Abakpa et al., IJAVMS, Vol.

7, Issue 4, 2013: 135-141

ANTI-TRYPANOSOMAL EFFECT OF THE AQUEOUS EXTRACT OF MORINGA

OLEIFERA LEAF AND ASSOCIATED HAEMATOLOGICAL CHANGES IN RATS
INFECTED WITH TRYPANOSOMA BRUCEI
1

Abakpa, S.A.V., 1Daramola, O.O, 2Takeet, M.I 2Akande, A.F. and 3Oyewusi, J. A.
1
Department of Veterinary Medicine and Surgery,
2
Department of Veterinary Microbiology and Parasitology,
3
Veterinary Teaching Hospital,
College of Veterinary Medicine,
Federal University of Agriculture, Abeokuta.
Correspondence: drabakpasimon@yahoo.com
ABSTRACT

The effect of the aqueous extract of Moringa oleifera leaf on parasitaemia and associated
haematological changes were investigated in 36 albino rats of both sexes that were experimentally
infected with Trypanosoma brucei (T.brucei). The effect of the Moringa oleifera extract was compared
with that of Diminazene aceturate. The rats were divided into six groups (A, B, C, D, E and F). Groups
A was not infected and was given normal saline orally. Group B was infected but not treated. Group C
was infected and treated with diminazene aceturate (Nozomil) at the dose rate of 3.5mg/kg. Groups
D, E and F were infected and treated with Moringa oleifera at dose rates of 100mg/kg, 300mg/kg and
500mg/kg, respectively. Diminazene aceturate completely clear the parasitemia day 7, post therapy.
The 100mg/kg dose of Moringa oleifera failed to clear the parasitaemia but insignificantly reduced
(P>0.05) it and remained static until they all died. There was no sinificant difference (P>0.05) between
the rats that were treated with 300mg/kg and those treated with 500mg/kg of Moringa oleifera.
Moringa oleifera only prolonged the lifespan of the rats. There was a significant decrease (P<0.05) in
the haematocrit, erythrocyte count, haemoglobin concentration and platelet count of all the T. brucei
infected groups while, the white blood cells and granulocytes were significantly increased (P<0.05).
We concluded that Moringa oleifera is not trypanocidal but, has anti-trypanosomal effect, and has
insignificant effect on haemopoiesis. Trypanosomosis has significant effect on haemopoiesis.
Key words: Anti-trypnosome, Diminazine aceturate, haematology, Moringa oleifera, rats

INTRODUCTION
Medicinal properties of plants have been investigated in the light of recent scientific developments
throughout the world, due to their potent pharmacological activities, low toxicity and economic
viability, when compared with synthetic drugs1. Interest in medicinal plants as a re-emerging health
aid in the maintenance of personal health and well-being has been fuelled by rising costs of
prescription drugs, and the bio-prospecting of new plant-derived drugs2. The antioxidant effect of
medicinal plants has been suggestive of playing an important role in maintaining physiological levels
of oxygen and hydrogen peroxide and eliminating peroxides generated from inadvertent exposure to

