PD 528/08 Rev.1

PD 528/08 Rev.1 (F): Towards sustainable indigenous Mahogany production in : Phase II, refining the silviculture “tool kit” and practical training for industrial-foresters and community farmers

Antimicrobial potential and phytochemical properties of four species of African mahogany

Ofori E. 1, 2 Opuni-Frimpong E.1 and Amissah-Eshun, R.2 1) Forestry Research Institute of Ghana 2) Accra Polytechnic

ABSTRACT

Traditional healers’ use different species of parts for healing purposes, however, very little scientific research to support claims of effectiveness have been undertaken. The objective of this study was to determine the phytochemical properties in four species of African mahogany ( grandifoliola, , and ) and to determine its antimicrobial potential. Standard procedures were followed in the extraction and analysis of both aqueous and ethanolic extracts of stem bark of the African mahogany. The study identified the presences of Glycoside, Tannins, Saponins, Anthraquinones in the stem bark of all four species of the African mahogany, Terpenes are absent in Khaya anthotheca but was present in the other species. Aqueous and Ethanolic extracts were evaluated for potential antimicrobial activities against medically important bacterial and fungal strain. An antimicrobial activity was determined in the extracts using agar disc diffusion method. At a concentration of 20mg/ml, the zone of inhibition (mm) ranges from 13.5±0.50 – 24.0±0.00, for both extracts of African mahogany whiles ciprofloxacin (10µg/ml) was also in the range of 14.0±0.00 – 36.0±1.00 against all organism tested. The results showed a broad spectrum of activity against Staphylococcus aureus, Streptococcus pyogenes, Bacilius subtilise, Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi and Candida albicans. The results also identified ethanolic extract to have higher zone of inhibition than that of the aqueous extract. The analysis was carried out in the Pharmacognosy department of Kwame Nkrumah University of Science and Technology.

1.0 INTRODUCTION

1.1 BACKGROUND Herbal medicines have been known to man for centuries. Therapeutic efficacy of many indigenous for several disorders has been described by practitioners of traditional medicine. Antimicrobial properties of medicinal plants are being increasingly reported from different parts of the world (Ramasamy et al., 2009). The healing of diseases in Ghana and other part of the world make use of numerous plants species as a source of medicine. Some of these plants and their parts used in the healing processes are fairly known. Information on the use and the properties of these plants are kept secret by these traditional healers (Dokosi, 1969). About 61% of new drugs developed between 1981 and 2002 were based on natural products and they have been very successful, especially in the areas of infectious disease and cancer (Cragg, 2005). The World Health Organization estimates that plant extracts or their active constituents are used as folk medicine in traditional therapies of 80% of the world's population (Shaik et al., 1994). The harmful microorganisms can be controlled with drugs and these results in the emergence of multiple drug-resistant bacteria and it has created alarming clinical situations in the treatment of infections. The pharmacological industries have produced a number of new antibiotics; resistance to these drugs by microorganisms has increased. In general, bacteria have the genetic ability to transmit and acquire resistance to synthetic drugs which are utilized as therapeutic agents (Towers et al., 2001). Natural products of higher plants may give a new source of antimicrobial agents with possibly novel mechanisms of action (Shahidi, 2004). The effects of plant extracts on bacteria have been studied by a very large number of researchers in different parts of the world (Reddy et al., 2001). In an effort to expand the spectrum of antibacterial agents from natural resources, four species of African Mahogany were studied. These species include , Khaya ivorensis, Khaya anthotheca and Khaya senegalensis of the family . They are found in the humid forest of , ranging from Cote d’Ivoire, through Ghana and to (Irvine, 1961). K. ivorensis naturally occurs mainly within the moist semi-deciduous forest zone of Ghana, the distribution of K. anthotheca spans across three ecological zones (moist semi- deciduous forest zone, dry semi-deciduous forest zone and the transitional zone between dry semi-deciduous forest zone and Savannah zone) (Ofori et al., 2007). Khaya grandifoliola is naturally found in the dry Semi-deciduous and the moist Semi-deciduous zone (Irvine, 1961). These two ecological zones are characterized by high rainfall, short dry season and rich soil nutrient. These zones also have a wider range of environmental conditions which Khaya grandifoliola is adapted to in the natural forest (Hall and Swaine, 1981). Khaya senegalensis is also a transitional and Savannah zone tree species

1.2 PROBLEM STATEMENT In Ghana and in most developing countries, extracts from indigenous plants species are employ for the treatment of several diseases. The herbal medicines produced from these extracts are very important for the economies of these countries. The traditional knowledge of employing these local plants for medicine are not normally supported by scientific information and also not usually written down (Dokosi, 1966). This work seeks to determine and document the effect of extracts from the African mahogany on some clinically important microbes.

