSO) δ = 165.03 (C═O), 162.58 (d, J = 241 Hz),
142.97 (C13 triazole), 140.51 (d, J = 11 Hz), 131.80 (d, J = 9 Hz),
125.59 (C17 triazole), 122.00, 121.75, 118.68, 115.44, 111.24,
110.86, 110.75 (d, J = 21 Hz), 106.52 (d, J = 26 Hz), 52.64, 38.65, and
14.27. IR (KBr, ʋmax) = 3,292 (N–H str.), 3,140 (C–H str., triazole),
3,035 (C–H str., aromatic), 1,691 (C═O str.), 1,614 (N–H bending),
1,556, 1,514, and 1,470 (C═C str., aromatic) cm−1. HRMS (ESI): m/z
calculated for [M+H]+ C19H17N6OF 365.1526; found 365.1529.
N‐(2‐Chlorophenyl)‐2‐{4‐[(2‐methyl‐1H‐benzo[d]imidazol‐1‐yl)‐
methyl]‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4b)
Yellow color, yield: 63%, M.P.: 156°C, 1H NMR (400 MHz, DMSO)
δ = 10.58 (s, 1H, N–H), 8.18 (s, 1H, C–H triazole), 7.65 (d, J = 8.1 Hz,
1H), 7.58 (d, J = 8.8 Hz, 2H), 7.52 (d, J = 7.4 Hz, 1H), 7.38 (d, J = 8.8 Hz,

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DESWAL ET AL.
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2‐{4‐[(2‐Methyl‐1H‐benzo[d]imidazol‐1‐yl)methyl]‐1H‐1,2,3‐triazol‐
1‐yl}‐N‐(p‐tolyl)acetamide (4f)
128.67, 128.02, 126.68, 126.52, 126.23, 126.04, 125.62 (C17
triazole), 123.70, 123.00, 122.11, 122.03, 121.82, 110.53, 52.51,
38.61, and 14.19. IR (KBr, ʋmax) = 3,246 (N–H str.), 3,143 (C–H
str., triazole), 3,055 (C–H str., aromatic), 1,691 (C═O str.), 1,616
White color, yield: 74%, M.P.: 130–132°C, 1H NMR (400 MHz,
DMSO) δ = 10.34 (s, 1H, N–H), 8.17 (s, 1H, C–H triazole), 7.64 (d,
J = 8.1 Hz, 1H), 7.52 (d, J = 7.2 Hz, 1H), 7.44 (d, J = 8.4 Hz, 2H),
7.22–7.09 (m, 4H), 5.53 (s, 2H), 5.26 (s, 2H), 2.67 (s, 3H, CH3), and
2.25 (s, 3H, CH3). 13C NMR (100 MHz, DMSO) δ = 164.30 (C═O),
152.18, 142.94 (C13 triazole), 142.76, 136.41, 136.34, 135.40,
133.20, 129.74, 125.56 (C17 triazole), 121.96, 121.74, 119.65,
118.64, 110.51, 52.62, 38.57, 20.91, and 14.23. IR (KBr, ʋmax) = 3,255
(N–H str.), 3,136 (C–H str., triazole), 3,037 (C–H str., aromatic), 1,693
(C═O str.), 1,618 (N–H bending), 1,556, 1,512, and 1,454 (C═C str.,
aromatic) cm−1. HRMS (ESI): m/z calculated for [M+H]+ C20H20N6O
361.1777; found 361.1759.
(N–H bending), 1,556, 1,508, and 1,465 (C═C str., aromatic) cm−1
.
HRMS (ESI): m/z calculated for [M+H]+ C23H20N6O 397.1777;
found 397.1773.
