Synthesis and molecular docking study of some 5,6-dichloro-2-cyclopropyl-1H-benzimidazole derivatives bearing triazole, oxadiazole, and imine functionalities as potent inhibitors of urease

Article abstract of DOI:10.1016/j.bmcl.2017.05.019

A new series of benzimidazole compounds including hydrazinecarbothioamide, 1,2,4-triazole, 1,3,4-oxadiazole and imine function were synthesized starting from 5,6-dichloro-2-cyclopropyl-1H-benzimidazole. All of the benzimidazole derivatives exhibited good urease inhibitor activity. Compound 6a proved to be the most potent showing an enzyme inhibitory activity with an IC₅₀?=?0.06?μM. Molecular docking studies were also conducted on enzyme extracted from Jack bean urease to identify the binding mode of the newly synthesized compounds.

Full text of DOI:10.1016/j.bmcl.2017.05.019

Accepted Manuscript
Synthesis and molecular docking study of some 5,6-dichloro-2-cycloprop-
yl-1H-benzimidazole derivatives bearing triazole, oxadiazole, and imine func-
tionalities as potent inhibitors of urease
Emre Menteşe, Hakan Bektaş, Bahar Bilgin Sokmen, Mustafa Emirik, Demet
Çakır, Bahittin Kahveci
PII:
S0960-894X(17)30499-7
DOI:
http://dx.doi.org/10.1016/j.bmcl.2017.05.019
BMCL 24964
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
10 March 2017
4 May 2017
5 May 2017
Please cite this article as: Menteşe, E., Bektaş, H., Sokmen, B.B., Emirik, M., Çakır, D., Kahveci, B., Synthesis and
molecular docking study of some 5,6-dichloro-2-cyclopropyl-1H-benzimidazole derivatives bearing triazole,
oxadiazole, and imine functionalities as potent inhibitors of urease, Bioorganic & Medicinal Chemistry Letters
(2017), doi: http://dx.doi.org/10.1016/j.bmcl.2017.05.019
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Synthesis and molecular docking study of some 5,6-dichloro-2-cyclopropyl-
1H-benzimidazole derivatives bearing triazole, oxadiazole, and imine
functionalities as potent inhibitors of urease
Emre Menteşea,*, Hakan Bektaşb, Bahar Bilgin Sokmenb, Mustafa Emirika, Demet Çakırb, Bahittin
Kahvecic
aDepartment of Chemistry,Art and Science Faculty, Recep Tayyip Erdogan University, Rize, Turkey
bDepartment of Chemistry, Faculty of Arts and Sciences, Giresun University, 28049, Giresun, Turkey;
cDepartment of Nutrition and Dietetics, Faculty of Health Sciences, Karadeniz Technical University, Trabzon,
Turkey
E-mail: emre.mentese@erdogan.edu.tr
Fax: +904642234019
ABSTRACT
A new series of benzimidazole compounds including hydrazinecarbothioamide, 1,2,4-triazole, 1,3,4-
oxadiazole and imine function were synthesized starting from 5,6-dichloro-2-cyclopropyl-1H-
benzimidazole. All of the benzimidazole derivatives exhibited good urease inhibitor activity.
Compound 6a proved to be the most potent showing an enzyme inhibitory activity with an IC50 = 0.06
µM. Molecular docking studies were also conducted on enzyme extracted from Jack bean urease to
identify the binding mode of the newly synthesized compounds.
Keywords: Benzimidazole, 1,2,4-Triazole, Schiff base, Docking study, Antiurease activity.

