N-Phenyl indole derivatives as AT1 antagonists with anti-hypertension activities: Design, synthesis and biological evaluation

Article abstract of DOI:10.1016/j.ejmech.2016.03.021

The design, synthesis, in vitro and in vivo evaluation of 6-substituted benzimidazole with 1, 4-disubsituted or 1, 5-disubsituted indole derivatives as novel angiotensin II receptor antagonists are outlined. Radioligand binding assays showed that several 6-substituted benzimidazole derivatives displayed high affinities binding to the angiotensin II type 1 receptor at the same order of magnitude to telmisartan. The biological evaluation on spontaneously hypertensive rats showed that 2-[4-[[2-n-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-1H-indol-1-yl]benzoic acid, 1c, could cause significant decrease on MBP in a dose dependent manner. Its maximal response lowered 53 mmHg of MBP at 5 mg/kg and 64 mmHg of MBP at 10 mg/kg after oral administration, and the significant antihypertensive effect lasted beyond 24 h, which was better than both losartan and telmisartan. A study designed to determine acute toxicity showed that 1c had low acute toxicity with no significant changes in the weight and no obvious untoward reactions. The encouraging results make 1c an effective and durable anti-hypertension drug candidate and deserve further investigation for therapeutic application.

Full text of DOI:10.1016/j.ejmech.2016.03.021

Accepted Manuscript
N-Phenyl Indole Derivatives as AT Antagonists with Anti-Hypertension Activities:
1
Design, Synthesis and Biological Evaluation
Weibo Zhu, Xiaolu Bao, He Ren, Yajing Da, Dan Wu, Fuming Li, Yijia Yan, Li Wang,
Zhilong Chen
PII:
S0223-5234(16)30195-7
10.1016/j.ejmech.2016.03.021
EJMECH 8446
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 17 November 2015
Revised Date: 7 March 2016
Accepted Date: 9 March 2016
Please cite this article as: W. Zhu, X. Bao, H. Ren, Y. Da, D. Wu, F. Li, Y. Yan, L. Wang, Z.
Chen, N-Phenyl Indole Derivatives as AT Antagonists with Anti-Hypertension Activities: Design,
1
Synthesis and Biological Evaluation, European Journal of Medicinal Chemistry (2016), doi: 10.1016/
j.ejmech.2016.03.021.
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ACCEPTED MANUSCRIPT
Graphical abstract

ACCEPTED MANUSCRIPT
N-Phenyl Indole Derivatives as AT1 Antagonists with
Anti-Hypertension Activities: Design, Synthesis and Biological
Evaluation
Weibo Zhu1, Xiaolu Bao2, He Ren1, Yajing Da1, Dan Wu1, Fuming Li1ꢀYijia Yan3, Li
Wang1, Zhilong Chen1*
1Department of Pharmaceutical Science and Technology, College of Chemistry and
Biology, Donghua University, Shanghai 201600, China
2State Key Laboratory for Modification of Chemical Fibers and Polymer Materials,
College of Materials Science and Engineering, Donghua University, Shanghai 201620,
P.R.China
3Shanghai Xianhui Pharmaceutical Co., Ltd, Shanghai 200433, China
*Corresponding Author: Zhi-long Chen, Department of Pharmaceutical Science &
Technology, College of Chemistry and Biology, Donghua University, 2999 North
Renmin Road, Shanghai, 201620, China.
Phone: 86-21-67792654, Email: zlchen1967@yahoo.com
Abbreviations used
RAS, renin angiotensin system; AII, angiotensin II; ARBs, angiotensin II receptor
blockers;
FDA,
US
Food
and
Drug
Administration;
DMAP,
4-(dimethylamino)pyridine; NMR, nuclear magnetic resonance; HRMS,
high-resolution mass spectrometry; ESI, electrospray ionization; MS, mass
spectrometry; DMSO, dimethylsulfoxide; DMF, N,N-dimethylformamide; THF,
tetrahydrofuran; AIBN, azodiisobutyronitrile; NBS, bromosuccinimide; VSMC,
vascular Smooth Muscle Cell; IC50, half maximal inhibitory concentration; SHR,
spontaneously hypertensive rats; MBP, mean blood pressure; DMEM, Dulbecco’s
Modified Eagle’s Medium; FBS, fetal bovine serum.

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Abstract
The design, synthesis, in vitro and in vivo evaluation of 6- substituted benzimidazole
with 1, 4-disubsituted or 1, 5-disubsituted indole derivatives as novel angiotensin II
receptor antagonists are outlined. Radioligand binding assays showed that several 6-
substituted benzimidazole derivatives displayed high affinities binding to the
angiotensin II type 1 receptor at the same order of magnitude to telmisartan. The
biological evaluation on spontaneously hypertensive rats showed that
2-[4-[[2-n-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl
]-1H-indol-1-yl]benzoic acid, 1c, could cause significant decrease on MBP in a dose
dependent manner. Its maximal response lowered 53 mmHg of MBP at 5 mg/kg and
64 mmHg of MBP at 10 mg/kg after oral administration, and the significant
antihypertensive effect lasted beyond 24 h, which was better than both losartan and
telmisartan. A study designed to determine acute toxicity showed that 1c had low
acute toxicity with no significant changes in the weight and no obvious untoward
reactions. The encouraging results make 1c an effective and durable anti-hypertension
drug candidate and deserve further investigation for therapeutic application.
Keywords: AT1 Antagonists, 6- substituted benzimidazole, anti-hypertension, in vitro
activity, in vivo activity, acute toxicity

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1. Introduction
Hypertension is recognized as one of the leading risk factors for human
morbidity and mortality. On a worldwide basis hypertension has been ranked third as
a cause of disability adjusted life years [1]. The renin angiotensin system (RAS) plays
an important role in the regulation of blood pressure and electrolyte/fluid homeostasis
[2]. This system consists of a cascade of enzymatic reactions, leading to the production
[3]
of angiotensin II (AII)
which is the final site of action in the RAS. RAS elicits
multiple biological effects such as increase in blood pressure, vascular contraction and
modulation of central drinking behavior [4]. It is widely accepted that angiotensin II
type 1 (AT1) receptor accounts for a majority of the cardiovascular effects evoked by
[5]
AII. Losartan (Fig.1) is the first non-peptide orally active AT1 receptor antagonist
.
Since then, a series of angiotensin II receptor blockers (ARBs) were used in clinics
for the treatment of hypertension. These ARBs share a common mechanism of action,
which block the effects of angiotensin II by selectively antagonizing the AT1 receptor
[6]. Recent investigations suggest that ARBs are also useful for treating congestive
heart failure (CHF) and also reducing the cardiovascular disease associated with
cardiac and vascular remodeling.
Fig.1 Non-peptide angiotensin II receptor antagonists.
Indole derivatives have been extensively studied as a pharmacophore group
because of their wide potential bioactivities [7-9]. Poss [10] developed a series of AII
receptor antagonists that contain N-phenyl indole group. BMS-180560 (Fig.1) was
the most potent analog identified in the series, which could cause sustained reduction
in blood pressure for over 72 h. In our preliminary work [11], we used N- phenyl indole
group instead of biphenyl system as a strategy to generate new potent and long lasting

