Discovery of potent and selective acetylcholinesterase (AChE) inhibitors: acacetin 7-O-methyl ether Mannich base derivatives synthesised from easy access natural product naringin

Article abstract of DOI:10.1080/14786419.2017.1340280

Naringin, as a component universal existing in the peel of some fruits or medicinal plants, was usually selected as?the material to synthesise bioactive derivates since it was easy to gain with low cost. In present investigation, eight new acacetin-7-O-methyl ether Mannich base derivatives (1–8) were synthesised from naringin. The bioactivity evaluation revealed that most of them exhibited moderate or potent acetylcholinesterase (AChE) inhibitory activity. Among them, compound 7 (IC₅₀ for AChE?=?0.82?±?0.08?μmol?L^?1, IC₅₀ for BuChE?=?46.30?±?3.26?μmol?L^?1) showed a potent activity and high selectivity compared with the positive control Rivastigmine (IC₅₀ for AChE?=?10.54?±?0.86?μmol?L^?1, IC₅₀ for BuChE?=?0.26?±?0.08?μmol?L^?1). The kinetic study suggested that compound 7 bind to AChE with mix-type inhibitory profile. Molecular docking study revealed that compound 7 could combine both catalytic active site (CAS) and peripheral active site (PAS) of AChE with four points (Trp84, Trp279, Tyr70 and Phe330), while it could bind with BuChE via only His 20.

Full text of DOI:10.1080/14786419.2017.1340280

Natural Product Research
Formerly Natural Product Letters
ISSN: 1478-6419 (Print) 1478-6427 (Online) Journal homepage: http://www.tandfonline.com/loi/gnpl20
Discovery of potent and selective
acetylcholinesterase (AChE) inhibitors: acacetin
7-O-methyl ether Mannich base derivatives
synthesised from easy access natural product
naringin
Hao-ran Liu, Xue Men, Xiao-hui Gao, Lin-bo Liu, Hao-qun Fan, Xin-hua Xia &
Qiu-an Wang
To cite this article: Hao-ran Liu, Xue Men, Xiao-hui Gao, Lin-bo Liu, Hao-qun Fan, Xin-hua Xia
& Qiu-an Wang (2017): Discovery of potent and selective acetylcholinesterase (AChE) inhibitors:
acacetin 7-O-methyl ether Mannich base derivatives synthesised from easy access natural product
naringin, Natural Product Research, DOI: 10.1080/14786419.2017.1340280
To link to this article: http://dx.doi.org/10.1080/14786419.2017.1340280
View supplementary material
Published online: 15 Jun 2017.
Submit your article to this journal
View related articles
View Crossmark data
Full Terms & Conditions of access and use can be found at
http://www.tandfonline.com/action/journalInformation?journalCode=gnpl20
Download by: [University of New Mexico]
Date: 16 June 2017, At: 03:56

Natural Product research, 2017
https://doi.org/10.1080/14786419.2017.1340280
SHORT COMMUNICATION
Discovery of potent and selective acetylcholinesterase
(AChE) inhibitors: acacetin 7-O-methyl ether Mannich base
derivatives synthesised from easy access natural product
naringin
Hao-ran Liua, Xue Mena, Xiao-hui Gaob, Lin-bo Liua, Hao-qun Fana, Xin-hua Xiab and
Qiu-an Wanga
acollege of chemistry and chemical engineering, hu’nan university, changsha, china; bcollege of Pharmacy,
hu’nan university of chinese Medicine, changsha, china
ABSTRACT
ARTICLE HISTORY
received 19 december 2016
accepted 13 May 2017
Naringin, as a component universal existing in the peel of some
fruits or medicinal plants, was usually selected as the material to
synthesise bioactive derivates since it was easy to gain with low
cost. In present investigation, eight new acacetin-7-O-methyl ether
Mannich base derivatives (1–8) were synthesised from naringin.
The bioactivity evaluation revealed that most of them exhibited
moderate or potent acetylcholinesterase (AChE) inhibitory activity.
Among them, compound 7 (IC50 for AChE = 0.82 0.08 μmol•L−1, IC50
for BuChE = 46.30 3.26 μmol•L−1) showed a potent activity and high
selectivity compared with the positive control Rivastigmine (IC50 for
AChE = 10.54 0.86 μmol•L−1, IC50 for BuChE = 0.26 0.08 μmol•L−1).
The kinetic study suggested that compound 7 bind to AChE with
mix-type inhibitory profile. Molecular docking study revealed that
compound 7 could combine both catalytic active site (CAS) and
peripheral active site (PAS) of AChE with four points (Trp84, Trp279,
Tyr70 and Phe330), while it could bind with BuChE via only His 20.
KEYWORDS
Naringin; Mannich base;
acetylcholinesterase
inhibitors; alzheimer’s
diseases; molecular docking
CONTACT Qiu-an Wang
wangqa@hnu.edu.cn
supplemental data for this article can be accessed at https://doi.org/10.1080/14786419.2017.1340280.
© 2017 Informa uK limited, trading as taylor & Francis Group