1.3 JUSTIFICATION Antibiotics are one of the most important drugs in fighting bacterial infections and have greatly benefited the health of human since their introduction. However, over the past few decades, the health benefits of antibiotics are under threat as many commonly used antibiotics have become less and less effective against certain bacterial infections. This is due to emergence of drug- resistant bacteria. It is therefore essential to search for newer drugs to which bacteria will have lesser resistance. In many developing countries, traditional medicine is one of the primary healthcare systems (Farnsworth, 1993). Drugs derived from natural sources play significant role in the prevention and treatment of human diseases. Scientific knowledge on the use of indigenous plant such as African Mahogany is limited, there is therefore the need to investigate the antimicrobial properties of African Mahogany extracts for proper use in the treatment of illness.

1.4 RESEARCH OBJECTIVES The research objectives were: 1. Determination of the phytochemical properties of the African Mahogany (Khaya grandifoliola, Khaya ivorensis, Khaya anthotheca and Khaya senegalensis). 2. Determination of the antimicrobial potential on some clinical important microbes.

1.5 RESEARCH QUESTION Can African Mahogany be a potential source of antimicrobial substance against disease causing organisms?

3.0 METHODOLOGY

3.1 COLLECTION OF PLANT MATERIALS The bark of stem of the four (4) African Mahogany species (Khaya grandifoliola, Khaya ivorensis, Khaya anthotheca and Khaya senegalensis) were harvested from moist semi- deciduous forest zone, dry semi-deciduous forest zone and the transitional zone between dry semi-deciduous forest zone and Savannah zone of Ghana. The samples were air-dried and then send to Kwame Nkrumah University of Science and Technology in department of Pharmacognosy for extraction and analysis.

3.2 PREPARATION OF PLANT EXTRACT The extraction of the African Mahogany was carried out using known standard procedures (Harbome, 1973). The plant material was dried in shade and powdered in a mechanical grinder. About 25.0g of the powder from the plant material was first extracted with 900 ml petroleum ether at temperature 60-80°C for 72 hours, followed by 900 ml of ethanol by using a Soxhlet extractor. The extracts were filtered using Whatman filter paper (No.1) while hot and concentrated under vacuum and reduced pressure using rotary evaporator, and dried in a desiccator to obtained dried powdered extracts. The aqueous extract of each sample was prepared by soaking 15g of the dry powdered sample in 100ml of double distilled water (DD) for 24 hours. The extract centrifuged for 10 min. at 2500 rpm, then filtered using Wattman filter paper No. 42.

3.3 PRELIMINARY PHYTOCHEMICAL SCREENING The extracts were subjected to preliminary phytochemical testing to detect for the presence of different chemical groups of compounds. The petroleum ether and the ethanol plant extracts were screened for the presence of saponins, tannins, alkaloids, flavonoids, triterpenoids, steroids, glycosides, anthraquinones, coumarin, as described in literatures, (Khandelwal, 2009 and Kumar et al, 2009). One gram of dry powdered extract was dissolved in 5ml DD. A standard procedure for identification of the various classes of active chemical constituent was used (Abdul-Aziz, 2005). Alcoholic extraction are similar to aqueous extraction expect using 70% ethanol instead of DD H2O (Okogum, 2000).

The preliminary detection of some chemical composition were performed such as flavonids that l0g of dry powdered extract (A) was dissolved in 5ml of ethanol 95% filtered; then Aliquot of 10ml of KOH 50% was added to 10ml ethanol 50% (B), equal volumes from (A) and (B) solution were mixed, a yellow color was developed indicating the presence of flavonoid (Jaffer et al, 1983).