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antihypertensive agents.
Telmisartan (Fig.1) is the most recently marketed drug as an AT1 receptor
antagonist. It has a 24-hour antihypertensive efficacy and causes reductions in clinic
BP similar to those newer agents in its class. [12]
Herein, we report on the design, synthesis, and biological activity of a novel
class of potent non-peptide AT1 receptor antagonists, in which the biphenyl moiety of
telmisartan is replaced with N- phenyl indole group. Some of these designed
compounds have been authorized as Chinese patent [13]. The dominant conformations
of the designed compounds 1c, 2e, and 5b as well as telmisartan were presented in
Fig.2 using computer software Spartan 10. According to their overlay conformations,
it is indicated that they align well with each other.
Fig.2 Energy-minimized conformations of 1c, 2e, 5b, telmisartan and their overlay
conformations.
2. Results and discussion
2.1 Chemistry
We developed a general and useful synthesis of indole derivatives with a variety
of substituents. In short, this procedure comprised of two coupling parts, one gave the
bromide and the other gave the heterocyclic amine. We initially synthesized the halide
component which was coupled to the heterocyclic fragment to construct the target
molecule.
The synthesis of indole bromides (11a-b) is outlined in Scheme 1. N-benzoyl
protected methylindoles (10a-b) were prepared from the corresponding commercially
available methylindoles (9a-b) by reacting with benzoyl chloride. They were then

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brominated with NBS to give the indole bromides (11a-b). The bromides 11a-b were
then used in subsequent synthetic transformations.
Scheme 1. (a) benzoyl chloride, Et3N, DMAP, DCM; (b) NBS, AIBN, CCl4, reflux; (c)
benzoyl chloride, Et3N, DMAP, DCM; (d) NBS, AIBN, CCl4, reflux.
The synthesis of 1a−e and 2a-e is outlined in Schemes 2 and 3. The substituted
benzimidazoles (7a-e) were prepared from the commercially available methyl
4-amino-3-methylbenzoate (6) by four steps. The phenylamine (6) was reacted with
alkyl acyl chloride in DCM, nitrated with concentrated nitric acid, hydrogenated with
hydrogen and finally cyclized in acetic acid to produce benzimidazoles (7a-e). The
obtained benzimidazoles were converted to the corresponding bis-benzimidazole
derivatives (8a-e) by reacting with 2M NaOH, and cyclized with N-methylbenzene- 1,
2-diamine in the presence of polyphosphorous acid. Each bis-benzimidazole
derivatives (8a-e) was reacted with an indole bromide (11a or 11b) to generate
bis-benzimidazole indole derivatives (12a-e, 15a-e). After deprotecting the benzoyl
group, the obtained indole derivatives were alkylated with 2-fluorobenzonitrile to
produce benzonitrile (14a-e, 17a-e); which were then hydrolyzed with 5 M NaOH in
methanol to yield the final products, 1a−e and 2a-e.

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Scheme 2. Reagents and reaction conditions: (a) RCOCl, Et3N, DCM; (b) H2SO4, HNO3, -5 oC;
(c) Raney Ni, H2; (d) CH3COOH, reflux; (e) NaOH, MeOH, reflux; (f) PPA,
N-methylbenzene-1,2-diamine, 150 oC; (g)NaH, THF, 50 oC; (h) NaOH, H2O, CH3OH, reflux; (i)
K2CO3, 2-fluorobenzonitrile, DMF, reflux; (j) NaOH, H2O, CH3OH, reflux.

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N
N
N
R
N
N
N
N
N
R
a
b
8a-e
+ 11b
N
O
N
H
16a-e
N
15a-e
N
N
N
N
N
N
R
R
N
d
c
N
N
COOH
CN
2a-e
17a-e
2a: R=Me
2b: R=Et
2c: R=n-Pr
2d: R=n-Bu
2e: R=n-pentyl
Scheme 3. Reagents and reaction conditions: (a) NaH, THF, 50 oC; (b) NaOH, H2O, CH3OH,
reflux; (c) K2CO3, 2-fluorobenzonitrile, DMF, reflux; (d) NaOH, H2O, CH3OH, reflux.
The tetrazole 3 was synthesized by treating benzonitrile 14c with sodium azide and
chlorotributylstannane at 160 oC in DMF, as shown in Scheme 4.
Scheme 4. Reagents and reaction conditions: (a) Bu3SnCl, NaN3, DMF, reflux.
The preparation of 4a-c and 5a-c is outlined in Schemes 5 and 6, which is similar
to 1a-e and 2a-e. The commercially available 2-bromopyridine (18) was reacted with
3-methylphenylmagnesium bromide (19) to give the coupling product, 20; which was
then nitrated with concentrated nitric acid and then hydrogenated to produce amino
compound. Acylation of the resulting amino compound with alkyl acyl chloride in
DCM, nitration, hydrogenation, and formation of the benzimidazole were performed

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using the same procedures as described for the preparation of 8a-e to give the
benzimidazole derivatives (21a-c). Coupling the obtained benzimidazole derivatives
with indole bromide (11a-b), deprotection, alkylation, and hydrolyzation of the
benzonitrile were also performed using the same procedures as described for the
preparation of 1a-e and 2a-e to give the final products, 4a-c and 5a-c.
N
b,c,d,e,f,g
a
R
+
N
H
Mg
Br
N
Br
N
N
21a: R=Et
22b: R=n-Pr
23c: R=n-Bu
18
19
20
21a-c
N
N
N
R
R
N
N
N
k
h, i, j
N
N
CN
COOH
4a: R=Et
4b: R=n-Pr
4c: R=n-Bu
4a-c
22a-c
Scheme 5. Reagents and reaction conditions: (a) FeBr3, N2, t-BuOMe; (b) H2SO4, HNO3, -5oC;
o
(c) Raney Ni, H2; (d) RCOCl, Et3N, DCM; (e) H2SO4, HNO3, -5 C; (f) Raney Ni, H2; (g)
o
CH3COOH, reflux; (h) 11a, NaH, THF, 50 C; (i) NaOH, H2O, CH3OH, reflux; (j) K2CO3,
2-fluorobenzonitrile, DMF, reflux; (k) NaOH, H2O, CH3OH, reflux.
o
Scheme 6. Reagents and reaction conditions: (a) 11b, NaH, THF, 50 C; (b) NaOH, H2O,
CH3OH, reflux; (c) K2CO3, 2-fluorobenzonitrile, DMF, reflux; (d) NaOH, H2O, CH3OH, reflux.
The aforementioned synthetic route was not amenable to scale-up, where the
preparation of heterocyclic amine could be in high yield, but the bromide fragment
suffered from low yields coupled with complex manipulative procedures. For this

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reason, we developed a new method for the preparation of bromide fragment: Indole
-4 -formaldehyde (or indole -5- formaldehyde) was selected as the starting material to
avoid large amounts of by-products compared to the benzylic methyl group. Here, we
used indole -4 -formaldehyde to present the modified approach. The preparation of 1,
5- disubstituted indole compounds was in a similar manner.
The modified procedure is shown in Scheme 7. Indole-4-formaldehyde was
coupled with 2- fluorobenzonitrile to give benzonitrile compound (26); which was
then hydrogenated and brominated to generate the halide component (28). After
substitution with bis-benzimidazole derivatives (8a-e), the obtained benzonitrile
compounds (14a-e) were hydrolyzed to produce carboxylic acid compounds (1a-e).
The total yield was 36.3%, while the former method was 32.9%.
Scheme 7. Reagents and reaction conditions: (a) K2CO3, DMF, reflux; (b) NaBH4, EtOH, THF; (c)
HBr/AcOH (35%), DCM; (d) K2CO3, DMF; (de) NaOH, H2O, CH3OH, reflux.
Another issue was that, after coupling with the heterocyclic amines, the
synthesized benzonitrile compounds showed low solubility in 50% methanol in water
with a low hydrolysis rate. To improve the reaction, we adjusted the order of the
reactions (Scheme 8). The benzyl alcohol compound (27) was first hydrolyzed to give
benzoic acid compound (29), which was then esterified to form ethyl benzoate
compound (30). After brominating the ester, the bromide fragment was reacted with
the heterocyclic amines in DMF. Without separation after reaction, 1-2 mL 2 M

