question: Flavonoids are polyphenolic compounds that are ubiquitous in nature. They possess varied promising properties for medical use. Quercetin (3,3',4',5,7-pentahydroxyflavone) and apigenin (4',5,7-trihydroxyflavone) are two representative flavonoids, both of which have been reported to possess antibacterial activity by acting on multiple targets. Here, we determined that d-alanine:d-alanine ligase (Ddl) is another new target for quercetin and apigenin. Kinetic analysis indicated that these two flavonoids function as reversible inhibitors that are competitive with the substrate ATP of Ddl, whereas they are non-competitive with the other substrate d-Ala. The fact that quercetin showed lower 50% inhibitory concentration (IC(50)) and inhibitor binding constant (K(i)) values than apigenin against both the Helicobacter pylori Ddl and the Escherichia coli DdlB implies that the two additional hydroxyls on the flavone skeleton of quercetin in structure might facilitate its inhibitory activity and binding affinity to Ddl. This work is expected to help shed more light on the potential antibacterial mechanism of flavonoids.