Development of coumarin–benzofuran hybrids as versatile multitargeted compounds for the treatment of Alzheimer’s Disease

Article abstract of DOI:10.1111/cbdd.13316

Alzheimer’s disease (AD), the most common cause of dementia, is a neurodegenerative disorder characterized by progressive deterioration of memory and cognition. The evidenced multifactorial nature of AD has been considered the main reason for the absence of cure so far. Therefore, the development of novel hybrids to treat the disease is very much essential. Focusing on this, a novel series of coumarin–benzofuran hybrids have been designed and screened as anti-Alzheimer’s disease agents. The strategy is to obtain an effective mimetic of donepezil, which is acetylcholinesterase inhibitor. Herein, the two main scaffolds namely coumarin and benzofuran are known pharmacophore moieties and we have performed their molecular design, pharmacokinetic descriptor studies for drug-likeliness. Further, in vitro studies such as antioxidant capacity, acetylcholinesterase (AChE) inhibition and amyloid-β (Aβ) self-aggregation inhibition have also been performed. Most importantly, these studies revealed that the newly synthesized hybrids can be versatile and promising drug-like moieties as efficient anti-AD agents.

Full text of DOI:10.1111/cbdd.13316

DR RANGAPPA S. KERI (Orcid ID : 0000-0001-6262-8379)
Article type : Research Article
Development of Coumarin and Benzofuran Conjugated Hybrids as the Versatile Multi-
targeted Compounds for the Treatment of Alzheimer’s Disease
1
2
1,
Asha Hiremathad , Karam Chand , and Rangappa S. Keri *
1
Centre for Nano and Material Sciences, Jain University, Jain Global Campus, Kanakapura,
Ramanagaram, Bangalore - 562112, India.
2
Department of Chemistry, Umeå University, 90187 Umeå, Sweden.
*
Corresponding author: Dr. Rangappa S. Keri, Organic and Medicinal Chemistry Laboratory, Centre for Nano
and Material Sciences, Jain University, Jain Global Campus, Jakkasandra post, Kanakapura Road, Ramanagara
District, Karnataka-562112, INDIA.
Tel.: +918027577199
E-mail: keriphd@gmail.com, sk.rangappa@jainuniversity.ac.in
Abstract
Alzheimer’s disease (AD), the most common cause of dementia, is a neurodegenerative
disorder characterized by progressive deterioration of memory and cognition. The evidenced
multi-factorial nature of AD has been considered the main reason for the absence of cure so
This article has been accepted for publication and undergone full peer review but has not
been through the copyediting, typesetting, pagination and proofreading process, which may
lead to differences between this version and the Version of Record. Please cite this article as
doi: 10.1111/cbdd.13316
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far. Therefore, the development of novel hybrids to treat the disease is very much essential.
Focusing on this, a novel series of coumarin-benzofuran conjugated hybrids have been
designed and screened as anti-Alzheimer’s disease agents. The strategy is to obtain an
effective mimetic of donepezil, which is acetylcholnesterase inhibitor. Herein, the two main
scaffolds namely coumarin and benzofuran are known pharmacophore moieties and we have
performed their molecular design, pharmacokinetic descriptor studies for drug likeliness.
Further, In vitro studies such as antioxidant capacity, acetylcholinesterase (AChE) inhibition
and amyloid-β (Aβ) self-aggregation inhibition have also been performed. Most importantly,
these studies revealed that the newly synthesized hybrids can be versatile and promising drug
like moieties as efficient anti-AD agents.
Keywords: Alzheimer’s disease; Coumarin; Benzofuran; Acetylcholinesterase inhibitors.
Introduction
Despite much progress has been done in the area of drug discovery, Alzheimer's
disease (AD) still remains an unresolved challenge in modern medicine and it is being a big
st
threat to people's health in the 21 century. Currently more than 26 million people are
suffering from AD worldwide and the number of patients are expected to increase by 4-fold
[1,2]
in 2050.
