Abstract: The ultrasound-assisted extraction of flavonoids from leaves of Helianthus tuberosus L. was optimized. The extract obtained was evaluated for its antimicrobial activity.Based on the results from single-factor tests, the extraction conditions were further optimized using the response surface methodology. It was found that when 40% ethanol was applied as the solvent in a substrate:solvent ratio of 1:40 to extract the leaves for 20 min at 64℃, the yield and efficiency of the process could be maximized to reach a flavonoid extraction rate of 1.47 mg·g^-1. The flavonoids obtained showed an excellent antibacterial activity on Escherichiacoli, but not on Staphylococcus aureus or Bacillus subtilis.