DEPRESSANT, ANTICONVULSANT AND ANTIBACTERIAL ACTIVITIES OF HIPPOCRATEA AFRICANA

Inter. J. of Phytotherapy / Vol 4 / Issue 3 / 2014 / 144-153.

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e - ISSN - 2249-7722

Print ISSN - 2249-7730

International Journal of Phytotherapy

www.phytotherapyjournal.com

DEPRESSANT, ANTICONVULSANT AND ANTIBACTERIAL

ACTIVITIES OF HIPPOCRATEA AFRICANA

*Jude E. Okokon1, Koofreh Davies

2, Bassey S. Antia

3, Patience J. Okokon

1

1Department of Pharmacology and Toxicology Faculty of Pharmacy, University of Uyo, Uyo, Nigeria.

2Department of Physiology, Faculty of Basic Medical Sciences, University of Uyo. 3Department of Chemistry, Faculty of Sciences, University of Uyo, Uyo, Nigeria.

INTRODUCTION

Hippocratea africana (Willd.) Loes.ex Engl.

(Celastraceae) syn. Loeseneriella africana (Willd.)

N.Halléis a green forest perennial climber without hairs

(glabrous), reproducing from seeds [1]. It is commonly

known as African paddle-pod. The Ibibio tribe of Nigeria

calls it „Ebaenangenang‟. The plant is widely distributed

in tropical Africa. The root of the plant is used

traditionally by the Ibibios of the Niger Delta region of

Nigeria in the treatment of various ailments such asfever,

convulsion, malaria, body pains, diabetes and diarrhea

[2]. Ethnobotanical survey revealed that decoction of the

plant‟s root is also use as an antidote or antipoison to treat

liver and inflammatory diseases such as jaundice and

hepatitis [3-5].The plant (root) has been reported by

Okokon et al [2] to possess in vivo anti-plasmodial

activity. Other biological activities include; anti-

inflammatory and analgesic [6], anti-diarrheal, antiulcer

[7], anti-diabetic and hypolipidemic activities

Corresponding Author:-Jude E. Okokon Email: [email protected]

ABSTRACT

Hippocratea africana (Willd.) Loes.ex Engl. (Celastraceae) is used locally to treat convulsion and

microbial infections. The root extract of Hippocratea africana was evaluated for depressant effect and antimicrobial

activity.Depressant activity was evaluated using open field, force swimming, tail suspension tests and

phenobarbitone-induced sleeping time. Anticonvulsant activity was testedagainst pentylenetetrazol and

aminophylline-induced convulsions. The root extract/fractions were screened for antimicrobial activity against

some typed and pure cultures of bacterial and fungal species using Plate -hole diffusion methodon Mueller – Hinton

agar (MHA) for bacteria and Sabouraud Dextrose Agar (SDA) for the fungi. Minimum Inhibitory Concentrations

(MICs) of active test samples were determined. The root extract decreased significantly (p<0.01) the line crossing,

walling and rearing activities in open field test and increased (p<0.001) the immobility time in force swimming and

tail suspension tests. The root extract significantly (p<0.001) shortened the onset time of sleep and prolonged the

duration of sleep induced by phenobarbitone sodium. The root extract and fractions (200 – 600 mg/kg) were found

to delay significantly (p<0.05 – 0.001) the onset of tonic/clonic convulsion and prolonged the time of death of the

treated mice in PTZ- and aminophylline induced convulsions. The crude extract, chloroform and aqueous fractions

were active against Staph aureus, and B. subtilis. P. aeruginosa was sensitive to aqueous fraction only. The extract

and fractions were inactive against any fungal isolate. The root extract of H. africana has depressant,

anticonvulsant, sedating and antibacterial activities.

Key words: Hippocratea africana, Depressant, Anticonvulsant, Antibacterial, Convulsion, Sedation.

http://www.phytotherapyjournal.com/


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[8], cytotoxicity against HeLa cells, anti-oxidative burst

and anti-leishmanial activities [9] and hepatoprotective

activity [10]. In this study, we report the anticonvulsant

and antibacterial activities of the root extract of

Hippocratea africana.

