Antileishmanial activity of novel indolyl-coumarin hybrids: Design, synthesis, biological evaluation, molecular docking study and in silico ADME prediction

Article abstract of DOI:10.1016/j.bmcl.2015.12.085

In present work we have designed and synthesized total twelve novel 3-(3-(1H-indol-3-yl)-3-phenylpropanoyl)-4-hydroxy-2H-chromen-2-one derivatives 13(a-l) using Ho³⁺doped CoFe₂O₄nanoparticles as catalyst and evaluated for their potential antileishmanial and antioxidant activities. The compounds 13a, 13d and 13h were found to possess significant antileishmanial activity (IC₅₀value = 95.50, 95.00 and 99.00 μg/mL, respectively) when compared to the standard sodium stibogluconate (IC₅₀= 490.00 μg/mL). The compounds 13a (IC₅₀= 12.40 μg/mL), 13d (IC₅₀= 13.49 μg/mL), 13g (IC₅₀= 13.24 μg/mL) and 13l (IC₅₀= 13.74 μg/mL) had shown good antioxidant activity when compared with standards butylated hydroxy toluene (IC₅₀= 16.5 μg/mL) and ascorbic acid (IC₅₀= 12.8 μg/mL). After performing molecular docking studies, it was found that compounds 13a and 13d had potential to inhibit pteridine reductase 1 enzyme. In silico ADME pharmacokinetic parameters had shown promising results and none of the synthesized compounds had violated Lipinski's rule of five. Thus, suggesting that compounds from the present series can serve as important gateway for the design and development of new antileishmanial as well as antioxidant agent.

Full text of DOI:10.1016/j.bmcl.2015.12.085

Accepted Manuscript
Antileishmanial activity of novel indolyl-coumarin hybrids: Design, synthesis,
biological evaluation, molecular docking study and in silico ADME prediction
Jaiprakash N. Sangshetti, Firoz A. Kalam Khan, Abhishek A. Kulkarni,
Rajendra H. Patil, Amol. M. Pachpinde, Kishan S. Lohar, Devanand B. Shinde
PII:
S0960-894X(15)30398-X
http://dx.doi.org/10.1016/j.bmcl.2015.12.085
BMCL 23446
DOI:
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
2 September 2015
19 December 2015
24 December 2015
Please cite this article as: Sangshetti, J.N., Kalam Khan, F.A., Kulkarni, A.A., Patil, R.H., M. Pachpinde, Amol.,
Lohar, K.S., Shinde, D.B., Antileishmanial activity of novel indolyl-coumarin hybrids: Design, synthesis, biological
evaluation, molecular docking study and in silico ADME prediction, Bioorganic & Medicinal Chemistry Letters
(2015), doi: http://dx.doi.org/10.1016/j.bmcl.2015.12.085
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Antileishmanial activity of novel indolyl-coumarin hybrids: Design, synthesis, biological
evaluation, molecular docking study and in silico ADME prediction
Jaiprakash N. Sangshetti a,*, Firoz A. Kalam Khan a, Abhishek A. Kulkarni a, Rajendra H. Patil b,
, Amol. M. Pachpinde c, Kishan S. Lohar d Devanand B. Shinde e
a
Dr. Rafiq Zakaria Campus, Y.B. Chavan College of Pharmacy, Aurangabad 431001, (M.S.),
India
b Department of Biotechnology, Savitribai Phule Pune University, Pune 411007, (M.S.), India
c Department of Chemistry, Jawahar Art Science and Commerce College, Andur, Osmanabad
413603, (M. S.), India
d
Materials Research Laboratory, Srikrishna Mahavidyalaya Gunjoti, Omerga, Osmanabad
413 613, (M. S.), India
e Shivaji University, Vidyanagar, Kolhapur 416 004, (M.S.), India
*Corresponding author. Tel./fax: +91 240 2381129.
E-mail address: jnsangshetti@rediffmail.com
ABSTRACT
In present work we have designed and synthesized total twelve novel 3-(3-(1H-indol-3-yl)-3-
phenylpropanoyl)-4-hydroxy-2H-chromen-2-one derivatives 13(a-l) using Ho3+ doped
CoFe2O4 nanoparticles as catalyst and evaluated for their potential antileishmanial and
antioxidant activities. The compounds 13a, 13d and 13h were found to possess significant
antileishmanial activity (IC50 value= 95.50, 95.00 and 99.00 μg/mL, respectively) when
compared to the standard sodium stibogluconate (IC50= 490.00 μg/mL). The compounds 13a
(IC50= 12.40 μg/mL), 13d (IC50= 13.49 μg/mL), 13g (IC50= 13.24 μg/mL) and 13l (IC50=
13.74 μg/mL) had shown good antioxidant activity when compared with standards butylated
hydroxy toluene (IC50= 16.5 μg/mL) and ascorbic acid (IC50= 12.8 μg/mL). After performing
molecular docking studies, it was found that compounds 13a and 13d had potential to inhibit
pteridine reductase 1 enzyme. In silico ADME pharmacokinetic parameters had shown
promising results and none of the synthesized compounds had violated Lipinski’s rule of five.
Thus, suggesting that compounds from the present series can serve as important gateway for
the design and development of new antileishmanial as well as antioxidant agent.
Keywords: Indolyl-coumarin hybrids; Ho3+ doped CoFe2O4 nanoparticles; Antileishmanial;
Antioxidant; Molecular docking studies.
1