Glycosides detected by a portion of the aqueous filtrate of each plant extracts, 1ml was placed in a clean test tube, treated with 2ml fehling reagent [1ml of fehling A + 1ml of fehling B (Vogle, 1974)] placed in a water path for 10 minutes till boiling; then cooled. The appearance of red- brown precipitate indicated the presence of saccharides. Retreated by adding 1ml of the aqueous plant extract to 5ml [Benedict’s reagent (Vogle, 1974)]. Appearance of red precipitate indicated the presence of saccharides. Addition of few drops of [kedde reagent (Al-Shahat, 1986)],

confirmed the presence of the glycoside. Since violet-blue coloration in each extract indicated the presence of glycoside (Al-Anzy, 2004).

Phenolic compound detected by a portion of the aqueous filtrate of each plant extract, 5ml was added to 1-2 drops of 1% of ferric chloride. A blue-green coloration in each extract indicated the presence of phenolic compounds (Harbom, 1973).

Tannins detected by dried powder sample 0.5g were boiled in 20ml DD H2O in test tubes; then filtered. A few drops of 0.1g ferric chloride (FeCl3) were added to the filtrate. A blue-black or brownish-green precipitate confirmed the presence of Gallic tannins or catechol tannins respectively (Trease and Evans, 1987).

Alkaloids detected by a small portion 0.2ml of the extract was stirred and placed in 5ml 1% of HCl on a steam bath, then 1ml of the filtrate was treated with 1-3 drops of Mayer’s reagent and appearance of white precipitate was evidence for the presence of alkaloid (Trease and Evans, 1987).

Terpenoids detected by 5ml of each extract was mixed with 2ml chloroform. 3ml of conc. sulfuric acid (H2SO4) was carefully added to form a layer. A reddish-brown coloration of the interface was formed to prove the presence of terpenoid (Trease and Evans, 1989).

Steroid detected by ethanolic extracts of each sample 0.5ml with 2ml (H2SO4) were added to 2ml acetic acid anhydrate. Color change from violet to blue or green in samples indicating the presence of steroids (Trease and Evans, 1989).

3.4 TEST MICROORGANISMS AND GROWTH MEDIA

The following microorganisms; Staphylococcus aureus (MTCC 96), Streptococcus pyogenes (MTCC 442), Bacillius subtilise (NTCC10073), Escherichia coli (MTCC 443), Pseudomonas aeruginosa (MTCC 424) Salmonella typhi and fungal strains, Candida albicans (MTCC 227) were chosen based on their clinical and pharmacological importance (McCracken, 1983). The bacterial strains obtained from the department of Microbiology, KNUST, were used for evaluating antimicrobial activity. The bacterial and fungal stock cultures were incubated for 24 hours at 37°C on nutrient agar and potato dextrose agar (PDA) medium respectively, following refrigeration storage at 4°C. The bacterial strains were grown in Mueller-Hinton Agar (MHA) plates at 37°C (the bacteria was be grown in the nutrient broth at 37°C and maintained on nutrient agar slants at 4°C), whereas the yeasts and molds were grown in Sabouraud dextrose agar and PDA media, respectively, at 28°C. The stock cultures were maintained at 4°C.

In vitro antibacterial and antifungal activities were examined from the aqueous and ethanol (hydroalcohol) extracts. Antibacterial and antifungal activities of plant part extracts against six pathogenic bacteria (three Gram-positive and negative) and one pathogenic fungi were investigated by the agar disk diffusion method (Alzoreky, 2003), (Bauer et al, 1966). Antimicrobial activity testing was carried out by using agar cup method. Each purified extracts were dissolved in sterile double distilled water. For the determination of zone of inhibition, pure Gram-positive, Gram-negative, and fungal strains were taken as a standard antibiotic for comparison of the results. All the extracts were screened for their antibacterial and antifungal

activities against the Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Streptococcus pyogenes, Salmonella typhi, Bacillius subtilise and the fungi Candida albicans. The sets of four dilutions (2.5, 5.0, 10, and 20 mg/ml) of the African Mahogany extract and standard drugs were prepared in double-distilled water using nutrient agar tubes. Mueller-Hinton sterile agar plates were seeded with indicator bacterial strains (108 cfu) and allowed to stay at 37°C for 3 hours. Control experiments were carried out under similar condition by using ciprofloxacin for antibacterial activity and antifungal activity as standard drug. The zones of growth inhibition around the disks were measured after 24 hours of incubation at 37°C for bacteria and fungi. The sensitivities of the microorganism species to the plant extracts were determined by measuring the sizes of inhibitory zones (including the diameter of disk) on the agar surface around the disks.

