Novel racemic tetrahydrocurcuminoid dihydropyrimidinone

analogues as potent acetylcholinesterase inhibitors

Sarawalee Arunkhamkaew, Anan Athipornchai, Nuttapon Apiratikul, Apichart Suksamrarn,
Vachiraporn Ajavakom

Department of Chemistry and Center of Excellence for Innovation in Chemistry, Faculty of Science, Ramkhamhaeng University, Bangkok 10240, Thailand
a r t i c l e i n f o
Article history:
Received 31 October 2012
Revised 11 March 2013
Accepted 21 March 2013
Available online 1 April 2013
Keywords:
Alzheimers disease
Acetylcholinesterase inhibitor
Curcumin
Dihydropyrimidinone
a b s t r a c t
The synthesis of racemic tetrahydrocurcumi n- (THC-), tetrahyd rodemethoxycurcumin- (THDC-) and tet-
rahydrobis demethoxycurcumin- (THBDC-) dihydropyrimidinon e (DHPM) analogues was achieved by uti-
lizing the multi-component Biginelli reaction in the presence of copper sulphate as a catalyst. The
evaluation of acetylcholinesterase inhibitors for Alzheimers disease of these compounds showed that
they exhibited higher inhibitory activity than their parent analogues. THBDCDHPM demonstrated the
most potent inhibitory activity with an IC
50
value of 1.34 0.03 lM which was more active than the
approve d drug galanthamine (IC
50
= 1.45 0.04 lM).
2013 Elsevier Ltd. All rights reserved.
Alzheimers disease (AD) is a progressive neurogener ative disor-
der of the central nervous system, affecting approximat ely 18 mil-
lion people worldwide and this number is estimated to grow up to
34 million people in 13 years time.
1
This disease is one of the most
common forms of dementia , which is determined clinically by
multiple cognitive decits including memory loss, emotional dis-
turbance, personali ty changes and nally the total eradication of
the nervous system.
1
This dreadful disease is particularly charac-
terized by the loss of cholinergic neurons which make an essential
acetylcholin e (ACh) neurotransmit ter in the neocortex and hippo-
campus. The deciency of a cholinergic neurotrans mitter, espe-
cially the decline of the level of ACh neurotransmit ter in the
brain, is intimately involved in memory loss in AD. The deactiva-
tion or terminat ion of ACh by acetylcholinest erase (AChE) within
the synapse itself together with the formatio n of extracellular
amyloid plaque (amyloid-b) and intracellular neurobrillary tan-
gles are generally thought to be associated with the reduction in
the release of neurotransmit ters.
2
To the present date many re-
search approaches to the discovery of the effective AChE inhibitor
(AChEI) for the treatment of AD have been reported.
3
Among them,
there are some important examples such as the rst generation of
AChE inhibitor (AChEI), tacrine (Cognix),
4
which is rarely used
nowadays because of its potential liver toxicity, donepezi l (Air-
cept),
5
rivastigm ine (Exelon)
6
and galantha mine (Reminyl),
7
which have been proved as effective drugs in treatment of AD. De-
spite these great efforts, no drug that can provide the complete
protectio n of neurons has ever been found. However, effective
AD treatments are realized by blocking ACh breakdown by use of
AChEI and thus prolonging ACh action. We are therefore intereste d
to develop a novel AChEI by modifying curcuminoid natural prod-
ucts, which are an abundant resource in Asian countries.
Curcumin oids can be obtained from rhizomes of turmeric, Cur-
cuma longa L. (Zingiberaceae).
8
By possessing attractive biologica l
activities including antioxidant, anti-inammatory, anticance r,
anti-HIV and recently discovered anti-Alzhei mers, curcumin oids,
particular ly the three main components; curcumin (1), demethoxy-
curcumin (2) and bisdemetho xycurcumin (3), have been drawing
an enormous amount of attention from researchers.
9
However , its
hydrogen ated curcumin oids, tetrahyd rocurcumin (THC, 4), tetra-
hydrodem ethoxycurcumi n (THDC, 5) and tetrahydrob isdemthoxy -
curcumin (THBDC, 6), have been evaluated as antioxidant s and a
biomarker in a clinical study.
10,11
The evaluation of these curcumi-
noid congeners (46) as the AChEI is therefore very interesting.
Also, the comparison between AChE inhibitory activities of curc-
uminoids (13) and THCs (46) on AChE may provide an important
discovery for the cure of AD (Table 1). The Ellman method was uti-
lized for the test of the AChE inhibitory activities of both curcumi-
noids (13) and THCs (46).
12
According to the results, each THC
exhibited approximat ely twofold better IC
50
compared to that of
its curcuminoid pair. This preliminary evaluation demonstrat ed
that the absence of double bond and methoxy group within the
curcumin oid structure tends to increase in the inhibitory activity.
