Novel chromanone-dithiocarbamate hybrids as multifunctional AChE inhibitors with β-amyloid anti-aggregation properties for the treatment of Alzheimer's disease

Article abstract of DOI:10.1016/j.bioorg.2019.103027

By connecting chromanone with dithiocarbamate moieties through flexible linkers, a series of hybrids as novel multifunctional AChE inhibitors have been designed and synthesized. Most of these compounds displayed strong and excellently selective inhibition

Full text of DOI:10.1016/j.bioorg.2019.103027

Accepted Manuscript
Novel chromanone-dithiocarbamate hybrids as multifunctional AChE inhibitors
with β-amyloid anti-aggregation properties for the treatment of Alzheimer’s
disease
Neng Jiang, Jiaoli Ding, Jing Liu, Xiaona Sun, Zhipeng Zhang, Zhongxia Mo,
Xiao Li, Hong Yin, Weizhong Tang, Sai-Sai Xie
PII:
S0045-2068(19)30594-2
https://doi.org/10.1016/j.bioorg.2019.103027
103027
DOI:
Article Number:
Reference:
YBIOO 103027
Bioorganic Chemistry
To appear in:
Received Date:
Revised Date:
Accepted Date:
22 April 2019
30 May 2019
30 May 2019
Please cite this article as: N. Jiang, J. Ding, J. Liu, X. Sun, Z. Zhang, Z. Mo, X. Li, H. Yin, W. Tang, S-S. Xie, Novel
chromanone-dithiocarbamate hybrids as multifunctional AChE inhibitors with β-amyloid anti-aggregation
properties for the treatment of Alzheimer’s disease, Bioorganic Chemistry (2019), doi: https://doi.org/10.1016/
j.bioorg.2019.103027
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Novel chromanone-dithiocarbamate hybrids as multifunctional AChE inhibitors
with β-amyloid anti-aggregation properties for the treatment of Alzheimer’s
disease
Neng Jiang‡, a,b, Jiaoli Ding‡, a, Jing Liuc, Xiaona Sunb, Zhipeng Zhanga, Zhongxia
Mob, Xiao Lib, Hong Yine, Weizhong Tang*, d, Sai-Sai Xie**, a
aNational Pharmaceutical Engineering Center for Solid Preparation in Chinese Herbal
Medicine, Jiangxi University of Traditional Chinese Medicine, Nanchang, PR China
bDepartment of Pharmacy, Affiliated Tumor Hospital of Guangxi Medical University,
Nanning, PR China
cSchool of Pharmacy, Jiangxi University of Traditional Chinese Medicine, Nanchang,
PR China
dDepartment of Gastrointestinal Surgery, Affiliated Tumor Hospital of Guangxi
Medical University, Nanning, PR China
eCollege of Chemistry & Pharmacy, Northwest A&F University, Yangling, PR China
*Authors for correspondence: tangweizhong@gxmu.edu.cn
**Authors for correspondence: xiesaisainanchang@hotmail.com
‡These authors contributed equally to this work.

Abstract: By connecting chromanone with dithiocarbamate moieties through flexible
linkers, a series of hybrids as novel multifunctional AChE inhibitors have been
designed and synthesized. Most of these compounds displayed strong and excellently
selective inhibition to eeAChE as well as potent inhibition to self- and AChE-induced
Aβ aggregation. Among them, compound 6c showed the best activity to inhibit
eeAChE (IC50 = 0.10 μM) and AChE-induced Aβ aggregation (33.02% at 100 μM),
and could effectively inhibit self-induced Aβ aggregation (38.25% at 25 μM). Kinetic
analysis and docking study indicated that compound 6c could target both the CAS and
PAS, suggesting that it was a dual binding site inhibitor for AChE. Besides, it
exhibited good ability to penetrate the BBB and low neurotoxicity in SH-SY5Y cells.
More importantly, compound 6c was well tolerated in mice (2500 mg/kg, po) and
could attenuate the memory impairment in a scopolamine-induced mouse model.
Overall, these results highlight 6c as a promising multifunctional agent for treating
AD and also demonstrate that the dithiocarbamate is a valid scaffold for design of
multifunctional AChE inhibitors.
Keywords: Alzheimer’s disease; chromanone; dithiocarbamate; acetylcholinesterase;
multifunctional agent.

1. Introduction
Alzheimer’s disease (AD) is a complex neurodegenerative disease that leads to severe
menace to human health characterized by memory loss, cognitive deficits and
behavioral abnormalities. [1]. Many AD patients lack daily self-care ability and need
long term home care, which increases patients’ family burden and influences the
development of the society to a great extent. The reports from the Alzheimer’s
Association indicate that the number of AD patients is now nearly 47 million and the
number is projected to increase to 100 million by 2050 all over the world [2]. The
fast-growing population of AD patients is considered to be severe and urgent health
issue for society.
It has been more than one hundred years since the first report about AD. However, the
etiopathogenesis of AD is still unclear, and the AD treatment remains a challenge for
the pharmaceutical community. Many factors and targets including deficits of
acetylcholine (ACh), amyloid-β (Aβ) peptide deposits and hyperphosphorylated tau
protein, etc. are regarded to be associated with the initiation and development of AD.
[3]. Over the past few decades, a lot of academic institutions and pharmaceutical
companies make great efforts to develop novel chemical molecular and biological
medicines for the treatment of AD. Unfortunately, most of these clinical trial anti-AD
drugs have been ended up with failure [4-5]. To date, the current clinical treatment of
AD mainly relies on the cholinergic hypothesis [6-7]. This hypothesis asserts that the
decline of ACh levels in brain leads to memory and cognitive impairments of AD
patient, and inhibiting the ACh metabolism can improve the ACh level, which

alleviate these symptoms. Based on this notion, four of the five drugs approved by the
FDA are cholinesterase inhibitors (Tacrine, Donepezil, Galantamine and Rivastigmine)
(Figure 1).
Acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) are two types of
cholinesterases that can break down ACh in the central nervous system (CNS) [8].
However, in contrast to AChE, the pathological mechanism of BuChE in AD is not
yet completely identified. Although a few studies suggest that BuChE may be also
beneficial to AD treatment, BuChE mainly distributes in peripheral tissues, so the
inhibition of BuChE may cause adverse effects in the peripheral system [9]. For
instance, tacrine, one of the first drugs approved for AD treatment with both
inhibitory activities of AChE and BuChE, was withdrawn due to the serious
hepatotoxicity and other side-effect in the peripheral system [10, 11]. Therefore,
development of selective AChE inhibitors is the more promising approach for AD
treatment. The crystallographic structure of AChE shows that the enzyme has two
binding active sites, which are connected by a nearly 20 Å deep narrow gorge. The
two binding active sites are a peripheral anionic site (PAS) near the entry of the gorge
and a catalytic active site (CAS) at the bottom of the gorge [12]. Studies show that the
structural features of AChE inhibitors should contain three parts: a CAS binding
moiety, a PAS binding moiety and a linker between these two moieties to fulfil the
structural requirements of AChE [13].
Apart from the cholinergic hypothesis, another hypothesis called the amyloid cascade
hypothesis suggests that Aβ peptide is also a causative factor in the course of AD

