Study of the Antimicrobial effect of Melia azedarach L . plant حبحبسلا تابنل هيموثرجلا دض ةيلاعفلل هسارد

This study was aimed to evaluate the antimicrobial activity of the leaf extracts of Melia azedarach L. plant against different pathogenic microorganisms such as (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, klebsilla sp., Candida albicans). Two plant extracts (Aqueous and ethanolic) under four different concentrations (25, 50, 75, 100) mg/ml were used by Agar-well diffusion method. Chemical detection of extract showed that the extracts contain tannins, flavonoids, terpins, steriods, alkaloids, and saponins. Aqueous leaf extract showed no effect against all tested microorganisms at (25, 50, 75, 100) mg/ml concentration except Candida albicans which was sensitive to 100mg/ml concentration. While Ethanolic (80%) leaf extract showed sensitivity on Staphylococcus aureus at 100mg/ml concentration. صلختسملا حبحبسلا تابن قارولا يلوحكلاو يئاملا صلختسملل ةيطيبثتلا ةردقلا مييقت وه هساردلا هذه نم فدهلا Melia azedarach لثم هضرمملا ةيرهجملا ءايحلاا ومن ىلع (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, klebsilla sp , ( Candida albicans ، ) زيكارتلابو 25،50،75،100 لم/مغلم( رفحلا ةقيرط لامعتساب كلذو . تابنلل يئايميكلا فشكلا يرجا تابكرملا ىلع تابنلا ءاوتحا جئاتنلا ترهظاو ةلاعفلا صلختسملا يف تاديوريتسلاو تانيبرتلاو تانيناتلا لثم يلوحكلا صلختسملا اما ، يئاملا يوتحي تلاو تادينوفلافلاو تانيناتلا ىلع يتسلاو تانيبر ور كلاذكو تادي ةريمخلا ادع ام همدختسملا ةيرهجملا تانئاكلا ىلع يئاملا صلختسملل طيبثت يا رهظي مل . تاينوباصلا Candida albicans زيكرتلل اهسسحت تدبا دقف 100 لم/مغلم ، امنيب ايرتكبلا ترهظا Staphylococcus aureus اهسسحت يلاعلا زيكرتلل 100 نم لم/مغلم صلختسملا لا يلوحك 80 لوناثيا% Introduction The demand for herbal medicinal plant is growing very fast in recent years, because of the patient’s immunity to medicine and its side effects [1]. The screening of plant extracts and plant products for antimicrobial activity has shown that higher plants represent potential source of antibiotic prototypes [1]. Herbal medicines are dietary supplements that contain herbs, either singly or in mixture. An herb also is a plant or plant part used for its scent, flavor, and/or therapeutic properties. Products made from botanical parts that are used to maintain or improve health are called herbal supplements, botanicals, or phytomedicines [2]. Melia azedarach Linn. belongs to (Meliaceae) family, commonly known as “Persian Lilac”, is one of these important plants which contain a number of medicinally compounds [3]. M. azedarach is used orally and topically as an antiparasitic and antifungal agent [4,5]. Chemical composition reveals the presence of alkaloids,

The demand for herbal medicinal plant is growing very fast in recent years, because of the patient's immunity to medicine and its side effects [1].The screening of plant extracts and plant products for antimicrobial activity has shown that higher plants represent potential source of antibiotic prototypes [1].Herbal medicines are dietary supplements that contain herbs, either singly or in mixture.An herb also is a plant or plant part used for its scent, flavor, and/or therapeutic properties.Products made from botanical parts that are used to maintain or improve health are called herbal supplements, botanicals, or phytomedicines [2].Melia azedarach Linn.belongs to (Meliaceae) family, commonly known as "Persian Lilac", is one of these important plants which contain a number of medicinally compounds [3].M. azedarach is used orally and topically as an antiparasitic and antifungal agent [4,5].Chemical composition reveals the presence of alkaloids, tannins, meliotannic acid, benzoic acid, vanillic acid, and others [6,7].The aim of this current work was to evaluate the antimicrobial potential of M. azedarach extracts in different solvents against some pathogenic microorganisms.