Thus, much attention is required to design and develop novel alternative agents
which can provide possible complete cure from AD with lesser side effects. There are some
disease-modifying drugs available and most of them are AChE inhibitors (tacrine, donepezil,
rivastigmine and galantamine) and only one N-methyl-D-aspartate receptor (NMDAr)
antagonist (memantine). However, these drugs can display limited therapeutic success as they
[
3]
can delay the disease growth over short period of time. Apart from choline loss, there are
[
4]
number of complex processes such as β-secretase, which involve in the first step of the
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pathway leading to the production and deposition of amyloid-β peptide. This in turn leads to
the formation of intracellular neurofibrillary tangles due to formation of amyloid-beta (Abeta)
[
5]
[6]
aggregation
as well as dyshomeoatasis of redox-active metal ions
and increased
[
7]
oxidative stress, which are considered to be responsible for the development of this
neurodegenerative disease. The progression of these multiple factors is simultaneously
believed to be the main reason for the absence of an effective cure. Therefore, the molecules
which can tackles these multiple problems together could be potentially effective for the
treatment of AD. To find out new anti-AD agents, various approaches have been developed,
such as the inhibition of enzymes (eg AChE and monoamineoxidase (MAO-B)), which are
[
8]
responsible for degradation of cholinergic and dopaminergic neurotransmitters, inhibition
[
9]
of other pathogenic processes, namely the anti-aggregation of beta amyloid (Aβ) peptide
[
10]
[11]
and β-secretase
as well as the modulation of the oxidative stress
and the related
[
12, 13]
transition metal dysregulation
has been attempted separately through “one-compound
one-target” strategy, while very limited efforts has been made to deal these targets
simultaneously.
To deal the several targets of AD simultaneously, we have proposed the dual-acting
[7, 14-17]
multitargeted molecules as anti-AD agents
and accordingly developed the benzofuran
[
18-20]
(
donepezil component known for AChE inhibition
) and coumarin (known for the AChE
[
19-21]
[22, 23]
inhibition
antioxidant and anti-Aβ self-aggregation
) based hybrids to provide new
class of anti-AD molecules as shown in Fig. 1. The coupling of these two selected
pharmacophores was designed through amidic linkage of aliphatic chain and adequate length
of alkyl linker was optimized through molecular docking simulations, so that designed
molecules can fit comfortably in the cavity of acetylcholinesterase (TcAChE) enzyme and
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provide maximum interaction with catalytic active site (CAS) and the peripheral anionic site
(PAS) to induce maximum AChE inhibition.
In this context, the aim of this study is to develop a novel class of multi-functional
hybrid compounds as anti-AD drug candidates that composed of multiple molecules into one
single-molecular entity and to improve their activity associated with various targets of AD to
enhance therapeutic efficacy of the developed anti-AD drug.
Materials and methods
Molecular modeling
To perform the ligand-enzyme docking studies the X-ray crystallographic structure of
Torpedo californica-AChE (TcAChE) complexed with an inhibitor (donepezil, Dnp) was
[
25]
taken from RCSB Protein Data Bank (PDB entry 1 EVE) in order to be used as enzyme
model. This structure was chosen because of the above referred similarity between the
original inhibitor (Donepezil) and herein studied ligands, namely between the
benzylpiperidine and the indanone moieties of donepezil and corresponding coumarin and
benzofuran moieties of the ligands. Further, molecular simulations and docking were
[21]
performed as previously described method.
Prediction of pharmacokinetic properties
To analyze the potential of the new compounds as new anti-AD drugs, a brief
prediction on pharmacokinetic properties was performed in silico. Parameters such as the
lipo-hydrophilic character (clogP), blood-brain barrier partition coefficient (log BB), the
ability to be absorbed through the intestinal tract (Caco-2 cell permeability) and CNS activity
were calculated. The ligands were built and minimized as previously mentioned for the
docking studies. The structures were submitted to the calculation of these relevant
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[
30]
pharmacokinetic proprieties and descriptors using QikProp v. 2.5 . These predictions are
for orally delivered drugs and assume non-active transport.
Inhibition of AChE
The enzymatic activity of TcAChE was measured using an adaptation of the method
[17, 28, 31,]
previously described.