MATERIALS AND METHODS

Plant materials

Fresh roots of H.africana were collected in

August, 2014 at a forest in Uruan, Akwa Ibom State,

Nigeria. The plant was identified and authenticated by

Dr. Margaret Bassey, a taxonomist in the Department of

Botany, University of Uyo, Uyo. Nigeria. Herbarium

specimen was deposited atFaculty of Pharmacy

Herbarium.

Microorganisms

Typed and pure cultures of some bacterial and

fungal species were obtained from Pharmaceutical

microbiology unit of the Department of Pharmaceutics

and Pharmaceutical Technology, Faculty of Pharmacy,

University of Uyo, Uyo and maintained at 40oC on

nutrient agar plates before use.

Extraction

The root materialswere washed and shade-dried

for two weeks. The dried plants‟ materials were further

chopped into small pieces and reduced to powder. The

powdered material was soaked in 70% ethanol. The liquid

filtrate was concentrated and evaporated to dryness in

vacuo 40C using rotary evaporator. The crude ethanolic

extract (10 g) was partitioned with a 50:50 mixture of

distilled water and chloroform. The aqueous fraction was

evaporated to dryness in a water bath at 60OC and the

chloroform fraction air-dried. The ethanolic extract, the

aqueous and chloroform fractions were stored at -4OC

until used.

Animals

The animals (Swiss albino mice of either sex)

that were used for these experiments were obtained from

University of Uyo animal house. The animals were

housed in standard cages and were maintained on a

standard pelleted feed (Guinea feed) and water ad libitum.

Permission and approval for animal studies were obtained

from College of Health Sciences Animal Ethics

committee, University of Uyo.

Evaluation of Depressant activity

Open Field test

Rats were randomly divided into groups of 5 rats

each and treated as follows for 5 days before open field

test; control (normal saline, 2 ml/kg p.o.), imipramine

(5.0 mg/kg, p.o.) and ethanolic root extract of

Hippocratea africana (200, 400 and 600 mg/kg, p.o.).

The open-field arena was made of acrylic (transparent

walls and black floor, 30 × 30 × 15 cm), divided into nine

squares of equal areas. The open field was used to

evaluate the exploratory activity of the animal [11]. The

observed parameters were the number of squares crossed

(with the four paws) and number of grooming and rearing,

recorded for 5 min testing period.

Forced Swimming test

Mice were randomly divided into groups of 5

mice each and treated as follows for 5 days before the

behavioural test; control (normal saline, 2 ml/kg p.o.),

imipramine (5.0 mg/kg, p.o.) and ethanolic root extract of

Hippocratea africana (200, 400 and 600 mg/kg, p.o.). For

assessing antidepressant activities, we employed the

method described by Porsolt et al [12,13]. The

development of immobility when mice were placed inside

an inescapable cylinder filled with water reflects the

cessation of persistent escape-directed behavior. Briefly,

mice were individually placed in a circular tank (46 cm

tall × 20 cm in diameter) filled with tap water (25 ± 1°C)

to a depth of 20 cm and left there for 5 min. During this

period, the behavior of the animals was recorded by an

observer. Mice were considered immobile when remained

floating without struggling and making only slight

movements necessary to maintain the head above the

water.

Tail Suspension test (TST)

Mice of either sex were randomly divided into

groups of 5 mice each and treated as follows for 5 days

before open field test; control (normal saline, 2 ml/kg

p.o.), imipramine (5.0 mg/kg, p.o.) and ethanolic root

extract of Hippocratea africana (200, 400 and 600 mg/kg,

p.o.). The total duration of immobility induced by tail

suspension was measured according to the methods

described by Steru et al [14]. Briefly, mice both

acoustically and visually isolated were suspended 50 cm

above the floor by adhesive tape placed approximately

1 cm from the tip of the tail. Immobility time was

recorded during a 6 min period. Mice were considered

immobile only when they hung passively and were

motionless.