pathology [14, 15]. Aβ peptide consists of 39-43 amino acid residues, which is a
proteolytic product of the amyloid precursor protein (APP). Biochemical studies
reveal that the Aβ peptides can form monomers, oligomers and large Aβ plaques,
which are toxic to neuronal and mitochondrial functions, ultimately leading to
neurodegeneration and cognitive dysfunction [16]. Aβ1-40 and Aβ1-42 are the two main
isoforms of Aβ peptides. However, the latter is more prone to aggregate into fibrils
and more toxic to neuronal cells than the former [17]. Thus, inhibition of Aβ1-42
aggregation seems to be also important for AD treatment.
In fact, recent studies indicate that there is a close relationship between AChE and Aβ
aggregation. The AChE can promote Aβ aggregation and amyloid fibril formation
through its PAS [18, 19]. The interaction of Aβ and the PAS of AChE produces stable
AChE-Aβ complexes, and the complexes are more toxic than Aβ peptide aggregates.
Thus, developing dual binding site inhibitors, which can bind simultaneously to the
CAS and PAS of AChE, appears to be a very promising strategy for AD treatment,
because they can not only improve cognition by inhibiting AChE, but also slow the
Aβ aggregation contribute to AD pathogenesis [20].
Dithiocarbamate is a versatile pharmacophore in drug design, which has been
attracting considerable interest due to their diverse activities. The reported biological
activities of dithiocarbamate derivatives include anti-cancer, anti-bacteria and
carbonic anhydrase inhibition, etc [21-23]. However, compounds containing a
dithiocarbamate moiety were rarely reported to treat neurodegenerative disease, such
as AD. Recently, our group found that some dithiocarbamate derivatives could inhibit

AChE through the interactions between their dithiocarbamate moieties and the CAS
of AChE, and several of them also showed potential ability to inhibit self-induced Aβ
aggregation [24, 25]. Therefore, our recent research interests mainly focus on
developing dithiocarbamate derivatives as new AChE inhibitors with additional Aβ
anti-aggregation properties for the treatment of AD. Chromanones, a group of
naturally occurring compounds, are ubiquitous in nature, especially in plants. [26].
Recent years, they received much attention due to their wide use in designing
compounds for the treatment of AD. Some chromanone derivatives have been
demonstrated to possess potent AChE inhibitory activity, and the molecular docking
studies suggest that chromanone moiety can exert AChE inhibitory effect through
interacting with the PAS of AChE [27, 28].
Marketed AChE inhibitors
HO
O
O
O
NH2
O
Et
N
NMe2
O
O
Me
O
N
N
N
Tacrine
Donepezil
Rivastigmine
Galantamine
Design strategy
O
O
S
S
S
NH
O
OH
O
O
S
NH
n
Dithiocarbamate
7-hydroxychroman-4-one
CAS Binding
PAS Binding
Multifunctional AChE inhibitors
Figure 1 Marketed AChE inhibitors for the treatment of AD and design strategy for
chromanone-dithiocarbamate hybrids.
In light of the potential roles of dithiocarbamate and chromanone moieties mentioned

above in design of AChE inhibitors, in present work, our group attempted to connect
these two moieties by a flexible linker to produce a series of new chromanone-
dithiocarbamate hybrids as multifunctional AChE inhibitors for AD treatment (Figure
1). All designed compounds were evaluated in vitro to determine their ability to
inhibit AChE. The compounds with good AChE inhibitory activity and BBB
permeability were also tested to inhibit self- and AChE -induced Aβ aggregation.
Finally, the most promising compound fished out from the above assays was selected
for further studies including kinetic and molecular modeling analysis of its binding
mode with AChE, acute toxicity, cytotoxicity in SH-SY5Y neuroblastoma cells and
the neuroprotective effects on scopolamine-induced cognitive impairment in mice.
2. Results and Discussion
2.1 Chemistry
The syntheses of the target compounds are depicted in Scheme 1. The commercially
available resorcinol was reacted with 3-chloropropionic acid in the presence of
trifluoromethanesulphonic acid to obtain the intermediate 3, which was then cyclized
in aqueous NaOH to produce 7-hydroxychroman-4-one (4). Reaction of compound 4
with corresponding α, ω-dibromoalkanes using K2CO3 in acetone provided
intermediates 5a-d. Finally, following our previously reported method [24], the target
compounds 6-18 were accomplished in good yields by reacting compounds 5a-d with
the CS2, appropriate secondary amines and TEA in DMF.

O
O
O
OH
O
a
Cl
b
+
Cl
OH
OH
HO
OH
3
OH
1
4
2
O
O
O
6a-d
7a-18a
7b-18b
a
b-m
b-m
n = 3-6 R =
n = 4 R =
n = 5 R =
S
c
O
S
R
O
O
Br
n
n
5a-d
n = 3, 4, 5, 6
R=
a=
N
N
N
N
N
N
f=
e=
c=
i=
d=
b=
OH
N
N
N
N
N
N
g=
h=
j=
k=
l=
N
O
N
O
m=
N
Scheme 1 Synthesis of the target compounds 6-18. Reagents and conditions: (a) 3-
chloropropanoic acid, CF3SO3H, 80 oC, 30 min; (b) 2 M NaOH (aq), 5°C to r.t., 6 M
H2SO4 (aq), 2 h. (c) α, ω-dibromoalkanes, K2CO3, acetone, reflux, 4 h; (d) appropriate
secondary amines, CS2, TEA, DMF, r.t., 12 h.
2.2 In vitro eeAChE and eqBuChE inhibition studies
The inhibitory activities of the target compounds 6-18 against electric eel
acetylcholinesterase (eeAChE) and equine serum butyrylcholinesterase (eqBuChE)
were evaluated by the Ellman’s method. For comparison purposes, the marketed
drugs, donepezil and tacrine, were taken as reference compounds [29]. The inhibition
data are reported in Table 1 as IC50 (μM) or, for poorly active compounds, as the
percentage of inhibition at 10 μM. It can be seen from the table that all compounds