ANTI-TRYPANOSOMAL EFFECT OF THE AQUEOUS EXTRACT OF MORINGA OLEIFERA LEAF

xenobiotics and drugs. Among myriad of plants, Moringa oleifera, commonly called the drumstick, is
a tree native to India, but has been planted and domesticated in many other countries, including
Nigeria. A remarkable surge of interest in chemoprevention research has led to the identification of
many phytochemicals of dietary origin as effective potential chemopreventive agents3. The crude
aqueous extract of the dried leaves contain the same chemical components like that of the fresh leaves
except for the absence of steroids and terpenes4. Recent approaches to alternative therapeutic agents
for treatment of trypanosomosis have focused on plants such as Moringa oleifera and other natural
products5. Trypanosomosis is a hemoprotozoan disease of animals and humans. The effect of
Trypanosoma on certain haematological parameters was studied using dogs and the results showed
decrease in hemoglobin (Hb) concentration and packed cell volume (PCV), while erythrocytes
sedimentation rate was increased6. Despite the high prevalence rate of trypanosomosis, only a few
drugs are available for its treatment. These drugs are old, toxic, expensive and not readily available 7 In
addition, resistances to these major drugs as well as multiple drug resistant populations have been
described for different species of the parasite8,9. Numerous relapses after treatment have also been
reported10.There is urgent need to seek for new sources of therapeutic agents that will address these
shortcomings5. These problems together with the associated expensive nature of most drugs in
developing nations with poor economic status, including endemic nation like Nigeria, make it
pertinent to search for new, better and cheaper trypanocides/trypanostats. This study was to evaluate
the effect of Moringa oleifera in rats experimentally infected with Trypanosoma brucei and the
associated haematological changes.
MATERIALS AND METHODS
Trypanosome parasites
Parasites used were T. brucei isolated from naturally infected cattle. The parasite was identified as T.
brucei by microscopy and specie specific identification carried out using TBR 1&2 primers set for
Polymerase Chain Reaction (PCR).
Experimental Animals
Thirty-six sexually matured albino rats of both sexes with the mean weight of 1802.4g were used.
They were housed in a group of six in a metal cages bedded with wood shavings. Pelleted grower
ration (Guinea feeds Ltd, Benin, Nigeria) and water were provided for the rats ad-libitum The protocol
for this study was approved by the Ethical Committee of the College of Veterinary Medicine, Federal
University of Agriculture, Abeokuta, Nigeria.
Extraction of Moringa oleifera
The leaves of the plant were obtained from from Abeokuta, Nigeria. The leaves were air dried and
mashed. The mashed powder (weighing 450g) was soaked in distilled water (1,000mls) for 24 hours
before extraction. The aqueous solution was filtered using Wartmans filter paper (size 1). The filtrate
was concentrated by evaporation on on a water bath at 600 C. The brownish residue obtained weighing
18.4g, was kept in an airtight bottle in a refrigerator at 4C until used.

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ANTI-TRYPANOSOMAL EFFECT OF THE AQUEOUS EXTRACT OF MORINGA OLEIFERA LEAF

Experimental design
The rats were divided into six (6) groups (A, B, C, D, E and F) with six (6) rats per group. The rats in
Group A were neither infected with T. Brucei nor treated with either Moringa oleifera or Diminzine
aceturate. They given normal saline orally. Group B was infected with T. brucei, but received no
treatment. Group C was infected with T. brucei and treated with diminazine aceturate. The rats in
groups D, E and F were infected with T. brucei and treated with Moringa oleifera extract 100mg/kg,
300kg/kg and 500mg/kg body weight respectively.
Infection of rats with the parasite
Blood samples were collected from an infected rat into Phosphate Buffer Saline. The experimental rats
in groups B, C, D, E, & F were infected intra-peritoneally with the organism at approximately 1 x 103
tryps/ml of blood and the level of pasitaemia was monitored daily using rapid matching method11
Administration of diminazene aceturate and Moringa oleifera
Group C received a single dose of diminazene aceturate intramuscularly at the dose rate of 3.5mg/kg
body weight. Groups D, E and F received the aqueous extract of Moringa oleifera leaf orally once
daily at the dose rate of 100mg/kg, 300mg/kg and 500mg/kg body weight respectively until
termination of the experiment.
Haematological analysis
Blood samples were collected through the medial canthus of the rats on day 13 post infection into
EDTA bottle for complete blood count, using auto-haemo-analysing machine (Mindray).
Statistical Analysis
Data collected was presented as Mean SEM, and analyzed using analysis of variance (ANOVA) and
Dunnet multiple comparism, using Graphpad (Graphpad software Inco., California). .
Results
The dynamism of the parasitaemia in the infected rats in this work is shown in Table 1 below.
Parasitaemia was observed in all the experimentally infected groups of rats on day 3 post-infection
(pi). Peak parasitaemia was established in all the infected rats on day 6 (Table 1). Group A, (negative
control) showed no parasitaemia. The parasitaemia in group B (positive control) remained high until
day 14 when the last rat in the group died. The infected rats treated with diaminazene aceturate (group
C) showed a progressive significant decrease (P<0.001) in the level of parasitaemia from day 3 post
treatment (pt) (day 9 pi on the table) until day 7 pt (day 13pi on the table) when it was completely
cleared. All group C rats survived after treatment with Diminazene aceturate. The parasitaemia in the
infected rats in group D were treated with 100mg/kg of Moringa oleifera remained high without any

