Synthesis, molecular docking and antimalarial activity of phenylalanine-glycine dipeptide bearing sulphonamide moiety

Article abstract of DOI:10.1016/j.molstruc.2021.131201

Ten novel phenylalanine-glycine dipeptide sulphonamide conjugate were synthesized and characterized using ¹HNMR, ¹³CNMR, FTIR and HRMS spectroscopic techniques. The in silico studies predicted better interactions of compounds with target protein residues and a higher dock score in comparison with standard drugs. The in vivo antimalarial study, hematological study, liver and kidney function test were evaluated on the synthesized compounds. Compounds 7h, 7i and 7j inhibited the parasite by 34.5–60.2% on day 4 of after-treatment exposure. Compound 7j inhibited the multiplication of the parasite by 60.2% on day 4 of after-treatment which was comparable with that of the standard drug with 68.8% inhibition at same day of after-treatment exposure.

Full text of DOI:10.1016/j.molstruc.2021.131201

Journal of Molecular Structure 1246 (2021) 131201
Contents lists available at ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstr
Synthesis, molecular docking and antimalarial activity of
phenylalanine-glycine dipeptide bearing sulphonamide moiety
a,b,∗
b
c
b
Babatunde.S. Aronimo
, Uchechukwu.C. Okoro , Rafat. Ali , Collins.U. Ibeji ,
b
b
James.A. Ezugwu , David.I. Ugwu
a
Department of Chemistry Kogi State College of Education (Technical), Kabba, Kogi State, Nigeria
b
Department of Pure and Industrial Chemistry, University of Nigeria, Nsukka 410001, Enugu State, Nigeria
c
Department of Chemistry, Indian Institute of Technology, Kanpur 208016, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Ten novel phenylalanine-glycine dipeptide sulphonamide conjugate were synthesized and characterized
using 1HNMR, 13 CNMR, FTIR and HRMS spectroscopic techniques. The in silico studies predicted better
interactions of compounds with target protein residues and a higher dock score in comparison with stan-
dard drugs. The in vivo antimalarial study, hematological study, liver and kidney function test were eval-
uated on the synthesized compounds. Compounds 7h, 7i and 7j inhibited the parasite by 34.5–60.2% on
day 4 of after-treatment exposure. Compound 7j inhibited the multiplication of the parasite by 60.2% on
day 4 of after-treatment which was comparable with that of the standard drug with 68.8% inhibition at
same day of after-treatment exposure.
Received 26 April 2021
Revised 17 July 2021
Accepted 26 July 2021
Available online 2 August 2021
Keywords:
Synthesis
Molecular docking
Antimalarial
©
2021 Elsevier B.V. All rights reserved.
Phenylalanine
Glycine
Dipeptide Sulphonamide
1
.0. Introduction
present artemisinin-based combination therapy is the most effec-
tive drug in the treatment of malaria, however, artemisinin resis-
tance has been observed in some countries [4]. This call for the de-
velopment of new antimalarial drug. The challenge now is to iden-
tify new classes of drugs like peptide based drugs to combat the
disease and imped drug resistance [5].
Malaria is a common and life-threatening disease which is
caused by parasites that are transmitted to people through the bite
of infected female Anopheles mosquitoes [1]. Malaria is caused by
the protozoan parasite plasmodium. Four species of the genus plas-
modium cause all malarial infections in human being. In addition,
one species, Plasmodium knowlesi that naturally affects animal has
recently been recognized to be the cause of zoonotic malaria in hu-
mans [2]. The WHO’s malaria report 2018, revealed that 228 mil-
lion people had malaria cases with an estimated 405,000 deaths.
The African region was home to 93% of malaria cases and 94% of
malaria deaths [1]. Children under the age of five years are the
most vulnerable group affected by malaria and they accounted for
Peptides have been reported as good chemotherapeutic agent
and can also serve as excipient in drug delivery system to over-
come tissue and cellular membrane barriers [6]. It has been at-
tracting considerable attention because of their potential utility in
pharmaceutical [7]. Peptides have some advantages as a drug can-
didate over alternative molecules because of its low toxicity, high
affinity, strong specificity and adequate tissue penetration [7].
Sulphonamides are class of medicinally important molecules
that possesses varying pharmacological activities like antimicro-
bial, anticancer, anti-inflammatory, antioxidant, diuretic, carbonic
anhydrase inhibition [8-14] among many others. They are used in
drug design because of their low cost, and toxicity, chemical and
metabolic stability, enhanced crystallinity, carboxyl bio-isosterism
and high level of biological activity [15].
67% of all the malaria deaths worldwide in 2018 [1].
In recent past, there has been emergence of resistance towards
many of the anti-malarial drugs like chloroquine, sulfadoxine-
pyrimethamine, quinine, mefloquine and piperaquine [3]. At
∗
Dipeptide-sulphonamide hybrids have been reported as a good
human carbonic anhydrase inhibitor [16], antimalarial [17] and an-
tioxidant agents [18].
Corresponding author at: Department of Chemistry Kogi State College of Educa-
tion (Technical), Kabba, Kogi State, Nigeria.
E-mail
addresses:
aronimot2@gmail.com
(Babatunde.S.
Aronimo),
uchechukwu.okoro@unn.edu.ng (Uchechukwu.C. Okoro), rafatali@iitk.ac.in (Rafat.
Ali), ugochukwu.ibeji@unn.edu.ng (Collins.U. Ibeji), james.ezugwu@unn.edu
Computational techniques have been used in drug discovery so
that high cost and time required for wet experiment will be re-
(James.A. Ezugwu), izuchukwu.ugwu@unn.edu.ng (David.I. Ugwu).
https://doi.org/10.1016/j.molstruc.2021.131201
022-2860/© 2021 Elsevier B.V. All rights reserved.
0

Babatunde.S. Aronimo, Uchechukwu.C. Okoro, Rafat. Ali et al.
Journal of Molecular Structure 1246 (2021) 131201
duced. The increase in agreement between computational and wet
experiments demonstrates the former as a good alternative to the
later. Although, no drug has stirred from computer to market, com-
putational method has been used in the development of FDA ap-
proved drugs. Therefore, both methods complement each other in
drug development [19-20].
