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Antiplasmodial effect and sub-acute toxicity of
alkaloid, flavonoid and phenolic extracts of Sida
acuta leaf on Plasmodium berghei-infected animals
Daniel Anuoluwa Adesina , Sherifat Funmilola Adefolalu , Ali Audu Jigam &
Bashir Lawal
To cite this article: Daniel Anuoluwa Adesina , Sherifat Funmilola Adefolalu , Ali Audu Jigam &
Bashir Lawal (2020) Antiplasmodial effect and sub-acute toxicity of alkaloid, flavonoid and phenolic
extracts of Sida�acuta leaf on Plasmodium�berghei-infected animals, Journal of Taibah University
for Science, 14:1, 943-953, DOI: 10.1080/16583655.2020.1790912
To link to this article: https://doi.org/10.1080/16583655.2020.1790912
© 2020 The Author(s). Published by Informa
UK Limited, trading as Taylor & Francis
Group
Published online: 13 Jul 2020.
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JOURNAL OF TAIBAH UNIVERSITY FOR SCIENCE
2020, VOL. 14, NO. 1, 943–953
https://doi.org/10.1080/16583655.2020.1790912
Antiplasmodial effect and sub-acute toxicity of alkaloid, flavonoid and
phenolic extracts of Sida acuta leaf on Plasmodium berghei-infected animals
Daniel Anuoluwa Adesina a, Sherifat Funmilola Adefolalu a, Ali Audu Jigam aand Bashir Lawal a,b
aDepartment of Biochemistry, Federal University of Technology Minna, Minna, Nigeria; bPhD Program for Cancer Molecular Biology and
Drug Discovery, Taipei Medical University and Academia Sinica, Taipei, Taiwan
ABSTRACT
The antiplasmodial and safety profile of alkaloid, flavonoid and phenol extracts of Sida acuta
(300 and 600 mg/kgbw) on hepatic and renal integrity of rats were investigated. Alkaloid,
flavonoid and phenol extracts produce parasitaemia suppression of 50.83%, 33.50% and 64.64%,
respectively. Sub-chronic administration of the phytochemicals caused marked (p<0.05) dose-
dependent increase in erythrocytic and leucocytic indices while serum ALP, AST, ALT, albumin,
urea and creatinine concentrations compared well (p>0.05) with the controls. Total proteins,
sodium and chloride levels were altered in rats treated with 300 mg/kgbw of the phytochemicals.
The integrity of hepatocytes and renal cells increases with increase extract concentrations and no
degenerative changes were observed in all treatment groups. However, flavonoid extract caused
severe renal vacuolation at 300 mg/kgbw which cleared out at 600 mg/kgbw. Conclusively, phe-
nol exhibited higher antiplasmodial activities and safety profile and thus could be considered a
potential candidate for the development of a new drug.
ARTICLE HISTORY
Received 28 November 2019
Revised 5 May 2020
Accepted 17 June 2020
KEYWORDS
Sida acuta; alkaloid;
flavonoid; phenolic;
antiplasmodial activity;
sub-acute toxicity
1. Introduction
Malaria is a public health and life-threatening infectious
disease caused by a parasitic protozoan of the Plasmod-
ium genus and transmitted via the bite of Anopheles
mosquito [1]. Malaria is one of the major tropical dis-
eases with a global estimate of 219 million incidence
and ∼445,000 malarial deaths in 2017 [2]. The burden
of malaria is the greatest in Africa, representing 90%
of the estimated malaria-associated death [3]. It is the
greatest cause of mortality and hospitalization among
children <5 years of age in Sub-sahara Africa [4]. The
malaria incidence or burden is increasing rapidly, due to
the increase in parasite resistance to conventional drugs
[5] and side effects associated with conventional anti-
malarial drugs [6]. This global concern has, therefore,
called for an urgent need to search for new antimalarial
agents particularly from medicinal plant extracts which
are the potential source of new affordable and effective
antimalarial agents. This idea originated from the his-
torical use of medicinal plants in malaria treatment by
traditional healers. Moreover, artemisinin and quinine,
standardized antimalarial drugs, were derived from Cin-
chona species and Artemisia annua, respectively, thus
suggesting that other medicinal plant could also serve
as reservoir for new effective antimalarial agents [7].
A number of plants have been reportedly used in the
traditional management of malaria and have received
ample scientific validation in both in vivo and in vitro
models [8]. One of such plants is Sida acuta.
Sida acuta Burm.f (Malvaceae) is one of those plants
commonly used by traditional healers for the man-
agement of some health problems [9]. This plant is a
branched, erect, perennial herb of about 1.5 m height
[10]. S. acuta has been reportedly used as an antipyretic,
stomachic, antipyretic and diaphoretic. It is used as a
tonic and astringent and for treatments of urinary, bile,
hepatic and nervous disorders [11,12]. Pharmacologi-
cally, S. acuta has been reported for inhibitory activi-
ties against Anopheles stephensi [13], antioxidants [14],
antiulcer [15], wound healing [16] cardioprotective [17],
antidiabetic [18] and antibacterial activities [19,20]. Pre-
vious studies on the crude extract of S. acuta have
shown markedly antimalarial activities in both in vitro
(IC50 0.05 µg/ml compared to <0.042 µg/ml for chloro-
quine) and in vivo models [21,22].