2‐{4‐{[2‐(Methylthio)‐1H‐benzo[d]imidazol‐1‐yl]methyl}‐1H‐1,2,3‐
triazol‐1‐yl}‐N‐phenylacetamide (4j)
White color, yield: 74%, M.P.: 138–140°C, 1H NMR (400 MHz,
DMSO) δ = 10.41 (s, 1H, N–H), 8.13 (s, 1H, C–H triazole), 7.60
(d, J = 8.7 Hz, 1H), 7.56 (t, J = 8 Hz, 1H), 7.54 (d, J = 6.3 Hz, 2H),
7.32 (t, J = 7.9 Hz, 2H), 7.21–7.13 (m, 2H), 7.08 (t, J = 7.4 Hz, 1H),
5.46 (s, 2H), 5.28 (s, 2H), and 2.74 (s, 3H, CH3). 13C NMR
(100 MHz, DMSO) δ = 164.50 (C═O), 152.68, 143.50 (C13 tria-
zole), 142.28, 138.82, 136.57, 129.34, 125.57 (C17 triazole),
124.25, 122.06, 122.02, 119.74, 118.00, 110.21, 52.68, 39.21,
and 14.93. (KBr, ʋmax) = 3,267 (N–H str.), 3,142 (C–H str., tria-
zole), 3,051 (C–H str., aromatic), 1,691 (C═O str. amide), 1,662
N‐(4‐Methoxyphenyl)‐2‐{4‐[(2‐methyl‐1H‐benzo[d]imidazol‐1‐yl)‐
methyl]‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4g)
Yellow color, yield: 84%, M.P.: 124–126°C, 1H NMR (400 MHz,
DMSO) δ = 10.30 (s, 1H, NH), 8.18 (s, 1H, C–H triazole), 7.64
(d, J = 7.6 Hz, 1H), 7.52 (d, J = 7.6 Hz, 1H), 7.47 (d, J = 8.9 Hz, 2H),
7.23–7.10 (m, 2H), 6.90 (d, J = 9.0 Hz, 2H), 5.53 (s, 2H), 5.25 (s, 2H),
3.33 (s, 3H, CH3), and 2.67 (s, 3H, CH3). 13C NMR (100 MHz, DMSO)
δ = 164.03 (C═O), 155.99, 152.18, 142.94 (C13 triazole), 142.73,
135.40, 131.94, 125.56 (C17 triazole), 121.97, 121.76, 121.21,
118.64, 114.42, 110.53, 55.63, 52.57, 38.58, and 14.24. IR (KBr,
(N–H bending), 1,604, 1,558, and 1,442 (C═C str., aromatic) cm−1
.
HRMS (ESI): m/z calculated for [M+H]+ C19H18N6SO 379.1341;
found 379.1342.
N‐(2‐Chlorophenyl)‐2‐{4‐{[2‐(methylthio)‐1H‐benzo[d]imidazol‐1‐
yl]methyl}‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4k)
ʋmax) = 3,251 (N–H str.), 3,140 (C–H str., triazole), 3,034 (C–H str.,
aromatic), 1,691 (C═O str.), 1,610 (N–H bending), 1,560, 1,510, and
1,460 (C═C str., aromatic) cm−1. HRMS (ESI): m/z calculated for
[M+H]+ C20H20N6O2 377.1726; found 377.1467.
White color, yield: 67%, M.P.: 160–162°C, 1H NMR (400 MHz,
DMSO) δ = 10.57 (s, 1H, N–H), 8.13 (s, 1H, C–H triazole),
7.61–7.53 (m, 4H), 7.39–7.37 (m, 2H), 7.19–7.16 (m, 2H), 5.47
(s, 2H), 5.30 (s, 2H), and 2.74 (s, 3H, CH3). 13C NMR (100 MHz,
DMSO) δ = 164.76 (C═O), 152.69, 143.46 (C13 triazole), 142.30,
137.79, 136.55, 127.82, 125.63 (C17 triazole), 122.05, 121.24,
117.99, 110.29, 52.62, 39.15, and 14.90. IR (KBr, ʋmax) = 3338
(N–H str.), 3,134 (C–H str., triazole), 3,061 (C–H str., aromatic),
1,687 (C═O str.), 1,604 (N–H bending), 1,552, 1,492, and 1,446
(C═C str., aromatic) cm−1. HRMS (ESI): m/z calculated for [M+H]+
C19H17N6SClO 413.0951; found 413.0943.