2-(4-methylphenylsulfonamido)-N-(2-oxo-2-(phenylamino)ethyl)-3-
phenylpropanamide (7a)
O
−1
Yield 66.5%, Mp=175–176 C. FTIR (KBr, cm ): 3350 (N–H),
3062 (C-H aromatic), 2925 (C–H aliphatic), 1664, 1600 (2C=O),
1537, 1498, 1445 (C=C), 1321, 1158 (2S=O), 1091, 1030 (C–N),
1
1224 (SO N). H-NMR (400 MHz, CDCl ) δ 8.60 (s, 1H, NH), 7.65
2
3
Based on the numerous applications of sulphonamide, reported
side effects of current antimalarial drug and emerging challenges
associated with multidrug resistance malaria parasites, there is
need for the synthesis of novel compounds as potential anti-
malarial agents with less side effect and improved selectivity [16].
In view of these challenges, we designed the synthesis of novel
sulphonamide-dipeptide moieties with possible and improved an-
timalarial potential. We report herein the synthesis and antimalar-
ial properties of ten novel phenylalanine derived sulfonamides
conjugates. The design of this work was based on the reported
antimalarial activity of benzenesulphonamides containing dipep-
tide of alg-gly [17], valine-dipeptide [21] and val-val dipeptides
sulphonamide conjugates [18] and the need to develop newer
chemotherapeutic agents that will overcome the reported emerg-
ing resistance against artemisinin based therapy.
(d, J = 7.3 Hz, 2H, ArH), 7.41 (d, J = 7.9 Hz, 3H, ArH), 7.30 (t,
J = 7.9 Hz, 2H, ArH), 7.09–7.21 (m, 6H, ArH), 6.86 (d, J = 7.3
Hz, 2H), 5.17 (s, 1H, NH), 4.39 (q, J = 8.1 Hz, 1H, CH), 3.82
(dt, J = 17.1, 4.9 Hz, 2H, CH ), 3.16 (dd, J = 14.3, 4.6 Hz, 1H,
2
CHa of CH ), 2.71 (dd, J = 14.3, 10.1 Hz, 1H, CHb of CH ), 2.40
2
2
13
(d, J = 12.8 Hz, 3H, CH ). C-NMR (400 MHz, CDCl ) δ 171.58,
3
3
167.45 (C=O), 144.28, 137.78, 135.02, 134.28, 129.91, 129.00, 128.98,
128.88, 127.35, 127.25, 124.56, 120.29 (twelve aromatic carbons),
58.68, 44.07, 38.11, 21.66 (four aliphatic carbons). HRMS-ESI (m/z):
+
calcd. for C24H25N NaO S [M+Na] 474.15; found 474.15.
3
4
N-(2-(4-fluorophenylamino)-2-oxoethyl)-2-(4-
methylphenylsulfonamido)-3-phenylpropanamide (7b)
Yield 67.4%, Mp=76–77 °C. FTIR (KBr cm 1), 3351 (N–H), 3065
−
(C-H aromatic), 2926 (C-H aliphatic), 1662, (C=O), 1535, 1509,
1
455, 1410 (C=C), 1324, 1158 (2S=O), 1030, 1019 (C–N), 1215
1
(
SO N), 1092 (C-F). H-NMR (400 MHz, CDCl ) δ 8.70 (s, 1H, NH),
2 3
2.0. Material and methods
7
.59-7.63 (m, 2H, ArH), 7.54-7.58 (m, 1H, NH), 7.38 (d, J = 8.5
Hz, 2H, ArH), 7.07-7.19 (m, 5H, ArH), 6.97 (t, J = 8.9 Hz, 2H,
ArH), 6.86 (d, J = 7.3 Hz, 2H, ArH), 5.30–5.42 (m, 1H, NH),
All chemicals reagents and solvents used were obtained from
Aldrich, Merch, Fluka, Avra, SD Fine and Alfa Aesar and used with-
out purification. 1H-NMR and 13C-NMR spectra were recorded on
Jeol 400MHz spectrometers in CDCl3 and DMSO-d6 using TMS as
internal standard. FT-IR Spectroscopy of the compounds were run
in PerkinElmer Spectrum version 10.03.06 and the bands presented
in wavenumber. The mass spectroscopy was carried out using mi-
cro TOF electrospray time of flight mass spectrometer (Aerodyn Re-
seach Inc USA). Melting points were determined in open capillary
tubes, using a Rolex melting-point apparatus and are uncorrected.
All experiments were carried out at Prof Sandeep Verma Labora-
tory, Department of Chemistry, Indian Institute of Technology, Kan-
pur, India. All reactions were monitored by thin layer chromatog-
raphy (TLC) on precoated silica gel 60 F254 (mesh); sports were vi-
sualized under UV light
4.41 (q, J = 8.3 Hz, 1H, CH), 3.78-3.86 (m, 2H. CH ), 3.16 (dd,
2
J = 14.0, 4.3 Hz, 1H, CHa of CH ), 2.56–2.72 (m, 1H, CHb of
2
13
CH ), 2.36-2.41 (m, 3H, CH ). C-NMR (400 MHz, CDCl ) δ 171.51,
2
3
3
1
67.36 (C=O), 160.72, 144.44, 134.88, 133.97, 133.84, 129.96, 129.81,
129.08, 128.80, 127.44, 127.24, 121.99, 121.91, 115.66, 115.44 (aro-
matic carbons), 58.72, 43.91, 38.03, 21.67(four aliphatic carbons).
HRMS-ESI found value is (m/z): calcd. for C24H25FN O S [M+H]
3
4
470.15; found 470.16.
N-(2-(4-chlorophenylamino)-2-oxoethyl)-2-(4-
methylphenylsulfonamido)-3-phenylpropanamide (7c)
Yield 72.5%, Mp=92–93 °C. FTIR (KBr cm 1), 3344 (N–H), 3063
−
(
C–H aromatic), 2925 (C–H aliphatic), 1664, 1597 (2C=O), 1539,
1
493, 1400 (C=C), 1305, 1158 (2S=O), 1091, 1030 (C–N), 1244
1
(
SO N), 722 (C–Cl). H-NMR (400 MHz, CDCl ) δ 8.72 (s, 1H, NH),
2 3
2
.1. Synthesis of substituted benzenesulphonamoyl alkanamides (3)
7
.61 (d, J = 9.2 Hz, 2H, ArH), 7.51–7.54 (m, 1H, NH), 7.38 (d,
[
22]
J = 8.5 Hz, 2H, ArH), 7.23 (t, J = 9.8 Hz, 2H, ArH), 7.08-7.18 (m,
5
4
H, ArH), 6.85 (d, J = 7.3 Hz, 2H, ArH), 5.30-5.33 (m, 1H, NH),
The modified methods of Ugwu et al., 2017 was used for the
.42 (q, J = 8.3 Hz, 1H, CH), 3.73-3.85 (m, 2H, CH ), 3.17 (dd,
2
synthesis of the substituted benzenesulphonamoyl alkanamides.