Leishmaniasis is a vector-borne fatal parasitic disease caused by different species of the
genus Leishmania protozoan and transmitted to humans by the bite of female phlebotomine
sand fly and mainly affects people living below poverty line. World Health Organization
(WHO) classified leishmaniasis as a major tropical disease that ranks second after malaria.1-8
It is estimated that 12 million people worldwide are infected by over 20 different species of
Leishmania and about 350 million people living in the endemic areas are at the risk to be
9, 10
infected.
No effective vaccine is available against leishmaniasis, so chemotherapy is the
only effective way to treat all forms of the disease.11, 12 However, the drugs currently used for
the treatment of human cutaneous and visceral leishmaniasis are toxic; having severe adverse
reactions (such as pancreatitis, pancytopenia, reversible peripheral neuropathy,
nephrotoxicity, cardiotoxicity, bone pain, myalgia etc.) and resistance growing towards them
13, 14
had also questioned their use.
Therefore, development of novel, effective and safe
antileishmanial agents, with reduced side effects, is a major priority for health researchers.
Free radicals can be defined as reactive chemical species having a single unpaired electron
in an outer orbit. 15 The majority of free radicals that damages biological systems are oxygen-
free radicals (reactive oxygen species), the by-products of aerobic organism’s cellular process
which generally initiates autocatalytic reactions so that molecules to which they react are
themselves converted again into free radicals which in turn mediates chain reaction to
16, 17
damage (oxidative stress) cell structures like proteins, nuclear substances etc.
This
oxidative damage accumulates during the life cycle of humans and has been implicated in
aging (neurodegeneration) and age related diseases such as cardiovascular disease,
18-20
Alzheimer's disease, cancer, and other similar chronic conditions.
Thus, the discovery
and development of novel synthetic radical scavengers attained great importance in organic
chemistry.
Our present work entails the synthesis of such derivatives which has a skeleton that
contains an orderly arrangement of coumarin and indole cores fused with chalcone like
structure (Fig. 1). The rationale behind using these three scaffolds in our design is based on
the diversity of these moieties in nature as well as in various pharmaceuticals (alone or in
combination). For example: 2H-chromene-2-ones, the oxygenated heterocycles (natural
coumarins) are found to be present in numerous natural products with wide range of
biological activities.21, 22 Iranshahi et al. reported that a prenylated sesquiterpene coumarin
Umbelliprenin (1), present as one of the components in the extract of Ferula szowitsiana
(Apiaceae) roots, found to possess significant antileishmanial activity with an IC50 value of
2