Anticonvulsant activity

Pentylenetetrazol induced Convulsion

Anticonvulsant effect of the extract was assessed

using a modified method of Vellucci and Webster [15] on

overnight fasted mice. The mice were divided into seven

groups of six animals each and treated with 200, 400 and

600 mg/kg of the root extract, 400 mg/kg of chloroform

and aqueous respectively, phenytoin, 40 mg/kg one hour

before induction of convulsion. Seizure was induced in

each set of mice with PTZ (70 mg/kg i.p). Control group

received normal saline. The onset of Clonic/tonic

convulsion and the mortality rate was recorded and

compared with the respective control group. The ability of


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the plant extract to prevent or delay the onset of the hind

limb extension exhibited by the animals was taken as an

indication of anticonvulsant activity [16].

Aminophylline-induced Convulsion

The extract and fractions were evaluated for

aminophylline –induced convulsion using the method of

Juliet et al [17].The mice were divided into seven groups

of six animals each and treated with 200, 400 and 600

mg/kg of the extract 400 mg/kg of chloroform and

aqueous respectively, phenytoin, 40 mg/kg one hour

before induction of convulsion. Seizure was induced

using aminophylline (280 mg/kg,i.p). The animals were

observed for 120 mins after the administration of AMPH

and the following parameters were noted:

1. Time to onset of myoclonic jerks in mins.

2. Time to onset of tonic convulsions in mins.

3. Time to death during experimental time of 120 mins.

4. Number of mice dead/alive at 24 hours.

Effect on phenobarbitone –induced sleeping time of

rats

The crude ethanolic extract was evaluated for

effect on phenobarbitone sodium sleeping time of rats.

The rats were divided into five groups of five rats each

(n=5). The extract (200, 400 and 600 mg/kg) was

administered to various groups of rats, diazepam (2

mg/kg) was given to the reference group and the control

group was given distilled water (10 ml/kg). After 30 min

the groups were treated with phenobarbitone sodium (40

mg/kg,i.p). The onset and the duration of sleep were noted

and recorded in minutes.

Evaluation of antibacterial and antifungal activities of

the extract

Plate-hole diffusion test

The evaluation of antimicrobial activity of the

extract/fractions were carried out by the Plate -hole

diffusion method [18] on Mueller – Hinton agar (MHA)

for bacteria and Sabouraud Dextrose Agar (SDA) for the

fungi. Solutions of the extract and fractions were prepared

in 10% Tween 80 to concentrations of 100, 50, 25 and

12.5mg/ml.

The innocula of the microorganisms were

prepared separately from 12h broth cultures (Mueller-

Hinton broth for bacteria and the Sabouraud dextrose

broth for the fungi) and incubated at 37C. All culture

media and distilled water were sterilized at 121C for 15

min in an autoclave. These innocula were diluted with

sterilized distilled water to obtain a density corresponding

approximately to 0.5 of McFarland standard turbidity

scale (108 colony forming unit “CFU” per ml for the

bacteria and 103 spores per ml for fungi)[18]. 0.5 ml of

each innoculum was introduced into the corresponding

fluid agar medium homogenized and 25 ml of it poured

into sterile plastic petridishes. The petridishes were

allowed on the flat slab top for the medium to solidify

within 30 min. A standard cork borer of 5mm in diameter

was used to cut four equidistant uniform wells per plate

on the surface of different plates into which was added

50l solution of each extract/fraction at varying

concentration 12.5, 25, 50 and 100 mg/ml. The reference

drugs were Gentamicin, batch 20070402 (0.4 mg/ml) and

Nystatin batch 04D05 (500 g/ml). The plates were

incubated at 37C for 24 and 48h for the bacteria and

fungi respectively. The antimicrobial activity was

evaluated by measuring the zone of inhibition around the

hole. Each test concentration had three replications. The

results were recorded as the mean diameter of the zones

of growth inhibition surrounding the discs [19].