The details is found in the supporting document.
J = 14.3, 4.6 Hz, 1H, CHa of CH ), 2.68 (dd, J = 14.0, 10.4 Hz, 1H,
2
13
CHb of CH ), 2.37-2.42 (m, 3H, CH ). C-NMR (400 MHz, CDCl ) δ
2
3
3
1
71.38, 167.34 (C=O), 144.40, 136.32, 134.70, 133.76, 129.87, 129.37,
2
.2. General procedure of preparing compound (6a-j) [23]
129.00, 128.84, 128.67, 127.37, 127.15, 121.35 (twelve aromatic car-
bons), 58.60, 43.88, 37.90, 21.58 (four aliphatic carbons). HRMS-ESI
The methods of Sharm and Soman 2016, Ugwuja et al., 2019
−
(
m/z): calcd. for C24H23ClN O S [M-H] 484.11; found 484.11.
3
4
and Ezugwu et al., 2020 were adopted for this synthesis with little
modifications. See supporting document for details.
N-(2-(4-bromophenylamino)-2-oxoethyl)-2-(4-
methylphenylsulfonamido)-3-phenylpropanamide (7d)
Yield 66.7%, Mp=94–95 °C. FTIR (KBr cm 1), 3347 (N–H), 3063
−
2
.3. General procedure of preparing (7a-j)
mixture of alkanamide (3, 1.84 mmol), 1-ethyl-3-(3-
(
C–H aromatic), 2925 (C–H aliphatic), 1662, 1597 (2C=O), 1532
A
1489, 1455, 1396 (C=C), 1305, 1158 (2S=O), 1091, 1073 (C–N),
1
dimethylamino carbodiimide hydrochloride EDC.HCl (0.53 g, 2.76
mmol), 1-hydroxybenzotriazole HOBt (0.248 g, 1.84 mmol), triethy-
lamine, compounds (6a-j), 1.53 mmol) in dichloromethane (50 mL)
was stirred at room temperature for 16 h and monitored using TLC.
Upon completion of the reaction, the mixture was washed with
water (2 × 20 mL), brine (1 × 10 mL), dried over sodium sulphate.
The crude product was obtained after evaporating the solvent un-
der reduced pressure and then purified by column chromatography
using silica gel (3% MeOH/DCM).
1244 (SO N), 551 (C–Br). H-NMR (400 MHz, CDCl ) δ 8.69 (s,
2
3
1H, NH), 7.55 (d, J = 8.5 Hz, 2H, ArH), 7.48 (d, J = 7.3 Hz,
1H, NH), 7.37 (q, J = 4.3 Hz, 4H, ArH), 7.07–7.20 (m, 5H, ArH),
6.84 (d, J = 7.3 Hz, 2H, ArH), 5.25 (d, J = 4.3 Hz, 1H, NH),
4.41 (q, J = 8.3 Hz, 1H, CH), 3.76–3.83 (m, 2H, CH ), 3.16 (dd,
2
J = 14.0, 4.3 Hz, 1H, CHa of CH ), 2.55-2.70 (m, 1H, CHb of
2
13
CH ), 2.34-2.41 (m, 3H, CH ). C-NMR (400 MHz, CDCl ) δ 171.43,
2
3
3
167.42 (C=O), 144.55, 136.94, 134.75, 133.76, 131.90, 130.00, 129.15,
128.76, 127.52, 127.27, 121.79, 117.15 (twelve aromatic carbons),
2

Babatunde.S. Aronimo, Uchechukwu.C. Okoro, Rafat. Ali et al.
Journal of Molecular Structure 1246 (2021) 131201
5
8.67, 43.98, 37.99, 21.70 (four aliphatic carbons). HRMS-ESI (m/z):
129.00, 128.91, 128.78, 127.34, 127.24, 125.39, 120.94, 117.40 (four-
teen aromatic carbons), 58.63, 44.14, 38.17, 21.64 (four aliphatic
+
calcd. for C24H25BrN O S [M+H] 530.07; found 530.08.
3
4
+
carbons). HRMS-ESI (m/z): calculated for C25H28N O S [M+H]
3
4
2-(4-methylphenylsulfonamido)-N-(2-oxo-2-(p-tolylamino)
466.18; found 466.18.
ethyl)-3-phenylpropanamide (7e)
Yield 68.9%, Mp=76-77 °C. FTIR (KBr cm−1), 3346 (N–H), 3063,
N-(2-(2,6-dimethylphenylamino)-2-oxoethyl)-2-(4-
methylphenylsulfonamido)-3-phenylpropanamide (7i)
3
030 (C–H aromatic), 2924 (C–H aliphatic), 1660, 1601 (2C=O),
Yield 92.7%, Mp=173-174 °C. FTIR (KBrcm 1), 3352 (N–H), 3030
−
1
515, 1498, 1454,1407 (C=C), 1319, 1159 (2S=O),1092, 1031 (C–N),
1
2 3
(C–H aromatic), 2927 (C–H aliphatic), 1662, 1599 (2C=O), 1515,
1
248 (SO N). H-NMR (396 MHz, CDCl ) δ 8.52 (s, 1H, NH),
1
455, (C=C), 1329, 1159 (2S=O), 1092, 1032 (C–N), 1237 (S0 N).