exhibit very weak or inactive inhibitory activity for eqBuChE, which suggest that our
present compounds are obviously selective inhibitors to eeAChE. Of these
compounds, compounds 6b and 6c exhibited the most potent inhibitory activity for
eeAChE, showing IC50 values of 0.12 and 0.10 μM, respectively, which were 4.5
times stronger than that of tacrine.
Table 1. Inhibition of eeAChE and eqBuChE by compounds 6-18.
O
S
O
O
S
R
n
Compd.
n
R
eeAChEa
eqBuChE Inhibition (%)b
IC50(μM)/Inhibition (%)
0.37 ± 0.07
6a
6b
3
4
5
6
4
5
4
5
4
5
4
5
4
5
4
5
a
a
a
a
b
b
c
c
d
d
e
e
f
15.03 ± 3.32
16.26 ± 2.45
14.38 ± 7.63
7.67 ± 1.21
14.15 ± 2.46
21.37 ± 10.05
n.a.c
0.12 ± 0.01
6c
0.10 ± 0.03
6d
0.92 ± 0.26
7a
3.99 ± 1.09
7b
45.43 ± 8.79%
0.24 ± 0.04
8a
8b
0.23 ± 0.03
18.93 ± 6.17
n.a.c
9a
0.59 ± 0.12
9b
0.54 ± 0.11
5.18 ± 1.03
23.02 ± 9.84
24.73 ± 4.51
13.84 ± 5.67
n.a.c
10a
10b
11a
11b
12a
12b
19.25 ± 2.68%
32.21 ± 9.15%
12.00 ± 4.05%
26.22 ± 6.34%
5.11 ± 0.99
f
g
g
21.92 ± 8.72
9.97 ± 1.06
3.02 ± 0.75

13a
13b
4
5
4
5
4
5
4
5
4
5
4
5
-
h
h
i
35.40 ± 3.64%
44.04 ± 10.20%
1.63 ± 0.78
8.79 ± 1.14
12.73 ± 5.13
6.19 ± 2.68
10.11 ± 1.42
5.67 ± 0.96
21.37 ± 11.35
n.a.c
14a
14b
i
0.94 ± 0.13
15a
j
1.16 ± 0.51
15b
j
0.51 ± 0.04
16a
k
k
l
18.16 ± 2.57%
13.74 ± 3.56%
13.20 ± 3.61%
23.15 ± 8.78%
10.15 ± 3.43%
10.32 ± 5.12%
0.042 ± 0.005
0.45 ± 0.03
16b
8.49 ± 2.61
5.79 ±1.49
17a
17b
l
11.53 ± 3.44
n.a.c
18a
m
m
-
18b
n.a.c
Donepezil
Tacrine
IC50 = 4.12 ± 0.87 μM
IC50 = 0.026 ± 0.002 μM
-
-
aThe 50% inhibitory concentration of eeAChE or percent inhibition with inhibitor at
10 μM (means ± SD of three experiments).
bThe percent inhibition of eqBuChE with inhibitor at 10 μM (means ± SD of three
experiments).
c
n. a. = no active. Compounds defined “no active” means that percent inhibition is
less than 5.0 % at a concentration of 10 μM in the assay conditions.
According to the previous studies, the linker length between the CAS and PAS
binding moieties plays a crucial role in AChE inhibition [30, 31]. Therefore, to
determine the optimal length in present study, compounds 6a-d with linker length
ranging from three to six carbon atoms (n = 3-6) were prepared in our initial step.
Biological evaluation indicated that compounds 6b and 6c presented more potent
inhibitory activities than other compounds, which suggested that the linkers

containing four or five carbon atoms seemed to be suitable for eeAChE inhibition.
After obtaining the optimal linker length, we next introduced different substituents to
the terminal group for exploring the SARs. As shown in the Table 1, introduction of a
methyl substituent to piperidine ring gave rise to a decrease in inhibiting eeAChE. All
compounds 7-9 bearing mono- or di-methyl substituents on piperidine ring showed
lower inhibitory activity than their no methyl analogues 6b and 6c. However, the
position of the methyl group on piperidine ring influenced the activity in different
degree. The position 4 seemed to be essential for the inhibitory activity. When it was
substituted by the methyl group, compounds 7a and 7b displayed a large drop in
eeAChE inhibition, especially for compound 7b, which contain a five-carbon atom
linker, showed a poor inhibitory activity, which was less than 50% at 10 μM. In
addition, a hydroxy group or a bulky group on this position of piperidine ring was also
not favorable for the inhibitory activity, as compounds 10a-b possessing a hydroxy
group and compounds 11a-b having a phenyl group exhibited a total loss activity on
eeAChE inhibition.
In order to further explore the SARs, the effects on replacing the piperidine group
with other secondary amines were also investigated. When the piperidine ring was
contracted to a pyrrole ring, the inhibitory activity was significantly reduced, as
compounds 12a and 12b only provided a micromolar inhibition for eeAChE.
Moreover, the inhibitory activities of compounds 13-15 substituted with alkyl amines
were continuously improved with gradually increasing the alkyl groups. Compounds
15a-b with a diisopropylamine group presented more potent inhibitory activity than

its counterparts 13a-b and 14a-b. Finally, replacement of the piperidine with 4-
substituted piperizine or with a N-methylbenzylamine could not improve the
inhibitory activity, as compounds 16-18 did not give any inhibition for eeAChE.
2.3 Kinetic study of eeAChE inhibition
To gain insight into the inhibition mode of the present compound for eeAChE,
compound 6c with the most potent activity for eeAChE inhibition was selected for
kinetic study. It can be seen from the Figure 2 that the Lineweaver-Burk reciprocal
plots show both increased slopes (decreased Vmax) and intercepts (higher Km) at
increasing concentration of the inhibitor, which indicated a mixed-type inhibition.
These results implied that compound 6c might be a dual binding site AChE inhibitor,
which could target the catalytic active site (CAS) and the peripheral anionic site (PAS)
of AChE simultaneously.
Figure 2. Kinetic study on the mechanism of eeAChE inhibition by compound 6c.
Overlaid Lineweaver-Burk reciprocal plots of eeAChE initial velocity at increasing
substrate concentration (0.05-0.50 mM) in the absence of inhibitor and in the presence

of 6c are shown. Lines were derived from a weighted least-squares analysis of the
data points.
2.4 Molecular modeling studies of AChE
To further confirm the binding mode of compound 6c with AChE, docking studies
were performed using a Molecular Operating Environment (MOE 2015.10) software
package. The structure of TcAChE co-crystallized with donepezil (PDB code 1EVE)
was used to establish the starting model of AChE. As shown in the Figure 3,
compound 6c fits well in the active site of AChE, which can occupy the entire
enzymatic CAS, mid-gorge site and PAS. The chromanone moiety located at the PAS
of the enzyme, forming a π-π stacking interaction with the indole ring of Trp 279
(3.48 Å). Similar to our previous result [24, 25], the piperidinyldithiocarbamate
moiety of compound 6c was bound to the CAS through hydrophobic interactions with
residues Trp 84, Asn 85, Ser122 and Gln 69. Moreover, the polymethylene chain was
folded in a conformation in the mid-gorge, which allowed it to establish hydrophobic
interactions with Phe330 and Tyr 334. All in all, this docking study demonstrated that
compound 6c was a dual binding site inhibitor of AChE, which was in agreement with
our result from kinetic analysis.

