Antimicrobial Activities of the Extracts and Fractions of Allanblackia floribunda
Allanblackia floribunda is a tree employed in Nigeria and other countries to treat skin disease and other microbial diseases. The ethanol extract, n-hexane, chloroform, ethyl acetate and butanol fractions of the leaves, stem bark and root bark were evaluated for antimicrobial activities against Staphylococcus aureus NCIB 8588, Bacillus subtilis NCIB 3610, Escherichia coli NCIB 86, Proteus vulgaris NCIB 67, Pseudomonas aeruginosa NCIB 950, Klebsiella pneumoniae NCIB 418, Candida albicans and Aspergilus flavus, using agar diffusion method to validate the ethnobotanical uses of the plant. Among the extracts, the ethanol extract of the leaf gave the most significant antibacterial activity. However, no extract showed antifungal activity. Generally, the fractions obtained from the extracts elicited better activity, including antifungal activity against C. albicans. The highest inhibitory effect was exhibited by leaf extract against Ps. aeruginosa NCIB 950, while the ethyl acetate fraction of the stem bark gave the least inhibitory effect against B. subtilis NCIB 3610. The plant extract and fractions produced inhibition zone range between 5 and 35 mm.
Allanblackia floribunda Oliver (Clausiaceae), commonly known as
Tallow tree reproduces by seeds. Its occurrence is limited to tropical
Africa, but centred mostly on the lowland rainforests (Van Rompaey, 2003).
The nuts of the plant produce fine oil taken by the members of local
communities in Tanzania to relieve rheumatism (Anonymous, 2004). Also,
in Cameroon, the stem bark of the plant mixed with Capsicum frutescens
or Solanum anguivi is used for the treatment of cough (Betti, 2004).
In Gabon, the bark is pounded and rubbed on the body to relieve painful
conditions. There also, a decoction is taken for dysentery and as a mouthwash
for toothache and, in Côte D`Ivoire, for stomach pains (Steentoft,
1988). In Ghana, the bark is used for medicinal purposes such as toothache,
diarrhoea and as a pain reliever (Abiww, 1990). The bark decoction of
the stem and root is also used in Central African Republic and West Africa
to treat toothache, dysentery and as an analgesic (Lewis and Elvin-Lewis,
In Akwa Ibom State of Nigeria, the leaves as well as the bark of the
stem and root of the plant are used by the local communities to treat
dysentery, diarrhoea, skin diseases and some other microbial diseases.
The fruits and the seed kernels produce a hard white fat (Cunningham,
1993). The use of the fat in soap production has been suggested (Foma
and Abdala, 1985). A new prenlated xanthone known as Allanxanthone A and
some other known xanthones have been identified from the stem bark of
A. floribunda and have also been shown to have cytotoxic activity
against KB cell line (Nkengfack et al., 2002).
This study aimed at evaluating the leaves, stem bark and root bark of
A. floribunda for antimicrobial activities against Staphylococcus
aureus NCIB 8588, Bacillus subtilis NCIB 3610, Escherichia
coli NCIB 86, Proteus vulgaris NCIB 67, Pseudomonas aeruginosa
NCIB 950, Klebsiella pneumoniae NCIB 418, Candida albicans
and Aspergilus flavus, in order to validate the ethnobotanical
use of this plant for microbial diseases.
MATERIALS AND METHODS
Extraction: The fresh leaves (1 kg), stem bark (1 kg) and root
bark (1 kg) of Allanblackia floribunda were collected in June,
2006, air-dried for a week and reduced to powder. The powder (300 g) was
macerated in 1 L of EtOH-H2O (1:1) for 72 h. The liquid extract
obtained was concentrated to dryness in vacuo at 40°C to yield
dry ethanol extract (40-50 g).
Phytochemical screening: Applying the methods of Sofowora
(1993) and Trease and Evans (2002), the dry ethanol extract of
each part was subjected to phytochemical screening to reveal the presence
of its secondary metabolites.