23
13.3 µM against L. major promastigotes. Other cores such as indole and chalcone were
also found to have antileishmanial properties.24-26 Representative examples of them includes:
an indole alkaloid Corynantheine (2), present in the bark of Corynanthe pachyceras
(Rubiaceae) exhibited antileishmanial activity against L. major promastigotes with an IC50
value of about 3 µM, reported by Staerk et al. 27 Kharazmi et al. reported that Licochalcone
A (3), an oxygenated chalcone, inhibits in vitro growth of both L. major and L. donovani
promastigote form. Its IC50 value was found to be in the range 2.5-4 µg/ml against L. major
promastigotes.28 These scaffolds also have been well reported for antioxidant activity (Fig.
2). Wu et al. reported that they had studied antioxidant properties of coumarin ingredients
present in the crude extract, fractions and ingredients of Cortex Fraxini (4) and the obtained
results were found to be satisfactory.29 Marques et al. reported that best antioxidant results
were obtained for the indole derivatives such as tryptophan (5) and tryptamine (6) with IC50s
of 3.50± 0.4 and 6.00± 0.60 µM, respectively.30 Butein (7), a naturally occurring substance
having chalcone backbone is one of the major active components of Dalbergia odorifera T.
Chen (Fabaceae) and also found in heartwood of Acacia, has been reported to have
antioxidant properties. 31, 32
3

Figure 1. Natural compounds reported for antileishmanial activity and design of target
compounds 13(a-l).
4

Figure 2. Various natural derivatives reported for antioxidant activity.
33
Here, in continuation of our work on synthesis of bioactive molecules, we report the
design and synthesis of a series of of some new combi-hetrocycles 13(a-l) having
combination of the coumarin, and indole linked via chalcone like chain and same were
evaluated in vitro antileishmanial activity. Since, all the three cores were also found to
possess antioxidant activities; we therefore performed their antioxidant screening too using
DPPH-radical scavenging method. In order to clarify and understand the probable binding
mode of synthesized compounds 13(a-l) for their antileishmanial activity, the compounds
were docked for possible targets for antileishmanial activity. We had also predicted their
ADME properties in silico to suggest the suitability of any of the new compounds for further
drug development.
The acetylated compound i.e. 3-acetyl-4-hydroxy-2H-chromen-2-one (9) was synthesized
by refluxing commercially available 4-hydroxy-2H-chromen-2-one (8) (25 mmol) into acetic
o
acid (133.3 mmol) in presence of phosphorous oxychloride (3 mL) at 250 C for about 3.5
hours in an oil bath. 34 The 3-acetyl-4-hydroxy-2H-chromen-2-one (9) was then subjected for
reaction with different aromatic aldehydes 10(a-l) (both 0.01 mol) in presence of 40% NaOH
using ethanol as a solvent to form different substituted chalcones 11(a-l). These chalcones
11(a-l) (0.01 mol) and indole 12 (0.01 mol) were used for synthesis of titled compounds
5

13(a-l) using 5 mol % of HoxCoFe2-xO4 (x= 0.05) as a novel magnetically recoverable and
reusable catalyst (Scheme 1) in ethanol. Ho3+ doped CoFe2O4 nanoparticles were prepared
35
and characterized by our group
(Supporting data, Fig. S1). In recent times, several
transition metal oxides in the form of nanoparticles were employed as recyclable catalysts for
36
many reactions.
In general, nanomaterials with natural morphologies containing higher
37
surface area as reactive sites allow them to act as effective catalysts for organic synthesis.
These nanoparticles have provided a simplified isolation procedure for the product, with
small amounts of catalyst, affording easy recovery and recyclability of the catalyst. In some
cases, recovery of the nanoparticles from the reaction mixtures is so difficult that
conventional techniques such as filtration or centrifugation are not enough for an efficient
recovery. To overcome this issue, the use of magnetic nanoparticles has emerged as a viable
solution; their insoluble and paramagnetic nature enables easy and efficient separation of the
catalysts from the reaction mixture with an external magnet. Recently, we have reported the
synthesis of 2,4,5 -triaryl-1H-imidazoles using Ho3+ doped CoFe2O4 nanoparticles. 38
Scheme 1. Snthesis of indolyl-coumarin hybrids 13(a-l). Reagents and conditions: (a) AcOH,
POCl3, Reflux; (b) 40% NaOH, EtOH; (c) EtOH, 5 mol % of HoxCoFe2-xO4 (x= 0.05)
nanoparticles
6