Abakpa et al., IJAVMS, Vol. 7, Issue 4, 2013: 135-141

ANTI-TRYPANOSOMAL EFFECT OF THE AQUEOUS EXTRACT OF MORINGA OLEIFERA LEAF

statistically significant change (P>0.05) till all the rats died (Table 1). The parasitemia in the rats
treated with 300 mg/ kg of the Moringa oleifera extract (group E) decreased significantly (P<0.05) on
days 12 pi and 13pi (P<0.001) and started rising again until the last rats in the group died on day 19 pi
(Table 1). The parasitemia in the rats treated with 500 mg/ kg of the Moringa oleifera extract (group
F) progressively decreased significantly (P<0.05) on day 11 dpi, day 12 (P<0.001), 13 dpi (P<0.001)
and began to rise again until day 18 dpi when the last rat in the group died (Table 1). The rats in
groups E and F survived till day 19 and 18 dpi of the experiment respectively (Table 1).
Table 1: Parasitaemia profile in control rats, rats treated with Diminazine aceturate ( Nozomil), and the rats treated with
Moringa oleifera extracts.
Control

DPI=DA
Y POST
INFECT
ION
0-2
3
4
5
6*
7
8
9
10
11
12
13
14
15
16
17
18
19
20

abcde

Grou
pA
Nega
tive
Contr
ol
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00

B
Posi
tive
Cont
rol
0
0.00
2
0.20
3
0.32
6
0.20
6
0.00
6
0.00
6
0.00
a
6
0.00
a
6
0.00
a
6
0.00
a
6
0.00
a
6
0.00
a
6
0.00

Nozo
mil
treate
d rats
C

Moringa oleifera treated rats

3.5
mg/kg

100m
g/kg

300m
g/kg

500m
g/kg

0
0.00
3
0.32
4
0.32
6
0.20
6
0.00
6
0.00
6
0.22
a
5
0.12
b
4
0.12
c
2
0.36
d
1
0.12
e
0
0.00
e
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00
0
0.00

0
0.00
2
0.20
4
0.20
5
0.32
6
0.00
6
0.00
6
0.00
a
6
0.00
a
6
0.22
a
6
0.22
a
6
1.62
a
5
0.00
a
5
0.00

0
0.00
2
0.20
4
0.20
5
0.32
6
0.00
6
0.00
6
0.00
a
6
0.00
a
6
0.00
a
6
0.00
b
5
0.22
c
4
0.12
b
5
0.43
a
6
0.33
a
6
0.33
a
5
0.33
a
6
0.00
a
6
0.00

0
0.00
3
0.32
4
0.20
5
0.32
6
0.00
6
0.00
6
0.00
a
6
0.00
a
6
0.22
b
5
0.22
c
4
0.12
d
3
0.24
c
4
0.37
b
5
0.27
a
6
0.35
a
6
0.00
a
6
0.00

Values in the columns with different superscripts are significantly different at between P<0.001 - P<0.05.

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ANTI-TRYPANOSOMAL EFFECT OF THE AQUEOUS EXTRACT OF MORINGA OLEIFERA LEAF

*Treatment day

The white blood corpuscles (WBC) were increased in all the infected groups with a significant
difference (P<0.05) between the untreated rats (group B) and the negative control (group A) or the
treated groups (Table 2). The granulocytes were increased all the infected but with significant
difference (P < 0.05) between the untreated rats and the rats treated with diminazene aceturate or those
treated with 300mg/kg of moringa oleifera extract (Table 2). The lymphocytes and monocytes were
increased in all the infected groups without a significantly difference (P > 0.05). The red blood cells
(RBC), haematocrit and platelet counts were significantly decreased (P < 0.05) compared to that of the
negative control