N‐(4‐Fluorophenyl)‐2‐{4‐[(2‐methyl‐1H‐benzo[d]imidazol‐1‐yl)‐
methyl]‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4h)
Gray color, yield: 85%, M.P.: 180–182°C, 1H NMR (400 MHz,
DMSO) δ = 10.51 (s, 1H, N–H), 8.14 (s, 1H, C–H triazole),
7.72–7.38 (m, 4H), 7.29–7.07 (m, 4H), 5.47 (s, 2H), 5.29 (s, 2H),
and 2.74 (s, 3H, CH3). 13C NMR (100 MHz, DMSO) δ = 164.51
(C═O), 158.69 (d, J = 239 Hz), 152.68, 143.46 (C13 triazole),
142.29, 136.55, 135.23 (d, J = 3 Hz), 125.63 (C17 triazole), 122.03
(d, J = 2 Hz), 121.47 (d, J = 8 Hz), 117.99, 115.98 (d, J = 22 Hz),
110.30, 52.56, 39.15, and 14.89. (KBr, ʋmax) = 3,331 (N–H str.),
3,136 (C–H str., triazole), 3,070 (C–H str., aromatic), 1,690 (C═O
str. amide), 1,616 (N–H bending), 1,563, 1,508, and 1,477 (C═C
str., aromatic) cm−1. HRMS (ESI): m/z calculated for [M+H]+
C19H17N6OF 365.1526; found 365.1525.
2‐{4‐{[2‐(Methylthio)‐1H‐benzo[d]imidazol‐1‐yl]methyl}‐1H‐1,2,3‐
triazol‐1‐yl}‐N‐(2‐nitrophenyl)acetamide (4l)
Yellow color, yield: 65%, M.P.: 210–212°C, 1H NMR (400 MHz,
DMSO) δ = 10.70 (s, 1H, N–H), 8.11 (s, 1H, C–H triazole), 7.98
(dd, J = 8.2, 1.3 Hz, 1H), 7.72 (dd, J = 7.1, 1.4 Hz, 1H), 7.69
(d, J = 1.5 Hz, 1H), 7.59 (dd, J = 6.2, 2.7 Hz, 1H), 7.57–7.51 (m, 1H),
7.45–7.37 (m, 1H), 7.22–7.12 (m, 2H), 5.46 (s, 2H), 5.37 (s, 2H), and
2.73 (s, 3H, CH3). 13C NMR (100 MHz, DMSO) δ = 165.33 (C═O),
152.68, 143.46 (C13 triazole), 142.85, 142.37, 136.55, 134.63,
130.79, 126.36, 126.02, 125.61, 125.53, 122.04 (C17 triazole),
117.98, 110.29, 52.39, 39.13, and 14.89. IR (KBr, ʋmax) = 3,329 (N–H
str.), 3,142 (C–H str., triazole), 3,068 (C–H str., aromatic), 1,720
(C═O str.), 1,602 (N–H bending), 1,548, 1,492, and 1,442 (C═C str.,
aromatic) cm−1. HRMS (ESI): m/z calculated for [M+H]+ C19H17N7O3S
424.1192; found 424.1182.
2‐{4‐[(2‐Methyl‐1H‐benzo[d]imidazol‐1‐yl)methyl]‐1H‐1,2,3‐
triazol‐1‐yl}‐N‐(naphthalen‐1‐yl)acetamide (4i)
White color, yield: 66%, M.P.: 145–147°C, 1H NMR (400 MHz,
DMSO) δ = 10.43 (s, 1H, N–H), 8.23 (s, 1H, C–H triazole), 8.11
(d, J = 9.0 Hz, 1H), 7.95 (d, J = 9.3 Hz, 1H), 7.79 (d, J = 8.2 Hz, 1H),
7.65 (t, J = 8.6 Hz, 2H), 7.58–7.47 (m, 4H), 7.22–7.11 (m, 2H), 5.54
(s, 2H), 5.49 (s, 2H), and 2.67 (s, 3H, CH3). 13C NMR (100 MHz,
DMSO) δ = 165.53 (C–O), 142.97 (C13 triazole), 134.15, 133.08,

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N‐(2‐Acetylphenyl)‐2‐{4‐{[2‐(methylthio)‐1H‐benzo[d]imidazol‐1‐yl]
methyl}‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4m)
(C–H str., aromatic), 1,687 (C═O str.), 1,614 (N–H bending), 1,556, 1,514,
and 1,460 (C═C str., aromatic) cm−1. HRMS (ESI): m/z calculated for [M
+H]+ C20H20N6SO2 409.1447; found 409.1404.