Figure 3. (a) 3D docking model of compound 6c with TcAChE. Atom colours: cyan-
carbon atoms of compound 6c, grey-carbon atoms of residues of TcAChE, dark blue-
nitrogen atoms, red-oxygen atoms, yellow-sulfur atom. (b) 2D schematic diagram of
docking model of compound 6c with TcAChE. The figure was prepared using the
ligand interactions application in MOE. The green dashed line represents the
interaction between the protein and the ligand.
2.5 Blood-brain barrier (BBB) permeation assay
Figure 4. Lineal correlation between experimental and reported permeability of
commercial drugs using the PAMPA-BBB assay. Pe (exp.) = 0.9075 Pe (bibl.) -

0.2016 (R2 = 0.9654).
The ability to cross the blood-brain barrier (BBB) is a crucial factor for successful
CNS drugs [32]. Thus, we performed a parallel artificial membrane permeation assay
of blood-brain barrier (PAMPA-BBB) to investigate whether the present compounds
could penetrate the BBB. This assay stablished by Di et al. is a simple and rapid
method to predict passive BBB permeation [33]. Compounds 6a-d, 8a-b, 9a-b, 14b
and 15b with sub-micromolar inhibitory activity for AChE were selected for test.
After comparing experimental permeabilities of 9 commercial drugs with reported
values (Table 2), a plot of experiment data versus the bibliographic values gave a
good linear correlation: Pe (exp.) = 0.9075 Pe (bibl.) - 0.2016 (R2 = 0.9654) (Figure
4). From this equation and taking into account the limits established by Di et al. for
BBB permeation, we classified compounds as follows：
(a) ‘CNS+’ (high BBB permeation predicted): Pe (10-6 cm/s) > 3.43
(b) ‘CNS-’ (low BBB permeation predicted): Pe (10-6 cm/s) < 1.61
(c) ‘CNS±’ (low BBB permeation predicted): 1.61 < Pe (10-6 cm/s) < 3.43
According to the measured permeabilities shown in Table 2, all compounds exhibited
the Pe values higher than 3.43, which indicated that these selected compounds could
cross the BBB and exert the biological effects in the CNS.
-6
Table 2. Permeability Pe (×10 ) in the PAMPA-BBB assay for 9 commercial drugs
in the experiment validation.
Pe (×10 -6 cm/s)
Commercial
Bibliographya
Experimentb
Compd.

drugs
Testosterone
Estradiol
17.0
12.0
9.3
6.5
5.1
1.9
1.3
1.02
0.1
-
14.8 ± 0.85
9.83 ± 0.73
9.52 ± 0.62
7.55 ± 0.54
3.56 ± 0.62
0.63 ± 0.06
1.12 ± 0.24
0.21 ± 0.02
0.17 ± 0.01
-
6a
6b
6c
7.55 ± 0.64
7.67 ± 0.82
7.83 ± 0.51
7.62 ± 0.98
8.14 ± 0.73
7.96 ± 0.61
7.24 ± 0.39
8.02 ± 0.76
7.26 ± 0.36
7.91 ± 0.45
Progesterone
Chlorpromazine
Corticosterone
Hydrocortisone
Caffeine
6d
8a
8b
9a
Atenolol
9b
14b
15b
Theophylline
-
a Taken for Ref. [33]
b Experimental data are expressed as mean ± SD from three independent experiments,
using PBS: EtOH (70:30) as solvent.
2.6 Inhibition studies of self-induced Aβ aggregation
As all selected compounds could cross the BBB, we next tested their ability to inhibit
amyloid-β (Aβ) aggregation. The abilities of compounds to inhibit self-induced Aβ1-42
aggregation was evaluated using a thioflavin T (ThT) fluorometric assay [34, 35].
Curcumin, a well-known natural inhibitor for Aβ1-42 self-aggregation, was taken as
reference compound. From the results summarized in Table 3, it can be seen that
these compounds exhibit moderate to good potencies (15.04-38.56% at 25 μM)
compared with curcumin (38.59% at 25 μM). Among these compounds, compounds
6b, 6c and 9a (38.56%, 38.25%, and 38.38% at 25 μM, respectively) exhibited the
best inhibitory activity for Aβ1-42 aggregation, which were comparable to that of
curcumin.

Table 3. Inhibition of AChE-induced and self-induced Aβ1-42 aggregation by selected
compounds.
Compd.
Inhibition of Aβ1-42 aggregation (%)
Self-induceda
AChE-inducedb
19.67 ± 1.02
30.46 ± 2.19
33.02 ± 3.55
14.75 ± 2.53
27.17 ± 2.16
29.32 ± 3.10
23.84 ± 2.14
25.13 ± 3.27
15.25 ± 1.03
18.22 ± 2.04
27.13 ± 1.93
-
6a
18.43 ± 2.87
38.56 ± 5.62
38.25 ± 3.84
20.11 ± 2.02
36.24 ± 1.96
35.76 ± 5.37
38.38 ± 2.18
37.24 ± 3.44
15.04 ± 1.10
19.11 ± 0.93
-
6b
6c
6d
8a
8b
9a
9b
14b
15b
Donepezil
Curcumin
38.59 ± 2.17
a
Inhibition of self-induced Aβ1-42 aggregation (means ± SD of three experiments).
The thioflavin-T fluorescence method was used, and the measurements were carried
out in the presence of 25 μM inhibitor.
b Inhibition of AChE-induced Aβ1-42 aggregation (means ± SD of three experiments).
The thioflavin-T fluorescence method was used, and the concentration of the tested
inhibitor was 100 μM.
2.7 Inhibition studies of AChE-induced Aβ aggregation
Accumulating evidences indicate that the PAS of AChE can bind to Aβ and promote
the formation of amyloid fibrils. Therefore, inhibition of AChE, especially for the
inhibition of PAS of AChE, may reduce Aβ aggregation. Since the studies above have

proved that our present compounds were able to bind both CAS and PAS of AChE,
compounds 6a-d, 8a-b, 9a-b, 14b and 15b were also selected to detect their inhibitory
activities against AChE-induced Aβ1-42 aggregation. The inhibitory activities were
also determined by the ThT method, and a dual binding site inhibitor of AChE,
donepezil, was chosen as reference compound [36, 37]. As shown in the Table 3,
most compounds exhibit effectively inhibitory activity for AChE-induced Aβ1-42
aggregation with inhibition rate ranging from 14.75 to 33.02% at 100 μM in
comparison to donepezil (27.13%). Compounds 6b and 6c possessing higher
inhibitory activity for eeAChE also showed stronger activity to inhibit the aggregation.
Compound 6c displayed the strongest inhibitory activity in this series with inhibition
rate of 33.02%, which was more potent than that of donepezil. After completing all
above biological evaluations, compound 6c with good BBB permeability and potent
inhibitory activity to eeAChE and self- and AChE-induced Aβ aggregation was
selected as a promising compound for further study.
2.8 Cytotoxicity of human neuroblastoma SH-SY5Y cells
Figure 5. Cytotoxicity of compound 6c and donepezil on human neuroblastoma cells
SH-SY5Y. SH-SY5Y cells were incubated with different concentrations of compound