The percentage inhibition of the enzyme activity due to the
presence of increasing test compound concentration was calculated by the following Eq. (1)
ꢆ
ꢇ
ꢀ
ꢁꢂꢃꢃꢄꢅ ꢉꢂꢃꢃꢊ
(1)
ꢆ
ꢈ
i o
in which v is the initial reaction rate in the presence of inhibitor and v is the initial rate of
the control reaction. The inhibition curves were obtained by plotting the percentage of
enzymatic inhibition versus inhibitor concentration, and a calibration curve was obtained
from which the linear regression parameters were obtained.
Inhibition of self-induced Aβ1-42 aggregation
To study the Aβ42 aggregation inhibition, a reported method, based on the
[
17, 25,
fluorescence emission of thioflavin T (ThT), was followed as reported on the literature.
3
2, 33]
Firstly, Aβ42 (10 µL) samples and the tested compounds (10 µL) were diluted with the
phosphate buffer to a final concentration of 40 µM (Aβ) and 80 µM (compounds), and then,
they were incubated for 24 h at 37 °C, without stirring. The percent inhibition of the self-
induced aggregation due to the presence of the test compound was calculated by the Eq. (2),
i 0
in which IF and IF correspond to the fluorescence intensities, in the presence and the
absence of the test compound, respectively, minus the fluorescence intensities due to the
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respective blanks. The reported values were obtained as the mean ±SEM of duplicate of two
different experiments.
ꢋꢌ
ꢇ
ꢀ
ꢁꢂꢃꢃꢄꢅꢋꢌ ꢉꢂꢃꢃꢊ
(2)
ꢈ
Radical scavenging activity
The antioxidant activity was evaluated by the DPPH method previously described.
The absorbance was measured at 517 nm against the corresponding blank (methanol). The
[34]
[35].
antioxidant activity was calculated by Eq. (3).
ꢎꢏꢐꢐꢑꢒꢎꢓꢔꢕꢖꢗꢘ
ꢍꢍꢁ
ꢉꢂꢃꢃ
(3)
ꢎꢏꢐꢐꢑ
The tests were carried out in triplicate. The compound concentrations providing 50%
of antioxidant activity (EC50) were obtained by plotting the antioxidant activity against the
compound concentration.
Results
The synthetic strategy of novel coumarin-benzofuran (CA-BF) hybrids was
accomplished by the convergent approach of three steps; which mainly involved the synthesis
of the two main intermediate coumarin acid (CA) (8) and benzofuran amine (BF) (5) in two
steps, and their coupling in third step, as shown in Fig. 2. Firstly, the synthesis of BF esters
was carried out by condensation of (un)substituted salicylaldehyde with ethylchloroformate
in DMF under basic and refluxing conditions to yield BF intermediates 3(a,b); where the
subsequent condensation with alkyl amines of chain length (n = 0-2) in methanol at room
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temperature yielded the desired benzofuran amines 5(a-f). In second step, the synthesis of CA
was achieved by condensing the (un)substituted salicyaldehyde with meldrum’s acid in water
[
24].
under refluxing condition to give desired intermediate 8(a,b). Finally, in third step, the
coupling of two intermediates; 8(a,b) and 5(a-f) was achieved by using T P as carboxylic
3
acid activator in presence of N-methylmorpholine (NMM) base in dichloromethane (DCM) at
room temperature, which led to the formation of the novel coumarin-benzofuran (CA-BF)
hybrid molecules 9(a-l). 1. Experimental details and characterization for the synthesized
molecules is provided as supplementary data (see Fig. S1 to Fig. S12).
Molecular design and docking studies were performed. The strategy for the
conjugation of coumarin and benzofuran scaffolds endeavored to provide the hybrids with
multi-targeting ability, particularly AChE inhibition, antioxidant and anti-Aβ aggregation
capacity. These two main scaffolds are known potent molecules to hit the targeted enzymes.