Gray color, yield: 53%, M.P.: 140–142°C, 1H NMR (400 MHz, DMSO)
δ = 11.33 (s, 1H, N–H), 8.22 (d, J = 8.3 Hz, 1H), 8.16 (s, 1H, C–H
triazole), 7.99 (d, J = 6.7 Hz, 1H), 7.65–7.54 (m, 3H), 7.28–7.17
(m, 3H), 5.48 (s, 2H), 5.41 (s, 2H), 2.74 (s, 3H, CH3), and 2.57 (s, 3H,
CH3). 13C NMR (100 MHz, DMSO) δ = 202.73 (C═O), 165.53 (C═O,
amide), 152.70, 143.46 (C13 triazole), 142.57, 138.27, 136.56,
134.56, 131.97, 125.60 (C17 triazole), 125.47, 124.28, 122.04,
122.02, 121.49, 117.97, 110.31, 53.23, 39.18, 29.06, and 14.91. IR
(KBr, ʋmax) = 3,318 (N–H str.), 3,136 (C–H str., triazole), 3,066 (C–H
str., aromatic), 1,762 (C═O str. ketone), 1,687 (C═O str. amide),
1,581 (N–H bending), 1,521, 1,487, and 1,419 (C═C str., aromatic)
N‐(4‐Fluorophenyl)‐2‐{4‐{[2‐(methylthio)‐1H‐benzo[d]imidazol‐1‐
yl]methyl}‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4q)
Gray color, yield: 84%, M.P.: 170–172°C, 1H NMR (400 MHz, DMSO)
δ = 10.52 (s, 1H, N–H), 8.18 (s, 1H, C–H triazole), 7.64 (d, J = 7.5 Hz,
1H), 7.57 (dd, J = 9.1, 5.0 Hz, 2H), 7.52 (d, J = 7.5 Hz, 1H), 7.16 (ddt,
J = 15.1, 9.8, 4.9 Hz, 4H), 5.53 (s, 2H), 5.28 (s, 2H), and 2.67 (s, 3H,
CH3). 13C NMR (100 MHz, DMSO) δ = 164.53 (C═O), 158.69
(d, J = 239 Hz), 142.98 (C13 triazole), 135.23 (d, J = 3 Hz), 125.57 (C17
triazole), 121.84 (d, J = 21 Hz), 121.47 (d, J = 7 Hz), 118.65, 115.98 (d,
J = 22 Hz), 110.51, 52.57, 38.57, and 14.23. IR (KBr, ʋmax) = 3,261
(N–H Str.), 3,145 (C–H str., triazole), 3,037 (C–H str., aromatic), 1,691
(C═O str. amide), 1,622 (N–H bending), 1,571, 1,508, and 1,462 (C═C
cm−1
.
HRMS (ESI): m/z calculated for [M+H]+ C21H20N6SO2
421.1447; found 421.1445.
N‐(3‐Fluorophenyl)‐2‐{4‐{[2‐(methylthio)‐1H‐benzo[d]imidazol‐1‐
yl]methyl}‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4n)
str., aromatic) cm−1 HRMS (ESI): m/z calculated for [M+H]+
.
C19H17N6SOF 397.1247; found 397.1249.
