2.6.6. Antibacterial Activity against *P. Intermedia* and *S. Mutans*

The antibacterial activity was determined using the microtiter plate assay, where the MIC was calculated; against *Streptococcus mutans* (ATCC 25175) and *Prevotella intermedia* (ATCC 25611). In short, a 96-well plate was supplemented with 0.1 mL of Brain Heart Infusion broth for *S. mutans* and Tryptone Soy Broth for *P. intermedia*. To that, 100 μL of plant extract dissolved in 2.5% DMSO (50 mg/mL) and 100 μL of Chlorhexidine (CHX) dissolved in water (1.25 mg/mL) were serially diluted down to give concentrations ranging from 0.048 to 12.5 mg/mL and 4.8 <sup>×</sup> 10−<sup>3</sup> to 0.625 mg/mL, for the plant extract and positive control respectively. After which 24 h old *S. mutans* and *P. interdia* inocula (3 <sup>×</sup> 108 CFU/mL), grown at 37 ◦C under anaerobic conditions in an anaerobic jar containing Anaerocult®A, were added. The plates were then incubated for a further 24 h, followed by the addition of 20 μL of PrestoBlue® viability reagent. In addition to CHX, DMSO (2.5%) and the respective media were used as negative controls [21,22].
