Synthesis and insight into the structure-activity relationships of 2-phenylbenzimidazoles as prospective anticancer agents

Article abstract of DOI:10.1039/d0ra02282a

In order to explore and develop new anticancer agents, three series of 2-phenylbenzimidazoles,15-46, were condensed under simple and mild conditions using sodium metabisulfite as an oxidation agent and another series,47-55, were obtainedviaa reduction rea

Full text of DOI:10.1039/d0ra02282a

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Synthesis and insight into the structure–activity
relationships of 2-phenylbenzimidazoles as
prospective anticancer agents†
Cite this: RSC Adv., 2020, 10, 20543
Thi-Kim-Chi Huynh,ab Thi-Hong-An Nguyen,a Thi-Cam-Thu Nguyenac
ab
*
and Thi-Kim-Dung Hoang
In order to explore and develop new anticancer agents, three series of 2-phenylbenzimidazoles, 15–46,
were condensed under simple and mild conditions using sodium metabisulfite as an oxidation agent and
another series, 47–55, were obtained via a reduction reaction using sodium borohydride. All the
compounds synthesized were evaluated for their in vitro anticancer activities against three human
cancer cell lines. The novel compound 38 was found to be the most potent multi cancer inhibitor
against A549, MDA-MB-231, and PC3 cell lines (IC50 values 4.47, 4.68 and 5.50 mg mLꢀ1, respectively). In
addition, compound 40 exhibited the best IC50 value of 3.55 mg mLꢀ1 against the MDA-MB-231 cell line.
The results demonstrated that introducing a new substituent to compounds 37–55 could improve their
antiproliferative activities.
Received 11th March 2020
Accepted 10th May 2020
DOI: 10.1039/d0ra02282a
rsc.li/rsc-advances
antihypertensive, and antiparasitic properties.3–12 In particular,
some of the commercial anticancer drugs contain the benz-
1. Introduction
imidazole scaffold (Fig. 1), so benzimidazole is one of the most
promising structures used against several human cancer cell
lines. Many studies have reported that benzimidazole deriva-
tives that contain functional groups at the 1, 2, 5 and/or 6-
positions on their skeleton show the most efficient anticancer
activity.13–17 However, previous studies18–21 that evaluated the
antiproliferative activity of 2-phenylbenzimidazoles had a small
number of desired products which led to a lack of conclusion on
the structure–activity relationship (SAR), the effect of the pres-
ence of substituents into both the C-5/6 and C-2 positions on
benzimidazole structure and also the bioactivity. In addition,
there are two general pathways to synthesize benzimidazole
derivatives. The rst method involves the condensation of o-
phenylenediamines and carboxylic acids (or their derivatives
such as nitriles, chlorides and orthoesters) in the presence of
acid as we have previously reported.22 The second one is con-
ducted by using o-phenylenediamines in combination with
aldehydes instead of the carboxylic acids and oxidation agents
that should have been required for this process.23–30 According
to our latest publication, a large number of benzimidazoles
were synthesized from o-phenylenediamines and aldehydes
under simple and mild conditions using sodium metabisulte
(Na2S2O5) as the oxidative reagent and a solvent which was
a mixture of ethanol : water (9 : 1 v/v) in high yields.30 Thus, it
was recognized that this pathway is convenient to synthesize
benzimidazoles that contain a variety of substituents on the C-2,
C-5 and C-6 positions of the benzimidazole scaffold without
undesired products. Continuing with our plans to develop new
anti-tumor agents,22,31,32 in this research, we planned to
Cancer is a phenomenon that involves abnormal cell growth
invading or spreading to other parts of the organism. There are
more than 100 types of cancers which affect humans and that
has led to the discovery of many options for cancer treatment
such as chemotherapy, surgery, radiation therapy, targeted
therapy, immunotherapy, and so on. In particular, chemo-
therapy is required in the treatment, either alone or in combi-
nation with the other therapies depending on type of cancer, its
location, and its grade. For example, chemotherapy could
reduce the size of an inoperable tumor, which could then be
operated on in the future. However, the increase in multidrug
resistance (MDR) to chemotherapy is a contributing factor that
inuences the treatment of cancer1 and side effects can also
limit the effectiveness of anticancer drugs such as nausea,
arthralgia, fatigue, osteoporosis, and so on.2 Therefore, the
exploration and development of new anticancer drugs that have
better efficacy and fewer side effects are required.
Benzimidazole structures are well-known heterocyclic
compounds that possess a wide range of antiviral, anti-
histaminic,
antifungal,
antiallergic,
anticoagulant,
aInstitute of Chemical Technology – VAST, 01 Mac Dinh Chi Str., Dist. 1, Ho Chi Minh
City, Vietnam. E-mail: hoangthikimdung@gmail.com
bGraduate University of Science and Technology – VAST, 18 Hoang Quoc Viet Str., Cau
Giay Dist., Hanoi, Vietnam
cTon Duc Thang University, 19 Nguyen Huu Tho Str., Dist. 7, Ho Chi Minh City,
Vietnam
† Electronic supplementary information (ESI) available. See DOI:
10.1039/d0ra02282a
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carbonyl group in compounds 37–46 was reduced to a corre-
sponding hydroxyl group in the presence of NaBH4 in methanol
to give the desired benzimidazole derivatives 47–55 with yields
of 45% to 98%. The structures of the synthesized compounds
are shown in the Table 1. All the synthesized compounds 15–55
were carefully characterized using FTIR, 1H-NMR, 13C-NMR
spectroscopic and ESI-HRMS spectrometric methods. The
FTIR bands at 3256, 3025, 1613 cmꢀ1 conrmed the presence of
–NH, C–H and C]N functionalities, respectively, of the most
1
potent compound 38. The H-NMR spectrum showed a multi-
plet at d 6.95–7.94 ppm conrming the presence of aromatic
protons and benzimidazole protons. Similarly, in the 13C-NMR
spectrum of compound 38, the carbonyl carbon was at
d 195.58 ppm and the remaining carbons all belonged to
aromatic carbons which appeared in the range of d 113.53–
Fig.
structure.
1 Some anticancer drugs that contain the benzimidazole
synthesize a large number of new 2-phenylbenzimidazole
derivatives with a diverse range of substituents on the C-2, C-5
and C-6 positions via condensation of o-phenylenediamines
and various aldehydes under the conditions mentioned previ-
ously and then to elucidate their SAR against three cancer cell
lines: A549, MDA-MB-231, and PC3. The aim was that the
presence of substituents such as carbonyl and hydroxyl groups
in the C-5 position together with substituents at the C-2 position
would affect the bioactivity of targeted products and their
presence could signicantly enhance the anticancer activity of
the novel compounds.
1
157.79 ppm. The fashion in the FTIR, H-NMR, and 13C-NMR
spectra similar patterns were observed for the other
compounds 37–46. Moreover, the appearance of carbon which
was eliminated from a carbonyl group (compounds 37–46) to
form a hydroxyl group (compounds 47–55) was recognized by
13C-NRM high eld signals at d 73.77–74.58 ppm. The HRMS
(ESI) spectra of all the compounds showed the pseudo-
molecular ion [M + H]+ or [M ꢀ H]ꢀ peaks corresponding to
their respective molecular weights. The spectroscopic data
(FTIR, 1H-NMR, 13C-NMR and ESI-HRMS) of all the newly
synthesized benzimidazoles 24, 35, 39, 41, 43–45 and 47–55
were in agreement with the corresponding structures illustrated
in the Experimental section.
