The alteration in metabolism or transport resulted from concomitant medications and some foods can change the known safety and efficacy of a drug.The pharmacokinetic interactions between an investigational new drug and other drugs should be defined during drug development, as part of an adequate assessment of the drug's safety and effectiveness.The DDI study generally begins with in vitro studies to determine whether a drug is a substrate, inhibitor, or inducer of metabolizing enzymes.In vitro drug interaction data can be used in guiding clinical interaction studies.We developed an in vitro drug-drug interaction assay applied the validated method to assess the antofloxacin DDI risk.The study includes four parts: ① the incubation of antofloxacin in human live microsome (HLM) with the presence of NADPH or UDPGA were carried out and LC-MS/MS were used to examine the possible metaboliste, however, no significant metaboliste was detected after 30 or 60 min of incubation in these two reaction system.