7
.38-7.51 (m, 5H, ArH), 7.07–7.19 (m, 7H, ArH), 6.85 (d, J = 7.9
2
1
H-NMR (400 MHz, CDCl ) δ 8.01 (s, 1H, NH), 7.57 (d, J = 5.5 Hz,
Hz, 2H, ArH), 5.25-5.29 (m, 1H, NH), 4.34 (q, J = 8.3 Hz, 1H,
3
1
6
H, NH), 7.35 (d, J = 8.5 Hz, 2H, ArH), 7.04–7.17 (m, 8H, ArH),
.88 (d, J = 6.7 Hz, 2H, ArH), 5.39 (d, J = 5.5 Hz, 1H, NH), 4.36
CH), 3.77-3.83 (m, 2H, CH ), 3.13 (dd, J = 14.2, 4.5 Hz, 1H,
2
CHa of CH ), 2.70 (dd, J = 13.9, 10.3 Hz, 1H, CHb of CH ),
2
2
1
3
(dd, J = 17.1, 7.3 Hz, 1H, CH), 3.82–3.97 (m, 2H, CH ), 3.14 (dd,
2
.28–2.40 (m, 6H, 2CH ).
C-NMR (101 MHz, CDCl ) δ 171.53,
2
3
3
J = 14.0, 4.3 Hz, 1H, CHa of CH ), 2.73 (dd, J = 14.0, 9.8 Hz, 1H,
1
67.26 (C=O), 144.22, 135.19, 135.07, 134.34, 134.14, 129.89, 129.46,
28.99, 128.90, 127.33, 127.25, 120.33 (twelve aromatic carbons),
2
CHb of CH ), 2.40 (d, J = 10.4 Hz, 3H, CH ), 2.22 (d, J = 7.9 Hz, 6H,
1
2
3
13
2
CH ). C-NMR (400 MHz, CDCl ) δ 171.57, 167.69 (C=O), 144.20,
5
8.66, 44.01, 38.13, 21.65, 20.98 (five aliphatic carbons). HRMS-
3
3
+
137.60, 135.64, 135.06, 133.45, 129.86, 129.02, 128.89, 128.20,
27.49, 127.37, 127.17(twelve aromatic carbons), 58.68, 43.59, 38.18,
1.65, 18.45 (five aliphatic carbons). HRMS-ESI (m/z): calcd. For
ESI (m/z): calcd. for C25H28N O S [M+H] value is 466.18; found
3
4
1
4
66.18.
2
+
C26H30N O S [M+H] 480.20; found 420.20.
2
-(4-methylphenylsulfonamido)-N-(2-morpholino-2-oxoethyl)-3-
3
4
phenylpropanamide (7f)
N-(2-(4-methoxyphenylamino)-2-oxoethyl)-2-(4-
Yield 70.6%, Mp=142–143 °C. FTIR (KBr cm 1), 3293 (N–H),
−
methylphenylsulfonamido)-3-phenylpropanamide (7j)
Yield 70.2%, Mp=77–78 °C. FTIR (KBr cm 1), 3352 (N-H), 3064
2
921, 2857 (C–H aliphatic), 1638, (C=O), 1530, 1498, 1437 (C=C),
−
1
1
333, 1161 (2S=O), 1093, 1034 (C–N), 1245 (SO N). H-NMR (400
2
(
C–H aromatic), 2928 (C–H aliphatic), 1660, 1601 (2C=O), 1512,
MHz, CDCl ) δ 7.52 (d, J = 7.9 Hz, 2H, ArH), 7.15-7.18 (m, 6H,
3
1
455, 1442, 1414 (C=C), 1325, 1159 (2S=O), 1032, (C–N), 1246
ArH), 6.96 (dd, J = 7.6, 1.5 Hz, 2H, ArH), 5.31 (d, J = 7.9 Hz, 1H,
1
(
SO N). H-NMR (400 MHz, CDCl ) δ 8.57 (s, 1H, NH), 7.53–
2 3
NH), 3.85-4.02 (m, 3H, CH&CH ), 3.62-3.68 (m, 6H,3CH ), 3.37 (t,
2
2
7
.63 (m, 3H, ArH), 7.40 (d, J = 8.5 Hz, 2H, ArH), 7.07–7.21 (m,
J = 4.9 Hz, 2H, CH ), 2.88–3.03 (m, 2H, CH ), 2.40 (s, 3H, CH ).
2
2
3
5
H, ArH), 6.84 (dd, J = 18.6, 8.2 Hz, 4H, ArH), 5.45 (d, J = 4.9
13
C-NMR (400 MHz, CDCl ) δ 170.63, 166.22(C=O), 143.52, 136.27,
3
Hz, 1H, NH), 4.35 (q, J = 8.1 Hz, 1H, CH), 3.80–3.85 (m, 2H,
CH ), 3.74 (d, J = 11.0 Hz, 3H, OCH ), 3.14 (dd, J = 14.0, 4.3
135.57, 129.72, 129.26, 128.79, 127.23, 127.16(eight aromatic car-
2
3
bons), 66.74, 66.39, 58.02, 44.89, 42.40, 41.35, 38.87, 21.64 (eight
Hz, 1H, CHa of CH ), 2.71 (dd, J = 14.0, 10.4 Hz, 1H, CHb of
2
aliphatic carbons). HRMS-ESI (m/z): calcd. for C22H27N NaO S
3
5
13
CH ), 2.32–2.45 (m, 3H, CH ) C-NMR (400 MHz, CDCl ) δ 171.50,
M+Na]+ 468.16; found 468.16.
2
3
3
[
1
67.15(C=O), 156.53, 144.27, 135.02, 134.26, 130.91, 129.90, 129.02,
28.88, 127.35, 127.26, 121.95, 114.09 (twelve aromatic carbons),
1
2-(4-methylphenylsulfonamido)-N-(2-(naphthalen-1-ylamino)-2-
5
8.67, 55.50, 43.90, 38.10, 21.65(five aliphatic carbons). HRMS-ESI
oxoethyl)-3-phenylpropanamide (7g)
+
(
m/z): calcd. For C26H30N O S [M+H] 482.17; found 482.18.