[
18, 36]
Recently, benzofuran derivatives studied as potent and dual binding AChE inhibitors
and coumarins are naturally occurring phytochemicals with wide range of biological
activities and these can be synthesized synthetically and these showed potent AChE
inhibition and further, functionalization of the core moiety and coupling with other
pharmacological molecule result in improved activities and neuroprotective effects. Hence
the coupling of these two pharmacophores (coumarin and benzofuran) may improve the
[
19]
inhibitory capacity by interacting in the PAS of enzyme gorge. Further, the introduction of
the methoxy group either on coumarin and benzofuran rings was found to improve the AChE
[
37, 20]
inhibitory activity.
The docking simulations of the ligand into active site of the enzyme
(
Torpedo Californica variant (TcAChE)), were performed to find the optimal chain length
between these scaffolds to get fit into the empty site of AChE enzyme. Molecular docking
simulations demonstrated that two to four methylene linkers could endow adequate length to
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facilitate the dual mode interaction with two molecular fragments coumarin acid (CA) and
[37]
benzofuran amine (BF) within the AChE gorge.
The pharmacokinetic profiles of the synthesized compounds were simulated using the
QikProp program, v. 2.5, in order to study the drug-likeness of the compounds and their
ability to penetrate into the important membranes such as the blood-brain barrier
[
26]
(
BBB). Accordingly, the following parameters were calculated; (i) the octanol-water
partition coefficient (clogP), which indicates the ability to cross the BBB (log BB), (ii) the
capacity to be absorbed through the intestinal tract to the blood (Caco-2 cell permeability),
[
38]
and (iii) the verification of Lipinski’s rule of five
for the drug-likeness properties as the
potential anti-AD agents. The calculated values are given in Table S1, in which all the
compounds with zero violation of Lipinski’s rule. This promises the drug likeliness and these
can be orally active molecules.
These hybrids were aimed for multi-functionality, so firstly these series of novel
hybrid compounds 9(a-l) were studied for their antioxidant properties based on their
[
34]
interaction with 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical ions. The results are
given in Table 1. All the compounds have demonstrated lower radical scavenging activity
than the known antioxidant Trolox (EC50 = 15.6 µM). However, these exhibited moderate to
improved antioxidant properties in comparison with the reference drug deferiprone (174 µM).
[
21]
Thus, the presence of methoxy group series improved the antioxidant capacity.
Further,
TcAChE enzymatic inhibition properties of the synthesized hybrids 9(a-l) were analyzed
[
31]
using the method described in literature.
These hybrids 9(a-l) showed the improved AChE
inhibition capacity as given in Table 1. In continuation, these novel hybrids were tested for
the Inhibition of self-induced Aβ1-42 aggregation. The in vitro anti-amyloidogenic activity of
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synthesized compounds was studied using thioflavin-T (ThT) assay in order to monitor and
quantify the aggregation of Aβ42 synthetic peptide into peptide fibrils. These activities were
monitored by the fluorescence emission of ThT with the absorption and emission peak at 446
nm and 490 nm, respectively. The anti-amyloidogenic activity was carried out by incubating
the Aβ42 in the presence and absence of the synthesized hybrids and the results are given in
[
32, 33]
Table 1.
Discussion
As shown in Fig. 3 the molecular design and modeling revealed that the docked
hybrid molecules revealed that the designed molecules can have the good binding ability with
amino acid of both CAS and PAS through π-π stacking, Hydrogen-bond (H-bond) and
electrostatic interaction as similar to the model ligand, which is donepezil. In general,
docking simulations gave insights into the compounds interaction with original ligand and
enzyme. For example, the compound 9d interacts with Trp84 in the CAS and Trp279 in the
PAS through π-π stacking, and also oxygen atom of coumarin and -NH hydrogen establishes
the H-bonding with Tyr121 (2.65 Å and 2.27 Å respectively). Hence, the compound is found
to be a promising AChE inhibitor (Fig. 3a).
Similarly, the simulation study of compound 9h revealed its interactions with the
Trp84 and Trp279 amino acids. Interestingly, amidic oxygen of benzofuran ring forms the H-
bond with Tyr121 (2.58 Å; Fig. 3b) resulted into enhancement of interactions with enzyme.
