Two synthetic compounds, MHY1383, azo-resveratrol and MHY1387, 5-[4-hydroxy-3,5-methoxybenzy]-2-thioxodihydropyrimidine-4,6[1H,5H]-dione have been reported to have an anti-biofilm effect onPseudomonas aeruginosaat very low concentrations (1–10 pM). Here, we investigated the anti-biofilm effects of these compounds in various bacteria. We found that MHY1383 significantly inhibitedEscherichia coli,Bacillus subtilis, andStaphylococcus aureusbiofilm formation at 1 pM, 1 nM, and 10 nM, respectively. MHY1387 also inhibited the biofilm formation ofE. coli,B. subtilis, andS. aureusat 1 pM, 10 nM, and 100 pM, respectively. Both MHY1383 and MHY1387 showed medium-dependent anti-biofilm effects onSalmonella entericaat high concentrations (10 μM). We also tested the susceptibility to antibiotics by measuring the minimum inhibitory concentration (MIC) in various bacteria. WhenP. aeruginosa,E. coli,B. subtilis,S. enterica, andS. aureuswere treated with MHY1383 or MHY1387 in combination with four different antibiotics, the MICs of carbenicillin againstB. subtilisandS. aureuswere lowered more than two-fold by the combination with MHY1387. However, in all other combinations, the MIC changed within two-fold. The results of this study suggest that MHY1383 and MHY1387 are effective anti-biofilm agents and can be used at very low concentrations against biofilms formed by various types of bacteria. We also suggest that even if a substance that inhibits biofilm is used together with antibiotics, it does not necessarily have the effect of lowering the MIC of the antibiotics.