Study On The Metabolites Of Enrofloxacin In Carp

Carp is widely cultivated because of its high nutritional value,low cost of breeding,and easy digestion absorption by the human body.However,due to water pollution,excessive density of culture and other reasons in the process of carp breeding,it is easy to cause fish pollution and damage,resulting in death,and the use of antibiotics can effectively prevent and treat fish diseases.Enrofloxacin is the most common antibiotic,has strong antibacterial effect,and is not easy to cross-react with other antibiotics and is widely used in aquaculture.But the unreasonable use or abuse of antibiotics may cause veterinary drug residues.The residues of veterinary antibacterial drugs may cause certain potential harm to human health,which may cause cancer,teratogenicity and mutagenicity.The research and detection of enrofloxacin in fish and its metabolites can not only understand the residuals of the original drugs and metabolites,but also provide a basis for further study on the environmental and ecological effects of metabolites,and provide guidance for improving the aquaculture environment,timely diseases control,and reasonable judgment of the drug withdrawal period.To achieve complete treatment of aquatic diseases,and not to use antibiotics,to ensure environmental safety,food safety and to ensure the testing of export domestic sales process.In this experiment,carp was used as the research object.Firstly,the pretreatment was optimized.Secondly,the conditions of high performance liquid chromatography tandem mass spectrometry were optimized,and the metabolic products and metabolism of enrofloxacin in carp were studied by optimized method.The research results are as follows:(1)Determine the best pretreatment method:the sample is vortexed by anhydrous sodium sulfate,and the acidified acetonitrile is dissolved and ultrasonically extracted.After centrifugation of the solution,the supernatant was rotary evaporated to dryness.The residue was dissolved in 0.1%formic acid water and acetonitrile,and the mixture was vortexed and extracted,and then allowed to stand for separation.The upper layer was discarded,and the lower layer was passed through a 0.2?m filter.The method is simple and convenient to operate,uses less reagents,takes less time,and has an average recovery rate of 90.17%in muscle,an average recovery rate of 90.44%in the liver,and an average recovery rate of 89.22%in the kidney.(2)Optimize the elution procedure,ion source,capillary positive,collision energy and other conditions of LC-MS-MS to obtain the best liquid quality conditions:0.1%formic acid aqueous solution-acetonitrile as mobile phase,0.2 mL/min flow rate,10?L injection volume,gradient elution method,mass spectrometry using atmospheric piezoelectric spray ion source(ESI),positive ion detection mode.The results showed that enrofloxacin had a good linear relationship in the range of 0.0050~3.000?g/mL,and the correlation coefficient was R~2=0.9995.The minimum detection limit is 0.0050?g/mL.(3)The carp used in this experiment were administered intraperitoneally with enrofloxacin,and the enrofloxacin in the muscle,liver and kidney of enrofloxacin was treated by Triple Quad Liquid Chromatograph-Mass.Its metabolites were identified,and the metabolism of enrofloxacin and its metabolites in different tissues and at different times of carp was analyzed.According to the mass spectrometry,the excimer ion of enrofloxacin was[M+H]~+m/z 360.200,and four metabolites were identified,which were enrofloxacin deethylated metabolite ciprofloxacin m/z 332.200,enrofloxacin decarboxylation metabolite m/z 316.900,enrofloxacin deethylene metabolite m/z334.000 and enrofloxacin dehydrated metabolite m/z 342.200.(4)After enrofloxacin enters the carp,most of them are in the form of the original drug,and a small part is metabolized.The main metabolite is ciprofloxacin,followed by the decarboxylation metabolite of enrofloxacin.After administration,the concentration of enrofloxacin in the muscle,liver and kidney of the carp increased rapidly.The drug concentration began to show a slow downward trend after reaching the peak in the carp.At this time,the drug concentration of the liver and the kidney was higher than that of the muscle.