Determination of minimum inhibitory concentrations

(MIC) using macrodilution method

The Minimum Inhibitory Concentrations (MICs)

of test samples found to be active by the diffusion test

were determined based on the macrodilution method [18]

with some modifications as follows. The test

extract/fractions were dissolved in 10% Tween 80 to give

a stock concentration of 100mg/ml and serially diluted

(two-fold) in a seres of test tubes to a working

concentration ranging from 1.560 to 100mg/ml using

nutrient broth supplemented with 10% glucose and 0.05%

phenol red (colour indicator). These were later inoculated

with 0.2ml suspension of the test organisms. Microbial

growth was determined by observing for color change in

the tube (red to yellow when there is growth). The lowest

concentration that showed no change of color was

considered as the MIC.

Statistical Analysis and Data Evaluation

Data obtained from this work were analyzed

statistically using one way ANOVA followed by a post

test (Turkey-Kramer multiple comparison test).

Differences between means was considered significant at

1% and 5% level of significance, that is P ≤ 0.01and 0.05

RESULTS

Open field test

Administration of root extract of H. africana

(200 – 600 mg/kg) for 5 days caused significant (p<0.05 –

0.01) reduction in the frequency of line crossing with the

low doses of the extract (200 and 400 mg/kg) when

compared to control. Highest dose of the extract (600

mg/kg) did not affect the locomotor activity of the rats.

The standard drugs, imipramine (5 mg/kg), caused a

significant (p<0.001) increase in the locomotor activity of

the rats as evident in the frequency of the line crossing

(Figure 1).

The root extract of H. africana (200 – 600

mg/kg) caused significant (p<0.001) increase in walling

frequency of the rats at high doses (400 and 600 mg/kg)

when compared to control. The low dose (200 mg/kg) had


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no effect on the locomotor activity of the rats. The

standard drug, imipramine (5 mg/kg), produced a

significant (p<0.001) increase in the walling frequency of

the animals.(Figure 2).

The root extract of the H. africana (200 – 600

mg/kg) caused significant (p<0.001) non dose –

dependent reduction of the rearing frequency of rats

administered with the extract for five days. However, the

standard , imipramine (5 mg/kg), exerted a significant

(p<0.001) increase in the rearing frequency when

compared to control.(Figure 3).

Effect on Force Swimming Test

Administration of the ethanolic root extract of H.

africana (200 – 600 mg/kg) to rats for five days did not

show any significant (p>0.05) effect on the behaviour of

the rats during force swimming test when it was

compared to control. However, standard drug, imipramine

(5 mg/kg) produced a significant (p<0.001) reduction in

the immobility time of the rats when compared to control

(Figure 4).

Effect on Tail Suspension Test

The activities of rats pretreated for five days with

root extract of H. africana (200 – 600 mg/kg) was not

affected during tail suspension test when it was compared

to control. The lower doses (200 and 400 mg/kg) exerted

noticeable reductions in the immobility time though

insignificant (p>0.05) when compared to control. The

standard drug, imipramine (5 mg/kg), exerted a

significant (p<0.001) reduction of the immobility time of

the rats when compared to control (Figure 5).

Anticonvulsant activity Hippocratea africana root

extract on aminophylline- induced convulsion

The administration of Hippocratea africana root

extract and fractions (200 – 600 mg/kg) raised the

threshold of seizure and caused a significant (p<0.05 –

0.001) delay in the onset of seizure induced by

aminophylline. The delay was significant (p<0.05 –

0.001) when compared with the control and in a dose

dependent fashion. The activities of the extract and

fractions were higher than that of the standard drug,

phenytoin. The extract/fractions also caused a significant

(p<0.001) prolongation of the time of death of the mice

when compared to control though less than that of the

standard drug, phenytoin. The aqueous fraction

demonstrated a higher anticonvulsant potential than the

chloroform fraction (Table 1).