4.0 RESULTS AND DICUSSION

4.1 RESULTS

The extracts from the African Mahoganies were analyzed and came out with the results presented below.

Table 1: Preliminary detection of secondary compounds in four species of African Mahogany

RESULTS

TEST Khaya anthotheca Khaya Khaya Khaya ivorensis grandifoliola senegalensis Aqueous Ethanol Aqueous Ethanol Aqueous Ethanol Aqueous Ethanol Extract Extract Extract Extract Extract Extract Extract Extract

Glycoside +++ ++ ++ + + + ++ ++ Flavonoids ------

Tannins + + + + + + + + Saponins ++ + ++ - ++ ++ ++ ++ Steriods ------Terpenes - - ++ + ++ + + +

Alkaloids ------Anthraquinones + - + - + + + -

key: (+) = Presence (+ = low, ++ = moderate, +++ = high); (-) = Absence

Table 2: Antimicrobial Zone of Inhibition (mm) Activity of Aqueous and Ethanol Extract from Stem Back of Khaya grandifoliola.

Concentration (mg/ml) Control Test Isolates Extracts 2.5 5 10 20 (cipro) 10 µg/ml

Staphylococcus Aqueous 0.0 ± 0.00 0.0 ± 0.00 12.5 ± 0.50 14.5 ± 0.50 16.0 ± 1.00 aureus Ethanol 11.5 ± 0.50 13.5 ± 0.50 16.0 ± 0.00 20.0 ± 0.00 19.0 ± 1.00 Streptococcus Aqueous 0.0 ± 0.00 0.0 ± 0.00 12.5 ± 0.50 14.0 ± 0.00 16.5 ± 0.50 pyogenes Ethanol 11.5 ± 0.50 11.5 ± 0.50 14.0 ± 0.00 18.5 ± 0.50 19.5 ± 0.50

Candida Aqueous 0.0 ± 0.00 0.0 ± 0.00 13.5 ± 0.50 14.5 ± 0.50 20.5 ± 0.50 albicans Ethanol 12.5 ± 0.50 12.5 ± 0.50 15.0 ± 1.00 16.5 ± 0.50 19.5 ± 0.50 Pseudomonas Aqueous 0.0 ± 0.00 11.5 ± 0.00 13.5 ± 0.50 14.5 ± 0.50 26.5 ± 1.5 aeruginosa Ethanol 11.5 ± 0.50 14.0 ± 0.00 16.5 ± 0.50 18.5 ± 0.50 21.0 ± 2.00

Salmonella Aqueous 0.0 ± 0.00 12.0 ± 0.00 12.5 ± 0.50 13.5 ± 0.50 20.5 ± 0.00 typhi Ethanol 12.0 ± 0.00 13.0 ± 0.00 16.5 ± 0.50 17.5 ± 0.50 19.5 ± 1.00

Escherichia Aqueous 0.0 ± 0.00 0.0 ± 0.00 14.0 ± 0.00 15.5 ± 0.50 20.0 ± 0.00 coli Ethanol 12.0 ± 0.00 13.0 ± 0.00 16.5 ± 0.50 17.5 ± 0.50 19.0 ± 1.00

Bacilus subtilis Aqueous 0.0 ± 0.00 0.0 ± 0.00 12.5 ± 0.50 13.5 ± 0.50 14.5 ± 0.50 Ethanol 13.0 ± 1.00 13.0 ± 1.00 15.5 ± 0.50 19.0 ± 1.00 19.0 ± 1.00

Cipro = Ciprofloxacin

Table 3: Antimicrobial Zone of Inhibition (mm) Activity of Aqueous and Ethanol Extract from Stem Back of Khaya ivorensis.

