2.6.5. Determination of Minimum Inhibitory Concentration (MIC)

MIC values were defined as the lowest concentration of each natural product, which completely inhibited microbial growth. The results were expressed in milligrams per milliliter [19]. MIC was determined as describe by [20] with some modifications.

The MIC value of compound and CME were determined by 10 serial dilutions (500, 250, 125, 62.5, 31.25, 15.62, 7.81, 3.95, 1.98, 0.99 and 0.49 µg/mL). Equal volumes of test compound (dilution) and nutrient broth were mixed in each test tube. Inoculum (20 µL, 108 cfu) was also added in each tube. The tubes were incubated aerobically at 37 °C for 24 h. Two control tubes for each dilution were maintained. One included test compound (each dilution) and growth medium while second tube contained growth medium, physiological saline and inoculum. MIC was determined as the lowest concentration of the compound permitting no visible growth (no turbidity) when compared with the control tubes [20].