Introduction

The use of medicinal plants in the treatment of diseases is as old as the diseases themselves. Ijomah et al., (1997) noted that herbal medicine is the earliest form of medicine. Herbal medicine involves the use of medicinal plants in the treatment of disease. According to Sofowora (1993), a medicinal plant is a plant whose one or more of its parts or organs contains substances that can be useful for therapeutic purposes or which are precursors for the synthesis of useful drugs.

Mitracarpus scaber is a medicinal plant widely employed in traditional medicine mainly in west African countries. The plant belongs to the family Rubiaceae and it grows in many tropical countries where it is usually identified as trees shrubs and woody climbers. Many of its herbaceous forms are found in temperate countries. The plant is annual in nature. In the Middle Belt area of Nigeria, it starts sprouting in July/August, while in the southern part of Nigeria, it is usually available much earlier (Ekpandu et al., 1999).

In Nigeria and some other west African countries, the plant is used for the treatment of headache, toothache, amenorrhoea, hepatic diseases, venereal diseases, leprosy, skin diseases including eczema and scabies (Dalziel 1956, Kerharo and Adam 1974).

Staphylococcus aureus is a bacterium that causes food poisoning and many pus forming disorders such as boils, and abscesses on the defective skin and other epithelial surfaces. It also causes secondary infections such as peritonitis, cystitis and meningitis (Duguid et al., 1980, Azoro and Gaffa 2005). Many strains of Staphylococcus and other bacteria species are known to have become resistant to penicillin and many other newly introduced antibiotics (Ruskin 1991). It is mainly because of this that researchers had intensified efforts towards the discovery of cheaper and more stable therapeutic agents for the treatment of human diseases that are of bacteria origin. Since the leaf of Mitracarpus scaber has been reported to be effective in the treatment of many human skin diseases, this work was therefore undertaken to determine the activities of this leaf extract against Staphylococcus aureus in vitro.

Materials and Methods

The leaves of Mitracarpus scaber were collected from grain farms within Bauchi metropolis after it has been identified by a competent Botanist.

Methanol and Aqueous Extraction

The collected plant leaves were carefully washed, dried and then pounded into powder using mortar and pestle as descried by Dada et al., (1987).

Finer particles of the powder was obtained by blending in a milling machine. About 50g of the powder was packed into a Sohxlet apparatus and extracted exhaustively with methanol and the removal of the solvent was by distillation over water bath. Thereafter, concentrated extracts were put inside labeled universal bottles.

Also, another 50g of the powder was added to 200ml of distilled water and was heated at 70⁰C in a water bath for two hours. After heating, the mixture was cooled and filtered. The filtrate was further concentrated to reduce the volume.

Assay of the Extracts for Antibacterial Activity

Inocula taken from the preserved test organism were cultured on nutrient broth and allowed to grow at 37⁰C for 48 hours. Then cultures were taken from it and spread evenly on the surface of nutrient agar plates.

Then dilutions of both the aqueous and methanol extracts were done by adding 25ml, 50ml and 75ml each of the different types of extracts into 75ml, 50ml and 25ml of distilled water respectively in separate sterile beakers to obtain 75%, 50% and 25% concentrations of the extract. Also, the beakers containing the undiluted portion of each of the types of extract were labeled as undiluted.

Table 1: Antibacterial activity of the extract on Staphylococcus aureus (determined by the diameter of the zone of inhibition on agar plates)

Table 2: Phytochemical analysis of the leaf extracts of Mitracarpus scaber

Thereafter, separate sterilized 6mm filter paper discs were soaked into the different dilutions of the extracts and also in the undiluted samples. Theses soaked filter paper discs were carefully placed on the surfaces of the cultures in the different inoculated plates. The plates were incubated at 37⁰C for 24 hours. After this period of incubation, the plates were examined for clear zones of growth inhibition around the soaked filter discs containing the different dilutions of the two types of extracts.

Phytochemical Analysis

The two types of extracts were evaluated for the presence of carbohydrates (sugars), alkaloids, tannins, glycosides, sterols, saponins, flavonoids and resins.

Test for Carbohydrates (Sugars) Molisch’s Test

Few drops of molisch’s reagent were added separately to two (2) test tubes each containing 2mls of the methanol and aqueous extracts respectively. Then 1ml of concentrated sulphuric acid was added slowly down the sides of the inclined tubes so that the acid forms a layer beneath. A redish brown solution in the methanol extract indicates the presence of sugar while in the aqueous extract a light brown colour indicates the presence of sugar.

Test for Alkaloids

Test for Saponins

2mls of each extract in different test tubes were vigorously shaken for two minutes and observed for frothing which indicates presence of saponins. The emulsion test method for the presence of saponins was also employed. And in doing this, 5 drops of olive oil was added to 3mls of each extract in test tubes, and the mixture was vigorously shaken and observed for the development emulsion, which indicates the presence of saponins.

Test for Sterols using the Liebermann Burcharel Reaction

1ml of concentrated sulphuric acid was added to 1ml of each of the extracts and observed for the development of a red colour which indicates the presence of sterols.

Test for Flavonoids

A little amount of magnesium powder (2g) and a few drops of concentrated hydrochloric acid were added to 3ml of each of the extract and observed for red colouration, which indicates the presence of flavonoids.

Test for Resins

5ml of copper acetate solution was added to 5ml of each of the extracts. The resulting solution was shaken vigorously and allowed to separate, then observed for the development of a green colour, which indicates the presence of resins.

Results and Discussion

1ml of 1% HCl was added to 3ml of each of the extracts and observed for the development of a brown precipitate which indicates the presence of alkaloids.

Test for Glycosides

10mls of 5% sulphuric acid was added to 1ml of each of the extracts in a test tube. The mixture was heated in a boiling water for 15 minutes. 10ml of Fehling’s solution (5ml. each of solution A and B) was added and the mixture was boiled for one (1) minute and observed for a red precipitate which indicates the presence of glucosides.

The result of antibacterial activity of the different concentrations of the two types of extracts on Staphylococcus aureus is given in Table 1 below and that of the phytochemical analysis of the extract is given in Table -2.

The result in Table 1 above shows that both the methanol and aqueous extracts of the leaf of Mitracarpus scaber were very active against the test organism Staphylococcus aureus in vitro. The methanol extract gave a range of 5.0-5.8 zone of inhibition, while the aqueous extract gave a range of 3.5- 4.5mm both in the different dilutions used.

This very high levels of growth inhibition exhibited by both the methanol and aqueous extracts proves that the leaf of this plant can be used effectively in the treatment of human skin diseases caused by Staphylococcus aureus.

According to Dalziel (1956) and Kerharo and Adam (1974), the leaf of Mitracarpus scaber is used in the treatment of leprosy, eczema and other skin diseases in West African countries.

Results of the phylochemical analysis shows that carbohydrates (sugars), tannins, glycosides, saponins, flavonoids; alkaloids and resins were present. Sterols were however absent in both types of extracts. Some of these compounds are known to be among the antimicrobial agents commonly found in medicinal plants (Germano et al., 1999, Ali Emmanuel et al., 2003, Otiko et al., 1987, Sofowora 1991, Sofowora and Sainsbury 1981).

Conclusions

Our investigation has revealed that the extracts from the leaves of Mitracarpus scaber have very high levels of antibacterial effect on Staphylococcus aureus in vitro. This confirms that human skin infections caused by Staphylococcus aureus can be treated effectively with the leaf of this plant more so when some of the antimicrobial agents usually found in many medicinal plants are also present in the leaf extracts of this plant.

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