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NaOH aqueous solution was added to the reaction mixture and stirred for another 1-2
h at room temperature which eliminated the ester to give the final products. This
method skipped intermediate compounds (14a-e, 17a-e, 22a-c, 23a-c) with low
solubility in 50% methanol in water and also shortened the reaction time.
Scheme 8. Reagents and reaction conditions: (a) NaOH, ethylene glycol, H2O, reflux; (b)
KHCO3, bromoethane, DMF; (c) HBr/AcOH (35%), DCM; (d) NaH, DMF; (e) NaOH, H2O.
2.2 Biological evaluation
2.2.1 Radioligand binding assays
The affinity of the prepared compounds, losartan and telmisartan for the AT1
receptor was determined using vascular smooth muscle cells in a competition
radioligand binding assay with [125I]-AII and expressed as IC50 values. The results
(Tables 1 and 2) showed that most of the tested compounds displayed high affinity for
AT1 receptor. The specific binding of 125I-AII was inhibited in
a
concentration-dependent manner by 1a-e, 2a-e, 3, 4a-c, 5a-c, losartan and telmisartan,
as exemplified by 1c in Fig. 3.
The results showed that all the prepared compounds had better receptor affinity in
vitro than losartan while compounds 1b-d, 2c-e, 3, and 5a were better than
telmisartan.

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Fig.3 Inhibitory effects of 1c, losartan and telmisartan on the specific binding of 125I-Ang II to
AT1 receptors in VSMCS.
Table 1. IC50 and Ki value of the tested compounds (1a-e, 2a-e, 3, losartan, telmisartan)
Compound
R
Mp(oC)
Formula
IC50 ± SEM
(nM)
Ki (nM)
1a
Me
Et
263-265
262-264
264-266
264-267
265-267
262-264
263-265
264-266
265-267
266-269
212-214
183-185
C33H27N5O2
C34H29N5O2
C35H31N5O2
C36H33N5O2
C37H35N5O2
C33H27N5O2
C34H29N5O2
C35H31N5O2
C36H33N5O2
C37H35N5O2
C35H31N9
7.46±0.16
1.83±0.19
0.36±0.18
2.61±0.26
11.30±5.89
8.25±3.72
13.06±2.19
3.76±1.19
2.23±0.54
0.92±0.03
3.39±1.06
4.66±0.78
1.14±0.23
0.23±0.17
1.63±0.19
7.06±4.14
5.13±1.25
8.16±1.37
2.35±0.91
1.39±0.34
0.67±0.15
2.12±0.65
1b
1c
n-Pr
n-Bu
n-pentyl
Me
1d
1e
2a
2b
Et
2c
n-Pr
n-Bu
n-pentyl
/
2d
2e
3
Losartan
/
C22H23ClN6O
20.09±0.11
3.80±0.22
13.06±0.07
2.75±0.17
Telmisartan
/
260-262
C33H30N4O2

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Table 2. IC50 and Ki value of the tested compounds (4a-c, 5a-c, losartan, telmisartan)
Compound
R
Mp(oC)
242-244
Formula
IC50 ± SEM (nM)
3.55 ±0.11
4.70±0.29
Ki (nM)
2.57±0.09
3.40±0.21
4.08±0.15
0.84±0.25
2.37±0.13
4.06±0.14
13.06±0.07
2.75±0.17
4a
Et
C31H26N4O2
4b
n-Pr
n-Bu
Et
243-245
245-247
241-243
242-244
243-245
C32H28N4O2
C33H30N4O2
C31H26N4O2
C32H28N4O2
C33H30N4O2
4c
5a
5.63±0.24
1.17±0.32
5b
n-Pr
n-Bu
/
3.27±0.18
5c
5.61±0.24
Losartan
Telmisartan
183.5-184.5 C22H23ClN6O
260-262 C33H30N4O2
20.09±0.11
3.80±0.22
/
2.2.2 In vivo antihypertensive activities
In spontaneously hypertensive rats (SHR), the effects of prepared compounds (5,
10 mg/kg), losartan (10 mg/kg) and telmisartan (10 mg/kg) on the mean blood
pressure (MBP) in vivo after oral administration were shown in Fig.4. It was observed
that under the dose of 5 mg/kg or 10 mg/kg, 1a, 1d, 1e, 2b and 5c could not decrease
the blood pressure significantly compared with the negative control group. 1b, 2c, 2d,
3, 4a, 4b and 4c could decrease blood pressure but with no significant improvement.
2a, 2e, 5a and 5b had similar anti-hypertensive effects with losartan at the dose of 10
mg/kg, but weaker than telmisartan. The maximal reduction of MBP of 2a, 2e, 5a, 5b
and losartan were 44 mmHg, 53 mmHg, 42 mmHg, 43mmHg and 43 mmHg,
respectively. 1c caused significant decrease on MBP in a dose dependent manner. The
maximal response of 1c (10 mg/kg ) was observed at 6 h after dosing, it lowered by
64 mmHg of MBP, and the significant antihypertensive was sustained for over 24 h.

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Even at dose of 5 mg/kg, 1c had similar anti-hypertensive effects with telmisartan at
the dose of 10 mg/kg. Compared with telmisartan, 1c had a higher and longer
antihypertensive effect at the same dose.

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Fig.4 Effects of prepared compounds, losartan and telmisartan on mean blood pressure (MBP) in
spontaneously hypertensive rats. *, ** Significant difference from the control, pꢁ0.05 and pꢁ0.01,
respectively.
2.2.3 Acute toxicity test
The acute toxicity assay showed that compound 1c had low acute toxicity with
the LD50 value of 526.203 mg/kg and the 95% confidence interval was
466.342-593.656 mg/kg which was more than 260-fold of suggested therapeutic dose
(Table 3). There were no physiological abnormalities detected within in mice after
administered with dose of 307.2 and 384 mg/kg respectively. However,
hyperthyroidism appeared 6 h later after administration in other groups, while some
mice died. There were no significant changes in the weight of the surviving mice after
2 weeks of observation and no obvious untoward reactions.

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Table 3. The lethal dose (LD50) of compound 1c determined by acute toxicity
test.
Dose
(mg/kg)
Log
(dose)
Mortality
(%)
LD50 (mg/kg) and 95%
confidence interval
(mg/kg)
307.20
384.00
480.00
600.00
750.00
937.00
2.49
2.58
2.68
2.78
2.88
2.97
0
10
50
60
90
100
526.203
（466.342—593.656）
3. Conclusions
A series of 6- substituted benzimidazole with 1, 4-disubsituted or 1, 5-disubsituted
indole derivatives were designed and synthesized. Their in vitro AII antagonistic
activities as well as in vivo antihypertensive activities were evaluated. 6-Pyridine
benzimidazole derivatives (4a-c, 5a-c) showed higher receptor affinity with lower
antihypertensive activities in vivo than losartan. Bis-benzimidazole derivatives (1a-e,
2a-e, 3) exhibited a high AT1 receptor affinity, while their antihypertensive activities
in vivo varied from each other. Among them, 1c, 2a, 2e, 3 exhibited high
antihypertensive activities in vivo; but others showed no significant activities. The
carboxylic acid 1c (AT1 IC50=0.36 nM) was found to be the most potent AT1
antagonist in the N-phenyl indole derivatives with a long lasting antihypertensive
activity. The study of acute toxicity suggested that 1c also had a positive safety profile.
In addition, a simple method for synthesis of 1, 4-disubstituted and 1, 5-disubstituted
indole compounds was developed. The process was also optimized for large scale
production at a kilogram level.
In summary, 1c was the most promising drug candidate among these synthesized
compounds. It could be considered as a novel effective anti-hypertension drug
candidate with long-lasting blood pressure-lowering effect, and deserve further
investigation.

