Ethnobotanical Leaflets 14: 766-73. 2010.
Screening of Antimicrobial Ethanolic Extract of Peristrophe bicalyculata
O.E. Giwa, O.E. Seyifunmi, B. L. Adewumi, V. T. Adebote and A.O. Aladejimokun
Department of Science
Laboratory Technology, Rufus Giwa Polytechnic, Owo,
Issued: June 1, 2010
The ethanolic extract from Peritrophe bicalyculata leaves was
evaluated for the presence of phytochemicals and its antimicrobial activity
in vitro against selected bacteria and fungi using the antibiotic gentamycin
as control. The extract showed the presence of secondary metabolites such as
alkaloid, saponin, tannin and steroid. It also inhibited the growth of the
tested microorganisms at different concentrations. However stronger in-vitro
activity was recorded against Staphylococcus aureus, Klebsiella spp,
Pseudomonas aeruginosa, Aspergillus
Key words: Phytochemicals, Antimicrobial, Inhibition, Ethnomedicine, In-vitro, Concentration.
vegetables play a very important role in our diet and nutrition since they
are the major sources of not only raw fibers but also essential nutrients,
vitamins and minerals. Some of these vegetables are common in
The importance and awareness of nutrition and medicinal properties of some plant in public health issues has resulted in the increase demand of knowledge in the nutrient, phytochemical and antimicrobial properties of food (Chinma and Igyor, 2007). Green leafy vegetables are rich source of carotene, ascorbic acid, riboflavin, folic acids and minerals like calcium, iron and phosphorus. (Fasuyi, 2006).
The presence of some phytochemicals determines the antimicrobial
properties of various plants. They give plants its colour, flavour, odour and
are part of the defense system (disease resistance). According to Liu (2004),
phytochemicals are bio actives, non nutrient plants compounds in fruits,
vegetables, grains and other plants food that has been linked to reduce the
risk of major degenerative disease.
shows that the greatest sources of these photochemicals are fruits and
vegetables. (Onyeka and Uwambeke, 2007). Most leafy vegetable constitute an
indispensable constituent of human diet in Africa generally and Western part
This research work was therefore, aimed at determining the presence or otherwise of some phytochemicals and also antimicrobial properties of ethanolic extracts of Peristrophe bicalyculata leaves on selected microorganisms in order to determine its medicinal use.
Materials and Methods
Source of Materials
Fresh leaves of Peritrophe bicalyculata were collected from
�Obasoto� in Owo township,
Extraction of Plant Materials
The pulverized fresh leaves (100g) were extracted with 95% ethanol by cold maceration for 24 hours at room temperature (25oC). The extract was filtered and evaporated to dryness over water bath after concentration with rotor vapour to minimal volume.
Determination of alkaloids
A measured weight of the sample was dispensed into 10% acetic acid solution in ethanol to form a ratio of 1:10. The mixture was allowed to stand for 4 hours at 28oC. It was later filtered with filter paper and the filtrate was treated with drop wise addition of aqueous NH4OH until the alkaloid was precipitated, this was washed with 10% ammonia solution and dried in the oven at 80oC.
Determination of flavonoids
5g of the sample was boiled in 50ml of 2M HCl solution for 30 minutes under reflux. It was allowed to cool, then filtered through filter paper and the filtrate was treated with equal volume of ethyl acetate.
Determination of tannin
A 5g portion of the sample was dispensed in 50ml of distilled water and mixed properly. This was allowed to stand for 30 minutes at 28oC before it was filtered. 2ml of the plant extract was dispensed into a 50ml volumetric flask. Similarly, 2ml standard solution and 2ml of distilled water were put in separate volumetric flask. The reagent was added to each of the flask, and 2.5ml of saturated Na2CO3 solution was also added, the total content of the flask was made up to 50ml with distilled water and incubated at 28oC for 90 minutes. A spectrophotometer set at 260nm wavelength was used to measure the respective absorbance using the reagent blank to calibrate the instrument.