Yield 75.2%, Mp=85–86 °C. FTIR (KBr cm−1), 3345 (N–H), 3060
3
4
(
C–H aromatic), 2927 (C–H aliphatic), 1662, 1599 (2C=O), 1531,
2
.4. In vivo antimalarial test
1
1
499, 1445 (C=C), 1328, 1159 (2S=O), 1091, (C–N). H-NMR (400
MHz, CDCl ) δ 8.83 (s, 1H, NH), 7.97 (d, J = 8.5 Hz, 1H, NH),
3
The method of Peter et al. [24] and Kalra et al. [25] were
7
.81 (dd, J = 20.4, 7.6 Hz, 2H, ArH), 7.70 (d, J = 7.9 Hz, 1H,
adopted for antiplasmodial activity with some modification. Forty-
eight infected mice were randomly divided into twelve groups
ArH), 7.37-7.56 (m, 7H, ArH), 7.04-7.15 (m, 4H, ArH), 6.87 (d,
J = 7.3 Hz, 2H, ArH), 5.30-5.35 (m, 1H, NH), 4.34 (q, J = 7.9 Hz,
(
four mice per group). Group 1 to 10 composed the treatment
1
H, CH), 3.90–4.08 (m, 2H, CH ), 3.11 (dd, J = 14.0, 4.9 Hz, 1H,
2
group and was given 50 mg/kg body weight. While group 11 and
12 served as the positive control (received commercial samples
of Artemether/ Lumefantrin combination) and the negative con-
trol respectively. The inoculum was prepared from a donor mouse
with a minimum of peripheral parasiemia 20% by cardiac punc-
ture in EDTA-coated tube. Five days after the inoculation of the
mice with parasite, percentage parasitaemia was determined and
treatment with the synthesized compound (7a-j) was started and
this was done for three days. All the compounds and the standard
drug (Artemether/ Lumefantrin) were administered orally using a
standard intragastric tube. For all the parasitaemia determination,
blood samples were collected from the mice through the retrobul-
bar plexus of the median canthus of the mice eye.
CHa of CH ), 2.79 (dd, J = 14.3, 9.5 Hz, 1H, CHb of CH ), 2.36
2
2
13
(
d, J = 9.8 Hz, 3H, CH ). C-NMR (400 MHz, CDCl ) δ 172.16,
3
3
1
68.33 (C=O), 144.03, 135.08, 134.83, 134.18, 132.10, 129.81, 128.99,
28.91, 128.48, 127.79, 127.25, 127.14, 126.41, 126.14, 125.57, 121.90,
21.78 (seventeen aromatic carbons), 58.54, 44.54, 38.32, 21.58
four aliphatic carbons). HRMS-ESI (m/z): calcd. for C28H28N O S
1
1
(
[
3
4
M+H]+ 502.18; found 502.8.
2-(4-methylphenylsulfonamido)-N-(2-oxo-2-(m-tolylamino)
ethyl)-3-phenylpropanamide (7h)
Yield 89.9%, Mp=175–176 °C. FTIR (KBr cm−1), 3347 (N–H),
3
060, 3030 (C–H aromatic), 2924, 2860 (C–H aliphatic), 1662, 1615
(
2C=O), 1596, 1547, 1492, 1454, 1434 (C=C), 1322, 1158 (2S=O),
1
1
091, 1031 (C–N), 1259 (SO N). H-NMR (400 MHz, CDCl ) δ 8.59
2.5. Haematological analysis
2
3
(
s, 1H, NH), 7.53-7.56 (m, 1H), 7.50 (s, 1H, NH), 7.30–7.42 (m,
3
5
3
2
H, ArH), 7.07–7.20 (m, 6H, ArH), 6.89 (q, J = 7.1 Hz, 3H, ArH),
.48 (d, J = 4.9 Hz, 1H, NH), 4.31 (q, J = 7.9 Hz, 1H, CH), 3.81-
Whole blood used for the tests was collected from the mice
through the retrobulbar plexus of the median canthus of the mice
eye. Packed Cell Volume (PCV), Haemoglobin (Hb) levels and Red
blood cell (RBC) count were determined before and after the treat-
ment to determine the effect of the various treatments on the pa-
rameters.
.86 (m, 2H, CH ), 3.13 (dd, J = 14.0, 4.9 Hz, 1H, CHa of CH2),
2
.72 (dd, J = 14.3, 10.1 Hz, 1H, CH of CH ), 2.42 (d, J = 23.2
b
2
13
Hz, 3H, CH ), 2.30 (s, 3H, CH ). C-NMR (400 MHz, CDCl ) δ
1
71.55, 167.30 (C=O), 144.22, 138.82, 137.65, 135.05, 134.46, 129.90,
3
3
3
3

Babatunde.S. Aronimo, Uchechukwu.C. Okoro, Rafat. Ali et al.
Journal of Molecular Structure 1246 (2021) 131201
2
.6. Liver function tests (LFTs)
3.1. In vivo antimalarial
The methods of Reitman et al. [26] for the determination of as-
To assess the in vivo antimalarial activity, the synthesized com-
pounds were tested against P. berghei (NK-65) which was obtained
from National Institute of Medical Research, Yaba Lagos, Nigeria.
The university of Nigeria ethical committee for the use of animal
gave approval for this project. Artemether/ Lumefantrine was used
as the standard drug for the antimalarial activity. The percentage
partate aminotransferase (AST), Alanine transaminase (ALT) and Al-
kaline phosphate (ALP) were used.
2
.7. Kidney function test
(
%) inhibition of the parasite multiplication was calculated by com-
Kidney Function tests carried out on the blood of the mice that
paring the treated group with untreated group using the following
formula [17, 18, 21].
were fed with the sulphonamide derivative were creatinine and al-
bumin adopting the modified method of Kaplan et al. [27].
%
Inhibition = Mean % parasiteamia of before-treatment -
Mean % parasiteamia of after-treatment x 100
2
.8. Molecular docking method
Mean % parasiteamia of before-treatment
Molecular docking was carried out to determine the binding
Compounds that reduced parasitaemia by at least 40% were
considered active, whereas those that reduce parasitaemia by 30-
potential of ten synthesized compounds against plasmepsin II. 3D
crystal structure PDB access code: 4Z22[28] was retrieved from
4
0% were considered partially active while less than 30% were in
˚
protein data bank with resolution of 2.65 A. This PDB code was
active [17, 18, 21]. From data in Table 1, some of the synthesized
compounds were active at the 50 mg/kg dose when compared with
the standard drug. Specifically, compound 7j had percentage inhi-
bition of 60.2 against P. berghei which was close to 68.8% obtained
from the standard drug. Compounds 7h and 7i were partially ac-
tive while the rest were inactive. So these three compounds could
be considered for further studies. Structure activity relationship
study reveals the most potent compound 7j had a Methoxy-group
at the 4-position. The 3-methyl derivative was at the distance sec-
ond position. The order of activity is 4-methoxy> 3-methy>2,6-
dimethyl>4-Chloro>4-methyl>Naphthalyl>benzene>4-Bromo. The
least active was 4-Fluoro derivative (7b). The trend of activity sug-
gests that highly electron donating group at position 4 will posi-
tively influence the actimalarial properties of the derivatives. We
also observed that aromaticity influenced the activity positively as
expressed in the compounds 7a, 7f, and further increase of aro-
matic ring increased activity as in compound 7 g.
selected based on the resolution of the crystal structure and Plas-
mepsin II is an aspartic protease encoded by P. falciparum (1 L.A.
