New isoleucine derived dipeptides as antiprotozoal agent: Synthesis, in silico and in vivo studies.

Article abstract of DOI:10.1016/j.molstruc.2021.130017

The increasing emergence of malaria drug-resistant parasites and the deficiency in effective chemotherapy for trypanosomiasis represents a huge challenge in infectious disease treatment in tropical regions. As regards to developing effective antiprotozoal agents, ten new ile-gly dipeptide sulphonamide derivatives were synthesized by condensing compound (10) with (8a-j)using peptide coupling reagents. Compounds11b, 11i and 11j were most potent in clearing Trypanosome brucei in mice with 11b showing comparable activity with diminazene aceturate. In the antimalarial study, 11b was the most active compound, even better than the standard. Molecular docking result suggests good interaction between the reported compounds and the target protein. The results of haematological analysis, liver and kidney function tests showed that the compounds had no adverse effect on the blood and organs. Compound 11b stands out amongst the derivatives haven shown better activity in both the antimalarial and antitrypanosomal assay.

Full text of DOI:10.1016/j.molstruc.2021.130017

Journal of Molecular Structure 1231 (2021) 130017
Contents lists available at ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstr
New isoleucine derived dipeptides as antiprotozoal agent: Synthesis,
in silico and in vivo studies.
Ogechi C. Ekoha,c,∗, Uchechukwu C. Okorob, Rafat Alic, David I. Ugwub, Sunday N. Okaford,
James A. Ezugwub
a Department of Industrial Chemistry, Evangel University, Akaeze, Ebonyi State
b Department of Pure and Industrial Chemistry, University of Nigeria, Nsukka, Nigeria
c Department of Chemistry, Indian Institute of Technology, Kanpur, India
d Department of Pharmaceutical and Medicinal Chemistry, University of Nigeria, Nsukka, Nigeria
a r t i c l e i n f o
a b s t r a c t
Article history:
The increasing emergence of malaria drug-resistant parasites and the deficiency in effective chemother-
apy for trypanosomiasis represents a huge challenge in infectious disease treatment in tropical regions.
As regards to developing effective antiprotozoal agents, ten new ile-gly dipeptide sulphonamide deriva-
tives were synthesized by condensing compound (10) with (8a-j)using peptide coupling reagents. Com-
pounds11b, 11i and 11j were most potent in clearing Trypanosome brucei in mice with 11b showing com-
parable activity with diminazene aceturate. In the antimalarial study, 11b was the most active compound,
even better than the standard. Molecular docking result suggests good interaction between the reported
compounds and the target protein. The results of haematological analysis, liver and kidney function tests
showed that the compounds had no adverse effect on the blood and organs. Compound 11b stands out
amongst the derivatives haven shown better activity in both the antimalarial and antitrypanosomal assay.
Received 29 September 2020
Revised 18 January 2021
Accepted 23 January 2021
Available online 28 January 2021
Keywords:
Dipeptide
Sulphonamide
Isoleucine
Antitrypanosomal
Antimalaria
© 2021 Elsevier B.V. All rights reserved.
Molecular docking
1. Introduction
deadly if untreated. Melarsoprol (3) and eflornithine (4) are used
for treatment at this stage. However, the few registered drugs are
In the tropical regions of the world, protozoan parasites cause
severe diseases with human African trypanosomiasis (HAT) and
malaria being at the top of the list. The noted toxicities of current
antitrypanosomaldrugsandthe worldwide resurgence of malaria,
accompanied by the springing up of widespread drug-resistant
protozoan parasites motivated this research.HAT otherwise refers
to as sleeping sickness is caused by infections of Trypanosome bru-
cei rhodesiense and T.b. gambiense in humans in Sub-Saharan Africa,
and has remained a disease with no effective treatment. About
500,000 people are infected with sleeping sickness leading to the
death of almost 100,000 people every year [1]. Recent research
progress suggests that a vaccine against the disease is far from be-
ing successful leaving chemotherapy as the only available means of
controlling trypanosomiasis. There are only 4 registered drugs at
present for the treatment of HAT. Pentamidine (1) and suramin (2)
are used for the treatment of the disease in an early stage. How-
ever, in the second stage of the disease, the trypanosomes must
have invaded the central nervous system (CNS) and this become
associated with severe side effects such as poor oral availability,
high cost, toxicity, lack of efficacy and long treatment regimen [2].
Because of this, there is an urgent need to develop new, cheap and
safe alternative chemotherapy against trypanosomiasis.
Malaria is usually caused due to infection with Plasmodium spp
and is one of the most devastating diseases in developing coun-
tries. The World Health Organization (WHO) malaria report in 2017
[3] estimated about 216 million cases of malaria and 445,000
deaths worldwide with 90% of cases and 91% of deaths affect-
ing the African region [4]. Numerous highly active drugs such as
quinine, chloroquine, mefloquine, artesunate and their analogues
are available for the treatment of malaria, but unfortunately, sig-
nificant resistance to almost all these drugs has been developed
even to the ‘‘last resort’’artemisinin-derivatives, first cases of de-
layed clinical efficacy havebeen reported [5]. The main contribu-
tors to widespread resistance in malaria parasite are an inappro-
priate use of antimalaria drugs and the use of monotherapies or
substandard and counterfeit medicine [6]. However, the surfacing
of extensive resistance to the available antimalaria drugs accompa-
nied by a worldwide resurgence of malaria underscores the need
to develop new antimalarial agents that are efficient, safe and syn-
thetically economical [7,8].
∗
Corresponding author.
E-mail addresses: ekohogechi@gmail.com, ogeh15@yahoo.com (O.C. Ekoh).
https://doi.org/10.1016/j.molstruc.2021.130017
0022-2860/© 2021 Elsevier B.V. All rights reserved.

