2.3. Antifungal and Antibiofilm Formation Susceptibility Test

The anti-biofilm activity of dendrons for inhibition of biofilm formation was performed as previously described [9]. An inoculum of C. albicans was adjusted to a density equivalent to 0.5 McFarland standard (1 × 10⁶ cells/mL, Grant Instruments, Royston, UK) in RPMI 1640 medium (Sigma-Aldrich) with morpholinepropanesulfonic acid (MOPS, Sigma-Aldrich) and 2% glucose (Scharlab, Barcelona, Spain) (RPMI + MOPS + GLU). Then, 50 μL of the suspension was inoculated in 96-well microtiter plates containing two-fold serial dilutions of the dendrons ranging from 2 to 512 mg/L. Controls were included in all experiments: un-inoculated medium and dendron free medium. Plates were sealed with Parafilm^® (Bemis, Neenah, WI, USA) and incubated for 48 h at 37 °C. Then, resazurin colorimetric assay (Section 2.6) and drop plate method (Section 2.7) were used to determine the minimum biofilm inhibitory concentration (MBIC) and the minimum fungicidal concentration (MFC), respectively. The MBIC was defined as the lowest concentration at which no growth (resazurin reduction) was observed [9]. The MFC was defined as the lowest concentration for which no growth was observed when plating 5 μL suspension of each well, and, in consequence, a biofilm was not formed.