Synthesis and Pharmacological Activity of C(2)-Substituted Benzimidazoles

Article abstract of DOI:10.1007/s11094-019-01979-0

Series of new compounds containing 2-pyrrolidinomethyl and 2-piperazinomethyl groups and 2-morpholino and 2-morpholinoethylamine substituents were synthesized. The antiglycation activities of 2-(morpholin-4-yl)-1-(propen-2-en-1-yl)-1H-benzimidazole and 2-(morpholin-4-yl)-1-[2-(morpholin-4-yl)ethyl]-1H-benzimidazole were comparable to that of aminoguanidine. Some compounds exhibited moderate antiserotonin, anti-angiotensin, antioxidant, and dipeptidyl peptidase-4 (DPP-4) inhibitory effects.

Full text of DOI:10.1007/s11094-019-01979-0

DOI 10.1007/s11094-019-01979-0
Pharmaceutical Chemistry Journal, Vol. 53, No. 3, June, 2019 (Russian Original Vol. 53, No. 3, March, 2019)
SYNTHESIS AND PHARMACOLOGICAL ACTIVITY
OF C(2)-SUBSTITUTED BENZIMIDAZOLES
O. N. Zhukovskaya,1,* A. A. Spasov,2,3 D. S. Yakovlev,2,3 V. A. Kosolapov,2 D. V. Mal’tsev,2,3
A. S. Morkovnik,1 V. A. Babkova,2 A. A. Brigadirova,2,3 Ya. V. Agatsarskaya,2,3 A. S. Taran,2
M. V. Miroshnikov,2 K. T. Sultanova,2 V. I. Kornilov,2 and V. A. Anisimova1
Translated from Khimiko-Farmatsevticheskii Zhurnal, Vol. 53, No. 3, pp. 10 – 15, March, 2019.
Original article submitted March 23, 2018.
Series of new compounds containing 2-pyrrolidinomethyl and 2-piperazinomethyl groups and 2-morpholino
and 2-morpholinoethylamine substituents were synthesized. The antiglycation activities of 2-(morpholin-4-
yl)-1-(propen-2-en-1-yl)-1H-benzimidazole and 2-(morpholin-4-yl)-1-[2-(morpholin-4-yl)ethyl]-1H-benz-
imidazole were comparable to that of aminoguanidine. Some compounds exhibited moderate antiserotonin,
anti-angiotensin, antioxidant, and dipeptidyl peptidase-4 (DPP-4) inhibitory effects.
Keywords: benzimidazole, synthesis, serotonin receptor, antiglycation and antioxidant activity.
2-Dialkylaminobenzimidazoles were previously scree-
ned for antagonist activity toward serotonin (5-HT) receptors
[1]. In continuation of this research, compounds with substi-
tuted piperazines and pyrrolidine as the amine bonded to
benzimidazole either through a methylene linker or directly
to it were prepared. Considering that these amines are
pharmacophores in compounds exhibiting antihistamine,
antiplatelet, and several other types of activity [2 – 10], all
products were tested in various in vitro pharmacological
models including 5-HT - (serotonin receptor type 3),
3
5-HT - (serotonin receptor type 2A), and AT -antagonistic
2A
1
(angiotensin receptor type 1); antiglycation; DPP-4 inhibi-
tory; and antioxidant activity.
1
Institute of Physical and Organic Chemistry, Southern Federal University,
Compounds I and II were prepared by nucleophilic sub-
stitution of the sulfonic acid in benzimidazole-2-sulfonic
acid (VI and VII) by morpholine [1] followed by alkylation
of 1-(2-benzimidazole)morpholine (V) by allyl bromide and
4-(2-chloroethyl)morpholine hydrochloride using a modified
Rostov-on-Don, 344090 Russia.
2
Volgograd State Medical University, Ministry of Health of the RF,
Volgograd, 400131 Russia.
3
Volgograd Medical Scientific Center, Volgograd, 400131 Russia.