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Sulphonamides have been the centre of drug structures as they
are quite stable and well-tolerated in human beings [9]. Sulfadox-
ine, sulfadiazine and sulfalene are effective malaria drugs that pos-
sesssulphonamide groups attached to a heterocyclic ring. In addi-
tion, several sulphonamide derivatives have been reported as an-
timalarial [10], antibacterial [9], anticancer [11], antidiabetic [12],
antitrypanosomal [13,14] agents. Sulphonamide is an important
pharmacophore and its coupling with other moieties such as pep-
tides always affords new biologically active compounds [15]. Pep-
tides are amongst the most versatile bioactivemolecules and play
crucial roles in the human body and other organisms [16], because
of their good solubility, permeability and bioavailability. Short pep-
tides incorporatesulphonamide or heterocyclic moieties also ex-
hibit numerous biological properties such as carbonic anhydrase
inhibitory [17], chemotactic activity [18], antimicrobial [19,20] and
antioxidant [21] activities. However, Ugwuja et al.,(2019) [22] re-
cently reported Glycine derived dipeptide sulphonamides having
interesting antimalaria and antibacterial properties. Again Val-Val-
derived dipeptides as an antimalaria and antioxidant agent was
also reported [23]. These reports encouraged us to seek a solu-
tion to the problems of multidrug resistance, insecticide-resistant
malaria parasite and lack of adequate chemotherapeutic agent in
managing trypanosomiasis.
(C = O), 141.5, 132.8, 129.4, 127.0 (aromatic carbons), 60.5, 37.3,
24.8, 15.8, 11.4 (aliphatic carbons). HRMS-ESI (m/z) for C12H17 NO4S:
272.0950 (M + H)+, calculated, 272.0951.
2.3. Synthesis of compounds (7a-j)
The procedure according to (Sharma and Soman, 2016)
[24] with little modification was used for the synthesis of these
compounds. 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide hy-
drochloride EDC.HCl (0.53 g, 2.7 mmol), 1-hydroxybenzotriazole
HOBt (0.25 g, 1.84 mmol), triethylamine TEA(2.19 mmol) and
amines (1.84 mmol) were added to a solution of Boc-glycine (8,
0.32 g, 1.84 mmol) in dichloromethane DCM (20 mL) at 0 °C. The
resulting mixture was allowed to stir at room temperature for 16–
19 h and was being monitored using TLC. Upon completion, the
mixture was diluted with DCM, washed with 1 N HCl (50 mL),
5% sodium bicarbonate solution(50 mL) and brine (50 mL). It was
dried using anhydrous sodium sulphate and the solvent was re-
moved under diminished pressure to give the crude product which
was further purified by column chromatography using silica gel
and 5% methanol/dichloromethane. The spectra data are available
as supporting information.
In the interest of discovering new antiprotozoal agents, and
to exploit the synergistic activity coming up from the success-
ful combination of sulphonamide and peptides in drug molecule,
we report here the synthesis of ten new benzenesulphonamide
derivatives incorporating dipeptide moiety with interesting antit-
rypanosomal and antimalarial activity.
2.4. Synthesis of compounds (11a-j)
Compounds (7a-j) were deprotected by stirring in 50% triflu-
oroacetic acid in dichloromethane (TFA/DCM (1:1)) for 40 min
as monitored by TLC, the solvent was then evaporated to give
the substituted acetamides(8a-j). To a solution of compound 6
(1.84 mmol) in DCM (20 mL), EDCl (2.7 mmol), HOBt (1.84 mmol)
and TEA (1.84 mmol) were added at 0 °C and after stirring for
10 min, compounds (8a-j)was also added. The reacting mixture
was further stirred at room temperature for 16 - 19 h as being
monitored using TLC. On completion of the reaction, it was di-
luted with DCM, washed with 1 N HCl (50 mL), 5% sodiumbi-
carbonate solution(50 mL) and brine (50 mL). It was dried us-
ing anhydrous sodium sulphate and the solvent was removed un-
der diminished pressure to give the crude product which was fur-
ther purified by column chromatography using silica gel and 5%
methanol/dichloromethane to give the pure product (11a-j).
2. Material and methods
2.1. Instrumentation
All reagents were of analytical grade and were procured fro-
mAvra, Spectrochem, Aldrich, Merck, SRL, SD fine and Fluka. They
were used without further purification. Silica plates that were used
for thin-layer chromatography were purchased from Avra, spots
were visualized under UV light and in the oven with ninhydrin.
Purification by column chromatography was achieved using Merck
silica gel (60 – 120 mesh). Proton and carbon-13 NMR were carried
outusing Jeol 400 MHZ or 500 MHZin dimethyl sulfoxide(DMSO)-
d6 and chemical shifts presented in part per million (ppm) with
reference to tetramethylsilane. Micro TOF electronspray time of
flight (ESI-TOF) mass spectrometer (AerodynReseachInc.USA) was
used for mass determination, with formate as calibrant. FT-IR spec-
tra of the derivatives were achieved using PerkinElmer spectrum
version 10.03.06 and the bands are given in wavenumber. Melt-
ing points were determined using a glass capillary tube on Stu-
art melting point apparatus and were uncorrected. All experiments
were carried out at Prof. Sandeep Verma Laboratory, Department
of Chemistry, Indian Institute of Technology Kanpur, India.
2.4.1. N-(2-(4-Chlorophenylamino)−2-oxoethyl)−3-methyl-2-
(phenylsulfonamido)pentanamide (11a)
Yield 90%, mp 142–144 °C. FTIR (KBr,cm−1): 3337, 3248 (NH),
–
–
3060 (C H aromatic), 2970, 2928, 2876 (C H aliphatic), 1677, 1640
=
–
(2C O), 1595, 1526, 1493 (C = C), 1388, 1343 (SO2), 1166, (SO2 N),
1204, 1093, 1063 (C N), 729 (C Cl). 1HNMR (400 MHz, DMSO–d6)
δ 9.92 (s, 1H, NH), 8.28 (d, J = 9.8 Hz, 1H, NH), 7.83 (t, J = 9.5 Hz,
1H, NH), 7.74 (d, J = 6.7 Hz, 2H, ArH), 7.52 (dd, J = 29.9, 7.9 Hz,
5H, ArH), 7.33 (d, J = 8.5 Hz, 2H, ArH), 3.64–3.71 (m, 1H, CH), 3.55
(d, J = 9.8 Hz, 2H, CH2), 1.49–1.61 (m, 1H, CH), 1.32–1.49 (m, 1H,
CHa of CH2), 0.94–1.09 (m, 1H, CHb of CH2), 0.72 (dd, J = 17.1,
6.7 Hz, 6H, 2CH3). 13C NMR (DMSO–d6, 100 MHz) ∂: 171.0,167.9,
–
–
=
2.2. Synthesis of 3-methyl-2-(phenylsulphonamido)pentanoic acid
(2C O), 141.4, 138.3, 132.7, 129.2, 127.3, 127.2, 121.0 (aromatic car-
(10)
bons), 61.0, 42.9, 37.2, 24.7, 15.5, 11.0 (aliphatic carbons). HRMS-ESI
(m/z) for C20H24ClN3O4S: 438.1253 (M + H)+, calculated, 438.1249
(M + H)+.
The procedure by Ugwu et al. [14] with little modifications was
adopted for the synthesis of compound 10. The detail of the pro-
cedure is available as supporting information.
2.4.2. N-(2-(4-Bromophenylamino)−2-oxoethyl)−3-methyl-2-
(phenylsulfonamido)pentanamide (11b)
Yield 85%, mp 146–147 °C.FTIR (KBr,cm−1): 3333, 3249 (NH),
3060 (C H aromatic), 2969, 2929, 2875 (C H aliphatic), 1680, 1640
Yield 94%, mp, 148.00–148.60 °C. FTIR (KBr,cm−1): 3295 (NH),
–
–
3066(C H aromatic), 2968, 2936, 2883 (C H aliphatic), 1699 (C = O
of carboxylic acid). 1H NMR (400 MHz, DMSO–d6) δ: 12.54 (s, 1H,
OH), 8.01 (d, J = 9.2 Hz, 1H, NH), 7.72–7.74 (m, 2H, ArH), 7.54 (dt,
J = 23.8, 7.0 Hz, 3H, ArH), 3.48–3.56 (m, 1H, CH), 1.57–1.64 (m, 1H,
CH), 1.25–1.33 (m, 1H, CHa of CH2), 1.00–1.07 (m, 1H, CHb of CH2),
0.68–0.75 (m, 6H, 2CH3). 13C NMR (DMSO–d6, 100 MHz) δ: 172.6
–
–
=
–
(2C O), 1592, 1526, 1490 (C = C), 1343, 1286 (SO2), 1166, (SO2 N),
1213, 1093, 1072 (C N), 594 (C-Br). 1H NMR (400 MHz, DMSO–
–
d6) δ 9.92 (s, 1H, NH), 8.22 (t, J = 5.5 Hz, 1H, NH), 7.84 (d,
J = 8.5 Hz, 1H, NH), 7.72–7.75 (m, 2H, ArH), 7.44–7.55 (m, 7H,
2

