Design, synthesis and bioevaluation of tricyclic fused ring system as dual binding site acetylcholinesterase inhibitors

Article abstract of DOI:10.1016/j.bioorg.2018.10.035

Due to recently discovered non-classical acetylcholinesterase (AChE) function, dual binding-site AChE inhibitors have acquired a paramount attention of drug designing researchers. The unique structural arrangements of AChE peripheral anionic site (PAS) and catalytic site (CAS) joined by a narrow gorge, prompted us to design the inhibitors that can interact with dual binding sites of AChE. Eighteen homo- and heterodimers of desloratadine and carbazole (already available tricyclic building blocks) were synthesized and tested for their inhibition potential against electric eel acetylcholinesterase (eeAChE) and equine serum butyrylcholinesterase (eqBChE). We identified a six-carbon tether heterodimer of desloratadine and indanedione based tricyclic dihydropyrimidine (4c) as potent and selective inhibitor of eeAChE with IC₅₀ value of 0.09 ± 0.003 μM and 1.04 ± 0.08 μM (for eqBChE) with selectivity index of 11.1. Binding pose analysis of potent inhibitors suggest that tricyclic ring is well accommodated into the AChE active site through hydrophobic interactions with Trp84 and Trp279. The indanone ring of most active heterodimer 4b is stabilized into the bottom of the gorge and forms hydrogen bonding interactions with the important catalytic triad residue Ser200.

Full text of DOI:10.1016/j.bioorg.2018.10.035

Accepted Manuscript
Design, Synthesis and Bioevaluation of Tricyclic Fused Ring System as Dual
Binding Site Acetylcholinesterase Inhibitors
Saba Tahir Tanoli, Muhammad Ramzan, Abbas Hassan, Abdul Sadiq,
Muhammad Saeed Jan, Farhan A. Khan, Farhat Ullah, Haseen Ahmad, Maria
Bibi, Tariq Mahmood, Umer Rashid
PII:
S0045-2068(18)30545-5
DOI:
https://doi.org/10.1016/j.bioorg.2018.10.035
YBIOO 2569
Reference:
Bioorganic Chemistry
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Revised Date:
Accepted Date:
4 June 2018
12 October 2018
17 October 2018
Please cite this article as: S. Tahir Tanoli, M. Ramzan, A. Hassan, A. Sadiq, M. Saeed Jan, F.A. Khan, F. Ullah, H.
Ahmad, M. Bibi, T. Mahmood, U. Rashid, Design, Synthesis and Bioevaluation of Tricyclic Fused Ring System as
Dual Binding Site Acetylcholinesterase Inhibitors, Bioorganic Chemistry (2018), doi: https://doi.org/10.1016/
j.bioorg.2018.10.035
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Design, Synthesis and Bioevaluation of Tricyclic Fused Ring System
as Dual Binding Site Acetylcholinesterase Inhibitors
Saba Tahir Tanoli 1, Muhammad Ramzan 2, Abbas Hassan 3, Abdul Sadiq 4,
Muhammad Saeed Jan 4, Farhan A. Khan 1, Farhat Ullah 4, Haseen Ahmad 3, Maria
Bibi 1, Tariq Mahmood 1, Umer Rashid 1,*
1 Department of Chemistry, COMSATS University Islamabad, Abbottabad Campus
22060, Pakistan
2 Department of Chemistry, Hazara University, Mansehra 21120, Pakistan
3 Department of Chemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan
4 Department of Pharmacy, University of Malakand, Chakdara 18000 Dir (L), Pakistan
Corresponding author. *E-mail: umerrashid@ciit.net.pk; umer_rashid39@hotmail.com (Umer
Rashid);
Tel. Off.: +92 (992) 383591-6
1

Abstract
Due to recently discovered non-classical acetylcholinesterase (AChE) function, dual binding-site
AChE inhibitors have acquired a paramount attention of drug designing researchers. The unique
structural arrangements of AChE peripheral anionic site (PAS) and catalytic site (CAS) joined by
a narrow gorge, prompted us to design the inhibitors that can interact with dual binding sites of
AChE. Eighteen homo- and heterodimers of desloratadine and carbazole (already available
tricyclic building blocks) were synthesized and tested for their inhibition potential against
electric eel acetylcholinesterase (eeAChE) and equine serum butyrylcholinesterase (eqBChE).
We identified a six-carbon tether heterodimer of desloratadine and indanedione based tricyclic
dihydropyrimidine (4c) as potent and selective inhibitor of eeAChE with IC50 value of 0.09±
0.003 μM and 1.04± 0.08 μM (for eqBChE) with selectivity index of 11.1. Binding pose analysis
of potent inhibitors suggest that tricyclic ring is well accommodated into the AChE active site
through hydrophobic interactions with Trp84 and Trp279. The indanone ring of most active
heterodimer 4b is stabilized into the bottom of the gorge and forms hydrogen bonding
interactions with the important catalytic triad residue Ser200.
Key words. Dual binding site inhibitors, tricyclic fused rings, desloratadine, nonclassical AChE
function, butyrylcholinesterase
2

1. Introduction
Alzheimer disease is characterized by the acetylcholine (ACh)-mediated abrupt blockade of
cortical cholinergic neuron population and the deposition of extracellular amyloid-β (Aβ) and tau
proteins into plaques and neurofibrillary tangles [1-3]. The inactivation of ACh is a unique
process carried out by two cholinesterases; acetylcholinesterase (AChE) and
butyrylcholinesterase (BChE), resulting in release of choline and acetate [4-6]. Thus, the
inhibition of both cholinesterases (acetylcholinesterase AChE and butyrylcholinesterase BChE)
is found to be the only effective therapeutic approach for AD up till now. Only five drugs have
been approved for its medication since 1993. Four out of five fall under the class cholinesterase
inhibitors (ChEIs): i.e tacrine (1), donepezil (2), rivastigmine (3) and galantamine (4). Only one
drug memantine (5) is an N-methyl-D-aspartate (NMDA) antagonist [1, 7-8]. More
disappointing fact is that all the current drugs, approved for AD treatment lack the disease-
modifying abilities, and are mainly symptomatic. None of the approved AD drug have the ability
to revert the damage or at least slow down the disease progression [9].
It is evident in some earlier studies that the ligands bind with peripheral anionic site (PAS) of
AChE e.g. propidium, prevented AChE-induced Aβ aggregation. This non-classical or non-
cholinergic function is distinct from active site inhibitors [10-13]. Due to this non-classical
AChE function, dual binding-site AChE inhibitors have acquired a paramount attention of
medicinal chemistry and drug designing researchers due to increased number of ligand-target
interactions. Dual binding site inhibitors interacted with both the catalytic anionic site (CAS) at
the bottom of the gorge (Trp84), and with the peripheral anionic site (PAS), near its entrance
(Trp279), via aromatic π-staking interactions [14-15]. Donepezil shows dual-binding site
interactions among FDA approved AChEIs [16]. Dual binding site inhibitors must have some
3