6c or donepezil (12.5 – 100 µM) for 24 h. The results are shown as the percentage of
viable cells after treatment with compound 6c or donepezil vs untreated control cells.
Date are expressed as mean ± SD from three independent experiments.
To check the potential toxicity of compound 6c to nerve cells, its cytotoxicity was
evaluated using the human neuroblastoma SH-SY5Y cell line, and donepezil was
taken as reference compound. After treating the cells with 6c or donepezil at different
concentrations (12.5, 25, 50 and 100 μM) for 24 h, the cell viability was determined
by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium (MTT) assay. [38] As shown
in Figure 5, similar to donepezil, compound 6c did not cause any toxicity to SH-
SY5Y cells even at the highest concentration, which indicated that it might be a safe
multifunctional inhibitor for AD treatment.
2.9 Acute oral toxicity studies
To further investigate the safety property of compound 6c, an acute toxicity
evaluation was performed in KM mice (18-25 g, n = 10 per group, half male and half
female) [24, 25, 39]. After delivering compound 6c to mice with various dosages (625,
1250 and 2500 mg/kg) by oral administration, no abnormal behavior and acute
toxicity, such as death, marked weight loss, and drastically altered consumption of
water or food, were detected during the experimental period. In addition, the possible
toxic damage on organs of the mice was also examined after sacrificing these animals
on the 14th day. The results indicated that no toxic effect was observed on heart, liver
or kidneys, which demonstrated that compound 6c was well tolerated at a dose up to

2500 mg/kg.
2.10 In vivo step-down passive avoidance assay
Given the excellent results obtained from above experiments, we next performed a
step-down passive avoidance test to determine whether compound 6c could improve
memory impairment in vivo. The scopolamine was used as an inducer to establish the
cognitive deficit mouse model and donepezil was selected as the positive control [40,
41]. It can be seen from the Figure 6 that model group show significantly reduced the
latency and increased the number of errors in comparison to the control group (###p <
0.001). When mice were treated with compound 6c, the step-down latency and
number of errors were reversed in a dose-dependent manner. The high dose group
(83.2 mg/kg) showed the longest latency (221.0) and the least number of errors (1)
among these groups, which were better than those of donepezil group (5 mg/kg, 200.9
s, 1.1). Meanwhile, the medium dose group (41.6 mg/kg) also exhibited the effects
comparable (201.3 s; 1.2) to those of donepezil. Finally, compared with model group,
although the low dose group (20.8 mg/kg) did not showed a significant difference, the
latency and number of errors were largely improved (155.2 s vs 123.0 s, 1.9 vs 2.7).
Altogether, the above results indicated that compound 6c could attenuate the cognitive
deficit through increasing the cholinergic activity in brain by inhibition of AChE.

Figure 6. Effects of compound 6c on the (a) latency (s) and (b) number of errors in
the step-down test by the scopolamine-induced cognitive impairment. The data shown
are mean ± SD (n = 8). ###P < 0.001 vs. control group, *P < 0.05, **P < 0.01 vs.
model group.
3. Conclusion
A series of new hybrid molecules by connecting chromanone with dithiocarbamate
moieties through flexible linkers have been designed and synthesized as
multifunctional AChE inhibitors for the treatment of AD. Biological assays indicated
that most of these compounds presented potent inhibitory activity and high selectivity
for eeAChE. Of these compounds, compound 6c stood out as the most potent inhibitor
to eeAChE, showing the IC50 values of 0.10 μM, which was 4.5-fold more potent than
that of tacrine. Kinetic analysis revealed that compound 6c was a mix-type inhibitor
of AChE. Further docking studies confirmed that 6c was a dual binding site inhibitor,
which could interact simultaneously with the PAS and CAS of AChE. Besides,
compound 6c also displayed the best activity to inhibit AChE-induced Aβ aggregation

and potent activity to inhibit self-induced Aβ aggregation. Meanwhile, it could cross
the BBB and showed good safety profiles in both cell and animal tests. At last, the
step-down passive avoidance assay indicated that compound 6c could significantly
ameliorate the cognitive dysfunction induced by scopolamine in mice (83.2 and 41.6
mg/kg, p.o.). Overall, the present results suggest that compound 6c is a potential
disease-modifying agent for treating AD, and the dithiocarbamate moiety could be
considered as a valid scaffold for design of multifunctional AChE inhibitors.
4. Experimental section
4.1 Chemistry
All chemicals and solvents were obtained from commercial sources and were used
without purification. Reactions were monitored using thin-layer chromatography
(TLC) on glass-packed precoated silica gel plates and visualized with a UV lamp.
Column chromatography was performed using silica gel (200-300 mesh) purchased
from Qingdao Haiyang Chemical Co. Ltd. Melting points were measured on an XT-4
micro melting point apparatus and were uncorrected. 1H NMR spectra (500 MHz) and
13C NMR spectra (126 MHz) were recorded on a Bruker ACF-600 spectrometer at
room temperature using CDCl3 or DMSO-d6 as the solvent. Chemical shifts (δ) are
reported in parts per million (ppm) using the tetramethylsilane (TMS) as internal
standard. The coupling constants J are presented in hertz (Hz). The ESI-MS spectra
were obtained on LCQ Fleet LC-MS System (Thermo Fisher Scientific, USA). The
high-resolution mass spectra (HRMS) were recorded with a TripleTOF 5600 System
(AB SCIEX). The purity (>95%) of all final compounds was determined by HPLC,

