Determining Antimicrobial Activity of PA Emulsions

Minimal inhibitory concentration of the emulsions was determined by the modified 96 well plate Resazurin assay^{23 (link)}. Briefly, stock solutions of the PA emulsions were serially diluted into Iso-sensitest broth (Oxoid Ltd., Hants, UK) containing Resazurin (Biotium, Hayward, CA) (0.001 g/ml) and inoculated with ca. 5 Log CFU/ml of Salmonella. Serial dilutions were performed to obtain PA concentrations ranging from 500 mM to 2.73 mM of each emulsion. The 96 well plates were incubated for 24 h at 37 °C and the wells containing the serial dilutions of the emulsions were observed for a change in color from blue to pink indicating bacterial growth. The lowest dilution at which no bacterial growth was determined (blue well) was considered the minimum inhibitory concentration. The MICs of the individual surfactants at concentrations of 1%, 0.1% and 0.01% (w/v) and PA without surfactant that was suspended in water by vortexing for 1 min were determined as comparative controls using the modified 96 well plate Resazurin assay. Each test was performed three times and an average of the three MIC values were used.