To optimize the reaction conditions, we studied a model reaction of chalcone 11a (0.01
mol) and indole 12 (0.01 mol) in ethanol as solvent using Ho3+ doped CoFe2O4 nanoparticles
as catalyst to synthesize compound 13a. We screened the nanoparticles at various loads such
as 0, 5, 10, 15 and 20 mol % (Supporting data, Table S1). When no catalyst was added for
model reaction, there was only small amount (yield 30 %) of product obtained after 180 min.
The use of 5 mol % nanaoparticle gave the compound 13a with 97 % yield in short reaction
time (45 min). The increase in amount of catalyst from 5 mol % to 10, 15 and 20 mol % did
not show any change in yield and time of reaction (Supporting data, Table S1). Therefore, 5
mol % of the catalyst Ho3+ doped CoFe2O4 nanoparticles was assumed to ensure the best
yield (97 %) in short reaction time (45 min). Thus, our results make the process under study
more attractive and interesting from the viewpoint of economy and simplicity. The recovery
and reusability of the catalyst was investigated in this reaction for model reaction (13a). After
completion of reaction (monitored by TLC), catalyst recycling was achieved by fixing the
catalyst magnetically at the bottom of the flask with a strong magnet, after which the solution
was taken off with a pipette, the solid washed twice with acetone and the fresh substrates
with solvent was introduced into the flask, allowing the reaction to proceed for the next run.
The catalyst was consecutively reused three times without any noticeable loss of its catalytic
activity (Cycle number and yield of 13a: 1, 97 %; 2, 97 %; 3, 93 %). So the catalyst was
found to be recyclable and reusable.
We further expanded our series and synthesized 12 novel derivatives of 3-(3-(1H-indol-3-
yl)-3-phenylpropanoyl)-4-hydroxy-2H-chromen-2-one 13(a-l). The reaction preceded
smoothly under mild reaction conditions (stirring at room temperature). The isolated yields of
synthesized compounds were in the range of 80-97 % and reactions were completed in about
45-60 min (monitored by TLC). Melting points were determined in open capillary tubes and
are uncorrected. The physical data for the compounds 13(a-l) are presented in Table 1. The
1
formation of synthesized compounds was confirmed by IR, H NMR, 13C NMR and Mass
spectral analysis.
7

Table 1
Physical data for 3-(3-(1H-indol-3-yl)-3-phenylpropanoyl)-4-hydroxy-2H-chromen-2-ones
13(a-l)
#
Entry
R
Molecular formula
(mw)
Reaction
%
M.P.*
Rf
time (min) Yield
value
0.74
0.66
H
2-Cl
C26H19NO4 (409)
C26H18ClNO4 (444)
C26H18ClNO4 (444)
C26H17Cl2NO4 (478)
C26H18FNO4 (427)
C27H21NO5 (439)
C28H23NO6 (470)
C28H23NO6(470)
C28H23NO6 (470)
C26H19NO5 (425)
C26H19NO6 (441)
C27H18N2O4 (434)
45
60
60
55
50
45
60
45
50
45
60
60
97
83
85
95
92
97
97
97
96
91
89
87
72-74
76-78
13a
13b
13c
13d
13e
13f
13g
13h
13i
4-Cl
118-120 0.68
124-126 0.61
2-Cl, 6-Cl
4-F
78-80
74-76
0.63
0.77
4-OCH3
2-OCH3, 4-OCH3
2-OCH3, 5-OCH3
3-OCH3, 4-OCH3
4-OH
118-120 0.79
128-130 0.78
126-128 0.83
58-60
92-94
80-82
0.69
0.65
0.70
13j
13k
13l
3-OH, 4-OH
4-CN
* Melting point uncorrected; # Solvent system: n-Hexane: Ethyl acetate (8:2).
The title compounds 13(a-l) were tested for their in vitro antileishmanial activity against a
culture of Leishmania donovani promastigotes (NHOM/IN/80/DD8). Parasite viability was
evaluated using a modified 3-(4,5-dimethylthiazol-2 yl)-2,5-diphenyl tetrazolium bromide
(MTT) assay wherein the amount of formazan produced is directly proportional to the
number of metabolically active cells.39 The concentration that decreased cell growth by 50%
(IC50) was determined by graphic interpolation and data obtained depicted in Table 2. Sodium
8