Phytoconstituents are natural secondary metabo-
lites in plants, responsible for the plant odour, colour
and its therapeutic potencies. Most of the phytocon-
stituents have antioxidant, antimicrobial and antimalar-
ial properties and they protect cells against oxidative
stress damage [23,24]. Screening of phytochemicals
from medicinal plants is, therefore, an important tool in
identifying active metabolites of medicinal and indus-
trial application. Previous phytochemical studies of S.
CONTACT Bashir Lawal bashirlawal12@gmail.com Department of Biochemistry, Federal University of Technology Minna, Nigeria, PhD Program
for Cancer Molecular Biology and Drug Discovery, Taipei Medical University and Academia Sinica, Taipei 111, Taiwan
© 2020 The Author(s). Published by Informa UK Limited, trading as Taylor& Francis Group
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted
use, distribution, and reproduction in any medium, provided the original work is properly cited.
944 D. A. ADESINA ET AL.
acuta have reported the presence of alkaloids (vasicine,
cryptolepine and ephedrine), phenolic compounds
(scopoletin, evofolin-A and B, 4-ketopinoresinol and
loliolide), polyphenol, flavonoids, coumarins, sapono-
sides, steroids (ecdysterone, β-sitosterol, stigmasterol,
ampesterol) and tannins as the major bioactive com-
ponents of the plant [14,19,25]. The present study
investigated the antimalarial potencies of alkaloid,
flavonoid and phenol extracts from Sida acuta against
P. bergehi–infected mice and also evaluated their
safety profile of on hepatic and renal integrity in
albino rats.
2. Materials and methods
2.1. Chemicals and reagents
Organic solvents (analytical grade) used for the extrac-
tion of the plant material (methanol, ethanol and n-
hexane) were products of Sigma Chemical Co St. Louis
M.O (USA). All analyses were performed with com-
mercial kits from Randox Laboratories Limited, United
Kingdom and Quimica Clinica Applicada, Spain. All
other chemicals used were also of analytical grade and
obtained from the Department of Biochemistry, Federal
University of Technology, Minna.
2.2. Plant materials
Sida acuta (broom weed) plant was randomly collected
in the environment of Zumba, Shiroro, Niger State. The
leaf was air-dried (36.5°C) for six days and was ground
into fine powder after which it was sieved and packaged
in air-tight containers.
2.3. Plasmodium parasite
P. berghei NK 65 chloroquine-sensitive strain was
obtained from the Institute for Medical Research and
Training, University of Ibadan, Nigeria and was main-
tained in the laboratory by serial passage in mice [3].
2.4. Experimental animals
Healthy albino animals (mice and rats) were procured
from Animal Breeding unit of Ahmadu Bello University
Zaria, Nigeria. The animals were maintained under stan-
dard laboratory conditions with access to commercial
feed pellets (growers) and water ad libitum. The princi-
ples, governing the use of laboratory animals as laid out
by the Federal University of Technology, Minna Com-
mittee on Ethics for Medical and Scientific Research
and also existing internationally accepted principles for
laboratory animal use and care as contained in the
Canadian Council on Animal Care Guidelines and Pro-
tocol Review and international standard (NIH Publica-
tion No. 85-23, 1985) were duly observed. The ani-
mals were fasted 12 h before the commencement of
any study.
2.5. Sample extraction
2.5.1. Alkaloid extraction
The extraction of the alkaloid was done by the continu-
ous extraction method using the Soxhlet apparatus, as
described by Gonzales and Tolentino [26]. Sida acuta-
powdered leaf (350 g) was moistened with 650 mL of
95% ethanol and alkalinified with 525 mL ammonia.
Following the overnight maceration, the sample was
extracted with ethanol; the extract was filtered and
concentrated in a water bath at 60°C. The crude alka-
loid was acidified with 1.0 N hydrochloric acid (30 mL),
filtered and the filtrate was alkalinified with ammo-
nia (40 mL), followed by chloroform (600 mL) partition-
ing. Extraction was continued with chloroform until the
last chloroform extract tested negative to Dragendorff’s
reagent. The extract was weighed and the percentage
yield (0.56%) was documented.
2.5.2. Flavonoid extraction
Flavonoid extraction was carried out according to the
method of Yahaya [27]. Sida acuta (175 g)-powdered
leaf sample was defatted with n-Hexane (250 mL) using
a Soxhlet extractor. The extraction was carried out for
six hours at 65°C. After the extraction, the thimbles were
driedinanovenat50
°C. The extracted marc was further
extracted with methanol (250 mL). The extract obtained
was then evaporated (40°C) using water bath to yield
concentrated flavonoids (6.78%).
