Synthesis, characterization and antimalarial activity of isoquinoline derivatives

Article abstract of DOI:10.1007/s00044-020-02642-0

This paper presents synthesis of novel decorated isoquinolines that can be used as antimalarial agents. Two series of compounds, isoquinoline phenyl and isoquinoline triazole derivatives, were synthesized via the Suzuki–Miyaura and Sharpless–Fokin reactio

Full text of DOI:10.1007/s00044-020-02642-0

MEDICINAL
CHEMISTRY
RESEARCH
Medicinal Chemistry Research
https://doi.org/10.1007/s00044-020-02642-0
ORIGINAL RESEARCH
Synthesis, characterization and antimalarial activity of isoquinoline
derivatives
1
Artitiya Thiengsusuk2 Opa Vajragupta3 Phunuch Muhamad4
●
●
●
Sewan Theeramunkong
Received: 5 April 2020 / Accepted: 23 September 2020
© Springer Science+Business Media, LLC, part of Springer Nature 2020
Abstract
This paper presents synthesis of novel decorated isoquinolines that can be used as antimalarial agents. Two series of
compounds, isoquinoline phenyl and isoquinoline triazole derivatives, were synthesized via the Suzuki–Miyaura and
Sharpless–Fokin reactions respectively. The final compounds were investigated for their antimalarial activity in cell-based
experiments. In the series of isoquinoline phenyl derivatives, compound 6 exhibited interesting antiplasmodial activity
against chloroquine resistant (K1) and chloroquine sensitive (3D7) strains of P. Falciparum with IC50 1.91 0.21 μM and
2.31 0.33 μM, respectively. In the series of isoquinoline-triazole derivatives, compound 15 was the most active against
resistant (K1) and sensitive (3D7) strains of P. falciparum with IC50 4.55 0.10 μM and 36.91 2.83 μM, respectively. The
respective resistance indices of compounds 6 and 15 against K1 and 3D7 were 0.83 and 0.12. Compound 15 was
promisingly effective against the resistant strain. The results of this study provided useful contributions for further lead
discovery and lead modification of modern rational drug design.
●
●
●
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Keywords Antimalarial activity Plasmodium falciparum Chloroquine resistant Isoquinoline 1,2,3-triazole
Abbreviations Pfmdr1 Plasmodium falciparum multidrug resistance 1
P.
Plasmodium
Pfcrt
Plasmodium falciparum chloroquine resistant
transporter
Introduction
Malaria is a major disease and public health concern. It is
mainly prevalent in Africa, South-East Asia and the Eastern
Mediterranean. Malaria is caused by Plasmodium parasite.
The main transmission vector is the female Anopheles mos-
quito [1, 2]. Mosquitoes taking a blood meal on infected
individuals ingest parasites into the midgut. Subsequently, the
parasites move into the mosquito’s salivary glands and sub-
sequently infect a human host [3–6]. Humans can be infected
with one or more of the following five strains: P. falciparum,
P. vivax, P. malariae, P. ovale and P. knowlesi [4–7]. Cur-
rently, P. falciparum and P. vivax are the prevalent protozoan
species causing high mortality, P. falciparum being the par-
ticular most virulent species [2]. The clinical symptoms of
malaria are mainly caused by schizont rupture and destruction
of erythrocytes. Major symptoms of malaria are fever, chills,
headaches, and diaphoresis [8].
Supplementary information The online version of this article (https://
doi.org/10.1007/s00044-020-02642-0) contains supplementary
material, which is available to authorized users.
* Sewan Theeramunkong
sewan@tu.ac.th
1
Thammasat University Research Unit in Drug, Health Product
Development and Application (DHP-DA), Department of
Pharmaceutical Sciences, Faculty of Pharmacy, Thammasat
University, Bangkok, Pathumthani 12120, Thailand
2
Center of Excellence in Pharmacology and Molecular Biology of
Malaria and Cholangiocarcinoma, Chulabhorn International
College of Medicine, Thammasat University, 99 Moo 18
Phahonyothin Road, Klongluang, Bangkok, Pathumthani 12120,
Thailand
3
Department of Pharmaceutical Chemistry, Faculty of Pharmacy,
Mahidol University, 447 Sri-Ayuthaya Road, Rajathevi,
Bangkok 10400, Thailand
Currently, the commercially available antimalarial drugs
are quinoline derivatives, artemisinin derivatives, antifolates
and some antimicrobial agents [9]. Antimalarial agents
principally aim to inhibit one or two stages in the life cycle
of the parasite. Among these drugs, quinine and chloroquine
4
Drug Discovery and Development Center, Office of Advanced
Science and Technology, Thammasat University, 99 Moo 18
Phahonyothin Road, Klongluang, Bangkok, Pathumthani 12120,
Thailand

Medicinal Chemistry Research
are the oldest drugs and the first choice for treating all
species of malaria [10]. An artemisinin-based combination
therapy based regimen could be applied where chloroquine
is ineffective in eradiacating malaria [10]. However, the
emergence of drug resistance to available antimalarial drugs
is a serious problem [11]. Two of the five human malaria
parasite species, P. falciparum and P. Vivax, are frequently
resistant. The incidence of chloroquine resistance of
P. falciparum is a major problem for malarial control. Both
artemisinin and its partner drugs are also threatened, espe-
cially in Africa [2]. On the other hand, P. malariae or
P. ovale remain unknown for current drug resistance [4–6].
Genetic mutations or genetic malfunction on the major
pathways of drug resistance, i.e., enzymes or transporters,
are the main factors leading to drug resistance [12]. The
examples of multidrug resistant strains of P. falciparum are
Dd2, K1 and W2 [13]. The most common cause of chlor-
oquine resistance is derived from mutation in the Pfmdr1
and Pfcrt genes in P. falciparum [14, 15].
To afford better malaria control, many researchers have
sought new compounds or to modify the molecular structure
of available current drugs such as chloroquine [10, 16]. The
mechanism of chloroquine is inhibition of hemozoin for-
mation. Normally, the parasites digest hemoglobin to
release free heme which is toxic to the parasites themselves.
However, parasites have a self-protective mechanism,
crystallizing heme to hemozoin which is insoluble and
harmless for the parasites [17, 18]. Chloroquine is able to
prevent the formation of hemozoin. In particular, the current
proposed mechanism is accumulation of chloroquine in
acidic digestive vacuoles of intraerythrocytic trophozoites
by a weak base mechanism [19]. At physiological pH,
chloroquine can exist in three forms: unprotonated, mono-
protonated, and diprotonated forms. Only uncharged
chloroquine can permeate the erythrocyte membrane and
finally reach the vacuole. In this vacuole, displaying acid
condition chloroquine molecules become protonated and,
since membranes are not permeable to charged species, the
drug accumulates into the acidic digestive vacuole [20, 21].
Another mechanism of chloroquine, the inhibition of heme
polymerase in the food vacuole of the malaria trophozoite,
results in a lack of essential amino acids and leads to
parasite death. There have been several studies on synthesis
of chloroquine derivatives including their metal complexes
to optimize the bioactivity of synthesized compounds [10].
