Antibacterial assay
Staphylococcus aureus strain is purchased from ATCC (25923).S. aureus was cultured at +37°C in a Mueller Hinton Agar (BD) for 24 hours. Then on colony was transferred to 10 mL of Mueller Hinton Broth (BD) for 24 hours at +37°C with agitation for preculture step. The absorbance is read at 620 nm and we adjusted DO value to 0.001 to perform our assay, corresponding to a density of 8.105CFU.mL-1. As cytotoxicity assay, we tested PDA-PAR30 NPs in solution and also Gel- PDA-PAR30 composite hydrogel. For solution part, we kept the same ratio (samples:bacteria) in medium as 1 to 9, final volume is 100 µL (10µL of NPs for 90 µL of bacteria in medium). Samples with bacteria were incubated for 24 hours at +37°C with agitation. Final absorbance were read at 620 nm in order to quantify bacteria growth or inhibition. For gelatin hydrogels part, samples were directly produced in 48 wells-plate and sterilized by UV light exposure. After preculture of S. aureus, we added 300 µL of suspension with OD=0.001 then we incubated for 24 hours at +37°C with agitation. OD was read at 620 nm. We compared PDA NPs and PDA-PAR30 NPs. In solution, OD of samples (PDA and PDA-PAR30) with bacteria were subtracted to OD the same samples without bacteria to remove PDA absorption (brown solution). All results were compared to a normal growth condition named “Bacteria”, defined as 100% growth. We also tested Tris buffer (in solution or hydrogel) as a negative control. We used antibiotics (Cefotaxime 0.1 µg.mL-1 + Tetracycline 10 µg.mL-1) as positive control. We also used PAR30 (10 µg.mL-1) without PDA as positive control. All samples were performed in triplicates.