*
e-mail: zhukowskaia.ol@yandex.ru
N
N
RHal
NR1
2
NR1R2
R
N
N
R
H
N
N
I, II
V
SO3H
N
NHR1R2
N
N
H
NR1R2
VII
SO3H
N
III, IV
VI
NR1R2 = N(CH2CH2)O (I, II, V), (CH2)4N (III), NH(CH2)2N(CH2CH2)2O (IV).
R = All, (CH2)2N(CH2CH2)2O.
201
0091-150X/19/5303-0201 © 2019 Springer Science+Business Media, LLC

202
O. N. Zhukovskaya et al.
R4
R4
R4
N
NHR1R2
C6H6
N
N
N
N
SO2Cl
CHCl3
CH2NR1R2
CH2Cl
CH2OH
N
R3
R3
R3
NR1R2=N(CH2)4 (VIII, IX), N(CH2CH2)2N(CH2)2OH (X, XI), N(CH2CH2)2NC6H3(CH3)2-2,5 (XII), N(CH2CH2)2NCH(C6H4F-4)2 (XIII).
R3 = R4 = H (VIII), CH3 (IX, X, XIV), R3 = CH3, R4 = OCH3 (XI, XII, XIII, XV).
literature method [11]. The reaction of sulfonic acid VII and
morpholine occurred smoothly to afford V although the reac-
tions with pyrrolidine and 4-(2-aminoethyl)morpholine
formed resins from which final products could not be puri-
fied by recrystallization or column chromatography because
of low chromatographic mobility. Resinous products could
be avoided by changing the order of the synthesis, i.e., VII
was first alkylated by benzyl chloride [12] and then substi-
tuted by the amine.
enough that the temperature of the reaction mixture was (°C.
The starting compound dissolved. The sodium salt of
2-(morpholin-4-yl)benzimidazole precipitated. The mixture
was stirred at room temperature for 15 – 20 min, cooled to
–2°C, treated with allyl bromide (1.03 mL, 12 mmol), and
stirred for 40 min at 0 – 5°C and for 1.5 h at 50°C. The or-
ganic layer was separated. The aqueous layer was treated
with Me CO (10 mL). The combined Me CO solution was
2
2
evaporated. The solid was treated with cold H O
2
Compounds VIII-XIII were prepared by nucleophilic
substitution of Cl in 2-(chloromethyl)benzimidazole XIV by
various substituted piperazines and pyrrolidine. Intermedi-
ates were synthesized via condensation of o-phenylenedia-
mine with glycolic acid in the presence of HCl followed by
treatment of the 2-(hydroxymethyl)benzimidazoles with
thionyl chloride under acidic conditions [13].
(20 – 30 mL). The precipitate was filtered off after 30 min,
rinsed on the filter with several portions of cold (5 – 8°C)
H O, and dried. Yield of I, 1.94 g (80%). I was crystallized
2
from CCl , dried in air, dissolved in i-PrOH and EtOH, and
4
acidified with HCl in PrOH to pH 1. mp of hydrochloride
221 – 223°C. C H N O × HCl. PMR spectrum (600 MHz,
14 17
3
DMSO-d ), d, ppm: 3.56 – 3.58 (t,
J
4.41 Hz, 4H,
6
PMR spectra of I-IV were characterized by sets of com-
mon resonances. Thus, protons of morpholine bonded di-
rectly to benzimidazole appeared as a triplet (3.56 –
3.89 ppm). Protons in II-IV with morpholine in a 2-morpho-
lin-4-ylethyl chain also appeared as a triplet (2.49 –
3.34 ppm). Pyrrolidine protons in III and IX appeared as
broad singlets (1.95 and 3.37 ppm). The piperazine protons
in X and XI appeared as broad singlets (2.35 – 3.14 ppm).
Thus, the goal of the present work was to synthesize and
study the pharmacology of benzimidazoles containing pyrro-
lidine, morpholine, or piperazine groups in the 2-position.
