Toxicological Effects of Oral Consumption of Aqueous Leaves ...

Dutse Journal of Pure and Applied Sciences (DUJOPAS) Vol. 3 No. 1 June 2017

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Toxicological Effects of Oral Consumption of Aqueous

Leaves Extract of Senna occidentalis on Experimental

Animals (Albino Rats)

Bello Aminu Bello Department of Biochemistry,

Federal University Dutse,

Jigawa State

Jalaluddin Awllia Khan Department of Biochemistry,

King Abdul-Aziz University, Jeddah,

Kingdom of Saudi Arabia

Ibrahim Khalil Adam Department of Biochemistry,

Federal University Dutse,

Jigawa State

Muhammad Inuwa Hassan Hematology Department,

Muhammad Abdullahi Wase Specialist Hospital,

Kano Nigeria

Fatima Sulaiman Abdullahi Food science and Technology,

K.U.S.T. Wudil, Kano State

Abstract enna occidentalis is among the shrubs that are widely distributed in the temperate or warm

regions across the globe. The leaves of this plant were reported to exhibit abroad

pharmacological effects including; antibacterial, antimalarial, antifungal as well as

anticancer activities. This study accounts for the toxicological effects of oral administration of

the aqueous Senna occidentalis leaves extract on organ body weight ratio, some biochemical,

hematological as well as histopathological parameters in male albino rats. The results indicate

a significant decrease in organ body weight ratio for liver, kidney, spleen, pancreas, lungs and

heart in the test groups. The result also revealed a significant increase in the hematological

S


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parameters, indicating that, the extract burst the hematopoietic capacity of the animals. The

administration of low dosage of the extracts does not remarkably alter the liver as well as the

renal function parameters studied as compared to control group, indicate that the extract does

not pose any liver and renal damage. Therefore, the aqueous leaves extract of Senna occidentals

can be considered as hematopoietic stimulator and does not have effect on vital organs.

Key words: Senna occidentalis, aqueous extract, toxicity, liver function, renal function

Introduction

The plant Senna occidentalis (Fabaceaecaesalpiniaceae) known commonly as coffee senna is widely

grown in warm areas of the world, with exception of Australasia. The plant is a shrub in nature

and was reported to be a native of America and mainly used for landscape flowering purposes

(Hussein, 2003). Coffee senna is also reported to be used as a substitute to coffee. The seeds of the

shrub are brewed into the coffee-like beverage and used for the treatment of asthma. The

pharmacological effect of the leaf extracts were previously reported to have a broad spectrum of

antibacterial, antimalarial and antifungal activities (Caceves et al., 1991; Perez and Suarez, 1997;

Tona et al., 1999). The leaves extract were also reported to possess antimutagenic (Sharma et al.,

2000), antiplasmodial (Tona et al., 2004), anticarcinogenic (Vashishtha et al., 2009), and

hepatoprotective activity. It was also been used for the treatment of stomach disorders,

rheumatism, and some liver diseases (Sara et al., 1994; Jafri et al., 1999). The leaves are commonly

used as a leaf vegetable and are eaten either raw or in a mixture with coconut, chilli, and onion

(Selvam, 2007; Vashishtha et al., 2009). Currently, in Kano, Nigeria the leaves of Senna occidentalis

are sold in market for its medicinal values. The leaves were either soaked and boiled in water and

taken for the treatment of typhoid fever, malaria and some form of cancers or the fresh leaves are

rubbed on the tumor. In view of the above, there is an urgent need to assess the toxicological

effect of the aqueous extract of the leaves in order to validate its traditional uses.

Materials and Methods

Collection of Plant Materials and Extraction

The leaves of Senna occidentalis(Fabaceaecaesalpiniaceae) were obtained from a farm in Wudil

town, Kano state, Nigeria, in the Month of January, 2016. The leaves of the plant were identified

and the identity of the leaves was authenticated in the Department of Botany, Ahmadu Bello

University Zaria, Kaduna State, Nigeria, by Mr. Namadi Sunusi. The leaves were given a voucher

number of 1047 and was deposited in their Herbarium for future reference.

The leaves were dried under shade in the absence of sunlight for a period of two weeks and were

grinded to powder using mortar and pestle. The dried powdered leaves (400g) was homogenized

with hot (70oC) distilled water (200 cm3) and kept for two days with regular stirring. The aqueous

mixture formed was filtered using Whatman no. 1 filter paper and then, the filtrate was


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evaporated or concentrated using rotary evaporator. The concentrated extract was then freeze-

dried and the dried powder of aqueous leaves extract was obtained. The dried powdered extract

was kept in refrigerator at -20oC for until use.