conducted on a Shimadzu LC-20AT series system, TC-C18 column (4.6 mm × 150
mm, 5 μm), eluted with MeOH/H2O = 85/15, at a flow rate of 1.0 mL/min.
4.2 3-chloro-1-(2,4-dihydroxyphenyl)propan-1-one (3)
To a stirred mixture of resorcinol (5.0 g, 45.4 mmol) and 3-chloropropionic acid (4.9
g, 45.4 mmol), trifluoromethanesulfonic acid (13.6 g, 90.8 mmol) was added. After
o
heating the mixture at 80 C for 30 min, the reaction mixture was allowed to cool to
room temperature and poured into ice cooled water (100 mL). Then, the mixture was
extracted with Cl2CH2. The combined organic phase was washed with brine, dried
over anhydrous Na2SO4 and concentrated under vacuum to afford crude product as
orange oil. Without further purification, the crude product was directly used in next
step.
4.3 7-hydroxychroman-4-one (4)
To a cooled 2 N aqueous NaOH was added crude compound 3. The resulting solution
o
was stirred at room temperature for 2 h and then cooled to 5 C. The solution was
acidified with 6 M H2SO4 to pH = 2. The mixture was extracted with ethyl acetate.
The combined organic phase was washed with brine, dried over anhydrous Na2SO4
and concentrated under vacuum to afford crude product as brown solid. Yield 62%;
brown solid; 1H NMR (500 MHz, CDCl3) δ 7.83 (d, J = 8.7 Hz, 1H, Ar-H), 6.54 (dd,
J = 8.7, 2.3 Hz, 1H, Ar-H), 6.41 (d, J = 2.3 Hz, 1H, Ar-H), 4.57 – 4.46 (m, 2H, -
OCH2-), 2.84 – 2.72 (m, 2H, -COCH2-); 13C NMR (126 MHz, CDCl3) δ 191.4, 164.2,
163.2, 129.7, 115.4, 110.7, 103.3, 67.4, 37.5.
4.4. General procedure for synthesis of compounds 5a-d.

Compound 4 (5.0 mmol) and anhydrous K2CO3 (1.4 g, 10 mmol) were suspended in
acetone (15 mL). Then, suitable α, ω-dibromoalkanes (50 mmol) was added, and the
mixture was stirred under reflux for 4 h. When the reaction was completed, the
reaction mixture was filtered, and the filtrate was concentrated under reduced pressure.
The obtained residue was purified by silica gel chromatography with PE/EA (16:1) as
eluent to give compounds 5a-d.
4.4.1. 7-(3-bromopropoxy)chroman-4-one (5a)
Yeild 75%; white solid; m.p. 86-88 oC; 1H NMR (500 MHz, CDCl3) δ 7.81 (d, J = 8.8
Hz, 1H, Ar-H), 6.54 (dd, J = 8.8, 2.3 Hz, 1H, Ar-H), 6.37 (d, J = 2.3 Hz, 1H, Ar-H),
4.59 – 4.39 (m, 2H, -OCH2-), 4.09 (t, J = 6.4 Hz, 2H, -OCH2-), 3.60 (t, J = 6.0 Hz, 2H,
13
-CH2Br), 2.86 – 2.67 (m, 2H, -COCH2-), 2.35 – 2.29 (m, 2H, -CH2-). C NMR (126
MHz, CDCl3) δ 190.6, 165.3, 163.8, 129.0, 115.3, 110.3, 101.3, 67.4, 67.1, 37.5, 34.9,
30.1.
4.4.2 7-(4-bromobutoxy)chroman-4-one (5b)
Yeild 69%; white solid; m.p. 58-60 oC ; 1H NMR (500 MHz, CDCl3) δ 7.80 (d, J = 8.8
Hz, 1H, Ar-H), 6.54 (dd, J = 8.8, 2.3 Hz, 1H, Ar-H), 6.37 (d, J = 2.3 Hz, 1H, Ar-H),
4.49 (t, J = 6.4 Hz, 2H, -OCH2-), 4.00 (t, J = 6.0 Hz, 2H, -OCH2-), 3.46 (t, J = 6.5 Hz,
2H, -CH2Br), 2.87–2.64 (m, 2H, -COCH2-), 2.07 – 2.01 (m, 2H, -CH2-), 1.96 – 1.91
(m, 2H, -CH2-).13C NMR (126 MHz, CDCl3) δ 190.6, 165.3, 163.8, 129.0, 115.4,
110.2, 101.3, 67.4, 67.3, 37.5, 33.3, 29.4, 27.7.
4.4.3. 7-((5-bromopentyl)oxy)chroman-4-one (5c)

Yeild 62%; white solid; m.p. 56-58 oC ; 1H NMR (500 MHz, CDCl3) δ 7.81 (d, J = 8.8
Hz, 1H, Ar-H), 6.55 (dd, J = 8.8, 2.3 Hz, 1H, Ar-H), 6.37 (d, J = 2.3 Hz, 1H, Ar-H),
4.60 – 4.38 (m, 2H, -OCH2-), 3.98 (t, J = 6.3 Hz, 2H, -OCH2-), 3.42 (t, J = 6.7 Hz, 2H,
-CH2Br), 2.87 – 2.58 (m, 2H, -COCH2-), 1.95 – 1.89 (m, 2H, -CH2-), 1.84 – 1.78 (m,
2H, -CH2-), 1.60 (p, J = 7.6 Hz, 2H, -CH2-).13C NMR (126 MHz, CDCl3) δ 190.6,
165.4, 163.9, 128.9, 115.3, 110.3, 101.3, 68.1, 67.4, 37.5, 33.6, 32.4, 28.2, 24.8.
4.4.4. 7-((6-bromohexyl)oxy)chroman-4-one (5d)
Yeild 67%; white solid; m.p. 54-56 oC; 1H NMR (500 MHz, CDCl3) δ 7.81 (d, J = 8.8
Hz, 1H, Ar-H), 6.55 (dd, J = 8.8, 1.7 Hz, 1H, Ar-H), 6.37 (d, J = 1.9 Hz, 1H, Ar-H),
4.50 (t, J = 6.4 Hz, 2H, -OCH2-), 3.97 (t, J = 6.8 Hz, 2H, -OCH2-), 3.41 (t, J = 6.7 Hz,
2H, -CH2Br), 2.74 (t, J = 5.9 Hz, 2H, -COCH2-), 1.88 (p, J = 6.5 Hz, 2H, -CH2-), 1.80
(p, J = 6.4 Hz, 2H, -CH2-), 1.55 – 1.42 (m, 4H, -CH2-).13C NMR (126 MHz, CDCl3) δ
190.6, 165.5, 163.9, 129.0, 115.3, 110.3, 101.3, 68.3, 67.5, 37.5, 33.9, 32.7, 28.9, 28.0,
25.3.
4.5 General procedure for synthesis of compounds 6-18.
To a solution of the secondary amine (1.2 mmol) and TEA (131 mg, 1.3 mmol) in
DMF (2 mL) was added CS2 (99 mg, 1.3 mmol) dropwise. After stirring the mixture
for 5 min, appropriate chromanone derivatives 5a-d (1.2 mmol) dissolved in DMF (3
mL) was added, and the mixture was stirred at room temperature for 12 h. After
completion, the mixture was diluted with 20 mL of water and extracted with ethyl
acetate. The combined organic phase was washed with water (15 mL × 3), dried over
anhydrous Na2SO4, concentrated under vacuum, and purified by silica gel