The metalloenzyme urease (urea amidohydrolase EC 3.5.1.5) is a nickel-containing enzyme
that catalyzes the hydrolysis of urea to ammonia and carbon dioxide. Urease, widely distributed in
nature, and it is found in a variety of plants, fungi, algae and bacteria.1,2 Urease is involved in the
pathogenesis of hepatic encephalopathy, hepatic coma urolithiasis, pyelonephritis, ammonia and
urinary catheter encrustation.3 It is also a major cause of pathologies induced by Helicobacter pylori
(H. pylori) as this allows bacteria to survive at the low pH of the stomach and hence plays an
important role in producing peptic and gastric ulcers.4 In the near past, a number of compounds have
been proposed as urease inhibitors to reduce environmental problems and to enhance the uptake of
urea nitrogen by plants and health problems.5 Many urease inhibitors have been investigated in the
past decades, such as imidazoles, phosphorodiamidates and hydroxamic acid derivatives, but most of
these compounds are very unstable or toxic to allow their use in vivo. Therefore, the search is still on
for novel urease inhibitors with promising levels of activity.6
It is known that benzimidazoles are among the most important heterocyclic systems that
show many different pharmaceutical properties. Recently, some of benzimidazole derivatives have
been used as drugs in human and veterinary medicine.7 In addition, many biological activities related
to benzimidazole derivatives have been described: anthelmintic,8 antiurease,9 antimicrobial,10
cytotoxic properties,11 antitumor activity,12,13 antibacterial activity,14 anti-MRSA and anti-VRE
agents,15 anticancer agents,16 anti-viralagents17 and anti tubercular activity.18 Moreover, previous
studies show that 5,6-dichloro benzimidazole derivatives have some biological activities.19-23
Several literature reports of various heterocyclic compounds containing cyclopropyl
substituent that are very important for their pharmacological effects. Introduction of cyclopropyl
group can potentially increase inhibitory activity.24-27 Also, literature survey reveals that Schiff bases
are important in diverse field's chemistry has been because of their biological properties.28
Furthermore, substituted acylhydrazide Schiff bases have shown a broad range of bioactivities like
antibacterial and antiurease.29,30
Therefore, preparations of new benzimidazole derivatives containing different functional
groups have received an increasing attention from synthetic chemists. In this study, some novel
benzimidazole derivatives have been synthesized and their antiurease activities have been
investigated with molecular docking study.
In the current study, we have synthesized some benzimidazole derivatives starting from iminoester
hydrochloride and have been investigated for their antiurease activity. Starting material, iminoester

hydrochloride (1) was prepared according to the known method.31 The reaction of compound 1 with
4,5-dichloro-o-phenylenediamine afforded 5,6-dichloro-2-cyclopropyl-1H-benzo[d]imidazole (2).32,33
The reaction of compound 2 with ethyl bromoacetate in the presence of dry potassium carbonate
yielded ethyl 2-(5,6-dichloro-2-cyclopropyl-1H-benzo[d]imidazol-1-yl)acetate (3). The compound 4
was prepared by reacting 3 with hydrazine monohydrate in ethanolic solution. The ring closure
reaction of 4 with carbon disulfide in presence of KOH resulted in the formation of compound 5.
Thiosemicarbazides were prepared by the nucleophilic addition of acid hydrazides (5a-b) to methyl
isothiocyanate (for 6a) or ethyl isothiocyanate (for 6b). Intramolecular cyclization of
thiosemicarbazides (6a-b) in the presence of 2N NaOH solution under reflux resulted in the
formation of 1,2,4-triazole derivatives 7a-b. We have also converted compound 4 to some schiff
bases (8a-g) by the reaction of acetohydrazide derivative (4) with corresponding aldehydes. The
synthetic path of the target compounds was illustrated in Scheme 1. The structures of the new
1
compounds were confirmed by H-NMR, 13C-NMR spectroscopy and mass spectra. For the
spectroscopic investigations of newly synthesized compounds are accordance with the proposed
structure.
Scheme 2. The synthetic path of the target compounds.

The inhibition effect of urease activity was shown in Table 1. All of the benzimidazole derivatives
exhibited good urease inhibitor activity. Lower IC50 values indicate higher enzyme inhibitor activity.
Compound 6a which has methyl group on thiosemicarbazide part proved to be the most potent
showing compound an enzyme inhibitory activity with an IC50 = 0.06 µM. Thiourea showed lower
activity than compounds 2, 6a-b, 7a-b, 8b, 8d, 8e, 8f bearing thiosemicarbazide, triazole and imine
function at positionon-1 on the benzimidazole nucleus. It may be concluded from the results than
benzimidazoles bearing open-chain thiosemicarbazide (6a-b), in general, are more potent that their
mercapto-1,2,4-triazole (7a-b) counterparts.
Table 1. The antiurease activities and docking score of benzimidazole derivatives.
Antiurease IC50
Compounds
Docking Score
(μM)*
2
0.17±0.011
1.12±0.08
1.92±0.33
1.34±0.28
0.05±0.002
0.28±0.006
0.17±0.072
0.38±0.083
2.07±0.31
0.29±0.030
1.49±0.29
0.06±0.008
0.22±0.089
0.14±0.034
0.15±0.019
0.56±0.17
-6.424
-5.741
-6.353
-7.34
3
4
5
6a
-8.968
-8.34
6b
7a
-4.775
-7.701
-5.591
-6.339
-8.916
-7.133
-7.044
-7.815
-5.843
-6.11
7b
8a
8b
8c
8d
8e
8f
8g
Thiourea
*Values were the means of three replicates ± Standard deviation (SD).

