







































_____________________________________________________________________________________________________ 
 
*Corresponding author: E-mail: meshackkugama@gmail.com; 
 
Cite as: Y.A., Onaolapo, Mazadu, R.M., Baraya, K.Y., Fasuyi, F.H., Irhue, A.E., Ahmed, B., Hassan, A., Shehu, A.A., Bala, 
S.B., Kasim, S.N., and Kugama, M.A. 2024. “Effect of Alstonia Boonei and Morinda Lucida on Renal Histology of Wistar Rats 
Infected With Trypanosoma Brucei Brucei”. Asian Journal of Immunology 7 (1):292-301. 
https://doi.org/10.9734/aji/2024/v7i1152. 

 
 

Asian Journal of Immunology 
 
Volume 7, Issue 1, Page 292-301, 2024; Article no.AJI.127386 
 

 
 

 

 

Effect of Alstonia boonei and Morinda 
lucida on Renal Histology of Wistar 

Rats Infected with Trypanosoma brucei 
brucei 

 
Onaolapo, Y.A. a, Mazadu, R.M. a, Baraya, K.Y. a, 

Fasuyi, F.H. b, Irhue, A.E. a, Ahmed, B. a, Hassan, A. a, 
Shehu, A.A. a, Bala, S.B. a, Kasim, S.N. c 

 and Kugama, M.A. a* 

 
a Nigerian Institute for Trypanosomiasis Research, Federal Ministry of Science, Technology and 

Innovation, Nigeria. 
b National Biotechnology Development Agency, Bioresource Development Center, Onipanu, 

Ogbomoso, Oyo State, Nigeria. 
 c Department of Applied Biology, College of Science and Technology, Kaduna Polytechnich, Nigeria. 

 
Authors’ contributions 

 
This work was carried out in collaboration among all authors. All authors read and approved the final 

manuscript. 
 

Article Information 
 

DOI: https://doi.org/10.9734/aji/2024/v7i1152  
 

Open Peer Review History: 
This journal follows the Advanced Open Peer Review policy. Identity of the Reviewers, Editor(s) and additional Reviewers,  

peer review comments, different versions of the manuscript, comments of the editors, etc are available here: 
https://www.sdiarticle5.com/review-history/127386 

 

 
Received: 02/10/2024 
Accepted: 05/12/2024 
Published: 20/12/2024 

 
  

Original Research Article 

https://doi.org/10.9734/aji/2024/v7i1152
https://www.sdiarticle5.com/review-history/127386


 
 
 
 

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ABSTRACT 
 

Animal African trypanosomiasis is a protozoan disease caused by trypanosomes and transmitted 
through the bite of an infected tsetse fly. The disease has an adverse effect to the economy of the 
affected areas and drugs used for the treatment of the disease are faced with several challenges 
ranging from resistance to the parasite to high level of toxicity. Our study aimed at evaluating the 
effect of Alstonia boonei and Morinda lucida plant extracts on the renal histology of wistar rats 
infected with Trypanosoma brucei brucei. Phytochemical screening of the methanol extracts 
indicated the presence of alkaloids, saponins, tannins, flavonoids, carbohydrates, phenols and 
steroids/terpenes, while glycosides were not detected. Only four of the eight constituents tested for 
were detected in the chloroform leaf extract of A. boonei, namely, alkaloids, tannins, flavonoids, 
and carbohydrates. Alkaloids, tannins falvonoids, and carbohydratses were detected in both the 
chloroform and methanol extract of Morinda lucida. Histological lesions of the infected-untreated 
rats revealed severe cellular degeneration in the renal cortex of T. brucei infected rats. 
Furthermore, the glomerular tufts were shrunken leaving a large Bowman’s. The renal 
photomicrographs of rat treated with 500mg/kgbwt of Alstonia boonei revealed that the renal cortex 
presented apparently normal histological features of Malpighian renal corpuscle containing 
glomerulus and Bowman’s space with slightly shrunken glomerulus. The proximal convoluted 
tubules had narrow lumina and were lined with cuboidal cells with rounded vesicular basal nuclei. 
Groups treated with 1000mg/kgbwt revealed erythrocytic remains of degenerated renal cells with 
scanty inflammatory cells in the cortical region of kidney of the wistar rats. However, the 
glomerulus, the proximal and distal convoluted tubules presented normal histological features. The 
results indicated that treatment with graded doses of methanol extract of A. boonei elicited varying 
effects on the visceral organs of the infected animals. It was noted that Alstonia boonei ameliorated 
the effect of the infection on the kidney of the infected animals with the histology sections being 
comparable with those treated with the reference anti-trypanosomal drug (Diminazine aceturate) 
and the uninfected rats. However, Morinda lucida had little or no ameliorative effect on the kidney 
of wistar rats infected. Therefore, we recommend that extracts of Astonia boonei should be 
characterized and active components responsible for the ameliorative effect be detected and 
elucidated structurally. 
 