2. Results and discussion
2.1. Chemistry
2.2. In vitro anticancer activities and structure–activity
relationships (SAR)
The preparation of the target 2-phenylbenzimidazoles 15–55 is
shown in Scheme 1. Firstly, the ortho-phenylenediamine deriv-
atives were sequentially condensed with various benzaldehydes All the synthesized benzimidazole derivatives 15–55 were tested
that contain different substituents (–OH, –OCH3, –NO2, for their cytotoxicity in vitro against human cancer cell lines:
–N(CH3)2, –CF3, –I, –O-CH2-Ph). This reaction was carried out A549 (human lung adenocarcinoma epithelial cell line), MDA-
under mild conditions using a mixture of solvents: ethanol : - MB-231 (human breast cancer cell line) and PC3 (human pros-
water (ratio of 9 : 1 v/v) and Na2S2O5 as an oxidation agent to tate cancer cell line), using camptothecin as a standard drug
produce a series of benzimidazoles, 15–46, that have substit- and the IC50 values (mg mLꢀ1) are detailed in Table 2. Notably,
uent groups such as hydro, chloro, phenyl methanonyl at the 5, the tested compounds showed a moderate to good anti-
6 positions, with yields ranging from 32% to 98%. Next, the proliferative activity on selected cancer cell lines. From the
Scheme 1 Synthesis of different 2-phenyl benzimidazole derivatives 15–55.
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Table 1 Structures of the different substituents (X, Y, Z, R1, R2, R3, R4) used for the synthesis of benzimidazoles 15–55 and their yields (%)
Cpd no.
X
Y
Z
R1
R2
R3
R4
Yield (%)
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
H
H
H
H
H
H
H
H
H
H
H
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Cl
Ph-CO–
Ph-CO–
Ph-CO–
Ph-CO–
Ph-CO–
Ph-CO–
Ph-CO–
Ph-CO–
Ph-CO–
Ph-CO–
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
—
OH
H
H
H
OCH3
H
H
OH
H
H
H
OCH3
H
H
H
OCH3
H
H
OH
H
H
H
OCH3
H
H
H
OCH3
H
H
OH
H
H
OCH3
H
H
H
OCH3
H
H
OH
H
H
OCH3
H
H
H
H
OH
OCH3
H
OCH3
N(CH3)2
H
H
OH
–O-CH2-Ph
H
H
OH
OCH3
H
OCH3
N(CH3)2
H
H
OH
–O-CH2-Ph
H
H
H
H
H
H
OCH3
OCH3
H
H
H
I
H
H
H
H
H
OCH3
OCH3
H
H
H
I
H
H
H
H
H
OCH3
H
H
H
I
H
H
68
54
41
63
84
81
81
67
83
98
48
85
78
78
87
82
90
73
32
52
85
68
75
78
84
84
83
98
54
53
98
76
56
72
45
84
92
64
53
98
57
H
NO2
CF3
H
H
OH
H
H
H
OCH3
H
H
NO2
CF3
H
H
OH
H
H
H
H
H
NO2
CF3
H
H
OH
H
H
H
H
H
OH
OCH3
OCH3
N(CH3)2
H
H
OH
–O-CH2-Ph
H
H
Ph-CH(OH)–
Ph-CH(OH)–
Ph-CH(OH)–
Ph-CH(OH)–
Ph-CH(OH)–
Ph-CH(OH)–
Ph-CH(OH)–
Ph-CH(OH)–
Ph-CH(OH)–
—
—
—
—
—
—
—
—
H
H
H
OCH3
H
H
I
H
OH
OCH3
OCH3
N(CH3)2
H
OH
–O-CH2-Ph
CF3
H
H
OCH3
H
overall results, there were 11 compounds such as 16, 19, 21, 22,
The difference in the electron donating and electron with-
23, 24, 28, 38, 45, 47, and 49 which displayed IC50 values less drawing groups in the 2-phenyl ring affected the activity of the
than 100 mg mLꢀ1 on the three cancer cell lines. There were ve synthesized compounds and this effect was found to be related
compounds (23, 25, 28, 38, 39), two compounds (23, 38) and one to the substituents in the C-5 position. To clarify this point, the
compound (38) that displayed IC50 values of less than 8 mg mLꢀ1 activities of compounds containing the hydroxyl group (electron
on the MDA-MB-231, PC3, and A549 cell lines, respectively. donating group) were compared to those of compounds con-
From a scientic point of view, the MDA-MB-231 human breast taining the triuoromethyl group (electron withdrawing group)
cancer cell line was found to be more sensitive towards the at the R1-position such as 15 vs. 23 (containing one chloro atom
synthesized 2-phenylbenzimidazoles than the PC3 and A549 cell at the C-5 position), 26 vs. 34 (containing two chloro atoms at
lines.
the C-5 and C-6 positions), 37 vs. 44 (containing a carbonyl
group at the C-5 position) and 47 vs. 53 (containing a hydroxyl
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Table 2 Anti-proliferative activity (IC50, mg mLꢀ1) of 41 benzimidazole
derivatives against three cancer cell lines, determined by MTT assay
the anticancer effect of 37/47 more than that of 44/53 which
consisted of a triuoromethyl group (IC50 values of compounds
37, 47, 44 and 53 were 44.67, 14.79, >100 and 54.95 mg mLꢀ1
,
IC50 ꢁ SD (mg mLꢀ1
)
respectively).
Cpd no.
A549a
MDA-MB-231b
PC3c
To demonstrate that the chloro atoms on the benzimidazole
skeleton make a signicant contribution to the anticancer
activity, compounds 15–25 and 26–36 were synthesized that
contain one and two chloro atoms in the 5- and 6-positions,
respectively. The compounds containing one chloro atom at the
5-position such as 21, 23 and 25 reached IC50 values of 10.7, 7.08
and 6.61 mg mLꢀ1 with the MDA-MB-231 cell line, respectively.