Anticonvulsant Activity of Hippocratea africana root

extract on PTZ- induced convulsion The pretreatment of mice with root extract and

fractions of Hippocratea africana (200 - 600 mg/kg)

delayed the onset of tonic –clonic convulsion induced by

the administration of petylenetetrazol in a dose dependent

manner. These delays were only significant (p<0.05 –

0.001) at the highest dose of the extract (400 mg/kg)

when compared to control. The chloroform fraction which

exerted the most significant (p<0.001) delay could not

prolong the time of death of the mice significantly

(p>0.05) as the aqueous fraction which had a significant

(p<0.05) protection activity. The prolongation produced

by the highest dose of the extract (600 mg/kg) was more

than that of the standard drug, phenytoin (Table 2).

Effect of Hippocratea africana root on phenobarbitone

induced sleeping time of rats

Administration of the root extract of H. africana

(200 – 400 mg/kg) to rats shortened considerably the time

for onset of sleep. The extract also prolonged the duration

of sleep significantly (p<0.001) when compared to

control. However, the effect was lower than that exerted

by the standard drug (Table 3).

Table 1. Anticonvulsant activity Hippocratea africana root extract on aminophylline- induced convulsion

Drug Extract Dose

(mg/kg)

Latency of

clonic

convulsion (s)

Latency of

Tonic

convulsion (s)

Convulsion

% Mortality Time of Death

Control

(normal saline) 0.2ml 197.7 14.53 281.3 19.93 100 100 385.4 12.28

Hippocratea africana

root extract

200 339.6 6.73 b

430.1 23.10 b 100 100 1323.5 56.69

c

400 366.2 1.79,b 500.3 35.10

b 100 100 1532.6 48.07

c

600 390.6 41.86c 617.5 31.75

c 100 100 3336.4 54.32

c

Chloroform fraction 400 224.5 17.70a 340.6 17.83

a 100 100 1246.5 16.46

c

Aqueous fraction 400 298.2 21.16a 418.8 54.55

b 100 100 1749.2 68.47

c

Phenytoin 40 252.6 10.13 c 380.2 41.66

a 100 100 2566.6 23.18

c

Data are represented as mean SEM. significant at aP< 0.01, bP<0.001 when compared to control. (n=6)


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Table 2. Anticonvulsant Activity of Hippocratea africana root extract on PTZ- induced convulsion

Data are represented as mean SEM .significant at aP < 0.05;bp<0.01; p<0.001 when compared to control.( n=6)

Table 3. Effect of Hippocratea africana root extract on sleeping time of rats

Drug Extract Dose (mg/kg) Onset of Sleep (min) Duration of Sleep (min

Control (normal saline) 0.2ml 8.97 1.53 71.30 3.93

Hippocratea Africana stembark extract

200 7.12 1.86 85.29 3.28

400 6.58 1.74 98.05 3,86

600 6.95 1.97 114.72 3.29

Diazepam 2 4.96 1.13 160.32 2.66b

Data are represented as mean SEM. significant at aP< 0.05, bp<0.01, cp<0.001 when compared to control. (n=6)

Table 4. Antimicrobial activity of Hippocratea africana root extract and fractions

Fraction/

Drug/

Microorganism

Zone of Inhibition in mm

Conc/ml

mg/ml

Ethanolic

crude Chloroform Aqueous

Streptomycin

0.4 mg/ml

Nystatin

500µg/ml

E. coli

ATCC 10418

100

50

25

12.5

-

-

-

-

-

-

-

-

-

-

-

35.5 -

Staph .aureus

NCTC 6571

100

50

25

12.5

20.0

18,0

15,0

14.0

26.0

24.0

24.0

22.0

20.0

18.0

16.0

14.0

35.0 -

B. subtilis

NCTC 8853

100

50

25

12.5

18.0

16.0

-

-

25.0

25.0

23.0

22.0

22.0

20.0

18.0

16.0

31.0 -

Pseudomonasaeruginosa

ATCC 27853

100

50

25

12.5

-

-

-

-

-

-

-

-

20.0

18.0

-

-

45.0

-

-

-

Klebsiellapneumoniae

100

50

25

12.5

-

-

-

-

-

-

-

-

-

-

-

-

45.0

Salmonella typhi

100

50

25

12.5

-

-

-

-

-

-

-

-

-

-

-

-

50.0

Candida albicans

100

50

25

12.5

-

-

-

-

-

-

-

-

-

-

-

-

-

30.5

Tineacapitus

100

50

25

12.5

-

-

-

-

-

-

-

-

-

-

-

-

- 25.0

Drug Extract Dose

(mg/kg)