2.5.3. Phenolic extraction
Dried and powdered leaves of Sida acuta (100 g) were
solubilized in methanol:water 240:60 mL (80:20, v/v)
and homogenized at room temperature (36.5°C) for 1 h
30 min. The solution was filtered with Whatman filter
paper, using a separatory funnel under vacuum. The fil-
trate was then evaporated using a water bath at 40°C to
obtain the free phenol (5.24%) extract [28].
2.6. Acute toxicity test
The acute toxicity studies were conducted as per the
Organization for Economic Cooperation and Develop-
ment (OECD) guidelines 425 [29]. Five female mice (6–8
weeks) were fasted for three hours and orally given a
dose of 5000 mg/kg bw of the phytochemical extract.
The mice were observed for physical signs first for three
hours and further for 72 h. The experiment was termi-
nated after 2 weeks of observations.
JOURNAL OF TAIBAH UNIVERSITY FOR SCIENCE 945
2.7. Antiplasmodial screening (curative test)
Evaluation of the curative potential of the extract was
done using the method described by Ryley and Peters
[30]. On the first day (D0), standard inoculums of about
1×107P. berghei-infected red blood cells were injected
intraperitoneally into the mice. Seventy-two hours later,
eight groups (consisting of three mice each) were set
up. Groups A and B were treated with 300 mg/kg bw and
600 mg/kg bw of the alkaloid extract, groups C and D
were treated with 300 mg/kg bw and 600mg/kg bw of
the phenol extract, while groups E and F were treated
with 300 mg/kg bw and 600 mg/kg bw of the flavonoid
extract, respectively. Chloroquine (25 mg/kg bw) was
given to the positive control group (group G) and dis-
tilled water to the negative control group (group H). The
drugs/extracts were given once daily for 5 days. Each
day of treatment, blood samples were collected from
the tail prick of each mouse and thin smears were pre-
pared and stained with 10% Giemsa solution. The dried
slides were examined under the microscope at x100
magnification and parasitaemia level was determined
by counting the parasites in a field [23]. Percentage par-
asitaemia and percentage suppression were calculated
using the following formula.
Calculation
%Suppression =
Parasitaemia in
negative control
−parasitaemia in
treated group
Parasitaemia in
negative control
×100
% Parasitaemia =
Number of parasitized
red blood cell
Total red
blood cell counted
×100
2.8. Sub-acute toxicity
Sub-acute toxicity was carried out according to OECD
guideline 410 [31]. Both sexes of the albino rats
(100–170 g) were divided into seven groups (5 animals
per group). 300 mg/kg bw and 600 mg/kg bw of each
phytochemical extract were administered orally to six
groups and the seventh group (positive control) was
given distilled water. All treatments were administered
orally with the aid of intubation cannula once daily for
21 days. All the animals were observed for clinical signs
at the time of onset, duration of these symptoms, if any,
were recorded. Body weights of the rats in all groups
were recorded once before the start of dosing and once
weekly.
2.8.1. Collection of blood, preparation of serum and
tissue homogenate
The methods described by Yusuf et al. [32] were used
for blood sample and liver collections. At the end of
the experiment (on the 21st day), the blood samples
were collected from overnight fasted rats (only water
allowed) by retro-orbital bleeding into heparinized and
non-heparinized tubes for haematological analysis and
biochemical analyses. Blood samples for biochemical
study were centrifuged at 3000 r.p.m. for 15 min, after
which the serum was transferred into a plain sample
bottle. The liver and kidney were carefully harvested
and stored in formalin for histological analysis.
2.8.2. Biochemical parameters
Spectrophotometric methods were used for the deter-
mination of aspartate transaminase and alanine trans-
aminase activities, as described by Reitman and Frankel
[33]. Urea in serum/plasma is hydrolyzed to ammonia
in the presence of urease. The ammonia is measur-
able photometrically by Berthelot’s reaction [34]. Cre-
atinine was analyzed as the amount of coloured com-
plex formed when alkaline solution reacts with picric
acid [35]. Determination of total protein was done, as
described by Weichselbaum [36]. Albumin measure-
ment was done with Bromocresol Green according to
the methods of Doumas et al. [37]. Sodium concen-
tration was estimated, as described by Maruna [38],
while serum chloride concentration was determined, as
described by Tietz [39].
2.8.3. Determination of haematological parameters
The haematological components including haemo-
globin (Hb), packed cell volume (PCV), red blood
cells (RBC), mean corpuscular volume (MCV), mean
corpuscular haemoglobin (MCH), mean corpuscular
haemoglobin concentration (MCHC), white blood cells
(WBC), platelate count (PLC), differential count (lym-
phocytes and neutrophils) and RBC Distribution Width
Count (RDWC) were determined using the automated
haematologic analyzer SYSMEX KX 21, a product of
SYSMEX Corporation, Japan employing the methods
described by Dacie and Lewis [40].
2.8.4. Histopathological studies
Ten percent formalin solution was used to fix the kid-
ney and liver tissues. Staining was done using Harri’s
haematoxylin and eosin method. The dried slides were
mounted with Distyrene Plasticizer Xylene (DPX) moun-
tant and cover slips and examined under the micro-
scope to verify histological details [41].
