A stable phospholipid bilayer membrane was constructed on Au electrode surface by using a headgroup modified phospholipid and applied as a biomembrane model to investigate the interaction of an antibiotic polymyxin B with phosphatidylserine.Polymyxin B interacted with phosphatidylserine and destroyed the bilayer structure，resulting in increasing membrane permeation.The extent of interaction was dependent on polymyxin B concentration，reaction time and presence of cholesterol.The destroyed bilayer membrane could undergo self-recovery to some extent in KCl solution and the self-recovery was dependent on recovery time and concentration of polymyxin B.In addition，the interactions between PMB and different phospholipids were compared.Headgroup，alkyl chain length and saturation of phospholipid all affected the interaction.