Compound 8a having phenyl ring with no substituent had the lowest antiurease activity among all
the compounds tested and standards (IC50 = 2.07 µM). Compounds 8b, 8d-g containing halogen
atoms and hydroxyl group on phenyl ring showed excellent inhibitor activity. Among the Schiff base
derivatives, compound 8d bearing a 3-flouro, and 4-bromo groups on phenyl ring was found to be
the most active inhibitor when compared with the other Schiff bases. Compound 8e contain hydroxyl
group 2- position when compared with compounds 8f and 8g having hydroxyl group 2-position with
bromo (8f) and chloro (8g) substituents respectively at phenyl part. The inhibitory potential of
compounds 8f and 8g was found greater than 8e. This indicates that halogen atoms on the phenyl
ring enhance the activity. It is clear that electronic effect plays an important role in the inhibitor
activity of compounds 8a-g. We have demonstrated that the potency of the benzimidazole
derivatives into inhibit urease.
In order to give an explanation and understanding of potent inhibitory activity, docking poses of the
most potent compounds 6a and 8d at the binding site of Jack bean urease were illustrated. The
docking scores of the studied compounds with 3LA4 were given in Table 1. The docking scores have a
good agreement with the experimental results, in general. The newly synthesized benzimidazole
derivatives have the capability to bind to active sites of urease enzyme. Thus, the hydrolysis of
enzyme was stopped and activity of enzyme was inhibited.
According to the in vitro results, compound 8d was the most active inhibitor of Jack bean urease. The
binding model of compound 8d was shown in Fig. 1. The docking studies showed that the best
orientation of compound 6a in the active pocket was formed by hydrogen bonding with the amino
acid residues of Arg439, Asp494. Furthermore, phosphate, PO4, group of the enzyme has an
important contribution to binding energy with forming hydrogen bond and the salt bridge through
the NH moiety of ligand. The amino acids Val591, Ile411, Ala436, Ala440, Met588 and Leu589 form
hydrophobic interaction with compound 8d. Fig. 2 represents the most interacting conformation of
8d in the active side of the enzyme at the electrostatic surface.

Fig.1. 2D interaction diagram of compound 8d in the active pocket.
Fig. 2. The predicted binding mode of compound 8d at electrostatic surface

Fig. 3 shows the docking pose of compound 6a. The prominent interactions of compound 6a with
enzyme are three hydrogen bond interactions and salt bridge by means of PO4 anion of enzyme. The
docking study predicted that compound 6a binds to enzyme at the binding site forming a hydrogen
bond with Arg439 and Asp494 residue through the -NH and -OH groups of benzimidazole and other
interactions involve the salt bridge with Asp494 and PO4 groups of the enzyme. Van der Waals
interactions were established with Ala636, Ala440, Ile411, Met637 and Leu523 amino acid residues in
hydrophobic pocket. Figure 4 represents the most interacting conformation of 6a in the active side of
the enzyme at the electrostatic surface.
Fig. 3. 2D interaction diagram of compound 6a in the active pocket.

Fig. 4. The predicted binding mode of compound 6a at electrostatic surface
In conclusion, a new series of benzimidazole compounds including hydrazinecarbothioamide, 1,2,4-
triazole, 1,3,4-oxadiazole and imine function were synthesized and evaluated for their antiurease
activities. All of the benzimidazole derivatives exhibited good urease inhibitor activity. Compound 6a
which has methyl group on thiosemicarbazide part proved to be the most potent showing compound
an enzyme inhibitory activity with an IC50 = 0.06 µM. Among the Schiff base derivatives, containing
halogen atoms on phenyl ring was found to be the most active inhibitor compared with other Schiff
bases. The molecular docking results show that the H-bonding is more important than other factors
3-
in urease inhibition mechanism. PO4 anion has a significant role as hydrogen bond acceptor in the
active region of urease enzyme. The results could be inspiration for further structural modifications
and investigations of potential antiurease activity within heterocyclic compounds.
Conflict of interest
The authors confirm that this article has no conflict of interest.
A. Supplementary data

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