2.9. Statistical analysis
The analysis was performed using SPSS statistical pack-
age for WINDOWS (version 21.0; SPSS Inc, Chicago).
946 D. A. ADESINA ET AL.
Data were expressed as the Mean ±SEM of three deter-
minations. Results were subjected to ANOVA followed
by DMRT. Statistically significance was considered at
p<0.05.
3. Results
3.1. Acute toxicity
There was no death recorded in animals dosed with
alkaloid, flavonoid and phenolic extracts of S. acuta leaf
at 5000 mg/kg bw. No toxic symptom was observed
in any of the experimental animals. All animals in the
extract-treated groups were normal and did not display
any observable signs of toxicity on the skin, breathing,
food intake, water consumption, postural patterns and
hair loss.
3.2. In vivo antiplasmodial eect of the
phytochemicals
The % parasitaemia of P. bergei-infected mice treated
with alkaloid, flavonoid and phenolic extracts of S. acuta
are presented in Figures 1and 2. The negative con-
trol (infected untreated mice) had the highest % par-
asitaemia of 60.33 ±4.05. Alkaloid extract (300 mg/kg
bw) gave the lowest % parasitaemia and highest %
suppression of 25.00 ±7.00 and 58.56%, respectively,
while phenolic extract (600 mg/kg bw) gave the low-
est % parasitaemia and highest % suppression of
21.33 ±2.84 and 64.64%, respectively. However, chloro-
quine (25 mg/kg bw) had the loweR % parasitaemia
and higheR % suppression of 14.00 ±1.00 and 76.79%,
respectively when compared with the extracts-treated
groups.
There was loss of body weight in all the treated and
untreated mice (Table 1). The PCV of untreated mice as
well as those treated with the chloroquine (25 mg/kg
bw) and phytochemicals at 300 mg/kg bw had a loss
of PCV after treatments. Only mice treated with the
Tab le 1. Effect of alkaloid, flavonoid and phenolic extracts of S.
acuta leaf on the bodyweight of Plasmodium berghei-infected
mice.
Groups
Weight
before (g)
Weight
after (g)
Weight
gain/Loss (g)
Alkaloid 300 mg/kg bw 27.02 ±2.93b26.97 ±2.86b−0.05
Flavonoid 300 mg/kg bw 29.85 ±1.51b29.04 ±1.37c−0.81
Phenol 300 mg/kg bw 28.56 ±0.71b27.29 ±2.06c−1.27
Alkaloid 600 mg/kg bw 28.07 ±2.01b21.50 ±1.98a−6.57
Flavonoid 600 mg/kg bw 28.82 ±1.76b25.41 ±2.10b−3.41
Phenol 600 mg/kg bw 27.76 ±1.56b26.06 ±2.35b−1.7
Chloroquine 25 mg/kg bw 25.02 ±3.25ab 25.38 ±1.87b0.36
Negative control 25.96 ±0.75ab 22.94 ±0.29a−3.02
Note: Values are mean ±SEM of 3 determinations. Values along the same
column with different superscripts are significantly different (p<0.05).
flavonoids and phenols at 600 mg/kg bw gain PCV of
1.74 and 6.61%, respectively, after treatments (Table 2).
3.3. Sub-chronic eect of avonoid, alkaloid and
phenolic extracts of S. acuta leaf
3.3.1. Haematological parameters
Red Blood Cell was significantly increased (p<0.05) in
rats treated with 600 mg/kg bw of alkaloids and phenol
extracts when compared with other treatment groups
and the control group. The result also showed a signif-
icant (p<0.05) increase in Hb, MCH, MCHC, PCV and
WBC in all the treatment groups (alkaloid, flavonoid and
phenol extracts of S. acuta leaf) when compared with
the control group. No significant difference (p>0.05)
was noted in MCV, LY, PLC and RDWC when compared
with the control group (Table 3).
3.3.2. Biochemical parameters
Serum aspartate transaminase (AST), alanine transam-
inase (ALT), alkaline phosphatase (ALP), albumin, urea
and creatinine concentrations were not significantly dif-
ferent (p>0.05) in all the treatment groups (alkaloid,
flavonoid and phenol extracts of S. acuta) when com-
pared with the control group, while sodium was signif-
icantly lowerd (p<0.05) in all treatment groups when
Figure 1. In vivo antiplasmodial effect of alkaloid, flavonoid and phenolic extracts (300 mg/kg bw) of S. acuta leaf on Plasmodium
berghei-infected mice.
JOURNAL OF TAIBAH UNIVERSITY FOR SCIENCE 947
Figure 2. In vivo antiplasmodial effect of alkaloid, flavonoid and phenolic extracts (600 mg/kg bw) of S. acuta leaf on Plasmodium
berghei-infected mice.
Tab le 2. Effect of alkaloid, flavonoid and phenolic extracts of
S. acuta leaf on packed cell volume of Plasmodium berghei-
infected mice.