CH -N-CH ), 3.78 – 3.80 (t, J 4.59 Hz, 4H, CH -O-CH ),
2
2
2
2
4.83 – 4.84 (t, J 1.98 Hz, 2H, N -CH ), 5.22 – 5.33 (m, 2H,
Het.
2
CH ), 6.06 – 6.12 (m, 1H, CH), 7.33 – 7.38 (m, 2H,
2
5.6-H ), 7.47 (d, J 7.44 Hz, 7-H ), 7.74 (d, J 7.74 Hz,
Bzm
Bzm
4-H ), 14.12 (s, 1H, N+H).
Bzm
2-(Morpholin-4-yl)-1-[2-(morpholin-4-yl)ethyl]-1H-b
enzimidazole (II) was prepared analogously.
Yield 81%, mp 127 – 128°C (CH CN). C H N O
2
3
17 24
4
PMR spectrum (300 MHz, CDCl ), d, ppm: 2.49 – 2.52 (t, J
3
4.50 Hz, 4H, CH -N-CH ), 2.74 – 2.79 (t, J 7.05 Hz, 2H,
2
2
N
-CH ), 3.31 – 3.34 (t, J 4.80 Hz, 4H, CH -O-CH ),
2 2 2
exocycl
3.65 – 3.68 (t, J 4.65 Hz, 4H, CH -N-CH ), 3.86 – 3.89 (t, J
2
2
EXPERIMENTAL CHEMICAL PART
4.65 Hz, 4H, CH -O-CH ), 4.13 – 4.17 (t, J 7.05 Hz, 2H,
2
2
N
-CH ), 7.18 – 7.28 (m, 3H, H
), 7.62 – 7.63 (m, 1H,
Arom
Het.
2
PMR spectra of synthesized compounds were recorded
on Unity-300 (Varian, USA) and Avance-600 spectrometers
(Bruker, USA). Residual resonances of deuterated solvents
(DMSO-d and CDCl ; d 2.49 and 7.24 ppm, respectively)
H
).
Arom
1-Benzyl-2-(pyrrolidin-1-yl)-1H-benzimidazole hy-
drochloride (III). 1-Benzylbenzimidazole-2-sulfonic acid
(VI, 2.88 g, 10 mmol) [12] in pyrrolidine (5 mL, 60 mmol)
in a sealed ampul or autoclave was heated at 150 – 160°C on
an oil bath for 5 – 6 h, cooled, and poured into ice water. The
6
3
were used as internal standards. Melting points were mea-
sured on a Fisher-Johns melting point apparatus (Fisher Sci-
entific, USA). Elemental analyses used classical methods
(Table 1) [14] and agreed with those calculated for C, H, N,
and Cl. The course of reactions and purity of products were
monitored by TLC (Al O activity grade III plates, CHCl
resulting precipitate was filtered off, rinsed with H O, and
2
dried. The obtained 1-benzyl-2-(pyrrolidin-1-yl)-1H-benz-
imidazole was passed through an Al O layer (1.5 ´ 20 cm)
2
3
2
3
3
with elution by CHCl . The eluate was evaporated. The col-
elution, I development in a humid chamber).
3
2
orless crystals were dissolved in the minimal amount of
2-(Morpholin-4-yl)-1-(prop-2-en-1-yl)-1H-benzimida
Me CO and acidified with HCl in PrOH.
zole hydrochloride (I). A suspension of 1-(2-benzimida-
2
Yield 1.93 (70%), mp 231 – 232°C. C H N × HCl.
zole)morpholine (2.03 g, 10 mmol) in Me CO (15 mL) was
18 19
3
2
PMR spectrum (600 MHz, DMSO-d ), d, ppm: 1.94 – 1.96
stirred vigorously at room temperature and treated dropwise
with 85% aqueous KOH (1.32 g, 20 mmol) (2 mL) slow
6
(q, J 3.27 Hz, 4H, CH -CH ), 3.72 – 3.74 (t, J 6.6 Hz, 4H,
2
2

Synthesis and Pharmacological Activity
203
CH -N-CH ), 5.64 (s, 2H, N -CH ), 7.23 – 7.51 (m, 3H,
7.47 (s, 1H, 4-H ), 7.58 (d, J 7.74 Hz, 1H, 7-H ), 10.32
2
2
Het.