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
1
ArH), 3.66 (dd, J = 16.8, 5.8 Hz, 1H, CH), 3.54 (dd, J = 16.5,
6.1 Hz, 2H, CH2), 1.52 (q, J = 11.0 Hz, 1H, CH), 1.34–1.45 (m, 1H,
CHa of CH2), 1.01 (td, J = 13.9, 7.1 Hz, 1H, CHb of CH2), 0.67–
0.82 (m, 6H, 2CH3). 13C NMR (DMSO–d6, 100 MHz) δ 171.0,167.9,
–
–
1214, 1107, 1093, 1075, 1032 (C N, C O). H NMR (400 MHz,
DMSO–d6) δ 9.60 (s, 1H, NH), 8.19 (t, J = 5.8 Hz, 1H, NH), 7.69–7.74
(m, 2H, NH and ArH), 7.42–7.61 (m, 6H, ArH), 6.83 (d, J = 9.2 Hz,
2H,ArH), 3.61–3.66 (m, 4H, OCH3 and CH), 3.50–3.55 (m, 2H, CH2),
1.46–1.61 (m, 1H, CH), 1.33–1.46 (m, 1H, CHa of CH2), 0.99 (td,
J = 14.0, 6.5 Hz, 1H, CHb of CH2), 0.65–0.72 (m, 6H, 2CH3). 13C
=
(2C O), 141.4, 138.3, 132.7, 132.2, 129.2, 127.3, 121.4 (aromatic car-
bons), 61.0, 42.9, 37.2, 24.8, 15.5, 11.0 (aliphatic carbons). HRMS-ESI
(m/z) for C20H24BrN3O4S: 482.0747 (M + H)+, calculated, 482.0744
(M + H)+.
=
NMR (DMSO–d6, 100 MHz) ∂: 170.9, 167.2, (2C O), 155.7, 141.3,
132.7, 132.4, 129.2, 127.2, 121.0, 114.4 (aromatic carbons), 61.1, 55.7,
42.9, 37.5 24.7, 15.2, 11.1 (aliphatic carbons). HRMS-ESI (m/z) for
C21H27N3O5S: 434.1743 (M + H)+, calculated, 434.1744 (M + H)+.
2.4.3. 3-Methyl-N-(2-oxo-2-(p-tolyamino)ethyl)-2-
(phenylsulfonamido)pentanamide (11c)
Yield 79%, mp 140–142 °C. FTIR (KBr,cm−1): 3334, 3244 (NH),
2.4.7. N-(2-(4-Fluorophenylamino)−2-oxoethyl)−3-methyl-2-
(phenylsulfonamido)pentanamide (11 g)
–
–
3060 (C H aromatic), 2967, 2929, 2875 (C H aliphatic), 1671, 1640
Yield80%, mp 151–153 °C.FTIR (KBr,cm−1): 3335, 3246 (NH),
=
–
(2C O), 1597, 1531 (C = C), 1389, 1344 (SO2), 1167, (SO2 N), 1215,
1146, 1065, 1021 (C N). 1H NMR (400 MHz, DMSO–d6)δ 9.66 (s,
–
–
–
3061 (C H aromatic), 2971, 2931, 2877 (C H aliphatic), 1674, 1642
1H, NH), 8.20 (t, J = 5.5 Hz, 1H, NH), 7.85 (d, J = 8.5 Hz, 1H,
NH), 7.73–7.75 (m, 2H, ArH), 7.55 (d, J = 7.3 Hz, 1H, ArH), 7.47
(dd, J = 8.2, 6.4 Hz, 2H, ArH), 7.41 (d, J = 8.5 Hz, 2H, ArH),
7.07 (d, J = 8.5 Hz, 2H, ArH), 3.50–3.68 (m, 3H, CH and CH2),
2.20 (s, 3H, CH3-Ar), 1.51–1.60 (m, 1H, CH), 1.42 (q, J = 10.6 Hz,
=
–
(2C O), 1534, 1512 (C = C), 1389, 1344 (SO2), 1167, (SO2 N), 1221,
1145, 1095 (C N), 1062 (C-F). 1H NMR (400 MHz, DMSO–d6) δ 9.99
–
(s, 1H, NH), 8.28 (t, J = 5.8 Hz, 1H, NH), 7.88 (d, J = 9.2 Hz, 1H,
NH), 7.73 (d, J = 7.3 Hz, 2H, ArH), 7.44–7.57 (m, 5H, ArH), 7.09
(t, J = 8.9 Hz, 2H, ArH), 3.50–3.67 (m, 3H, CH and CH2), 1.56 (m,
1H, CH), 1.40 (m, 1H, CHa of CH2), 0.98–1.04 (m, 1H, CHb of CH2),
0.65–0.73 (m, 6H, 2CH3). 13C NMR (DMSO–d6, 100 MHz) ∂: 170.9,
1H, CHa of CH2), 0.98–1.05 (m, 1H, CHb of CH2), 0.66–0.74 (m,
13
=
6H, 2CH3) C NMR (DMSO–d6, 100 MHz) ∂: 171.0,167.9, (2C O),
141.4, 136.8, 132.7, 129.6, 129.2, 127.2, 119.4 (aromatic carbons),
61.0, 42.9, 37.2, 24.7, 20.9, 15.5, 11.0 (aliphatic carbons). HRMS-
ESI (m/z) for C21H27N3O4S: 418.1809 (M + H)+, calculated, 418.1795
(m + H)+.
=
167.7, (2C O), 141.4, 135.8, 132.7, 129.2, 127.2, 121.2, 121.1, 115.9,
115.7 (aromatic carbons), 61.2, 42.8, 37.2 24.8, 15.5, 11.1 (aliphatic
carbons). HRMS-ESI (m/z) for C20H24FN3O5S: 444.1378 (M+Na)+,
calculated, 444.1364 (M+Na)+
2.4.4. 3-Methyl-N-(2-(naphthalene-1-ylamino)−2-oxoethyl)−2-
(phenylsulfonamido)pentanamide (11d)
2.4.8. N-(2-(3-Chlorophenylamino)−2-oxoethyl)−3-methyl-2-
(phenylsulfonamido)pentanamide (11 h)
Yield 91%, mp 150–152 °C. FTIR (KBr,cm−1): 3329, 3240 (NH),
Yield 90%, mp 142–144 °C. FTIR (KBr,cm−1): 3313, 3239 (NH),
–
–
3061 (C H aromatic), 2964, 2930, 2877 (C H aliphatic), 1675, 1638
–
–
3064 (C H aromatic), 2961, 2929, 2875 (C H aliphatic), 1676, 1640
=
–
(2C O), 1531, 1505 (C = C), 1389, 1340 (SO2), 1167, (SO2 N), 1207,
=
–
(2C O), 1596, 1530, 1481 (C = C), 1388, 1344 (SO2), 1166, (SO2 N),
1094, 1063 (C N). 1H NMR (400 MHz, DMSO–d6) δ 9.80 (s, 1H,
–
–
1214, 1145, 1093, 1075 (C N), 732 (C Cl). 1H NMR (400 MHz,
DMSO–d6)δ 9.98 (s, 1H, NH), 8.22 (t, J = 5.5 Hz, 1H, NH), 7.84 (d,
J = 8.5 Hz, 1H, NH), 7.73 (d, J = 7.3 Hz, 3H, ArH), 7.44–7.55 (m,
3H, ArH), 7.27–7.38 (m, 2H, ArH), 7.05–7.07 (m, 1H, ArH), 3.66 (dd,
J = 16.5, 5.5 Hz, 1H, CH), 3.53 (dd, J = 16.8, 7.0 Hz, 2H, CH2), 1.49–
1.55 (m, 1H, CH), 1.41 (q, J = 6.7 Hz, 1H, CHa of CH2), 0.97–1.04
(m, 1H, CHb of CH2), 0.66–0.74 (m, 6H, 2CH3). 13C NMR (DMSO–d6,
–
NH), 8.32 (d, J = 5.5 Hz, 1H, NH), 8.01 (t, J = 3.4 Hz, 1H, NH),
7.90 (d, J = 9.2 Hz, 2H, ArH), 7.75 (d, J = 7.3 Hz, 3H, ArH), 7.63
(d, J = 7.3 Hz, 1H, ArH), 7.44–7.54 (m, 6H, ArH), 3.54–3.89 (m, 3H,
CH and CH2), 1.38–1.60 (m, 2H, CH and CHa of CH2), 0.96–1.06 (m,
1H, CHb of CH2), 0.65–0.74 (m, 6H, 2CH3). 13C NMR (DMSO–d6,
=
100 MHz) ∂: 171.0,168.5, (2C O), 141.4, 134.2, 133.7, 132.7, 129.2,
128.6, 127.2, 126.6, 126.3, 126.1, 125.8, 123.2,121.9 (aromatic car-
bons), 61.1, 42.9, 37.2, 24.7, 15.5, 11.0 (aliphatic carbons). HRMS-ESI
(m/z) for C24H27N3O4S: 454.1808 (M + H)+, calculated, 454.1795
(M + H)+.
=
100 MHz) ∂: 171.1, 168.2, (2C O), 141.4, 140.8, 133.6, 131.1, 129.2,
127.2, 123.5, 118.9, 117.9 (aromatic carbons), 61.0, 42.9, 37.2 24.7,
15.5, 11.1 (aliphatic carbons). HRMS-ESI (m/z) for C20H24ClN3O4S:
460.1071 (M+Na)+, calculated, 460.1068 (M+Na)+.
2.4.5. 3-Methyl-N-(2-oxo-2-(phenylamino)ethyl)−2-
(phenylsulfonamido)pentanamide (11e)
2.4.9. 3-Methyl-N-(2-oxo-2-(m-tolylamino)ethyl)−2-
(phenylsulfonamido)pentanamide (11i)
Yield 90%, mp 146–148 °C. FTIR (KBr,cm−1): 3330, 3245 (NH),
Yield 76%, mp 144–146 °C.FTIR(KBr,cm−1): 3316, 3236 (NH),
–
–
–
–
3060 (C H aromatic), 2967, 2931, 2875 (C H aliphatic), 1674, 1640
3061 (C H aromatic), 2969, 2929, 2875 (C H aliphatic), 1673, 1638
=
–
=
–
(2C O), 1600, 1537, 1499 (C = C), 1391, 1346 (SO2), 1166, (SO2 N),
(2C O), 1614, 1537, 1489 (C = C), 1389, 1344 (SO2), 1166, (SO2 N),
1
1212, 1145, 1093, 1073 (C N). 1H NMR (400 MHz, DMSO–d6) δ 9.68
–
–
1215, 1145, 1093, 1065 (C N). H NMR (400 MHz, DMSO–d6)δ 9.76
(s, 1H, NH), 8.21 (t, J = 5.8 Hz, 1H, NH), 7.84 (d, J = 8.5 Hz,
1H, NH), 7.73–7.75 (m, 2H, ArH), 7.46–7.56 (m, 5H, ArH), 7.27 (t,
J = 7.9 Hz, 2H, ArH), 7.00 (t, J = 7.3 Hz, 1H, ArH), 3.67 (dd, J = 16.8,
5.8 Hz, 1H, CH), 3.55 (dt, J = 16.5, 4.6 Hz, 2H, CH2), 1.51–1.59 (m,
1H, CH), 1.36–1.46 (m, 1H, CHa of CH2), 0.98–1.05 (m, 1H,CHb of
CH2), 0.66–0.75 (m, 6H, 2CH3). 13C NMR (DMSO–d6, 100 MHz) ∂:
(s, 1H, NH), 8.20 (d, J = 5.5 Hz, 1H, NH), 7.85 (d, J = 8.5 Hz, 1H,
ArH), 7.74 (d, J = 7.3 Hz, 2H, ArH), 7.46–7.54 (m, 3H, ArH), 7.30–
7.36 (m, 2H, ArH), 7.14 (t, J = 7.6 Hz, 1H, NH), 6.82 (d, J = 7.3 Hz,
1H, ArH), 3.63–3.67 (m, 1H, CH), 3.51–3.54 (m, 2H, CH2), 2.23 (s,
3H, CH3-Ar), 1.50–1.58 (m, 1H, CH), 1.36–1.47 (m, 1H, CHa of CH2),
1.01 (dd, J = 22.0, 14.6 Hz, 1H, CHb of CH2), 0.66–0.74 (m, 6H,
13
=
=
171.0, 167.7, (2C O), 141.3, 139.2, 132.7, 129.3, 127.1, 123.8, 119.5
2CH3). C NMR (DMSO–d6, 100 MHz) ∂: 170.9, 167.8, (2C O),
(aromatic carbons), 61.1, 42.9, 37.1 26.1,24.7, 15.2, 11.1 (aliphatic car-
bons). HRMS-ESI (m/z) for C20H25N3O4S: 404.1645 (M + H)+, cal-
culated, 404.1639 (M + H)+.
141.4, 139.2, 138.5, 132.7, 129.2, 129.1, 127.2, 124.5, 120.0, 116.7
(aromatic carbons), 61.1, 42.9, 37.2, 24.8, 21.7, 15.6, 11.1 (aliphatic
carbons). HRMS-ESI (m/z) for C21H27N3O4S: 418.1801 (M + H)+, cal-
culated, 418.1795 (M + H)+.
2.4.6. N-(2-(4-Methoxyphenylamino)−2-oxoethyl)−3-methyl-2-
(phenylsulfonamido)pentanamide (11f)
2.4.10. N-(2-(2,6-Dimethylphenylamino)−2-oxoethyl)−3-methyl-2-
(phenylsulfonamido)pentanamide (11j)
Yield 89%, mp 140–141 °C. FTIR (KBr,cm−1): 3324, 3244 (NH),
3061 (C H aromatic), 2961, 2933, 2877 (C H aliphatic), 1669, 1639
Yield 81%, mp 148–149 °C. FTIR (KBr,cm−1): 3258 (NH), 3064
–
–
=
–
– –
(C H aromatic), 2965, 2930, 2876 (C H aliphatic), 1641, 1594
(2C O), 1599, 1536, 1515 (C = C), 1343, 1252 (SO2), 1166, (SO2 N),
3