The substituted chloroquine derivatives were reported as
having interesting antimalarial activity. Azoles are an
important class of heterocyclic compounds containing
nitrogen with various biological activities [22, 23]. The
linker of azoles with the core quinoline structure was known
to increase its antimalarial activities [10]. A plausible reason
for this was heme complexation through azoles and pre-
sented antimalarial activity [23].
The aim of this study is to find new compounds with
antimalarial properties, particularly the ability to tackle the
chloroquine resistant Plasmodium strains. Naturally, we
maintained the core structures which preserve nitrogen
atoms in the ring, similar to quinoline, but we decided to
alter the position of nitrogen atoms probing the antimalarial
activity of isoquinolines and attempt to synthesize quino-
lines containing a phenyl or triazole ring as shown in Fig. 1.
We decorated the isoquinoline scaffold with two reactions,
namely (i) the Suzuki–Miyaura reaction and (ii) the
Sharpless–Fokin reaction to afford two series; isoquinoline-
phenyl and isoquinoline-triazole (Fig. 1).
The modification of the two series is displayed in Fig. 2.
All synthesized compounds were programmed to examine
biological activity in two strains, the chloroquine-sensitive
strain (3D7) and the chloroquine resistant strain (K1) Plas-
modium falciparum. We determined the resistance index
(Ri) as the ratio of the IC50 value of the resistant strain to that
of the sensitive strain. The resistance indices of K1 and 3D7
were calculated and eventually assessed for any relationship
between structures and activities of compounds.
Results and discussion
Chemical synthesis
To generate new compounds, we rationalized the structure of
the isoquinoline core structure and modified it in two series
(Fig. 2), isoquinoline phenyl derivatives 1–13 and iso-
quinoline triazole derivatives 14–32. Isoquinoline phenyl
derivatives were prepared using the Suzuki–Miyaura reaction
(Scheme 1). This cross-coupling reactions were in which
commercially available 4-bromoisoquinoline and different
aryl boronic acid in the presence of palladium as a catalyst
Fig. 1 Concept of structural modification
Fig. 2 Modified structures of the two drug series a isoquinoline-phenyl
and b isoquinoline-triazole

Medicinal Chemistry Research
Table 1 Percentage yield and antimalarial activity of isoquinoline
phenyl derivatives against chloroquine sensitive (3D7) and
chloroquine resistant (K1) strains of P. falciparum and resistance
index (Ri)
Scheme 1 Synthesis of compounds 1–13 via the Suzuki–Miyaura
reaction
Yield
(%)
Resistance index
(Ri)
Antimalarial activity
Entry
R1
3D7
IC50 (µM)
K1
IC50 (µM)
1
2
81
20
26.05 1.99
27.11 2.24
52.44 6.95
37.38 1.40
2.01
Scheme 2 Synthesis of compounds 14–32 via the click reaction
under basic conditions. For isoquinoline triazole derivatives,
we proceeded to synthesize by using the click reaction
(Scheme 2). The archetypical click chemistry is a two-
component reaction in which an alkyne and an azide react in
the presence of copper(I) catalyst to give rise to a 1,4-dis-
ubstituted 1,2,3-triazole. We therefore use different azide
compounds bearing the desired triazoles. The azide com-
pounds were prepared as described in Utsintong et al. [24].
The average percentage yields of isoquinoline phenyl
derivatives, compounds 1–13 were more than 70 (Table 1).
Compound 2 obtained the lowest percentage yield (20%)
possibly due to the presence of an unprotected hydroxyl
group. In the series of isoquinoline triazole derivatives,
compounds 14–32, the 1,2,3-triazole ring was easily obtained
via azide-alkyne cycloaddition in the presence of a small
catalytic amount of copper (I) giving the desired products
with percentage yields ranging from 19 to 88% (Table 2). All
synthesized compounds were purified by column chromato-
1.38
1.05
3
4
92
84
30.97 2.40
47.77 5.35
32.49 1.49
30.47 2.86
0.64
5
6
91
73
30.01 3.61
2.31 0.33
22.33 2.27
1.91 0.21
0.74
0.83
7
8
79
63
36
78
42.01 6.99
19.78 3.41
0.47
1.10
1.45
0.64
203.73
31.41
223.29
25.70
1
9
29.98 4.10
73.55 7.63
35.59 0.82
43.58 3.92
47.31 3.33
39.52 7.75
graphy and characterization by FTIR and H-NMR. Full
characterization was performed on the compounds most
active against P. falciparum. The typical aromatic ring band
at 3100–3000, 1600–1450 cm−1 in IR and signal at around
10
1
δ9.20–8.00 ppm in H NMR spectra of all derivatives con-
firm the presence of isoquinoline ring. The presence of trai-
zole ring in compounds 14–32 was confirmed by the
observed 1H singlet at δ 8.3 ppm in 1H NMR spectra and the
signal at around δ 133 ppm in 13C NMR. Compound 6
conatins m-OCH3 and is confirmed by signal at δ 3.85 ppm
(3H, singlet) in 1H NMR along with signals at δ 55.3 ppm in
13C NMR. In the mass spectrum of compound 6 [M+H]+
peak was observed confirming the corresponding structure.
The complete spectral data along with the assignments are
provided in the Experimental section and partial spectra data
are provided in the Supplementary section.
11
33
1.11
12
13
78
25
41.60 3.93
21.86 2.14
14.40 0.61
19.53 3.57
0.35
0.89
Ri (resistance index) is the ratio of the IC50 of the K1 strain to the IC50
of 3D7 strain
are summarized in Tables 1–3. All synthesized compounds
were active against the 3D7 strain of P. falciparum, with
IC50 from 2.31 to 364.93 µM. The activity of all synthesized
compounds against the K1 strain of P. falciparum with IC50
values from 1.91 to 1104.51 µM. Compound 6 was the most
Antimalarial activity of synthetic compounds
The IC50 values against P. falciparum of individual syn-
thesized compounds and four standard antimalarial drugs

Medicinal Chemistry Research
Table 2 Percentage yield and antimalarial activity of isoquinoline
triazole derivatives against 3D7 and K1 strains of P. falciparum and
resistance index (Ri)
active of all 32 compounds with an IC50 value of 2.31
0.33 µM and 1.91 0.21 µM against 3D7 and K1, respec-
tively. The less potent compounds were compound 16 with
an IC50 value of 364.93 16.79 µM against 3D7 and com-
pound 24 with an IC50 value of 1104.51 133.17 µM
against K1.
Fidock and Kalra categorized antimalarial activity into
three classes: potent activity if the IC50 is <1 µM, moder-
ately potent if the IC50 is between 1 and 5 µM, and weak
activity if the IC50 is >5 µM [25, 26]. Under this classifi-
cation, only compound 6 was a moderate antimalarial agent
against both strains, while compound 15 was as moderate
antimalarial agent against the resistant strain.