Phytochemical Analysis

The total alkaloid, saponin, flavonoid, tannin and phytate of the dried powdered aqueous leaves

extract of Senna occidentalis were determined using standard methods (Sofowora, 1993).

Experimental Animals

The experimental animals used in this work are albino rats and were purchased from animal

house of the Department of Zoology, Bayero University Kano, Nigeria. A total of 25 adult male

rats aged 6-8 weeks and weighing 100-160g, were housed under a standard laboratory living

conditions with 12 hours dark/light, with full access to standard animal feed (Pelletised growers

feed, Zawan roundabout, Jos, Plateau State, Nigeria) and water ad libitum was also provided.

The albino rats were divided into five groups of five rats each based on weight and concentration

of extract to be taken and were used after one week of acclimatization.

The albino rats used were fasted for a period of 15 hours (5.00pm- 8.00 am) before the beginning

of the experiments but were all allowed free access to water. These animals were handled with

strict compliance to international guidelines as reported by the Canadian council on the care and

uses of experimental animals (1993).

Toxicological Studies

The rats were divided into five groups of five animals each, a concentration of 25mg/kg body

weight was received by group one, 50mg/kg/day received by group two, 100mg/kg/day

received by group three and group four received 200mg/kg/day. The fifth group serves as

control and received only 1ml of physiological saline daily.

Administration of Plant Extract and Collection of Blood Samples

The aqueous leaves extract of S. occidentalis was found to bcompletely soluble in water and

corresponding daily dose of each group was orally administered to the rats for a period of six

weeks. At the end of the experimental period, under a mild chloroform anesthesia, all the rats

were sacrificed and 5cm3 blood samples were collected from each rat with aid of cervical

decapitation. One cm3 of the blood was immediately taken out and mixed with anticoagulant

(EDTA) for hematological analysis. The remaining 4 cm3 of the blood were allowed to clot and

then centrifuged at 3000 rpm for 10 minutes. The serum was separated from the cells and was

used to determine the liver and renal function’s parameters (Biochemical analysis). The liver,

kidney, pancreas, spleen, lungs and heart of each rat were carefully dissected (Mikel, 1994) from

the animal and the remnant’s blood was wiped out with filter paper and weighed. At the end of


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dissection the liver and kidneys after weighing were immediately immersed in 10% neutral

buffered formalin for histopathological studies.

Determination of Relative organ weight

The organ weight in relation to body weight of the rats is the percentage of the ratio of absolute

organ weight to the body weight of animal on sacrifice day (Yakuba et al., 2008).

Determination of Hematological Parameters

The percentage hematocrits (HCT%) was determined according to method described by

Sanderson (1981), while the RBC, Total WBC and platelet counts were determined according to

standard methods.

Determination of Biochemical Parameters

The liver function indices such as Aspartate aminotransferase (AST), Alanine aminotranferase

(ALT), and Alkaline phosphatase (ALP) were determined using Randox kits as described by the

manufacturer, while the total and direct bilirubin were determined by the method described by

Jendrassik and Grof (1938).

The renal function parameters determined include the serum concentration of chloride

(Schoenfeld and Lowellen, 1964), Bicarbonate (Van slyke method), urea (Veniamin and Vakirtzi,

1970), Creatinine (Jaffe, 1886), uric acid (caraway method)while the sodium and potassium ion

concentrations were determined by flame photometry.

Histopathology Test

The biopsies of the liver and kidney of the albino rats were fixed in 10% formal saline, dehydrated

with ascending grades of alcohol, cleared with toluene, infiltrated with molten paraffin wax. The

microtome sections (approx. 5mm) were stained with hematoxylin and eosin (H and E) technique

and structure was examined under electron microscope (Auwioro, 2010).

Statistical Analysis of Data

The results of this work were all expressed as Mean values ±SD and the statistical variations

among the groups were evaluated by one way analysis of variance (ANOVA) at p value of less

than 0.05.

Results

The result of the phytochemical screening, hematological test parameters, organ body weight

ratio and liver and renal function test parameters are presented in the form of mean and standard

deviation of three determinations as shown in Table 4. The histopathological analysis is in the

form of stained picture of the respective test tissue section and are shown in Figure 6.


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Phytochemical Screening

The aqueous leaves extract of Senna occidentalis was tested for some common phytochemical

constituents, the result revealed the presence of saponins, flavonoids, tannins, alkaloids and

phytate as shown in Table 1.