Further, on evaluating the docking simulations studies for the compounds of longer chain
length (n=2) reveals no hydrogen bonding however, π -π stacking interaction of these
compounds with Trp84 and Trp279 were sufficient to provide highest AChE inhibition
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capacity among all the compounds. The biological studies revealed that the higher chain
linker (n=2) and methoxy substituted compounds were tend to show enhanced AChE
inhibition as compared to the rest of the hybrids, which could be due to the electrostatic
interaction and π-π stacking with Trp84 and Trp279 in dual site of CAS and PAS (Fig. 3c).
Further, the drug likeliness of these hybrids were performed by pharmacokinetics
properties, it was found that these series of compounds possess the clogP values ≤5,
molecular weight ≤500, low negative log BB values (ranging from -1.9 to -0.689), which
indicated their relatively improved blood-brain permeability which was found to be in the
range between -0.3 and +1.0, which well lies in the acceptable range of Lipinski’s rule. In
addition, the obtained values also showed promising results in Caco-2 permeability, ranging
from 968 to 116 nm/s (≥500 considered as good permeability constant). However, some of
the compounds (9a, 9d, 9g, 9j, and 9l) showed lesser Caco-2 permeability, which may be due
to low intestinal absorption or less hydrophilicity of the molecules. However the other
compounds exhibited greater values, which indicate that the absorption through the intestinal
to the blood is possible.
Additionally, biological evaluation of these novel hybrids was demonstrated. The
results are indicated in Table 1. Initially, radical scavenging activity performed and these
have shown moderate to good capacity towards free radicals. Interestingly, the compounds
9
d, 9j, and 9l (EC50 = 347, 359 and 445 µM respectively) with methoxy and shorter linker
showed highest antioxidant property among all the hybrids and moderate antioxidant capacity
may attribute to the less potency of molecules to transfer the electrons to the DPPH radical.
Further, AChE inhibition of these novel hybrids was performed; the results are presented in
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Table 1. It showed improved inhibition activity towards the targeted enzyme. In particular, it
was observed that the compounds with methoxy substitution with chain length n=2 (9j)
showed the highest inhibitory capacity (IC50) of 0.18 µM, which may be due to the good π-π
stacking in both active sites (CAS and PAS) with Trp84, Trp279 and electrostatic attraction
between the Tyr121 (see Fig. 3c), which may help to inhibit the enzyme activity. Similarly,
the compounds 9d, 9h, 9j, and 9l, which have the methoxy group either at the benzofuran or
coumarin ring, were found to exhibit improved AChE inhibitory capacity of 0.27, 0.32, 0.18
and 0.224 µM, respectively as compared to the hybrids 9a, 9e, 9g, and 9i, as these
compounds did not contain any methoxy substituents. Such observations essentially revealed
that the positive effect of methoxy substitution increases the inhibitory capacity of the
molecules as corroborated in the docking studies. Methoxy group is forming better
interaction with Trp279 in PAS which improves the inhibitory activity. Simultaneously
adequate linker is essential to fit the molecule into the enzyme gorge. Finally, the ThT assay
performed to test the percentage of Aβ self-aggregation inhibition. The obtained results
revealed that all the compounds tend decrease the ThT fluorescence associated with the Aβ
fibril binding, which signified the inhibition of the Aβ self-aggregation. These hybrids
exhibited Aβ self-aggregation inhibition in the range of 16.5-72.1%. Notably, the compounds
(
9g and 9h) with chain linker (n = 1) and methoxy substitution showed the highest inhibition
against Aβ plaques formation, which may be due to the good π-π stacking in the PAS which
result in successfully inhibit the aggregation.
Further, the compounds 9d and 9l showed the similar aggregation properties even
though they were different in chain lengths, which attributed to the presence methoxy group
in both hybrids and able to inhibit the aggregation. However, some of these hybrids are very
effective against the Aβ self-aggregation, which could be due to the appropriate methylene
chain linker (n=1). The results are given in Table 1.
