The use of plants as medicine is an ancient practice common to all societies
especially the African society. This practice continues to exist in the
developing nations. It is on this basis that researchers keep on working
on medicinal plants in order to produce/develop the best for physiological
uses as medicines (Usman and Osuji, 2007).
The plant Cissus cornifolia (Bak.) Planch is a species of the
genera Cissus that belongs to the family Vitaceae. It is an annual,
sub-erect herb with height of about 1.3 m from the permanent woody root
base. The plant is distributed in the rocky suburbs and bush Savannah
in Ghana and Northern Nigeria. The plant is locally called Riigarbirii
(robe of the monkey) or Tsùwààwùùn
birii among the Hausa speaking people of Northern Nigeria (Burkill,
The plant has wide array of uses in African Traditional Medicine amongst
which is its being used by the Fulani of Northern Nigeria as a remedy
for gonorrhea when taken with native natron while the leaf-sap is used
among the Tanganyika as a sedative in cases of mental derangement; the
root-decoction is also used for malaria, septic tonsil and pharyngitis
(Burkill, 2000). The objectives of this study aimed at verifying the folkloric
claim on the leaf of this plant as a remedy against cases of mental derangement
using various scientific models.
MATERIALS AND METHODS
Plant material: The leaves of Cissus cornifolia (Bak.)
Planch was collected from Kufena Village, Zaria, Kaduna State Nigeria;
in the month of July 2007. The herbarium sample was identified by Mallam
Musa, M. of the Herbarium Unit, Department of Biological Sciences, Ahmadu
Bello University, Zaria-Nigeria, where a voucher specimen (No. 024) was
Preparation/extraction of plant material: The leaves of Cissus
cornifolia was air-dried for seven days under shade and then ground
into fine powder. About 250 g of the powdered leaves was extracted with
2000 mL of 80% methanol in H2O (v/v) using the maceration technique.
The extract was concentrated under reduced pressure to afford a dark green
mass which weighed 32.5 g (13.0% w/w) as crude methanol extract.
Animals: The pharmacological experiments were conducted using
adult Swiss Albino mice of either sex weighed between 20-26 g obtained
from Animal house, Department of Pharmacology and Clinical Pharmacy, ABU
Zaria-Nigeria. The animals were maintained on a Standard Animal Feeds
obtained from Excel Feeds Plc (Kaduna, Nigeria) and allowed food and water
ad libitum. The animals were housed in a standard cage at room
temperature in a 12 h light/dark cycle (6 am - 6 pm) and then allowed
to acclimatize with the laboratory environment for at least five days
prior the commencement of the experiments.
Drugs: Diazepam - DZ (Pfizer, USA) and Chloroform, Ethylacetate,
Methanol (Fluka-Aldrich) solvents were obtained respectively from a pharmacy
and chemical retail stores in Samaru-Zaria, Nigeria. The drug/chemicals
were freshly prepared to the desired concentration with appropriate solvent
just before use.
Phytochemical screening: The qualitative phytochemical analysis
of the crude methanolic extract of Cissus cornifolia was carried
out in order to ascertain the presence of its constituents employing standard
conventional protocols (Sofowora, 1993; Trease and Evans, 2002).
Thin layer chromatography (TLC): The methanolic leaf extract of
C. cornifolia was subjected to TLC examination using the solvent
system: Ethylacatate:Chloroform:Methanol:Water in the ratio 15:8:4:1 as
the developing solvent and then sprayed with Gibb`s reagent.
Acute toxicity studies: Acute toxicity (LD50) was determined
using the method of Lorke (1983). The study was divided into 2 phases.
Nine mice were used in the first phase in 3 divided groups of 3 each.
