Effect of Saccocalyx satureioides essential oil on the growth of Ascochyta rabiei 

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... and plant pathogenic fungi. They are needed for reduction in the use of chemical in agriculture increases interest in the possibility of the application of essential oils to control plant pathogens [7]. A good number of essential oils are reported to be effective against many phytopathogenic fungi [20]. Therefore, the purpose of this work was to evaluate the fungitoxic effect in vitro of Saccocalyx satureioides on the phytopathogenic fungus Ascochyta rabiei . The aerial parts of Saccocalyx satureioides were harvested during morning hours in June 2005, in El Masrane near Djelfa, Algeria. Plants were at the flowering stage, at the beginning of seed formation. The leaves were separated from the stems and air-dried. This material was then hydrodistilled (4 h) using a Clevenger type apparatus, according to the European Pharmacopoeia. The essential oil, obtained in a yield of 1.5% (w/w), was dried over anhydrous sodium sulphate and stored in hermetically sealed coloured vials at 4°C before analysis. Essential oil analysis were done by GC/MS [14]. The oil was first dissolved in water containing tween 80 (10% v/v) at a solution concentration of 10 % (w/v). Two ml of this solution was added to 28 ml of Czapek Dox Cation Liquid Medium (CDCLM) medium (Oxoid) (30 ml in 250 ml flask) and a series a dilution was made, 0.6% (6 mg/ml), 0.3% (3 mg/ml), 0.15% (1.5 mg/ml), 0.075% (0.75 mg/ml), 0.0375 % (0.375 mg/ml),0.01875 % (0.1875 mg/ml) with three replicates for each dilution. The medium was inoculated with 30 ml solution (10 7 spores /ml) of Tk21 isolates and incubated at 20 °C for 14 days. After incubation the mycelium was recolted by filtering through four layers of muslin and dry in for 72 h. to measure the dry weight. The solution was partitioned three times with 1/3 rd volume ethyl acetate, and taken to dryness on a rotary evaporator at < 35 °C. The residue was dissolved in 2 ml methanol and analysed by analytical high performance liquid chromatography on a Philips HPLC equipped with a diode array detector essentially according to Hamid and Strange [11] except that the solvent system consisted of water 60%, methanol 20.1%, tetrahydrofuran 18.1% and acetonitrile 1.8% (v/v/v/v) which was pumped at a flow rate of 1 ml/min. The stationary phase was an ODS column (Spherisob ODS 2; 150 x 4.5 mm diam.; Jones Chromatography, Glamorgan, UK) which was protected by a guard column (20 x 4.6 mm diam.). After 14 days the mycelium was collected and the dry weight measured. A. rabiei did not grow at a final concentration of 6 mg/ml, 3 mg/ml and 1.5 mg/ml. At 0.75 mg/ml, 0.375 mg/ml there was little growth of the fungus, 0.08 μg/ml and 0.042 μg/ml respectively but no solanapyrone A produced (figure 1). The growth of A. rabiei was reduce by 50% at a the essential oil concentration of 0.0187 mg/ml with the production of 6μg/ml of solanapyrone A. However in the fungus culture filtrates with the concentration of 6 mg/ml, 3 mg/ml and 1.5 mg/ml of the essential oil, a new compound appeared at the HPLC at 10 mn 30 sec compared with the solanapyrone A which elutes at nearly 14 minutes (figures 2, 3,4 and 5). The inhibition of the the growth of A. rabiei by S. saturoides essential oil my be attributed to its majors contents such as α -terpineol, thymol and borneol. Forty-two compounds, representing 94.1% of the total oil, were identified in the essential oil of S. satureioides . The main constituents were α -terpineol (35.9%), thymol (15.6%) and borneol (12.4%), followed by p cymene (7.2%), camphene (4.3%) and γ -terpinene (4.0%) [14]. The 1/10 diluted diluted essential oil of this plant inhibited completely the growth of Candida albicans, Aspergillus fumigatus and Aspergillus niger [14]. Many other researchers related the correlettion between the chemical composition of the essential oils and their antifungal activity. The essential oil of Thymus eriocalyx and Thymus X-porlock inhibited the growth and the aflatoxin production of Aspergillus parasiticus at a concentration of 250 ppm [3]. The essentials oils of Thymus vulgaris has been reported to inhibit fungal growth. Their fungistatic activity has been attributed to the presence of thymol, at 50.06% in the oil tested [23]. The essential oil of Thymus numidicus , endemic species of East Algeria, inhibited completely the growth of Candida albicans , and its antifungal effect was 31 fold more important that hymus T. vulgaris . There was a positive correlation between antifungal activity and phenols (thymol and carvacrol) content [10]. In ...