stibogluconate and pentamidine were used as standard drugs. Compounds 13(a-l) showed
varying degrees of antileishmanial activities with IC50 ranging between 95.00 to 287.50
μg/mL. Amongst all tested compounds 13a, 13d and 13h were found to be most promising
compounds showing IC50 value of 95.50 μg/mL, 95.00 μg/mL and 99.00 μg/mL, respectively
when compared with sodium stilbogluconate. All the synthesized compounds showed better
activity than standard sodium stibogluconate (IC50 = 490 μg/mL) against L. donovani
promastigotes.
Table 2
Evaluation of Antileishmanial and antioxidant activities.
Entry Antileishmanial activity IC50 Antioxidant activity IC50
Pteridine reductase 1
(μg/mL)
95.50
102.00
125.00
95.00
155.50
174.00
249.50
99.00
212.00
287.50
231.00
101.75
490.00
5.50
(μg/mL)
12.40
39.62
46.35
13.49
47.35
16.21
13.24
19.02
63.16
58.91
71.08
13.74
*
Binding energy (kcal/mol)
-83.39
-79.01
-78.42
-84.82
-76.98
-76.84
-73.60
-82.77
-76.49
-72.15
-74.63
-80.55
*
13a
13b
13c
13d
13e
13f
13g
13h
13i
13j
13k
13l
STD1
STD2
STD3
STD4
*
*
16.50
12.80
*
*
*
*
IC50 represents the mean values of three replicates; standard errors were all within 10% of the
mean; (*) denotes not performed; STD1: Sodium stibogluconate; STD2: Pentamidine; STD1:
Butylated hydroxytoluene; STD4: Ascorbic acid
9

Structural-activity relationship (SAR) revealed that coumarin, indolyl and chalcone like
cores are important for antilishmanial activity (Table 2). Modification of the parent
compounds with various substituents on phenyl ring such as halogen, hydroxyl, methoxyl,
amino and cyano were performed to explore the SAR of these synthesized compounds 13(a-
l). Compound 13a with no substitution on phenyl ring showed good antileishmanial activity
(IC50=95.50 μg/mL). Introduction of 2-Cl 13b (IC50=102.00 μg/mL) and 4-Cl 13c
(IC50=125.00 μg/mL) on phenyl ring showed significant antileishmanial activity. Intoduction
of two chloro groups i.e. 2,6-dichloro 13d (IC50=95.00 μg/mL) on phenyl ring further
increased antileishmanial potency and found to be most promising from the synthesized
series. Furthermore, phenyl ring containing 4-F 13e (IC50=155.50 μg/mL) was found to
reduce activity when compared with 13a, 13b, 13c and 13d. Therefore, there may be chances
of interference of electronegativity of atoms and position of atoms with the activity.
Introduction of 4-OCH3 13f (IC50=174.00 μg/mL) on phenyl ring was found to reduce the
activity. Replacement of 4-OCH3 13f with 4-OH 13j (IC50=287.00 μg/mL) led to decrease in
antileishmanial activity about 2 folds. Compounds 13g (IC50=249.50 μg/mL) and 13i
(IC50=212.00 μg/mL) with 2,4-dimethoxy and 3,4-dimethoxy substitutions on phenyl ring,
respectively further reduced the activity. But introduction of 2,5-dimethoxy 13h (IC50=99.00
μg/mL) on phenyl ring was found to be increase in activity by 2 folds when compared with
other methoxy substituted derivatives 13f, 13g and 13i. The replacement of 3,4-dimethoxy
13i with 3,4-dihydroxy 13k (IC50=231.00 μg/mL) did not lead to any significant change in
activity. The introduction of 4-CN group on phenyl ring had also shown good activity
(IC50=101.75 μg/mL) as compared to most promising compounds 13a, 13d and 13h. As
observed from activity data, compounds 13b, 13c, 13d, 13e and 13l with electron
withdrawing groups substituents at phenyl ring are more effective than compounds 13f, 13g,
13i, 13j and 13k with electron donating substituent at phenyl ring.
Antioxidant activities of the synthesized compounds 13(a-l) were measured using the 2,2-
diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay.40 The percent of inhibition (I %)
of free radical production from DPPH was calculated by equation: % of scavenging= [(A
control- A sample)/A blank] ×100. Where ‘A control’ is the absorbance of the control
reaction (containing all reagents except the test compound) and ‘A sample’ is the absorbance
of the test compound. DPPH radical scavenging activity is the most commonly used method
for screening the antioxidant activities of various natural as well as synthetic antioxidants.
Butylated hydroxytoluene (BHT) and ascorbic acid (AA) have been used as standard drugs
10

