Metabolism Of Fluoroquinolones Antibiotics In Liver Microsomes Of Crucian Carp(Carassius Ayratus)

Fluoroquinolone antibiotics remaining in the environment can enter the organism through ingestion,breathing,skin contact,etc.,and metabolic transformation occurs in the organism,which may produce more toxic metabolites and affect their ecological toxicity.At present,most of the research on the biological metabolism of FQs is about its metabolic transformation in vivo,and there are few studies on its in vitro metabolism.In the in vivo experiment,the external environment,body health and other factors interfered greatly.In vitro experiments can not only reduce the complex interference factors in in vivo experiments,but also directly observe the metabolic conversion process of enzymes to pollutants,and it is easier to find active intermediates in the metabolic process.Therefore,in this study,enrofloxacin and enoxacin were selected as the target compounds,we investigated their metabolic transformation process in vitro.The main research contents and results are listed below:?1?Under the conditions of in vitro incubation,the metabolic kinetics of ENR and ENO were explored.The results showed that when the exposure concentrations of ENR and ENO were 1 mg/L,5 mg/L and 10 mg/L,the in vitro metabolism of ENR and ENO both met the first-order kinetic equation.When 1 mg/L was selected as the exposure concentration of ENR and ENO,the maximum values of the elimination rate constants k of ENR and ENO were 0.00295min^-1 and 0.00404 min^-1,respectively,and the shortest half-lives t_1/2 were 235.0 min and 171.6min,respectively.When the exposure concentrations of ENR and ENO were the same,the value of ENO elimination rate constant was higher than that of ENR,indicating that under the conditions of in vitro metabolism,ENO was more easily metabolic transformed than ENR.?2?UPLC-MS/MS was used to detect and analyze the metabolic transformation products of ENR and ENO in vitro.There were three routes for in vitro metabolic transformation of ENR,which were hydroxylation,deethylation and piperazine epoxidation.Defluorination and hydroxylated ethyl methylation were the three ENO metabolic transformation pathways.EPR technology was used to detect the reactive oxygen species during the in vitro metabolism of ENR and ENO.The results showed that ENR and ENO were accompanied by the production of·OH,O₂·^-and ¹O₂ during the in vitro metabolic transformation.In particular,ROO·was generated during the in vitro metabolism of ENR,indicating that ENR may be more ecologically toxic than ENO.?3?During in vitro incubation,ENR and ENO have certain inhibitory effects on ECOD,AH and ERND enzyme activities,where at a significant level?=0.05,ECOD and ERND enzyme activities have a significant correlation with ENR concentration,Pearson correlation coefficient were-0.617 and-0.540,respectively.ECOD and ERND enzyme activities had no significant correlation with ENO concentration;AH enzyme activity had no significant correlation with ENR and ENO concentration.The results of in vitro metabolic inhibition experiments shown that CYP1A and CYP3A were the key enzymes for the in vitro metabolic conversion of ENR,and CYP2E1 was the key enzyme for the in vitro metabolic conversion of ENO.