The Study Of Pharmacodynamics In Vitro And In Vivo And Pharmacokinetics Of Florfenicol In Acipenser Baeri

Florfenicol is a new antibiotic of chlormycetin, which is used widely by fish farming. Florfenicol amine is the main active metabolite of florfenicol. Florfenicol and florfenicol amine are common detection objects in EU and China in aquatic products. The thesis will be provided theoretical basis for fish farming by pharmacodynamic and pharmacokinetic of florfenicol against Acipenser baeri.The minimal inhibitory concentration(MIC),minimal bactericidal concentration(MBC), growth curve, kinetic curve and influence of environmental factors(bacterial inoculum, concentration of magnesium ion, calf serum and pH of medium) were measured by2-fold serial dilution method. The results showed that MIC, MBC and MBC/MIC of florfenicol against Yersinia ruckeri were0.5μg/mL,1.0μg/mL and2. It showes that the florfenicol against Yersinia ruckeri has good bacteriostatic and bactericidal effect. According to the logarithmic growth phase, Yersinia ruckeri was inoculated in the peptone medium afterlst hour.It lasted approximately7h,8h,and the concentration of the bacterial was exceed108CFU/mL. The growth velocity was getting to slow after the8th hour and stabilize after12th hour.The killing rate of8MIC florfenicol againgst Yersinia ruckeri was better than4MIC,2MIC and1MIC separately. The killing rate of florfenicol was8MIC>4MIC>2MIC>1MIC. The PAE of florfenicol is concentration-dependent manner.When the concentration of florfenicol is2MIC,4MIC and8MIC,PAE was3.71±0.11h、4.54±0.27h、5.52±0.23h separately. The effect of florfenicol against Yersinia ruckeri in vitro is obvious by pH value, bacterial inoculum volume and blood volume of culture conditions effect.Patho-exemplar of Acipenser baeri infected Yersinia ruckeri was made in the experiment. The experiment was done when the Acipenser baeri was infected24h ago. The infected Acipenser baeri were divided three dose group(high concentration of50mg/Kg, Middle concentration of20mg/Kg, low concentration of5mg/Kg). Each group were treated once a day and3days consecutive. The three dose group degrade death rate of Acipenser baeri.The effective rateof high concentration of50mg/Kg and middle concentration of20mg/Kg can achieve100%. Yersinia ruckeri is highly sensitive to florfenicol, which is consistent with the test result in vitro. We established the methods of florfenicol and florfenicol amine in plasma, muscle, liver and kidney of Acipenser baeri by high performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Samples was prepared by alkality acetidin, then defated by N-hexane. Samples was separated by chromatographic column of ACQUIT UPLC BEH-C18.and gradient dilution by acetonitrile and water. The results indicated, there were linear regression correlation when florfenicol and florfenicol amine in10～1000ng/mL concentration, correlation coefficient is0.999; the minimum detection limits were0.1ng/mL and0.5ng/mL; the average recovery rate were85%～110%and80%～98%; Within day and days average variation coefficient are less than15%, providing a reliable analysis method for florfenicol and florfenicol amine.The pharmacokinetics and tissue distribution of florfenicol, related metabolites (florfenicol amine) in Acipenser baeri were studied in vivo using high performance liquid chromatography with tandem mass spectrometric (HPLC-MS/MS). In this study, oral administration of florfenicol was performed in the Acipenser baeri at a dosage of15mg/kg body weight at22℃, and the blood plasma, liver, kidney and muscle were then prepared to measure the concentration of florfenicol and its metabolite (florfenicol amine) using HPLC-MS/MS method. The results indicated that the concentration time course of florfenicol and florfenicol amine can be described by a two-compartment open model with first order absorption in vivo. The maximal concentration (Cmax)of florfenicol in blood plasma was3.4μg/mL, Time-point of maximal concentration(Tpeak) was2.943h, volume of distribution (V/F) was3.267L/kg, half-life of elimination (t1/2β) was31.21h, Cmax (FFA)/Cmax(FF) and AUCFFA/AUCFF were5.44%,20.73%;Florfenicol and florfenicol amine extensively distributed and had a similar regularity in several tissues, which had a higher concentration in liver and kidney at first. The results indicate that florfenicol have following features in Acipenser baeri:rapid absorption, high peak concentration, slow elimination and wide range of tissue distribution, and mainly metabolized or eliminated in the form of initial drugs.According to the results of pharmacological and pharmacokinetic, We suggest that the suitable doseage of florfenicol against Yersinia ruckeri was20mg/kg,and dosing interval time was24h.