 

Keywords: Trypanosomiasis; parasitaemia; phytochemicals; toxicity. 
 

1. INTRODUCTION 
 

Animal trypanosomosis has a serious effect of 
ravaging both lower and higher ruminants in sub-
saharan Africa and is caused by a protozoan 
parasite of the genus Trypanosoma [1]. The 
species responsible for this disease known as 
nagana in Africa are Trypanosoma vivax, 
Trypanosoma congolense and to a lesser extent 
Trypanosoma brucei brucei. The disease is 
transmitted through the bite of an infected tsetse 
fly of the Glossina specie. It is implicative in 
causing severe anaemia, weight loss, reduced 
productivity, infertility and abortion, with death 
occurring in some animals during the acute and 
chronic phase of the infection [2]. 
 

In Nigeria, studies of ethno medicinal plants used 
in the traditional management of trypanosomiasis 
indicated both significant in vitro and in vivo anti-
trypanosomal activity [3]. The country is richly 
endowed with indigenous plants, which are used 
in herbal medicine to cure diseases and heal 
injuries, some of these plants are used as food or 

medicine. These indigenous plants exhibit a wide 
range of biological and pharmacological activities 
such as anticancer, anti-inflammatory, diuretic, 
laxative, antispasmodic, antihypertensive, 
antidiabetic and antimicrobial functions. It is 
generally assumed that the active medicinal 
constituents contributing to these protective 
effects are the phytochemicals, vitamins and 
minerals. Some of these secondary metabolite 
can act singly or in synergy to bring about 
healing [4]. 
 

Currently, homidium chloride, isometamidium 
and diminazene aceturate are used for the 
treatment of this disease. Each of these drugs 
has one or more of these challenges: expensive, 
highly toxic, need parenteral administration and 
parasites increasing resistance. Also, attempt to 
produce vaccine for the prevention of 
trypanosomiasis proves abortive because of the 
parasite changing its antigenic identity. 
Therefore, the need for alternative new 
molecules that are safe, effective and affordable 
is urgent [5].  



 
 
 
 

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Scientific studies revealed that natural products 
derived from plants offer novel possibilities to 
obtain new drugs that are active against 
trypanosomes and investigation of 
antitrypanosomal activity of traditionally used 
plants has been a major area of concern [5,6]. 
There is need for the development of new agents 
to complement the existing drugs for the 
treatment of African trypanosomiasis. Alstonia 
boonei and Morinda lucida are used as traditional 
remedy for several infectious and noninfectious 
diseases including antihelminthic, anti-
inflammatory, analgesic/pain-killing, 
antimalaria/antipyretic, antidiabetic (mild 
hypoglycaemic), antiprotozoan and antibiotic 
properties [7]. These justify the need to explore 
these plants as potent drugs for trypanosomiasis. 
 

2. MATERIALS AND METHODS 
 

2.1 Plants Materials  
 

The plant materials were harvested in the 
morning in, Zaria Local Government Area of 
Kaduna State. It was authenticated in the 
herbarium, Department of Biological Science, 
Kaduna State University and given the voucher 
number (A765 and Q567) for Alstonia boonei and 
Morinda locida respectively. It was air dried at 
room temperature, pulverized in a mortal using 
pestle to obtained fine powder. 
 