2
Bzm
Bzm
H
), 7.28 – 7.32 (m, 2H, H
), 7.35 – 7.38 (m, 2H,
Arom
(br.s, 1H, N+H). mp 123 – 125°C.
Arom
2-{4-[(5-Methoxy-1-methyl-1H-benzimidazol-2-yl)me-
thyl]piperazin-1-yl}ethanol (XI).
H
), 7.42 (d, J 7.98 Hz, 1H, H
), 13.5 (br.s, 1H, N+H).
), 7.51 (d, J 7.86 Hz,
Arom
Arom
1H, H
Arom
Yield 81%, mp 142 – 143°C (CH CN). C H N O .
1-Benzyl-N-[2-(morpholin-4-yl)ethyl]-1H-benzimida-
zol-2-amine dihydrochloride (IV) was prepared analo-
gously to III.
3
16 24
4
2
PMR spectrum (300 MHz, CDCl ), d, ppm: 2.52 (m, 11H,
3
CH
+ CH
+ OH), 3.59 (t, J 5.4 Hz, 2H, CH ),
2endocycl.
2exocycl. 2
3.78 (s, 2H, N-CH ), 3.83 (s, 3H, CH ), 3.85 (s, 3H, OCH ),
Yield 71%, mp 248 – 250°C. C H N O×2HCl. PMR
3
2
3
20 24
4
6.91 (dd, J 3.69 Hz, 1H, 6-H ), 7.21 (t, J 4.05 Hz, 2H,
spectrum (300 MHz, CDCl ), d, ppm: 3.18 – 4.49 (m, 6H,
Bzm
3
4.7-H ). 2-{4-[(5-Methoxy-1-methyl-1H-benzimidazol-2-
CH -N-CH , CH ), 3.91 – 4.04 (m, 6H CH -O-CH , CH ),
Bzm
2
2
2
2
2
2
yl)methy]piperazin-1-yl}ethanol hydrochloride, mp 236 –
237°C.
7.23 – 7.35 (m, 7H, H ), 7.41 – 7.51 (m, 2H, H ), 9.69
arom
arom
(s, 1H, NH), 11.25 (br.s, 1H, N+H), 14.2 (br.s, 1H, N+H).
2-(Pyrrolidin-1-ylmethyl)-1H-benzimidazole dihydro-
chloride (VIII). A solution of IIIa (4.16 g, 25 mmol) and
pyrrolidine (4.1 mL, 50 mmol) in dioxane (27 mL) was
refluxed over NaOH for 3 h. The dioxane (20 mL) was dis-
tilled off. The reaction mixture was cooled and treated with
2-{[4-(2, 5-Dimethylphenyl)piperazin-1-yl]methyl}-5-
methoxy-1-methyl-1H-benzimidazole (XII).
Yield 79.5%, mp 156 – 157°C (CH CN). C H N O.
3
22 28
4
PMR spectrum (300 MHz, CDCl ), d, ppm: 2.24 (s, 3H,
3
CH Ph), 2.86 (c, 3 H, CH ), 2.67 (br.s, 4H, CH ),
2endocycl
3
3Ph
2.90 (t,
J
4.65 Hz, 4H, CH
), 3.86 (s, 5H,
2endocycl
H O (50 mL). The precipitate was filtered off, rinsed on the
2
N-CH +C-CH ), 3.88 (c, 3H, OCH ), 6.79 (d, 2H, H ), 6.95
filter with H O (50 mL), and dried. The obtained
3
2
3
Ph
2
(dd, J 3.79 Hz, 1H, 6-H ), 7.06 (d, J 7.8 Hz, 1H, H ),
2-(pyrrolidin-1-ylmethyl)-1H-benzimidazole was crystal-
av
Bzm
Ph
7.22 – 7.24 (m, 2H, 4.7-H ).
lized from MeCN (mp 147 – 148°C), dissolved in the mini-
Bzm
2-({4-[bis(4-Fluorophenyl)methyl]piperazin-1-yl}me-
thyl)-5-methoxy-1-methyl-1H-benzimidazole (XIII).