chromatography with PE/EA (10:1) as eluent to give the desire compounds 6-18.
4.5.1 3-((4-oxochroman-7-yl)oxy)propyl piperidine-1-carbodithioate (6a)
Yield 85%; white solid; m. p. 127- 129 oC; 1H NMR (500 MHz, CDCl3) δ 7.76 (d, J =
8.8 Hz, 1H, Ar-H), 6.51 (dd, J = 8.8, 2.4 Hz, 1H, Ar-H), 6.34 (d, J = 2.4 Hz, 1H, Ar-
H), 4.45 (t, J = 6.4 Hz, 2H, -OCH2-), 4.23 (br s, 2H, -NCH2-), 4.03 (t, J = 6.1 Hz, 2H,
-OCH2-), 3.81 (d, J = 8.0 Hz, 2H, -NCH2-), 3.41 (t, J = 7.0 Hz, 2H, -SCH2-), 2.67 (t, J
13
= 6.4 Hz, 2H, -COCH2-), 2.18 – 2.12 (m, 2H, -CH2-), 1.63 (br s, 6H, -(CH2)3-). C
NMR (126 MHz, CDCl3) δ 194.1, 189.5, 164.2, 162.8, 127.9, 114.3, 109.3, 100.3,
66.4, 65.8, 51.8, 50.3, 36.4, 32.3, 28.7, 23.3; ESI-MS m/z: 365.96 [M+H]+; HRMS:
calcd for C18H24NO3S2 [M+H]+ 366.1192, found 366.1189. HPLC purity, 98.39%.
4.5.2 4-((4-oxochroman-7-yl)oxy)butyl piperidine-1-carbodithioate (6b)
o
1
Yield 82%; White solid; m. p. 95-97 C; H NMR (500 MHz, CDCl3) δ 7.83 (d, J =
8.8 Hz, 1H, Ar-H), 6.57 (dd, J = 8.8, 2.4 Hz, 1H, Ar-H), 6.39 (d, J = 2.3 Hz, 1H, Ar-
H), 4.51 (t, J = 6.5 Hz, 2H, -OCH2-), 4.31 – 4.29 (m, 2H, -NCH2-), 4.02 (t, J = 6.5 Hz,
2H, -OCH2-), 3.89 (br s, 2H, -NCH2-), 3.38 (t, J = 6.5 Hz, 2H, -SCH2-), 2.75 (t, J =
6.5 Hz, 2H, -COCH2-), 1.94 – 1.87 (m, 4H, -(CH2)2-), 1.73 – 1.68 (m, 6H, -
(CH2)3-).13C NMR (126 MHz, CDCl3) δ 195.7, 190.7, 165.5, 163.9, 129.0, 115.3,
110.4, 101.3, 67.9, 67.5, 53.1, 51.4, 37.6, 36.7, 28.4, 25.6, 24.5; ESI-MS m/z: 380.03
[M+H]+; HRMS: calcd for C19H26NO3S2 [M+H]+ 380.1348, found 380.1346. HPLC
purity, 97.32%.
4.5.3 5-((4-oxochroman-7-yl)oxy)pentyl piperidine-1-carbodithioate (6c)

o
1
Yield 87%; white solid; m. p. 70-71 C; H NMR (500 MHz, CDCl3) δ 7.82 (d, J =
8.8 Hz, 1H, Ar-H), 6.57 (dd, J = 8.8, 2.4 Hz, 1H, Ar-H), 6.38 (d, J = 2.4 Hz, 1H, Ar-
H), 4.50 (t, J = 6.4 Hz, 2H, -OCH2-), 4.25 (br s, 2H, -NCH2-), 3.98 (t, J = 6.4 Hz, 2H,
-OCH2-), 3.91 (br s, 2H, -NCH2-), 3.32 (t, J = 6.5 Hz, 2H, -SCH2-), 2.74 (t, J = 6.0 Hz,
2H, -COCH2-), 1.85 – 1.56 (m, 12H, -(CH2)6-).13C NMR (126 MHz, CDCl3) δ 195.8,
190.6, 165.5, 163.8, 128.9, 115.2, 110.3, 101.2, 68.2, 67.4, 52.9, 51.4, 37.5, 36.9, 28.6,
25.4, 24.4; ESI-MS m/z: 394.07 [M+H]+; HRMS: calcd for C20H28NO3S2 [M+H]+
394.1505, found 394.1506. HPLC purity, 98.55%.
4.5.4 3 6-((4-oxochroman-7-yl)oxy)hexyl piperidine-1-carbodithioate (6d)
Yield 87%; yellow oil; 1H NMR (500 MHz, CDCl3) δ 7.76 (d, J = 8.8 Hz, 1H, Ar-H),
6.50 (dd, J = 8.8, 2.4 Hz, 1H, Ar-H), 6.32 (d, J = 2.4 Hz, 1H, Ar-H), 4.44 (t, J = 6.4
Hz, 2H, -OCH2-), 4.21 (d, J = 6.6 Hz, 2H, -NCH2-), 3.91 (t, J = 6.5 Hz, 2H, -OCH2-),
3.84 (br s, 2H, -NCH2-), 3.24 (t, J = 7.5 Hz, 2H, -SCH2-), 2.68 (t, J = 6.5 Hz, 2H, -
COCH2-), 1.80 – 1.60 (m, 10H, -(CH2)5-), 1.44 – 1.41 (m, 4H, -(CH2)2-).13C NMR
(126 MHz, CDCl3) δ 194.9 , 189.6, 164.5, 162.8, 127.8, 114.1, 109.3, 100.2, 67.3,
66.4, 51.8, 50.3, 36.4, 36.0, 27.8, 27.7, 24.6, 23.3; ESI-MS m/z: 408.01[M+H]+;
HRMS: calcd for C21H30NO3S2 [M+H]+ 408.1661, found408.1672. HPLC purity,
98.83%.
4.5.5. 4-((4-oxochroman-7-yl)oxy)butyl 4-methylpiperidine-1-carbodithioate (7a)
Yield 87%; white solid; m. p. 60 - 61oC; 1H NMR (500 MHz, DMSO-d6) δ 7.68 (d, J
= 8.8 Hz, 1H, Ar-H), 6.63 (dd, J = 8.8, 2.3 Hz, 1H, Ar-H), 6.53 (d, J = 2.3 Hz, 1H,
Ar-H), 4.50 (t, J = 6.4 Hz, 2H, -OCH2-), 4.06 (t, J = 6.0 Hz, 2H, -OCH2-), 3.29 (br s,