2.2 Experimental Animals 
 
Sixty (60) wistar rats were used for this study. 
The animals were purchased from the animal 
house of Nigerian Institute for Trypanomiasis 
Research, Kaduna. They were kept in a standard 
rat cage, fed with standard pellet diet with water 
ad libitum for thirty (30) days. 
 

2.3 Infection of Animals with 
Trypanosomes  

 
The parasite Trypanosoma brucei brucei was 
obtained from Nigerian Institute for 
Trypanosomiasis Research, Kaduna. The 
animals were inoculated with 1000 parasite per 
meal of blood intraperitoneally. The number of 
parasite was compared with Harbert and 
Lumsden rapid matching method [8].  
 

2.4 Extraction from Plant Materials 
 

Extraction was done using Soxhlet apparatus. 
One thousand grams (1000g) of the powdered 
plant was divided into five hundred (500g) each. 

2,000ml of each of methanol and chloroform 
were used for the extraction of each of the 
powdered plant. The solvent was recovered and 
concentrated in vacuoe to obtain the extract.  
  

 2.5 Phytochemical Analysis of the Plant 
Parts  

 

The phytochemical analysis was carried out 
according to the method of Sofowora. The 
presence of tannins, saponins, flavonoids, 
glycosides, anthraquinones, carbohydrate, 
phenols, terpens and alkaloids was tested 
qualitatively [9]. 
 

2.6 Histopathological Examination of 
Harvested Organs 

 

The kidney specimens from each rat was stored 
immediately in 10%v/v formalin in normal saline 
after gross histological examination and 
dehydrated using increasing Concentrations of 
isopropyl alcohol (80e100%). Paraffin sections at 
5mm thickness was made from the paraffin 
embedded organs using a Leica rotary 
microtome (Bright B5143 Huntington, England). 
This was followed by routine staining with 
hematoxylin and eosin which involved the 
process of deparaffinization, hydration, staining, 
rinsing and clearing in xylene. Slides was viewed 
under light microscope with photomicrographs 
taken with a Leica DM750 Camera Microscope 
(400 X) [10]. Histopathological lesions were 
scored using semi-quantitative approach as 
follows:  
 

0 for normal, 1 (1%e30%) for mild, 2 (31%e70%) 
for moderate, and 3 (>70%) for severe. 
 

2.7 Statistical Analysis of Data  
 

Values of the data obtained from the study was 
summarized and expressed data analysis was 
performed using Statistical Package for Social 
Science (SPSS) 2018 version 23.0.  
 

3. RESULTS 
 

3.1 Phytochemical Constituents  
 
Screening of Extracts of Alstonia boonei 

and Morinda lucida Using Methanol 
and Chloroform as Solvents 

 

The results of the phytochemical constituents 
and screening of the methanolic leaf extract of 
Alstonia boonei indicated the presence of seven 



 
 
 
 

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constituents, namely, alkaloids, saponins, 
tannins, flavonoids, carbohydrates, phenols and 
steroids/terpenes, while glycosides were not 
detected. Only four of the eight constituents 
tested for were detected in the chloroform leaf 
extract of A. boonei, namely, alkaloids, tannins, 
flavonoids, and carbohydrates. Four 
phytoconstituents namely, alkaloids, tannins, 
flavonoids, and carbohydrates were detected in 
the methanol extract of Morinda lucida. In the 
chloroform extract of M. lucida, only three of the 
eight phytoconstituents were detected namely, 
alkaloids, tannins, and flavonoids; saponins, 
glycosides, carbohydrates, phenols, and steroids 

were not detected. All three constituents that 
were present in the chloroform extract of M. 
lucida were detected in relatively high quantities. 
Glycosides were not detected in all the solvents 
leaf extracts of the plants, while carbohydrates 
were detected, albeit in relatively minute 
quantities, in the methanol and chloroform 
extracts of A. boonei and in the methanol extract 
of M. lucida. Phenols as well as terpenes were 
detected in the methanol extract of A. boonei; 
none was detected in the chloroform extract of 
the plant nor in either the methanol or chloroform 
extracts of M. lucida (Table 1). 