Yield 92.4%, mp 156 – 157°C (CH CN). C H F N O.
mal amount of Me CO, and acidified with HCl in PrOH to
2
pH 1.
Yield 65%, mp 234 – 236°C. C H N × 2HCl. PMR
3
27 28
2
4
12 15
3
PMR spectrum (300 MHz, CDCl ), d, ppm: 2.35 (br.s, 4H,
spectrum (300 MHz, DMSO-d ), d, ppm: 1.95 (br.s, 4H,
3
6
CH
), 2.53 (br.s, 4H, CH
), 3.78 (s, 2H, C-CH ),
CH -CH
), 3.39 (br.s, 4H, CH NCH
), 4.80 (s,
2-pyrrolid.
2endocycl
2endocycl 2
2
2-pyrrolid.
2
3.83 (s, 3H, N-CH ), 3.83 (s, 3H, ÎCH ), 4.2 (s, 1H, CH),
2H, C-CH ), 7.42 – 7.45 (q, J 3.09 Hz, 2H, H ),
3
3
2
arom
6.89 (m, 5H, 7-H
+ 4H ), 7.2 (m, 2H, 6.4-H ),
Ph Bzm
7.74 – 7.78 (q, J 3.09 Hz, 2H, H ), 9.5, 11.8 (br.s, 2H,
N+H).
Bzm
arom
7.29 – 7.34 (m, 4H, H ).
Ph
1,5-Dimethyl-2-(pyrrolidin-1-ylmethyl)-1H-benzimi-
dazole hydrochloride (IX). A solution of XIV (0.9 g,
4.6 mmol) and pyrrolidine (0.76 mL, 9.2 mmol) in anhy-
drous C H (8 mL) was refluxed for 3.5 h and cooled.
EXPERIMENTAL PHARMACOLOGICAL PART
The properties of 10 compounds (I-IV and VIII-XIII)
were studied for the pharmacological part of the work. The
compounds were converted to the hydrochloride or dihydro-
chloride salts in HCl-saturated i-PrOH to improve the water
solubility.
6
6
Pyrrolidine hydrochloride was extracted by H O (30 mL).
2
The C H fraction was separated, dried over anhydrous
6
6
Na SO , evaporated to the minimal volume, and passed
2
4
through a layer of Al O (h = 2 cm; d = 3 cm) with elution
2
3
Antiglycation activity was determined in vitro by the lit-
erature method [15]. The reaction mixture contained glucose
(500 mmol) and bovine serum albumin (BSA, 1 mg/mL) in
phosphate buffer (pH 7.4). BSA was incubated before the
specific fluorescence of glycated BSA was measured on an
F-7000 fluorescence spectrometer (Hitachi, Japan) at
by CHCl . The eluate was evaporated. The solid was dis-
3
solved in the minimal amount of Me CO and acidified with
2
HCl in PrOH to pH 1.
Yield 70%, mp 207 – 208°C (C H OH). C H N HCl.
2
5
14 19
3
PMR spectrum (300 MHz, DMSO-d ), d, ppm: 1.95 (br.s,
6
4H, CH -CH
), 2.48 (s, 3H, C-CH ), 3.37 (br.s, 4H,
3
2
2-pyrrolid.
l
= 370 nm and l = 440 nm. The reference compound
em
CH NCH
), 4.01 (s, 3H, N-CH ), 4.89 (s, 2H, CH ),
3 2
ex
2
2-pyrrolid.
was the known non-enzymatic glycosylation inhibitor
aminoguanidine (Sigma, USA) [16]. Antiglycation activity
of the compounds was calculated vs. the fluorescence of con-
trol samples.