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Table 1
Table 2
Physiochemical properties calculation of the new analogs.
Binding free energy (࢞G kcal/mol) of 11a-j.
Cpd. No. HBA HBD NoRB Log p TPSA
MW
Lipinski violation
Antitrypanosomiasis
Antimalaria
11a
11b
11c
11d
11e
11f
4
4
4
4
4
5
4
4
4
4
3
3
3
3
3
3
3
3
3
3
9
3.37
3.58
3.08
4.00
2.78
2.74
2.93
3.41
3.12
3.37
104.37 437.948
104.37 482.399
104.37 417.530
104.37 453.563
104.37 403.503
113.60 433.529
104.37 421.493
104.37 437.948
104.37 417.530
104.37 431.557
0
0
0
0
0
0
0
0
0
0
Compounds
2EWG: Scoring
function-London dG
−7.94
1CET: Scoring
function-London dG
−6.14
9
9
11a
9
11b
−5.54
−6.14
9
11c
−7.32
−6.26
10
9
11d
−7.31
−6.08
11 g
11h
11i
11e
−7.52
−5.57
9
11f
−8.16
−5.89
9
11 g
−7.09
−5.78
11j
9
11h
−5.99
−6.10
11i
−6.43
−5.95
HBA = hydrogen bond acceptor, HBD = hydrogen bond donor, NoRB = number of
rotatable bond, TPSA = topological polar surface area, MW = molecular weight,
logp = octanol-water partition coefficient.
11j
−7.73
−6.29
Native ligand
Standard drug
−6.32
−5.67
−5.27
−5.52
Standard drug: chloroquine for antimalaria and melarsoprol for antitrypanosomal.
=
–
(2C O), 1524, 1466 (C = C), 1381, 1344 (SO2), 1166, (SO2 N), 1225,
1094, 1072, 1044 (C N). 1H NMR (400 MHz, DMSO–d6) δ 9.05 (s,
–
2.6.1. In vivo antitrypanosomal test
1H, NH), 8.27 (t, J = 5.8 Hz, 1H, NH), 7.88 (d, J = 8.5 Hz, 1H, NH),
7.73–7.75 (m, 2H, ArH), 7.51 (dt, J = 28.1, 7.3 Hz, 3H, ArH), 7.01
(s, 3H, ArH), 3.71 (dd, J = 16.5, 6.1 Hz, 1H, CH), 3.51–3.61 (m, 2H,
CH2), 2.08 (s, 6H, 2CH3-Ar), 1.50–1.61 (m, 1H, CH), 1.31–1.42 (m,
The method described and used by Tekaet al [25]for the in
vivoantitrypanosomal study against Trypanosomabrucei with some
modification was employed for the experimental design and treat-
ment of mice. In brief, heavily infected donor mice obtained
from the Department of Veterinary Anatomy, Faculty of Veterinary
Medicine, University of Nigeria, Nsukka were subjected to retro-
orbital puncture through the medial canthus of the eye and blood
was collected and instantly diluted with PBS to serve as inocu-
lum. Healthy albino mice were distributed at random into thirteen
groups of four mice per group, 2000 trypanosomes/mice in 0.2 mL
of blood was given intraperitoneally to all the mice except the ones
in group 13. [26].The animals were allowed to grow parasitaemia
(approximately 107) and the percentage parasitaemia was deter-
mined before the animals began to receive treatment. The synthe-
sized compounds (100 mg/kg body weight) were orally adminis-
tered to animals in groups 1–10, DiminazeneAceturate (7 mg/kg)
was given to the mice in group 11, group12 was infected but not
treated and group 13 was neither infected nor treated. The formu-
lations were given to the groups orally every morning for 7 days
and the degree of parasitaemia was determined by wet blood film
prepared from tail blood at X40 magnification. The number of par-
asites seen was counted using the method described by Herbert
and Lumsden [27] and the results expressed as the log of an abso-
lute number of parasites per mL of blood.
1H, CHa of CH2), 0.97–1.05 (m, 1H, CHb of CH2), 0.64–0.71 (m, 6H,
13
=
2CH3). C NMR (DMSO–d6, 100 MHz) δ:171.1, 167.6 (2C O), 141.3,
135.7, 135.2, 132.7, 129.3, 128.1, 127.2, 126.9 (aromatic carbons),
61.0, 42.4, 37.2, 24.7, 18.6, 15.5, 11.1 (aliphatic carbons). HRMS-ESI
(m/z) for C22H29N3O4S: 432.1956 (M + H)+, calculated 432.1952
(M + H)+.
2.5. In silico studies
2.5.1. Physiochemical evaluation
The physiochemical properties, including molecular weight
(MW), octanol/water partition coefficient (Log P(o/w)), hydrogen
bond donor (HBD), hydrogen bond acceptor (HBA), topological po-
lar surface area (TPSA) and number of rotatable bond (NoRB) of
the new derivatives were calculated using molinspiration and the
drug-likeness was evaluated using Lipinski’s rule of five (Table 1).
2.5.2. Molecular docking study
Molecular docking studies were carried out to have a better un-
derstanding of the interaction of the synthesized compounds at
molecular level with pathogenic organisms. Two receptors were
used for this study: T. brucei farnesyl diphosphate synthase (PDB
Code: 2EWG) for antitrypanosomal study and lactate dehydroge-
nase (PDB ID: 1CET) from Plasmodium falciparum for antimalar-
ial study. The co-crystallized inhibitor for each receptors 1CET
and 2EWG are chloroquine and minodronate respectively. The
3D crystal structures with their co-crystallized ligands for the
drug targets were obtained from protein data bank repository
(https://www.rcsb.org/). The downloaded 3D structures were pre-
pared in Discovery Studio Visualizer 4.1 in which the needed
chains were selected while the multiple ligands and non-protein
parts were deleted. ACD/ChemSketch 2015 version was used to
draw the 2D structures of the synthesized compounds. The com-
pounds were docked against the druggable targets. The binding en-
ergy of the compounds was calculated using London dG scoring
function and results are presented in Table 2.
2.6.2. In vivo antimalarial assay
Samples of chloroquine-sensitive Plasmodium berghei(NK-65)
was obtained from the National Institute of Medical Research
(NIMR), Yaba, Lagos, Nigeria. The method by Peter et al. [28]for
the antiplasmodial assay against Plasmodium berghei infection with
some modifications was adopted for the experimental design and
treatment of mice.In brief, about fifty-two infected mice of both
sexes weighing 18–24 g obtained from the Department of Veteri-
nary Anatomy, Faculty of Veterinary Medicine, University of Nige-
ria, Nsukka were randomly divided intogroups (test groups and
three control groups) of four mice each.The inoculum was prepared
from a donor mouse with rising parasitaemia of about 45%. Five
days after infection, percentage parasitaemia was determined and
animals begin to receive treatment. For each of the synthesized
compound (11a-j), animals received a daily oral dose of 100 mg/kg
body weight for four consecutive days, Artemetherlumefantrine
(4/24 mg/kg body weight) was given to the mice in the positive
control group, the negative control group was not treated while the
other group was not infected. On the fifth day,Giemsa-stained thin
blood smears were prepared from the tail of each animal to deter-
mine parasitaemia and percentage inhibition [29]. The percentage
2.6. Biological studies
Animal use The use of animal for this study was approved by
the University of Nigeria ethical committee for animal use for the
PhD research work of Ekoh Ogechi Chinelo PG/PhD/15/78,254.
4