Gil, P. A.Valiente,P.G.Pascutti andT.Pons,Computational perspectives
into plasmepsins structure-function relationship: implications to
inhibitors design, J. Trop. Med., 2011, 657483.)
AutoDock tools 1.5.4 was used to determine the grid box size
for the potential binding site [29]. The structure of the compounds
were optimized with Gaussian 09 [30]. The determined dimension
˚
was X= 24 Y= 24 Z= 24 with 1.00 A as the grid spacing. Lamar-
ckian genetic algorithm method was applied to obtain optimum
binding site for the ligand [31]. Gasteiger charges were computed
using the AutoDock Tools graphical user interface supplied by MGL
Tools [32]. The co-crystal ligand was also docked into the active
site of the plasmepsin to validate the docking.
3.0. Results and discussion
The first step in synthesis of the compounds involved
3.2. Liver function analysis
the reaction of substituted benzenesulphonyl chloride with L-
phenylalanine in an alkaline medium to give substituted ben-
zenesulphonamide [3]. The reaction of Boc-glycine, and amines in
the presence of peptides coupling regents, 1-ethyl-3-(3-dimethyl
amino propyl carboxiimide hydrochloride, 1-hydroxybenzotriazole
and Triethylamine in dichloromethane provided the carbmate
derivatives of glycine which on further reaction with 30% trifluo-
roacetic acid (TFA) gave the TFA salt of unprotected amides (6a-
j).The coupling reaction between the solid TFA salt of unprotected
amides with the (N-phenylsulfonyl) phenylalanine) afforded the
desired products (7a-j). The structure of the compounds (7a-j) was
confirmed by characterization using FTIR, 1HNMR, 13CNMR and
High resolution mass spectroscopy. The presence of sharp bands
Liver function tests are a group of blood tests that are used to
determine inflammation and damage to the liver [17]. The liver
function parameters evaluated in this research are AST, ALT and
ALP. The results of LFT (Table 2) showed that the administration
of 50 mg/kg of the tested compounds did not lead to substantial
increase or decrease in the levels of liver parameters. The result
showed that the compounds were not toxic to the animals.
3.3. Kidney function analysis
This is used to determine how well the kidneys are performing.
The data in Table 3 revealed that there is no significant change
in the serum level of urea and creatinine of mice fed with 50
mg/kg of the reported derivatives when compared with the refer-
ence drug (Artemether/ Lumefantrin). The result showed that the
compounds were not toxic to the animals.
between 3344 and 3352 cm 1, 1597 and 1664 cm
−
−1
are assigned
to NH and C=O respectively. In the 1H NMR of 7a, the prominent
NH resonance of the sulfonamide part of the dipeptide hydride was
observed at δ5.17 ppm. Other amide NH resonances of the com-
pounds appeared at δ7.09–7.21 ppm with aromatic protons and at
δ8.60 ppm region as multiplet and singlet peaks, respectively. In
the 13CNMR spectrum, two peaks at 167.45 and 171.58 ppm for
the carbonyl carbons of the amide groups, twelve peaks ranging
from 120.29 to 144.28 ppm for aromatic carbons and four peaks
ranging from 21.66 to 58.68 ppm for aliphatic carbons confirmed
the formation of 7a, which was supported by High resolution mass
Table 4 presents the results of the heamatological parameters.
Though there is a reduction in the value of RBC, PCV and HB, it
was observed that they were not substantial when compared with
the standard (Artemether/ Lumefantrin). The result showed that
the compounds were not toxic to the animals.
3.4. Molecular studies
+
spectrometer spectrum with a peak at m/z 474.15 for [M+Na] .
All other compounds were in agreement with their structures. The
spectra and other important data are found in the supplementary
information.
The results obtained from the molecular docking of com-
pounds 7a-7j with Plasmodium falciparum plasmepsin II are pre-
sented in Table 5. The results obtained showed that 7j had the
4

Babatunde.S. Aronimo, Uchechukwu.C. Okoro, Rafat. Ali et al.
Journal of Molecular Structure 1246 (2021) 131201
Scheme 1. The synthesis of dipeptides bearing sulphonamide. (i) Na2CO3, DCM/H2O, HCl, 0°°C, r.t, pH 2, 4°h (ii) EDC. HCl, HOBt, TEA, DCM, amines, 16 h (iii) 10% TFA in
DCM (iv) EDC.HCl, HOBt, TEA, 16 h.
Table 1
Percentage inhibition of parasite in mice.
Compounds No.
% parasitaemia Before treatment
% Parasitaemia After treatment
% inhibition
7
7
7
7
7
7
7
7
7
7
a
b
c
d
e
f
60.5
61.3
53.8
60.3
51.5
55.0
55.3
62.5
58.0
61.0
56.0
49.8
57.0
39.7
49.0
39.3
49.0
68.0
38.3
38.0
24.3
17.5
17.7
7.0
26.2
18.7
23.7
10.9
−23.0
38.7
34.5
60.2
68.8
g
h
i
j
Artemether/ Lumefantrin
Table 2
Liver function test.
μmol/l
Total
μmol/l
IU/L
ALP
IU/L
ALT
IU/L
AST
Direct
Group
Bilirubin
Bilirubin
7
7
7
h
i
46.3
45.7
48.3
20.6
30.5
33.5
22.5
32.5
8.5
22.4
18.3
24.3
23.6
7.0
9.0
5.3
j
8.0
10.0
8.9
Artemether/
Lumefantrin
20.0
highest binding energy of (−6.87) kcal mol–1) compared to the
co-crystalized ligand and standard drug even though all com-
pounds had relatively low binding energy. This observation agrees
with the In vivo antimalarial studies. Hydrogen bonding is a cru-
cial marker indicating functional stability and binding of ligands
to crucial amino acids. Fig. 1 revealed that SER118 an amino
acid around the active site formed hydrogen bonding with the
atoms of 7j. Fig. 1 also revealed Pi-sigma, Van der Waals inter-
actions and carbon hydrogen bond interactions. These interactions
could proffer better insight in proposing the mechanism of in-
hibition. The docking simulations was authenticated by docking
the co-crystallized ligand against the protein under study. The
5

Babatunde.S. Aronimo, Uchechukwu.C. Okoro, Rafat. Ali et al.