Latency of

clonic

convulsion (s)

Latency of

tonic

convulsion (s)

Convulsion

%

Mortality

%

Time of Death

Control

(normal saline) 0.2ml 35.0 1.73 72.2 2.00 100 100 128.6 12.65

Hippocratea africana

root extract

200 48.6 7.17 75.0 9.50 100 100 438.1 37.04 b

400 53.0 4.50 80.6 6.18 100 40 404.5 71.52 a

600 83.0 5.85 c 97.6 9.28 a 100 40 793.4 33.35 c

Chloroform fraction 400 87.6 6.25 c 135.6 8.25c 100 100 177.6 23.02

Aqueous fraction 400 55.5 3.84 102.0 2.16a 100 100 344.6 23.02 a

Phenytoin 40 53.3 3.84 84.5 2.72 100 100 436.7 18.16b


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Table 5. Minimum inhibitory concentration of ethanolic crude root extract and fractions of H. africana (mg/ml).

Organism Crude Extract/ Fraction

Ethanolic Crude Extract Aqueous Chloroform

Staph. aureus NCTC 6571 10 6.5 3.125

Bacillus subtilis NCTC 8853 30 7.0 3.125

Pseudomonas aureginosa ATCC 27853 - 30 -

Figure 1. Bar diagram showing the effect of Hippocratea

africana root extract on line crossing frequency of rat

Results are represented as mean ± SEM with n = 5 in each group. *p<0.05,

***P < 0.001 when compared with control group.


africana root extract on walling frequency of rat

Results are represented as mean ± SEM with n = 5 in each group. ***P <

0.001 when compared with control group.


africana root extract on rearing frequency of rat.

Results are represented as mean ± SEM with n = 5 in each group. * *p<0.01, ***P < 0.001 when compared with control group.

Figure 4. Bar diagram representing the immobility

duration (in min) of mice in force swimming test

Results are represented as mean ± SEM with n = 5 in each group.

Figure 5. Bar diagram representing the immobility duration (in min) of mice in Tail suspension test

Results are represented as mean ± SEM with n = 5 in each group


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Effect of the crude extract and fractions of H. Africana

on some microorganisms

Table 4 shows the diameters of the zones of

inhibition exhibited by ethanolic crude extract and

fractions at various concentrations employed. The crude

extract, chloroform and aqueous fractions showed a

narrow spectrum of activity against gram positive Staph

aureus (NCTC 6571) and Bacillus subtilis (NCTC 8853).

Pseudomonas aeroginosa (ATCC 27853) was the only

gram negative bacteria tested that was sensitive to the

aqueous fraction. The chloroform fraction had the highest

activity against both Staph aureus and Bacillus subtilis

followed by aqueous fraction and the crude extract. Other

tested organisms were not sensitive to either the crude

extract or fractions..The activities of the crude

extract/fractions were not comparable to that of the

standard, streptomycin. The crude extract and the

fractions were inactive against all fungal species tested.

The results of minimum inhibitory concentrations

(MIC)of the crude / fraction are shown in Table 5. The

lowest MICs of chloroform fraction (3.125 mg/ml) were

recorded against B.subtilis (NCTC 8853) and Staph.

Aureus (NCTC 6571). While the MICs of aqueous

fraction against B. subtilis and Staph aureus were 7.0 and

6.5 mg/ml respectively. Aqueous fraction also inhibited P.

aeruginosa with MIC value of 30.0 mg/ml.