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4. Experimental
4.1 Chemistry
All chemical reagents were purchased from commercial suppliers and were used
without further purification. Yields of the purified products were not optimized.
Melting points (MP) were measured on an electro thermal melting point apparatus and
1
were not corrected. H NMR spectra were measured on a Bruker 400 MHz
spectrometer using TMS (Me4Si) as internal standard. ESI-MS spectra were recorded
on a Micromass triple quadrupole mass spectrometer. Column chromatography was
performed using silica gel H (300- 400). All final compounds were purified by
preparative HPLC on reverse phase column using 4.6*150 mm, WondaSil C18-WR
5ꢀm column under 0.1% formic acid with 2 mmol ammonium acetate in 100% high
purity water. All final compounds had a purity level greater than 95% based upon
HPLC, and 1H NMR.
4.1.1 General procedure for the synthesis of 7a-e:
Methyl 4-amino-3-methylbenzoate (6) (10.00 g, 60.58 mmol) was acylated with
alkyl acyl chloride (90.87 mmol) and triethylamine (16.79 mL, 121.16 mmol) in
DCM at 0 oC. The resulting amide was reacted with fuming nitric acid in sulfuric acid
o
(60%) at -20 C. The resulting nitro-compound was reduced with hydrogen (5 bar)
o
and Raney Ni (0.79 g, 13.34 mmol) in methanol at 100 C. The resulting amino
compound was dissolved in glacial acetic acid and heated under reflux for 1 h. After
evaporation of the acetic acid, water was added and the pH was adjusted to 9 by
addition of concentrated ammonia. This solution was extracted with ethyl acetate (3×
100 mL). The combined organic layers were washed with aqueous NaHCO3 solution
and dried over MgSO4. After filtration, the solvent was removed under reduced
pressure.
4.1.1.1 Methyl 2, 4-dimethyl-1H-benzo[d]imidazole-6-carboxylate (7a).
The above general procedure was followed by using acetyl chloride. The residue
1
was purified by CC to give the product as yellow oil. Yield: 49.5%. H NMR(400
MHz, CDCl3) δ: 8.10 (s, 1H), 7.96 (s, 1H), 3.96 (s, 3H), 2.70 (s, 3H), 2.63 (s, 3H).
MS(ESI): [M+H]+ calcd 205.1; found 205.1.
4.1.1.2 Methyl 2-ethyl-4-methyl-1H-benzo[d]imidazole-6-carboxylate (7b).
The above general procedure was followed by using propionyl chloride. The
1
residue was purified by CC to give the product as yellow oil. Yield: 56.2%. H
NMR(400 MHz, CDCl3) δ: 8.15(s, 1H), 7.88 (s, 1H), 3.93(s, 3H), 2.81(q, 2H), 2.63(s,
3H), 1.45(t, 3H). MS(ESI): [M+H]+ calcd 219.1; found 219.1.

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4.1.1.3 Methyl 2-propyl-4-methyl-1H-benzo[d]imidazole-6-carboxylate (7c). [14]
The above general procedure was followed by using butyryl chloride. The
1
residue was purified by CC to give the product as yellow oil. Yield: 41.4%. H
NMR(400 MHz, CDCl3) δ: 8.1 (s, 1H), 7.8 (s, 1H), 3.9 (s, 3H), 2.9 (t, 2H), 2.4 (s, 3H),
1.9 (m, 2H), 1.0 (t, 3H). MS(ESI): [M+H]+ calcd 233.1; found 233.1.
4.1.1.4 Methyl 2-butyl-4-methyl-1H-benzo[d]imidazole-6-carboxylate (7d).
The above general procedure was followed by using valeryl chloride. The
1
residue was purified by CC to give the product as yellow oil. Yield: 47.1%. H
NMR(400 MHz, CDCl3) δ: 8.23(s, 1H), 7.69(s, 1H), 3.93(s, 3H), 2.75(t, 2H), 2.51(s,
3H), 1.75(m, 2H), 1.35(m, 2H), 0.94(t, 3H). MS(ESI): [M+H]+ calcd 247.1; found
247.1.
4.1.1.5 Methyl 2-pentyl-4-methyl-1H-benzo[d]imidazole-6-carboxylate (7e).
The above general procedure was followed by using hexanoyl chloride. The
1
residue was purified by CC to give the product as yellow oil. Yield: 52.5%. H
NMR(400 MHz, CDCl3) δ:8.26(s, 1H), 7.63(s, 1H), 3.89(s, 3H), 2.76(t, 2H), 2.53(s,
3H), 1.73(m, 2H), 1.37(m, 4H), 0.94(t, 3H). MS(ESI): [M+H]+ calcd 261.2; found
261.3.
4.1.2 General procedure for the synthesis of 8a-e:
A solution of an appropriate ester (25.01 mmol) in methanol (25 mL) was added
to a solution of NaOH (2.0 g) in water (25 mL), and the mixture was heated under
reflux for 2 h. After evaporation of methanol, the pH was adjusted to 4-5 by addition
of aqueous citric acid. The precipitated solid was filtered, washed with ethanol and
dried to yield carboxylic acid. The resulting compound was dissolved in
polyphosphoric acid (10 mL) at 150 oC. N-methyl-o-phenylenediamine
dihydrochloride (3.65 g, 18.8 mmol) was added to the mixture for 4 times in 4 h.
After stirring at 150 oC for 10 h, the mixture was cooled and then poured into ice
water (30 mL). The pH was adjusted to 10 by addition of concentrated ammonia (ice
cooling). The precipitated solid was filtered off, dried, and boiled in ethyl acetate (300
mL). After cooling, the precipitated solid was filtered off, washed with diethyl ether,
and dried to give the product as white solid.
4.1.2.1 2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimidazole (8a).
8a was prepared by following the above general procedure. Yield: 55.2%. MP:
o
190-192 C. 1H NMR (400 MHz, CDCl3) δ: 7.79(t, 2H), 7.35-7.44(m, 4H), 3.86(s,
3H), 2.80(s, 3H), 2.61(s, 3H). MS(ESI): [M+H]+ calcd 277.2; found 277.2.
4.1.2.2 2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole (8b).