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Scheme 1. Synthetic route to the new dipeptide sulphonamide derivatives.
of parasite inhibition was estimated using the following equation:%
inhibition = [(A-B)/A] × 100
3. Results and discussion
A corresponds to the average parasitaemia of the untreated
group, B corresponds to the average parasitaemia of the treated
group.
3.1. Chemistry
3-Methyl-2-(phenylsulphonamido)pentanoic acid (10) was syn-
thesized in excellent yield (96.97%) by reactingL-isoleucine (9)
with benzenesulphonyl chloride in the presence of sodium car-
2.6.3.Haematological. analysis
Twenty-four hours following the final treatment, the mice were
slain by cervical dislocation and the blood samples were gath-
ered by heart puncture. The haematological parameters carried out
were packed cell volume (PVC),haemoglobin(HB) and red blood
cell (RBC) count, the blood samples were gathered into EDTA bot-
tles and analysed by automated machine (Automated CBC analyzer:
Sysmex KX-21).
bonate at
0 °C for 4 h.Boc-protected acetamides(7a-j) were
synthesized by reacting commercially available Boc-glycine (5)
with substituted anilines (6a-j) in the presence of 1-ethyl-
3-(3-dimethylaminopropyl)carbodiimide hydrochloride EDCl, 1-
hydroxybenzotriazole HOBT and triethylamine TEA. Compounds
7a-jwere deprotected by the addition of 50% TFA in DCM to give
8a-j which on further reaction with compound (10) in the presence
of peptide coupling reagents EDCl, HOBT and TEA furnished the de-
sired dipeptide sulphonamides (11a-j, scheme 1) which were char-
acterized using FTIR,1H NMR, 13C NMR and high-resolution mass
spectroscopy (HRMS) Scheme 2.
2.6.4. Liver function tests (LFTs)
The standard laboratory procedure [30] was used for liver func-
tion test evaluation. The liver function tests performed with the
blood of the mice fed with the new compounds were Aspartate
Aminotransferase (AST), Alanine Transaminase (ALT) and Alkaline
Phosphatase (ALP)
3.1.1. Spectral characterization
The FTIR spectra of the dipeptide derivatives showed two
–
2.6.5. Renal or kidney function test
strong N H bands between 3337 and 3236 cm−1. The two C = O
Kidney function tests performed in this study were serum urea,
creatinine and uric acid, adopting a standard method [31].
amide carbonyl bands appeared between 1680 and 1594 cm−1. The
–
SO2 N bands appeared between 1167 and 1166. These bands are an
5