Concentration (mg/ml) Control Test Isolates Extracts 2.5 5 10 20 (cipro) 10 µg/ml

Staphylococcus Aqueous 11.5 ± 0.50 12.5 ± 0.50 13.5 ± 0.50 14.5 ± 0.50 17.5 ± 0.50 aureus Ethanol 13.5 ± 0.50 15.0 ± 0.00 15.5 ± 0.00 17.5 ± 0.50 31.5 ± 0.50

Streptococcus Aqueous 11.5 ± 0.50 12.5 ± 0.50 12.5 ± 0.50 14.5 ± 0.50 15.5 ± 0.50 pyogenes Ethanol 15.5 ± 0.50 16.0 ± 0.00 17.5 ± 0.50 19.5 ± 0.50 32.5 ± 2.50

Candida Aqueous 0.0 ± 0.00 0.0 ± 0.00 12.5 ± 0.50 14.5 ± 0.50 15.5 ± 0.50 albicans Ethanol 14.5 ± 0.50 15.5 ± 0.50 16.5 ± 0.50 20.5 ± 0.50 29.5 ± 0.50 Pseudomonas Aqueous 11.5± 0.50 13.5 ± 0.50 14.5 ± 0.50 15.5 ± 0.50 30.5 ± 0.50 aeruginosa Ethanol 15.0 ± 0.50 16.5 ± 0.50 19.5 ± 0.50 20.5 ± 0.50 33.0 ± 2.00

Salmonella Aqueous 0.0 ± 0.00 13.5 ± 0.50 13.5 ± 0.50 13.5 ± 0.50 23.0 ± 1.00 typhi Ethanol 14.5 ± 0.00 15.0 ± 0.50 16.5 ± 0.50 20.5 ± 0.50 27.5 ± 2.5

Escherichia Aqueous 11.5 ± 0.50 12.5 ± 0.50 13.5 ± 0.50 15.5 ± 0.50 23.0 ± 1.00 coli Ethanol 13.5 ± 0.50 16.5 ± 0.50 18.5 ± 0.50 20.5 ± 0.50 29.5 ± 0.50

Bacilus subtilis Aqueous 11.5 ± 0.50 12.5 ± 0.50 13.5 ± 0.50 15.5 ± 0.50 16.5 ± 0.50 Ethanol 14.5 ± 0.50 14.5 ± 0.50 15.5 ± 0.50 20.5 ± 0.50 27.5 ± 2.5

Cipro = Ciprofloxacin

Table 4: Antimicrobial Zone of Inhibition (mm) Activity of Aqueous and Ethanol Extract from Stem Back of Khaya senegalensis.

Concentration (mg/ml) Control Test Isolates Extracts 2.5 5 10 20 (cipro) 10 µg/ml

Staphylococcus Aqueous 14.5 ± 0.50 16.5 ± 0.50 17.5 ± 0.50 20.5 ± 0.50 33.5 ± 1.5 aureus Ethanol 14.5 ± 0.50 15.0 ± 0.00 17.5 ± 0.00 19.5 ± 0.50 25.5 ± 0.50

Streptococcus Aqueous 12.0 ± 0.00 13.0 ± 0.00 13.5 ± 0.50 14.5 ± 0.50 30.5 ± 0.50 pyogenes Ethanol 13.0 ± 0.00 13.5 ± 0.50 17.0 ± 1.00 17.0 ± 1.00 23.0 ± 1.00

Candida Aqueous 0.0 ± 0.00 13.5 ± 0.50 15.5 ± 0.50 16.5 ± 0.50 30.5 ± 0.50 albicans Ethanol 11.0 ± 0.00 10.5 ± 0.50 14.5 ± 0.50 15.0 ± 1.00 20.5 ± 0.50 Pseudomonas Aqueous 12.5± 0.50 13.5 ± 0.50 14.5 ± 0.50 15.5 ± 0.50 29.5 ± 0.50 aeruginosa Ethanol 12.5 ± 0.50 13.5 ± 0.00 14.5 ± 0.50 15.5 ± 0.50 22.0 ± 0.00

Salmonella Aqueous 12.0 ± 0.50 13.5 ± 0.50 14.5 ± 0.50 16.5 ± 0.50 29.5 ± 0.50 typhi Ethanol 10.5 ± 0.50 11.5 ± 0.50 13.5 ± 0.50 16.0 ± 0.00 21.5 ± 0.50

Escherichia Aqueous 12.0 ± 0.00 13.0 ± 0.00 13.5 ± 0.50 15.0 ± 0.50 29.5 ± 0.50 coli Ethanol 14.5 ± 0.50 15.5 ± 0.50 16.5 ± 0.50 17.5 ± 0.50 23.0 ± 1.00

Bacilus subtilis Aqueous 12.5 ± 0.50 13.0 ± 0.50 13.5 ± 0.50 15.5 ± 0.50 28.5 ± 0.50 Ethanol 13.5 ± 0.50 18.0 ± 0.50 18.0 ± 1.00 19.5 ± 0.50 26.5 ± 0.50

Cipro = Ciprofloxacin

Table 5: Antimicrobial Zone of Inhibition (mm) Activity of Aqueous and Ethanol Extract from Stem Back of Khaya anthotheca.