In addition, the resistance indices (Ri) of K1 and 3D7
were calculated by finding the ratio between the IC50 of the
resistant strain (K1) to that of the sensitive strain (3D7). A
high index value represents the resistance of the tested
compound in the sensitive strain. One-third of the thirty-two
compounds were equipotent against both 3D7 and
K1 strains. Compounds 12, 15, 16 and 25 displayed lower
Ri values than reference standards (compound 12: Ri 0.35,
15: Ri 0.12, 16: Ri 0.26, 25: Ri 0.24) (Tables 1–3). Com-
pound 15 was the most potent against K1 strains, with an
IC50 of 4.55 0.1 µM (Fig. 3).
Regarding the isoquinoline-phenyl series, the results
displayed that the position of substituted groups on the
phenyl ring connected to the isoquinoline was very
important. Compared with compound 1, substitution with
one donor group (–OCH3) in the meta position on phenyl
ring as compound 6 resulted in a 10 and 25-fold increment
in activity against P.falciparum 3D7 and K1 respectively.
In contrast, the potency of analogues substituted with
donor groups (–OCH3, –OH, –SH) or a withdrawing
group (–SO2CH3) in the ortho/para position displayed a
distinct reduction of activity. This substituition showed
improper orientation resulting in activity decrement. In
addition, disubstition of methoxy groups at meta orien-
tation resulted in a 10–20-fold decrement in anti-
plasmodial activity. This study demonstrated the possible
potential of modification on the phenyl ring in increasing
antimalarial activity.
Antimalarial activity
Yield
(%)
Resistance
index (Ri)
Entry
X
Y
R2
R3
3D7
IC50 (µM)
K1
IC50 (µM)
14
C
N
H
35
42
57.96 11.39
39.11 1.71
0.67
0.12
15
16
17
C
C
C
N
N
N
H
H
H
36.91 2.83
4.55 0.10
73
43
364.93 16.79 95.76 15.57
0.26
0.68
122.81 4.27
62.02 5.05
83.18 6.50
47.72 3.67
18
C
N
H
25
0.77
19
20
C
C
N
N
H
H
88
19
NA
139.98 13.91 205.98 17.13
1.47
21
22
C
C
N
N
H
H
41
38
57.15 5.05
12.51 2.02
58.10 5.95
14.28 2.97
1.02
1.14
23
24
C
C
N
N
H
H
74
23
35.81 1.76
51.95 5.85
1.45
34.53 2.96 1104.51 133.17
31.99
25
C
N
H
36
47.46 0.82
11.19 1.72
0.24
26
27
C
C
N
C
H
H
32
33
181.36 23.08
36.97 45.90
91.65 7.78
42.19 40.46
0.51
1.14
When considering the isoquinoline-triazole series
without a methylene bridge between the 1,2,3-triazole and
the phenyl ring, the change of substituted position from
para- to meta- on the phenyl ring had a significant effect
on antiplasmodial activity against both 3D7 and K1 strains
as shown in the resistant index values (Table 2). The
results show that the para-substituted chlorine in com-
pound 15 was the preferred group on phenyl ring but meta-
substitution in compound 16 resulted in a tenfold decre-
ment in activity against both 3D7 and K1 strains. Fur-
thermore, the isoquinoline-triazole series with one
methylene bridge and substitution at p-position of the
28
29
C
C
C
C
H
H
83
30
160.25 10.85
83.87 18.05
65.12 6.93
91.69 34.70
0.41
1.09
30
31
32
N
N
N
C
C
C
Cl
Cl
Cl
39
39
32
130.77 17.24 154.98 14.13
1.19
NA
91.79 6.66
72.05 7.55
0.78
Values are mean SDs of three times repetition
NA Not applicable, / excessively high values/undeterminable

Medicinal Chemistry Research
Table 3 Antimalarial activity of
four current drugs against 3D7
and K1 strains of P. falciparum
and resistance index (Ri)
Compound
Antimalarial activity
Resistance index (Ri)
3D7 IC50 (μM)
K1 IC50 (deM)
Chloroquine (CQ)
Quinine (Q)
0.007 0.001
0.098 0.015
0.161 0.014
0.303 0.013
23
3.09
0.41
0.47
Mefloquine (MQ)
Dihydroartemisinin (DHA)
0.029 0.003
0.012 0.002
0.00456 0.0012
0.00216 0.0005
Resistance index between 3D and K1 strains
of some synthesized compounds and drugs
0.80
0.70
0.60
0.50
0.40
0.30
0.20
0.10
0.00
12 14 15 16 25 CQ
Q DHA MQ
Compound
Fig. 3 Resistance index and chemical structures of some synthesized compounds
phenyl ring still maintained activity. Compound 22 with
tert-butyl was the most active for antimalarial activity on
both strains.
growth was observed during a 60–72 h incubation period
(Fig. 4). Compound 6 inhibited parasite growth and
multiplication in both 3D7 and K1 strains since h12 at the
ring stage. On the other hand, under negative control,
parasites still developed to the trophozoite stage. Parasite
shrinkage and death were observed after 12 h incubation.
Compound 15 had markedly different antimalarial activ-
ity against 3D7 and K1 strains with higher specific
activity against K1 than 3D7. In addition, compound 15
was able to delay parasite growth since h24, with early
schizonts observed in the control sample, whereas the
compound-exposed parasite remained at the trophozoite
stage and gradually developed to schizonts at h60 and h72
in K1 and 3D7 respectively. Furthermore, compound 15
may have affected hemozoin formation because abnormal
hemozoin crystals were found since h36 (Fig. 4, shown
with arrows).
Finally, to understand the importance of the isoquioline
ring, this ring was replaced with a naphthyl group. Com-
pounds 27, 28 and 29 were compared in activity to similar
compounds (22, 15 and 25). All modified derivatives
demonstrated obviously decrease antimalarial activity in
both strains. Furthermore, we attempted to mimic CQ
structure by replacing the isoquinoline ring with a 7-
chloroquinoline ring as presented in compound 30, 31 and
32. The results exhibited the effect of the 7-chloroquinoline
ring in decreasing their corresponding activity in both
strains.
Parasite growth and morphological change cells
following exposure to compounds
The results displayed the effects of parasite exposure to
compounds 6 and 15 for 60 h. Both compounds 6 and 15
could inhibit Plasmodium falciparum time-dependent
growth compared to the negative control. However, com-
pound 15 was less potent than CQ and DHA. Furthermore,
compound 6 and CQ completely inhibited growth of the
3D7 strain while DHA completely inhibited parasite growth
The morphology change of 3D7 and K1 P. falciparum
following exposure to selected compounds (compounds 6
and 15) at IC90 were determined. CQ and DHA were used
as a positive control and evaluated at IC100. The experi-
ment with no compound exposure was determined as a
negative control. The effect of compounds on parasite

Medicinal Chemistry Research
Fig. 4 Morphology of
Plasmodium falciparum after
compound exposure 6. Two
standard antimalarials,
chloroquine (CQ) and
dihydroartemisinin (DHA), were
used as the positive control. Red
arrows pointed to abnormal
hemozoin crystals found in K1
after 36 h exposure
100.00
80.00
60.00
40.00
20.00
0.00
in the K1 strain at 6 h of testing exposure (data not pre-
sented in chart) (Fig. 5).