DISCUSSION

In this study, evaluation of the effect of ethanolic

root extract on central nervous system was carried out in

rats using different models; Open field test, tail

suspension test and force swimming test. The root extract

(200 – 600 mg/kg) was found to cause significant dose

dependent reductions in the frequency of line crossing and

rearing activities of the pretreated rats, while the walling

frequency was significantly (p<0.001) increased at higher

doses. However, it had no significant effect on the

immobility time of the rats in force swimming and tail

suspension tests.

Monitoring of locomotor activity of animals has

been used in assessing effect of drug on the CNS. An

increased movement is a measure of the level of

excitability of the CNS [20] and its decrease may be

intimately related to sedation resulting from depression of

the CNS [21]. Central nervous system stimulants are

known to increase locomotor activity, while agents with

depressant activity cause reduction in movements [22].

The root extract was found to increase significantly

walling activity during open field test suggesting

stimulatory effect on the CNS which may have been

masked in other behaviours. However, it is noteworthy

that several established antidepressants decrease

locomotor activity [23].

The root extract was found to have no effect on

immobility time of rats during force swimming and tail

suspension tests. Psycho stimulants may also reduce

immobility in FST and TST models, but in contrast to

antidepressants, these cause marked motor stimulation in

locomotor activity test. In case of minor or major

tranquilisers, immobility was not affected but there was a

reduction in motor activity [24].

Forced swimming and tail suspension tests are

two of the most commonly used animal models of

depression for antidepressant screening. In the forced

swimming test, the development of immobility when mice

are placed into an inescapable cylinder of water reflects

the cessation of persistent escape-directed behavior [25].

The tail suspension test is based on the fact that animals

subjected to the short-term, inescapable stress of being

suspended by their tail, will develop an immobile posture.

Various antidepressants are able to reverse the immobility

and promote the occurrence of escape related behavior.

Both models of depression are widely used to screen new

antidepressants. These tests are quite sensitive to major

antidepressant drugs including tricyclics, serotonin-

specific reuptake inhibitors, MAO inhibitors, and atypical

antidepressant [26].

Forced swimming and tail suspension tests

which represent the behavioural despair model, claimed to

reproduce a condition similar to human depression [27].

The tests are based on the observation that animals,

following initial escape oriented movements, develop an

immobile posture when placed in an inescapable chamber.

The immobility is thought to reflect either a failure of

persistence in escape-directed behaviour (i.e. behavioural

despair) or the development of passive behaviour that

disengages the animal from active forms of coping with

stressful stimuli. It is well known that clinically effective

antidepressants (such as imipramine) typically increase

the swimming efforts of the animal seeking a solution to

the problem and, therefore, they decrease the duration of

immobility in the forced swimming test. This was

observed in this study.

However, the results of this study suggest that

the root extract exhibited depressant activity with a weak

psychomotor stimulation. Phytochemical constituents

such as flavonoids have been implicated in depressant

action on the CNS [28], while polyphenols especially

flavonoids like quercetin and rutin have also been

reported to exhibit antidepressant effect [29]. The root

extract of H. africana have been reported to contain

thujene, sabinene as well as 3, 4, 5-trimethoxy phenol.

These phytochemical constituents may be responsible for

the observed activity of the root extract in this study

The evaluation of anticonvulsant and

antimicrobial activities of root extract and fractions of

Hippocratea africana was also carried out in this study.

Pretreatment of the mice with the root extract and

fractions (aqueous and chloroform) of Hippocratea

africana (200 – 600 mg/kg) was found to significantly


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delay the onset of tonic/clonic convulsions and prolonged

the time of the death of the treated mice against

pentylenetetrazol and aminophylline induced convulsions.

The aqueous fraction was observed to have the highest

activity.