Groups PCV before (%) PCV after (%)
PCV
Gain/Loss
Alkaloid 300 mg/kg bw 40.18 ±7.37b36.00 ±6.85b−4.18
Flavonoid 300 mg/kg bw 50.17 ±5.26c45.57 ±4.42c−4.59
Phenol 300 mg/kg bw 50.23 ±5.37c33.56 ±9.31b−16.67
Alkaloid 600 mg/kg bw 43.76 ±3.29b42.22 ±1.90c−1.54
Flavonoid 600 mg/kg bw 41.52 ±5.16b43.26 ±1.17c+1.74
Phenol 600 mg/kg bw 44.91 ±2.05b38.30 ±2.59b+6.61
Chloroquine 25 mg/kg bw 54.96 ±3.13c40.82 ±2.65c−14.14
Negative control 36.17 ±7.41a23.83 ±2.53a−12.34
Note: Values are mean ±SEM of three determinations. Values along
the same column with different superscripts are significantly different
(p<0.05).
compared with the control group. Total protein concen-
tration was significantly (p<0.05) lower in rats treated
with 300 mg/kg bw of flavonoid extract, while chlo-
ride was significantly (p<0.05) higher in rats treated
with 300 mg/kg bw of alkaloid and phenol extracts than
other treatment groups and the control group (Table 4).
3.3.3. Bodyweight
The effects of the administration of alkaloid, flavonoid
and phenolic extracts of S. acuta on the bodyweight
of albino rats are shown in Table 5. The bodyweight
gains of the rats administered with alkaloid, flavonoid
and phenol extracts of S. acuta were not significantly
(p>0.05) different from each other but were signifi-
cantly (p<0.05) lower than those of the control group.
3.4. Histopathology
3.4.1. Effect of phytochemical extracts of S. acuta
leaf on the kidney histopathology of albino rats
The histological features as regards to renal vacuola-
tion, integrity of renal corpuscles, degenerative changes
and general appearances of kidney in the albino rats
treated with alkaloid, flavonoid and phenol extracts
of S. acuta are presented in Tables 6and 7. There
were no renal vacuolation in rats treated with phe-
nol and alkaloid extracts of Sida acuta when com-
pared with the control group. However, rat treated
with flavonoid extract had severe renal vacoulation at
300 mg/kg bw (Table 6) which clears out at 600 mg/kg
bw (Table 7) when compared with the control group.
The integrity of renal corpuscles is better as the
administered doses increases. There were also mild
degenerative changes in rats treated with 300 mg/kg
bw phenol and flavonoids; however, normal cellu-
lar kidney appearance was observed in all treatment
groups.
3.4.2. Effect of phytochemical extract of S. acuta
leaf on the liver histopathology of albino rats
The histological features, as regards to central vein,
integrity of hepatocyte, degenerative changes and gen-
eral appearances of liver in albino rats treated with
alkaloid, flavonoid and phenol extracts of S. acuta,
are presented in Tables 8and 9. Severe conges-
tion with flavonoid at 300 mg/kg bw (Table 8)which
cleared out at 600 mg/kg bw (Table 9) when compared
with the control group was observed. The integrity
of hepatocytes is better as the administered doses
increase; no any degenerative changes were observed
in all treatment groups. Also, normal hepatocellu-
lar appearance was observed in all treatment groups
(Figures 3and 4).
4. Discussion
Phytochemicals are secondary metabolites of plants
known to exhibit diverse pharmacological and bio-
chemical effects on living organisms [42]. The acute tox-
icity of the alkaloids, flavonoids and phenols was mea-
sured to obtain information regarding the oral safety of
the secondary metabolites. Interestingly, it was discov-
ered that all phytochemicals tested had LD50 >than
5000 mg/kg bw, thus exhibiting a wide safety margin
upon acute oral administration. However, Konaté et al.
948 D. A. ADESINA ET AL.
Tab le 3. Effects of alkaloid, flavonoid and phenolic extracts of S. acuta leaf on haematological parameters of albino rats.