White color, yield: 64%, M.P.: 160–162°C, 1H NMR (400 MHz,
DMSO) δ = 10.68 (s, 1H, N–H), 8.15 (s, 1H, C–H triazole), 7.61 (s, 1H),
7.55 (s, 2H), 7.38 (d, J = 14 Hz, 1H), 7.28 (d, J = 14 Hz, 1H), 7.18 (s,
2‐{4‐{[2‐(Methylthio)‐1H‐benzo[d]imidazol‐1‐yl]methyl}‐1H‐1,2,3‐
triazol‐1‐yl}‐N‐(naphthalen‐1‐yl)acetamide (4r)
2H), 6.93 (s, 1H), 5.47 (s, 2H), 5.32 (s, 2H), and 2.75 (s, 3H, CH3). 13
C
White color, yield: 83%, M.P.: 151–153°C, 1H NMR (400 MHz,
DMSO) δ = 10.39 (s, 1H, N–H), 8.18 (s, 1H, C–H triazole), 8.13
(d, J = 8 Hz, 1H), 7.97–7.95 (m, 1H), 7.80 (d, J = 8.2 Hz, 1H), 7.69
(d, J = 8 Hz, 1H), 7.63–7.53 (m, 4H), 7.49 (t, J = 12 Hz, 1H), 7.19–7.16
(m, 2H), 5.49 (s, 2H), 5.48 (s, 2H), and 2.74 (s, 3H, CH3). 13C NMR
(100 MHz, DMSO) δ = 165.50 (C═O), 152.69, 143.47 (C13 triazole),
142.31, 136.57, 134.16, 133.15, 128.66, 128.00, 126.65, 126.48,
126.17, 126.03, 125.66 (C17 triazole), 123.04, 122.02, 117.99,
110.31, 52.54, 39.18, and 14.90. IR (KBr, ʋmax) = 3,340 (N–H str.),
3,136 (C–H str., triazole), 3,057 (C–H str., aromatic), 1,732 (C═O str.
amide), 1,666 (N–H bending), 1,558, 1,502, and 1,442 (C═C str.,
aromatic) cm−1. HRMS (ESI): m/z calculated for [M+H]+ C23H20N6SO
429.1498; found 429.1496.
NMR (100 MHz, DMSO) δ = 165.02 (C═O), 162.58 (d, J = 243 Hz),
152.69, 143.46 (C13 triazole), 142.32, 140.52 (d, J = 11 Hz), 136.55,
131.09 (d, J = 9 Hz), 125.66, 122.03 (C17 triazole), 117.99, 115.42,
110.74 (d, J = 21 Hz), 110.30, 106.49 (d, J = 26 Hz), 52.62, 39.13, and
14.89. IR (KBr, ʋmax) = 3,251 (N–H str.), 3,140 (C–H str., triazole),
3,034 (C–H str., aromatic), 1,691 (C═O str.), 1,610 (N–H bending),
1,560, 1,510, and 1,460 (C═C str., aromatic) cm−1. HRMS (ESI): m/z
calculated for [M+H]+ C19H17N6SOF 397.1247; found 397.1249.
2‐{4‐{[2‐(Methylthio)‐1H‐benzo[d]imidazol‐1‐yl]methyl}‐1H‐1,2,3‐
triazol‐1‐yl}‐N‐(p‐tolyl)acetamide (4o)
White color, yield: 64%, M.P.: 124–126°C, 1H NMR (400 MHz,
DMSO) = 10.34 (s, 1H, N–H), 8.13 (s, 1H, C–H triazole), 7.61–7.42
(m, 4H), 7.20–711 (m, 4H), 5.46 (s, 2H), 5.26 (s, 2H), 2.74 (s, 3H, CH3),
and 2.25 (s, 3H, CH3). 13C NMR (100 MHz, DMSO) = 164.27 (C═O),
152.68, 143.47 (C13 triazole), 142.25, 136.58, 133.21, 129.73, 125.61
(C17 triazole), 122.05, 119.66, 117.99, 110.33, 52.62, 39.16, 20.90,
and 14.90. IR (KBr, ʋmax) = 3,271 (N–H Str.), 3,136 (C–H str., triazole),
3,059 (C–H str., aromatic), 1,691 (C═O str.), 1,612 (N–H bending),
1,554, 1,514, and 1,452 (C═C str., aromatic) cm−1. HRMS (ESI): m/z
calculated for [M+H]+ C20H20N6OS 393.1498; found 393.1484.
|
4.2
Pharmacological assays
|
4.2.1
α‐Amylase inhibition assay
The activity was measured according to the published protocol
with slight modifications.[39] The growing material was prepared
by dissolving soluble starch (1 g) in 50 ml of 0.4 M NaOH and then
heating it at 100°C for 5 min. The solution was cooled on crushed
ice, and the pH of the solution was settled at 7 with 2 M HCl.