4H, -NCH2-), 2.71 (t, J = 6.4 Hz, 2H, -COCH2-), 1.83 – 1.72 (m, 8H, -(CH2)4-), 1.24
(s, 1H, -CH-), 1.19 – 1.04 (m, 2H, -CH2-), 0.92 (s, 3H, -CH3). 13C NMR (126 MHz,
DMSO-d6) δ 194.5, 190.6, 165.3, 164.0, 128.7, 115.4, 110.6, 101.9, 68.2, 67.7, 52.1,
50.5, 37.5, 36.4, 30.6, 28.2, 25.7, 21.7; ESI-MS m/z: 394.05 [M+H]+; HRMS: calcd
for C20H28NO3S2 [M+H]+ 394.1505, found 394.1516. HPLC purity, 96.43%.
4.5.6. 5-((4-oxochroman-7-yl)oxy)pentyl 4-methylpiperidine-1-carbodithioate (7b)
o
1
Yield 87%; white solid; m. p. 60-62 C; H NMR (500 MHz, CDCl3) δ 7.81 (d, J =
8.8 Hz, 1H, Ar-H), 6.55 (dd, J = 8.8, 2.3 Hz, 1H, Ar-H), 6.37 (d, J = 2.3 Hz, 1H, Ar-
H), 4.49 (t, J = 6.4 Hz, 2H, -OCH2-), 3.97 (t, J = 6.4 Hz, 2H, -OCH2-), 3.30 (br s, 2H,
-NCH2-), 3.13 – 3.05 (m, 2H, -NCH2-), 2.73 (t, J = 6.4 Hz, 2H, -COCH2-), 1.85 – 1.68
(m, 8H, -(CH2)4-), 1.60 – 1.55 (m, 2H, -CH2-), 1.26 (br s, 3H, -CHCH2-), 0.96 (s, 3H,
-CH3). 13C NMR (126 MHz, CDCl3) δ 195.9, 190.6, 165.5, 163.8, 128.9, 115.2, 110.3,
101.2, 68.2, 67.4, 52.1, 50.4, 37.5, 37.0, 31.0, 28.6, 28.5, 25.4, 21.3. ESI-MS m/z:
408.08 [M+H]+; HRMS: calcd for C21H30NO3S2 [M+H]+408.1661, found 408.1660.
HPLC purity, 96.16%.
4.5.7. 4-((4-oxochroman-7-yl)oxy)butyl 2-methylpiperidine-1-carbodithioate (8a)
o
1
Yield 87%; white solid; m. p. 46-47 C; H NMR (500 MHz, CDCl3) δ 7.85 (d, J =
8.8 Hz, 1H, Ar-H), 6.59 (dd, J = 8.8, 2.4 Hz, 1H, Ar-H), 6.41 (d, J = 2.4 Hz, 1H, Ar-
H), 4.53 (t, J = 6.0 Hz, 2H, -OCH2-), 4.05 (t, J = 6.0 Hz, 2H, -OCH2-), 3.41 (t, J = 7.0
Hz, 2H, -NCH2-), 3.15 (br s, 1H, -NCH-), 2.77 (t, J = 7.2 Hz, 2H, -COCH2-), 1.98 –
1.88 (m, 4H, -(CH2)2-), 1.80 – 1.63 (m, 8H, -(CH2)4-), 1.29 (d, J = 7.0 Hz, 3H, -CH3).
13C NMR (126 MHz, CDCl3) δ 195.9, 190.6, 165.4, 163.8, 128.9, 115.2, 110.3, 101.2,

67.8, 67.4, 54.0, 37.4, 36.4, 28.3, 25.4, 18.7. ESI-MS m/z: 394.05 [M+H]+; HRMS:
calcd for C20H27NO3S2Na [M+Na]+ 416.1325, found 416.1339. HPLC purity, 95.99%.
4.5.8. 5-((4-oxochroman-7-yl)oxy)pentyl 2-methylpiperidine-1-carbodithioate (8b)
o
1
Yield 87%; white solid; m. p. 46-48 C; H NMR (500 MHz, CDCl3) δ 7.76 (d, J =
8.8 Hz, 1H, Ar-H), 6.50 (dd, J = 8.8, 2.4 Hz, 1H, Ar-H), 6.32 (d, J = 2.3 Hz, 1H, Ar-
H), 4.44 (t, J = 6.5 Hz, 2H, -OCH2-), 3.93 (t, J = 6.4 Hz, 2H, -OCH2-), 3.25 (t, J = 7.5
Hz, 2H, -NCH2-), 3.06 (br s, 1H, -NCH-), 2.69 (t, J = 6.5 Hz, 2H, -COCH2-), 1.80 –
1.65 (m, 8H, -(CH2)4-), 1.62 – 1.49 (m, 6H, -(CH2)3-), 1.21 (d, J = 7.0 Hz, 3H, -CH3).
13C NMR (126 MHz, CDCl3) δ 195.1, 189.5, 164.5, 162.8, 127.8, 114.1, 109.2, 100.2,
67.1, 66.3, 52.7, 36.4, 35.7, 27.5, 27.4, 24.4, 17.6; ESI-MS m/z: 408.05 [M+H]+;
HRMS: calcd for C21H30NO3S2 [M+H]+408.1662, found 408.1681. HPLC purity,
98.53%.
4.5.9. 4-((4-oxochroman-7-yl)oxy)butyl 2,6-dimethylpiperidine-1-carbodithioate (9a)
o
1
Yield 87%; white solid; m. p. 81-82 C; H NMR (500 MHz, CDCl3) δ 7.85 (d, J =
8.8 Hz, 1H, Ar-H), 6.59 (dd, J = 8.9, 2.3 Hz, 1H, Ar-H), 6.41 (d, J = 2.3 Hz, 1H, Ar-
H), 4.53 (t, J = 6.4 Hz, 2H, -OCH2-), 4.05 (t, J = 5.9 Hz, 2H, -OCH2-), 3.48 – 3.32 (m,
2H, -N(CH)2-), 2.77 (t, J = 6.4 Hz, 2H, -COCH2-), 2.00 – 1.52 (m, 12H, -(CH2)6-),
1.37 (d, J = 7.0 Hz, 3H, -CH3), 1.33 (d, J = 7.1 Hz, 3H, -CH3). 13C NMR (126 MHz,
CDCl3) δ 196.6, 190.6, 165.4, 163.8, 128.9, 115.2, 110.3, 101.2, 67.8, 67.4, 53.6, 52.8,
37.4, 36.4, 30.4, 30.2, 28.3, 25.3, 19.8, 18.7, 14.0. ESI-MS m/z: 408.03 [M+H]+;
HRMS: calcd for C21H30NO3S2 [M+H]+ 408.1662, found 408.1680. HPLC purity,
95.48%.