Measurement of minimum inhibitory concentration (MIC) of chalcone derivatives

MIC was measured in a 96-well plate through the microdilution broth method. H. pylori strain ATCC 43504 was incubated for 2-3 days at 37°C and 10% CO₂ on a Blood Agar plate. After liquid culture in Brucella broth containing 10% horse serum, it was seeded at 5×10⁵ CFU/well. The chalcone derivatives were treated by 2-fold dilution from 200 μM to 12.5 μM. Amoxicillin (Sigma-Aldrich) was used as a reference compound at 2 μg/mL. Then, it was incubated for 72 h under the conditions of 10% CO₂ and 37°C, and optical density (OD) was measured at 550 nm. After subtracting the OD value of blank (only Brucella broth) from each well, the MIC was determined using the following equation: (OD₅₅₀ of wells that contained the compound/OD₅₅₀ of the compound-free well)×100.