structural moieties that can enter the gorge and can interact with the aromatic residues of CAS
linked through a linker or a spacer that lies in the neck which is a narrow part of active site of
AChE and other side of a molecule interact with amino acid residues present in PAS. Dual
binding site inhibitors can be analogues, hybrids and homo-dimers or hetero-dimers linked
through different linkers [17-24].
Tacrine (1), composed of a tricyclic fused ring system, is a potent AChE inhibitor with its
potency in submicromolar range. Bis-tacrine (2) is novel compound consisting of homodimer of
tacrine with heptylene linker reported as dual binding site inhibitor. Many tricyclic fused ring
systems has been reported in literature as potent AChE/BChE inhibitors. Apart from tacrine and
physostigmine analogues, quinolizidinyl analogous of bi- and tricyclic fused ring system with
inhibition of ChEs in submicromolar range [25]. Tin et al. reported multipotent derivatives of
phenothiazine and phenoselenazine targeting cholinergic, oxidative stress and amyloid pathways
[26]. The structures of the potent tricyclic systems are shown in Figure 1.
a)
HN
N
NH
N
N
NH2
2
1
H
H
b)
N
N
O
7
N
N
2
C
N
N
OH
H
HN
Cl
3, IC50=17 nM
4, IC50=20 nM
O
O
S
N
N
Cl
H
n
HN
N
H
H3C
O
CH3
HN
( )m
S
OAc
O
OAc
N
R
H3C
CH3
N
5, IC50 = 22 nM
7, IC50= 0.840 M
6, IC50= 260 M
4

Figure 1: (a) Tricyclic fused ring antidepressants; (b) tricyclic fused ring inhibitors of AChE
Recently, our group has reported dihydropyrimidine and quinazoline, a bicyclic fused ring
system, based AChE and BChE inhibitors [17, 27-28]. In continuation of our previous work,
series of tricyclic fused ring system-based homo- and hetero-bivalent inhibitors was designed to
monitor the dual binding site interactions into the AChE active site gorge. To this end, we
planned to use already available tricyclic fused systems; desloratadine (8) and carbazole (9) in
order to improve binding interactions. Desloratadine (8) is an aromatic drug having a benzene
and pyridine ring fused with a seven membered cyclic ring. A piperidine ring is also present
connected with a seven membered ring via a double bond. The calculated volume of
desloratadine from online software Molinspiration (http://www.molinspiration.com/cgi-
bin/properties) is 283.78 A3. It was envisioned that this ring would result in the enhanced dual
binding site interactions. Desloratadine is considered as potent antihistamine. Structural
modification of desloratadine resulted in many compounds having antihistaminic and anti-
inflammatory, antagonist of arginine vasopressin V2 [29-30]. On the other hand, the effect of a
relatively small tricyclic carbazole (9, Volume=157.01 A3) was also investigated. Carbazole
contains a fused ring system with two benzene rings fused on either side with a five membered
nitrogen-heterocycle
5

Figure 2: Desloratadine (8) and carbazole (9) as starting structures of current research
2. Results and discussion
2.1. Chemistry
2.1.1. Synthesis of benzyl/benzoyl derivatives of desloratadine
Benzyl derivatives of desloratadine (1a-e) were synthesized by reaction of desloratadine (8) with
substituted benzyl chlorides (10-13) and 4-chlorobenzoyl chloride (14) in the presence of
triethylamine as base and THF as solvent as shown in Scheme 1.
Cl
Cl
TEA / DCM
Reflux
Cl
N X
X
NH
+
R
N
N
R
10-14
8
1a-e
R (10-13) = -H, -4-Cl, -4-OH, -4-CH3; X= -CH2
R (14) = -4Cl; X = -C=O
Scheme 1: Synthesis of benzyl derivatives of desloratadine (1a-e)
6

2.1.2. Synthesis of homodimers of desloratadine and carbazole.
We synthesized homo- and heterodimers of selected tricyclic fused rings containing drug(s)
(desloratadine) / carbazole using suitable linkers (rigid and flexible) (Schemes 2-3). In general
strategy, for the synthesis of homodimers, 2 equivalents of desloratadine (8) or carbazole (9)
were reacted with reactants 15-19 to yield 2a-e and 3a-e (Scheme 2). Mannich bases 2a and 3a
were synthesized by using 2 equivalents of 8 or 9 and formalin (15, 40 %) in ethanolic solution
at room temperature. While, for the synthesis of 2b-e and 3b-e, desloratadine and carbazole were
reacted with each of bromoacetyl bromide (16), dibromo- (17), diiodo hexane (18) and triazine
(19) in dimethylformamide (DMF) using potassium carbonate as base at 70-80 ˚C (Scheme 2).
7

X R
16-19
Y
N
N
N
K2CO3 / DMF
NH
N
R
N
2
15, EtOH
Stir, rt, 14 hrs
2a-e
Cl
Cl
2a
Cl
8
X R
Y
16-19
K2CO3 / DMF
N R
N
2
N
15, EtOH
Stir, rt, 14 hrs
H
3a-e
9
3a
2a, 3a = -CH2-
2b, 3b = -CH2-CO-
2c, 3c = -(CH2)2-
2d, 3d = -(CH2)6-
R=
2e, 3e = -6-methoxy-1,3,5-triazine-
O
O
O
N
N
Br
Br
H
H
I
I
Br
6
Br
Cl
N
Cl
15
16
17
18
19
Scheme 2: Synthesis of bis-desloratadine (2a-e) from 8 and bis-carbazole derivatives (3a-d) from
9
2.1.3. Synthesis of heterodimers of desloratadine
8