Hematological Indices

The result for the hematological test revealed that, there were significant differences in the values

for WBC, RBC,HCT% and PLT% for all groups when compared to their corresponding values in

control group (p<0.05)

Liver Function Test

The result in the form of mean and standard deviation of the liver function indices (Aspartate

aminotransferase, Alanine aminotransferase, alkaline phosphatase and bilirubin (total and

direct)) were shown in Table 4. There was significant difference in AST of group II, III and IV

when compared with control group (p<0.05) but no significant difference exist between group I

and control group (p>0.05).

The result of this work also shows a significant difference in values of AST and ALT for all the

test groups when compared with their corresponding values in the control group (p<0.05) with

exception in AST values for group I in which no significant difference exist when compared to

corresponding values in control group (p>0.05).

The values for ALP in group I, II and IV were found to be significantly different when compare

to the corresponding values in control group (p<0.05) but those from group III were found to be

significantly similar to those of control group as depicted in Table 4 respectively.

The values for total and direct bilirubin across the test groups found indicate a significant

variation when compared to the corresponding values in control group (p<0.05) with exception

for the values of total bilirubin in group II and IV (P>0.05) in which no significant difference exist

when compared to their corresponding values in control group as shown in Table 4.

Renal Function Indices

The result for the renal function indices shows that, there were significant differences in the

values of Na, K and chloride for all the test groups when compared to the corresponding value

in control group (p<0.05) with exception for the values of Na and chloride from group IV and K

from group I as shown in Table 5 respectively.

Histopathology of Liver and Kidney

The result for liver and kidney histology revealed that, there were no remarkable liver and renal

damage for the liver and renal section from the group I, II and III that received mild doses of the

extract but mild hepatocyte degeneration and renal tubule dilatation were detected in group IV

that received the highest extract concentration.


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Table 1: The quantitative phytochemical constituents of aqueous leaves extract of Senna occidentalis

Phytochemicals (%) Mean ± SD

Phytate (%) 0.18±0.01

Alkaloids (%) 3.99±0.13

Tannins (%) 1.05±0.07

Flavonoids (%) 76.92±0.44

Saponins (mg/L) 13.17±0.04

Table 2: Effect of aqueous leaves extract of Senna occidentalis on organ body weight ratio in albino rats

Organ Control group

(Normal saline)

Group I

(25mg/kg)

Group II

(50mg/kg)

Group III

(100mg/kg)

Group IV

(200mg/kg)

Liver

4.01

3.29*

3.30*

3.29*

2.96*

Kidney 0.74 0.67* 0.65* 0.70 0.67*

Spleen 0.46 0.41* 0.49* 0.36* 0.35*

Pancreas 0.38 0.32* 0.30* 0.32* 0.30*

Lungs 2.06 1.13* 1.15* 1.04* 1.21*

Heart 0.48 0.41* 0.46 0.47 0.41*

Note: Values are expressed as mean of three determinations ±SD, * stance for significant difference from the control

(p<0.05)

Table 3: Effect of aqueous leaves extract of Senna occidentalis on some Hematological parameters in albino rats

Conc. of extract

(mg/kg) WBC x109/L RBCX1012/L HCT % PLT X109/L

Group I (25)

9.2±0.21 *

4.6±0.04*

32.5±1.41*

374.5±4.95*

Group II (50)

9.7±0.74*

6.2±0.03*

42.7±0.64*

1214.5±0.70*

Group II (100)

10.9±0.54*

11.3±0.69*

66.2±1.69*

866.0±5.65*

Group IV (200)

11.9±0.08*

15.2±1.23*

93.0±0.14*

1350.0±7.07*

Control

3.2±0.21

8.4±0.03

63.6±0.78

942.5±0.71


(p<0.05)


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Table 4: Liver function indices for albino Rats administered with aqueous leave extract of Senna occidentalis

Conc. Extract

(mg/kg)

AST

(U/L)

ALT

(U/L)

ALP

(IU/L)

Bilirubin Total

(mg/dl)

Bilirubin Direct

(mg/dl)

Group I (25)

22.0±2.83 27.5±0.71* 34.0±1.41* 7.5±0.71* 2.75±0.35*

Group II (50)

25.0±0.07* 30.5±0.78* 34.5±0.71* 8.5±2.12* 3.02±0.24*

Group III (100) 29.5±3.54* 41.0±0.55* 46.0±1.41 10.5±0.71 3.5±0.71*

Group IV

(200) 36.5±2.12* 43.0±0.71* 50.0±4.24* 12.0±0.99 3.5±2.12*

Control 22.9±0.14 69.5±3.54 46.0±5.66 10.5±2.12 1.2±0.28


(p<0.05)

Table 5: Renal function indices for albino Rats administered with aqueous leave extract of Senna occidentalis

Conc.