The results demonst rated that curcuminoids (13) and THCs
(46) displayed potential inhibitory activity against AChE,
however , the activity was not high when compared to that of the
0960-894X/$ - see front matter 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.03.069

Corresponding author. Tel.: +66 2 319 5112; fax: +66 2 310 8401.
E-mail address: a_vachiraporn@ru.ac.th (V. Ajavakom).
Bioorganic & Medicinal Chemistry Letters 23 (2013) 28802882
Contents lists available at SciVerse ScienceDi rect
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reference AChEI, galanthamine. According to the molecular model-
ing studies reported by Tomassoli et al. some dihydropyrimi dinone
(DHPM) derivatives have been reported as the potent compound
against AChE.
13
These six-membered heterocyclic analogues also
have wide range of biological properties such as antibacterial , anti-
fungal, anti-inamatory, calcium channel blocker, and anti-cancer
activity.
14
It is therefore of interest to combine the THCs (46)
with the DHPM units, which may lead to the novel drug candidat e
with much improved AChE inhibitory activity. In order to do so, a
synthetic strategy was designed to construct the DHPM building
block out of the THC unit. Observation of the curcumin structure
led us to consider the 1,3-dicarbonyl moiety of THC as one compo-
nent for a multi-comp onent cyclocondens ation reaction.
15
The rep-
resentative Biginelli reaction, the acid-cata lyzed three-comp onent
condensation of an aldehyde, a urea and a 1,3-dicarbonyl com-
pound, is capable of creating the DHPM derivatives.
16
Though sul-
furic acid can be traditionally used as the acid catalyst in this
reaction, various Brnsted and Lewis acids, for example p-TsOH,
Sr(OTf)
2
, Mn(OAc)
2
2H
2
O, CuCl
2
2H
2
O and CuSO
4
5H
2
O, have been
proven to be very effective.
17
In this work, after screening several
Lewis acids as catalysts for the Biginelli reaction of THC,
CuSO
4
5H
2
O gave acceptably promising results (Scheme 1). The
divalent copper metal may have the coordina ting property with
the 1,3-dicarbonyl group and stabilizes the enol form generate d
in situ which prompts to react with other two components to pro-
vide the DHPM product.
In our initial investigations, the THC readily available through
the catalytic hydrogen ation reaction of curcumin was subjected
to Biginelli reaction incorporating with urea and various aromatic
aldehydes to create a number of THCDHPM analogues (Table 2,
compounds 815). It is worth noticing that all cyclized products
are racemic mixtures. The reaction of THC (4), urea and benzalde-
hyde in the presence of CuSO
4
5H
2
O under reuxing temperature
in EtOH gave the smooth cyclocondensati on to the correspond ing
THCDHPM derivative 8 in good yield. Various substituents such
as nitro, hydroxy, and methoxy groups were introduced onto the
aromatic aldehydes to provide a series of THCDHPM derivatives
915, in order to evaluate the substituent effect on their biological
activities. THCDHPMs (11 and 12) bearing a hydroxy or methoxy
group at the para-position of the starting aromatic aldehyde satis-
factorily revealed high inhibitory activities of AChE with IC
50
val-
ues of 17.31 0.17 and 14.28 0.09, respectively. Encouraged by
these results, the synthesis of THDCDHPMs (16 and 17) from
THDC (5), benzaldehy de and 4-methoxyb enzaldehyde were carried
out in a similar manner to that of the preparation of THCDHPMs
(815). The mixtures of structural isomers from both THDC
DHPMs (16 and 17) were obtained. The regioisomer s of each of
compounds 16 and 17 were separated using normal phase HPLC
to give 16a and 16b, 17a and 17b, respectively, and biologically
Table 2
Synthesis and AChE inhibitory activity of racemic THCDHPM analogues
Compound R
1
R
2
R
3
Yield % IC
50
(lM)
8 H OCH
3
OCH
3
74 36.33 0.50
9 3-NO
2
OCH
3
OCH
3
59 ND
c
10 4-NO
2
OCH
3
OCH
3
67 ND
c
11 4-OH OCH
3
OCH
3
37 17.31 0.17
12 4-OCH
3
OCH
3
OCH
3
44 14.28 0.09
13 2,4-OCH
3
OCH
3
OCH
3
27 ND
c
14 2,5-OCH
3
OCH
3
OCH
3
48 ND
c
15 3-OCH
3
-4-OH OCH
3
OCH
3
44 56.42 0.58
16 H OCH
3
or H H or OCH
3
39
b
5.36 0.20
16a H OCH
3
H 10.65 0.58
16b H H OCH
3
20.94 0.54
17 4-OCH
3
OCH
3
or H H or OCH
3
21
b
2.89 0.10
17a 4-OCH
3
OCH
3
H 5.78 0.64
17b 4-OCH
3
H OCH
3
9.10 0.16
18 H H H 55 2.04 0.11
19 4-OCH
3
H H 25 1.34 0.03
7 Galanthamine
a
1.45 0.04
a
Reference compound.
b
Yield given for the mixture (1:1) of two isomers.
c
Not determined.