2
H.-R. LIU ET AL.
1. Introduction
In recent years, a lot of tertiary amine derivatives originated from natural products were
synthesised and evaluated as acetylcholinesterase (AChE) inhibitors for potential application
in the treatment of Alzheimer’s diseases (Li et al. 2013; Silva et al. 2014; Sang et al. 2015).
According to the experiences in medicinal chemistry, amino moiety in drugs seemed to
improve physico-chemical properties or enhance bioactivity. The Mannich reaction is one
of important ways to introduction amino moiety for the natural compounds (Roman 2015).
In the present study, a series of acacetin-7-O-methyl ether Mannich-base derivatives were
synthesised from naringin, which is a low-cost and easy access natural product abundant
existing in China and South-east Asia. Then the effects of them in inhibiting AChE, BuChE or
carboxylesterase (CaE) were evaluated, followed by molecular docking study in further.
2. Results and discussion
2.1. Chemistry
Acacetin 7-O-methyl ether Mannich-base derivatives 1–8 were synthesised via a four steps
procedure as described in Scheme 1.
According to our previous procedure (Wu et al. 2012), the acacetin and acacetin-7-O-
methyl ether could be obtained by dehydrogenation, regioselective methylation, hydrolysis
of glycosidic bond from naringin. Then the synthesis of acacetin or acacetin-7-O-methyl
ether Mannich-base derivatives was conducted in the presence of formaldehyde, aliphatic
or alicyclic amines. The structures of the new synthesised compounds were confirmed by
1H NMR, 13C NMR and ESI-MS(+) analyses. The purity of new synthesised compounds was
more than 98.0% with the assay by high-pressure liquid chromatography.
2.2. Bioactivity evaluation in inhibiting AChE and BuChE
The results for the inhibitory effects against AChE are shown inTable 1. The variation of amine
groups markedly influenced the activity in inhibiting AChE. According to the data of com-
pounds 1–4, the order of inhibiting potency of them against AChE is: dimethylamino
group > diethylamino group > dipropylamino group > dibutylamino group. But there is a
significant difference among compounds 5–8. Compound 7 (IC50 = 0.82 0.08 μmol•L−1)
possessed the highest activity, but compound 6 had poor activity (IC50 > 500 μmol•L−1).
Compound 8 had a comparable activity with compound 5. It seemed that the introduction
of morpholine group markedly decreased the inhibitory activity for AChE. Among them
compound 7 had a highest selectivity in inhibiting AChE over BuChE (ratio: 56.5).
All the compounds exhibited high-specific inhibition against cholinesterase over CaE
(IC50 > 500 μmol/L) compared with the control drug Rivastigmine (for CaE IC50 = 31.10
1.20 μmol/L).
2.3. Kinetic assay
The linear Lineweaver–Burk equation was applied and the graphical analysis of the steady-
state inhibition data of compound 7 was shown in Figure S1. According to the results, the
kinetic profile of compound 7 exhibited a mixed-type inhibition. It suggested that compound