Partition chromatography: The dry crude ethanol extract (30 g)
of each plant part was dissolved in 40 mL of distilled water and successively
partitioned with n-hexane (50 mL x3), chloroform (50 mL x3), ethyl acetate
(50 mL x3) and n-butanol (50 mL x3) to yield their respective fractions.
The fractions were separately concentrated to dryness in vacuo
to give dry residues.
Test organisms: The bacteria used in this study were typed cultures
obtained from the Department of Microbiology, Obafemi Awolowo University,
Ile-Ife, Osun State, Nigeria, while the fungi were clinical isolates collected
from the same source. The bacteria: Staphylococcus aureus NCIB
8588, Bacillus subtilis NCIB 3610, Escherichia coli NCIB
86, Proteus vulgaris NCIB 67, Pseudomonas aeruginosa NCIB
950 and Klebsiella pneumoniae NCIB 418 were sustained on nutrient
agar (Oxoid) slant at 4°C prior to use. However, the fungi Candida
albicans and Aspergilus flavus were sustained on Sabouraud`s
Dextrose Agar (Oxoid) slants at 4°C before use.
Antimicrobial test: The dry ethanol extract and the dry fractions
were evaluated against the test microorganisms using agar-gel diffusion
method described by Alade and Irobi (1993). The ethanol extract and the
fractions were redissolved in Dimethyl sulphoxide (DMSO). The ethanol
extract and the fractions were tested at concentration levels of 40 and
80 mg mL-1. Fixed volumes (150 μL) of the ethanol extract,
fractions and DMSO were separately introduced into equidistant wells bored
on the surface of the agar and Sabouraud`s plates, which had been previously
inoculated with one of the test organisms. A well containing a standard
drug, chloramphenicol was made in the bacteria plates, while the fungal
plates had a hole containing Nystatin as standard drug.
The bacteria were incubated at 37°C for 24 h, while the fungi were
incubated at 25°C for seven days. The presence of zones of inhibition
surrounding the wells was taken as an evidence of antimicrobial activity.
The extracts of the leaves, stem bark and root bark showed various classes
of compounds (Table 1) inherent in the plant. The extracts of the three
parts contained tannins and cardiac glycosides in high concentration,
while flavonoids and terpenes occurred in moderate concentration. Anthraquinones,
phlobatannins and alkaloids were absent in all the parts, while saponins
were abundant in the leaves and absent in others.
||Phytochemical screening results of the plant parts of Allanblackia
|+++: High concentration; ++: Moderate concentration;
- : Absent
The extracts of the leaves, stem bark and root bark exhibited varying
degrees of antimicrobial effects, with leaves showing the highest antibacterial
activity and the root bark, the least (Table 2). It is noteworthy that
none of the extracts was effective against the test fungi. Also, E.
coli was resistant to all the extracts. The inhibitory effect of the
leaf extract against Ps. aeruginosa was much better than that of
Chloramphenicol. The purification of the extracts by partition chromatography
showed some improvement on the antimicrobial activity of all the extracts.
For instance, the ethyl acetate fraction of the leaf extract elicited
an improvement on the activity of the extract, though fungal activity
was still lacking. With the exception of aqueous fraction, all the other
fractions generally showed improved activity than the stem bark extract.
This improved activity included fungal effect against C. albicans.
However, n-hexane fraction gave the highest activity. This pattern is
similar for root bark extract.
The result of this study showed that the extracts and fractions of the
leaves, stem bark and root bark of Allanblackia floribunda gave
good inhibitory effects against all the test microorganisms except A.
The result also revealed that the leaf extract gave more significant
inhibitory effects than those of stem bark and root bark. However, all
the extracts exhibited only antibacterial effects.