Conclusion
H6
H12
H24
H36
H48
H60
In this study, the antimalarial activity of novel synthesized
isoquinoline derivatives was evaluated. Compound 6
exhibited antiplasmodial activity compared to reference
standard antimalarial drugs. Although compound 6 only
displayed moderate antimalarial activity, it was active
against both chloroquine sensitive and chloroquine resistant
strains. In addition, compound 15 showed interesting
selective activity against resistant strains over sensitive
strains. Overall this work provides knowledge for further
study about the discovery of antimalarial agents, which
selectively work on resistant strains.
3D7 or K1- CTRL
3D7-DHA, 50 nM
3D7-15, 652 uM
K1-CQ, 1000 nM
K1-6, 212 uM
100.00 100.00 100.00 100.00 100.00 100.00
0.00
100.00
0.00
14.67
66.45
0.00
0.00
17.68
0.00
0.00
17.95
0.00
0.00
19.75
2.64
0.00
22.61
2.40
0.00
0.00
0.00
0.00
0.15
0.00
K1-15, 163 uM
100.00 100.00 100.00
19.19
30.69
35.75
Fig. 5 Percentage growth of Plasmodium falciparum after exposure to
compound 6 at 212 µM concentration and 15 at 163 µM concentration
for 60 h. Two standard antimalarial drugs, chloroquine (CQ) at 100 µM
concentration and dihydroartemisinin (DHA) at 50 nM concentration
were used as a positive control. The vehicle was used as a negative
control (CTRL)

Medicinal Chemistry Research
Experimental section
Chemistry
H-5), 7.63 (m, 2H, H-6, H-7), 7.42 (t, J = 5.7 Hz, 1H, H-5′),
7.09–6.99 (m, 3H, H-2′, H-4′, H-6′), 3.85 (s, 3H, O-CH3);
13C-NMR (75 MHz, CDCl3, 25 °C) δ 159.7 (C-3′), 152.0 (C-
1), 142.7 (C-1′), 138.3 (C-3), 134.2 (C-6), 130.6 (C-4), 129.6
(C-5′), 128.4 (C-4a), 127.9 (C-8), 127.2 (C-7), 124.8 (C-8a),
122.6 (C-5), 115.8 (C-6′), 113.4 (C-4′), 108.4 (C-2′), 55.3
(O-CH3); HRMS (ESI) m/z calculated: C16H13NO: 235.0997
found: 236.1076 (M+H)+.
Unless otherwise noted, all reagents, chemicals and cata-
lysts were purchased from commercial sources (Sigma-
Aldrich; USA, Fluorochem; UK). Some solvents had to be
dried by distillation under nitrogen and used as freshly
prepared solvents. All other solvents were used as supplied
without further purification.
4-(4-(azepan-1-ylmethyl)phenyl)isoquinoline (12)
Instrumentation
To a solution of commercially available 4-bromoisoquinoline
(0.070 g, 0.34 mmol) and 4-(homopiperidine)methyl)phe-
nylboronic acid (0.127 g, 0.40 mmol) in 4 ml of DMF,
the tetrakis(triphenylphosphine) palladium (0.039 g,
0.017 mmol) and an aqueous solution of sodium carbonate
(0.11 g 1.01 mmol) were added and stirred. The reaction was
then warmed at 65 °C for overnight. The mixture was diluted
with water and extracted with ethyl acetate (×3). The com-
bined organic fractions were washed with brine (×1), dried
over sodium sulfate and concentrated under vacuum. The
crude product was purified by column chromatography by
using hexane/ ethyl acetate (7:3) as eluent, to give 0.083 g of
a yellow liquid (78%); Rf = 0.29 (Hexane/EtOAc 8:2); IR
1H NMR and 13C NMR spectra were recorded at 500 MHz
and 125 MHz, respectively (Bruker Biospin, USA). 1H
NMR chemical shifts are referenced to TMS or CDCl3 (0;
7.26 ppm). 13C NMR was referenced to CDCl3 (77.0 ppm).
Mass spectra and high-resolution mass spectra (HRMS)
were measured using the electrospray ionization (ESI)
technique (Bruker MicrOTOF, USA). Infrared spectra were
performed by Fourier Transformed infrared spectrometer
(Simadzu, Japan). Purification was carried out on column
chromatography using silica gel 60 (Merck, 230–400 mesh).
Chemical shifts are reported in part per million (ppm) and
spectral data are represented in the following order: chemical
shift; multiplicity (s = singlet, d = doublet, t = triplet, q =
quartet, dd = doublet of doublets, td = triplet of doublets,
m = multiplet); coupling constant (J, Hz); number of protons.
Column chromatography was performed on silica gel Merck
Kieselgel 70–230 mesh ASTM. Thin layer chromatography
(TLC) was carried out on 5 × 20 cm plates with a layer
thickness of 0.25 mm (Merck Silica gel 60 F254). Elemental
Analysis (C, H, N) of the target compounds are within 0.4%
of the calculated values unless otherwise noted.
1
(KBr) 3090, 2986, 2822, 1622, 1587, 1397, 1234 cm−1; H-
NMR (300 MHz, CDCl3, 25 °C) δ 9.26 (s, 1H, H-1), 8.51 (s,
1H, H-3), 8.01 (d, J = 7.9, 1H, H-8), 7.66 (m, 3H, H-5, H-6,
H-7), 7.50 (m, 4H, H-2′, H-3′), 3.77 (s, 2H, -Ph-CH2-N), 2.72
(m, 4H, H-2″), 1.68 (m, 8H, CH2-CH2); 13C-NMR (75 MHz,
CDCl3, 25 °C) δ 151.9 (C-1), 142.8 (C-4′), 134.2 (C-3),
132.2 (C-6), 132.0 (C-4), 130.9 (C-4a), 130.5 (C-3′, C-5′),
130.0 (C-7), 129.2 (C-2′, C-6′), 128.4 (C-8), 127.9 (C-8a),
127.2 (C-1′), 124.9 (C-5), 62.3 (CH2-N), 55.6 (C-1″), 27.8
(C-2″), 27.1 (C-3″); HRMS (ESI) m/z calculated: 316.1939
found: 317.2042 (M+H)+.