7.35 (d, J 8.4 Hz, 1H, 6-H ), 7.58 (s, 1H, 4-H ), 7.73 (d,
Bzm
Bzm
J 8.4, 7-H ), 11.8 (br.s, 1H, N+H).
Bzm
Compounds X-XIII were synthesized analogously.
2-{4-[(1, 5-Dimethyl-1H-benzimidazol-2-yl)methyl]pi-
perazin-1-yl}ethanol hydrochloride (X).
DPP-4 inhibitory activity was evaluated in vitro using a
modified literature method [17]. A solution of the tested
compound (10 mL, 10 mmol) was mixed with Tris-HCl
buffer (50 mL, 0.1 M, pH 8.4) and human plasma (40 mL).
The mixture was incubated beforehand at 37°C for 5 min,
treated with reaction substrate (100 mL, 1 mmol, Gly-Pro-p-
nitroaniline; Sigma, USA), and incubated at 37°C for
Yield 90%, mp 233 – 235°C (C H OH). C H N O ×
2
5
16 24
4
HCl. PMR spectrum (600 MHz, DMSO-d ), d, ppm: 2.43 (s,
6
3H, C-CH ), 2.71 (s, 2H, N
-CH ), 2.95 (s, 2H,
3
endocycl
2
N
-CH ), 3.14 (c, 4H, N
-CH ), 3.44 (c, 2H,
endocycl
2
endocycl
2
CH ), 3.75 (c, 2H, CH ), 3.76 (c, 3H, N-CH ), 3.99 (c, 2H,
2
2
3
C-CH ), 5.5 (br.s, 1H, OH), 7.21 (d, J 8.04 Hz, 1H, 6-H ),
2
Bzm

204
O. N. Zhukovskaya et al.
15 min. The yellow color that appeared from release of 4-nit-
roaniline was assayed at 405 nm using a universal automated
photometer (ELx800, BioTec, USA). The reference com-
pound was sitaglyptin (Merck Sharp & Dohme, Nether-
lands).
centrator, Russia). Spasmogenic effects were induced by
5-hydroxytryptamine (Sigma, USA) at a concentration of
1 mM. Tested compounds were studied at the equimolar con-
centration (1 mM). The reference drug was ketanserine
(Sigma, USA). Contractions were recorded using a sin-
gle-chamber system for working with isolated organs and tis-
sues (Ugo Basile S. R. L., Italy) and LabScribe 3.0™ soft-
ware (iWorx Systems, Inc., USA). The activity level was as-
sessed from the change of control and test measurements.
Antioxidant activity was studied in vitro using an
ascorbate-dependent lipid peroxidation (LPO) model [18].
The substrate was rat-liver homogenate (4%). The reaction
was initiated by ascorbic acid (50 mM; Chemapol, Czech
Rep.). The oxidation rate was determined from the accumu-
lation of malondialdehyde using the reaction with thiobarbi-
turic acid (Fluka, Switzerland). Optical density of the col-
ored product in a 10-mm cuvette was measured at 532 nm on
a PD-303 UV spectrophotometer (APEL, Japan). Com-
pounds were studied at a concentration of 1 ìM. Activities of
compounds were evaluated in percent vs. a sample without
the compound. The reference drug was dibunol (Merck, Ger-
many).
Antagonist activity for angiotensin AT -receptors was
1
studied in vitro using isolated portal vein from laboratory rats
of both sexes according to the literature method [21] with
modifications [22] and Krebs buffer solution (pH 7.4) with
constant oxygenation with 95% O (Armed 7F-3A O gener-
2
2
ator, Russia) and thermostatting at 24°C. Compound activi-
ties were evaluated from the degree of suppression of spas-
mogenic effects by angiotensin II (0.01 mM, Sigma, USA).