Extract

(mg/kg)

Sodium

(mEq/L)

Potassium

(mEq/L)

Chloride

(mEq/L)

Bicarbonate

(mEq/L)

Urea

(mg/dl)

Creatinine

(mg/dl)

Group I

(25)

156.5±3.53* 9.8±1.20 70.5±3.53* 8.55±0.071* 5.9±0.28* 25.8±2.12*

Group II

(50) 160.5±0.71* 10.1±1.13* 69.0±1.41* 11.1±1.20* 5.25±0.071* 34.0±4.24*

Group III

(100) 161.5±0.71* 10.6±2.40* 64.5±2.83* 10.65±4.9* 6.2±0.64* 30.3±0.71*

Group IV

(200) 162±1.41 13.2±2.89* 65.5±0.71 8.1±1.06* 5.9±0.28* 42.5±7.07*

Control

164.5±0.71 9.7±0.21 65.5±2.12 12.5±0.71 5.5±0.50 25.1±0.07


(p<0.05)


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Figure 1: Effect of aqueous leaves extract of Senna occidentalis on the histology of liver and kidney of the albino rats

Discussion

The results of the phytochemical screening, organ body weight ratio, hematology, biochemical

and histopathological analysis of the aqueous leave extract of Senna occidentalis are descibed. The

result of the phytochemical analysis of aqueous leave extract of Senna occidentalis revealed the

presence of the following constituents; alkaloids (4%), tannins (1.1%), flavonoids (76.9%),

saponins (13.2%) and phytate (0.2). In primary healthcare system, the use of traditional medicine

especially in rural and developing states for the curing of disease plays an important role because

of the general belief that, herbs are natural and are always safe (Seyyed et al., 2015). The oral

consumption of any herbal preparation for the sake of curing any aliment without any proper

prescription coupled with limited scientific studies on the safety of the traditional medicine raises

a strong concern on their possible toxicity (Saad et al., 2006).

The leaves extract is rich in some phytochemicals which may be responsible for its used in treating

diseases and that, at low concentration, the aqueous leave extract does not cause any toxic effect

on the organs and tissues of the albino rats used.These phytochemicals; alkaloids, tannins,

flavonoids, saponins and phytate couldaccount for the traditional use of the extract in managing

some diseases. This finding is supported by the fact that, alkaloids and their synthetic derivatives

are used as anti-analgesic, antimalarial, antiseptic as well as anti-bacteria (Evans, 2002). The

finding is also in line with the fact that, saponins are used as antifungal and antibacterial (Okwu

and Emenika, 2006). Therefore, the presence of alkaloids, saponins, flavonoids and tannins in the


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aqueous extract of Senna occidentalis can qualify the extract to have natural antibiotic (Dimaro et

al., 2007), as well as anti-inflammatory properties (Quiroga et al.,2001).

The organ body weight ratio is defined as the ratio of the organ weight to that of the body weight

of the organism. This ratio gives an idea of whether the extract administered interact with the

various organs favorably or unfavorably. The result of the organ body ratio revealeda gradual

decrease in the ratio with increase in extract concentration. This may be as a result of unfavorable

interaction (cellular constriction and inflammation) of the extract or its component with the

respective organs. This result is not in agreement with the fact that, aqueous leave extract of

Passiflora edulis posed no any effect on the ratio in experimental animals (Devaki et al., 2012;

Schmidst et al., 2007)

The effect of aqueous leaves extract of Senna occidentalis on WBC, RBC, HCT% and PLT counts of

albino rats were studied. The result reveals a significant increase in all the parameters determined

with increase in the concentration of the extract administered as compared to the control group.

These clearly indicate that, the aqueous leave extract may possess component that stimulates the

hemopoeisis in the albino rats. The rate of stimulation increases in dose-dependent manner. The

result is not in line with that reported by Appidi et al. (2009) that, oral administration of aqueous

leave extract of Hermania incana results in mild decrease in some hematological index in rats.

Some aqueous leave extract of such as Passiflora edulis produce no any change in hematological

parameters of rats (Devaki et al., 2012).