for the comparison of antioxidant activity and the observed results are summarized in Table
2. According to the DPPH assay, compounds 13a (IC50= 12.40 μg/mL), 13d (IC50= 13.49
μg/mL), 13f (IC50= 16.21 μg/mL), 13g (IC50= 13.24 μg/mL) and 13l (IC50= 13.74 μg/mL)
showed promising radical scavenging activities when compared with synthetic antioxidant
BHT (IC50= 16.5 μg/mL). The compound 13a (IC50= 12.40 μg/mL) showed equipotent
activity as that of natural antioxidant ascorbic acid (IC50= 12.80 μg/mL). Compounds 13b
(IC50= 39.62 μg/mL), 13c (IC50= 46.35 μg/mL), 13e (IC50= 47.35 μg/mL), 13h (IC50= 19.02
μg/mL), 13i (IC50= 63.16 μg/mL), 13j (IC50= 58.19 μg/mL) and 13k (IC50= 71.08 μg/mL)
were found to be inactive when compared with standard antioxidants.
From the antioxidant activity data (Table 2), the synthesized compounds had shown
moderate to good antioxidant activity. The compound 13a (IC50= 12.40 μg/mL) with no
substitution on phenyl ring was found most potent for antioxidant properties when compared
to standard drugs BHT (IC50= 16.50 μg/mL) and ascorbic acid (IC50= 12.80 μg/mL). The
introduction of 2-Cl 13b (IC50=39.62 μg/mL) and 4-Cl 13c (IC50=46.32 μg/mL) on phenyl
ring led to reduced in antioxidant activity by 3-4 fold when compared with compound 13a.
But introduction of 2,6-dichloro 13d (IC50=13.49 μg/mL) led to increase in activity and was
more potent when compared with standard BHT. Replacement of 4-Cl with 4-F 13e
(IC50=47.35 μg/mL) did not show any change in antioxidant activity. The methoxyl (-OCH3)
substituted compounds 13f (4-OCH3, IC50= 16.21 μg/mL), 13g (2,4-OCH3, IC50= 13.24
μg/mL) and 13h (2,5-OCH3, IC50= 19.02 μg/mL) was found to be show good antioxidant
activity except compound with 3,4-OCH3 13i (IC50=16.50 μg/mL) when compared with
standard drugs. The replacement of 4-CH3 13f with 4-OH 13j (IC50= 58.91 μg/mL) led to
decrease in antioxidant activity by 3 folds. The compound 13k with 3,4-OH on phenyl ring
was found to most inactive compounds of the synthesized series with IC50= 71.08 μg/mL.
The 4-CN substitution on phenyl ring (compound 13l) showed remarkable antioxidant
activity (IC50=13.74 μg/mL) more than that of standard BHT (IC50= 16.50 μg/mL) and nearly
same activity as that of ascorbic acid (IC50=12.80 μg/mL).
Cytotoxic study was performed on HeLa cell line and evaluated by 3-(4,5-dimethylthiazol-
2-yl)-2,5-diphenyltetrazolium bromide (MTT) method.41 The cells were seen under the
microscope (Ziess, Germany) at 10X magnification. The compound 13d from the synthesized
series was found to be most promising showing good antileishmanial (IC50= 95.00 μg/mL)
and antioxidant (IC50= 13.49 μg/mL) activities. Therefore, cytotoxic study against HeLa cell
11