Groups RBC Hb MCH MCHC PCV WBC MCV PLC N L RDWC
Alkaloid 300 mg/kg bw 6.75 ±0.35a13.05 ±0.35b19.50 ±0.50b30.50 ±0.50b42.00 ±2.0b9.95 ±0.55b63.50 ±1.50a387.50 ±24.50a7.00 ±1.00a73.50 ±2.50a16.50 ±0.50a
Flavonoid 300 mg/kg bw 6.75 ±0.45ab 13.45 ±0.85b18.50 ±1.50b28.50 ±0.50ab 43.00 ±5.0b8.10 ±3.00b64.50 ±3.50a525.00 ±7.00a10.50 ±1.50b60.00 ±18.00a16.50 ±0.40a
Phenol 300 mg/kg bw 6.75 ±0.25ab 12.55 ±0.50b19.50 ±0.50b29.00 ±1.00b40.00 ±0.0b10.15 ±0.05b65.50 ±0.50a416.50 ±17.50a10.50 ±6.50b64.50 ±18.50a16.95 ±0.15a
Alkaloid 600 mg/kg bw 7.60 ±0.00b13.75 ±0.75b18.00 ±1.00b31.00 ±1.00b43.00 ±3.0b8.05 ±0.05b60.00 ±0.00a382.00 ±71.00a10.00 ±0.00b80.00 ±2.00a17.12 ±0.12a
Flavonoid 600 mg/kg bw 6.95 ±0.15ab 13.10 ±0.30b18.00 ±2.00b30.00 ±0.00b39.50 ±0.5b5.60 ±0.50b61.50 ±2.50a522.00 ±99.01a6.00 ±1.00a76.00 ±4.00a17.30 ±0.40a
Phenol 600 mg/kg bw 7.75 ±0.55b14.15 ±0.75b20.50 ±0.50b31.50 ±0.50b43.50 ±3.5b9.20 ±1.80b62.50 ±3.50a514.00 ±27.00a6.00 ±1.00a72.50 ±3.50a19.00 ±0.40a
Positive Control 6.05 ±0.65a9.10 ±3.70a16.00 ±4.00a26.00 ±4.00a32.50 ±9.5a5.95 ±1.8a64.50 ±0.50a410.00 ±11.09a6.00 ±1.00a75.00 ±5.00a16.95 ±0.67a
Standard Value 7.2-10.1 10.36-13.9 13.0-19.0 26.5-58 33–50 10.2-13.8 46.5-65.0 385–610 20–70 20–70 15–20
Note: Hb-Haemoglobin, PCV-Packed Cell Volume, MCH-Mean Cell Haemoglobin, MCHC-Mean Cell Haemoglobin Concentration, RBC-Red Blood Cell Count, WBC-Total White Blood Cell Count, MCV-Mean Cell Volume, PLC-Platelet Count, N-
Neutrophils, L-Lymphocytes, RDWC-RBC Distribution Width. Values are mean ±SEM of 3 determinations. Values along the same column with different superscripts are significantly different (p<0.05). Standard Value Source: University of
Pennylsavania, School of Veterinary Medicine (2002).
Tab le 4. Effects of alkaloid, flavonoid and phenolic extracts of S. acuta leaf on biochemical parameters of albino rats.
Groups AST ALT ALP TP Albumin Sodium Chloride UREA Creatinine
Alkaloid 300mg 10.80 ±0.00a37.60 ±4.00a87.9 ±3.52a36.10 ±1.90b2.28 ±0.30a89.36 ±5.58a89.09 ±0.37bc 76.84 ±3.66a3.14 ±1.43a
Flavonoid 300mg 10.80 ±3.60a43.24 ±5.64a85.09 ±3.41a18.05 ±8.50a2.01 ±0.03a77.79 ±1.19a73.68 ±3.00ab 93.31 ±1.83a4.28 ±1.42a
Phenol 300mg 18.00±3.60ab 45.12 ±3.76a82.09 ±3.48a29.45 ±0.95ab 1.82 ±0.19a99.73 ±2.39a98.87 ±10.90c100.62 ±27.44a3.71 ±0.28a
Alkaloid 600mg 7.20±3.60a44.18 ±4.70a91.77 ±3.36ab 26.60 ±3.80ab 2.01 ±0.22a53.45 ±13.56a84.96 ±0.75abc 87.82±3.66a6.00 ±1.43a
Flavonoid 600mg 16.20 ±1.80ab 47.94 ±0.94a88.02 ±3.59a38.00 ±5.70b2.12 ±0.08a74.60 ±34.71a70.30 ±1.88a84.16 ±7.32a5.42 ±0.28a
Phenol 600mg 12.60 ±5.40a46.06 ±2.82a79.91 ±3.65a31.35 ±2.85ab 1.56 ±0.90a104.12 ±3.59a79.70 ±3.76ab 75.01 ±98.15a6.00 ±0.29a
Positive Control 18.00 ±0.00ab 46.06 ±2.82a80.67 ±3.79a34.09 ±0.11b1.56 ±0.38a176.73 ±15.56b75.18 ±1.50ab 84.16 ±7.32a6.00 ±2.57a
Standard Value 10–45 10–35 15–45 11–25 3.6-5.3 140–160 90–110 25–45 0.5-2.2
Note: Values are mean ±SEM of three determinations. Values along the same column with different superscripts are significantly different (p<0.05).
Standard Value Source: University of Pennylsavania, School of Veterinary Medicine (2002).
JOURNAL OF TAIBAH UNIVERSITY FOR SCIENCE 949
Tab le 5. Effect of alkaloid, flavonoid and phenolic extracts of S. acuta leaf on the bodyweight of
albino rats.
Groups Week 1 Week 2 Week 3 Total weight gain
Alkaloid 300 mg/kg bw 141.30 ±1.80bc 148.90 ±4.50bc 160.00 ±2.00c18.70 ±1.73a
Flavonoid 300 mg/kg bw 117.80 ±8.00ab 131.80 ±7.30b137.00 ±1.00ab 19.20 ±0.96a
Phenol 300 mg/kg bw 115.85 ±2.36ab 127.30 ±1.00ab 138.50 ±1.50ab 22.65 ±1.06ab
Alkaloid 600 mg/kg bw 126.40 ±4.10b134.30 ±20.80b144.50 ±19.50b18.10 ±0.95a
Flavonoid 600 mg/kg bw 121.50 ±6.20b126.65 ±11.95ab 134.00 ±19.00ab 12.50 ±0.02a
Phenol 600 mg/kg bw 93.35 ±2.95a100.75 ±0.95a113.00 ±4.00a19.65 ±1.31a
Positive Control 107.90 ±7.40ab 132.30 ±1.40b144.00 ±4.00b36.10 ±2.09b
Tab le 6. Effect of alkaloid, flavonoid and phenolic extracts of S. acuta (300 mg/kg bw) Leaf on the
kidney histopathology of albino rats.