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Structure-activity relationship (SAR) studies
The synthesized novel hybrids were further explored for their structure-activity
relationship properties as shown in Fig. 4. It was observed from the molecular simulations
studies that the compounds with shorter linker (n=0) possessed a good antioxidant activity,
while the compounds with longer chain linker showed the positive effects in the inhibition of
AChE and Aβ self-aggregation. Especially, the methoxy substitution showed significant
effects on the biological properties such as antioxidant, AChE inhibition and Aβ self-
aggregation inhibition. For example, the compound 9j showed the highest AChE inhibition
capacity of 0.18 µM among all the compounds and similarly, the compound 9h showed the
highest Aβ self-aggregation inhibition of 72.1%. On the other hand, the antioxidant capacity
was also found to be improved due to methoxy substitution, where the compounds 9d, 9j, and
9
l showed improved and almost similar scavenging properties among all the compounds
(EC50 µM) = 347, 359 and 445 µM, respectively).
Conclusion
In this study, the coupling of two pharmacophore molecules was demonstrated to
develop a versatile hybrid drug like moieties. A series of twelve novel hybrids were designed,
synthesized and evaluated for their biological activities as anti-AD agents. The designed
compounds were coupled by two main scaffolds (coumarin and benzofuran) which are
biologically active moieties, which could mimic the FDA approved drug-donepezil. The
molecular modeling simulations revealed that the coumarin and benzofuran could mimic
donepezil with linker size (n=0-2). Particularly, the compounds with two methylene chain
linkers were showed good interaction with original ligand by forming the H-bond (oxygen
atom of coumarin and –NH proton of methylene chain linker) with Tyr121. The longer chain
linker (n=2) was found to be efficiently conjugated in dual modes (CAS and PAS) with
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Trp84 and Trp279. The synthesized hybrids were further evaluated for antioxidant capacity,
AChE inhibition and Aβ self-aggregation inhibition. All hybrids showed moderate to good
antioxidant potential as compared to deferiprone and Trolox, among which the hybrid with
methoxy functional group with shorter linker (n=0) showed the highest DPPH radical
scavenging capacity. Similarly, these hybrids exhibited good AChE inhibition properties;
especially the compounds with methoxy substitution and longer chain linker showed the
highest AChE inhibitory capacity among all hybrids. Overall, it was realized that the
methoxy substitution and appropriate linker length has a positive effect on the inhibitory
activities. In conclusion, we have synthesized a series of moderate to good anti-AD
compounds with multi-targeting potential, by using a very simple approach and the structure
activity relationship information obtained from this study can be further useful for the
researcher working in similar area for the development highly active anti-AD agents.
Acknowledgement
Authors acknowledge Jain University for financial support and AH is thankful to Erasmus
NAMASTE consortium (unique grant number: NAMASTE_20140147) for the award of her
doctoral exchange fellowship.
Conflict of interest
The authors confirm that this article content has no conflicts of interest.
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List of figures and captions
Fig. 1. Design strategy of the novel coumarin-benzofuran (CA-BF) hybrids
Fig. 2. General method for the synthesis of the coumarin-benzofuran conjugated hybrids 9(a-
l)
Fig. 3. Docking results for the coumarin-benzofuran (CA-BF) hybrids with TcAChE and
superimpositions for illustration of structural differences: (a) superimposition of the original
ligand (Donepezil) (PDB entry 1EVE) (pink) and 9d (cyan); (b) superimposition of the
original ligand (pink) and 9h (cream); (c) superimposition of the original (pink) and 9j
(
yellow)
Fig. 4. Structure-activity relationship studies of coumarin-benzofuran conjugated hybrids
1
13
Fig. S1: (a) H NMR and (b) C NMR Spectra of N-(2-(benzofuran-2-carboxamido) ethyl)-
-oxo-2H-chromene-3-carboxamide (9a).
