| Florfenicol is a new generation of chloramphenicol veterinary synthetic antibiotics, its related researchs are still relatively weak. Due to lack of information on pharmacokinetic data and theoretical guidance, most of people often blindly use the drug, which not only affects the treatment, but also results in drug residues, affects the quality of aquatic products, it is not beneficial for aquaculture stable and healthy development. Industry of Eriocheir sinensis in our country has great prospects, to ensure the safety of food from animal origin, the detection methods of florfenicol and pharmacokinetics of florfenicol in Chinese mitten-handed crab(Eriocheir sinensis) should be studied, including:(1) Study on the determination of residues of florfenicol in aquatic products;(2) Study on the pharmacokinetics of florfenicol with different doses in the Chinesemitten-handed crab, Eriocheir sinensis;(3) Study on the pharmacokinetics of florfenicol with different temperature in the Chinesemitten-handed crab, Eriocheir sinensis.1. Two methods were developed for determination of florfenicol (FF) in fishery products. High-performance liquid chromatography with fluorescence detection (HPLC-FLD) used ethyl acetate as extractions, clean-up column was C18solid-phase extraction (SPE) cartridge, analysis column was reversed-phase column of Agilent TC-C18(250mm×4.6mm,5um), mobile phase were ammonium acetate (10mmol·L-1, pH3.5) and acetonitrile (55:45, by volume), the fluorescence detection with an excitation wavelength of225nm, the emission wavelength of280nm. In results, the linear ranges were0.1～5Sμg·mL-1of florfenicol, the correlation coefficient was more than0.999, at spiked level40-1000μg·kg-1for florfenicol, recoveries were88.6%～95.4%, with relative standard deviations from3.51%～8.28%. The detection limits were5.6μg·kg-1. High-performance liquid chromatography-tandem mass spectrometry used ethyl acetate as extraction, mobile phase methanolwere and water, identification was achieved by electrosp ray ionization (ESI) in negative mode using select reaction monitoring (SRM) by HPLC-MS\MS. In results, the linear ranges were1～100ng·mL-1of florfenicol, the correlation coefficient was more than0.999; the recoveries of florfenicol were86.6%～90.5%, with relative standard deviations from5.21%～7.28%. Compared the advantages and the disadvantages of the two methods and provided technique referrence for detecting of florfenicol residues in aquatic products.2. A pharmacokinetic study of florfenicol (FF) following by different doses of florphenicol (5.0,10,20mg·kg-1) with intramascular injection was carried out in the Chinese mitten-handed crab at17℃. Hemolymph, muscular and liver samples were obtained and determined by a high-performance liquid chromatography with fluorescence detection method, the concentration-time data derived from the experiment is processed with origin8.0and the corresponding pharmacokinetic parameters were calculated with3P97. The florfenicol level in hemolymph and muscular reached maximum momentarily, while those in liver appeared to be a maximum then declined at three administration doses. The kinetic profile of florfenicol in hemolymph, muscular and liver was conformed to a two-compartment model. The important parameters were listed. The absorption half-life (T1/2a) and the elimination half-lives (T1/2β) in hemolymph were0.04,0.21,0.87and0.60,1.48,3.29h, respectively, the total body clearance (CLs) was2.17,1.99and2.12mL·kg-1·h-1, the absorption half-life (T1/2α) and the elimination half-lives (T1/2β) in muscular were1.38,0.93,0.47h and10.39,31.78,23.91h, respectively, the total body clearance (CLs) was0.07,0.03and0.03mL·kg-1·h-1, the absorption half-life (T1/2α) and the elimination half-lives (T1/2β) in liver were0.69,0.07,0.12h and0.73,4.90,4.49h, respectively, the total body clearance (CLs) was1.79,2.55, and3.89mL·kg-1·h-1.3. The pharmacokinetics of florfenicol (FF) was investigated with a single dose of Smg·kg-1body weight after intramascular injection in Chinese mitten-handed crab (Eriocheir sinensis) at the water temperature of17℃and28℃. Hemolymph, muscular and liver samples were obtained and determined by a high-performance liquid chromatography with fluorescence detection method, the concentration-time data derived from the experiment is processed with origin8.0and the corresponding pharmacokinetic parameters were calculated with3P97. The kinetic profile of florfenicol in hemolymph, muscular and liver was conformed to a two-compartment model, the elimination half-lives (T1/2p) and the area under the concentration-time curve (AUC) in hemolymph were0.60h,2.30mg·mL-1·h, respectively, at17℃and0.41h,1.92mg·mL-1·h, respectively, at28℃; T1/2p and AUC in muscular were10.99h,69.36mg·mL-1·h, respectively, at17℃and73.89h, mL-1·h, respectively, at28℃; T1/2p and AUC in liver were0.73h,2.79mg·mL-1·h, respectively, at17℃and20.57h,11.mg·mL-1·h, respectively, at28℃. T1/2β in hemolymph were longer at the lower water temperature than those at the higher water temperature, however, T1/2β in muscular and liver were shorter at the lower water temperature than those at the higher water temperature; AUC in hemolymph was higher at the lower water temperature than those at the higher water temperature, however, AUC in muscular and liver was lower at the lower water temperature than those at the higher water temperature. The results showed that the pharmacokinetics of florfenicol in the various organs of Eriocheir sinensis is not same at different temperature, it illustrates that metabolic mechanism of Eriocheir sinensis is very complex. |
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