The design of desloratadine-carbazole heterodimers was carried out to analyze the effect of two
diverse tricyclic systems on the binding interactions. For the synthesis of desloratadine-carbazole
heterodimers, the tether length of two (rigid ethenone linker, 4a) and six carbon flexible linker
(4b) was selected. The reaction of desloratadine with bromoacetyl bromide (16) and diiodo
hexane (18) resulted in intermediates 20 and 21 respectively. These intermediates further react
with carbazole (9) in the presence of K2CO3 and DMF to give 4a and 4b respectively (Scheme
3).
N
N
O
O
K2CO3 / DMF
16
N
N
N
Br
9
N
4a
K2CO3 / DMF
Cl
Cl
20
NH
N
N
8
Cl
18
K2CO3 / DMF
N
N
9
N
I
Cl
Cl
4b
21
Scheme 3: Desloratadine-carbazole heterodimer 4a and 5a
Our initial pharmacological work to identify dihydropyrimidine (DHPM) scaffold as potent
inhibitors of cholinesterases led us to design a new tricyclic building block [17]. Thus, synthesis
of DHPM-based fused ring system (25) was planned through reaction of 1,3-indanedione (22) as
a 1,3-dicarbonyl precursor in the Biginelli reaction [31]. The synthesized 25 was then reacted
with intermediate 21 to synthesize target heterodimer 4c.
9

O
O
O
NH2
S
SnCl2 / ACN
Reflux
NH
H2N
N
S
O
H
H
22
23
24
25
Cl
N
K2CO3 / DMF
O
N
N
25
N
I
N
H
N
S
Cl
21
4c
Scheme 4: Synthesis of heterodimer 4-(4-methoxyphenyl)-2-thioxo-3,4-tetrahydro-5H-indeno[1,2-
d]pyrimidin-5-one and desloratadine (4c)
2.2. In vitro eeAChE and eqBChE inhibition studies
To investigate the potential of synthesized compounds, we evaluated their ability to inhibit
cholinesterases. Spectroscopic Ellman’s method was used for electric eel acetylcholinesterase
(eeAChE) and butyrylcholinesterase from equine serum (eqBChE) assays. Tacrine and donepezil
were used as reference compounds. The IC50 values of the parent desloratadine and carbazole
was also determined for comparison purpose. Table 1 enlist the in vitro AChE and BChE
inhibitory activities of the homodimers of desloratadine and carbazole as 50% inhibitory
concentration (IC50). The in vitro inhibitory data in Table 1 shows that all the benzyl derivatives
of desloratadine (1a-d) are moderately active against AChE and showed poor inhibition for
BChE. They are more active than parent desloratadine (AChE IC50=23.56±0.6 µM). Benzoyl
derivative 1e has shown good inhibition with IC50 value of 9.51±1.04 µM for AChE. Its IC50
value for BChE inhibition is 33.4±1.28 µM.
10

Homodimers of desloratadine 2a-e were emerged as potent inhibitors of AChE in low
micromolar to submicromolar range. Alkylene-linked bis-desloratadine (2d, 6 carbon linker)
with IC50 value of 0.19±0.01 µM emerged as the most potent AChE inhibitor of this series.
Compound 2d is found 38.66 times more selective for AChE than BChE (IC50 for BChE= 7.2 ±
0.18 µM). The inhibitory data in Table 1 revealed shortening of linker length led to decrease in
activity. Compound 2a with methylene linker exhibited the potency of 6.1±0.71 µM.
Compounds 2b and 2c with linker length of two carbon atoms showed varying potency.
Compound 2b with ethanone linker exhibited the potency of 6.43±0.1.19 µM. While, ethylene
linker containing compound 2c showed the potency of 1.5±0.88 µM. Compound 2e with a rigid
triazine linker have shown inhibitory potential with IC50 value of 6.6±0.43 µM. All of these
compounds also showed high selectivity for AChE than BChE.
In contrast, homodimers of carbazole 3a-e with almost same linker length were found to have
moderate eeAChE inhibition except 3c and 3d. In contrast to desloratadine homodimer with the
linker length of 6-carbon (2d), carbazole homodimer with same linker length (3d) showed 11
times less potency (IC50=2.1±0.09 µM). All the compounds of this series also exhibited poor
BChE inhibition (Table 1).
In the heterodimeric series (4a-c), a different influence of tricyclic system was observed. In
homodimer series of desloratadine, compound 2b with ethanone linker was almost 4 times potent
than bis-carbazole 3b (Table 1). The inhibition data for compounds 4a-c in Table 1 showed that
desloratadine-carbazole dimer (4a) with same spacer length showed IC50 value of 3.98 μM. The
biological profile displayed by heterodimer 4b (IC50=0.10 μM) have shown the influence of the
combination of small and large ring system. The observed activity of 4b is 1.8 and 20 times more
potent than respective homodimers (2d and 3d in Table 1). The most striking result of present
11

research is the inhibition data of 4c for both the tested enzymes. The introduction of indanone
tricyclic ring lead to the most potent inhibitor of eeAChE (IC50=0.09 μM) and eqBChE
(IC50=1.04 μM).
Table 1: ChEIs inhibition data of compounds homo- and heterodimers of desloratadine and
carbazole 1a-e, 2a-e, 3a-d and heterodimer of desloratadine 4a-c
IC50 (µM ±SEM)
Selectivity
Compound
Structures
Indexa
eeAChE
eqBChE
N
13.2 ± 0.78
118.5 ± 2.15
9.0
1a
1b
N
Cl
N
16.8 ± 0.71
121.3 ± 1.63
7.2
Cl
N
Cl
N
19.3 ± 1.09
109.4 ± 1.38
5.7
4.5
1c
HO
N
N
Cl
Cl
H3C
23.1±1.04
103.4±1.71
1d
N
12

N
Cl
9.51 ± 1.04
33.4 ± 1.28
3.5
1e
2a
N
Cl
O
Cl
N
6.1 ± 0.71
55.5 ± 1.21
9.1
N
N
N
Cl
N
N
N
N
6.4 ± 1.19
51.1 ± 1.11
8.0
2b
O
Cl
Cl
Cl
N
N
1.5 ± 0.88
12.0 ± 1.02
8.0
2c
N
N
Cl
N
Cl
N
0.19 ± 0.01
7.2 ± 0.18
38.0
2d
N
Cl
N
13

Cl
N
6.6 ± 0.43
75.1 ± 1.23
11.4
2e
N
N
O
N
N
Cl
N
N
CH3
N
N
34.9±1.61
25.4±1.98
80.2±1.76
2.3
4.4
3a
3b
O
N
112.2±1.43
N
N
7.3±1.10
2.1±0.09
98.2 ± 2.04
39.5±1.36
13.4
18.1
3c
N
N
3d
N
N
N
O
N
13.4±1.65
45.0±1.64
3.3
3e
N
CH3
N
14

Cl
N
N
4.0 ± 0.58
48.6 ± 1.37
12.1
43.0
4a
O
N
N
N
0.10±0.009
4.3± 0.08
4b
N
Cl
O
N
N
H
S
N
4
N
0.09 ± 0.003
1.04± 0.08
11.1
4c
Cl
Desloratadine
Carbazole
Donepezil
Tacrine
23.6 ± 0.56
58.6 ± 1.68
0.05 ± 0.01
0.4±0.019
34.1 ± 0.81
110.3 ± 2.3
5.4±0.27
1.4
1.8
108.0
0.15
0.06±0.009
a Selectivity Index = IC50 of BChE/ IC50 of AChE.
15