Unfortunately, for all three cancer cell lines of compounds, the
anti-tumor activity of compounds that have two chloro atoms in
the 5-, and 6-positions was considerably decreased and was less
than that of compounds with one chloro atom in the 5-position,
for example, 21 vs. 32, 23 vs. 34 and 25 vs. 36, but excluding
compound 28, which had an IC50 value of 6.92 mg mLꢀ1 for the
MDA-MB-231 cell line. In the further studies, other halogen
atoms (such as uoro or bromo atoms) will be investigated at
the 5-, and 6-positions in the benzimidazole structure so that
those products may improve the antiproliferative activity, for
example, against A549 cell lines, as reported previously in the
literature.21
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
>100
43.65 ꢁ 1.58
22.39 ꢁ 1.05
>100
>100
12.02 ꢁ 0.56
31.62 ꢁ 1.46
>100
44.67 ꢁ 1.69
11.48 ꢁ 0.74
9.12 ꢁ 0.55
38.02 ꢁ 1.89
45.71 ꢁ 1.75
39.81 ꢁ 1.03
>100
18.20 ꢁ 1.85
36.56 ꢁ 1.67
>100
>100
12.02 ꢁ 0.75
85.11 ꢁ 2.67
21.88 ꢁ 1.59
18.62 ꢁ 0.46
33.11 ꢁ 1.43
6.92 ꢁ 0.62
39.26 ꢁ 1.59
19.95 ꢁ 1.08
>100
>100
10.72 ꢁ 1.11
30.2 ꢁ 1.53
7.08 ꢁ 0.86
37.15 ꢁ 1.73
6.61 ꢁ 0.83
>100
>100
>100
>100
>100
64.57 ꢁ 2.68
6.92 ꢁ 0.98
60.26 ꢁ 2.12
>100
>100
>100
85.11 ꢁ 2.12
>100
>100
69.18 ꢁ 2.09
>100
31
32
33
34
83.18 ꢁ 2.05
52.48 ꢁ 2.34
>100
60.26 ꢁ 2.42
>100
>100
>100
>100
ND
45.71 ꢁ 1.56
>100
ND
35
ND
To nd prospective anticancer agents, the phenyl meth-
anonyl group was introduced into the benzene ring at the 5-
position and compounds 37–46 were obtained. Nevertheless,
this replacement lead to a decrease in the activity, compounds
39 and 42–46, when compared to other compounds that had
one chloro atom in the 5-position (compounds 17 and 21–25).
Surprisingly, there was a remarkable increase in the anti-
proliferative activity of benzimidazoles 38 and 40. In detail, the
IC50 value of compound 40 with the MDA-MB-231 cell line
reached 3.55 mg mLꢀ1 which had decreased dramatically from
more than 100 mg mLꢀ1 – the IC50 value of compound 18
(without a phenyl methanonyl group but with a chloro atom at
5-position). Specically, compound 38 acted as a potential multi
cancer inhibitor against A549, MDA-MB-231 and PC3 cell lines
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
60.26 ꢁ 1.25
>100
>100
>100
>100
44.67 ꢁ 1.69
4.68 ꢁ 0.42
>100
3.55 ꢁ 0.35
66.07 ꢁ 2.12
>100
4.47 ꢁ 0.33
>100
>100
>100
>100
5.50 ꢁ 0.28
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
42.66 ꢁ 1.62
46.77 ꢁ 1.50
54.95 ꢁ 1.96
>100
>100
>100
14.13 ꢁ 0.57
12.88 ꢁ 0.46
38.90 ꢁ 2.17
53.70 ꢁ 1.68
22.39 ꢁ 1.07
>100
14.79 ꢁ 1.08
>100
17.78 ꢁ 1.45
17.38 ꢁ 0.78
33.88 ꢁ 1.98
>100
30.2 ꢁ 2.04
>100
>100
>100
54.95 ꢁ 1.84
61.66 ꢁ 1.83
15.85 ꢁ 0.56
0.47 ꢁ 0.04
44.67 ꢁ 1.77
52
53
54
55
>100
with IC50 of 4.47 mg mLꢀ1, 4.68 mg mLꢀ1 and 5.50 mg mLꢀ1
,
47.86 ꢁ 1.73
>100
respectively.
>100
79.43 ꢁ 1.62
22.91 ꢁ 0.24
0.87 ꢁ 0.11
>100
0.2 ꢁ 0.06
To explore the effect of the presence of a carbonyl group in
compounds 37–42 and 44–46 on their bioactivity, the carbonyl
group was converted to a hydroxyl group by a reduction reaction
and produced the novel compounds 47–55. Basically, the
bioactivity on A549 and PC3 cell lines of compounds 47–55 was
better than that of compounds 37–46. This was shown by the
fact that there were six compounds (47–51 and 53) which dis-
played IC50 values less than 100 mg mLꢀ1 on the A549 cell line
instead of only two compounds (38 and 45). Conversely, that
modication gave no considerable change of the activity of
compounds 50–54, but compound 55 had an IC50 of 15.85 mg
mLꢀ1 with the MDA-MB-231 cell line.
In detail, the IC50 values of compound 48 on the three cancer
cell lines were slightly increased, in contrast to that of
compound 38 which did not have carbonyl group reduced to
a hydroxyl group. Considering the SARs of compound 47, 37, 26
and 15, the presence of substituent containing hydroxyl group
at the R1-position greatly increased the antiproliferative effect of
Camptothecind
a
b
Human lung adenocarcinoma epithelial cell line. Human breast
c
d
cancer cell line. Human prostate cancer cell line. Positive contrast
drug. All the values are showed as mean ꢁ SD in which each
treatment was calculated from at least three independent
experiments. ND ¼ not determined.
group at the C-5 position) on the MDA-MB-231 cell line and it
was realized that the activity of compounds 15/26 and 23/34 also
showed the same trend. The appearance of a hydroxyl group as
an electron donating group at the R1-position decreased the
activity of compound 15/26 in comparison to that of 23/34
containing triuoromethyl as a withdrawing electron group
(IC50 values of compounds 15, 26, 23 and 34 were 43.65 mg
mLꢀ1, >100, 7.08 and 45.71 mg mLꢀ1, respectively). In contrast to
this, introducing a hydroxyl group into the R1-position boosted
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compound 47, in comparison to those of the remained were recorded using electrospray ionization on a X500R QTOF
compounds (IC50 values ranged from 43.65 mg mLꢀ1 to more quadrupole time-of-ight (QTOF) system (Sciex).
than 100 mg mLꢀ1 on the three cancer cell lines). This was
shown by the fact that the IC50 values of compound 47 dropped
to 14.13, 14.79 mg mLꢀ1 and 17.78 mg mLꢀ1 with the A549, MDA-
4.3. General procedure for the synthesis of compounds 15–
MB-231 and PC3 cell lines, respectively. Aer the analysis of the
46
SARs, it was hypothesized that it was necessary to introduce the
carbonyl group into the benzimidazole structure at C-5 position A mixture of o-phenylenediamines 1–3 (2 mmol) and benzal-
and this adjustment markedly improved the anticancer activity dehyde derivatives 4–14 (2 mmol) were stirred in 10–20 mL of
of compounds 38 and 40.
ethanol and water (9 : 1 v/v) (until both dissolved) and then
Na2S2O5 (2 mmol) was added. The suspension was stirred
constantly at room temperature for 2 h and the reaction was
monitored by TLC. Aer completion of the reaction, the
precipitate was removed by ltering and the ltrate was evap-
orated under reduced pressure to obtain the crude product. The
crude product was washed three times with water and n-hexane,
and then dried in a vacuum at 80 ꢂC to achieve the nal product.
The purication was performed on a column chromatography
system or by crystallization from solvents with the desired
method as listed in the information for each product.
3. Conclusion
In summary, 41 2-phenylbenzimidazoles with various substit-
uents in the 5-, 6-positions and a 2-phenyl ring, including 17
novel derivatives, were synthesized under mild conditions using
a mixture of solvents: ethanol : water (9 : 1 v/v) to increase the
solubility of reactants (step 1) and NaBH4 in methanol (step 2)
and the products were obtained in high yields. All of synthe-
sized compounds were evaluated the antiproliferative activity
against A549, MDA-MB-231 and PC3 human cancer cell lines.