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Scheme 2. Isoleucine derived dipeptide sulfonamide derivatives.
indication of successful coupling of the acetamides(8a-j) with ben-
zenesulphonamide(10).
ties [32]. Drug-likeness has also been used as aparameter to pre-
dict the balance amongst the molecular propertiesof a compound
that influences its pharmacodynamics andpharmacokinetic proper-
ties [14] The absorption, distribution, metabolism, and excretion of
the drug can be optimized using the physicochemical parameters.
The physicochemical properties of the synthesized compounds
(11a-j) which are useful in the assessment of drug-likeness are
presented in Table 1. Lipinski’s rule of five helps to evaluate the
bioavailability for oral formulations. Lipinski’s rule states that, in
general, an orally active drug has no more than one violation of the
following criteria [33]: No more than 5 hydrogen bond donors (the
total number of nitrogen–hydrogen and oxygen–hydrogen bonds)
No more than 10 hydrogen bond acceptors (all nitrogen or oxygen
atoms). A violation of more than one parameter may be an indica-
tion of poor bioavailability. Table 1 reveals that compounds 11a-j
are in agreement with Lipinski’s rule of five, therefore all thecom-
pounds reported have a good balance between compound solubil-
ity and its penetration of the lipid bilayers and hence arelikely go-
ing to have good oral bioavailability.
In the 1H NMR spectra of the derivatives, the diagnostic peaks
at 3.51–3.61 ppm due to the interaction of CH2 and NH of glycine,
1.49–1.61 ppm due to the interaction of CH and CH2 of isoleucine
and 6.82–9.99 ppm due to NH protons and aromatic protons are
supportive of the target products formation.
The carbon-13 NMR showed all the peaks expected of success-
ful coupled products. The two C = O peaks appeared from 167.6–
171.1 ppm, the peaks assigned to aromatic carbons appeared be-
tween 155.7 and 114.4 ppm while the peaks assigned to aliphatic
carbons appeared between 61.1–11.0 ppm.
The high-resolution mass spectrometer (HRMS) peak of the
derivatives appeared either as M + H+ or M+Na+ adduct. The re-
sults corresponded with the calculated values. The spectra used for
the characterization of the new compounds are available as sup-
porting materials
3.1.1.1. In silico studies. Physicochemical properties Physicochem-
ical properties of compounds have been used by Medicinal
Chemists long ago to predict or estimate pharmacokinetic proper-
Molecular Docking Binding free energy is an indication of
the binding affinity of a ligand on the protein. It measures the
6

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Fig. 1. Examples of commercial antitrypanosomal agents.
Fig. 2. Stereoview of compound11f in the binding cavity of 2EWG.
inhibition of the protein or enzyme in the presence of both the
inhibitor. The free energy of binding is given as the dissociation
constant Kd which is the ratio of the concentration of the prod-
ucts (complexes) to reactants (protein and ligand). It shows the ex-
tent to which biological macromolecules interact with each other
or with various small molecules through noncovalent interactions
to form a specific complex. The higher the binding affinity, we will
expect a interactions between the biological macromolecules (drug
targets) and the ligands. These effective interactions will result in
the molecules eliciting biological activities. These predictions are
made through docking calculations.
out for further studies to enable us to understand its mechanism of
action. Likewise, there was no significant difference in the binding
affinity of the compounds and those of the standards against1CET.
Compound 11j has been chosen to represent the group for further
studies. Fig. 1
Figs. 2 and 5 show the stereo views of compounds 11f and
11j in the binding cavities of 2EWG and 1CET respectively. The
compounds fitted well into the binding cavities of the respective
targets thereby making extensive chemical interactions with their
amino acid residues. Fig. 3 shows the 2D representation of bind-
ing interactions of compound 11f with the amino acid residues
of 2EWG while Fig. 6 shows the 2D representation of the bind-
ing interactions of compound 11j with the amino acid residues of
1CET. There were numerous effective and significant chemical in-
teractions observed leading to high binding affinities of these com-
pounds with these targets. The detailed chemical interactions lead-
ing to high binding affinities are shown in Table 3 for 11f-2EWG
and Table 4 for 11j-1CET interactions. To understand the nature of
the atoms of the compounds interacting with various amino acid
residues, Figs. 4 and 7 show the numbering of compounds 11f and
11j respectively using Chem Draw.
Table 2 shows the binding free energy (kcal/mol) of the synthe-
sized compounds with the drug targets used, namely; T. brucei far-
nesyl diphosphate synthase complexed with minodronate (2EWG)
and Plasmodium falciparum lactate dehydrogenase complexed with
chloroquine (1CET). The compounds showed a reasonable binding
affinity with the targets. The molecular docking studies against
2EWG target showed that the compounds had more binding affin-
itythan the native ligand (minodronate) and the standard drug
(melarsoprol) for the treatment of trypanosomiasis. Compound 11f
showed the highest binding affinity with and hence was singled
7

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Fig. 3. 2-Dimensional representation of binding interactions of compound 11f with the amino acid residues of 2EWG.
Table 3
Summary of interactions between compound 11f and 2EWG.
˚
Receptor
Ligand
Distance (A)
Category
Typeof interaction
ARG50
O-10
2.42
2.41
1.72
1.93
2.65
2.85
2.44
3.68
2.78
3.33
5.45
5.48
4.15
5.11
4.74
Hydrogen Bond
Hydrogen Bond
Hydrogen Bond
Hydrogen Bond
Hydrogen Bond
Hydrogen Bond
Hydrogen Bond
Electrostatic
Conventional Hydrogen Bond
Conventional Hydrogen Bond
Conventional Hydrogen Bond
Conventional Hydrogen Bond
Conventional Hydrogen Bond
Carbon Hydrogen Bond
Carbon Hydrogen Bond
Pi-Cation
GLN96
O-10
ASP107
GLN252
ASP255
ASP175
CYS279
ARG112
LYS212
ASP273
PHE251
LEU100
LYS212
TYR216
LYS278
O-21
O-17
HN-22
H1–30
H3–30
Phenyl
methoxy phenyl
methoxy phenyl
S-7
Hydrogen Bond;Electrostatic
Electrostatic
Pi-Cation;Pi-Donor Hydrogen Bond
Pi-Anion
Other
Pi-Sulfur
C-15
Hydrophobic
Alkyl
methoxy phenyl
C-14
Hydrophobic
Alkyl
Hydrophobic
Pi-Alkyl
methoxy phenyl
Hydrophobic
Pi-Alkyl
3.2. Biological studies
3.2.1. In vivo antitrypanosomal activities
mals treated with 11b, 11i and 11j in which there was a continuous
reduction in the activity of trypanosomes and parasitaemia level
from day 1 to the final day of treatment. Compound 11b which
showed the best activity amongst the groups treated with the syn-
thesized derivatives has a comparable activity with the standard
drug (diminazeneaceturate).
Table 5 shows the in vivoantitrypanosomal activity of com-
pounds 11a-j on mice infected with T. bruccei. The result of the
study reveals that there were variations in the parasitaemia lev-
els of all the groups treated with the synthesized compounds and
diminazeneaceturate. The parasitaemia levels in the treated groups
are relatively low as compared to the infected untreated control.
However, compounds 11a-j displayed mild to moderate in vivo an-
titrypanosomal activity. The best results were obtained with ani-
3.2.2. In vivo antimalarial activities
The method by De Souza et al. [34] was employed for the
interpretation of the in vivo antimalarial activity of the syn-
thesized compounds against P. berghei (NK-65). Compounds that
8