4-(3-methoxyphenyl)isoquinoline (6)
4-(1-(4-chlorophenyl)-1H-1,2,3-triazol-4-yl)isoquinoline (15)
To a solution of commercially available 4-bromoisoquinoline
(0.080 g, 0.38 mmol) and 3-methoxyphenylboronic acid
(0.087 g, 0.57 mmol) in 4 ml of DMF, the tetrakis(triphe-
nylphosphine) palladium (0.022 g, 0.019 mmol) and an aqu-
eous solution of sodium carbonate (0.12 g 1.14 mmol) were
added and stirred. The reaction was then warmed at 65 °C for
overnight. The mixture was diluted with water and extracted
with ethyl acetate (×3). The combined organic fractions were
washed with brine (×1), dried over sodium sulfate and con-
centrated under vacuum. The crude product was purified by
column chromatography by using hexane/ethyl acetate
(8:2) as eluent, to give 0.065 g of a colorless liquid (73%);
Rf = 0.59 (Hexane/EtOAc 6:4); yield 73%; IR (KBr) 3053,
2937, 1602, 1484, 1226, 1160, 886 cm−1; 1H-NMR
(300 MHz, CDCl3, 25 °C) δ 9.23 (s, 1H, H-1), 8.48 (s, 1H,
H-3), 8.02 (d, J = 5.7 Hz, 1H, H-8), 7.93 (d, J = 6.0 Hz, 1H,
To a solution of 4-ethynylisoquinoline (0.05 g, 0.33 mmol)
and 1-azido-4-chlorobenzene (0.05 g, 0.33 mmol) in a
mixture of tert-butanol (1:1, 0.5 mL). Sodium ascorbate
(6.5 mg, 0.033 mmol) and copper sulfate pentahydrate
(0.8 mg, 0.0033 mmol) were added. The solution was stirred
at room temperature for 48 h. The mixture was diluted with
water and extracted with ethyl acetate (×3). The combined
organic fractions were washed with brine (×1), dried over
sodium sulfate and concentrated under vacuum. The crude
product was purified by column chromatography by using
hexane/ ethyl acetate (8:2) as eluent, to give 0.042 g of a
slightly yellow solid (42%); Rf = 0.15 (Hexane/EtOAc
7:3); m.p. 240–243 °C; IR (KBr) 3082, 3044, 1503, 1383,
1092, 990, 777 cm−1; 1H-NMR (300 MHz, CDCl3, 25 °C) δ
9.32 (s, 1H, H-1), 8.80 (s, 1H, H-3), 8.62 (d, J = 8.5, 1H,

Medicinal Chemistry Research
H-8), 8.33 (S, 1H, H-5’), 8.09 (d, J = 8.1, 1H, H-5),
7.86–7.83 (m, 3H, H-6, H-2″, H-6″), 7.72 (m, 1H, H-7),
7.60 (m, 2H, H-3″, H-5″); 13C-NMR (75 MHz, CDCl3,
25 °C) δ 153.4 (C-1), 143.0 (C-1′), 135.4 (C-3), 134.9 (C-
1″), 133.7 (C- 4″), 133.6 (C- 5′), 131.4 (C-4), 130.1 (C-6),
128.5 (C-4a), 128.1 (C-3″, C-5″), 127.7 (C-8), 124.4 (C-7),
121.8 (C-8a), 121.4 (C-2″, C-6″), 120.5 (C-5); HRMS (ESI)
m/z calculated: 306.0672 found: 307.0750 (M+H)+.
8.40 (s, 1H, H-5′), 8.24 (m, 1H, H-6″), 8.20 (m, 1H, H-4″),
8.11 (d, J = 8.1 Hz, 1H, H-5), 7.81–7.88 (m, 2H, H-6, H-
2″), 7.79 (d, J = 7.8 Hz, 1H, H-5″) 7.74 (m, 1H, H-7); 13C-
NMR (75 MHz, CDCl3, 25 °C) δ 153.5 (C-1), 145.8 (C-1′),
142.9 (C-3), 137.4 (C-6″), 133.2 (C-2″), 132.4 (C-4″),
131.6 (C-5′), 131.1 (C-6), 128.6 (C-4), 128.2 (C-5″), 127.8
(C-4a), 124.8 (C-1″), 124.6 (C-8), 123.7 (C-7), 121.3 (C-
8a), 120.3 (C-5), 117.3 (C≡N), 114.3 (C-3″); MS (ESI) m/z
calculated: 297.1014 found: 298.1089 (M+H)+.
4-(1-(3-chlorophenyl)-1H-1,2,3-triazol-4-yl)isoquinoline (16)
4-(1-benzhydryl-1H-1,2,3-triazol-4-yl)isoquinoline (25)
To a solution of 4-ethynylisoquinoline (0.05 g, 0.33 mmol)
and 1-azido-3-chlorobenzene (0.05 g, 0.33 mmol) in a
mixture of tert-butanol (1:1, 0.5 mL). Sodium ascorbate
(6.5 mg, 0.033 mmol) and copper sulfate pentahydrate
(0.8 mg, 0.0033 mmol) were added. The solution was stirred
at room temperature for overnight. The mixture was diluted
with water and extracted with ethyl acetate (×3). The
combined organic fractions were washed with brine (×1),
dried over sodium sulfate and concentrated under vacuum.
The crude product was purified by column chromatography
by using hexane/ ethyl acetate (8:2) as eluent, to give
0.073 g of a yellow solid (73%); Rf = 0.09 (Hexane/EtOAc
8:2); m.p. 236–240 °C; IR (KBr) 3119, 1624, 1599, 1499,
1057, 798 cm−1; 1H-NMR (300 MHz, CDCl3, 25 °C) δ 9.32
(s, 1H, H, H-1), 8.81 (s, 1H, H-3), 8.61 (d, J = 8.6 Hz, 1H,
H-8), 8.35 (s, 1H, H-5′), 8.10 (d, J = 8.1, 1H, H-5), 7.94 (m,
1H, H-2″), 7.86–7.79 (m, 2H, H-6, H-6″), 7.72 (m, 1H,H-
7), 7.58 (t, J = 7.6, 1H, H-5″), 7.52 (m, 1H, H-4″); 13C-
NMR (75 MHz, CDCl3, 25 °C) δ 153.4 (C-1), 145.4 (C-1′),
143.0 (C-3), 137.7 (C-1″), 135.8 (C-3″), 133.6 (C-5′), 131.4
(C-4), 131.0 (C-5″), 129.1 (C-6), 128.6 (C-4a), 128.1
(C-4″), 127.7 (C-8), 124.8 (C-7), 121.3 (C-6″), 120.9 (C-
2″), 120.5 (C-8a), 118.6 (C-5); HRMS (ESI) m/z calculated:
306.0672 found:307.0749 (M+H)+.
To a solution of 4-ethynylisoquinoline (0.05 g, 0.33 mmol)
and (azidomethylene)dibenzene (0.068 g, 0.33 mmol) in a
mixture of tert-butanol (1:1, 0.5 mL). Sodium ascorbate
(6.5 mg, 0.033 mmol) and copper sulfate pentahydrate
(0.8 mg, 0.0033 mmol) were added. The solution was stirred
at room temperature for overnight. The mixture was diluted
with water and extracted with ethyl acetate (×3). The
combined organic fractions were washed with brine (×1),
dried over sodium sulfate and concentrated under vacuum.