Compound 40 expressed a high selectivity towards the breast
cancer cell line MDA-MB-231 (IC50 ¼ 3.55 mg mLꢀ1). Whereas
compound 38 with the most potent anticancer activity showed
IC50 values of 4.47, 4.68 and 5.50 mg mLꢀ1 for the A549, MDA-
MB-231, and PC3 cell lines, respectively, and shows promise
as a new multi-cancer inhibitor. Benzimidazoles and the 2-
phenylbenzimidazole derivatives are also topoisomerase I
inhibitors.18,33–35 The studies to clarify the modes of action of the
synthesized 2-phenylbenzimidazoles binding on the topo I and
DNA complex are currently in progress.
2-(5-Chloro-1H-benzoimidazol-2-yl)-phenol (15): isolated by
silica gel column chromatography with chloroform : methanol
(98 : 2, v/v) to obtain a light yellow powder; yield: 68%; mp (ꢂC):
240–241; FTIR (KBr, n (cmꢀ1)): 3330 (N–H), 3057 (C–H), 1633 (C]
N), 1489 (C]C), 1257 (C–N), 738 (C–Cl); 1H-NMR (500 MHz,
DMSO-d6, d ppm): 7.02–8.06 (m, 7H, CHAr), 12.74 (s, 1H, OH),
13.23 (s, 1H, NH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 112.51–
157.79 (CHAr); ESI-HRMS (DMSO): m/z ¼ 245.0463 [M + H]+.
3-(5-Chloro-1H-benzoimidazol-2-yl)-phenol (16): light yellow
powder; yield: 54%; mp (ꢂC): 236–237; FTIR (KBr, n (cmꢀ1)):
3250 (N–H), 3060 (C–H), 1591 (C]N), 1466 (C]C), 1231 (C–N),
784 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm): 6.93–7.22 (m,
7H, CHAr), 9.75 (s, 1H, OH), 12.99 (s, 1H, NH); 13C-NMR (125
MHz, DMSO-d6, d ppm): 113.42–157.77 (CHAr); ESI-HRMS
(DMSO): m/z ¼ 245.0479 [M + H]+.
4. Experimental section
4.1. Materials
4-(5-Chloro-1H-benzoimidazol-2-yl)-phenol (17): slightly
brown powder; yield: 41%; mp (ꢂC): 259–260; FTIR (KBr, n
(cmꢀ1)): 3227 (N–H), 3070 (C–H), 1609 (C]N), 1461 (C]C),
All reagents and solvents were purchased from Acros Organics
(Belgium), Sigma-Aldrich (USA) and Xilong (China) and used
without further purication.
1
1251 (C–N), 729 (C–Cl); H-NMR (500 MHz, DMSO-d6, d ppm):
6.93–8.00 (m, 7H, CHAr), 10.04 (s, 1H, OH); 13C-NMR (125 MHz,
DMSO-d6, d ppm): 114.55–159.53 (CHAr); ESI-HRMS (DMSO): m/
4.2. Characterization of synthesized benzimidazoles
Reactions were monitored by thin-layer chromatography (TLC) z ¼ 245.0482 [M + H]+.
which was performed on silica gel 60 F254 plates (Merck). The
5-Chloro-2-(4-methoxy-phenyl)-1H-benzoimidazole
(18):
synthesized compounds were visualized by UV light (254 nm). All slightly yellow powder; yield: 63%; mp (ꢂC): 174–175; FTIR (KBr,
of the melting points (MPs) were determined on an IA 9000 series n (cmꢀ1)): 3007 (C–H), 1611 (C]N), 1492 (C]C), 1258 (C–N),
digital melting point apparatus (Electrothermal) and were 1180 (O–CH3), 738 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm):
uncorrected. The FTIR spectra were recorded using KBr pellets 3.38 (s, 3H, O–CH3), 6.67–7.67 (m, 7H, CHAr); 13C-NMR (125
with an Equinox 55 IR spectrometer (Bruker, Germany) and the MHz, DMSO-d6, d ppm): 55.30 (O–CH3), 114.38–160.88 (CHAr);
absorption bands were expressed in wavenumbers (cmꢀ1). The ESI-HRMS (DMSO): m/z ¼ 259.0633 [M + H]+.
1H-NMR and 13C-NMR spectra were recorded at 500 MHz (1H-
5-Chloro-2-(2,5-methoxy-phenyl)-1H-benzoimidazole
(19):
NMR) and 125 MHz (13C-NMR) on an AM0 FT-NMR spectrom- slightly yellow powder; yield: 84%; mp (ꢂC): 139–140; FTIR (KBr,
eter (Bruker) in (CD3)2SO. Tetramethylsilane (TMS) was used as n (cmꢀ1)): 3218 (N–H), 3008 (C–H), 1614 (C]N), 1490 (C]C),
an internal standard and the chemical shis were expressed in 1223 (C–N), 1176 (O–CH3), 740 (C–Cl); 1H-NMR (500 MHz,
d (ppm) and the coupling constants (J) in hertz (Hz). The signal DMSO-d6, d ppm): 3.80 (s, 3H, O–CH3), 3.98 (s, 3H, O–CH3),
multiplicities were expressed by standard abbreviations as 7.22–7.86 (m, 6H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm):
follows: s ¼ singlet, d ¼ doublet, t ¼ triplet, q ¼ quartet, dd ¼ 55.50, 56.17 (O–CH3), 113.50–153.18 (CHAr); ESI-HRMS (DMSO):
doublet of doublet, m ¼ multiplet. High-resolution mass spectra m/z ¼ 289.0734 [M + H]+.
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5-Chloro-2-(3,4,5-trimethoxy-phenyl)-1H-benzoimidazole
4-(5,6-Dichloro-1H-benzoimidazol-2-yl)phenol (28): yellow
(20): slightly gray powder; yield: 81%; mp (ꢂC): 233–234; FTIR powder; yield: 78%; mp (ꢂC): 209–210; FTIR (KBr, n (cmꢀ1)):
(KBr, n (cmꢀ1)): 3063 (C–H), 1588 (C]N), 1464 (C]C), 1240 (C– 3200 (OH), 3032 (C–H), 1609 (C]N), 1454 (C]C), 1250 (C–N),
N), 1127 (O–CH3), 731 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, 743 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm): 6.93–8.00 (m,
d ppm): 3.74 (s, 3H, O–CH3), 3.90 (s, 6H, 2 ꢃ O–CH3), 7.23–7.64 6H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm): 115.76–
(m, 5H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm): 56.04, 159.69 (CHAr); ESI-HRMS (DMSO): m/z ¼ 279.0088 [M + H]+.
60.11 (O–CH3), 104.00–153.21 (CHAr); ESI-HRMS (DMSO): m/z ¼
5,6-Dichloro-2-(4-methoxy-phenyl)-1H-benzoimidazole (29):
slightly yellow powder; yield: 87%; mp (ꢂC): 200–201; FTIR (KBr,
n (cmꢀ1)): 3019 (C–H), 1611 (C]N), 1493 (C]C), 1261 (C–N),
319.08466 [M + H]+.