Journal of Molecular Structure 1246 (2021) 131201
Fig. 1. 3D structure of (A) compound 7j in complex Plasmepsin II (highest binding affinity). (B) Descriptions showing the hydrogen bonding interactions with amino acid
residues around the binding pocket (ASP 214).
Table 3
Table 5
Kidney function test.
Binding energies of ligands in complex with antimalaria en-
zymes (plasmepsin II: 4Z22) obtained from molecular dock-
ing, using AutoDock 4.2.
Mmol/l
Urea
μmol/l
Group
Creatinine
Binding energies (kcal mol 1
−
)
Complexes
7
7
7
h
i
6.2
5.5
7.0
6.8
27.5
23.6
60.4
59.5
7a
7b
7c
7d
−4.17
−3.29
−4.11
−4.06
−5.22
−4.02
−4.22
−5.10
−5.22
−6.87
−6.13
−6.69
j
Artemether/
Lumefantrin
7
7
7
7
7
7
e
f
g
h
i
j
Co-crystalized ligand
Standard drug
˚
obtained RMSD value of 0.2 A revealed a structural and func-
tional stability of ligand-protein complex. The docking result ob-
tained in this work agrees with that obtained in the literature
(
D. I. Ugwu, U. C. Okoro, P. O. Ukoha, et al., Synthesis, character-
4. Conclusion
ization molecular docking and in vitro antimalarial properties of
new carboxamides bearing sulphonamide. Eur J Med Chem. 135
In conclusion, ten new L-phenylalanine derived dipeptide bear-
(2017)349–69.)
ing p-toluenesulphonamides were successfully synthesized and
Table 4
Heamotological test before and after treatment.
RBC (mm3)
PCV (%)
HB (g/dl)
Comp
Before
After
Before
After
Before
After
6
6
6
6
6
6
6
6
7
7
7
h
i
7.8 × 10
6.7 × 10
7.6 × 10
7.1 × 10
8.8 × 10
9.6 × 10
9.2 × 10
9.8 × 10
43
43
41
39
53
53
52
53
10.8
12.8
13.0
11.2
13.8
17.8
17.3
17.4
j
Artemether/ Lumefantrin)
6

Babatunde.S. Aronimo, Uchechukwu.C. Okoro, Rafat. Ali et al.
Journal of Molecular Structure 1246 (2021) 131201
characterized using spectroscopic techniques. The molecular dock-
ing results showed that the compounds interacted with the ac-
tive site of the protein target with good binding affinity using
conventional hydrogen bonding. In the antimalarial activity study,
compound 7j showed the most antimalarial activity with per-
centage inhibition of parasite growth of 60.2% comparable with
artemether/lumenfantrine (68.8%). The results of the haematolog-
ical analysis, liver and kidney function tests showed that there
were no significant changes in the parameters tested when com-
pared with the control. The physicochemical parameter predictions
indicate that the compounds would not pose oral bioavailability,
transport and permeability problems if developed further to drug
molecules The molecular docking studies showed good interaction
between the synthesized compounds and the protein targets for
antimalarial activities.
[11] R. Mahtab, A Srivastava, N. Gupta, S.K. Kushwaha, A. Tripathi, Synthesis of
novel 2-benzylbenzo[d]thiazole-6-sulfonamide derivatives as potential anti-in-
flammatory agents, J. Chem. Pharm. Sci. 7 (2014) 34–38.
[
12] M. Siddique, A.B. Saeed, S. Ahmad, N.A. Dogar, Synthesis and biological evalu-
ation of hydrazide based sulphonamides, J. Sci. Innovat. Res. 2 (2013) 628–634
Available online at http://www.jsirjounal.com .
[
13] M. Jainswal, P.V. Khadikar, C.T. Supran, Topological modeling of lipophilicity
diuretic activity and carbonic inhibition activity of benzenesulphonamides: a
molecular connectivity approach, Bioorg. Med. Chem. Lett. 14 (2004) 5666,
doi:10.1016/j.bmcl.2004.08.051.
[14] C.T. Supuran, Carbonic anhydrase: novel therapeutic applications for inhibitors
and activators, Nat. Rev. Drug Disc. 7 (2008) 168, doi:10.1038/nrd2467.
[15] G. Laudadio, E. Barmpoutsis, C. Schotten, L. Struik, S. Govaerts, L. Duncan.
Browne, T. Noël, Sulfonamide synthesis through electrochemical oxidative cou-
pling of amines and thiols, J. Am. Chem. Soc. 141 (2019) 5664–5668.
[
16] H. Küçükbay, N. Bu g˘ day, F. Zehra Küçükbay, A. Ageli, G. Bartolucci, T. Claudiu.
Supuran, Synthesis and human carbonic anhydrase I, II, VA, and XII inhibi-
tion with novel amino acid–sulphonamide conjugates, J. Enzyme Inhibit. Med.
Chem. 35 (1) (2020) 489–497 DOI: 10.1080/14756366.2019.1710503.
[
17] D.I. Ugwuja, U.C. Okoro, S.S. Soman, R. Soni, S.N. Okafor, D.I. Ugwu, New pep-
tide derived antimalaria and antimicrobial agents bearing sulphonamide moi-
ety, J. Enzyme Inhibit. Med. Chem. 34 (1) (2019) 1388–1399.
Declaration of Competing Interest
The authors declared no Conflict of Interest.
Acknowledgements
[
18] J.A. Ezugwu, U.C. Okoro, M.A. Ezeokonkwo, C.R. Bhimapaka, S.N. Okafor,
D.I. Ugwu, D.I. Ugwuja, Synthesis and biological evaluation of Val–Val
dipeptide–sulfonamide conjugates, Archiv. Der Pharm. 353 (7) (2020), doi:10.