ACCEPTED MANUSCRIPT
8b was prepared by following the above general procedure. Yield: 50.7%. MP:
191-194 oC. 1H NMR (400 MHz, CDCl3) δ:7.81(t, 2H), 7.31-7.45(m, 4H), 3.91(s, 3H),
2.90(q, 2H), 2.58(s, 3H), 1.33(t, 3H). MS(ESI): [M+H]+ calcd 291.2; found 291.1.
4.1.2.3 2-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole (8c).14
8c was prepared by following the above general procedure. Yield: 60.1%. MP:
o
193-195 C. 1H NMR (400 MHz, CDCl3) δ: 7.82 (d, 2H), 7.30-7.53 (m, 4H), 3.95 (s,
3H), 2.89 (t, 2H), 2.61 (s, 3H), 1.79 (m, 2H), 0.91 (t, 3H). MS(ESI): [M+H]+ calcd
305.2; found 305.1.
4.1.2.4 2-butyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole (8d).
8d was prepared by following the above general procedure. Yield: 48.2%. MP:
194-196 oC. 1H NMR (400 MHz, CDCl3) δ:7.82(t, 2H), 7.30-7.54(m, 4H), 3.96(s, 3H),
2.96(t, 2H), 2.57(s, 3H), 1.82(m, 2H), 1.45(m, 2H), 0.92(t, 3H). MS(ESI): [M+H]+
calcd 319.2; found 319.2.
4.1.2.5 2-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole (8e).
8e was prepared by following the above general procedure. Yield: 49.8%. MP:
o
196-198 C. 1H NMR (400 MHz, CDCl3) δ: 7.77(t, 2H), 7.27-7.44(m, 4H), 3.88(s,
3H), 2.72(t, 2H), 2.53(s, 3H), 1.66(m, 2H), 1.15-1.28(m, 4H), 0.75(t, 3H). MS(ESI):
[M+H]+ calcd 333.2; found 333.2.
4.1.3 N-Benzoyl-4-methylindol (10a).
10a was prepared by following the general procedure described by Dhanoa. [15]
4.1.3 N-Benzoyl-5-methylindol (10b).
10b was prepared by following the general procedure described by Dhanoa. [15]
4.1.4 (4-(bromomethyl)-1H-indol-1-yl)(phenyl)methanone(11a).
11a was prepared by following the general procedure described by Dhanoa. [15]
4.1.5 (5-(bromomethyl)-1H-indol-1-yl)(phenyl)methanone(11b).
11b was prepared by following the general procedure described by Dhanoa. [15]
4.1.6 General procedure for the synthesis of 12a-e, 15a-e:
A solution of an appropriate benzimidazole (5.03 mmol) and NaH (0.12 g, 5.53
o
mmol, 60 %) in 100 mL anhydrous THF was stirred for 30 min at 50 C. After
cooling to rt, a mixture of an appropriate bromide (6.04 mmol) in anhydrous THF (50
o
mL) was added dropwise to the solution. The solution was stirred for 3 h at 50 C.
Then the resulting mixture was poured into 30 mL ice water, and extracted with ethyl
acetate (50 mL×3). The combined organic layer was dried over MgSO4. After
filtration, the solvent was removed under reduced pressure and the residue was
purified by CC to give the product as white solid.

ACCEPTED MANUSCRIPT
4.1.6.1
[4-[[2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-1H-i
ndol-1-yl](phenyl)methanone (12a).
12a was prepared by following the above general procedure. Yield: 91.6%. MP:
o
1
213-215 C. H NMR (400 MHz, CDCl3): δ 8.33(d, 1H), 7.28-7.73(m, 13H), 6.76(d,
1H), 6.54(d, 1H), 5.68(s, 2H), 3.80(s, 3H), 2.76(s, 3H), 2.67(s, 3H). MS(ESI):
[M+H]+ calcd 510.2; found 510.3.
4.1.6.2
[4-[[2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-1
H-indol-1-yl](phenyl)methanone (12b).
12b was prepared by following the above general procedure. Yield: 88.3%. MP:
o
1
215-217 C. H NMR (400 MHz, CDCl3): δ 8.35(d, 1H), 7.31-7.86(m, 13H), 6.75(d,
1H), 6.53(d, 1H), 5.68(s, 2H), 3.86(s, 3H), 2.88(q, 2H), 2.76(s, 3H), 1.27(t, 3H).
MS(ESI): [M+H]+ calcd 524.2; found 524.3.
4.1.6.3
[4-[[2-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]
-1H-indol-1-yl](phenyl)methanone (12c).
12c was prepared by following the above general procedure. Yield: 91.9%. MP:
o
1
214-216 C. H NMR(400 MHz, CDCl3): δ 8.34(d, 1H), 8.04(s, 1H), 7.27-7.73(m,
12H), 6.74(d, 1H), 6.52(d, 1H), 5.67(s, 2H), 3.89(s, 3H), 2.89(t, 2H), 2.78(s, 3H),
1.85(m, 2H), 1.02(t, 3H). MS(ESI): [M+H]+ calcd 538.3; found 538.3.
4.1.6.4
[4-[[2-butyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-
1H-indol-1-yl](phenyl)methanone (12d).
12d was prepared by following the above general procedure. Yield: 90.9%. MP:
o
1
217-219 C. H NMR(400 MHz, CDCl3): δ 8.32(d, 1H), 7.31-7.80(m, 13H), 6.75(d,
1H), 6.55(d, 1H), 5.64(s, 2H), 3.84(s, 3H), 2.83(t, 2H), 2.71(s, 3H), 1.73(m, 2H),
1.43(m, 2H), 0.97(t, 3H). MS(ESI): [M+H]+ calcd 552.3; found 552.3.
4.1.6.5
[4-[[2-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]
-1H-indol-1-yl](phenyl)methanone (12e).
12e was prepared by following the above general procedure. Yield: 91.7%. MP:
o
1
218-220 C. H NMR(400 MHz, CDCl3): δ 8.35(d, 1H), 7.82(s, 1H), 7.74(d, 2H),
7.60(t, 2H), 7.54(t, 2H), 7.43(s, 2H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d, 1H), 6.73(d,
1H), 5.68(s, 2H), 3.74(s, 3H), 2.45(t, 2H), 2.36(s, 3H), 1.73(m, 2H), 1.37(m, 4H),

ACCEPTED MANUSCRIPT
0.94(t, 3H). MS(ESI): [M+H]+ calcd 566.3; found 566.3.
4.1.6.6
[5-[[2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-1H-i
ndol-1-yl](phenyl)methanone (15a).
15a was prepared by following the above general procedure. Yield: 93.8%. MP:
o
1
213-215 C. H NMR(400 MHz, CDCl3): δ 8.13(d, 1H), 7.73(s, 1H), 7.53(d, 2H),
7.38(t, 2H), 7.23(t, 2H), 7.13(s, 2H), 7.11(d, 2H), 7.08(t, 2H), 6.80(d, 1H), 6.59(d,
1H), 5.35(s, 2H), 3.67(s, 3H), 2.65(s, 3H), 2.64(s, 3H). MS(ESI): [M+H]+ calcd 510.2;
found 510.2.
4.1.6.7 [5-[[2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-
1-yl]methyl]-1H-indol-1-yl](phenyl)methanone (15b). 15b was prepared by
following the above general procedure. Yield: 90.2%. MP: 214-216 oC. 1H NMR(400
MHz, CDCl3): δ 8.35(d, 1H), 7.87(s, 1H), 7.76(d, 2H), 7.61(t, 2H), 7.55(t, 2H), 7.42(s,
2H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d, 1H), 6.73(d, 1H), 5.68(s, 2H), 3.74(s, 3H),
2.46(q, 2H), 2.30(s, 3H), 1.27(t, 3H). MS(ESI): [M+H]+ calcd 524.2; found 524.3.
4.1.6.8
[5-[[2-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]
-1H-indol-1-yl](phenyl)methanone (15c).
15c was prepared by following the above general procedure. Yield: 85.6%. MP:
o
1
214-217 C. H NMR(400 MHz, CDCl3): δ 8.35(d, 1H), 7.86(s, 1H), 7.78(d, 2H),
7.60(t, 2H), 7.54(t, 2H), 7.43(s, 2H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d, 1H), 6.73(d,
1H), 5.68(s, 2H), 3.74(s, 3H), 2.94(t, 2H), 2.78(s, 3H), 1.85(m, 2H), 1.04(t, 3H).
MS(ESI): [M+H]+ calcd 538.3; found 538.3.
4.1.6.9
[5-[[2-butyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-
1H-indol-1-yl](phenyl)methanone (15d).
15d was prepared by following the above general procedure. Yield: 87.6%. MP:
o
1
215-218 C. H NMR(400 MHz, CDCl3): δ 8.35(d, 1H), 7.83(s, 1H), 7.76(d, 2H),
7.61(t, 2H), 7.54(t, 2H), 7.43(s, 2H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d, 1H), 6.73(d,
1H), 5.68(s, 2H), 3.74(s, 3H), 2.41(t, 2H), 2.17(s, 3H), 1.73(m, 2H), 1.43(m, 2H),
0.97(t, 3H). MS(ESI): [M+H]+ calcd 552.3; found 552.3.
4.1.6.10
[5-[[2-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]
-1H-indol-1-yl](phenyl)methanone (15e).
15e was prepared by following the above general procedure. Yield: 90.3%. MP:

