25mm i.d., 0.25��m film thickness; Agilent Technologies, Hewlett-Packard, CA, USA) was used; the flow of the carrier gas (N2) was 1.6mL/min and the split ratio 60:1. Analyses were performed using the following temperature program: oven temps isotherm at 35��C for 10min, from 35 to 205��C at the rate of 3��C/min, and isotherm at 205��C selleck inhibitor over 10min. Injector and detector temperature were held, respectively, at 250 and 300��C.2.3. Gas Chromatography-Mass SpectrometryGC-MS analyses of essential oil volatile components were carried out on a gas chromatograph HP 5890 (II) coupled to a HP 5972 mass spectrometer (Agilent Technologies, Palo Alto, CA, USA) with electron impact ionization (70eV). A HP-5MS capillary column (30m��0.25mm, 0.25��m film thickness; Agilent Technologies, Hewlett-Packard, CA, USA) was used.
The column temperature was programmed to rise from 50��C to 240��C at a rate of 5��C/min. The carrier gas was helium with a flow rate of 1.2mL/min; split ratio was 60:1. Scan time and mass range were 1s and 40�C300m/z, respectively.2.4. Compounds IdentificationIdentification of essential oil volatile compounds was based on the calculation of their retention indices (RI) relative to (C8�CC22) n-alkanes with those of authentic compounds available in our laboratory. Further identification was made by matching their recorded mass spectra with those stored in the Wiley/NBS mass spectral library of the GC-MS data systems and other published mass spectra [16].2.5.
Screening of Antibacterial ActivityAntibacterial activity was analyzed by the disc diffusion method [17] against three human pathogenic bacteria including Gram-positive Staphylococcus aureus (ATCC 25923), and Gram-negative bacteria Escherichia coli (ATCC 35218) and Pseudomonas aeruginosa (ATCC 27853). All bacteria were grown on Mueller Hinton plate at 30��C for 18�C24h previous inoculation onto the nutrient agar. A loop of bacteria from the agar slant stock was cultivated in nutrient broth overnight and spread with a sterile cotton swap onto Petri dishes containing 10mL of API suspension medium and adjusted to the 0.5 McFarland turbidity standards with a Densimat (BioMerieux). Sterile filter paper disks (6mm in diameter) impregnated with 10��L of essential oil were placed on the cultured plates. After 1-2h at 4��C, the treated Petri dishes were incubated at 25 or 37��C for 18�C24h.
The antimicrobial activity was evaluated by measuring the diameter of the growth inhibition Drug_discovery zone around the discs. Each experiment was carried out in triplicate, and the mean diameter of the inhibition zone was recorded.2.6. Statistical AnalysesAll data are reported as means �� standard deviation of three samples. Statistical analysis was performed with STATISTICA. Differences were tested for significance by using the ANOVA procedure, using a significance level of P < 0.