2.4. Docking study
Docking study was carried out with Molecular Operating Environment (MOE. v2016.08)
software [32]. Only three apo-crystal structures of Electrophorus electricus acetylcholinesterase
(eeAChE) are present in protein data bank (PDB); 1C2O, 1C2B and 1EEA with a resolution
between 4.20–4.50 Å. In contrast, there are a number of crystal structures for Torpedo
californica AChE (TcAChE) with native ligands and good resolution. For current study, we
selected 2CKM (Resolution 2.15 Å) with alkylene-linked bis-tacrine (7 carbon linker) as native
ligand. Before the prediction of the docking poses of the synthesized compounds, we evaluated
the docking reliability by re-docking of native ligand AA7 (bis-tacrine). The computed RMSD
value of 0.892 Å was found within the acceptable criteria (< 2 Å). The binding mode analysis of
the highly active compounds revealed that they bind in the active site of TcAChE almost similar
to the bound inhibitor bis-tacrine (B7T). The overlaid binding poses of bis-desloratadine
homodimers 2c and 2d with B7T are shown in Figure 3a-c. They all are engaged in π-π
stacking interactions with aromatic residues of Trp84 and Trp279. The molecular modeling
results suggest that the most potent inhibitor (2d, IC50=0.187 ± 0.71) of the series 2 was docked
on TcAChE, resulting in almost similar binding pose to B7T. The inhibitor is well inserted into
the binding groove extending from bottom of the gorge to PAS (Figure 4a-b). It interacts
simultaneously with PAS and CAS residues. One of the tricyclic desloratadine ring of the
homodimer bound to CAS of eeAChE and forms π-π stacking interactions with Trp84 and
Phe330. The other tricyclic desloratadine ring is bound to the PAS and interacts with Tyr70 and
Trp279 (Figure 4c). For compound 2c bearing a shorter flexible linker (dimethylene), one of the
tricyclic ring located in CAS and by interacting with Trp84 and Phe330. While, the other ring
16

interacts with Trp279 through hydrogen bonding with pyridine nitrogen of desloratadine (Figure
4d-e).
Figure 3: (a) The overlaid ribbon diagram of B7T, 2c and 2d into the binding site of TcAChE (2CKM).
(b) Close-up 3-D depiction of superimposed bound conformations (line-model) of; B7T (red), 2c
(green) and 2d (blue). The key residues (Trp84 and Trp279) are shown in green sticks. (c). Two-
dimensional (2-D) depiction of superimposed ligands (generated by MOE).
17

Figure 4: (a-c) Three-dimensional (3-D) docking poses of compound 2d into the active site of 2CKM.
(a) Ribbon diagram; (b) Stick model. The active site residues are depicted in colored sticks. (c) Close-up
view of the docked-pose of 2d showing important residues. (d-e) Lowest-energy docking pose of 2e into
the active site (d) 3-D close-up view, (e) 2-D interaction plot.
Carbazole homodimers have shown moderate to poor inhibition potential against eeAChE. The
most active homodimer of carbazole is 3d with hexamethylene linker (IC50=2.07±0.09 μM).
Tricyclic carbazole ring inserted into the CAS and interacts with Trp84 at a distance of 3.81 Å
and with Phe330 (3.87 Å). Heterodimers (4a-c) also showed similar binding orientation to that of
B7T. The lowest-energy docking poses of 4b and 4c are shown in Figure 5a-b. The docking
pose of compound 4b is shown in Figure 5a. Desloratadine and carbazole rings forms strong π-π
stacking interactions with Phe330, Trp84 and Tyr70, Trp279 respectively. Small tricyclic ring,
carbazole, is accommodated in PAS region and forms π-π stacking interactions with Tyr70 and
Trp279. On the other hand, the docking pose of the most potent compound of current research 4c
18

establishes hydrogen bonding as well as π-π stacking interactions with the key residues (Figure
5b). The tricyclic indanone ring is well accommodated into the bottom of the gorge and its
carbonyl oxygen forms hydrogen bonding interactions with the important catalytic triad residue
Ser200. While, phenyl ring at 4-position forms π-π stacking interactions with Trp84. The binding
mode of important compounds are shown in Figures S-5 to S-10 (Supplementary
information).
Figure 4: Close-up depiction of the lowest-energy three-dimensional (3-D) docking poses of compounds
(a) 4b and (b) 4c.
3. Conclusions
Alzheimer disease is a brain disorder. Many inhibitors have been synthesized until now to the
cope the disease and only symptoms are controllable by the already available drugs in market.
This study aimed to design and synthesize some dual binding site (DBS) inhibitors and test their
activity against AChE/BChE. A series of 18 compounds were designed and synthesized having
tricyclic fused ring systems. The salient features from the in vitro AChE/BChE inhibition data
19

can be summarized as follows: Parent compounds desloratadine and carbazole exhibited poor
AChE and BChE inhibition. Benzyl/benzoyl derivatives of desloratadine (1a-e) also showed
poor inhibition of both enzymes. Tricyclic homodimers of desloratadine emerged as potent
inhibitors compared with the carbazole derivatives. Flexible methylene (ethyl or hexyl)
homodimers (Table 1, 2c, 2d and 3d) showed excellent AChE inhibition but poor selectivity for
BChE. Highly rigid triazine showed good to moderate AChE inhibition. Rigid ethenone linker
(2b and 3b) showed less inhibition than slightly flexible ethyl linker (2c and 3c). Indanedione
tricyclic heterodimer of desloratadine (4c) with a six-carbon tether displayed nanomolar potency.
Binding pose analysis of potent inhibitors suggest that tricyclic ring is well accommodated into
the AChE active site through hydrophobic interactions with Trp84 and Trp279. The indanone
ring of most active heterodimer 4b is stabilized into the bottom of the gorge and forms hydrogen
bonding interactions with the important catalytic triad residue Ser200.
4. Material and methods
4.1. General
All the reagents and solvents were purchased from standard commercial vendors and were used
without any further purification. Desloratadine and carbazole were purchased from Sigma
Aldrich. 1H and 13C-NMR spectra were recorded in deuterated solvents on a Bruker spectrometer
at 300 and 75 MHz respectively using tetramethylsilane (TMS) as internal reference. Chemical
shifts are given in δ scale (ppm). Melting points were determined in open capillaries using
Gallen Kamp melting point apparatus (MP-D). The progress of all the reactions was monitored
by TLC on 2.0 x 5.0 cm aluminum sheets pre-coated with silica gel 60F254 with a layer
thickness of 0.25 mm (Merck). LC-MS spectra were obtained using Agilent technologies 1200
20