Kidney Control
300 mg/kg bw
Phenolic
300 mg/kg bw
Flavonoid
300 mg/kg bw
Alkaloid
Renal vacuolation Nil Mild Severe Nil
Integrity of renal corpuscles Excellent Mild distortion Poor/severe distortion Good with no distortion
Degenerative changes Nil Mild Mild Nil
General appearances Excellent Good Mild distortion Excellent
Tab le 7. Effect of alkaloid, flavonoid and phenolic extracts of S. acuta (600 mg/kg bw) leaf on the kidney histopathology of albino
rats.
Kidney Control
600 mg/kg
bw Phenolic
600 mg/kg bw
Flavonoid
600 mg/kg
bw Alkaloid General remarks
Renal vacuolation Nil Nil Mild Nil Severe renal vacuolation with flavonoid
(300 mg) which clears out at 600 mg
when compared with control
Integrity of renal corpuscles Excellent Excellent Mild cellular integrity Good Integrity of renal corpuscles is better as the
administered doses increase
Degenerative changes Nil Nil Nil Nil Generally, degenerative changes seen at
lower doses
General appearances Excellent Excellent Good Excellent Generally normal cellular appearance
Tab le 8. Effect of alkaloid, flavonoid and phenolic extracts of S. acuta (300 mg/kg bw)leaf on the
liver histopathology of albino rats.
Liver Control
300 mg/kg bw
Phenolic
300 mg/kg bw
Flavonoid
300 mg/kg bw
Alkaloid
Central vein congestion Nil Nil Severe congestion Nil
Integrity of hepatocytes Excellent Mild distortion Poor/severe distortion Good with no distortion
Degenerative changes Nil Mild Mild to moderate Nil
General appearances Excellent Good Mild distortion Excellent
Tab le 9. Effect of alkaloid, flavonoid and phenolic extracts of S. acuta (600 mg/kg bw) leaf on the liver histopathology of albino rats.
Liver Control
600 mg/kg bw
Phenolic
600 mg/kg bw
Flavonoid
600 mg/kg bw
Alkaloid General remarks
Central vein congestion Nil Nil Mild Nil Severe congestion with flavonoid which clears
out at 600mg when compared with the
control group
Integrity of hepatocytes Excellent Excellent Good with improved integrity Good Integrity of hepatocytes is better as the
administered doses increase
Degenerative changes Nil Nil Nil Nil Generally, no degenerative changes seen.
General appearances Excellent Excellent Good Excellent Generally normal cellular appearance
[43] reported LD50 values of 3.2 g/kg for crude extract of
S. acuta in mice, thus suggesting negligible level of tox-
icity of the crude extract. Results of the present study
indicated that alkaloids, flavonoids and phenols exhib-
ited active antiplasmodial activity in accordance with
the reports of Carvalho [44] who stated that a com-
pound with reduction in parasitaemia ≥30% is consid-
ered active. This suggests that alkaloid, flavonoid and
phenolic extracts of S. acuta leaf may be considered as
a bioactive metabolite with potential for the develop-
ment of a novel antimalarial drug.
However, the higher % suppression of the parasite
produce by phenol extract (64.64% at 600 mg/kg bw)
compared to alkaloid and flavonoid extracts indicated
that the phenol extracts would be better antimalarial
than the other phytochemicals. Chloroquine has been
reported for the disruption of the parasite’s cell mem-
brane and induction of auto-parasite digestion via the
formation of FP-chloroquine complex which impaired
the haeme polymerization [45]. The lower percentage
suppression observed in the extract-treated group than
that of the chloroquine-treated group may be because
950 D. A. ADESINA ET AL.
Figure 3. Histomicrogram of the kidney of alkaloid, flavonoid and phenolic extracts of S. acuta leaf in albino rats. (Arrow indicates
renal vacuolation at 300 mg/kg bw which was corrected at 600 mg/kg bw).
Figure 4. Histomicrogram of the liver of alkaloid, flavonoid and phenol extracts of S. acuta leaf in albino rats. (Arrow indicates central
vein congestion at 300 mg/kg bw which was corrected at 600 mg/kg bw).
the phytochemicals in its current form and at the doses
administered had not accumulated sufficiently to bring
about considerable suppression [46]orithasalower
speed of action than that of chloroquine. This is similar
to the study of Builder [47] who reported 83% inhi-
bition of parasitaemia for chloroquine. Although the
activities demonstrated by each phytochemical extract
were lower than those of a standard drug (chloroquine)
(76.79%), it is possible that a combination of 2 or 3
of these phytochemicals would act synergistically to
exhibit higher antimalarial activities comparable to or
better than the chloroquine.