ACCEPTED MANUSCRIPT
o
1
217-220 C. H NMR(400 MHz, CDCl3): δ 8.35(d, 1H), 7.81(s, 1H), 7.76(d, 2H),
7.60(t, 2H), 7.53(t, 2H), 7.43(s, 2H), 7.34(d, 2H), 7.30(t, 2H), 7.29(d, 1H), 6.73(d,
1H), 5.68(s, 2H), 3.74(s, 3H), 2.45(t, 2H), 2.17(s, 3H), 1.73(m, 2H), 1.37(m, 4H),
0.94(t, 3H). MS(ESI): [M+H]+ calcd 566.3; found 566.3.
4.1.7 General procedure for the synthesis of 13a-e, 16a-e:
A solution of NaOH (0.8 g, 20 mmol) in 10 mL water was slowly added to the
solution of an appropriate N- benzoyl indole (3.72 mmol) in 10 mL methanol. The
mixture was refluxed for 3 h. After the reaction was completed, the solvent was
removed in vacuo. The residue was dissolved in 15 mL H2O and extracted with DCM
(10 mL×4). The organic layer was dried over MgSO4. After filtration, the solvent was
removed under reduced pressure and the residue was purified by CC to give the
product as white solid.
4.1.7.1
1-[(1H-indol-4-yl)methyl]-2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimida
zole (13a).
13a was prepared by following the above general procedure. Yield: 96.3%. MP:
o
1
221-223 C. H NMR (400 MHz, CDCl3): δ 8.5(brs, 1H), 7.83(s, 1H), 7.65(t, 2H),
7.41(d, 1H), 7.36(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.78(s, 3H), 2.04(s, 3H). MS(ESI): [M+H]+ calcd 406.2;
found 406.2.
4.1.7.2
1-[(1H-indol-4-yl)methyl]-2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzim
idazole (13b).
13b was prepared by following the above procedure. Yield: 95.2%. MP: 221-224
oC. 1H NMR (400 MHz, CDCl3): δ8.5(brs, 1H), 7.83(s, 1H), 7.62(t, 2H), 7.41(d, 1H),
7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H),
3.74(s, 3H), 2.46(q, 2H), 2.30(s, 3H), 1.27(t, 3H). MS(ESI): [M+H]+ calcd 420.2;
found 420.3.
4.1.7.3
1-[(1H-indol-4-yl)methyl]-2-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzi
midazole (13c).
13c was prepared by following the above general procedure. Yield: 97.9%. MP:
o
1
222-224 C. H NMR (400 MHz, CDCl3): δ 8.5(brs, 1H), 7.81(s, 1H), 7.60(t, 2H),
7.45(d, 1H), 7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.94(t, 2H), 2.78(s, 3H), 1.85(m, 2H), 1.04(t, 3H).

ACCEPTED MANUSCRIPT
MS(ESI): [M+H]+ calcd 434.2; found 434.3.
4.1.7.4 1-[(1H-indol-4-yl)methyl]-2-butyl-4-methyl-6-(1-methylbenzimidazol
-2-yl)benzimidazole (13d).
13d was prepared by following the above general procedure. Yield: 94.3%. MP:
o
1
223-225 C. H NMR (400 MHz, CDCl3): δ 8.5(brs, 1H), 7.84(s, 1H), 7.61(t, 2H),
7.45(d, 1H), 7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.41(t, 2H), 2.17(s, 3H), 1.73(m, 2H), 1.43(m, 2H),
0.97(t, 3H). MS(ESI): [M+H]+ calcd 448.3 ; found 448.3.
4.1.7.5
1-[(1H-indol-4-yl)methyl]-2-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzi
midazole (13e).
13e was prepared by following the above general procedure. Yield: 90.4%. MP:
o
1
224-226 C. H NMR (400 MHz, CDCl3): δ 8.5(brs, 1H), 7.81(s, 1H), 7.63(t, 2H),
7.48(d, 1H), 7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.45(t, 2H), 2.36(s, 3H), 1.73(m, 2H), 1.37(m, 4H),
0.94(t, 3H). MS(ESI): [M+H]+ calcd 462.3 ; found 462.3.
4.1.7.6
1-[(1H-indol-5-yl)methyl]-2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimida
zole (16a).
16a was prepared by following the above general procedure. Yield: 95.7%. MP:
o
1
221-223 C. H NMR (400 MHz, CDCl3): δ 7.83(d, 1H), 7.69(m, 1H), 7.52(m, 2H),
7.43(m, 2H), 7.37(m, 2H),7.32(s, 1H), 7.25(m, 1H), 6.50(brs, 1H), 5.52(s, 2H),
3.85(s,3H), 2.68(s, 3H), 2.04(s, 3H). MS(ESI): [M+1]+ calcd 406.2; found 406.4.
4.1.6.7
1-[(1H-indol-5-yl)methyl]-2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzim
idazole (16b).
16b was prepared by following the above general procedure. Yield: 93.4%. MP:
o
1
221-223 C. H NMR (400 MHz, CDCl3): δ8.5(brs, 1H), 7.82(s, 1H), 7.63(t, 2H),
7.45(d, 1H), 7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.46(q, 2H), 2.30(s, 3H), 1.27(t, 3H). MS(ESI): [M+H]+
calcd 420.2; found 420.3.
4.1.6.8
1-[(1H-indol-5-yl)methyl]-2-n-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)ben
zimidazole (16c).
16c was prepared by following the above general procedure. Yield: 80.1%. MP:

ACCEPTED MANUSCRIPT
o
1
222-224 C. H NMR (400 MHz, CDCl3): δ8.5(brs, 1H), 7.80(s, 1H), 7.65(t, 2H),
7.45(d, 1H), 7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.45(t, 2H), 2.78(s, 3H), 1.85(m, 2H), 1.04(t, 3H).
MS(ESI): [M+H]+ calcd 434.2; found 434.3.
4.6.7.9
1-[(1H-indol-5-yl)methyl]-2-n-butyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benz
imidazole (16d).
16d was prepared by following the above general procedure. Yield: 98.4%. MP:
o
1
223-225 C. H NMR (400 MHz, CDCl3): δ8.5(brs, 1H), 7.89(s, 1H), 7.68(t, 2H),
7.43(d, 1H), 7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.41(t, 2H), 2.17(s, 3H), 1.73(m, 2H), 1.43(m, 2H),
0.97(t, 3H). MS(ESI): [M+H]+ calcd 448.3; found 448.3.
4.6.7.10
1-[(1H-indol-5-yl)methyl]-2-n-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)ben
zimidazole (16e).
16e was prepared by following the above general procedure. Yield: 91.4%. MP:
o
1
223-225 C. H NMR (400 MHz, CDCl3): δ8.5(brs, 1H), 7.80(s, 1H), 7.65(t, 2H),
7.45(d, 1H), 7.31(t, 2H), 7.30(s, 1H),7.28(d, 1H), 7.27(d, 1H), 7.02(t, 1H), 6.50(d,
2H), 5.73(s, 1H), 3.74(s, 3H), 2.94(t, 2H), 2.78(s, 3H), 1.73(m, 2H), 1.37(m, 4H),
0.93(t, 3H). MS(ESI): [M+1]+ calcd 462.3; found 462.3.
4.1.7 General procedure for the synthesis of 14a-e, 17a-e:
2-Fluorobenzonitrile (0.5 g, 4.16 mmol) and K2CO3(0.77 g, 5.54 mmol) were
added to a solution of an appropriate bis- benzimidazole (2.77 mmol) in 50 mL DMF.
The mixture was refluxed for 5 h under nitrogen. After cooling to rt, the mixture was
added 30 mL ethyl acetate and washed with NaCl saturated solution (50 mL×5). The
organic phase was collected and dried over MgSO4. After filtration, the solvent was
removed under reduced pressure and the residue was purified by CC to give the
product as white solid.
4.1.7.1
2-[4-[[2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-1H
-indol-1-yl]benzonitrile (14a).
14a was prepared by following the above general procedure. Yield: 87.3%. MP:
o
1
231-233 C. H NMR(400 MHz, CDCl3): δ7.93(d, 1H), 7.82(s, 1H), 7.76(d, 2H),
7.71(s, 1H), 7.53(t, 1H), 7.48(s, 1H), 7.43(s, 1H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d,
1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.78(s, 3H), 2.04(s, 3H).