Table 2: Haematological profile of rats before and on day 13 post infection

Haematological response of rats on day 13 post infection
Parameters

Baseline Reference

Group A

Group B

Group C

Group D

Group E

Group F

15.99.32

a16 1.04

b61 15.83

a12 1.59

a27.1 5.80

a24.9 4.52

a27.6 4.52

11.54.20

11.4 1.02

52.9 14.54 17.4 1.05

21.1 4.74

18 9.21

21.9 4.31

0.70.25

0.9 1.31

1.5 0.33

0.5 0.09

1.0 0.25

0.85 0.45

0.8 0.15

5.21.21

a5.0 1.31

a6.6 1.05

b4 0.72

a5.1 0.83

b3.9 1.91

a4.9 0.79

RBC (x 10 /L)

8.83.42

b8.88 0.31 a5.25 0.70 a6.69 1.32

b8.07 0.97

a4.63 1.52

b8.14 0.97

HGB (g/L)

139.817.10

b139 4.58

b114 18.57

b125 18.24

a56.5 16.54

b120 14.03

42.82.13

b43.6 1.74 a25.1 2.55 b36.1 6.25

b41.4 0.07

a22.4 7.52

b41.3 4.34

511.532.42

b507 21.47 a222 55.18 b399.0 55.04 a172.0 35.20 b329.0 34.21 b453.0 81.8

WBC (x 109/L)
9

LYMPH(x 10 /L)
9

MONO (x 10 /L)
9

GRAN (x 10 /L)
12

HCT (%)
9

PLT (x 10 /L)

a76 7.89

A= uninfected group, B= Infected but untreated group, C = infected and treated with Diminazene aceturate, D =
infected and treated with 100mg/kg Moringa oleifera extract group, E = infected and treated with 300mg/kg
Moringa oleifera extract group and E = infected and treated with 500mg/kg Moringa oleifera extract group.
a,b

Values in the rows with different superscripts are significantly different at between P<0.001 P<0.05.
DISCUSSION
In this study, parasitaemia was observed in the rats on day 3 and peak and peaked day 6 post infection.
This is in agreement with the work of Ameh12 who reported parasitemia within 3 days and peak
parasitemia 5-6 days post infection. Trypanosomosis is mostly an acute disease that presents anaemia
as a major manifestation in various animals. In this study, the red blood cells (RBC), haematocrit

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ANTI-TRYPANOSOMAL EFFECT OF THE AQUEOUS EXTRACT OF MORINGA OLEIFERA LEAF

value, haemoglobin(HGB) concentration and platelet count of the infected but untreated rats decreased
significantly (P<0.05). This is consistent with the findings of different authors that reported decreased
in haematological values in rats13,14 , in dogs15 and in red fronted gazelles (Gazella rufifrons)16 in T.
brucei infection. The rats that received the aqueous extracts of Moringa oleifera or diaminazene
aceturate had an insignificant decrease (P>0.05) in their the red blood cells (RBC), haematocrit value,
haemoglobin (HGB) concentration and platelet count. This may probably be due to pathological
effects of the disease before the commencement of treatment, which failed to resolve within the period
of 7 days therapy. . In this study, although the doses of 100mg/kg, 300mg/kg and 500mg/kg produced
poor efficacy, the life span of the rats were prolonged but they died on days 15, 20 and 19 post
infection (ie, days 8, 13 and 12 post treatment) respectively. This finding correlates with that of
Atawodi17 who reported that the doses of 100mg/kg and 200mg/kg of Moringa oleifera produced poor
efficacy and the rats died day 10 post-infection. The lives of the rats that were prolonged in this study
could have probably been due to the effect of flavonoids which are scavengers of free fatty acids
which are the initiators of the pathological effects of trypanosomosis.
Conclusion
Based on our study, aqueous extract of Moringa oleifera has no trypanocidal effect but has the
potential to prolong the life span of rats due to its antioxidant activities. We also concluded that the
pathological effects of trypanosomosis as seen in the haematological result cannot be resolved
immediately after the clearance of parasitemia.
Recommendation
Further study is needed for the use of higher doses, frequency of use or otherwise effects of prolonged
use of the extracts of Moringa oleifera on trypanosomosis.
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