10 L of the inoculum and 100 L of each EO was added to the first
well of the microtiter plate. To study synergistic effects, we followed
the same protocol, but 50 L of E. globulus and 50 L of E.
urograndis (1:1) were used. Next, 100 L of the culture was successively
transferred to the next well, thereby obtaining a reduction of
50% after each transfer. The plates were then incubated at 37 ◦C for
24 h. After incubation, 10 L of 3% resazurin (Sigma®), was added
which is a redox indicator that has been used to assess viability
and bacterial contamination, and each dilution microtiter plate was
again incubated at 37 ◦C for 30 min. As a positive control for bacterial
growth, BHI broth and inoculum was added to the first well
of the microtiter plate. As a negative control for bacterial growth,
only BHI was added to the last well ofthe microtiter plate. The minimum
inhibitory concentration (MIC) of an antimicrobial agent is
defined as the lowest concentration of compound resulting in no
visible growth after 24 h of incubation.
2.5.