ACCEPTED MANUSCRIPT
MS(ESI): [M+H]+ calcd 507.2; found 507.3.
4.1.7.2
2-[4-[[2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]
-1H-indol-1-yl]benzonitrile (14b).
14b was prepared by following the above general procedure. Yield: 82.7%. MP:
o
1
231-233 C. H NMR(400 MHz, CDCl3): δ7.89(d, 1H), 7.82(s, 1H), 7.76(d, 2H),
7.70(s, 1H), 7.54(t, 1H), 7.48(s, 1H), 7.43(s, 1H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d,
1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.46(q, 2H), 2.30(s, 3H),
1.27(t, 3H). MS(ESI): [M+H]+ calcd 521.3; found 521.3.
4.1.7.3
2-[4-[[2-n-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]met
hyl]-1H-indol-1-yl]benzonitrile (14c).
14c was prepared by following the above general procedure. Yield: 88.3%. MP:
o
1
232-234 C. H NMR(400 MHz, CDCl3): δ7.89(d, 1H), 7.80(s, 1H), 7.78(d, 2H),
7.73(s, 1H), 7.53(t, 1H), 7.48(s, 1H), 7.43(s, 1H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d,
1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.94(t, 2H), 2.78(s, 3H),
1.85(q, 2H), 1.04(t, 3H). MS(ESI): [M+H]+ calcd 535.3; found 535.3.
4.1.7.4
2-[4-[[2-n-butyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]meth
yl]-1H-indol-1-yl]benzonitrile (14d).
14d was prepared by following the above general procedure. Yield: 87.6%. MP:
o
1
233-235 C. H NMR(400 MHz, CDCl3): δ7.86(d, 1H), 7.81(s, 1H), 7.75(d, 2H),
7.73(s, 1H), 7.53(t, 1H), 7.48(s, 1H), 7.43(s, 1H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d,
1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.41(t, 2H), 2.17(s, 3H),
1.73(q, 2H), 1.43(m, 2H), 0.97(t, 3H). MS(ESI): [M+H]+ calcd 549.3; found 549.3.
4.1.7.5
2-[4-[[2-n-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]met
hyl]-1H-indol-1-yl]benzonitrile (14e).
14e was prepared by following the above general procedure. Yield: 85.3%. MP:
o
1
234-236 C. H NMR(400 MHz, CDCl3): δ7.96(d, 1H), 7.85(s, 1H), 7.78(d, 2H),
7.73(s, 1H), 7.53(t, 1H), 7.48(s, 1H, Ph-H), 7.43(s, 1H), 7.34(d, 2H), 7.31(t, 2H),
7.28(d, 1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.45(t, 2H), 2.36(s,
3H), 1.73(q, 2H), 1.37(m, 4H), 0.94(t, 3H). MS(ESI): [M+H]+ calcd 563.3; found
563.3.
4.1.7.6

ACCEPTED MANUSCRIPT
2-[5-[[2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-1H
-indol-1-yl]benzonitrile (17a).
17a was prepared by following the above general procedure. Yield: 83.6%. MP:
o
1
231-233 C. H NMR(400 MHz, CDCl3): δ7.82(m, 2H), 7.53(m, 1H), 7.49(m, 1H),
7.43(s, 1H), 7.38(d, 1H), 7.34(s, 1H), 7.29(m, 3H), 7.26(m, 2H), 7.24(m, 1H), 7.02(d,
1H), 6.65(d, 1H), 5.50(s, 2H), 3.74(s, 3H), 2.76(s, 3H), 2.04(s, 3H). MS(ESI): [M+1]+
calcd 507.2; found 507.4.
4.1.7.7
2-[5-[[2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]
-1H-indol-1-yl]benzonitrile (17b).
17b was prepared by following the above general procedure. Yield: 82.6%. MP:
o
1
231-233 C. H NMR(400 MHz, CDCl3): δ7.89(d, 1H), 7.81(s, 1H), 7.78(d, 2H),
7.71(s, 1H), 7.51(t, 1H), 7.46(s, 1H), 7.42(s, 1H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d,
1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.46(q, 2H), 2.30(s, 3H),
1.27(t, 3H). MS(ESI): [M+H]+ calcd 521.3; found 521.3.
4.1.7.8
2-[5-[[2-n-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]met
hyl]-1H-indol-1-yl]benzonitrile (17c).
17c was prepared by following the above general procedure. Yield: 60.1%. MP:
o
1
232-234 C. H NMR(400 MHz, CDCl3): δ7.92(d, 1H), 7.80(s, 1H), 7.78(d, 2H),
7.73(s, 1H), 7.53(t, 1H), 7.48(s, 1H), 7.43(s, 1H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d,
1H), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.94(t, 2H), 2.78(s, 3H),
1.85(m, 2H), 1.04(t, 3H). MS(ESI): [M+H]+ calcd 535.3; found 535.3.
4.1.7.9
2-[5-[[2-n-butyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]meth
yl]-1H-indol-1-yl]benzonitrile (17d).
17d was prepared by following the above general procedure. Yield: 87.3%. MP:
1
233-235oC. H NMR(400 MHz, CDCl3): δ7.91(d, 1H), 7.83(s, 1H), 7.76(d, 2H),
7.71(s, 1H), 7.52(t, 1H), 7.48(s, 1H), 7.43(s, 1H), 7.34(d, 2H), 7.31(t, 2H), 7.28(d,
1HH), 7.02(t, 1H), 6.50(d, 2H), 5.73(s, 1H), 3.74(s, 3H), 2.41(t, 2H), 2.17(s, 3H),
1.73(m, 2H), 1.43(m, 2H), 0.97(t, 3H). MS(ESI): [M+H]+ calcd 549.3; found 549.3.
4.1.7.10
2-[5-[[2-n-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]met
hyl]-1H-indol-1-yl]benzonitrile (17e).
17e was prepared by following the above procedure. Yield: 89.4%. MP: 234-236