Group A received extract at a dose of 10 mg kg-1 body weight
intra-peritoneal (i.p.) while groups B and C received extract doses of
100 and 1000 mg kg-1 body weight i.p., respectively. The treated
animals were observed for 24 h; in the second phase 4 mice were divided
into 1 mouse each: Group D received extract at a dose of 600 mg kg-1
body weight i.p., groups E, F and G treated with the extract at a dose
of 1000, 1600 and 2900 mg kg-1 body weight i.p. respectively
and also observed for 24 h. Thereafter, the final LD50 value
computed as the square root of the product of the lowest lethal dose and
the highest non-lethal dose i.e., the geometric mean of consecutive doses
for which 0 and 100% survival rates were recorded.
Diazepam-induced sleep in mice: The method of Rakotonirina
et al. (2001), was adopted. Sleep potentiation time of the plant extract
was studied in group of mice that received DZ at a dose of 30 mg kg-1
body weight one hour after i.p. administration of extract 10, 20 and 40
mg kg-1 body weight and Normal saline; with 5 mice in each
group. The sleeping time was estimated as the time between disappearance
and recovery of the straightening reflex.
Head-dip test for exploratory behavioural pattern in mice: This
study was conducted using wooded apparatus measuring 40x40 cm with 16
evenly spaced holes (Perez et al., 1998). Mice were grouped into
5 of 5 mice each. Group I served as control treated with Normal saline
(10 mg kg-1 body weight i.p.), groups II - IV were treated
with extract at doses of 10, 20 and 40 mg kg-1 body weight
i.p., respectively; while those in group V received DZ 2 mg kg-1
body weight i.p. Thirty minutes after treatment, the mice were placed
singly on a board with 16 evenly spaced holes and the number of times
the mice dipped their heads into the holes during 5 min trial was counted.
Results were expressed as means for the various treatment groups at different
Beam walking assay (motor co-ordination) in mice: Mice were trained
to walk from a start platform along a ruler (80 cm long, 3 cm wide) elevated
30 cm above the bench by metal support to a goal box. Three trials were
performed for each mouse. The mice that have successfully walked along
the ruler were randomly divided into 5 groups of mice each. The first
group served as control treated with Normal saline (10 mL kg-1
body wt. i.p.), second, third and fourth groups were treated with extract
at doses of 10, 20 and 40 mg kg-1 body weight i.p., respectively;
while those in the fifth received DZ 2 mg kg-1 body weight
i.p. The beam was made of wood, 8 mm in diameter, 60 cm long and elevated
30 cm above the bench by metal supports. Thirty minutes post treatment;
each mouse was placed at one end and allowed to walk to the goal box.
Mice that fell were returned to the position they fell from with a maximum
time of 60 sec allowed on the beam. The number of foot slips was recorded
with the aid of a tally counter. The number of foot slips is measure of
motor coordination deficit (Stanley et al., 2005).
Statistical analysis: The results for the sleeping time, exploratory
behaviour and the beam walking assay were reported as mean ± SEM.
The mean values of the results from the control group were compared to
the mean values of groups treated with extract using the (Dunnett post
hoc test). The results were considered significant at p<0.001.
Phytochemical screening: The results from the preliminary qualitative
phytochemical analysis of the 80% methanolic leaf extract of C. cornifolia
revealed the presence of alkaloids, flavonoids, saponins, steroids/terpenoids,
tannins as shown in Table 1.
Thin layer chromatography (TLC): Thin layer chromatographic examination
using the solvent system (Ethylacatate:Chloroform:Methanol:Water in the
ratio 15:8:4:1) revealed 5 spots (Rf: 0.30, 0.36, 0.46, 0.64, 0.90). Upon
spray with Gibb`s reagent for the detection of stilbenoids, 3 spots (Rf:
0.30, 0.36, 0.46) tested positive by developing blue colour indicative
of stilbenoids zones.
Acute toxicity studies: The intra-peritoneal LD50 of
the methanolic leaf extract of C. cornifolia in mice was found
to be 775.0 mg kg-1 body weight i.p.