Contractions were recorded in a single-chamber system for
working with isolated organs and tissues (Ugo Basile S. R.
L., Italy) using LabScribe 3.0™ software (iWorx Systems,
Inc., USA). Compounds were studied at a concentration of
The blocking effect on serotonin 5-HT receptors was
3
studied using the published method [19]. The experiments
used preparations of isolated ilium from six mature male lab-
oratory guinea pigs (300 – 450 g) using Krebs buffer (pH
10 mM. The reference was the AT -antagonist losartan
1
7.4) with constant oxygenation with 95% O (Armed 7F-3A
(Sigma, USA).
2
O concentrator, Russia) and thermostatting at 37°C. The se-
Animals were kept and manipulated in the experiments
according to international recommendations of the European
Convention for the Protection of Vertebrate Animals Used
for Experimental and Other Scientific Purposes (1986) and
good laboratory practice rules for preclinical trials in the RF
according to “Good laboratory practice rules” (GOST R
33647 – 2015) and “On approval of good laboratory practice
rules” (Ministry of Health of the RF, Order No. 199n dated
April 1, 2016). Experiments were conducted according to
recommendations of the local ethics committee.
2
rotonin receptor agonist was 5-hydroxytryptamine (1 mM,
Sigma, USA). Pharmacological responses were recorded on
a system for working with isolated organs (Ugo Basile, It-
aly). All compounds were used at a concentration of 1 mM.
The reference drug was tropisetron (Sigma, USA). The de-
gree of receptor inhibition was assessed by comparing
spasmogenic effects measured for control and test speci-
mens.
5-HT antagonist activity was studied using serotonin-
2A
induced contractile activity of isolated rat uterine horn [20].
The experiments used De Jalon buffer (pH = 7.4, 24°C) with
constant oxygenation with 95% O (Armed 7F-3A O con-
Results were statistically processed using nonparametric
methods (check for normal distributions using the
Kolmogorov–Smirnov test as corrected by Lilliefors,
2
2
TABLE 1. Elemental Analyses of Benzimidazole Derivatives I-IV and VIII-XIII
Found, %
Calc., %
Halogen
Compound
Empirical formula
C
H
Halogen
12.50
–
N
C
H
N
59.93
64.35
68.70
58.49
52.37
63.14
58.97
62.92
72.31
69.92
6.34
7.47
6.23
6.22
6.09
7.46
7.57
7.74
7.59
5.92
14.84
17.54
13.21
13.5
60.10
64.53
68.89
58.67
52.54
63.26
59.16
63.13
72.50
70.11
6.48
7.65
6.42
6.40
6.25
7.58
7.76
7.95
7.74
6.10
12.67
–
15.02
17.71
13.39
13.68
15.32
15.81
17.25
18.41
15.37
12.11
I × HCl
II
C14H17N3O × HCl
C17H24N4O2
III
C18H19N HCl
11.11
17.16
25.72
13.23
10.72
–
11.29
17.34
25.89
13.35
10.91
–
IV × 2HCl
VIII × 2HCl
IX × HCl
X × HCl
XI
C20H24N4O × 2HCl
C12H15N3 × 2HCl
C14H19N3 × HCl
C16H24N4O × HCl
C16H24N4O2
15.15
15.69
16.45
18.20
15.16
12.01
XII
C22H28N4O
–
–
XIII
C27H28F2N4O
–
–

Synthesis and Pharmacological Activity
205
TABLE 2. Antiglycation, DPP-4 Inhibitory, and Antioxidant Activities of C(2)-Substituted Benzimidazoles