series high performance liquid chromatography comprising of G1315 DAD (diode array
detector) and ion trap LCMS G2445D SL. HPLC analysis were carried out on Schimadzu
LC20AT. Elemental analyses were conducted using a LECO-932 CHNS Analyzer (LECO
Corporation, USA).
4.1. General procedure for synthesis of benzyl/benzoyl derivatives of
desloratadine
To solution of desloratadine (8, 10 mmol) in dichloromethane (DCM, 30 mL) added Et3N and
o
stirred at 0 C for 10-15 minutes. To this solution, 10 mmol of benzyl/benzoyl bromide (21-25)
dissolved in 15 mL of DCM was added drop-wise and then heated to reflux until the completion
of reaction (TLC). After the completion of reaction, mixture was poured into ice, filtered and
recrystallized from ethanol.
4.1.1. 11-(1-benzylpiperidin-4-ylidene)-8-chloro-6,11-dihydro-5H-benzo[5,6]cyclohepta[1,2-
b]pyridine (1a)
1a was synthesized according to general procedure by using 8 and benzyl chloride (21). Rf 0.44
1
(n-hexane/ethyl acetate 6:1). Yield 73%, m.p. 156-155 ºC. HNMR (300 MHz, CDCl3): δ 8.39
(d, 1H, J=7.5 Hz, Ar-H), 7.48 (d, 1H, J=7.8 Hz, Ar-H), 7.20-7.15 (m, 4H, Ar-H), 7.10-7.06 (m,
5H, Ar-H), 3.66 (s, 2H, CH2), 3.10-3.01 (m, 2H, CH2), 2.83-2.73 (m, 2H, CH2), 2.61-2.59 (m,
2H, CH2), 2.40-237 (m, 2H, CH2), 2.18-2.08 (m, 4H, 2×CH2); 13CNMR (75 MHz, CDCl3):
153.2, 143.9, 137.5, 136.4, 133.5, 133.2(2C), 132.2, 129.9, 126.6(2C), 126.2(2C), 124.5,
124.2(2C), 123.7, 117.7, 114.2, 62.5, 57.5(2C), 31.7(2C), 29.9, 28.2.
2.1.2. 8-chloro-11-(1-(4-chlorobenzyl) piperidin-4-ylidene)-6, 11-dihydro-5H-benzo [5, 6]
cyclohepta [1, 2-b] pyridine (1b)
21

1b was synthesized according to general procedure by using 8 and 4-chlorobenzylchloride (22).
Rf 0.55 (n-hexane/ethyl acetate 6:1). Yield 74%, m.p. 158-161 ºC. 1HNMR (300 MHz, CDCl3):
δ 8.39 (d, 1H, J=7.5 Hz, Ar-H), 7.38 (d, 1H, J=7.8 Hz, Ar-H), 7.14-7.06 (m, 8H, Ar-H), 3.68 (s,
13
2H, CH2), 2.79-2.70 (m, 4H, 2×CH2), 2.45-2.25 (m, 8H, 4×CH2); CNMR (75 MHz, CDCl3):
153.1, 143.9, 137.3, 134.5, 133.2, 132.9 (2C), 132.0, 130.6, 129.0 (2C), 126.3(2C), 124.6, 124.2
(2C), 123.8, 117.7, 114.0, 62.4, 57.6 (2C), 31.7 (2C), 30.1, 28.4.
2.1.3. 4-((4-(8-Chloro-5H-benzo[5,6]cyclohepta[1,2-b]pyridin-11(6H)-ylidene)piperidin-1-
yl)methyl)phenol (1c)
1c was synthesized according to general procedure by using 8 and 4-hydroxybenzylchloride (23).
Rf 0.48 (n-hexane/ethyl acetate 6:1). Yield 63%, m.p. 160-163 ºC. 1HNMR (300 MHz, CDCl3):
δ 9.88 (brs, 1H, -OH), 8.42 (d, 1H, Ar-H), 7.48 (d, 1H, Ar-H), 7.18 (m, 4H, Ar-H), 6.98 (d, 2H,
Ar-H), 6.74 (d, 2H, Ar-H), 3.63 (s, 2H, CH2), 3.04-2.96 (m, 2H, CH2), 2.81-2.74 (m, 2H, CH2),
2.61-2.55 (m, 2H, CH2), 2.46-2.39 (m, 2H, CH2), 2.14-2.08 (m, 4H, 2×CH2); 13CNMR (75 MHz,
CDCl3): 153.3, 153.0, 144.1, 137.6, 133.4, 133.0 (2C), 131.1, 129.9, 128.8 (2C), 124.9 (2C),
123.9, 123.6, 118.1, 114.2, 113.4(2C), 62.8, 57.4(2C), 31.8 (2C), 30.1, 28.8.
2.1.5. 8-chloro-11-(1-(4-methylbenzyl) piperidin-4-ylidene)-6, 11-dihydro-5H-benzo [5, 6]
cyclohepta [1, 2-b] pyridine (1d)
1d was synthesized according to general procedure by using 8 and 4-methylbenzylchloride (24).
Rf 0.52 (n-hexane/ethyl acetate 6:1). Yield 72%, m.p. 171-173 ºC. 1HNMR (300 MHz, CDCl3):
δ 8.40 (d, 1H, J=7.2 Hz, Ar-H), 7.46 (d, 1H, J=7.8 Hz, Ar-H), 7.18 (m, 4H, Ar-H), 7.06 (d, 2H,
Ar-H), 6.94 (d, 2H, Ar-H), 3.63 (s, 2H, CH2), 2.99-2.89 (m, 2H, CH2), 2.75-2.68 (m, 2H, CH2),
2.58-2.52 (m, 2H, CH2), 2.45-2.39 (m, 2H, CH2), 2.27 (s, 3H, CH3), 2.14-2.08 (m, 4H, 2×CH2);
13CNMR (75 MHz, CDCl3): 154.0, 143.6, 137.5, 134.7, 133.5, 133.2(2C), 131.9, 129.8 (2),
22