2
1
13
Fig. S2: (a) H NMR and (b) C NMR Spectra of N-(2-(benzofuran-2-carboxamido)ethyl)-8-
methoxy-2-oxo-2H-chromene-3-carboxamide (9b)
1
13
Fig. S3: (a) H NMR and (b) C NMR Spectra of N-(2-(7-methoxybenzofuran-2-
carboxamido) ethyl)-2-oxo-2H-chromene-3-carboxamide (9c)
1
13
Fig. S4: (a) H NMR and (b) C NMR Spectra of N-(2-(7-methoxybenzofuran-2-
carboxamido)ethyl)-2-oxo-2H-chromene-3-carboxamide (9d)
1
13
Fig. S5: (a) H NMR and (b) C NMR Spectra of N-(3-(benzofuran-2-carboxamido) propyl)-
-oxo-2H-chromene-3-carboxamide (9e)
2
1
13
Fig. S6: (a) H NMR and (b) C NMR Spectra of N-(3-(benzofuran-2-carboxamido) propyl)-
-methoxy-2-oxo-2H-chromene-3-carboxamide (9f)
8
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1
13
Fig. S7: (a) H NMR and (b) C NMR Spectra of N-(3-(7-methoxybenzofuran-2-
carboxamido)propyl)-2-oxo-2H-chromene-3-carboxamide (9g)
1
13
Fig. S8: (a) H NMR and (b) C NMR Spectra of 8-methoxy-N-(3-(7-methoxybenzofuran-2-
carboxamido)propyl)-2-oxo-2H-chromene-3-carboxamide (9h)
1
13
Fig. S9: (a) H NMR and (b) C NMR Spectra of N-(4-(benzofuran-2-carboxamido)butyl)-2-
oxo-2H-chromene-3-carboxamide (9i)
1
13
Fig. S10: (a) H NMR and (b) C NMR Spectra of N-(4-(benzofuran-2-carboxamido)butyl)-
-methoxy-2-oxo-2H-chromene-3-carboxamide (9j)
8
1
13
Fig. S11: (a) H NMR and (b) C NMR Spectra of N-(4-(7-methoxybenzofuran-2-
carboxamido)butyl)-2-oxo-2H-chromene-3-carboxamide (9k)
1
13
Fig. S12: (a) H NMR and (b) C NMR Spectra of8-methoxy-N-(4-(7-methoxybenzofuran-
-carboxamido)butyl)-2-oxo-2H-chromene-3-carboxamide (9l)
2
Table S1: Summary of the calculated pharmacokinetic descriptors
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List of Tables and Captions
Table 1: In vitro activities of coumarin-benzofuran conjugated hybrids towards anti-oxidant,
AChE inhibition and Aβ42 self-aggregation
Aβ1-42 self-
aggreg
a
b
R
R
1
AChE Inhib
Antioxid.
c
Code
n
IC50(µM)±SD
0.82±0.04
EC50 µM
Inhib.(%)
23.6±1.3
32.4±1.8
44.5±2.4
64.5±3.9
---
9
9
9
9
9
9
9
9
9
9
9
9
a
b
c
d
e
f
0
0
0
0
1
1
1
1
2
2
2
2
--
--
--
H
H
846±0.18
523±0.18
776±0.16
347±0.19
847±0.14
509±0.1
771±0.17
525±0.20
525±0.20
359±0.17
670±0.21
445±0.20
--
H
MeO 0. 36±0.02
0. 46±0.05
MeO 0. 27±0.1
0. 65±0.05
MeO 0.42±0.02
0.62±0.04
MeO 0.32±0.1
0.98±0.05
MeO 0. 18±0.05
0. 42±0.06
MeO 0. 224±0.05
MeO
MeO
H
H
H
H
16.5±1.3
68.7±1.9
72.1±2.7
--
g
h
i
MeO
MeO
H
H
H
j
H
44.5±2.2
56.5±1.8
64.5±2.6
--
k
l
MeO
MeO
--
H
[
18]
Donepezil
DFP
--
--
--
0.031.0 ± .052
--
--
--
174±0.06
15.60.2
--
d
Trolox
--
--
a
[34] b
EC50 values for the DPPH assay . The values are mean of five independent experiments
c
±
SD . Inhibition of self-induced Aβ42 aggregation (in %) (40µM). The thioflavin-T
fluorescence method was used, and the measurements were carried out in the presence of an
c
inhibitor (80 µM). The values are the mean of two independent measurements in duplicate
[31] d
[21]
(SEM < 10%).
Literature value, EC50 15.6 µM.
This article is protected by copyright. All rights reserved.

This article is protected by copyright. All rights reserved.

This article is protected by copyright. All rights reserved.