1002/ardp.202000074.
[
19] P. Wang, M. Read, P.F. Sims, J.E. Hyde, Sulfadoxine resistance in the human
malaria parasite Plasmodium falciparum is determined by mutations in dihy-
dropteroate synthase and an additional factor associated with folae utilization,
Mol. Microbiol. 23 (1997) 979–986.
Samuel. T. Aromino gratefully acknowledge the assistance of
Prof Sandeep Verma, Department of Chemistry, Indian Institute of
Technology, Kanpur, India in providing the facilities for synthesis
and spectral characterization.
[20] W. Pornthanakasem, P. Riangrungroj, P. Chitnumsub, W. Ittarat, D. Kongkasuriy-
achai, C. Uthaipibull, Y. Yuthavong, U. Leartsakulpanich, Role of plasmodium
vivax dihydropteroate synthase polymorphisms in sulfa drug resistance, Agent
Chemother. 60 (2016) 4453–4463.
[
21] J.A. Ezugwu., U.C. Okoro., M.A. Ezeokonkwo, C.R. Bhimapaka, Synthesis of novel
valine-based dipeptide carboxamide bearing benzene sulfonamide moiety as
antimalarial agent, Commun. Phys. Sci. 5 (2) (2020) 176197.
Supplementary materials
[
22] D.I. Ugwu, U.C. Okoro, P.O. Ukoha, et al., Synthesis, characterization molecu-
lar docking and in vitro antimalarial properties of new carboxamides bearing
sulphonamide, Eur. J. Med. Chem. 135 (2017) 349–369.
Supplementary material associated with this article can be
found, in the online version, at doi:10.1016/j.molstruc.2021.131201.
[
23] R. Sharma, S .S. Soman, Design and synthesis of novel diamide derivatives of
glycine as antihyperglycemic agents, Synth. Commun. 46 (2016) 1307–1317.
24] W. Peters, J.H. Portus, B.L. Robinson, The chemotherapy of rodent malaria, XXII
the value of drug resistant strains of berghei in screening for blood schizonto-
cidal activity, Ann. Trop. Med. Parasitol. 69 (1975) 155–171.
Reference
[
[
1] www.who.int/news-room/fact-sheets/detail/malaria
[
2] H. Biruk, B. Sentayehu, Y. Alebachew, W. Tamiru, A. Ejigu, S. Assefa, In vivo
antimalarial activity of 80% methanol and aqueous bark extracts of terminalia
brownii Fresen. (Combretaceae) against plasmodium berghei in mice, Biochem.
Res. Int. (2020) 1–7 doi:10.1155/2020/97494103.
[25] B.S. Kalra, S. Chawla, P. Gupta, N. Valecha, Screening of antimalarial drugs: an
overview, Indian J. Pharmacol. 38 (2006) 5–12.
[26] S. Reitman, S.A. Frankel, Colorimetioc method for the determination of serum
glutamic oxaloacetic and glutamic pyruvic transaminase, Am. J. Clin. Pathol. 28
(1957) 56–63.
[
3] U. Farooq, R.C. Mahajan, Drug resistance in malaria, J. Vector Borne Dis. 41
(
3-4) (2004) 45–53.
[27] A. Kaplan, and L. L. Teeng, Selected Methods of Clinical Chemistry, W.R.
Faulkner and S. Meits. Washington, DC: AACC; 9 (1982) 357–63.
[28] D. Rasina, M. Otikovs, J. Leitans, R. Recacha, O.V. Borysov, L. Kanepe-Lapsa,
I. Domraceva, T. Pantelejevs, K. Tars, M.J. Blackman, Fragment-based discov-
ery of 2-aminoquinazolin-4 (3 H)-ones as novel class nonpeptidomimetic in-
hibitors of the plasmepsins I, II, and IV, J. Med. Chem. 59 (2016) 374–387.
[29] M.F. Sanner, Python: a programming language for software integration and de-
velopment, J. Mol. Graph. Model. 17 (1999) 57–61.
[
4] World Health Organization (2014). World Malaria Report
[
5] S. Sinha, B. Medhi, R. Sehgal, Challenges of drug-resistant malaria, Parasite 21
(
2014) 61, doi:10.1051/parasite/20140596.
6] J.J. Nestor Jr, The medicinal chemistry of peptides, Curr. Med. Chem. 16 (33)
2009) 4399–4418.
[
[
[
[
(
7] B.H. Sarmadi, A. Ismail, Antioxidative peptides from food proteins: a review,
Peptides 31 (10) (2010) 1949–1956, doi:10.1016/j.peptides.2010.06.0208.
8] S.A. Ali, A.J. Qasir, K.Y Saor, Synthesis of some sulfonamide derivatives with
expected antibacterial activity, Iraqi J. Mark. Rec. Cons. Prot. 1 (2009) 85–93.
9] H.S. Chandak, Synthesis of isoxazolylbenzenesulphonamide derived from
[30] M. Frisch, G. Trucks, H.B. Schlegel, G. Scuseria, M. Robb, J. Cheeseman, G. Scal-
mani, V. Barone, B. Mennucci, G. Petersson, Gaussian 09, Revision D. 01, Gaus-
sian, Inc., Wallingford CT, 2009.
N-[4-(2,3-dibromo-2-arylpropanoyl)phenyl]benzenesulphonamide,
Pharma. Chem. 4 (2012) 1054–1057.
10] M.M. Ghorab, M.S. Bashandy, M.S. Alsaid, Novel thiophene derivatives with
sulphonamides, isoxazole, benzothiazole, quinoline and anthracene moieties
as potential anticancer agents, Acta Pharm. 64 (2014) 419–431, doi:10.2478/
acph-2014-0035.
Der.
[31] B. Yang, F. Hao, J. Li, D. Chen, R. Liu, Binding of chrysoidine to catalase: spec-
troscopy, isothermal titration calorimetry and molecular docking studies, J.
Photochem. Photobiol. B: Biol. 128 (2013) 35–42.
[
[32] G.M. Morris, R. Huey, W. Lindstrom, M.F. Sanner, R.K. Belew, D.S. Goodsell,
A.J. Olson, AutoDock4 and AutoDockTools4: automated docking with selective
receptor flexibility, J. Comput. Chem. 30 (2009) 2785–2791.
7