Effects on diazepam-induced sleep in mice: The methanol leaf extracts
of C. cornifolia did not affect the onset of sleep, but significantly
(p<0.01) prolonged the duration of the DZ induced sleep at the dose
tested. The sleep time increased from 89.25 ± 3.19 min in the control
group to about 119.25 ± 12.33 to 249.50 ± 15.77 min in a
non-dose dependent manner in the groups treated with extract at dose of
40 and 10 mg kg-1 body weight i.p., respectively as shown in
Effects on exploratory behaviour sleep in mice: The extract treated
at a dose of 10, 20 and 40 mg kg-1 body weight i.p. exhibited
a significant reduction in the mean number of head dip responses in the
hole-board test compared with that of the control group; though the effects
observed was not dose-dependent. The extract (20 mg kg-1 body
weight i.p) significantly (p<0.001) reduced the effect as shown in
Effects on motor co-ordination (beam walking assay): The extract
under study showed no significant difference on the number of foot slips
compared to the control group in varying doses. However, the extract exhibited
a non significant reduction in the number of foot slips in a non-dose
dependent manner relative to the DZ treated group (Table
4). The group with high number of foot slips was considered to have
reduced motor coordination.
||Phytochemical constituents of the methanolic leaf extract
of Cissus cornifolia
|Key: + = Present
||Effects of Cissus cornifolia extract on diazepam-induced
sleep in mice
|Values are mean ± SEM; n = 5 in each group, *Significantly
different from control at p<0.001 (Dunnett post hoc test)
||Effects of Cissus cornifolia extract on exploratory
behaviour (head dip test) in mice
|Values are mean ± SEM; n = 5 in each group, #significantly
different from control at p<0.001 (Dunnett post hoc test)
||Effects of Cissus cornifolia extract on motor
coordination (beam walking assay) in mice
|Values are mean ± SEM; n = 5 in each group, δsignificantly
different from control at p<0.001 (Dunnett post hoc test)
This study reports preliminary qualitative chemical constituents and
some neuropharmacological activities of the methanolic leaf extract of
Cissus cornifolia in mice. The qualitative analysis of phytochemical
constituents of C. cornifolia revealed the presence of alkaloids,
flavonoids, saponins, steroids/terpenoids, stilbenoids and tannins. The
saponins and flavonoids have been reported by several researchers (Won
et al., 1980; Dubois et al., 1986; Amos
et al., 2001; Viswanatha Swamy et al., 2006;
Musa et al., 2006) to be responsible for sedative and likewise
to inhibit spontaneous motor activity in mice.
The extract, although at lower dose significantly (p<0.001) potentiated
the diazepam-induced sleep time in mice which is in agreement with earlier
studies of Rakotonirina et al. (2001) and Musa et al. (2006)
and thus confirms that the extract consists of substances with sleep inducing
(sedative) properties. Further neuropharmacological activity was demonstrated
by the significant reduction (p<0.001) compared with the negative control
in the exploratory behaviour in head dip test treated with extract (20
mg kg-1 body weight) which conforms with and had similar action
(insignificantly different) with CNS depressant drug (DZ). In support
to this, similar study reported by Viswanatha Swamy et al. (2006)
revealed that Cissus quadrangularis exhibited similar activity.
Therefore, a decrease in the hole board test for exploratory behaviour
by an extract is an indication of sedative properties (File and Wardill,
1975); hence, it is imperative to suggest that the extract possessed CNS
depressant activity as indicated by the decrease in exploratory behaviour
in mice as shown by the reduction in the head-dip test (Adzu, 2002; Viswanatha
Swamy et al., 2006).
The extract had no observable effect on motor coordination in beam assay
test when compared with the negative control, suggesting that the inhibition
effect observed in the other tests might be elicited centrally and not
due to a peripheral neuromuscular blockade (Perez et al., 1998).
It is therefore, possible that the sedative action of the extract was
Thus, it is pertinent to suggest that alkaloids, saponins and stilbenoids
detected in the plant extract could be responsible for some of these neuropharmacological
effects and that the extract is in support of the usage of the leaf-sap
as a remedy against mental derangement being expressed by its CNS depressant
and sedative properties.