128.8 (2C), 126.5 (2C), 125.5, 125.2, 123.5, 117.7, 114.2, 62.5, 57.3 (2C), 31.5 (2C), 30.1, 28.5,
22.3.
2.1.6.
13‐ chloro‐ 2‐ [1‐ (4‐ chlorobenzoyl)piperidin‐ 4‐ ylidene]‐ 4‐ azatricyclo
[9.4.0.03,8]penta- deca‐ 1(15),3(8),4,6,11,13‐ hexaene (1e)
1e was synthesized according to general procedure by using 8 and 4-chlorobenzoyl chloride (25).
1
Rf =0.35 (n-hexane/ethyl acetate 6:1). Yield 76%, m.p. 210-212 ºC. HNMR (300 MHz,
CDCl3): δ 8.56 (d, 1H, J=7.5 Hz, Ar-H), 7.86 (d, 2H, J=8.1 Hz, Ar-H), 7.51 (d, 1H, J=7.5 Hz,
Ar-H), 7.36 (d, 2H, J=8.1 Hz, Ar-H), 7.19-7.11 (m, 4H, Ar-H), 3.37-3.28 (m, 4H, CH2), 2.78-
2.69 (m, 2H, CH2), 2.31-2.22 (m, 2H, CH2), 2.00-1.95 (m, 4H, 2×CH2); 13CNMR (75 MHz,
CDCl3): 170.2, 153.8, 143.7, 137.4, 134.0, 133.4, 132.9 (2C), 132.0, 129.5, 129.0, 127.6 (2C),
126.4(2C), 124.9, 123.6, 117.5, 114.0, 47.3 (2C), 31.7(2C), 29.9.
4.2. Synthesis of homo-dimers of Desloratadine and carbazole (2a-e), (3a-d)
4.2.1. General procedure for the synthesis of 2a and 3a
To a solution of one equivalent of desloratadine (8, 10 mmol) or carbazole (9) in ethanol (15
mL), formalin 40% (26, 1.0 equiv. 10 mmol) was added and stirred for 10-15 minutes at room
temperature. in EtOH (15 mL). A solution of another on equivalent of desloratadine (0.6 mmol)
or carbazole in ethanol (5 mL) was then added drop-wise to it. The reaction was stirred and
monitored by TLC to completion. The mixture was then kept overnight in refrigerator. The
precipitates thus produced were filtered and recrystallized from ethanol to obtain 2a and 3a.
23

13-chloro-2-{1-[(4-{13-chloro-4-azatricyclo[9.4.0.0³,⁸ ] pentadeca-1(11),3(8), 4,6,12,14-
hexaen-2-ylidene}piperidin-1-yl) methyl]piperidin-4-ylidene}-4-azatricyclo [9.4.0.0³,⁸ ]
pentadeca-1(15),3(8),4,6,11,13-hexaene (2a)
2a was synthesized according to general procedure by using 8 and formalin (26). Yield= 70%;
1
m.p. 224-228℃; Rf = 0.58 (n-hexane/ethyl acetate 7:1). H NMR (300 MHz, CDCl3): 8.34 (d,
2H), 7.44 (d, 2H), 7.12 (m, 8H), 3.53 (s, 2H), 3.01-2.92 (m, 4H, 2×CH2), 2.78-2.71 (m, 4H,
2×CH2), 2.63-2.58 (m, 4H, 2×CH2), 2.38–2.32 (m, 4H, 2×CH2), 2.19–2.12 (m, 4H, 2×CH2),
13
2.00–1.92 (m, 4H, 2×CH2).; C NMR (75 MHz, CDCl3): 157.3 (2C), 144.9 (2C), 137.5(2C),
132.5 (2C), 132.2 (4C), 132.0 (2C), 131.2 (2C), 129.0 (2C), 126.5 (2C), 126.2 (2C), 123.7 (2C),
119.7 (2C), 114.2 (2C), 87.2, 55.6 (4C), 32.4 (2C), 32.0 (2C), 29.9(2C); LC-MS (m/z) [M+H+] =
633.3
Di(9H-carbazol-9-yl)methane (3a)
3a was synthesized according to general procedure by using 9 and formalin (26). Yield=77%; Rf
= 0.46 (n-hexane/ethyl acetate 7:1); m.p. 290-293℃; 1H NMR (300 MHz, CDCl3): δ 7.54 (d, 4H,
J=7.2 Hz, Ar-H), 7.47 (d, 4H, J=7.5 Hz, Ar-H), 7.33 (t, 4H, J=7.5 Hz, Ar-H), 7.24 (t, 4H, J=7.5
13
Hz, Ar-H), 6.50 (s, 2H, -CH2); C NMR (75 MHz, CDCl3): 137.1 (4C), 126.0(4C), 125.4(4C),
120.4(4C), 119.8(4C), 110.0(4C), 80.1; LC-MS found for C25H18N2 (m/z) = 347.2 [M+H+].
4.2.2. General procedure for the synthesis of bis-desloratadine (2b-e) and bis-
carbazole homodimers (3b-e)
A mixture of desloratadine/carbazole (2.0 eq; 8.686 mmol) and K2CO3 (2.0 eq; 17.373 mmol) in
DMF (10 mL) at 0˚C under N2. Stirred the mixture for 20 minutes and then add dihalo-
compounds (27-30, 1.0 eq;) in the reaction mixture drop wise at 0 ˚C. Then stay room
24

temperature and maintain the temperature at 70-80˚C and reflux the mixture for 5-6 hours. The
reaction was monitored by TLC. The extraction was done with water and chloroform (3 times in
50 ml/10ml ratio). Anhydrous sodium sulfate is added to dry off the mixture. The solvent was
evaporated and crude was obtained. Further, crude mixture was purified by silica gel column
chromatography (n-hexanes/ethyl acetate).
4.2.2.1. Bis-desloratadine derivatives (2b-e)
2-(4-{13-chloro-4-azatricyclo[9.4.0.0³,⁸ ]pentadeca-1(11),3(8),4,6,12,14-hexaen-2-ylidene}
piperidin-1-yl)-1-(4-{13-chloro-4-azatricyclo[9.4.0.0³,⁸ ]pentadeca-1(15),3(8),4,6,11,13-
hexaen-2-ylidene}piperidin-1-yl)ethan-1-one (2b)
2b was synthesized according to general procedure by using 8 and bromoacetyl bromide (27).
Purified by silica gel column chromatography (n-hexanes/ethyl acetate 5:1). Yield=69%; Rf =
0.43 (n-hexane/ethyl acetate 7:1); m.p. 215-219 ℃; 1H NMR (300 MHz, CDCl3): δ 8.29 (d, 2H, J
= 4.2 Hz, Ar-H), 7.43 (d, 2H, J = 6.9 Hz, Ar-H ), 7.09-7.01 (m, 8H), 3.43–3.20 (m, 4H), 3.16 (t,
4H, J=12.6 Hz), 3.01 (s, 2H), 2.88–2.76 (m, 8H), 2.59-2.35 (m, 8H).; 13C NMR (75 MHz,
CDCl3): 163.9, 157.4 (2C), 145.1 (2C), 137.6 (2C), 134.5 (2C), 134.0 (4C), 133.4 (2C),
131.6(2C), 131.1 (2C), 129.1 (2C), 126.5 (2C), 126.0 (2C), 123.4 (2C), 119.6 (2C), 114.1 (2C),
57.9 (2C), 55.5, 46.5(2C), 32.4(2C), 32.0(2C), 30.1(2C).; LC-MS found for C40H38Cl2N4O (m/z)
= 661.2 [M+H].
25