The fractions obtained from the extracts through partition chromatography
gave improved activity against test organisms, including activity against
C. albicans which the extracts failed to inhibit. A similar result
has been observed for the leaves of Heinsia crinita (Ajibesin et
al., 2003). This suggests that antimicrobial activity was increased
by purification of active constituents of the plant. Generally, all the
extracts and the fractions failed to inhibit A. flavus. However,
aqueous fraction did not show significant inhibitory effect against all
the test organisms except B. subtilis. This may be due to the fact
that virtually all the antimicrobial principles had been extracted from
the aqueous phase during fractionation. The best inhibitory effect was
elicited by the leaf extract against Ps. aeruginosa, while the
least was by ethyl acetate fraction of the stem bark against B. subtilis.
Furthermore, the antimicrobial activities of the extracts and the fractions
The presence of terpenes observed in the phytochemical screening may
be responsible for the enhanced effect of n-hexane fraction, since terpenes
are non-polar compounds located in non-polar fractions. In a similar fashion,
tannins, flavonoids and saponins may account for the improved activity
of the ethyl acetate and butanol fractions. The antimicrobial activity
of phenolics (tannins, flavonoids) and saponins has been established in
some plants (Burapadaja and Bunchoo, 1995; Adesina et al., 2000;
Binutu and Cordell, 2000; Pistelli et al., 2002; Mandal et al.,
||Antimicrobial activity of the extracts and fractions of the plant
parts on the test organisms
|L1 = Leaf ethanol extractB1 = Stem bark ethanol extractR1
= Root ethanol extract
L2 = Leaf n-hexane fractionB2 = Stem bark n-hexane fractionR2 = Root
L3 = Leaf chloroform fractionB3 = Stem bark chloroform fractionR3
= Root chloroform fraction
L4 = Leaf ethyl acetate fractionB4 = Stem bark ethyl acetate fractionR4
= Root ethyl acetate fraction
L5 = Leaf butanol fractionB5 = Stem bark butanol fractionR5 = Root
L6 = Leaf aqueous fractionB6 = Stem bark aqueous fractionR6 = Root
A = 80 mg mL-1Values are mean±SD (n = 4).- = No
zone of inhibition.
B= 40 mg mL-1*: p<0.01 with respect to control
C = Chloramphenicol (2 μg mL-1)
N = Nystatin (2 μg mL-1)
NA = Not applicable
DM = DMSO
The extracts of the plant and their fractions gave significant zones
of inhibition against the test organisms, thereby validating the ethnobotanical
claims on this plant as remedy for the treatment of infections and diseases
caused by these organisms. This result corroborates the reports on the
validation of claims from ethnobotanical survey on other plants used to
treat microbial diseases (Rajakaruna et al., 2002; Khan and Omoloso,
2002; Idowu et al., 2005; Rene et al., 2006; Ekpo et
al., 2007). From the result, the plant was found to be effective against
bacterial infections, but weak against fungal diseases. Further, the leaves
whose extract and fractions exhibited the best activity are recommended
for use in preference to other parts of the plant. Since activity improved
with purification, this encourages further investigation to be carried
out on the active fractions of the plant in order to isolate and characterize
the active constituents responsible for the improved activity recorded
in this study. The active constituents if isolated may show much better
activity, leading to the production of effective antibiotics.
Abbiw, D.K., 1990.
Useful Plants of Ghana: West African uses of Wild and Cultivated Plants. Intermediate Technology Publications and Royal Botanic Gardens Kew, London, ISBN-13: 9781853390432, Pages: 337
Adesina, S.K., O. Idowu, A.O. Ogundaini, H. Oladimeji, T.A. Olugbade, G.O. Onawunmi and M. Pais, 2000.
Antimicrobial constituents of the leaves of Acalypha wilkesiana
and Acalypha hispida
. Phytother. Res., 14: 371-374.Direct Link |
Ajibesin, K.K., J.B. Ekpo and N.D. Bala, 2003.
Comparative pharmacognostic and antimicrobial studies on leaves of two varieties of Heinsia crinita
. Global J. Med. Sci., 2: 49-57.Direct Link |
Alade, P.I. and O.N. Irobi, 1993.