Concentration (mg/ml) Control Test Isolates Extracts 2.5 5 10 20 (cipro) 10 µg/ml

Staphylococcus Aqueous 0.0 ± 0.00 12.0 ± 0.00 12.5 ± 0.50 13.5 ± 0.50 22.5 ± 0.50 aureus Ethanol 14.5 ± 0.50 15.5 ± 0.50 18.5 ± 0.50 20.5 ± 0.50 36.0 ± 1.00 Streptococcus Aqueous 0.0 ± 0.00 0.0 ± 0.00 0.0 ± 0.00 13.5 ± 0.50 25.5 ± 0.50 pyogenes Ethanol 15.0 ± 0.00 16.5 ± 0.50 18.5 ± 0.50 20.5 ± 0.50 31.0 ± 1.00

Candida Aqueous 11.5 ± 0.50 12.0 ± 0.00 13.5 ± 0.50 14.0 ± 0.00 15.5 ± 0.50 albicans Ethanol 15.5 ± 0.50 16.5 ± 0.50 18.5 ± 0.50 21.5 ± 1.00 27.5 ± 0.50 Pseudomonas Aqueous 0.0± 0.00 12.0 ± 1.00 12.5 ± 0.50 14.5 ± 0.50 23.5 ± 0.50 aeruginosa Ethanol 14.5 ± 0.50 15.5 ± 0.50 17.5 ± 0.50 19.5 ± 0.50 27.5 ± 0.50

Salmonella Aqueous 0.0 ± 0.00 0.0 ± 0.00 0.0 ± 0.00 12.5 ± 0.50 25.5 ± 0.50 typhi Ethanol 14.5 ± 0.50 16.5 ± 0.50 18.5 ± 0.50 20.5 ± 0.50 27.5 ± 0.50

Escherichia Aqueous 0.0 ± 0.00 0.0 ± 0.00 13.5 ± 0.50 15.0 ± 0.00 25.5 ± 0.50 coli Ethanol 15.5 ± 0.50 17.5 ± 0.50 19.5 ± 0.50 21.5 ± 0.50 33.5 ± 1.50

Bacilus subtilis Aqueous 0.0 ± 0.00 0.0 ± 0.00 0.0 ± 0.00 12.0 ± 0.00 14.0 ± 0.00 Ethanol 14.5 ± 0.50 17.5 ± 0.50 19.5 ± 0.50 24.0 ± 1.00 29.5 ± 0.50

Cipro = Ciprofloxacin

4.2 DICUSSION So many studies have reported the phytochemical constituents in the plant samples are known to be biologically active compounds and they are responsible for different activities such as antioxidant, antihelmentic, antimicrobial, antifungal, and anticancer (Abdelgaleil et,al., 2004) and (Ademola et, al., 2004). All secondary metabolite components displayed antimicrobial properties through different biological mechanisms. Most of the secondary metabolite components were isolated and identified in the polar plant crude extracts (Adesida et, al., 1971) and (Wu et, al., 2014). The phytochemical screening of both aqueous and ethanolic crude extracts from dry powder of stem barks samples of the African Mahoganies (Khaya grandifoliola, Khaya ivorensis, Khaya anthotheca and Khaya senegalensis) showed the presence of Glycoside, Tannins and Saponins. Terpenes were present in both aqueous and ethanolic extracts of Khaya grandifoliola, Khaya ivorensis, and Khaya senegalensis but absent in Khaya anthotheca. Anthraquinones is present in both Aqueous and ethanolic extracts of Khaya senegalensis but absent in the ethanolic extract of Khaya grandifoliola, Khaya ivorensis, and Khaya anthotheca. Flavonoids, Steriods and Alkaloids were absent in all of the fours species analyzed as shown in Table 1. Similar studies by Kankia and Zainab (2015) confirm absence of Steriods in extracts of\stem bark of Khaya senegalensis but dispute the presence of Alkaloids in the stem bark of the same species.