4-(5-Chloro-1H-benzoimidazol-2-yl)-N,N-dimethylaniline
(21): light yellow powder; yield: 81%; mp (ꢂC): 233–234; FTIR 1181 (O–CH3), 734 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm):
(KBr, n (cmꢀ1)): 3074 (C–H), 1611 (C]N), 1501 (C]C), 1367 (C– 3.85 (s, 3H, O–CH3), 7.13–8.11 (m, 6H, CHAr); 13C-NMR (125
1
N), 1127 (N–(CH3)2), 740 (C–Cl); H-NMR (500 MHz, DMSO-d6, MHz, DMSO-d6, d ppm): 55.39 (O–CH3), 114.49–161.14 (CHAr);
d ppm): 3.00 (s, 6H, 2 ꢃ CH3), 6.84–7.99 (m, 7H, CHAr); 13C-NMR ESI-HRMS (DMSO): m/z ¼ 293.02468 [M + H]+.
(125 MHz, DMSO-d6, d ppm): 79.11 (CH3), 111.74–153.55 (CHAr);
ESI-HRMS (DMSO): m/z ¼ 272.0949 [M + H]+.
5,6-Dichloro-2-(2,5-dimethoxyphenyl)-1H-benzo[d]imidazole
(30): yellow powder; yield: 82%; mp (ꢂC): 173–174; FTIR (KBr, n
5-Chloro-2-(2-nitro-phenyl)-1H-benzoimidazole (22): slightly (cmꢀ1)): 3086 (C–H), 1620 (C]N), 1483 (C]C), 1307 (C–N), 1169
yellow powder; yield: 67%; mp (ꢂC): 103–104; FTIR (KBr, n (cmꢀ1)): (O–CH3), 731 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm): 3.80
3197 (N–H), 3078 (C–H), 1613 (C]N), 1529 (C]C), 1348 (C–N), (s, 3H, O–CH3), 3.98 (s, 3H, O–CH3), 7.10–7.86 (m, 5H, CHAr),
1
753 (C–Cl); H-NMR (500 MHz, DMSO-d6, d ppm): 7.28–8.04 (m, 12.29 (s, 1H, NH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 55.52,
7H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm): 122.78–148.83 56.25 (O–CH3), 104.18–153.79 (CHAr); ESI-HRMS (DMSO): m/z ¼
(CHAr); ESI-HRMS (DMSO): m/z ¼ 274.0387 [M + H]+.
323.0368 [M + H]+.
5-Chloro-2-(2-triuoromethyl-phenyl)-1H-benzoimidazole
5,6-Dichloro-2-(3,4,5-trimethoxy-phenyl)-1H-benzoimidazole
(23): slightly gray powder; yield: 83%; mp (ꢂC): 173–174; FTIR (31): yellow powder; yield: 90%; mp (ꢂC): 252–253; FTIR (KBr, n
(KBr, n (cmꢀ1)): 3053 (C–H), 1620 (C]N), 1552 (C]C), 1433 (C– (cmꢀ1)): 3290 (N–H), 2990 (C–H), 1672 (C]N), 1412 (C]C),
N), 1129 (C–F), 769 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, 1375 (C–N), 1127 (O–CH3), 763 (C–Cl); 1H-NMR (500 MHz,
d ppm): 7.26–7.95 (m, 7H, CHAr); 13C-NMR (125 MHz, DMSO-d6, DMSO-d6, d ppm): 3.75 (s, 3H, O–CH3), 3.90 (s, 6H, 2 ꢃ O–CH3),
d ppm): 120.41–150.8 (CF3, CHAr); ESI-HRMS (DMSO): m/z ¼ 7.51–7.92 (m, 4H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm):
297.04084 [M + H]+.
4-(5-Chloro-1H-benzoimidazole-2-yl)-2-iodo-6-methoxy-
phenol (24): slightly gray powder; yield: 98%; mp (ꢂC): 204–205;
56.07, 60.13 (O–CH3), 104.18–153.79 (CHAr); ESI-HRMS (DMSO):
m/z ¼ 353.04543 [M + H]+.
4-(5,6-Dichloro-1H-benzo[d]imidazol-2-yl)-N,N-dimethylani-
FTIR (KBr, n (cmꢀ1)): 3453 (OH), 3098 (C–H), 1624 (C]N), 1539 line (32): yellow powder; yield: 73%; mp (ꢂC): 265–266; FTIR (KBr,
(C]C), 1464 (C–N), 1275 (O–CH3), 724 (C–Cl), 593 (C–I); 1H- n (cmꢀ1)): 3422 (NH), 3033 (C–H), 1612 (C]N), 1503 (C]C), 1372
NMR (500 MHz, DMSO-d6, d ppm): 3.93 (s, 3H, O–CH3), 7.21– (C–N), 1208 (N–(CH3)2), 735 (C–Cl); 1H-NMR (500 MHz, DMSO-d6,
8.1 (m, 5H, CHAr), 10.14 (s, 1H, NH); 13C-NMR (125 MHz, DMSO- d ppm): 3.01 (s, 6H, 2 ꢃ CH3), 6.84–7.96 (m, 6H, CHAr); 13C-NMR
d6, d ppm): 56.20 (O–CH3), 84.59 (CAr–I), 110.03–151.57 (CHAr); (125 MHz, DMSO-d6, d ppm): 39.33 (CH3), 111.72–154.68 (CHAr);
ESI-HRMS (DMSO): m/z ¼ 400.9527 [M + H]+.
ESI-HRMS (DMSO): m/z ¼ 306.0568 [M + H]+.
2-(4-(Benzyloxy)phenyl)-5-chloro-1H-benzoimidazole
(25):
5,6-Dichloro-2-(2-nitrophenyl)-1H-benzoimidazole (33): orange
brown powder; yield: 48%, mp (ꢂC): 170–171; FTIR (KBr, n powder; yield: 32%; mp (ꢂC): 218–219; FTIR (KBr, n (cmꢀ1)): 2924
(cmꢀ1)): 3030 (C–H), 1620 (C]N), 1552 (C]C), 1314 (C–N), 1129 (C–H), 1612 (C]N), 1526 (C]C), 1349 (C–N), 784 (C–Cl); 1H-NMR
(C–O–C ether), 769 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, (500 MHz, DMSO-d6, d ppm): 7.80–8.07 (m, 6H, CHAr), 13.38 (s,
d ppm): 5.2 (s, 2H, O–CH2), 7.19–7.60 (m, 12H, CHAr); 13C-NMR 1H, NH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 123.62–150.09
(125 MHz, DMSO-d6, d ppm): 69.37 (O–CH2), 115.22–159.93 (CHAr); ESI-HRMS (DMSO): m/z ¼ 307.9926 [M + H]+.
(CHAr); ESI-HRMS (DMSO): m/z ¼ 335.09497 [M + H]+.