The results of the effect of various concentrations of the extract on liver function parameters

indicated no significant difference in the value of AST in group I that received 25mg/kg extract

as compared to the control group. However, there was significant decrease in the values of ALT,

ALP and bilirubin as compared to these values from the control group. The values for all the liver

parameters studied were found to increase in a dose-dependent manner. Therefore, at low

concentration, the extract posed no any toxicity to liver. The increases in these values correspond

to the increase in membrane permeability of the liver caused by the toxicant. These result are in

line with the fact that, liver toxicity may lead to a compromised membrane integrity (Yakubu et

al., 2003). The compromised membrane integrity may also have negative effect on the metabolism

and regulation of amino acid by liver. The elevated levels of ALP with extract concentration may

also be due to the fact that the extract may possess component that can stimulate the enzyme

synthesis in the liver (Emelike and Dapper, 2013). The significant increase in total and direct

bilirubin in comparison with control is due to reduction of its uptake by the liver arising from the

toxicity of the extract at high doses. This is in line with the fact that liver damage may impaired

its function (Ashafa et al., 2010). The result of liver function test is generally in conformity with

the fact that, the liver markers may tend to increase due to toxic effect of high dose of aqueous

Hibiscus sabdariffa extract (Yakubu et al., 2001).


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The kidneys play a vital role in body homeostasis and are primary to the excretion of waste

products of metabolism. The regulation of intracellular volume, electrolyte and acid-base balance

depend solely on kidneys (Orisakwe et al., 2003), therefore, toxic insult of the kidneys could affect

the total body metabolism (Goldstein et al., 1999). The result of the possible effect of oral

administration of aqueous leave extract of Senna occidentalis was shown on Table 5, which

reveals that, there was significant change in serum levels of sodium, potassium, bicarbonate and

chloride as well as creatinine and urea as compared to that of control group. The assessment of

the effect of the oral administration on the level of the excretory metabolites such as the

electrolytes, urea and creatinine can aid in evaluating the integrity of the kidney functions

(Adebayo et al., 2003; Yakubu et al., 2003). Creatinine is a hydrolysed product of creatine

phosphate and its serum level is widely used to assess renal function while urea is a nitrogenous

waste product of protein and amino acid metabolism which can also be used as marker for renal

function test (Adedapo et al., 2009). The significant increase in the serum levels of creatinine and

urea at high concentration of the extract indicate a significant change as compared to the control

group. These suggest that, the integrity of the kidney is compromised by administering high dose

of the extract and the result is in line with the resultreported by Orisakwe et al. (2004) and

Abubakar et al. (2010).

The result of the present stud reveals a significant decrease in the level of serum sodium in the

groups that receives the various doses of the extract as compared to that for the control group.

These suggest that, the extract may possess component that stimulate the function of the sodium

pump and the extract do not contribute to the serum sodium level. This result did not agree with

the report by Adigun et al. (2006) that, aqueous extract of Hibiscus sabdariffa is rich in sodium

ion, implying that, rats consuming the Hibiscus sabdariffa may be increasing their oral sodium load

and hence the plasma level.

The serum potassium level increases significantly with increase extract concentration

administered as compared to that from the control group. This is in contrast with report on

aqueous Hibiscus sabdariffaby Ukoha et al. (2015), the serum potassium in rat decreases following

the oral administration.

The significant increase in serum level of chloride and bicarbonate following the administration

of aqueous leave extract of Senna occidentalis at various high doses may be a sign of tubular and

glomerular dysfunction. This observation is in line with the report of Ukoha et al. (2015) that,

serum chloride and bicarbonate increase with increase in doses of Hibiscus sabdariffa in rats.

The result of the effect of oral administration of various doses of aqueous leaves extract of Senna

occidentalis for a period of four weeks on liver and kidney tissues was assessed using the general

tissue structure studies (H and E) and the result was shown on Figure 6. It revealed that, there

was no much changes in the features of the liver and the kidney at low concentration of the

extract, little changes were detected at high extract concentration. The result is in conformity with


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the report of Seyyed et al. (2015) that, aqueous extract of Asafarida caused little degenerative

changes in hepatocyte but no prominent effect on the kidneys tissues of the rats. This is also in

agreeth with Ukaho et al. (2015), that chronic consumption of aqueous Hibiscussabdariffa calyx

might be toxic to kidneys of the rats.

Conclusion

The study of the toxicological effect of oral administration of aqueous leaves extract of Senna

occidentalis on organ body weight ratio, some hematological, biochemical and histopathological

parameters of male albino rats for a period of four weeks reveals that, at low concentration no

pronounced toxic effect was detected on the rats. Administration of high concentration of the

extract may cause some toxicity to the animals and that; the extract seems to stimulate the rate of

blood formation by the rats.

Competing interests

The author hereby declared that, no competing interest exists.


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