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Table 4
Summary of interactions between compound 11j and 2EWG.
˚
Receptor
Ligand
Distance (A)
Category
Typeof interaction
GLY99
THR101
ASP53
THR101
PHE100
ALA98
ILE54
O-21
2.09
2.75
2.38
2.97
5.96
3.64
4.56
4.86
5.38
5.00
5.02
4.04
4.29
4.43
Hydrogen Bond
Hydrogen Bond
Hydrogen Bond
Hydrogen Bond
Hydrophobic
Hydrophobic
Hydrophobic
Hydrophobic
Hydrophobic
Hydrophobic
Hydrophobic
Hydrophobic
Hydrophobic
Hydrophobic
Conventional Hydrogen Bond
O-9
Conventional Hydrogen Bond
NH-18
Conventional Hydrogen Bond
O-10
Carbon Hydrogen Bond
Pi-Pi Stacked
Alkyl
Phenyl
C-30
C-29
Alkyl
ILE119
VAL26
ILE54
C −29
Alkyl
C-30
Alkyl
C-30
Alkyl
PHE52
ILE54
C-30
Pi-Alkyl
Pi-Alkyl
Pi-Alkyl
Pi-Alkyl
2,6-dimethylphenyl
2,6-dimethylphenyl
2,6-dimethylphenyl
ALA98
ILE119
Fig. 4. Compound 11f showing the atom numbering.
Fig. 5. Stereoview of compound 11j in the binding cavity of 1CET.
9

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Fig. 6. 2-Dimensional representation of binding interactions of compound 11j with the amino acid residues of 1CET.
Fig. 7. Compound 11j showing the atom numbering.
reduced parasitaemia by 40% were considered active, whereas
tivity with artemether-lumenfantrine(79.89%). These compounds
should be considered for further studies because of their promising
activities.
those that reduced parasitaemia by 30–40% and by less than 30%
were considered partially active and inactive respectively. Apart
from 11c, all the compounds including the control drug reduced
the parasitaemia level by at least 40% (Table 6). However, com-
paring the compounds ability to inhibit parasitaemia to that of
the control drug, compounds 11b, 11aand 11i (%inhibition = 81.25,
77.80 and 76.23% respectively) showed better or comparable ac-
3.2.3. Haematological analysis
Evaluation of the complete blood count provides huge in-
formation on the haematological status in disease condition
[35]. Anaemia is usually assessed by evaluating the packed cell
10

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
Table 5
Table 9
Effects of compounds 11a-j on parasite count in mice
Kidney function test.
infected with T. brucei.
Compounds
Urea (mg/dl)
Creatinine(mg/dl)
Uric acid (mg/dl)
Parasitaemia level (log number/mL)
7d
12.4
11.7
12.0
12.4
0.6
0.6
0.5
0.5
3.5
3.3
3.4
3.2
Compounds
Day 1
Day 3
Day 5
Day 7
7i
11a
11b
11c
11d
11e
11f
11 g
11h
11i
6.70
7.12
7.11
7.20
6.93
7.01
6.99
7.22
7.01
7.00
7.23
6.98
6.77
4.50
8.32
7.06
5.30
7.01
6.29
7.36
5.23
6.09
5.84
7.29
5.31
3.05
6.38
4.39
4.33
5.20
3.00
5.00
3.42
3.25
2.37
8.45
5.91
0.60
7.82
5.05
3.10
5.00
5.04
4.49
2.01
2.23
0.52
8.88
7j
Control
3.2.5. Kidney function test
These are common laboratory tests used to evaluate how well
the kidneys are functioning. It is observed from Table 9 that there
is no significant change in the serum level of urea, creatinine and
uric acid of mice fed with 100 mg/kg of the reported derivatives
when compared with the control.
11j
DA
NTC
DA: Diminazeneaceturate, NTC: Non treated control.
4. Conclusion
Table 6
Ten new Ile-gly dipeptide sulphonamide derivatives have
been successfully synthesized using an efficient, versatile and
ecofriendly approach. The structures of the synthesized compounds
are consistent with spectral data. All the compounds were investi-
gated for their in vivo antitrypanosomal and antimalarial activities
in mice. The synthesized compounds showed mild to moderate an-
titrypanosomal activity except for compound 11b that had a com-
parable activity with diminazene aceturate at day 7 of the treat-
ment period. In the antimalarial activity study, compounds11b, 11f
and11i showed good antimalarial activity with percentage inhibi-
tion of parasite growth in the range of 73.63–77.80% comparable
with artemether/lumenfantrine (79.77%). The results of the haema-
tological 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 predic-
tions indicate that the compounds would not pose oral bioavail-
ability, transport and permeability problems if developed further
to drug molecules The molecular docking studies showed good in-
teraction between the synthesized compounds and the protein tar-
gets for antitrypanosomal and antimalarial activities. The synthe-
sized derivatives are promising drug candidates for trypanosomia-
sis and malaria. Furthermore, Compound 11b stands out amongst
the derivatives having shown good activity in both the antimalarial
and antitrypanosomal assay.
Percentage inhibition of P. berghei parasite in
mice.
Compounds
% Inhibition
11a
77.80
81.25
31.33
50.68
57.00
73.63
40.67
52.82
76.23
54.39
79.77
–
11b
11c
11d
11e
11f
11 g
11h
11i
11j
Arte lum
NTC
NIC
–
Arte lum: Artemetherlumefantrine, NTC: Non
treated control, NIC: Non infected control.
Table 7
Haematological analysis after treatment.
Compound
PCV (%)
HB (g/dL)
RBC (× 106/μL)
11a
36.9
37.6
37.9
38.2
11.4
12.4
12.0
12.8
7.2
7.4
6.8
7.6
11b
11i
Control
Declaration of Competing Interest
Table 8
Liver function tests.
The authors declare that there is no conflict of interest
compounds
AST (μL)
ALT (μL)
ALP (μL)
CRediT authorship contribution statement
11a
57.6
56.2
56.9
57.4
20.0
19.4
20.3
20.3
0.79
0.80
0.84
0.82
11b
11i
Ogechi C. Ekoh: Conceptualization, Methodology, Validation,
Control
Formal analysis, Investigation, Resources, Writing
- original
draft, Visualization, Project administration, Funding acquisition.
Uchechukwu C. Okoro: Conceptualization, Methodology, Writing -
review & editing, Supervision. Rafat Ali: Investigation, Writing - re-
view & editing, Supervision. David I. Ugwu: Investigation, Writing
- review & editing, Project administration. Sunday N. Okafor: Soft-
ware. James A. Ezugwu: Formal analysis.
volume (PCV), haemoglobin (HB), and red blood cell (RBC) count in
malaria patients [36]. Table 7 shows the haematological parameters
of P. berghei infected mice after treatment. The result revealed that
there is no significant decrease in the parameters of the treated
groups when compared with the control.
Acknowledgements
3.2.4. Liver function test
Liver function test is a group of blood tests that are used to
check how well the liver is working. They detect inflammation and
damage to the liver. The liver function test carried out in this re-
search is presented in Table 8. The result of this study shows that
the administration of 100 mg/kg of the synthesized compounds in
mice did not cause a significant increase or decrease in the serum
AST,ALT and ALP when compared with the control.
Authors gratefully acknowledge the assistance of Prof. Sandeep
Verma, Department of Chemistry, Indian Institute of Technology,
Kanpur, India in providing the facilities for synthesis and spec-
tral characterization. We are also thankful to the Department of
Veterinary Anatomy, Faculty of Veterinary Medicine, University of
Nigeria, Nsukka for the in vivo antitrypanosomal and antimalarial
study.
11

