Resistance to conventional antibiotics used in medicine is rising dangerously high worldwide, and could lead to more than 10 billion deaths per year by 2050 according to the World Health Organisation (WHO). New types of antibacterial molecules have to be developed and should be i) nontoxic to humans, ii) active against bacteria already resistant to conventional antibiotics, and iii) unable or less likely to cause resistance compared to conventional antibiotics. Natural plant flavonoids are known to be effective molecules, but their activity and selectivity have to be increased. Among them, aurones have attracted attention due to their good antimicrobial activity associated with limited toxicity. Various aurone derivatives were chemically synthetized and tested for their biological activity on microorganisms (bacteria and fungi) and for their toxicity on human cells. Whereas some derivatives were found to be inactive or active only on some strains, others were found to have a larger spectrum of activity with Minimum Inhibitory Concentration (MIC) values as low as 0.78 μM. Limited toxicity on human cells was found, giving high therapeutic indexes (> 100 and up to 600) for the best candidate. The data show that 5-acetamidoaurones are far more active and safer compared with 5-aminoaurones, and that benzyloxy and isopropyl substitutions at the B ring are the most promising strategy in the exploration of new antimicrobial aurones.