ACCEPTED MANUSCRIPT
1
oC. H NMR(400 MHz, CDCl3): δ7.84(m, 2H), 7.73(m, 1H), 7.53(m, 3H), 7.51(m,
2H), 7.49(m, 2H), 7.51(m, 2H), 7.12(m, 1H), 7.34(m, 1H), 7.02(d, 1H), 5.55(s, 2H),
3.76(s, 3H), 2.45(t, 2H), 2.36(s, 3H), 1.73(m, 2H), 1.37(m, 4H), 0.94(t, 3H). MS(ESI):
[M+1]+ calcd 563.3; found 563.3.
4.1.8 General procedure for the synthesis of 1a-e, 2a-e, 4a-c, 5a-c:
A solution of NaOH (5 g, 125 mmol) in 25 mL water was added to a solution of
an appropriate benzonitrile (0.75 mmol) in 25 mL methanol. The solution was
refluxed for 30 h. After evaporation of methanol, the pH was adjusted to 2 by adding
3 M hydrochloric acid solution. The mixture was extracted with ethyl acetate (50 mL
× 3). The organic phase was collected and dried over MgSO4. After filtration, the
solvent was removed under reduced pressure and the residue was purified by CC to
give the product as white solid.
4.1.8.1
2-[4-[[2,4-dimethyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]-1H
-indol-1-yl]benzoic acid (1a).
1a was prepared by following the above general procedure. Yield: 51.9%. MP:
263 - 265 oC. 1H NMR (400 MHz, DMSO): δ 13.14 (s, 1H), 7.90 (d, J = 7.1 Hz, 1H),
7.72-7.65 (m, 3H), 7.58-7.57 (m, 2H), 7.51-7.48 (m, 3H), 7.31-7.18 (m, 2H), 7.08 (d,
J = 37.3 Hz, 2H), 6.64 (s, 1H), 6.54 (s, 1H), 5.87 (s, 2H), 3.78 (s, 3H), 2.64 (s, 3H),
2.61 (s, 3H). 13C NMR (101 MHz, DMSO): δ 167.84, 154.49, 153.76, 143.09, 142.81,
137.53ꢀ137.51, 137.32, 137.04, 135.62, 132.64ꢀ131.00, 130.62, 128.80, 128.76,
128.55, 128.41, 126.79, 126.77, 123.54, 122.52, 122.47, 122.27, 119.12, 117.58,
110.88, 109.89, 109.63, 100.98, 45.67, 32.17, 16.90, 14.39. HRMS (ESI): m/z
calculated for C33H28N5O2 [M+H]+: 526.2243; found 526.2225.
4.1.8.2
2-[4-[[2-ethyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]methyl]
-1H-indol-1-yl]benzoic acid (1b).
1b was prepared by following the above general procedure. Yield: 51.6%. MP:
o
1
262-264 C. H NMR (400 MHz, DMSO): δ 13.08 (s, 1H), 7.94 (d, J = 7.4 Hz, 1H),
7.74 (t, J = 7.3 Hz, 1H), 7.69-7.62 (m, 2H), 7.61-7.56 (m, 2H), 7.53-7.48 (m, 3H),
7.29-7.19 (m, 2H), 7.09- 6.96 (m, 2H), 6.64 (d, J = 3.2 Hz, 1H), 6.52 (d, J = 6.1 Hz,
1H), 5.88 (s, 2H), 3.78 (s, 3H), 2.95 (q, J = 7.5 Hz, 2H), 2.66 (s, 3H), 1.34 (t, J = 7.5
Hz, 3H). 13C NMR (101 MHz, DMSO): δ 167.61, 158.01, 154.49, 143.08, 142.77,
137.74, 137.58, 137.32, 137.00, 135.67, 133.13, 131.15, 130.55, 128.94, 128.92,
128.69, 128.51, 126.74, 126.72, 123.53, 122.62, 122.52, 122.28, 119.11, 117.48,

ACCEPTED MANUSCRIPT
110.86, 109.76, 109.68, 101.10, 45.44, 32.15, 20.92, 16.98, 12.24. HRMS (ESI): m/z
calculated for C34H30N5O2 [M+H]+: 540.2400; found 540.2381.
4.1.8.3
2-[4-[[2-n-propyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]met
hyl]-1H-indol-1-yl]benzoic acid (1c).
1c was prepared by following the above general procedure. Yield: 55.7%. MP:
264-266 oC. 1H NMR (400 MHz, CDCl3): δ 8.39 (d, J = 6.7 Hz, 1H), 8.18 (d, J = 7.6
Hz, 1H), 7.65 (t, J = 6.9 Hz, 1H), 7.55 (t, J = 7.4 Hz, 1H), 7.46 (d, J = 7.6 Hz, 1H),
7.39-7.31 (m, 3H), 7.25-7.14 (m, 3H), 7.06 (d, J = 3.2 Hz, 1H), 6.96 (s, 1H), 6.49 (s,
1H), 6.08 (d, J = 14.9 Hz, 1H), 5.71 (d, J = 3.1 Hz, 1H), 5.39 (d, J = 14.7 Hz, 1H),
3.69 (s, 3H), 3.26 (t, 2H), 2.72 (s, 3H), 2.17-1.98 (m, 2H), 1.22 (t, J = 7.3 Hz, 3H).
13C NMR (101 MHz, CDCl3): δ 169.07, 156.46, 154.09, 143.04, 140.48, 138.73,
137.85, 135.17, 135.03, 132.20, 132.04, 132.01, 129.70, 128.70, 128.11, 127.74,
126.65, 125.80, 123.19, 123.13, 123.12, 122.41, 121.19, 119.73, 119.51, 111.48,
110.94, 109.37, 100.19, 49.14, 31.41, 30.26, 22.15, 17.02, 14.23. HRMS (ESI): m/z
calculated for C35H32N5O2 [M+H]+: 554.2556; found 554.2535.
4.1.8.4
2-[4-[[2-n-butyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]meth
yl]-1H-indol-1-yl]benzoic acid (1d).
1d was prepared by following the above general procedure. Yield: 50.8%. MP:
264-267 oC. 1H NMR (400 MHz, CDCl3): δ 8.40 (d, J = 6.7 Hz, 1H), 8.18 (d, J = 7.6
Hz, 1H), 7.64 (t, J = 7.6 Hz, 1H), 7.55 (t, J = 7.2 Hz, 1H), 7.46 (d, J = 7.7 Hz, 1H),
7.35-7.34 (m, 3H), 7.27- 7.15 (m, 3H), 7.06 (d, J = 3.2 Hz, 1H), 6.95 (s, 1H), 6.47 (s,
1H), 6.08 (d, J = 14.0 Hz, 1H), 5.70 (d, J = 2.8 Hz, 1H), 5.38 (d, J = 14.7 Hz, 1H),
3.70 (s, 3H), 3.27 (t, J = 9.0, 6.9 Hz, 2H), 2.72 (s, 3H), 2.09 - 2.02 (m, 2H), 1.67-1.60
13
(m, 2H), 1.08 (t, J = 7.4 Hz, 3H). C NMR (101 MHz, CDCl3): δ 169.09, 156.60,
154.18, 143.33, 140.45, 138.78, 137.84, 135.15, 135.11, 132.20, 132.09, 132.00,
129.64, 128.67, 128.09, 127.67, 126.67, 125.80, 123.16, 123.12, 123.01, 122.42,
120.97, 119.80, 119.62, 111.55, 110.95, 109.33, 100.22, 49.37, 31.40, 30.76, 28.31,
22.92, 17.00, 13.86. HRMS (ESI): m/z calculated for C36H34N5O2 [M+H]+: 568.2713;
found 568.2733.
4.1.8.5
2-[4-[[2-n-pentyl-4-methyl-6-(1-methylbenzimidazol-2-yl)benzimidazole-1-yl]met
hyl]-1H-indol-1-yl]benzoic acid (1e).
1e was prepared by following the above general procedure. Yield: 48.3%. MP: