5.5. Antimicrobial Activity Assay

Compounds were tested in a microtiter plate assay according to CLSI guidelines against E. coli DSM 30083, A. baumannii DSM 30007, P. aeruginosa DSM 50071, B. subtilis DSM 402, S. aureus DSM 20231, S. aureus ATCC 43300 (MRSA), and C. albicans, as described previously [33]. E. coli, A. baumannii, S. aureus, and B. subtilis were grown in Mueller Hinton broth, P. aeruginosa in cation-adjusted Mueller Hinton II, and C. albicans DSM1386 in Sabouraud broth. Compounds were dissolved in DMSO at 10 mg/mL. Starting from a 10 mg/mL stock solution, serial dilutions in culture media were prepared with the Tecan Freedom Evo 75 liquid handling workstation (Tecan, Männedorf, Switzerland), covering a range from 512 to 0.5 µg/mL. Assay volumes were 200 µL per well. After inoculating with 5 × 10⁵ bacteria/mL from late exponential cultures grown in the same media, cultures were incubated for 16–18 h at 37 °C. The lowest compound concentration inhibiting visible bacterial growth was recorded as MIC.