13-chloro-2-{1-[2-(4-{13-chloro-4-azatricyclo[9.4.0.0³,⁸ ]pentadeca-1(11),3(8),4,6,12,14-
hexaen-2-ylidene}piperidin-1-yl)ethyl]piperidin-4-ylidene}-4-azatricyclo[9.4.0.0³,⁸ ]
pentadeca-1(15),3(8),4,6,11,13-hexaene (2c)
2c was synthesized according to general procedure by using 8 and 1,2-dibromoethane (28).
1
Yield= 71%; Rf = 0.58 (n-hexanes/ethyl acetate 5:1); m.p. 250-254℃; H NMR (300 MHz,
CDCl3, TMS): δ 8.40 (d, 2H), 7.43 (m, 2H), 7.34 (m, 2H, Ar-H), 7.33 (m, 2H, Ar-H), 7.28 (m,
2H, Ar-H), 6.98 (m, 2H,), 3.08-3.00 (m, 4H), 2.83-2.75 (m, 4H), 2.65-2.56 (m, 4H), 2.48 (t, 4H,
J=7.2 Hz, 2CH2), 2.41-2.34 (m, 4H), 2.21-2.14 (m, 8H), 13C NMR (75 MHz, CDCl3): δ 157.3
(2C), 144.8 (2C), 137.5(2C), 132.3 (2C), 132.1 (4C), 131.9 (2C), 130.0 (2C), 129.0 (2C), 126.6
(2C), 126.1 (2C), 123.5 (2C), 119.7 (2C), 114.0 (2C), 55.0 (4C), 52.9 (2C), 32.1 (2C), 31.8 (2C),
29.8 (2C).; LC-MS found for C40H40Cl2N4 (m/z) [M+H]+ = 647.3
13-chloro-2-{1-[6-(4-{13-chloro-4-azatricyclo[9.4.0.0³,⁸ ] pentadeca1 (11),3(8), 4,6, 12,14-
hexaen-2-ylidene}piperidin-1-yl)hexyl]piperidin-4-ylidene}-4-azatricyclo
[9.4.0.0³,⁸ ]pentadeca-1(15),3(8),4,6,11,13-hexaene (2d)
2d was synthesized according to general procedure by using 8 and 1,6-diiodohexane (29). Yield=
74%; Rf = 0.55 (n-hexanes/ethyl acetate 6:1); m.p. 241-245℃; 1H NMR (300 MHz, CDCl3,
TMS): 8.48-8.46 (m, 2H, Ar-H), 7.54 (d, 2H, J=7.8 Hz, Ar-H), 7.19-7.05 (m, 8H, Ar-H), 3.01-
13
2.83 (m, 8H), 2.41-2.32 (m, 12H), 2.11-2.00 (m, 8H), 1.44-1.35 (m, 8H); C NMR (75 MHz,
CDCl3): 158.9(2C), 147.2 (2C), 140.5 (2C), 136.1(2C), 135.8 (4C), 135.5 (4C), 131.2 (2C),
129.5 (2C), 129.4 (2C), 128.5 (2C), 122.8 (2C), 115.9 (2C), 58.1(4C), 55.0(2C), 31.7 (4C),
29.1(2C), 28.9 (2C), 25.9 (2C), 22.5(2C).; LC-MS found for C44H48Cl2N4 (m/z) = 703.4 [M+H].
26

Synthesis
of
13-chloro-2-{1-[4-(4-{13-chloro-4-azatricyclo[9.4.0.0³,⁸ ]
pentadeca-
1(11),3(8),4,6,12,14-hexaen-2-ylidene}piperidin-1-yl)-6-methoxy-1,3,5-triazin-2-yl]
piperidin-4-ylidene}-4-azatricyclo [9.4.0.0³,⁸ ]pentadeca-1(15),3(8),4,6,11,13-hexaene (2e)
2e was synthesized according to general procedure by using 8 and 2, 4-dichloro-6-methoxy-
1
1,3,5-triazine (30). Yield= 73%; Rf = 0.57 (n-hexanes/ethyl acetate 6:1); m.p. 235-237 ℃; H
NMR (300 MHz, DMSO-d6): 8.34-8.31 (m, 2H, Ar-H), 7.57-7.55 (m, 2H, Ar-H), 7.28 (d, 2H,
Ar-H), 7.21-7.16 (m, 4H, Ar-H), 7.06 (d, 2H, J=8.1 Hz, Ar-H), 3.84 (s, 3H, -OCH3), 3.36-3.27
(m, 8H), 2.85-2.69 (m, 8H), 2.25-2.13 (m, 8H).; 13C NMR (75 MHz, DMSO-d6): 178.9 (2C),
169.6, 157.6 (2C), 145.1 (2C), 137.4 (2C), 132.5(2C), 132.1 (4C), 131.8 (2C), 131.2(2C), 129.0
(2C), 126.4 (2C), 126.2(2C), 123.5 (2C), 119.6 (2C), 114.2 (2C), 53.5, 52.0 (4C), 32.4 (2C), 32.0
(2C), 30.0 (2C).; LC-MS for C42H39Cl2N7O (m/z) 728.3 [M+H].
4.2.2.2. Bis-carbazole homodimers (3b-e)
1,2-di(9H-carbazol-9-yl)ethan-1-one (3b)
3b was synthesized according to general procedure by using 9 and bromoacetyl bromide (27).
1
Yield=77%; Rf = 0.62 (n-hexanes/ethyl acetate 5:1); m.p. 290-293℃; H NMR (300 MHz,
CDCl3): 7.83 (d, 2H, J=8.1 Hz, Ar-H), 7.52 (d, 2H, J=7.8 Hz, Ar-H), 7.45 (d, 2H, J=7.8 Hz, Ar-
13
H), 7.33 (d, 2H, J=7.5 Hz, Ar-H), 7.12-7.01 (m, 8H, Ar-H), 5.07 (s, 2H, -CH2).; C NMR (75
MHz, CDCl3): 198.1, 132.8(2C), 130.3(2C), 123.9(4C), 122.4(4C), 120.4(4C), 117.6(2C),
117.1(2C), 113.4(4C), 57.4; LC-MS found for C26H18N2 (m/z) = 375.2 [M+H].
27