NATURAL PRODUCT RESEARCH
3
Scheme 1. reagents and conditions: (a) I2/Py, reflux; (b) concentrated h2so4, etoh, reflux; (c) (ch3)2so4,
K2co3, acetone, 65°c; (d) 37% hcho, hNr1r2, ch3oh, hcl, 65–70°c.
7 could bind with the catalytic site and non-catalytic site of the AChE with different equilib-
rium constants, and the competitive constant (Ki) and non-competitive constant (Ki’) are
0.71 and 2.17 μmol/L, respectively (Table S2).
2.4. Molecular docking study
A molecular docking study was performed for compound 7 using Molecular Operating
Environment (MOE). According to the results in Figure S2, A, compound 7 exhibited multiple
point binding modes with AChE. The nitrogen of piperidine ring binds to the catalytic anionic

4
H.-R. LIU ET AL.
Table 1. effect of compounds 1–8 on ache and Buche. Ic50 (μmol•l−1).
−NR1R2
CaE
>500
AChE (IC50)
BuChE (IC50)
Selectivitya
Compound
1
3.54 0.12
25.90 0.71
7.32
2
3
>500
>500
7.94 0.09
12.4 0.13
38.9 1.85
1.56
2.59
15.02 1.31
4
>500
31.50 0.81
45.2 2.34
1.43
5
6
>500
>500
3.94 0.78
>500
76.8 4.63
58.6 2.21
19.5
<0.118
7
>500
0.82 0.08
3.50 0.17
10.54 0.86
46.3 3.26
32.4 1.42
0.26 0.08
56.5
9.26
8
>500
rivastigmine*
–
31.10 1.20
0.0247
adefined as the ratio of Ic50 for Buche/ache.
*used for a positive control.
site (CAS) via the π-cation interaction with Trp84 (4.21 Å), and the aromatic ring binds to
peripheral anionic site (PAS) via a π–π stacking interaction with Phe330 (4.26 Å) and Trp279
(4.07 Å).In addition, methoxyl group in benzene ring binds with Tyr70 and H2O (Figure S2,
B). However, compound 7 only binds to BuchE by His20 (4.15 Å). These result from molecular
docking may be partially explained the high selectivity of compound 7 for inhibiting BuChE
and AChE.
3. Conclusion
In the present study, eight Mannich-base derivatives were synthesised from naringin and
bioactivity evaluation showed that most of them had better activity than that of control
drug Rivastigamine in inhibiting AChE. Among them, compound 7 which possesses a pip-
erdine side chain had more potent bioactivity and higher selectivity than the other com-
pounds. It is a promising compound for the treatment of AD as a novel AChE inhibitor from
natural product.
Supplementary material
Experimental details related to this article are available online in supplementary material.
Disclosure statement
No potential conflict of interest was reported by the authors.

NATURAL PRODUCT RESEARCH
5
Funding
The present investigation was supported by the Innovative Investigation funds for graduated students
of Hu’nan province [grant number CX2015B324]; the Natural Science Foundation of Hu’nan Province
[grant number 2017JJ2050].
References
Li RS, Wang XB, Hu XJ, Kong LY. 2013. Design, synthesis and evaluation of flavonoid derivatives as
potential multifunctional acetylcholinesterase inhibitors against Alzheimer’s disease. Bioorg Med
Chem Lett. 23:2636–2641.
Roman G. 2015. Mannich bases in medicinal chemistry and drug design. Eur J Med Chem. 89:743–816.
Sang ZP, Qiang XM, LiY, Yuan W, Liu Q, ShiYK, Ang W, LuoYF, Tan ZH, DengY. 2015. Design, synthesis and
evaluation of scutellarein-O-alkylamines as multifunctional agents for the treatment of Alzheimer’s
disease. Eur J Med Chem. 94:348–366.
Silva T, Reis J, Teixeira J, Borges F. 2014. Alzheimer’s disease, enzyme targets and drug discovery
struggles: from natural products to drug prototypes. Ageing Res Rev. 15:116–145.
Wu Z, Cai SL, Fan WJ, Wang QA. 2012. Semisynthesis of bioactive flavonols and aurones from naringin.
Chin J Org Chem(Chinese). 32:1296–1302.