5,6-Dichloro-2-(2-triuoromethyl-phenyl)-1H-benzoimida-
5,6-Dichloro-2-(4-trimethoxy-phenyl)-1H-benzoimidazole (26): zole (34): slightly yellow powder; yield: 52%; mp (ꢂC): 266–267;
brown powder; yield: 85%; mp (ꢂC): 228–229; FTIR (KBr, n (cmꢀ1)): FTIR (KBr, n (cmꢀ1)): 3007 (C–H), 1632 (C]N), 1582 (C]C),
3331 (OH), 3098 (C–H), 1636 (C]N), 1488 (C]C), 1369 (C–N), 752 1314 (C–N), 1134 (C–F), 772 (C–Cl); 1H-NMR (500 MHz, DMSO-
1
(C–Cl); H-NMR (500 MHz, DMSO-d6, d ppm): 7.02–8.07 (m, 6H, d6, d ppm): 7.84–7.98 (m, 6H, CHAr), 13.14 (s, 1H, NH); 13C-NMR
CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm): 112.48–157.64 (125 MHz, DMSO-d6, d ppm): 115.13–150.68 (CF3, CHAr); ESI-
(CHAr); ESI-HRMS (DMSO): m/z ¼ 279.0069 [M + H]+.
HRMS (DMSO): m/z ¼ 331.0035 [M + H]+.
3-(5,6-Dichloro-1H-benzoimidazol-2-yl)phenol (27): white
4-(5,6-Dichloro-1H-benzoimidazol-2-yl)-2-iodo-6-methoxy-
powder; yield: 78%; mp (ꢂC): 260–261, FTIR (KBr, n (cmꢀ1)): phenol (35): isolated by silica gel column chromatography with
3228 (OH), 3032 (C–H), 1650 (C]N), 1469 (C]C), 1232 (C–N), chloroform : methanol (98 : 2) as a slightly yellow powder; yield:
7.18 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm): 6.94–7.82 (m, 85%; mp (ꢂC): 226–227; FTIR (KBr, n (cmꢀ1)): 3454 (OH), 3224
6H, CHAr), 9.76 (s, 1H, OH), 13.13 (s, 1H, NH); 13C-NMR (125 (NH), 2999 (C–H), 1699 (C]N), 1481 (C]C), 1279 (C–N), 1100
MHz, DMSO-d6, d ppm): 113.53–157.77 (CHAr); ESI-HRMS (O–CH3), 731 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm): 3.93
(DMSO): m/z ¼ 279.0088 [M + H]+.
(s, 3H, O–CH3), 7.76–8.11 (m, 4H, CHAr), 10.17 (s, 1H, OH), 13.08
(s, 1H, NH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 56.20 (O–
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CH3), 84.60 (CAr–I), 110.09–152.86 (CHAr); ESI-HRMS (DMSO): yield: 54%; mp (ꢂC): 194–195; FTIR (KBr, n (cmꢀ1)): 3216 (NH),
m/z ¼ 434.91745 [M + H]+.
3062 (C–H), 1600 (C]N), 1537 (C]C), 1286 (C–N); 1H-NMR (500
2-(4-Benzyloxy-phenyl)-5,6-dichloro-1H-benzoimidazole (36): MHz, DMSO-d6, d ppm): 7.57–8.09 (m, 12H, CHAr), 13.44 (s, 1H,
isolated by silica gel column chromatography with chlor- NH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 123.92–195.66
oform : methanol (98 : 2) as a slightly yellow powder; yield: (CHAr); ESI-HRMS (DMSO): m/z ¼ 344.1011 [M + H]+.
68%; mp (ꢂC): 262–263; FTIR (KBr, n (cmꢀ1)): 3291 (N–H), 3031
Phenyl(2-(2-(triuoromethyl)phenyl)-1H-benzo[d]imidazol-5-
(C–H), 1608 (C]N), 1494 (C]C), 1252 (C–N), 1174 (C–O–C yl)methanone (44): recrystallized from ethyl acetate to obtain
ether), 743 (C–Cl); 1H-NMR (500 MHz, DMSO-d6, d ppm): 5.20 (s, a brown solid; yield: 53%; mp (ꢂC): 114–115; FTIR (KBr, n (cmꢀ1)):
2H, O–CH2), 7.22–8.11 (m, 11H, CHAr), 13.06 (s, 1H, NH); 13C- 3279 (NH), 3076 (C–H), 1650 (C]N), 1554 (C]C), 1311 (C–N),
NMR (125 MHz, DMSO-d6, d ppm): 69.39 (O–CH2), 112.34– 1128 (C–F); 1H-NMR (500 MHz, DMSO-d6, d ppm): 7.58–7.99 (m,
160.18 (CHAr); ESI-HRMS (DMSO): m/z ¼ 369.05330 [M + H]+.
12H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm): 122.55–195.65
(CF3, CHAr); ESI-HRMS (DMSO): m/z ¼ 367.1052 [M + H]+.
(2-(4-Hydroxy-3-iodo-5-methoxyphenyl)-1H-benzoimidazol-5-
[2-(2-Hydroxy-phenyl)-1H-benzoimidazol-5-yl]-phenyl-
methanone (37): slightly yellow powder; yield: 75%; mp (ꢂC):
262–263; FTIR (KBr, n (cmꢀ1)): 3296 (N–H), 3025 (C–H), 1613 yl)(phenyl)methanone (45): yellow powder; yield: 98%; mp (ꢂC):
(C]N), 1446 (C]C), 1294 (C–N); 1H-NMR (500 MHz, DMSO-d6, 164–165; FTIR (KBr, n (cmꢀ1)): 3491 (OH), 3256 (NH), 3060 (C–
d ppm): 7.06–8.08 (m, 12H, CHAr), 12.68 (s, 1H, OH), 13.38 (s, H), 1616 (C]N), 1544 (C]C), 1281 (C–N), 1116 (O–CH3), 582
1H, NH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 112.51–195.45 (C–I); 1H-NMR (500 MHz, DMSO-d6, d ppm): 3.95 (s, 3H, O–CH3),
(CHAr); ESI-HRMS (DMSO): m/z ¼ 315.11349 [M + H]+.
7.57–8.15 (m, 10H, CHAr), 10.17 (s, 1H, OH); 13C-NMR (125 MHz,
DMSO-d6, d ppm): 56.21 (O–CH3), 84.65 (C–I), 110.12–195.46
[2-(3-Hydroxy-phenyl)-1H-benzoimidazol-5-yl]-phenyl-
methanone (38): yellow powder; yield: 78%; mp (ꢂC): 254–255; (CHAr); ESI-HRMS (DMSO): m/z ¼ 471.02093 [M + H]+.
FTIR (KBr, n (cmꢀ1)): 3445 (OH), 3256 (NH), 3025 (C–H), 1613
(2-(4-(Benzyloxy)phenyl)-1H-benzoimidazol-5-yl)(phenyl)
(C]N), 1461 (C]C), 1276 (C–N); 1H-NMR (500 MHz, DMSO-d6, methanone (46): slightly yellow powder; yield: 76%; mp (ꢂC):
d ppm): 6.95–7.94 (m, 12H, CHAr), 9.79 (s, 1H, OH); 13C-NMR 198–199; FTIR (KBr, n (cmꢀ1)): 3246 (NH), 3060 (C–H), 1612 (C]
(125 MHz, DMSO-d6, d ppm): 113.53–195.58 (CHAr); ESI-HRMS N), 1495 (C]C), 1248 (C–N), 1179 (C–O–C ether); 1H-NMR (500
(DMSO): m/z ¼ 315.11315 [M + H]+.