The crude product was purified by column chromatography
by using hexane/ethyl acetate (7:3) as eluent, to give
0.046 g of a white solid (39%); Rf = 0.20 (Hexane/EtOAc
7:3); m.p. 139–143 °C; IR (KBr) 3088, 3030, 1495, 1227,
1057, 901, 750 cm−1; 1H-NMR (300 MHz, CDCl3, 25 °C) δ
9.25 (S, 1H, H-1), 8.68 (s, 1H, H-3), 8.62 (d, J = 8.5 Hz,
1H, H8), 8.04 (d, J = 8.0 Hz, 1H, H-5), 7.84–7.74 (m, 2H,
H-6, H-5′), 7.68 (dd, J = 8.0, 1.1 Hz, 1H, H-7), 7.57–7.32
(m, 6H, H-3″, H-4″, H-5″), 7.57–7.32 (m, 5H, H-2″, H-6″,
CH); 13C-NMR (75 MHz, CDCl3, 25 °C) δ 152.4 (C-1),
144.1 (C-1′), 141.9 (C-3), 138.0 (C-1″), 133.7 (C-6), 131.6
(C-5′), 129.1 (2, C-3″, C4a), 128.8 (C-4a), 128.1 (2, C-8, C-
2″), 127.8 (C-7), 125.2 (C-4″), 122.7 (C-8a), 122.2 (C-5),
68.4 (CH); MS (ESI) m/z calculated: 362.1531 found:
363.1609 (M+H)+.
3-(4-(isoquinolin-4-yl)-1H-1,2,3-triazol-1-yl)benzonitrile (19)
1-(4-(tert-butyl)benzyl)-4-(naphthalen-1-yl)-1H-1,2,
3-triazole (27)
To a solution of 4-ethynylisoquinoline (0.05 g, 0.33 mmol)
and 3-azidobenzonitrile (0.047 g, 0.33 mmol) in a mixture
of tert-butanol (1:1, 0.5 mL). Sodium ascorbate (6.5 mg,
0.033 mmol) and copper sulfate pentahydrate (0.8 mg,
0.0033 mmol) were added. The solution was stirred at room
temperature for overnight. The mixture was diluted with
water and extracted with ethyl acetate (×3). The combined
organic fractions were washed with brine (×1), dried over
sodium sulfate and concentrated under vacuum. The crude
product was purified by column chromatography by using
hexane/ethyl acetate (8:2) as eluent, to give 0.085 g of a
white solid (88%); Rf = 0.17 (Hexane/EtOAc 7:3); m.p.
247–249 °C; IR (KBr) 3057, 2228, 1595, 1420, 1045,
To a solution of 4-ethynylisoquinoline (0.02 g, 0.131 mmol)
and
1-(azidomethyl)-4-(tert-butyl)benzene
(0.025 g,
0.131 mmol) in a mixture of tert-butanol (1:1, 0.5 mL).
Sodium ascorbate (2.6 mg, 0.013 mmol) and copper sulfate
pentahydrate (0.3 mg, 0.0013 mmol) were added. The
solution was stirred at room temperature for overnight. The
mixture was diluted with water and extracted with ethyl
acetate (×3). The combined organic fractions were washed
with brine (×1), dried over sodium sulfate and concentrated
under vacuum. The crude product was purified by column
chromatography by using hexane/ethyl acetate (8:2) as
eluent, to give 0.015 g of an amorphous white solid (33%);
Rf = 0.33 (Hexane/EtOAc 8:2); m.p. 158–160 °C; IR (KBr)
1
798 cm−1; H-NMR (300 MHz, CDCl3, 25 °C) δ 9.34 (s,
1H, H-1), 8.82 (s, 1H, H-3), 8.59 (d, J = 8.5 Hz, 1H, H-8)

Medicinal Chemistry Research
3119, 2963, 1458, 1360, 1221, 1053, 980, cm−1; H-NMR
(300 MHz, CDCl3, 25 °C) δ 8.36 (m, 1H, H-5), 7.85 (m,
2H, H-1, 8), 7.71–7.66 (m, 2H, H-5’, H-3), 7.54–7.47 (m,
3H, H-2, 6, 7), 7.41 (d, J = 8.0 Hz, 2H, H-3″, H-5″), 7.29
(d, J = 8.0 Hz, 2H, H-2″, H-6″), 5.58 (s, 2H, CH2), 1.30 (s,
9H, CH3); 13C-NMR (75 MHz, CDCl3, 25 °C) δ 152.0 (C-
4″), 147.3 (C-1′), 133.9 (C-4), 131.6 (C-8a), 131.1 (C-1″),
128.9 (C-5a), 128.4 (C-5′), 128.0 (C-8), 127.2 (C-5), 126.6
(C-1), 126.1 (C-7), 126.0 (C-2″), 125.5 (C-6), 125.3 (C-2),
124.4 (C-3″), 122.4 (C-3), 54.0 (CH2), 34.7 (CH), 31.3
(CH3); MS (ESI) m/z calculated: 341.1892 found:364.1785
(M+Na)+.
3057, 1599, 1498, 1450, 1051 cm−1; H-NMR (300 MHz,
1
1
CDCl3, 25 °C) δ 8.37 (m, 1H, H-5), 7.86 (m, 2H, H-1, H-8),
7.68 (m, 2H, H-2, H-3), 7.48 (m, 3H, H-1′, H-6, H-7), 7.39
(m, 6H, H-3″, H-4″, H-5″), 7.21 (m, 5H, H-2″, CH); 13C-
NMR (75 MHz, CDCl3, 25 °C) δ 146.7 (C-1′), 138.2 (C-4),
133.9 (C-1′), 130.1 (C-8a), 129.0 (C-3″), 128.9 (C-5′),
128.7 (C-4a), 128.5 (C-5), 128.2 (C-2′), 127.8 (C-8), 127.3
(C-1), 126.7 (C-6), 126.0 (C-7), 125.5 (C-4′), 125.3 (C2),
122.3 (C-3), 68.3 (CH); MS (ESI) m/z calculated: 361.1579
found: 384.1488 (M+Na)+.
4-(1-(4-(tert-butyl)benzyl)-1H-1,2,3-triazol-4-yl)-7-
chloroquinoline (30)
1-(4-chlorophenyl)-4-(naphthalen-1-yl)-1H-1,2,3-triazole (28)
To a solution of 7-chloro-4-ethynylquinoline (0.08 g,
0.49 mmol) and 1-(azidomethyl)-4-(tert-butyl)benzene
(0.092 g, 0.49 mmol) in a mixture of tert-butanol (1:1,
0.5 mL). Sodium ascorbate (9.7 mg, 0.049 mmol) and cop-
per sulfate pentahydrate (1.2 mg, 0.0049 mmol) were added.
The solution was stirred at room temperature for overnight.