line of compound 13d was carried out at concentration of 100 μg/mL. The compound 13d
had shown no cytotoxicity effect against HeLa cell lines to its tested concentration (Fig. 3).
13d
Figure 3. Cytotoxic study against HeLa cell line of most promising compound 13d.
Molecular docking study of the synthesized compounds 13(a-l) was performed to clarify
understand the probable binding interactions with various antileishmanial targets using
standard protocol implemented in VLife MDS 4.3 package.42 With this purpose, crystal
structures of various antileishmanial targets including, Pteridine reductase 1 (PDB ID:
2XOX),43 Adenine phosphoribosyltransferase (PDB ID: 1QCD),44 UMP synthase (PDB ID:
3QW4) 45 and MapK (PDB ID: 4QNY) 46 were obtained from the Protein Data Bank in order
to prepare protein for docking study. The docking study was performed using Vlife MDS 4.3
software by GRIP method. The compounds 13(a-l) have shown some significant binding
energy but did not show any binding affinity in the active site of Adenine phosphoribosyl
transferase and UMP synthase enzymes. The compounds have shown poor binding energy
with MapK enzyme (Supporting data, Table S2). But in turn, the compounds showed good
binding energy (-84.82 to -72.15 kcal/mol) toward Pteridine reductase 1 and docked well
into active pocket against this enzyme (Table 2). Pteridine reductase 1 (PTR1), a Leishmania
enzyme responsible for salvage of pteridines is potential target for chemotherapeutic
intervention.47 The binding mode of compounds 13a and 13d in active pocket of Pteridine
reductase 1 is shown in Fig. 4.
12

13a
13d
Figure 4. Docking study of compounds 13a and 13d with Pteridine reductase 1 (PDB ID:
2XOX). Ligands are shown in red color. Hydrogen bonds are shown in green color.
The binding energy of the compounds 13(a-l) and their antileishmanial activity showed the
corresponding results. The active compounds 13a and 13d showed lowest interaction energy
that is -83.39 kcal/mol and -84.82 kcal/mol, respectively. In case of compound 13a, the
13

compound was held in active site by forming various interactions with amino acid residues
like, THR12, GLY13, ALA15, LYS16, ARG17, HIS36, TYR37, HIS38, ARG39, SER40,
LEU66, ASN109, and SER111. The amino acid GLY13 (1.73 Å) had formed hydrogen
bonding with nitrogen of indole ring, and amino acids HIS38 (2.47 Å), ARG39 (1.58 Å) and
SER40 (1.92 Å) had formed hydrogen bondings with oxygen of chalcone like chain. In case
compound 13d, the compound was held in active site by forming various interactions with
amino acid residues like, THR12, SER14, ARG39, LEU66, SER67, ASN109, ALA110,
ASN147, SER111, LEU143, SER146, ASN147, ALA150, PRO151, and LEU188. The amino
acids SER146 (1.28 Å) and ASN147 (2.58 Å) had formed hydrogen bond with –OH of
coumarin ring. The chloro substituent of phenyl ring was held in active pocket by forming
van der waals interactions with amino acids SER111. On the basis of activity data and
docking result, it was found the compounds 13a and 13d had potential to inhibit pteridine
reductase 1 enzyme.
An in silico study of synthesized compounds 13(a-l) was performed for prediction of
ADME properties (Table 3). In this study, we calculated molecular volume (MV), molecular
weight (MW), logarithm of partition coefficient (miLogP), number of hydrogen bond
acceptors (n-ON), number of hydrogen bonds donors (n-OHNH), topological polar surface
48
area (TPSA), number of rotatable bonds (n-ROTB) and Lipinski’s rule of five
using
Molinspiration online property calculation toolkit.49 Absorption (% ABS) was calculated by:
% ABS= 109-(0.345×TPSA).50 From all these parameters (Table 3), it can be observed that
all titled compounds exhibited a good % ABS (66.30–80.26 %). it was observed that none of
the compounds violated Lipinski's rule of five, suggesting that the compounds 13(a-l) may be
developed as good drug candidates. A molecule likely to be developed as an orally active
drug candidate should show no more than one violation of the following four criteria: logP
(octanol-water partition coefficient) ≤5, molecular weight ≤500, number of hydrogen bond
acceptors ≤10 and number of hydrogen bond donors ≤5.51 Since all the synthesized
compounds 13(a-l) followed this criteria for orally active drug and therefore can be
developed as oral drug candidates.
14

