Studies Oncontrolof Thiamphenicol Residues In Fishery

Thiamphenicol (TAP) is a broad-spectrum antimicrobial agent, structurally similar to chloramphenicol, used widely in aquiculture. It is bacteriostatic for both Gram-positive and Gram-negative aerobes and for some anaerobes. Thus it is important to develop detection method for TAP analysis in fishery now. We have developed a rapid ELISA detection method according to ELISA theory; in addition, apparatus detection methods have been developed.The conjugation of TAP and bovine serum albumin (BSA) (TAP-BSA, the immunoantigen of TAP) was produced according to the method of isobutyl chloroformate. The conjugation was identified by ultraviolet scan, SDS-PAGE and animal immunization test. The results demonstrated that the conjugation was synthesized successfully.According to immune protocol, three New Zealand white rabbits were immuned by TAP-BSA. The serum antiserum titre is as highly as 1×106. After purified by saturated ammonium sulphate, ion exchange and immunoaffinity chromatography in turn, monospecific antibody was obtained and its average affinity constant is about 7.8×10-9 mol/L. Based on this purified antibody, an indirect competitive ELISA was developed. After optimized some parameters such as the pH and ionic strength of buffers, concentration of Tween-20 and so on, the minimum detection limit of this ELISA arrived 0.06 ng/mL, and the recovery body of fluid (urine and serum) and tissue is about 80 to 110 percent. Besides, the ELISA kit was evaluated through following items: reproducibility, sensitivity, precision, and accuracy. Moreover, it was compared with the gas chromatography. All the results convinced us that this ELISA KIT can be applied to actual detection of thiamphenicol.The methods for the analysis of TAP were developed and optimized by HPLC,HPLC/MS and GC/MS. The recovery of HPLC,HPLC/MS and GC/MS were78.6～95.4%, 72.6～99.8% and 85.6～101.5%respectively;and the limits of quantitation were 8.3μg/kg, 0.1μg/kg and 0.1μg/kg respectively .The RSD of the methods were less than 10%. The results demonstrated that these methods were satisfied the requirement of practice detection.We have studied the depletion rule of TAP in Lateolabras janopicus's serum and edible tissues by HPLC method. After both 5 mg/kg and 30 mg/kg oral single-dose administration , the peak serum were obtained 6h and 8h,mean residence time were 11.1 h and 11.9 h;the plasma concentration-time course of thiamphenicol can be describe by a one-compartment open model : C=10.812(e^-0.087t– e^-0.274t); the elimination half-time and absorption half-time were 5.589 h and 1.997 h. After both 5 mg/kg and 30 mg/kg for consecutive 5 days oral administration , the absorbed fraction and elimination rate of TAP remain constant ,this result...