Determination of Steroid
A measured weight of the sample was dispensed in 100ml freshly distilled water and homogenized in laboratory blender. This was filtered and was eluted with normal ammonium hydroxide solution� (PH 9). 2ml of the eluate was put into the test tube and mixed with 2ml of chloroform. 3ml of ice-cold acetic anhydride were added to the mixture in the flask and 2 drops of concentrated H2SO4 were added to cool. Standard sterol solution was prepared and spectrophotometer at 420nm was used to measure the absorbance.
The crude ethanolic extract of the fresh leaves was screened for antimicrobial activities against clinically isolates using Agar diffusion steak methods. The test organisms were prepared by sub culturing them overnight in culture media and incubated in a freshly prepared nutrient broth at 37oC for 3 hours. 100mg of the test extract was dissolved into 1ml to give a concentration of 100mg/ml of the extract. Clinically isolated bacteria (E. coli, Klebsiella spp, Staphylococcus aureus and Pseudomonas aeruginosa) were inoculated using steak method. 1ml of prepared extract was then introduced into 5mm hole in a solidified inoculated agar, bored with a cork borer and was placed at 540C. Gentamycin, a broad-spectrum antibiotic was used as standard.
Result and Discussion
The presence of alkaloids, tannin, steroid and flavonoid in the plant as shown in Table1 may be collectively or individually responsible for the observed antimicrobial activities. This result also corresponds with the results of phytochemicals of plants and fruit common in the region (Onyeka, and Uwambeke, 2007).
2 shows the result of the antimicrobial potency of crude ethanolic extract of
Peritrophe bicalyculata against some selected microorganisms. The
diameters of the zone of inhibition of this extract were compared with
Gentamycin. The effect of the ethanolic extract of the Peritrophe
bicalyculata was strongly effective against the test bacteria: Staphylococcus
aureus, Klebsiella spp., E. coli, and Pseudomonas aeruginosa, to be
2.1cm, 1.9cm, 2.0cm, 2.1cm zones of inhibition at concentration of 100mg/ml
respectively. The fungi: Aspergillus
MIC of the extract from the Peritrophe bicalyculata were 90g/ml, 90g/ml, 60g/ml and 60g/ml
for Staphylococcus aureus, Klebsiella spp., E. coli, and Pseudomonas
aeruginosa, respectively and 60g/ml, 90g/ml, 60g/ml for the fungi
isolates; Aspergillus niger, Aspergillus clavatu and Rhizopus stolonifer respectively.
The varied minimum inhibitory concentration revealed that the extract is
highly effective against E. coli, Pseudomonas aeruginosa,
This suggest that the species can be gainfully employed in the production of antibiotics as the low MICs means that only a small quantity of extract will be required to inhibit the organisms. The antimicrobial activities shown by Peritrophe bicalyculata is in line with the previous antimicrobial works on different plants analyzed in the region (Onyeka, and Uwambeke, 2007).
There is a need for further study to ascertain if the yield in this species will be increased by using stronger fractionating solvent such as ethyl acetone or methyl acetone. These solvents have been reported to be more vigorous than other solvents used in crude extraction of plants (Ajayeioba and Fadare, 2006). Efforts should also be made to quantify the identified phytochemicals.
In conclusion, the results from this study have shown the potency of the plant crude extract on the tested microorganisms, which is indication of the medicinal value of the plant extract. The extract compared favourably well with gentamycin that was used as control.
TABLE 1. Phytochemicals of crude extract of Peritrophe bicalyculata.
Secondary metabolite ���������������������� Peritrophe bicalyculata
Alkaloid �������������������������������������������������������� +
Saponin ��������������������������������������������������������� +
Tannin������������������������������������ ����������� ����������� +
Steroid ���������������������������������������������������������� +
Flavonoid ������������������������������������������������������ +
+ = Presence of secondary metabolite����� -� = Absence of secondary metabolite
TABLE 2. Minimum inhibitory concentration (MIC) of ethanolic extracts pertrophe bicalyculata on selected test isolates in (mg/ml) and the zone of inhitition measure in centimeter.
����������������������� ������������������������� Zones of inhibition (cm)
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