1,2-di(9H-carbazol-9-yl)ethane (3c)
3c was synthesized according to general procedure by using 9 and 1,2-dibromoethane (28).
1
Yield=79%; Rf = 0.51 (n-hexanes/ethyl acetate 5:1); m.p. 299-300℃; H NMR (300 MHz,
CDCl3): 7.71 (d, 4H, J=7.5 Hz, Ar-H), 7.48 (m, 8H, Ar-H), 7.25 (m, 4H, Ar-H), 4.25 (s, 4H, 2 x
13
CH2); C NMR (75 MHz, CDCl3): 132.7 (4C), 120.6(4C), 120.5(4C), 120.2(4C), 117.5 (4C),
108.0 (4C), 59.8 (2C).; LC-MS found for C26H20N2 (m/z) = 361.2 [M+H+].
9-(6-(9H-carbazol-9-yl)hexyl)-9H-carbazole (3d)
1
3d was synthesized according to general procedure by using 9 and 1,6-diiodohexane (18). H
NMR (300 MHz, CDCl3): 7.73 (d, 4H, J=7.5 Hz, Ar-H), 7.39-7.31 (m, 8H, Ar-H), 7.17-7.10 (m,
4H, Ar-H), 4.05 (t, 4H, J=6.6 Hz, CH2), 1.91-1.80 (m, 4H, CH2), 1.32 (t, 4H, J=6.6 Hz, CH2);
13C NMR (75 MHz, CDCl3): 132.8 (4C), 120.2 (4C), 119.6 (4C), 119.4 (4C), 118.0 (4C), 107.0
(4C), 56.1 (2C), 28.2 (2C), 25.7 (2C).; LC-MS found for C30H28N2 (m/z) = 417.3 [M+H].
9,9'-(6-methoxy-1,3,5-triazine-2,4-diyl)bis(9H-carbazole) (3e)
3e was synthesized according to general procedure by using 9 and 2, 4-dichloro-6-methoxy-
1
1,3,5-triazine (30). Yield=80%; Rf = 0.57 (n-hexanes/ethyl acetate 5:1); m.p. 319-321 ℃; H
13
NMR (300 MHz, CDCl3): 7.73 (d, 4H, J=7.5 Hz), 7.50 (m, 8H), 7.25 (m, 4H), 3.85 (s, 3H); C
NMR (75 MHz, CDCl3): 176.7 (2C), 172.8, 136.5 (4C), 124.4 (4C), 123.5 (4C), 122.2 (4C),
117.2 (4C), 107.2(4C), 52.4; LC-MS found for C28H19N5O (m/z) = 442.2 [M+H+].
4.3. Synthesis of desloratadine-carbazole hetero-dimers
4.3.1. Synthesis of
1‐ (9H‐ carbazol‐ 9‐ yl)‐ 2‐ (4‐ {13‐ chloro‐ 4‐ azatricyclo[9.4.0.03,8]pentadeca‐ 1(15),3(8
),4,6,11,13‐ hexaen‐ 2‐ ylidene}piperidin‐ 1‐ yl)ethan‐ 1‐ one (4a)
28

4a was synthesized according to general procedure described in Section 4.2.2 by using
desloratadine (8), carbazole (9) and bromoacetyl bromide (16). Yield=78.3%; Rf = 0.49; m.p.
1
241-243 ℃; HPLC purity=98.19 % (C18 RP, MeOH:H2O, 95:5). TR=8.339 min. H NMR (300
MHz, CDCl3): 8.41 (d, 1H, J=6.9 Hz, Ar-H), 7.73 (d, 1H, J=8.1 Hz, Ar-H), 7.50 (d, 2H, J=7.5
Hz, Ar-H), 7.40 (d, 2H, J=7.2 Hz, Ar-H), 7.06 (m, 8H, Ar-H), 3.21 (s, 2H, CH2), 3.06-2.94 (m,
4H, CH2); 2.55-2.32 (m, 4H, CH2), 2.19-2.12 (m, 4H, CH2).; 13C NMR (75 MHz, CDCl3): 168.4,
157.4, 143.9, 137.5, 136.8 (2C), 134.4, 134.2(2C), 133.8 (2C), 131.1, 126.0(2C), 123.5 (2C),
122.3(2C), 120.7, 119.1(2C), 117.6 (2C), 114.3, 112.8(2C), 57.5(2C), 55.2, 31.8(2C), 293, 28.5.
LC-MS found for C33H28ClN3O (m/z) = 518.2 [M+H+]. Analysis calcd: C, 76.51; H, 5.45; N,
8.11, Found C, 76.47; H, 5.43; N, 8.12.
2‐ {1‐ [6‐ (9H‐ carbazol‐ 9‐ yl)hexyl]piperidin‐ 4‐ ylidene}‐ 13‐ chloro‐ 4‐ azatricyclo[9.
4.0.03,8] pentadeca‐ 1(11),3(8),4,6,12,14‐ hexaene (4b)
4b was synthesized according to general procedure described in Section 4.2.2 by using 8, 9 and
1,6-diiodohexane (18). Yield= 74%; Rf = 0.49 (n-hexanes/ethyl acetate 9:1); HPLC purity=98.15
% (C18 RP, MeOH:H2O, 95:5). TR=6.27 min. m.p. 221-223 ℃; 1H NMR (300 MHz, DMSO-d6):
δ 8.71 (d, 1H, J=6.3 Hz, Ar-H), 7.69 (d, 1H, J=8.1 Hz, Ar-H), 7.42 (d, 4H, J=7.8 Hz, Ar-H),
7.20-7.10 (m, 8H, Ar-H), 3.88 (t, 2H, J=6.9 Hz, CH2), 299-2.85 (m, 4H, 2×CH2), 2.40-2.34 (m,
6H, CH2), 1.97 (t, 4H J=10.2 Hz, 2×CH2), 1.76-1.67 (m, 2H, 2×CH2), 1.44-1.30 (m, 6H,
2×CH2); 13C NMR (75 MHz, DMSO-d6): 157.8, 144.1, 141.2, 137.2, 136.8(2), 136.1, 135.0,
133.1, 132.8(2), 128.0, 127.4, 126.7, 123.4, 115.6, 122.4, 120.3(2), 119.6(2), 118.4(2), 116.9(2),
29