Antimicrobial activities of crude leaf extract of Acalypha wilkensiana
. J. Ethanopharmacol., 39: 171-174.CrossRef | PubMed |
African tree piques European interest. Spore, No. 111, World Agro Forestry Centre, pp: 7.
Betti, J., 2004.
An ethnobotanical study of medicinal plants among the baka pygmies in the dja biosphere reserve. Cameroon Afr. Study Monogr., 25: 1-27.Direct Link |
Binutu, O.A. and G.A. Cordell, 2000.
Gallic acid derivatives from Mezoneuron benthamianum
leaves. Pharm. Biol., 38: 284-286.Direct Link |
Burapadaja, S. and A. Bunchoo, 1995.
Antimicrobial activity of tannins from Terminalia citrina
. Planta Medica, 61: 365-366.Direct Link |
Cunningham, A.B., 1993.
African Medicinal Plants, Setting Priorities at the Interface Between Conservation and Primary Healthcare. People and Plants Working Paper, USA
Ajibesin, K.K., N. Rene, D.N. Bala and U.A. Essiett 2007.
Antimicrobial activities of the extracts and fractions of Triumfetta cordifolia
A. Rich Nig. J. Bot., 20: 181-186.Direct Link |
Foma, M. and T. Abdala, 1985.
Kernel oils of seven plant species of Zaire. J. Am. Oil Chem. Soc., 62: 910-911.
Idowu, P.A., J.O. Moody and H.A. Odelola, 2005.
Phytochemical and antimicrobial screening of three Nigerian medicinal plants used to treat infectious diseases traditionally. J. Pharm. Bioresour., 2: 116-119.Direct Link |
Khan, M.R. and A.D. Omoloso, 2002.
Antibacterial, antifungal activity of Harpullia petiolaris
. Fitoterapia, 73: 331-335.Direct Link |
Lewis, W.H. and M.P.H. Elvin-Lewis, 1977.
Medical Botany. John Wiley and Sons Ltd., New York
Mandal, P., S.P.S. Babu and N.C. Mandal, 2005.
Antimicrobial activity of saponins from Acacia auriculiformis
. Fitoterapia, 76: 462-465.Direct Link |
Nkengfack, A.E., G.A. Azebaze, J.C. Vardamides, Z.T. Fomum and F.R. van Heerden, 2002.
A prenylated xanthone from Allanblackia floribunda
. Phytochemistry, 60: 381-384.Direct Link |
Pistelli, L., A. Bertoli, E. Lepori, I. Morelli and L. Panizzi, 2002.
Antimicrobial and antifungal activity of crude extracts and isolated saponins from Astragalus verrucosus
. Fitoterapia, 73: 336-339.Direct Link |
Rajakaruna, N., C.S. Harris and G.H.N. Towers, 2002.
Antimicrobial activity of plants collected from Serpentine outcrops in Sri Lanka. Pharm. Biol., 40: 235-244.CrossRef | Direct Link |
Rene, N., O.H. Oladimeji, P.M.E. Ubulom and B.A.J. Ekpo, 2006.
Larvaecidal and antimicrobial potential of Anthocleista djalonensis
L. (Gentianaceae). Afr. J. Pharm. Res. Dev., 2: 97-101.
Sofowora, A., 1993.
Medicinal Plants and Traditional Medicine in Africa. 2nd Edn., Spectrum Books Ltd., Ibadan, Nigeria, ISBN-13: 9782462195, Pages: 289
Steentoft, M., 1988.
Flowering Plants in West Africa. 1st Edn., Cambridge University Press, Cambridge pp: 352
Trease, G.E. and W.C. Evans, 2002.
Pharmacognosy. 15th Edn., WB Saunders, London, ISBN: 8131200876, Pages: 406Direct Link |
Van Rompaey, R., 2003.
Distribution and Ecology of Allanblackia
species (Clusiaceae) in African Rain Forests with Special Attention to the Development of a Wild Picking System of the Fruits. Unilever Research Ecosystem, Vlaardingen, pp: 1-39