The mixture was diluted with water and extracted with ethyl
acetate (×3). The combined organic fractions were washed
with brine (×1), dried over sodium sulfate and concentrated
under vacuum. The crude product was purified by column
chromatography by using hexane/ ethyl acetate (9:1) as
eluent, to give 0.068 g of a colorless liquid; (39%); Rf =
0.17 (Hexane/EtOAc 9:1); IR (KBr) 3121, 2965, 1605,
To a solution of 1-ethynylnaphthalene (0.02 g, 0.128 mmol)
and 1-azido-4-chlorobenzene (0.02 g, 0.128 mmol) in a
mixture of tert-butanol (1:1, 0.5 mL). Sodium ascorbate
(2.5 mg, 0.013 mmol) and copper sulfate pentahydrate
(0.3 mg, 0.0013 mmol) were added. The solution was stirred
at room temperature for overnight. The mixture was diluted
with water and extracted with ethyl acetate (×3). The
combined organic fractions were washed with brine (×1),
dried over sodium sulfate and concentrated under vacuum.
The crude product was purified by column chromatography
by using hexane/ethyl acetate (8:2) as eluent, to give
0.032 g of a yellow solid (83%); Rf = 0.40 (Hexane/EtOAc
8:2); m.p. 149–151 °C; IR (KBr) 3125, 3063, 1593, 1500,
1233, 1049, 829 cm−1; 1H-NMR (300 MHz, CDCl3, 25 °C)
δ 8.40 (m, 1H, H-5), 8.21 (s, 1H, H-5′), 7.90 (m, 2H, H-1,
H-8), 7.79 (m, 3H, H-3, H-2”), 7.55 (m, 4H, H-6, H-7, H-
3″, H-5″); 13C-NMR (75 MHz, CDCl3, 25 °C) δ 147.8 (C-
1′), 135.6 (C-4), 134.6 (C-1″), 133.9 (C-4″), 131.1 (C-8a),
130.0 (C-4a), 129.3 (C-5’), 128.6 (C-3″), 127.4 (C-5), 126.9
(C-1), 126.2 (C-8), 125.4 (C-7), 125.3 (C-6), 123.8 (C-2),
121.7 (C-3), 120.4 (C-2″); MS (ESI) m/z calculated:
305.0720 found: 328.0626 (M+Na)+.
1458, 1219, 1047, 991, 723 cm−1 1H-NMR (300 MHz,
;
CDCl3, 25 °C) δ 8.79 (d, J = 4.4 Hz, 1H, H-2), 8.55 (d, J =
9.1 Hz, 1H, H-5), 8.05 (s, 1H, H-8), 7.90 (s, 1H, H-5′), 7.46
(d, J = 4.4 Hz, 1H, H-6), 7.44–7.33 (m, 3H, H-3, H-3″, H-
5″), 7.25 (d, J = 8.1 Hz, 2H, H-2″, H-6″), 5.57 (s, 2H, CH2),
1.26 (s, 9H, 3xCH3); 13C-NMR (75 MHz, CDCl3, 25 °C) δ
152.3 (C-2), 150.8 (C-4″), 149.1 (C-8a), 144.9 (C-1′), 136.3
(C-4), 135.6 (C-7), 131.1 (C-1″), 128.5 (C-5’), 128.1 (2) (C-
5, C-8), 127.5 (C-6), 126.3 (C-2″), 123.9 (C-3″), 123.5 (C-
4a), 120.3 (C-3), 54.2 (CH2), 34.7 (CH), 31.2 (CH3); MS
(ESI) m/z calculated: 376.1455 found: 377.1537 (M+H)+.
1-benzhydryl-4-(naphthalen-1-yl)-1H-1,2,3-triazole (29)
7-chloro-4-(1-(4-chlorophenyl)-1H-1,2,3-triazol-4-yl)
quinoline (31)
To
a
solution of 1-ethynylnaphthalene (0.024 g,
0.154 mmol) and (azidomethylene)dibenzene (0.032 g,
0.154 mmol) in a mixture of tert-butanol (1:1, 0.5 mL).
Sodium ascorbate (3.1 mg, 0.015 mmol) and copper sulfate
pentahydrate (0.4 mg, 0.0015 mmol) were added. The
solution was stirred at room temperature for overnight. The
mixture was diluted with water and extracted with ethyl
acetate (×3). The combined organic fractions were washed
with brine (×1), dried over sodium sulfate and concentrated
under vacuum. The crude product was purified by column
chromatography by using hexane/ethyl acetate (9:1) as
eluent, to give 0.017 g of a white solid (30%); Rf = 0.39
(Hexane/EtOAc 8:2); m.p. 161–163 °C; IR (KBr) 3088,
To a solution of 7-chloro-4-ethynylquinoline (0.08 g,
0.49 mmol) and 1-azido-4-chlorobenzene (0.075 g,
0.49 mmol) in a mixture of tert-butanol (1:1, 0.5 mL).
Sodium ascorbate (9.7 mg, 0.049 mmol) and copper sulfate
pentahydrate (1.2 mg, 0.0049 mmol) were added. The
solution was stirred at room temperature for overnight. The
mixture was diluted with water and extracted with ethyl
acetate (×3). The combined organic fractions were washed
with brine (×1), dried over sodium sulfate and concentrated
under vacuum. The crude product was purified by column
chromatography by using hexane/ ethyl acetate (9:1) as

Medicinal Chemistry Research
eluent, to give 0.066 g of an orange solid (39%); Rf = 0.20
(Hexane/EtOAc 7:3); m.p. 161–164 °C;IR (KBr) 3125,
90% N2 atmosphere. The parasite was maintained at 5%
parasitemia and subcultured every 48 h. Antimalarial
activity of the synthetic compounds was assessed using
SYBR Green I assay [28, 29]. The parasites were syncho-
nized by 5% sorbitol to obtain the ring stage of P. falci-
parum. The highly synchronous ring stage parasite culture
was prepared to a parasitemia of 2% and hematocrit of 1%.
The tested compound was dissolved in DMSO (vehicle) and
diluted for 50 times with the complete media to obtain the
first working concentration containing 2% DMSO. The
50 μL of parasite mixture was inoculated into the 50 μL pre-
dosed compounds in 96-well plates. The 100 µL final
volume contained a parasite with a parasitemia of 1% and
hematocrit of 0.5%, and maximum DMSO (vehicle) at 1%
for the highest compound concentration. The parasite
mixture was exposed to the following eight levels of con-
centration as testing compounds: 3.125, 6.25, 12.5, 25, 50,
100, 200 and 400 μM (at 37 °C in a 5% CO2, 5% O2, and
90% N2 atmosphere) for 48 h. The parasite DNA content
was stained by SYBR green I diluted in lysis buffer, and
fluorescence intensity was measured using a microplate
reader at excitation and emission wavelengths of 485 nm
and 530 nm, respectively. The experiment was repeated
three times and triplicated for each experiment. The IC50
values (concentrations that inhibit the parasite growth by
50%) were calculated from a log-dose–response curve
plotted using the CalcusynTM version 1.1 (BioSoft, Cam-
bridge, UK) and used as indicators of antimalarial activity.
