A microdilution broth susceptibility assay was used as
recommended by Natural Committee for Clinical Laboratory
Standard-NCCLS (1999) for the determination of the MIC. All tests
were performed in nutrient broth for bacteria and in potato dextrose
broth for yeast and fungi. Concentrations of 7.5, 10, 12.5, 15
and 20 g dry extract or essential oil were added to 1 ml nutrient
broth tubes containing 105 CFU/ml of live microorganism’s cells.
The tubes which contained 10 ml broth were incubated on an incubator
shaker as to evenly disperse the extracts and essential oils
throughout the broth in the tubes. The highest dilution (lowest concentration),
showing no visible growth, was regarded as MIC. Cells
from the tubes showing no growth were subcultured on nutrient
agar plates to determine if the inhibition was reversible or permanent.