MHz, DMSO-d6, d ppm): 5.23 (s, 2H, O–CH2), 7.24–8.17 (m, 2H,
CHAr, 17H), 13.11 (s, NH); 13C-NMR (125 MHz, DMSO-d6,
[2-(4-Hydroxy-phenyl)-1H-benzoimidazol-5-yl]-phenyl-
methanone (39): slightly yellow powder; yield: 84%; mp (ꢂC): d ppm): 69.41 (O–CH2), 115.31–195.53 (CHAr); HRMS (m/z):
295–296; FTIR (KBr, n (cmꢀ1)): 3421 (OH), 3250 (NH), 3063 (C– 405.16112 [M + H]+.
1
H), 1607 (C]N), 1454 (C]C), 1282 (C–N); H-NMR (500 MHz,
DMSO-d6, d ppm): 6.96–8.02 (m, 12H, CHAr), 12.77 (s, 1H, NH);
13C-NMR (125 MHz, DMSO-d6, d ppm): 115.44–195.20 (CHAr);
4.4. General procedure for the synthesis of the nal
ESI-HRMS (DMSO): m/z ¼ 315.11300 [M + H]+.
compounds 47–55
[2-(4-Methoxy-phenyl)-1H-benzoimidazol-5-yl]-phenyl-
The benzimidazoles 37–46 (1 mmol) in 5 mL of methanol were
2-neck round-bottom ask until they were
methanone (40): slightly yellow powder; yield: 84%; mp (ꢂC): 295– stirred in
a
296; FTIR (KBr, n (cmꢀ1)): 3273 (NH), 3061 (C–H), 1640 (C]N), completely dissolved, then 2 mmol of NaBH4 was added. The
1493 (C]C), 1318 (C–N), 1177 (O–CH3); 1H-NMR (500 MHz, reaction mixture was stirred at room temperature for 1 hand
DMSO-d6, d ppm): 3.85 (s, 3H, O–CH3), 7.16–8.16 (m, 12H, CHAr); monitored by TLC using a mixture of ethyl acetate : hexane
13C-NMR (125 MHz, DMSO-d6, d ppm): 55.39 (O–CH3), 114.53– (95 : 5 or 9 : 1 v/v) as eluting solvent. Then, the solvent was
195.53 (CHAr); ESI-HRMS (DMSO): m/z ¼ 329.12917 [M + H]+.
Phenyl-[2-(3,4,5-trimethoxy-phenyl)-1H-benzoimidazol-5-yl]-
evaporated under reduced pressure and the residue was washed
with water to obtain the product. The purication was per-
methanone (41): slightly brown powder; yield: 83%; mp (ꢂC): formed on a column chromatography system or by crystalliza-
220–221; FTIR (KBr, n (cmꢀ1)): 3094 (C–H), 1639 (C]N), 1585 tion from solvents using the desired method, as listed in the
1
(C]C), 1293 (C–N), 1177 (O–CH3); H-NMR (500 MHz, DMSO- information for each product.
d6, d ppm): 3.74 (s, 3H, OCH3), 3.90 (s, 6H, OCH3), 7.53–7.95 (m,
2-(5-(Hydroxy(phenyl)methyl)-1H-benzo[d]imidazol-2-yl)
10H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm): 56.09–60.18 phenol (47): isolated by silica gel column chromatography with
(O–CH3), 104.22–195.54 (CHAr); ESI-HRMS (DMSO): m/z ¼ chloroform : methanol (98 : 2) as a white powder; yield: 56%;
389.15027 [M + H]+.
mp (ꢂC): 161–162; FTIR (KBr, n (cmꢀ1)): 3361 (NH), 3056 (C–H),
1593 (C]N), 1486 (C]C), 1255 (C–N); 1H-NMR (500 MHz,
[2-(4-Dimethylamino-phenyl)-1H-benzoimidazol-5-yl](phenyl)
methanone (42): yellow powder; yield: 98%; mp (ꢂC): 216–217; DMSO-d6, d ppm): 5.86 (s, 1H, C–H), 5.94 (s, 1H, OH), 7.03–8.03
FTIR (KBr, n (cmꢀ1)): 3302 (NH), 3062 (C–H), 1607 (C]N), 1502 (m, 12H, CHAr), 13.12 (s, 2H, NH and OH); 13C-NMR (125 MHz,
(C]C), 1287 (C–N), 1198 (N–(CH3)2); 1H-NMR (500 MHz, DMSO- DMSO-d6, d ppm): 74.41 (C–H), 112.61–157.92 (CHAr); ESI-
d6, d ppm): 3.01 (s, 6H, CH3), 6.86–8.03 (m, 12H, CHAr); 13C-NMR HRMS (DMSO): m/z ¼ 315.1128 [M ꢀ H]ꢀ.
(125 MHz, DMSO-d6, d ppm): 39.33 (CH3), 111.79–195.54 (CHAr);
ESI-HRMS (DMSO): m/z ¼ 342.16067 [M + H]+.
3-(5-(Hydroxy(phenyl)methyl)-1H-benzo[d]imidazol-2-yl)
phenol (48): isolated by silica gel column chromatography with
chloroform : methanol (98 : 2) as a white powder; yield: 72%;
[2-(2-Nitrophenyl)-1H-benzoimidazol-5-yl](phenyl)
methanone (43): isolated by silica gel column chromatography mp (ꢂC): 146–147; FTIR (KBr, n (cmꢀ1)): 3240 (NH), 3061 (C–H),
with chloroform : methanol (98 : 2) as a slightly yellow powder; 1600 (C]N), 1472 (C]C), 1232 (C–N); 1H-NMR (500 MHz,
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DMSO-d6, d ppm): 5.81 (s, 1H, C–H), 5.85 (s, 1H, OH), 6.88–7.55
[2-(4-Benzyloxy-phenyl)-1H-benzoimidazol-5-yl]-phenyl-
(m, 12H, CHAr), 12.65 (s, 1H, NH), 9.66 (s, 1H, OH); 13C-NMR methanol (55): recrystallized from methanol to obtain a slightly
(125 MHz, DMSO-d6, d ppm): 74.46 (C–H), 108.85–157.67 brown solid; yield: 57%; mp (ꢂC): 126–127; FTIR (KBr, n (cmꢀ1)):
(CHAr); ESI-HRMS (DMSO): m/z ¼ 315.1136 [M ꢀ H]ꢀ.
4-(5-(Hydroxy(phenyl)methyl)-1H-benzo[d]imidazol-2-yl)
3353 (NH), 3030 (C–H), 1610 (C]N), 1492 (C]C), 1249 (C–N),
1
1130 (C–O–C ether); H-NMR (500 MHz, DMSO-d6, d ppm): 5.19
phenol (49): isolated by silica gel column chromatography with (s, 2H, O–CH2), 5.82 (s, 1H, OH), 7.20–8.1 (m, 17H); 13C-NMR (125
chloroform : methanol (98 : 2) as a so white powder; yield: MHz, DMSO-d6, d ppm): 69.35 (O–CH2), 74.58 (C–H), 115.13–
45%; mp (ꢂC): 147–148; FTIR (KBr, n (cmꢀ1)):3212 (NH), 3031 159.55 (CHAr); ESI-HRMS (DMSO): m/z ¼ 405.1595 [M ꢀ H]ꢀ.