Materials and Methods Preparation of leaf extracts
Healthy plant leaves were collected from the University of Al-Nahrain garden, in April 2011and 2012 and identified at the Department of Biology, University of Baghdad.The leaves washed thoroughly in running tap water, 100 grams of fresh leaves of M. azedarach plant were weighed and put in conical flask with 1000ml of distilled water to prepare the aqueous extract, then in water bath for 2hr at 40P o P C. The extract was filtered through whatman No.1 filter paper.The filtrate was concentrated in vacuum using rotary evaporator at 40P o P C then the extract was sterilized with millipore filter (o.22µm) [8].100gm of fresh leaves were blended with 500ml of alcohol (80%ethanol) and stored at room temp.for 5 days.The extract was filtered through cheesecloth then through whatman No.1 filter paper.The filtrate was concentrated by using rotary evaporator at 45P o P C. Both extracts were left at 4P o P C until used in the assay.The extraction was repeated three times.

Test microorganisms
Four strains of Gram-positive and Gram negative bacteria were used to estimate the antimicrobial activity were obtained from Biotechnology Research Center at Al-Nahrain University.The microbial strains used in the study were (Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, klebsilla sp.) and Candida albicans.

Antimicrobial assay
Muller Hinton Agar medium (MHA) was used for the antibacterial susceptibility study.The bacterial assay was performed by agar well diffusion method [9,10].(3.1 g/100 ml) of (MHA) was weighed and dissolved in 100 ml of distilled water, then sterilized by autoclaving and was allowed to cool at room temperature.The medium was poured into sterile Petri plates.The culture medium was inoculated with the given microorganisms by spreading the bacterial inoculums on the media.Microorganisms' concentration was 10P 2 P .Wells (6) mm were punched into the agar and filled with the extracts.Four different concentrations of plant extracts (25,50,75,100) mg/ml filled the wells separately.The plates were incubated at 37ºC for18 h. and the antibacterial activity was determined by measuring the diameters of inhibition zone in millimeter.Controls were maintained where pure solvents were used as negative controls instead of the extracts.The experiment was repeated three times and the mean values were presented.

Phytochemical analysis
The preliminary phytochemical screening of different extracts was done to ascertain the presence of bioactive components.The presence of alkaloids, flavonoids, tannins, terpenes, steroids, and saponins was determined [11,12], Table (1)P P showed the results The results showed also that aqueous leaf extract of M. azedarach did not possesse antibacterial activity against tested gram positive, and gram negative bacteria (Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli and Klebsiella pneumoniae) at all concentrations (25, 50, 75,100) mg/ml of aqueous and solvent leaf extract as shown in Table (2).While the ethanolic leaf extract of M. azedarach showed that it possessed antibacterial activity against Staphylococcus aureus bacteria at 100 mg/ml.The Pseudomonas aeruginosa, Escherichia coli and Klebsiella pneumonia were resistant to ethanolic leaf extract.While the ethanolic leaf extracts showed a slight effect on Candida albicans in aqueous leaf extracts Table (2).

Discussion
Aqueous extract did not show any antibacterial activity against all the bacteria studied.Similar results have been reported in the literature [13,14,15,16,17].The difference in sensitivity might be ascribed to the difference in morphological constitutions between Gram-positive and Gram-negative organisms.Many plant species present inhibition zones of differing diameters; however, size difference of the inhibition zone depends primarily upon many factors for e.g.diffusion capacity of substances (present in the extracts) in the agar medium, antimicrobial activity of diffused substances, growth and metabolic activity of microorganisms in the medium.Inhibition zone diameter can further be associated with polarities of substances which make up the tested extracts, and also with cell wall composition of tested organisms since Gram-positive bacteria present cell walls with lower lipid levels than do Gramnegative bacteria [18].P P