Anaemia is one of the recognized effects of malaria
that occurs due to the destruction of red blood cell.
Therefore, effective antimalariaL agents are expected to
reverse or prevent the parasite-induced anaemic con-
dition. The increases in PCV (1.74 and 6.61%) of mice
treated with the flavonoid and phenol extracts of S.
acuta (Table 2) are an indication of enhanced resis-
tance to erythrocyte haemolysis [48]. The flavonoids
and phenols, therefore, prevented the parasite-induced
anaemic condition in mice. The high loss of body weight
in mice treated with high dose (600 mg/kg bw) of
each phytochemical extract could be attributed to the
JOURNAL OF TAIBAH UNIVERSITY FOR SCIENCE 951
depressing effect of the phytochemical extracts on feed
uptake/appetite [49] with consequent effects on the
phytochemicals.
Hematological parameters are widely used markers
in assessing the toxicity or safety of a drug/extract as
well as assessing the health status of an animal [50]. The
significant increase in the WBC count in the rats treated
with alkaloid, flavonoid and phenolic extracts could be
an indication of leucopoetic potentials and possible
immunomodulatory properties of the phytochemical
extract which enhances the production of more WBC
[51]. This will enhance the antibody-generating poten-
tial of the animals via phagocytosis and will have high
resistance to infection and diseases [52] Similarly, the
significant (p<0.05) increase in erythrocytic indices,
including HGB, PCV, MCH, MCHC, in all the treatment
groups (alkaloid, flavonoid and phenolic extracts of S.
acuta) when compared with the control is an indication
of stimulation of erythropoiesis by the phytochemicals.
The phytochemicals must have enhanced the release of
erythropoietin in the kidney, a humoral mediator of RBC
production [53].
Analyses of biochemical parameters, such as transa-
minases, phosphatase, albumins and total proteins,
have been identified as a reliable indicator of organ
function and healthy or disease status of animals dur-
ing subacute administration of extracts or phytochemi-
cals to animals [54,55,56]. Alterations in serum levels of
these markers are indicators of hepatocellular impair-
ments, cell membrane impairment, liver hepatitis or
cirrhosis.
Interestingly, the serum aspartate transaminase
(AST), alanine transaminase (ALT), alkaline phosphatase
(ALP) and albumin concentrations were not significantly
altered by the treatment with alkaloid, flavonoid and
phenol extracts of S. acuta. This is an indication that
the administration of phytochemicals to the rats has
not compromised the integrity of the liver, i.e. the func-
tional integrity of the liver has been preserved. How-
ever, the alterations in total protein concentrations in
rats treated with 300 mg/kg bw of flavonoid extract
could be attributed to protein turnover during the
metabolism of the flavonoid, such an increase in total
proteins could lead to dehydration which is detrimental
to cellular homeostasis, thus effecting the health of the
animals [57].
Serum electrolytes, urea and creatinine, on the other
hand, indicate the function integrity of the kidney [55].
In this study, urea and creatinine concentrations were
not affected by the administration of the phytochem-
icals, thus suggesting that the functional capacity of
the kidney has not been compromised [58] However,
the significant alterations (p>0.05) in the concentra-
tions of sodium and chloride in the albino rats dosed
with the phytochemical extracts are an indication that
the functional integrity of the kidney as regards to this
metabolite has been compromised. Nevertheless, the
mild alterations recorded in the electrolyte may not be
of clinical relevance without histopathological backup
[59]. Fortunately, it was observed that the alkaloid and
phenol extracts of Sida acuta do not alter the normal cel-
lular architectures of the liver and kidney. In fact, the
integrity of the hepatocytes and renal corpuscles was
improved at higher doses of the alkaloid and pheno-
lic extracts better than the control group. It is, there-
fore, reasonable to conclude that the mild alteration
observed in some of the biochemical parameters could
be attributed to the initial metabolic adaptations of the
animals [60] to the phytochemicals and are not of clin-
ical significance as regards to the integrity of the liver
and kidney.
5. Conclusion
The study suggests that alkaloid, flavonoid and phe-
nolic extracts of Sida acuta leaf possess antiplasmodial
properties with phenolic extract (600 mg/kg bw) pos-
sessing the highest plasmodial suppression (64.64%)
among other extracts and close to the positive con-
trol (chloroquine) which caused plasmodial suppres-
sion of 76.79%. The phytochemicals were also found
to be safe as revealed by biochemical, haematological
and histopathological findings of sub-chronic toxicity
studies.
Disclosure statement
No potential conflict of interest was reported by the author(s).
ORCID
Daniel Anuoluwa Adesina http://orcid.org/0000-0002-6619-
3656
Sherifat Funmilola Adefolalu http://orcid.org/0000-0002-
6923-1304
Ali Audu Jigam http://orcid.org/0000-0001-6586-8087
Bashir Lawal http://orcid.org/0000-0003-0676-5875
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