Study On The Elimination Rule Of Enrofloxacin And Its Metabolites Residue In Largemouth Bass And Grass Carp

Enrofloxacin is a broad-spectrum and highly efficient antibiotic commonly used for treating diseases in aquatic animals.However,its non-standard use often results in a significant amount of residual drug in animal tissues,representing a severe risk to human health.Organ blood flow is a critical physiological parameter directly impacting the absorption,metabolism,distribution,and excretion of drugs in fish,thereby potentially interfering with the correct determination of withdrawal time（WT）for effective treatment.The purpose of this study was to explore the impact of organ blood flow on the elimination pattern of enrofloxacin（ENR）and its metabolite ciprofloxacin（CIP）in largemouth bass（Micropterus salmoides）and grass carp（Ctenopharyngodon idella）under actual aquaculture conditions.Findings may contribute to the development of adequate drug withdrawal periods and provide fundamental information for developing a physiologic pharmacokinetic model for cross-species extrapolation.The main conclusions of the study are summarized below:1.Organ blood flow in largemouth bass and grass carp was measured using a fluorescent microsphere method.Yellow-green microspheres were uniformly injected into both fish species through cardiac catheterization and a suction pump at a rate of 0.8m L/min.Reference blood samples were taken from the dorsal aorta,and after 5 minutes,fish tissues were collected with a surgical knife,pre-treated and measured using a fluorescence spectrophotometer.Results showed that under 25°C,compared to largemouth bass,grass carp had greater organ blood flow.In largemouth bass,the blood flow in the heart,swim bladder,kidney,liver,gills,skin,muscle,spleen,intestine,and other organs were 8.80,1.22,3.01,0.71,2.72,0.40,9.47,2.33,0.54,and 4.11（m L/min/g）,respectively.In grass carp,the blood flow in the heart,swim bladder,kidney,liver,gills,skin,muscle,spleen,intestine,and other organs were 9.55,2.06,10.3,6.92,6.70,3.72,5.78,1.00,2.81,and 6.04（m L/min/g）,respectively.These results reflect the biological characteristics and metabolic needs of different fish species,and are significant in understanding the physiological metabolism and regulatory mechanisms of fish.Moreover,this study also revealed that factors such as fish body size and metabolic habits affect organ blood flow,providing a reference for further research on metabolism and elimination of ENR and CIP in largemouth bass and grass carp.2.After establishing an efficient high-performance liquid chromatography（HPLC）detection method for ENR and CIP in plasma and tissue of largemouth bass and grass carp,the elimination pattern of ENR in the two fish species were studied and a preliminary risk assessment was conducted.ENR powder was added to ordinary feed to make a concentration of 20 mg/kg,and was fed continuously for 5 days at a rate of0.5%of the body weight of largemouth bass and grass carp,twice a day at 7:00 am and6:00 pm.Skin muscle,plasma,gills,kidney and liver tissues were collected at different times,pre-treated,and the concentrations of ENR and CIP were measured by HPLC-fluorescence.The WT of ENR in plasma and tissues were calculated using the WT1.4software.The T_1/2of ENR in skin muscle,plasma,gills,kidney and liver of largemouth bass were 7.13 d,4.81 d,6.31 d,7.52 d and 3.66 d,respectively,while those of CIP were 6.26 d,1.36 d,1.86 d,2.06 d and 7.27 d,respectively.The T_1/2of ENR in skin muscle,plasma,gills,kidney and liver of grass carp were 8.09 d,6.33 d,6.95 d,7.39 d and 5.35 d,respectively,while those of CIP were 8.96 d,11.89 d,12.04 d,7.27 d and7.49 d,respectively.This indicates that the T_1/2of CIP is longer than that of ENR,and that the liver is the main organ for metabolism and elimination due to its increased blood flow during the metabolism process.The homogeneity and linearity of regression line data were evaluated by Bartlett,Cochran,and F-tests on residual quantity data,and outliers were eliminated by establishing a standardized residual distribution with cumulative frequency distribution on a normal probability plot.ENR given at 20 mg/kg bw for five consecutive days resulted in a WT of 46 d,46 d,33 d,29 d and 20 d in the skin muscle,kidney,gills,plasma and liver of largemouth bass,respectively,according to Chinese and European regulations.For grass carp,the WT of ENR in skin muscle,plasma,gills,kidney,and liver were 51 d,49 d,44 d,43 d,and 35 d,respectively.The withdrawal time of largemouth bass and grass carp were after 46 and 51 days,respectively,and the HQ in plasma and all tissues was less than 1.The estimated daily intake（EDI）of ENR in largemouth bass skin muscle,plasma,gills,kidneys,and liver decreased to less than 10%of the acceptable daily intake（ADI）after 7,9,7,7,and 5days,respectively.The EDI of ENR in grass carp skin muscle,plasma,gills,kidneys,and liver decreased to less than 10%of the ADI after 28,28,14,28,and 28 days,respectively.These results indicate that the established withdrawal time can avoid human health risks caused by ENR and its metabolite residues in largemouth bass and grass carp.