Antiglycation activity
Antioxidant activity,
% inhibition,
DPP-4 inhibitory activity
(inhibition of BSA fluorescence),
Compound
(0.1 mmol) (M ± m)
1 mmol, D% (M ± m)
(1 mmol) (M ± m)
94.1 ± 1.26*#
89.7 ± 2.17*#
45.5 ± 1.29*#
54.4 ± 2.57*
–16.6 ± 0.34*#
–15.8 ± 0.76*#
0.5 ± 4.51#
– 9.4 ± 6.59#
–15.2 ± 2.71*#
–36.1 ± 11.08*#
70.7 ± 5.71*
…
5.3 ± 3.28#
7.6 ± 4.34#
9.7 ± 3.67*#
30.6 ± 10.0*#
6.6 ± 7.15#
–9.9 ± 13.15#
11.8 ± 3.08*#
0.01 ± 7.70#
2.1 ± 4.65#
–8.7 ± 3.35*#
…
2.1 ± 0.15*#
2.9 ± 0.50*#
0.0 ± 0.00
0.0 ± 0.00
0.0 ± 0.00
2.9 ± 0.06*#
0.0 ± 0.00
0.7 ± 0.06#
0.5 ± 0.04#
0.0 ± 0.00
…
I
II
III
IV
VIII
IX
X
XI
XII
XIII
Aminoguanidine
Sitaglyptin
Dibunol
…
–87.9 ± 1.61*
…
…
38.2 ± 1.01*
Differences statistically significant (p < 0.05) vs. * the control and # reference drug; …, not studied.
p < 0.05), e.g., Kruskal–Wallis criterion with a Dunn
post-test for multiple intergroup comparison and the Mann–
Whitney U-criterion for paired intergroup comparison in the
GraphPad Prism 5.0 program. Data were given as the means
errors of the mean.
RESULTS AND DISCUSSION
The new benzimidazoles showed different levels of
antiglycation activity in the conducted tests (Table 2). Thus,
3 of the 10 studied pharmaceutical substances inhibited BSA
TABLE 3. AT - and 5-HT
-Antagonist Activity of C(2)-Substituted Benzimidazoles
2A,3
1
5-HT3-Antagonist activity,
5-HT2a-Antagonist activity,
AT1-Antagonist activity
Compound
D% (1 mmol) (M ± m)
D% (1 mmol) (M ± m)
(10 mmol) (M ± m)
–0.7 ± 0.41#
–0.3 ± 0.20#
–1.9 ± 1.28#
–22.5 ± 2.50*#
–1.1 ± 1.10#
–2.6 ± 1.51#
–1.8 ± 0.95#
–0.7 ± 0.59#
…
–2.9 ± 0.7#
–3.0 ± 1.1#
–1.2 ± 0.2#
…
0.0 ± 0.00
0.0 ± 0.00
–27.5 ± 3.8*#
0.0 ± 0.00
0.0 ± 0.00
–15.9 ± 1.8*#
–3.3 ± 1.7#
0.0 ± 0.00
–52.5 ± 4.5*#
– 2.6 ± 1.3#
…
I
II
III
IV
–2.0 ± 1.2#
–0.7 ± 0.2#
–5.3 ± 0.6#
–2.4 ± 0.7#
–4.6 ± 1.0#
…
VIII
IX
X
XI
XII
–1.0 ± 0.55#
XIII
Ketanserine
Tropisetron
Losartan
…
–71.3 ± 3.5*
…
…
…
–30.2 ± 4.0*
…
–99.5 ± 0.2*
Differences statistically significant (p < 0.05) vs. * the control and # reference drug; …, not studied.

206
O. N. Zhukovskaya et al.
fluorescence by 50%. Compounds I and II were not only
comparable to aminoguanidine in antiglycation activity but
statistically significantly exceeded it. The activity of IV was
at the level of the reference drug. It is noteworthy separately
that all compounds that exhibited high levels of antiglycation
activity had morpholine as the 2-substituent. Compound III
with a pyrrolidine group showed moderate activity. The
antiglycation activities of the other compounds were weak or
absent.
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ACKNOWLEDGMENTS
The synthesis and development of pharmaceutical sub-
stances was financially supported by the Ministry of Educa-
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at the CUC of Southern Federal University.
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