 
Table 1. Phytochemical Screening of the Methanol and Chloroform Leaf Extracts of 

Alstonia boonei and Morinda lucida 
 

Phytochemical Constituents ABME ABCE MLME MLCE 

Alkaloids  
 

+++ 
+++ 

+++ 
+++ 

+++ 
+++ 

+++ 
+++ 

Saponins  +++ --- --- --- 
Tannins  +++ +++ +++ +++ 
Flavonoids  +++ +++ +++ +++ 
Glycosides  --- --- --- --- 
Carbohydrates  + + + --- 
Phenolic compounds +++ --- --- --- 
Terpenes ++ --- --- --- 
ABME: Alstonia boonei Methanol Extract; ABCE: Alstonia boonei Chloroform Extract; MLME: Morinda lucida 

Methanol Extract; MLCE: Morinda lucida Chloroform Extract 
+++: highly present; ++: moderately present; +: present; ---: undetected 

 

 
 

Plate 1. Photomicrographs of renal section from T. brucei infected rat administered 500 
mg/kgb.wt of methanol extract of A. boonei (Group A). The renal cortex presents apparently 
normal histological features of Malpighian renal corpuscle containing glomerulus (G) and 
Bowman’s space (arrowhead) with slight shrunken glomerulus. The proximal convoluted 

tubules (P) have narrow lumina and are lined with cuboidal cells with rounded vesicular basal 
nuclei. The distal convoluted tubules (D) have wider lumina and are lined with cuboidal cells. 

H&E x25 



 
 
 
 

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Plate 2. Photomicrographs of renal section of T. brucei brucei infected rats administered 1000 
mg/kgb.wt of methanol extract of A. boonei (Group B). The renal cortex shows what appeared 
to be erythrocytic (circle) remains of degenerated renal cells with scanty inflammatory cells 
(arrowhead). The glomerulus (G), proximal (P) and the distal convoluted tubules (D) present 

normal histological features. H&E x250 
 

 
 

Plate 3. Photomicrographs of renal section T. brucei brucei infected rats administered 500 
mg/kgb.wt of chloroform extract of M. lucida (Group C). The renal cortex shows severe atrophy 
of the glomerular tuft (G), with nearly all the tubular cells showing degeneration (arrow head). 
Note the focal aggregation of mononuclear cells around one of the glomerulus (circle). H&E 

x250 
 

3.2 Histopathological Studies of Kidney 
Organs of Trypanosoma brucei brucei 
Infected Wistar Rats Treated with Leaf 
Extracts of Alstonia boonei boonei 
and Morinda lucida  

 
Effect of 500 mg/kgb.wt of the methanol 
extract of Alstonia boonei on kidney 
histology of Trypanosoma brucei brucei 
infected rats: The renal photomicrographs of 
wistar rat in Group A revealed that the renal 
cortex presented apparently normal histological 

features of Malpighian renal corpuscle containing 
glomerulus and Bowman’s space with slightly 
shrunken glomerulus. The proximal convoluted 
tubules had narrow lumina and were lined with 
cuboidal cells with rounded vesicular basal 
nuclei. The distal convoluted tubules had wider 
lumina and were also lined with cuboidal cells 
(Plate 1). 
 

Effect of 1000 mg/kgb.wt of the Methanol Leaf 
Extract of Alstonia boonei on Kidney 
Histology of Trypanosoma brucei brucei 
Infected Wistar Rats: Erythrocytic remains of 



 
 
 
 

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degenerated renal cells with scanty inflammatory 
cells were seen in the cortical region of kidney in 
the rats in Group B. However, the glomerulus, 
and the proximal and distal convoluted tubules 
presented normal histological features (Plate 2). 