The exact mechanisms of seizures induced by

aminophylline appear to be diverse, multiple and

complex, and also unclear. Evidence suggests that

seizures induced by aminophylline, could be the result of

adenosine receptor antagonism or due to inhibition of

cerebral nucleotidase activity [30, 31], which lower the

adenosine content in the brain and eventually lead to a

process of disinhibition. However, report has it that di-

phenylhydantoin a potent inhibitor of adenosine uptake

was ineffective in preventing these seizures [32]. Apart

from non-specific adenosine receptor antagonism [33],

aminophylline is thought to have inhibitory influence on

adenosine synthesis. At higher doses inhibition of

phosphodiesterase activity including mobilization of

intracellular calcium ions from labile stores are said to be

implicated in AMPH-induced seizures [34, 35]. However,

a report by Ray et al., [36], has implicated oxidative stress

due to the generation of free radicals and reactive oxygen

species to be responsible for the seizures induced by

aminophylline.

Hippocratea africana root extract and fraction

which was observed to delay the onset of aminophylline

induced convulsion, has been reported to contain α-

thujene , sabinene and sesquiterpenes, which have been

implicated in the anticonvulsant activities of plants

[37,38]. These compounds may be responsible for the

observed anticonvulsant activity of the plant. Also,

Okokon et al. [10] reported the antioxidative property of

the root extract. The antioxidant activity of this root

extract may be responsible for the observed protection

against aminophylline induced convulsion.

According to De Sarro et al.,[39],

pentylenetetrazol (PTZ)is suggested to exert its

anticonvulsant effect by inhibiting the activity of gamma

aminobutyric acid (GABA) at GABAA receptors. Gamma

aminobutyric acid is the major inhibitory neurotransmitter

which is implicated in epilepsy. The enhancement and

inhibition of the neurotransmission of GABA will

attenuate and enhance convulsion respectively [40,41].

Phenobarbitone and diazepam, standard epileptic

drugs,have been shown to exert their antiepileptic effects

by enhancing GABA-mediated inhibition in the brain

[42,43]. These drugs are reported to antagonise PTZ-

induced convulsion [44] by enhancing GABA

neurotransmission. Phenytoin was unable to prevent PTZ-

induced seizure because it is thought to exert its

antiepileptic effect by blocking sodium ions into brain

cells thus inhibiting generation of repitative action

potential [42]. Since the root extract and fractions of

Hippocratea africana were able to delay PTZ – induced

convulsion it is probable that they may be interfering with

gabaergic mechanism(s) to exert its effect. Their

anticonvulsant activities are due to their phytochemical

components as reported above.

The ethanol root extract of H. africana was

found to significantly enhanced duration of the

phenobarbitone sodium -induced hypnotic effect, which

was observed in the shortening of time of onset of sleep

and prolonging the duration of sleep following its

administration suggesting a depressant activity on the

CNS. Substances which possess CNS depressant activity

either decrease the time for onset of sleep or prolong the

duration of sleep or both [45, 46]. A prolongation of the

phenobarbitone effect could involve a facilitation of

GABA mediated postsynaptic inhibition through

allosteric modification of GABAA receptors.

The root extract and fractions were found to

exert antibacterial activity against Gram positive S.

aureus and B. subtilis as well as Gram negative P.

aureginosa. This activity may have resulted from the

presence of phytochemical compounds such as

monoterpenes (thujene and sabinene) and sesquiterpenes

as reported above.

Compounds such as terpenes (mono and

sesquiterpenes) which have been implicated in

antibacterial activities of plants [47, 48] have been found

to be present in this extract. These compounds may have

been responsible for the antibacterial activity observed in

this study.

CONCLUSION

From the results of this study, the root extract/

fractions possess significant CNS depressant and

anticonvulsant activities as well as a considerable

antibacterial activity. It will be interesting to isolate and

characterised the active ingredient in this extract

CONFLICT OF INTEREST There is no conflict of interest.

ACKNOWLEDGEMENT The authors are grateful to Mr. Nsikan Malachy

Udo of Department of Pharmacology and Toxicology for

technical assistance.

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DEPRESSANT, ANTICONVULSANT AND ANTIBACTERIAL ACTIVITIES OF HIPPOCRATEA AFRICANA