O.C. Ekoh, U.C. Okoro, R. Ali et al.
Journal of Molecular Structure 1231 (2021) 130017
References
[20] N. Bug˘day, F.Z. Küçükbay, E. Apohan, et al., Synthesis and evaluation of novel
benzimidazole conjugates incorporating amino acids and dipeptide moieties,
Lett. Org. Chem. 14 (2017) 198–206.
[21] A. Sanchez, A. Vazquez, Bioactive peptides: a review, Food Qual. Saf. 1 (2017)
29–46.
[1] C. Burri, S. Nkunku, A. Merolle, et al., Efficacy of new,concise schedule for
melarsoprolin treatment of sleeping sickness caused by Trypanosomabru-
ceigambiense:a randomized trial, Lancet 355 (2000) 1419.
[22] D.I. Ugwuja, U.C. Okoro, S.S. Soman, et al., New peptide derived antimalarial
and antimicrobial agents bearing sulphoinamide moiety, Journal of Enzy. Inhibi
and Med. Chem. 34 (1) (2019) 1388–1399.
[2] M. Witschel, M. Rottmann, M. Kaiser, R. Brun, Agrochemicals against malaria,
sleeping sickness, leishmaniasis and Chagas disease, PloSNeglTrop Dis 6 (10)
(2012) el805.
[23] J.A. Ezugwu, U.C. Okoro, M.A. Ezeokonkwo, et al., Synthesis and biological
evaluation of Val–Val dipeptide–sulfonamide conjugates, Arch Pharma (2020)
e2000074.
[3] WHO. World malaria report. Geneva: World Health Organization; (2017)
[4] WHO. World malaria report. Geneva: World Health Organization; (2016)
[5] H. Noedl, L. Ghanthap, Y. Se, D. Socheat, et al., Artemisinin resistance in Cam-
bodia, Trop.Med.Int. Health 12 (2007) 69.
[6] World Health Organization (WHO). Global report on antimalarial drug effi-
cacy and drug resistance: 2000–2010. Available online: http://www.who.int/
malaria/en (accessed on 10 March 2011)
[24] R. Sharma, S.S. Soman, Design and synthesis of novel diamide derivatives of
glycine as antihyperglycemic agents, Synth Commun 46 (1) (2016) 307–317.
[25] F. Teka, T. Getachew, S. Workineh, Evaluation of in vivo antitrypanosomal activ-
ity of crude extracts of Artemisia abyssinica against a trypanosome congolense
isolate, BMC Comple. and Alterna. Med. 14 (2014) 117.
[7] K. Kaur, M. Jain, R.P. Reddy, R. Jain, Quinolines and structurally related hetero-
cycles as antimalarials, Eur. J. Med. Chem. 45 (2010) 3245–3264.
[8] C. Biot, K. Chibale, Novel approaches to antimalarial drug discovery, Infect. Dis-
ord.-Drug Targets 6 (2006) 173–204.
[26] A.C. Ene, S.E. Atawodi, D.A. Ameh, et al., Antitrypanosomal effects of petroleum
ether, chloroform and methanol extracts of Artemisia maciveraeLinn, Indian
expBiol 47 (9) (2009) 81–986.
[27] W.J. Herbert, .WH.R Lumsden, Trypanosome brucei. A rapid matching method
for estimating the host’s Parasitaemia, ExpParasitol. 40 (1976) 423–431.
[28] 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 schizon-
tocidal activity, Ann Trop Med Parasitol 69 (1) (1975) 55–71.
[29] P. Waako, B. Gumede, P. Smith, P. Folb, The in vitro and in vivo antimalarial
activity of Cardiospermumhalicacabum L. and MomordicafoetidaSchumch Et.
Thonn, J Ethno Pharmacol 99 (2005) 137–143.
[9] P.M. Shet, V.P. Vaidya, K.M. Mahadevan, et al., Synthesis, Characterisation and
antimicrobial Studies of novel Sulphonamides containing substituted naphtho-
furoyl group, Res. J. Chem. Sci. 3 (1) (2013) 15–20.
[10] B.D. Mistry, K.R. Desai, S.M Intwala, Synthesis of novel sulphonamidesas poten-
tial antibacterial, antifungal and antimalarial agent, Indian J.Chem. 54B (2015)
134.
[11] M.M. Ghorab, M.S. Bashandy, M.S. Alsaid, Novel thiophene derivatives with
sulphonamide, isoxazole, benzothiazole, quinoline and anthracene moieties as
potential anticancer agents, Acta Pharm. 64 (2014) 431.
[12] S. Jallow, A. Alabi, R. Sarge-Njie, et al., Virological Response to Highly Ac-
tive Antiretroviral Therapy In Patients Infected With Human Immunodeficiency
Virus Type2 (Hiv-2) and in Patients Dually Infected With Hiv-1And Hiv-2 in
the Gambia And Emergence Of Drug-resistant Variants, J. Clin. Microbiol. 47
(7) (2009) 2200–2208 PMID:19420165, doi:10.1128/JCM.01654-08.
[13] V.M. Papadopoulou, W.D. Bloomer, H.S. Rosenzweig, et al., Novel 3-ni-
tro-1H-1,2,4-triazole-based amides and sulfonamides as potential antitry-
panosomal agents, J. Med. Chem. 55 (11) (2012) 5554–5565.
[30] S. Reitman, S. Frankel, A colorimetioc method for the determination of serum
glutamic oxaloacetic and glutamic pyruvic transaminase, Am J ClinPathol 28
(1957) 56–63.
[31] A. Kaplan, L.L Teeng, in: W.R. Faulkner, S. Meits (Eds.), Selected methods of
clinical chemistry, 9, Washington, DC, AACC, 1982, pp. 357–363.
[32] M. Karlgren, C.A.S Bergstrom, How physicochemical propertiesof drug affect
their metabolism and clearance, in: AGE Wilson (Ed.), New horizons in predic-
tive drug metabolismand pharmacokinetics, 49, Cambridge, UK, Royal Society
of Chemistry, 2015, pp. 1–26.
[33] C.A. Lipinski, F. Lombardo, B.W. Dominy, P.J. Feeney, Experimental and compu-
tational approaches to estimate solubility and permeability in drug discovery
and development settings, Adv. Drug Deliv. Rev. 46 (1–3) (March 2001) 3–26,
doi:10.1016/S0169-409X(00)00129-0.
[34] N.B. De Souza, I.M. Andrade, et al., Blood shizonticidal activities of phenazines
and naphthoquinoidal compounds againstPlasmodiumfalciparum in vitro and
in mice malaria studies, Mem Inst Oswaldo Cruz 109 (2014) 546.
[35] T.B. Lathia, R. Joshi, Can haematological parameters discriminate malaria from
nonmalarious acute febrile illness in the tropics? Indian J. of Med.Sci. 58 (6)
(2004) 239–244.
[14] D.I. Ugwu, U.C. Okoro, N.K. Mishra, Synthesis, characterization and in vitro an-
titrypanosomal activities of new carboxamides bearing quinoline moiety, PLoS
ONE 13 (2018) 0191234 e.
[15] P. Jain, C. Sarravanan, S.K. Singh, (Sulphonamides: deserving class as MMP in-
hibitors, Eur. J. Med. Chem. 60 (2013) 89–100.
[16] T. Day, S.A Greenfield, Bioactivity of a peptide derived fromacetylcholinesterase
in hippocampal organotypiccultures, Exp Brain Res 155 (2004) 500.
[17] B. Nesrin, K. Zehra, K. Hasan, et al., Synthesis of novel dipeptide sulfonamide
conjugates with effective carbonic anhydrase I,II,IX and XII inhibitory proper-
ties, Bioorg. Chem. 81 (2018) 311–318.
[36] C.C. Mojisola, A. Akhere, M.A Omonkhua Olusegun, Effects ofAnogeissusleio-
carpusonhaematological parameters of mice infected withPlasmodiumberghei,
J Plant Stud 2 (2) (2013) 13–21.
[18] A. Mollica, M.P. Paradisi, K. Varani, et al., Chemotactic peptides: fMLF-OMe
analogues incorporating proline–methionine chimeras as N-terminal residue,
Bioorg. Med. Chem. 14 (2006) 2253–2265.
[19] M.A. Ibrahim, S.S. Panda, A.A. Oliferenko, et al., Macrocyclic peptidomimetics
with antimicrobial activity: synthesis, bioassay and molecularmodeling studies,
Org. Biomol. Chem. 13 (2015) 9492–9503.
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