Comparative Pharmacokinetics Of Florfenicol In Healthy And Vibrio Alginolyticus Infected Portunus Trituberculatus

With the development of the intensive aquaculture system model , the incidence and severity of Vibrio alginolyticus disease of Portunus trituberculatus are on the rise. This is a restrict factor for sustainable development,so some high-effective and low-poisonous and less-residual antimicrobial substance must be investigated for Portunus trituberculatus. Florfenicol, also called the fluorine Thiamphenicol, is a kind of chloramphenicols exclusively used in veterinary medicine, which has wide antibacterial spectrum and low toxicity and was completely absorbed and distributed. It has been widely intended for the treatment of infectious diseases in some aquatic animals.In this study, the pharmacokinetics and tissue distribution of florfenicol in the Vibrio alginolyticus diseased Portunus trituberculatus were investigated firstly, so as to appraise its feasibility and security,to provide the scientific basis for the standardization application of fishery drugs in aquaculture. The main results are:1. Pharmacokinetics of florfenicol in healthy Portunus trituberculatusThe concentrations of florfenicol were determined in haemolymph, muscle, hepatopancreas of Portunus trituberculatus following oral administration in a single dose of 25 mg/kg at (21±2)℃by using high performance liquid chromatography (HPLC).The results showed that the retention rime for florfenicol was 7.815min with no interference from endogenous components. The calibration curve exhibited excellent linearity over a rang of 0.05–10μg/mL(r=0.9995). The limit of detection(LOD) was 0.025μg/mL .The average recovery was 89.02±6.24% and the RSDs of intra-day and inter-day were all less than 5%. So the validated HPLC method was suitable for the measurement of florfenicol concentration. Florfenicol can unevenly distributed to all tissues, absorbed most quickly by hepatopancreas. Their concentration of florfenicol were all lower than 0.1μg/mL after oral administration for 7 days. The concentration-time data of haemolymph, muscle, hepatopancreas were all described by two-compartment open model with first-order absorb, described with C= 15.230e^{―0.253t}+0.696e^-0.015t-15.926e^-0.697t,C= 6.753e^{―0.670t}+1.445e^-0.014t-8.198e^-2.419t , C= 9.222e^{―3.068t} + 1.484e ^{- 0.017t} - 10.706e ^-15.886t, respectively. The main pharmacokinetic parameters were as follows: Haemolymph: T_peak=2.385h,T_1/2β=46.609h,C_max =5.980μg/mL,CLs=0.297L/kg·h,Vd=2.408L/kg,AUC=84.153μg/mL·h;Muscle:T_peak=0.816h,T_1/2β=50.236h,C_max=4.199μg/mL,CLs=0.224L/kg·h,Vd=3.957L/kg,AUC=111.396μg/mL·h;Hepatopancreas:T_peak=0.143h,T_1/2β=39.903h,C_max=6.324μg/mL,CLs=0.285L/kg·h,Vd=2.802L/kg, AUC=87.765μg/mL·h.So florfenicol could be absorbed quickly , distributed widely and eliminated slowly,which had an efficient effect of antibacterial in Portunus trituberculatus.2. Establishment of a Vibrio alginolyticus disease model for Portunus trituberculatusPortunus trituberculatus was infected artificially with Vibrio alginolyticus with a concentration of 107CFU/mL. The external symptoms and histopathological changes of infected crab were observed and the physio-biochemical indexes were determined at 0, 24, 48 and 72h post-injection. The results indicated that the pathological changes of the infected crab were obvious gradually at 24h post-injection and THCs,T-AOC,AKP,ACP and POD had significant changes. The main objection of drug prevention was early diseased population, so the indexes, such as THCs,T-AOC,AKP,ACP and POD, in the crab which was infected 24 hours can be used to determine whether the establishment of disease model was successful in this paper. The Vibrio alginolyticus disease model for Portunus trituberculatus which was established by this method was reproducible highly and convenient for observing changes of crabs.3. Pharmacokinetics of florfenicol in Vibrio alginolyticus diseased Portunus trituberculatusThe pharmacokinetic characteristics of haemolymph, muscle, hepatopancreas of Vibrio alginolyticus diseased Portunus trituberculatus in a single dose of 25mg/kg at (21±2)℃were all fitted with by two-compartment open model with first-order absorb, described with C = 18.071e―0.249t+1.711e-0.012t-19.782e-0.3933t,C= 19.012e―0.676t+1.440e-0.011t-20.452e-0.874t,C= 19.283e―3.286t + 1.427e -0.014t - 20.710e -4.363t, respectively. The main pharmacokinetic parameters were as follows: Haemolymph: Tpeak=3.700h,T1/2β=55.667h,Cmax=4.209μg/mL, CLs= 0.157L/kg·h, Vd=3.017L/kg, AUC=159.649μg/mL·h; Muscle: Tpeak=1.662h, T1/2β=63.349h,Cmax= 2.809μg/mL,CLs=0.248L/kg·h,Vd=4.366L/kg,AUC=136.329μg/mL·h;Hepatopancreas:Tpeak=0.331h,T1/2β=48.437h,Cmax=3.032μg/mL,CLs=0.183L/kg·h,Vd=4.043L/kg,AUC=100.843μg/mL·h.Campared with the healthy Portunus trituberculatus, the pharmacokinetic parameters of florfenicol in infected Portunus trituberculatus had great changes. The rate of absorption and elimination slowed, Tpeak and t1/2βprolonged, Cmax and CLs decreased,Vd and AUC increased. Supposed that the bioavailability was 100%,we suggested that the dose of clinical should be 50mg/kg for the first time and the maintenance dose should be 25mg/kg per 12hours. According to the maximum residue limit (MRL) of 0.1mg/kg in tissues,the withdrawal period should not be less than 10daysunder this experiment condition.