Four standard antimalarial drugs (chloroquine, CQ; quinine,
Q; mefloquine, MQ and dihydroartemisinin, DHA) were
used as a positive control.
1
2957, 1593, 1499, 1391, 1230, 1047, 991, 827 cm−1; H-
NMR (300 MHz, DMSO-d6, 25 °C) δ 9.61 (s, 1H, H-5″),
9.06 (d, J = 4.4 Hz, 1H, H-2), 8.90 (d, J = 9.1 Hz, 1H, H-5),
8.18 (s, 1H, H-8), 8.09 (d, J = 8.7 Hz, 2H, H-2″, H-6″), 7.94
(d, J = 4.4 Hz, 1H, H-6), 7.74 (m, 3H, H-3, H-3″, H-5″);
13C-NMR (75 MHz, CDCl3, DMSO-d6, 25 °C) δ 150.7 (C-
2), 150.0 (C-1′), 149.9 (C-8a), 139.4 (C-4), 136.0 (C-7),
135.5 (C-1″), 134.7 (C-4″), 129.9 (C-5′), 128.9 (C-5), 128.3
(C-8), 127.8 (C-3″), 125.3 (C-6), 122.6 (C-2″), 121.7 (C-
4a), 120.3 (C-3); MS (ESI) m/z calculated: 340.0283 found:
341.0363 (M+H)+.
4-(1-benzhydryl-1H-1,2,3-triazol-4-yl)-7-chloroquinoline (32)
To
a solution of 7-chloro-4-ethynylquinoline (0.08 g,
0.49 mmol) and (azidomethylene)dibenzene (0.102 g,
0.49 mmol) in a mixture of tert-butanol (1:1, 0.5 mL). Sodium
ascorbate (9.7 mg, 0.049 mmol) and copper sulfate pentahy-
drate (1.2 mg, 0.0049 mmol) were added. The solution was
stirred at room temperature for overnight. The mixture was
diluted with water and extracted with ethyl acetate (×3). The
combined organic fractions were washed with brine (×1),
dried over sodium sulfate and concentrated under vacuum.
The crude product was purified by column chromatography
by using hexane/ ethyl acetate (9:1) as eluent, to give 0.062 g
of an yellow solid (32%); Rf = 0.20 (Hexane/EtOAc 7:3); m.
p. 168–170 °C; IR (KBr) 3092, 3045, 2955, 1587, 1493,
1
1049, 878 cm−1; H-NMR (300 MHz, CDCl3, 25 °C) δ 8.88
(d, J = 4.3 Hz, 1H, H-2), 8.65 (d, J = 9.1 Hz, 1H, H-5), 8.13
(s, 1H, H-8), 7.83 (s, 1H, H-1′), 7.53 (m, 2H, H-3, H-6), 7.39
(m, 6H, 2 H-3″, 2 H-4″, 2 H-5″), 7.20 (m, 5H, CH, 2 H-2″, 2
H-6″); 13C-NMR (75 MHz, CDCl3, 25 °C) δ 149.9 (C-2),
148.2 (C-8a), 144.4 (C-1′), 137.6 (C-4), 137.2 (C-1″), 136.4
(C-7), 129.2 (C5′), 129.0 (C-3″), 128.7 (C-5), 128.1 (C-8),
127.9 (C-6), 127.8 (C-2″), 124.0 (C-4″), 123.8 (C-4a), 120.2
(C-3), 68.6 (CH); MS (ESI) m/z calculated: 396.1142 found:
397.1230 (M+H)+.
Observation of morphological change in cells following
exposure to the selected synthetic compounds
Synchronized P. falciparum 3D7 and K1 strains were used in
the experiment. The parasites were exposed to the separated
selected synthetic compounds and two standard antimalarial
drugs, chloroquine and DHA, which were used as a positive
control at IC100. The treated parasites were incubated in a 5%
CO2, 5% O2, and 90% N2 atmosphere at 37 °C for 72 h.
Blood films were prepared at the following time points: 6, 12,
24, 36, 48, 60, 72 h and stained with Giemsa (Biotechnical
Thai, Bangkok, Thailand). The morphology of the resulting
parasites was observed and captured under a light microscope
(×100, Leica Microsystems, Hesse, Germany) [30].
Biological assay
Assessment of in vitro antimalarial activity of synthetic
compounds
All synthesized compounds were evaluated for their in vitro
antiplasmodial activity against the P. falciparum
chloroquine-sensitive (3D7) and chloroquine-resistant (K1)
laboratory strains. The parasite was briefly cultured
according to the method laid out in [27], in O+ human
erythrocytes suspended in an RPMI culture medium sup-
plemented with 10% human B serum, 25 mM HEPES and
15 mg/ml gentamicin at 37 °C in a 5% CO2, 5% O2, and
Acknowledgements The authors would like to acknowledge the par-
tial financial support provided by Thammasat University Research
Fund under TU Research Scholar, Contract number 2/32.2557 and the
authors gratefully acknowledge Thammasat University Research Unit
in Drug, Health Product Development and Application (Project ID.
6305001), Thammasat University. In addition, we really appreciate
Prof Dr OV at Faculty of pharmacy, Mahidol university, Thailand for

Medicinal Chemistry Research
supporting some chemicals and thank to Prof Dr Kesara Na-
Bangchang at Center of Excellence in Pharmacology and Molecular
Biology of Malaria and Cholangiocarcinoma of Thammasat University
for providing suggestions and some biological equipment. In addition,
we would like to thank Drug Discovery and Development center for
some facilities for biological tests.
12. Cui L, Mharkurwa S, Ndiaye D. Antimalarial drug resistance:
literature review and activities and findings of the ICEMR net-
work. Am J Trop Med Hyg. 2015;93:57–68.
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drug discovery. Bioorg Med Chem Lett. 2013;23:2829–43.
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Author contributions ST participated in design of study, provided all
synthesized compounds and elucidated structure of all compounds and
interpret data, PM carried out the main biological experiment, parti-
cipated in image and performed the statistical analysis, AT assisted in
some biological study, OV helped to interpret the data and gave
comment in experiment and paper. ST and PM helped to draft the
paper. ST read and approved the final paper.
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Compliance with ethical standards
19. Slater AFG. Chloroquine: mechanism of drug action and resis-
tance in Plasmodium falciparum. Pharmac Ther. 1993;57:203–35.
20. Homewood CA, Warhurst DC, Peters W, Baggaley VC. Lyso-
somes, pH and the anti-malarial action of chloroquine. Nature.
1972;235:50–2.
21. Yayon A, Cabantchik ZI, Ginsburg H. Identification of the acidic
compartment of Plasmodium falciparum-infected human ery-
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EMBO J. 1984;3:2695–700.
Conflict of interest The authors declare that they have no conflict of
interest.
Publisher’s note Springer Nature remains neutral with regard to
jurisdictional claims in published maps and institutional affiliations.
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