Antimicrobial activity screening of both aqueous and Ethanolic extracts of all four species of African mahogany showed broad spectrum of activity. It can be seen from tables 2, 3, 4, and 5 that Ethanolic extract is the most active against the seven human pathogens tested.

Aqueous extract of stem back of Khaya grandifoliola (Table 2) was inactive at concentration of 2.5mg/ml and 5mg/ml against Staphylocuccus aureus, Streptococcus pyogenes, Candida albicans, Escherichia coli and Bacilus subtilis. It was also inactive at concentration of 2.5mg/ml to Pseudomonas aeruginosa and Salmonella typhi. Ethanol extract of Khaya gradifoliola was active at all concentrations against all the pathogens. Staphylocuccus aureus was more susceptible at 20mg/ml giving zone of inhibition of 20mm while ciprofloxacin (standard, 10 µg/ml) gave zone of inhibition of 19mm. Ethanol extract at 20mg/ml gave the same zone of inhibition to that of the standard (i.e. 19mm) against Bacilus subtilis.

There was more activity with aqueous extract of Khaya ivorensis (Table 3), however there was no activity against Candida albicans and Salmonella typhi at 2.5mg/ml, 5mg/ml and 2.5mg/ml respectively. Ethanol extract exhibited greater zone of inhibition among all the pathogen tested.

Candida albicans and Salmonella typhi were more affected by the aqueous extract as compared with its alcohol extracts `of Khaya senegalensis (Table 4). There was no activity against Candida albicans at 2.5mg/ml.

Streptococcus pyogenes, Bacilus subtilis and salmonella typhi were not affected by the aqueous extract of Khaya anthotheca (Table 5) at 2.5, 5, and 10mg/ml, also there was no activity by aqueous extract against Escherichia coli at 2.5 and 5mg/ml. Staphylococcus aureus and Pseudomonas aeruginosa exhibited no activity at a concentration of 2.5mg/ml of the aqueous extract. There was activity against all the pathogens of the ethanol extract.

Generally, the antimicrobial activity of plant crude extracts depends on the dose and the type of bacterial strains employed (Adebayo et, al., 2003). Also these antibacterial actions could be related to their chemical components in the crude extracts. The bioactive compounds such as tannins and saponins components were present in the crude extracts. However, these bioactive compounds were inducing antimicrobial activities (Zhang et, al., 2009). The amount of active components in the crude extract may be diluted or increased their concentrations by fractionation, because they have the ability to inactivate microbial activity, enzymes, cell envelope transport proteins, and so forth (Abdelgaleil et,al., 2004) and (Zhang et, al., 2009).

5.0 CONCLUSION AND RECOMMENDATION

5.1 CONCLUSION This study has provided the scientific basis for the traditional use of the African Mahogany. From the results obtained from this work, it can be concluded that the stem bark of Khaya anthotheca, Khaya grandifoliola, Khaya senegalensis, Khaya ivorensis contains Glycoside, Tannins, and Saponins. Terpene is present in all the species except Khaya anthotheca. There are also traces of Anthraquinones in all species tested. It has also been confirmed that the stem bark possesses a broad spectrum of antimicrobial activity against Staphylocuccus aureus, Streptococcus pyogenes, Candida albicans, Escherichia coli and Bacilus subtilis, Pseudomonas aeruginosa and Salmonella typhi.

The broad spectrum of antimicrobial activity exhibited by species of African Mahogany in this work may be attributed to the various active components present in it. The use of stem bark of African Mahogany as medicine by local folks suggests that they represent an economic and safe alternative to the treatment of diseases.

5.2 RECOMMENDATIONS In other to obtain maximum antimicrobial activity, I recommend that; 1. Hydroalcohol is used for the extraction of active components in the stem bark of African mahogany. 2. Only mature stem back should be harvested in other not to kill the plant. 3. Traditional healers should be educated on the proper way of harvesting stem bark for medicine. 4. Proper sun drying should be carried out in other to prevent fungal growth on the stem bark before preparation for medicinal use. 5. Further studies are required for the isolation and identification of individual active compounds and also in vivo studies are needed for better understanding of their mechanism of action as antimicrobials.

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