The percentage of metabolically active bacteria was determined by adding 20 µL of a 0.15 mg/mL resazurin (Sigma-Aldrich, St. Louis, USA) solution to 100 µL of the resuspended cells. This mixture was incubated at 37 °C for 2 h in a 96-well black plate in static conditions and the fluorescence was measured at the excitation/emission wavelengths of 550/590 nm, respectively, in an Infinite 200 M PRO microplate reader (Tecan, Hombrechtikon, Switzerland). The percentage of metabolically active cells was calculated using the Eq. (8):
where SC550/590 and GC550/590 correspond to the fluorescence measurements of the control without inoculation and the biofilm control without enzyme treatment, respectively. The experiments were performed in triplicate.
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