 
Effect of 500 mg/kgb.wt of the chloroform 
extract of M. lucida on kidney histology of T. 
brucei brucei infected rats: The renal cortex of 
the rat in Group C showed severe atrophy of the 
glomerular tuft; in addition, almost all the tubular 
cells showed degeneration. There was also focal 

aggregation of mononuclear cells around one of 
the glomeruli (Plate 3). 

 
Effect of 1000 mg/kgb.wt of the Chloroform 
Extract of Morinda lucida on Kidney 
Histology of Trypanosoma brucei brucei 
Infected Wistar Rats: Renal section from T. 
brucei brucei infected rats treated with 1000 
mg/kgb.wt of chloroform extract of M. lucida 
(Group D) showed features of cellular 
degeneration and shrunken glomerular tuft, as 
well as hemorrhage (Plate 4). 

 

 
 

Plate 4. Photomicrographs of renal sections T. brucei brucei infected rats administered 1000 
mg/kgb.wt of chloroform extract of M. lucida (Group D). The section is hemorrhagic (circle), 

showing features of cellular degeneration and shrunken glomerular tuft (G). H&E x250 

 

 
 

Plate 5. Photomicrographs of renal section T. brucei brucei infected rats administered 
Diminazine 3.5 mg/kgb.wt (Group E). The renal cortex shows apparently normal renal 

corpuscle of glomerulus (G), proximal and distal convoluted tubules (P and D) with few focal 
aggregated mononuclear infiltrated cells (circle). H&E x250 

 



 
 
 
 

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Effect of Diminazine aceturate (3.5 mg/kgb.wt) 
on kidney histology of T. brucei brucei 
infected rats: The renal cortex of rats in Group 
E showed apparently normal renal corpuscle of 
glomerulus, proximal and distal convoluted 
tubules, with few focal aggregated mononuclear 
infiltrated cells (Plate 5). 
 

Kidney histology of untreated Trypanosoma 
brucei brucei infected rats: Severe cellular 
degeneration was seen in the renal cortex of T. 

brucei infected rats that received no treatment 
(Group F). Furthermore, the glomerular tufts 
were shrunken leaving a large Bowman’s space 
(Plate 6). 

 
Kidney histology of uninfected rat: The renal 
cortex of uninfected rat (Group G) revealed 
apparently normal glomerulus, proximal and 
distal tubules, with few infiltrated inflammatory 
cells (Plate 7). 

 

 
 

Plate 6. Photomicrographs of renal section of untreated T. brucei brucei infected rats (Group 
F) The renal cortex is showing severe cellular degeneration and shrunken glomerular tufts (G) 

leaving a large Bowman’s space (BS). H&E x250 
 
Effect on Liver Histology: 
 

 
 

Plate 7. Photomicrographs of renal sections of uninfected-untreated (Group G). The renal 
cortex shows renal corpuscle with apparently normal glomerulus (G), proximal and distal 

tubules (P and D) with few infiltrated inflammatory cells (circles). H&E x250 



 
 
 
 

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4. DISCUSSION 
 

The result from the phytochemical screening 
obtained in this study revealed the presence of 
bioactive components in the aqueous and 
chloroform extracts of the leaves of A. boonei 
which is in agreement with the reports of [11] 
who noted the presence of phytochemicals in the 
water and methanol leaf extracts of A. boonei. 
[12] had also reported the presence of some 
bioactive agents in the methanol stem bark and 
leaf extracts of the plant extracts. These reports 
indicated that A. boonei is an excellent source of 
chemical moieties that could find important 
medicinal and pharmacological applications. 
Similarly, extracts of Morinda lucida showed an 
array of phytoconstituents including alkaloids, 
tannins, flavonoids, and carbohydrates. This also 
agreed with the reports of [13,14] who had 
previously reported the presence of bioactive 
agents in solvent extracts of the stem bark and 
root of M. lucida (saponins, tannins, anthracenes, 
flavonoids, alkaloids, carbohydrate and protein).  
 