Scheme 1. Synthesis of racemic THCDHPM analogues by a multi-component Biginelli reaction.
Table 1
Inhibitory activity IC
50
(lM) of curcumino ids and THCs on AChE
Compound Structure IC
50
(lM)
1
O O
H
3
CO
HO
OCH
3
OH
81.05 2.51
2
O O
HO
OCH
3
OH
37.74 0.16
3
O O
HO OH
18.27 0.09
4
O O
HO OH
OCH
3
H
3
CO
41.16 0.17
5
O O
HO OH
OCH
3
20.55 0.12
6
O O
HO OH
7.89 0.03
7 Galanthamine
a
1.45 0.04
a
Reference compound.
S. Arunkhamkaew et al. / Bioorg. Med. Chem. Lett. 23 (2013) 28802882 2881
evaluated together with their mixtures as AChEI. The THBDC (6)
was also introduce d to the Biginelli reaction with benzaldehyd e
and urea to furnish the THBDCDHPM analogue (18). As a methoxy
group in the aromatic ring of the aldehyde precursor is considered
to be essential for inhibitory activity, we therefore synthesized the
desired THBDCDHPM analogue (19) by treatment of THBDC (6)
with 4-methoxybenzal dehyde.
The preliminary study of the inhibitory potencies of synthesized
THCDHPM(8) showed a low inhibitory property against AChE and
in the same range IC
50
value of 36.33 0.50 lMwhen compared to
the correspond ing THC (IC
50
= 41.16 0.17 lM) (Table 2). From this
result, it was realized that the DHPM unit in the molecule essen-
tially enhances the activity. However , with low inhibitory values,
the 3- and 4-nitro phenyl THCDHPManalogues (9 and 10) showed
that the nitro group may have negative effect on the binding ability
with the enzyme. Other THCDHPM analogues (11 and 12) contain-
ing 4-OH or 4-OCH
3
phenyl group displayed better results with low
inhibitory activities and with IC
50
value of 17.31 0.17 lM and
14.28 0.09 lM, respectively, (entries 4 and 5). However, the intro-
duction of one more electron donating group in the aromatic moi-
ety did not increase the activities against AChE, as 2,4-OCH
3
-, 2,5-
OCH
3
- and 3-OCH
3
-4-OH-ph enyl THCDHPMs (1315) exhibited
poor inhibitory activities. A mixture isomers of THDCDHPM ana-
logue (17) bearing a 4-OCH
3
phenyl group demonst rated much bet-
ter activity with an IC
50
value of 2.89 0.10 lMthan that of THDC-
DHPM (16) without the 4-OCH
3
phenyl group (5.36 0.20 lM), as
well as in the case of both THBDCDHPMs (18 and 19), IC
50
value
of 2.04 0.11 lMand 1.34 0.03 lMwere obtained from the com-
pound without 4-OCH
3
phenyl group and with 4-OCH
3
phenyl
group, respectivel y. However, the individua l regioisomers 16a and
16b, 17a and 17b were less active than their respective mixture
16 and 17. This was probably due to the synergistic effect of the
regioisomer s in the mixture. From the screening test of AChEIs,
the methoxy substituent on the phenyl group originate d from alde-
hyde and an absence of methoxy group on curcuminoid component
seem to be crucially important for the inhibitory activity of these
compounds .
In conclusion, THC-, THDC- and THBDCDHPM analogues were
synthesized and evaluated as inhibitors of AChE. The inhibitory
potencies of the synthesized compounds against the AChE unveiled
that THBDCDHPM (19) bearing a 4-OCH
3
phenyl group has a
highly effective inhibitory activity of AChE
18
with an excellent
IC
50
value of 1.34 0.03 lM, which is slightly more potent than
that of galanthamine.
Acknowled gments
This work was supported by the Center of Excellence for Inno-
vation in Chemistry (PERCH-CIC), The Thailand Research Fund
and the Department of Chemistry, Faculty of Science, Ramkhamha-
eng Universit y. We would also like to acknowled ge the Depart-
ment of Chemistry , Faculty of Science, Chulalongk orn University
for HPLC separation.
Supplemen tary data
Supplement ary data associated with this article can be found, in
the online version, at http://dx .doi.org/10.1016/j .bmcl.2013.03.
069.
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