(C–H), 1612 (C]N), 1466 (C]C), 1278 (C–N); 1H-NMR (500
MHz, DMSO-d6, d ppm): 5.81 (s, 1H, CH), 5.84 (s, 1H, OH), 6.91–
4.5. Cytotoxicity assay
7.97 (m, 12H, CHAr), 12.53 (s, 1H, NH), 9.91 (s, 1H, OH); 13C-
All of the target compounds 15–55 were tested against A549
(human lung adenocarcinoma epithelial cell line), MDA-MB-231
(human breast cancer cell line) and PC3 (human prostate cancer
cell line). Dr Jeong-Hyung Lee (College of Natural Sciences,
Kangwon National University) provided the A549, MDA-MB-231
and PC3 cell lines to the Vietnam Academy of Sciences and
Technology (VAST). The benzimidazole derivatives 15–55 were
dissolved in DMSO to obtain various concentrations. Campto-
thecin and DMSO were used as the reference compound and
blank controls, respectively. All the cancer cells were grown in
RPMI 1640 supplemented with 10% fetal bovine serum, 100 U
mLꢀ1 of penicillin and 100 mg mLꢀ1 of streptomycin in a 5%
CO2 atmosphere for 48 h. Next, the cells were suspended in the
culture medium added to well plates at a concentration of 104
cells/well. Aer 24 h of incubation, the cells were treated with
various concentrations of the test compounds for a period of
72 h. Then, 0.5 mg mLꢀ1 of 3-(4,5-dimethylthiazol-2-yl)-2,5-
diphenyltetrazolium bromide (MTT) was added and incubated
at 37 ꢂC, in 5% CO2 for 4 h followed by the procedure described
by Mosmann.36 The formazan crystals were dissolved by adding
acidied isopropanol and mixed thoroughly to produce the
purple solution. This solution was spectrophotometrically
measured using a Multiskan™ microplate reader (Thermo-
Fisher Scientic) at 570 nm. Each concentration of tested
benzimidazoles was examined in triplicate. The percentage of
cell survival (CS) was calculated using the formula: CS (%) ¼ 100
ꢃ (ODsample ꢀ ODday 0)/(ODDMSO ꢀ ODday 0). The IC50 values (mg
mLꢀ1) were calculated for tested compounds, which had CS
values < 50% in the initial screening, using the non-linear
regression analysis.
NMR (125 MHz, DMSO-d6, d ppm): 74.58 (C–H), 115.60–158.99
(CHAr); ESI-HRMS (DMSO): m/z ¼ 315.1134 [M ꢀ H]ꢀ.
[2-(4-Methoxy-phenyl)-1H-benzoimidazol-5-yl]-phenyl-
methanol (50): recrystallized from methanol to obtain a slightly
yellow solid; yield: 84%; mp (ꢂC):157–158; FTIR (KBr, n (cmꢀ1)):
3410 (NH), 3082 (C–H), 1608 (C]N), 1485 (C]C), 1252 (C–N),
1
1177 (O–CH3); H-NMR (500 MHz, DMSO-d6, d ppm): 5.81 (m,
1H, C–H), 5.83 (m, 1H, OH), 3.83 (s, 3H, O–CH3), 7.10–8.08 (m,
12H, CHAr); 13C-NMR (125 MHz, DMSO-d6, d ppm): 55.29 (O–
CH3), 74.56 (C–H), 114.31–160.51(CHAr); ESI-HRMS (DMSO): m/
z ¼ 329.1291 [M ꢀ H]ꢀ.
Phenyl-[2-(3,4,5-trimethoxy-phenyl)-1H-benzoimidazol-5-yl]-
methanol (51): recrystallized from methanol to obtain a white
solid; mp (ꢂC): 231–232; yield: 92%, FTIR (KBr, n (cmꢀ1)): 3359
(NH), 3094 (C–H), 1632 (C]N), 1593 (C]C), 1263 (C–N), 1131
(O–CH3); 1H-NMR (500 MHz, DMSO-d6, d ppm): 3.79–3.94 (s,
9H, O–CH3), 5.93 (s, 1H, C–H), 7.22–7.88 (m, 10H, CHAr); 13C-
NMR (125 MHz, DMSO-d6, d ppm): 56.62–60.28 (O–CH3), 73.77
(C–H), 106.00–153.40 (CHAr); ESI-HRMS (DMSO): m/z
391.1660 [M + H]+.
¼
[2-(4-Dimethylamino-phenyl)-1H-benzoimidazol-5-yl]-
phenyl-methanol (52): recrystallized from methanol to obtain
a slightly yellow solid; yield: 64%; mp (ꢂC): 269–270; FTIR (KBr, n
(cmꢀ1)): 3312 (NH), 3010 (C–H), 1604 (C]N), 1514 (C]C), 1220
(C–N); 1H-NMR (500 MHz, DMSO-d6, d ppm): 3.06 (s, 6H, CH3),
5.89 (s, 1H, C–H), 6.93–8.15 (m, 12H, CHAr); 13C-NMR (125 MHz,
DMSO-d6, d ppm): 73.83 (CH3), 108.39–153.11 (CHAr); ESI-HRMS
(DMSO): m/z ¼ 342.1743 [M ꢀ H]ꢀ.
Phenyl(2-(2-(triuoromethyl)phenyl)-1H-benzo[d]imidazol-5-
yl)methanol (53): recrystallized from ethyl acetate to obtain
a so white powder; yield: 53%; mp (ꢂC): 154–155; FTIR (KBr, n
(cmꢀ1)): 3357 (NH), 3057 (C–H), 1593 (C]N), 1486 (C]C), 1255
(C–N), 1018 (C–F); 1H-NMR (500 MHz, DMSO-d6, d ppm): 5.84 (s,
1H, C–H), 5.87 (s, 1H, OH), 7.21–7.93 (m, 12H, CHAr), 12.65 (s,
1H, NH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 74.54 (C–H),
(CF3, CHAr); ESI-HRMS (DMSO): m/z ¼ 369.1207 [M + H]+.
4-[5-(Hydroxy-phenyl-methyl)-1H-benzoimidazol-2-yl]-2-iodo-
6-methoxy-phenol (54): recrystallized from methanol to obtain
a so white solid; yield: 98%; mp (ꢂC): 228–229; FTIR (KBr), n/
cmꢀ1: 3389 (NH), 3032 (C–H), 1629 (C]N), 1485 (C]C), 1294
(C–N), 511 (C–I); 1H-NMR (500 MHz, DMSO-d6, d ppm): 3.98 (s,
3H, O–CH3), 5.92 (s, 1H, OH), 7.22–8.33 (m, 10H), 10.81 (s, 1H,
OH); 13C-NMR (125 MHz, DMSO-d6, d ppm): 56.85 (O–CH3),
73.78 (C–H), 84.97 (C–I), 110.54–150.86 (CHAr); ESI-HRMS
(DMSO): m/z ¼ 471.0206 [M ꢀ H]ꢀ.
Conflicts of interest
There are no conicts to declare.
Acknowledgements
This research is funded by the Vietnam National Foundation for
Science and Technology Development (NAFOSTED) under grant
number 104.01-2017.335.
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