The present results indicated that the class of 
phytochemicals varied between the two plants 
(A. boonei and M. lucida) and between the 
solvent used for extracting a particular plant. 
According to [15], the metabolic profiles of 
medicinal plants are mainly controlled by genetic 
factors, adding that variations in the levels of 
phytochemicals may represent long-term 
ecological and evolutionary interactions. Thus, 
the genetic differences between the two plants. 
The present results indicated that T. brucei 
brucei infection in rats is characterized by several 
histopathological derangements of visceral 
organs (liver) which may impair their functionality 
and result in death of untreated animals. The 
histopathological abnormalities observed in the 
kidney of untreated T. brucei brucei infected rats 
were consistent with the reports of other authors 
who had reported the effect of Trypanosoma 
infections in animals. Also, [16] had noted 
alterations in kidney, liver, lungs and spleen of 
mice infected with T. brucei. In the study, the 
kidneys were characterized by congested 
intertubular spaces, and the lungs were seen 
with congestions of the blood vessels, interstitial 
expansion and emphysema in infected mice. 
Again, [17] who assessed the histological 
changes associated with infection with T. evansi, 
a phylogenetic close relative of T. brucei, noted 
marked changes in the visceral organs including 
cellular infiltration of the liver, myocardial 
necrosis, tubular necrosis of the nephrons and 
destruction of the Bowman’s capsule. The 
changes in the visceral organs seen in 

trypanosome have been attributed to any one or 
a combination of the following factors: 
mechanical damage, increased vascular 
permeability, deranged immunological 
mechanisms, and/or the activities of 
trypanosome toxins [17]. 
 

The results of the histology tissue indicated that 
treatment with graded doses of either the 
methanol extract of A. boonei or the chloroform 
extract of M. lucida elicited varying effects on the 
kidneys of the infected animals. It was noted that 
Alstonia boonei ameliorated the effect of the 
infection on the kidney of the infected animals 
with the histology sections being comparable 
with those treated with the reference anti-
trypanosomal drug (Diminazine aceturate) and 
the uninfected rats [18]. The M. lucida extract 
was shown to exhibit lower tissue protective 
effect against T. brucei brucei infection when 
compared with A. boonei: in the Morinda lucida 
treated animals, the kidneys revealed gross 
pathologies that were also seen in the untreated 
rats. From the aforementioned postulations by 
[17] on the probable mechanisms of tissue 
damage seen with trypanosome infection, it may 
be inferred that A. boonei act to prevent or 
ameliorate the tissue damages by inhibiting 
several of these mechanisms. The results of the 
phytochemical screening of the methanol extract 
of A. boonei and the chloroform extract of M. 
lucida indicated that A. boonei contained 
phenols, saponins, steroids, and terpenes, in 
addition to alkaloids, tannins, and flavonoids 
which were detected in M. lucida [19,20]. These 
differences might lend credence to the reno-
protective effects observed in the A. boonei 
treated rats. 
 

5. CONCLUSION 
 

Our findings revealed that crude extracts of 
Alstonia boonei and Morinda lucida contain 
variety of phytochemicals which have 
documented antimicrobial effects. The crude 
extract of Alstonia boonei demonstrated 
ameliorative in vivo effects on some histological 
lessions of wistar rats infected with Trypanosoma 
brucei brueci. 
 

CONSENT 
 

It is not applicable. 
 

ETHICAL APPROVAL   
 

Ethical clearance for the use of laboratory 
animals was obtained from Kaduna State 
Ministry of Agriculture and Forestry, Kaduna.  



 
 
 
 

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DISCLAIMER (ARTIFICIAL INTELLIGENCE) 
 

Author(s) hereby declare that NO generative AI 
technologies such as Large Language Models 
(ChatGPT, COPILOT, etc.) and text-to-image 
generators have been used during the writing or 
editing of this manuscript. 
 

COMPETING INTERESTS 
 

Authors have declared that no competing 
interests exist. 
 

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