Table 3
Pharmacokinetic parameters important for good oral bioavailability of synthesized compound
13(a-l).
Entry %ABS
TPSA
(A2)
n-
ROTB
MV
-
MW
miLogP
n-ON
acceptors donors
n-OHNH Lipinski’s
violations
Rule
-
-
-
<500
≤5
<10
5
<5
2
≤1
80.26
83.30
5
361.01 409.44
374.55 443.89
374.55 443.89
388.08 478.33
365.94 427.43
386.56 439.47
412.10 469.49
412.10 469.49
412.10 469.49
369.03 425.44
377.05 441.44
377.87 435.45
4.78
0
1
1
1
0
0
0
0
0
0
0
0
13a
80.26
80.26
80.26
80.26
77.08
73.89
73.89
73.89
73.28
66.30
72.05
83.30
83.30
5
5
5
5
6
7
7
7
5
5
5
5.41
5.46
6.04
4.95
4.84
4.83
4.83
4.43
4.30
3.81
4.54
5
5
5
5
6
7
7
7
6
7
6
2
2
2
2
2
2
2
2
3
4
2
13b
13c
13d
13e
13f
13g
13h
13i
83.30
83.30
92.53
101.77
101.77
101.77
103.53
123.76
107.09
13j
13k
13l
% ABS: percentage absorption, TPSA: topological polar surface area, n-ROTB: number of
rotatable bonds, MV: molecular volume, MW: molecular weight, milogP: logarithm of
partition coefficient of compound between n-octanol and water, n-ON acceptors: number of
hydrogen bond acceptors, n-OHNH donors: number of hydrogen bonds donors.
In conclusion, design, synthesis and biological of a novel series of 3-(3-(1H-indol-3-yl)-3-
phenylpropanoyl)-4-hydroxy-2H-chromen-2-one 13(a-l) has been presented. Use of 5 mol %
of Ho3+ doped CoFe2O4 nanoparticles as a catalyst helps in fast conversion of substituted
coumarin chalcones to desired products therefore proving its advantage. Good yields and
reusability of catalyst imparts further advantage of using it in the reaction. Thus, our results
made the process under study more attractive and interesting from the viewpoint of economy
and simplicity. Newly synthesized compounds were evaluated for both antileishmanial as
well as antioxidant activities. Based on the activity data, SAR for the series has been
15

developed. Compounds 13a (IC50= 95.50 μg/mL), 13d (IC50= 95.00 μg/mL) and 13h (IC50=
99.00 μg/mL) were found to possess significant antileishmanial activity when compared with
standard sodium stibogluconate (IC50= 490.00 μg/mL). Compound 13a (IC50=12.40 μg/mL)
with no substitution on phenyl ring was found to be most potent antioxidant when compared
to standard drugs BHT (IC50= 16.50 μg/mL) and ascorbic acid (IC50= 12.80 μg/mL). After
performing docking studies, on the basis of activity data and docking result, it was found the
compounds 13a and 13d had potential to inhibit pteridine reductase 1 enzyme. In silico
physicochemical pharmacokinetic parameters for synthesized compounds had shown
promising results. Thus, suggesting that the compounds from the present series can serve as
important gateway for the design and development of new antileishmanial agents along with
antioxidant activity.
Acknowledgements
The authors are thankful to Mrs. Fatma Rafiq Zakaria, Chairman, Maulana Azad
Educational Trust and Dr. Zahid Zaheer, Principal, Y.B. Chavan College of Pharmacy, Dr.
Rafiq Zakaria Campus, Aurangabad 431001 (MS), India for providing the laboratory facility.
The authors are also thankful Department of Biotechnology, Savitribai Phule Pune
University, Pune (MS), India for performing antileishmanial and antioxidant screening. The
authors are also thankful to SAIF, Punjab University, Chandigarh, India for providing NMR
spectra.
16

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19

Figure captions
Figure 1. Natural compounds reported for antileishmanial activity and design of target
compounds 13(a-l).
Figure 2. Various natural derivatives reported for antioxidant activity.
Figure 3. Cytotoxic study of most promising compound 13d against HeLa cell line.
Figure 4. Docking study of compounds 13a and 13d with Pteridine reductase 1 (PDB ID:
2XOX). Ligands are shown in red color. Hydrogen bonds are shown in green color.
20

Graphical Abstract
21