Black polystyrene 96 well microplate

The angiotensin-converting enzyme-I inhibition (iACE) assay was conducted following the procedure outlined by Coscueta et al. (2019) [52 (link)]. The assay was performed using a black polystyrene 96-well microplate (Nunc, Denmark), and fluorescence was monitored at 1 min intervals for 80 min using a Multidetection plate reader (Synergy H1, Vermont, USA) controlled by Gen5 Biotek software version 3.04. Each sample, blank, and control analysis was carried out in triplicate. The iACE was quantified as the concentration required to inhibit 50% of the enzymatic activity (IC₅₀). The IC₅₀ values were determined using non-linear modelling of the data obtained.

The oxygen radical absorbance capacity (ORAC) assay followed the protocol outlined by Coscueta et al. (2019) [52 (link)]. A black polystyrene 96-well microplate (Nunc, Roskilde, Denmark) was utilized, and measurements were taken using a Multidetection plate reader (Synergy H1, Vermont, USA) controlled by Gen5 Biotek software version 3.04. Fluorescence was monitored for 80 min at 1 min intervals. Each sample, standard, blank, or control analysis was duplicated. The final ORAC values were expressed as Trolox Equivalents (μmol TE).