Distilled water was added to make volume to 200 ml. A sample
solution was prepared in DMSO with different concentrations,
that is, 100, 50, 25, and 12.5 µg/ml. The growing material
(20 μl) and sample solutions (10 μl) were added in a microplate
well and the mixtures were incubated for 3 min at 37°C. Then,
10 μl of α‐amylase solution (50 μg/ml) was added and the mixture
was further incubated for 15 min at 37°C. The reaction was
stopped by adding 40 μl of 0.1 M HCl and then 100 μl of 1 mM
N‐(4‐Methoxyphenyl)‐2‐{4‐{[2‐(methylthio)‐1H‐benzo[d]imidazol‐1‐
yl]methyl}‐1H‐1,2,3‐triazol‐1‐yl}acetamide (4p)
Gray color, yield: 63%, M.P.: 137–139°C, 1H NMR (400 MHz, DMSO)
δ = 10.29 (s, 1H, N–H), 8.13 (s, 1H, C–H triazole), 7.61–7.45 (m, 4H),
7.1–7.15 (m, 4H), 5.46 (s, 2H), 5.25 (s, 2H), 3.72 (s, 3H, CH3), and 2.74
(s, 3H, CH3). 13C NMR (100 MHz, DMSO) δ = 164.00 (C═O), 155.99,
152.69, 143.46 (C13 triazole), 142.25, 136.55, 131.93, 125.61 (C17 tria-
zole), 122.05, 121.21, 117.98, 114.46, 110.31, 55.63, 52.56, 39.15, 14.89.
IR (KBr, ʋmax) = 3,290 (N–H str.), 3,147 (C–H str., triazole), 3,003

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iodine solution. The absorbances were recorded at 650 nm.
Acarbose was used as a standard drug. α‐Amylase activity was
expressed in % inhibition.
Compounds 4e, 4g, and 4n were docked in this active site using
AutoDock Vina.[47] The dimensions of the search box were as follows:
center_x = −12.5327806063, center_y = −5.97621363233, center_z =
−19.8977436728, size_x = 26.1196805853, size_y = 25.0, and size_z =
25.0. The results were analyzed using a Discovery Studio visualizer[48]
and PyMOL.[49]
% Inhibition = [1 − (Ab1–Ab2)/(Ab3–Ab4) × 100],
where Ab1 was the absorbance of solution holding starch and tested
compounds, Ab2 was the absorbance of solution holding amylase,
starch and tested compound, Ab3 was the absorbance of solution
holding starch, and Ab4 was the absorbance of solution holding
amylase and starch.
ACKNOWLEDGMENTS
Authors are highly thankful to DST‐PURSE ([SR/PURSE Phase
2/40[G]), New Delhi, and Dr. APJ Abdul Kalam CIL, GJUS&T, Hisar.
Mrs. Laxmi Deswal (SRF) acknowledges the University Grant Commis-
sion, New Delhi, India, for the award of JRF–SRF Fellowship with Sr.
No. 2121410071, Ref. No. 21/12/2014(ii) EU‐V, for financial assistance.
|
4.2.2
α‐Glucosidase inhibition assay
A modified version of the assay described by Xaio et al.[3] was followed. A
sample solution with various concentrations, that is, 100, 50, 25, and
12.5 µg/ml, was prepared in DMSO. A volume of 10 µl of the sample
solution was diluted with 120 µl of phosphate buffer (100 mM) of pH 6.9.
Then, 20 µl of 0.5 U/ml α‐glucosidase enzymes was added into each well
and the plates were incubated at 37°C for 15 min. Thereafter, 20 µl of
pNPG (4‐nitrophenyl‐α‐D‐glucopyranoside) in potassium phosphate
buffer (5 mM) was added to it, and the reaction mixture was further
incubated for 15 min at 37°C. The absorbance was measured at 405 nm.
For positive control, acarbose was used and DMSO (10 µl) was used at
the place of sample solution for negative control. The equation for
calculating percentage inhibition is as follows:
CONFLICT OF INTERESTS
The authors declare that there are no conflicts of interests.
ORCID
Vikas Verma
http://orcid.org/0000-0002-6648-1877
Chander P. Kaushik
Ashwani Kumar
http://orcid.org/0000-0001-7917-1280
http://orcid.org/0000-0002-8966-3569
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https://doi.org/10.1002/ardp.202000090