| Chloramphenicol(CAP) is an effective antibiotic that has widely been used to treat food-producing animals. In 1990s, the use of CAP was banned by the European Union (EU), USA, China and so on due to the potential health risk posed by its traces in food. The IAC-GC-μECD method of Chloramphenicol residue determination is established successfully in chicken muscles and livers in this Study.In this study, the immunoaffinity column is prepared successfully and appraised for its property. The mouse ascites against CAP is purified by saturated ammonium sulphate precipitation and dialysed. The concentration of monoclonal antibodies measured by spectrophotometer is 5.1mg/ml.The coupling ratio between monoclonal antibodies and Sepharose 4B is 85.21%.The active groups on the Sepharose 4B base materials are blocked and washed,stored.The dynamic column capacity determined by adding an amount of 20ml of CAP solution in PBS(500ng/ml) to the immunoaffinity column is 3265ng/ml and the specific column capacity is 766.5ng/mg.The recovery ratio of CAP in immunoaffinity column is 99.8%.The changed curve shows that the dynamic column capacity is decreased slightly after the column is used for 5 times and becomes 58%(1890ng/ml)of originality after 16 times,but it is stil meet the need of determianation.A method was developed for Clean-up of Chloramphenicol sample by immunoaffinity chromatography and determination by gas chromatography in broiler chicken muscles and livers.The tissues was extracted with ethyl acetate. The sample was defatted with hexane and Clean-up by immunoaffinity chromatography column, washed by Methanol, evaporated to dryness with nitrogen gas. Residue must be dry before adding Sylon N,O-Bis(trimethylsily) trifluoroacetamide(BFT). Added Sylon BFT. After derivatization, tolune and water were added immediately to the sample. The organic layer was carefully removed. 3μ1 analytes were determined by GC with micro electron capture detector (μECD).The recoveries of analytes at the fortified level of 0.05μg/kg, 0.1μg/kg, 0.5μg/kg, 1μg/kg, 5μg/kg, 10μg/kg range from 86.6%~96.9% in the muscle, and at the fortified level of 0.1μg/kg,0.5μg/kg, 1μg/kg, 5μg/kg, 10μg/kg, 20μg/kg range from 74.3%~96.1% in the liver, respectively. Coefficents of variation for distribution,CVd, ranged from 2.2%~5.9% in muscle, from 2.8%~3.9% in liver,respectively.And for precision CVr, ranged from 3.8%—9.2% in muscle, from 4.9% ~ 7.2% in liver, respectively. The limit of detection(LOD) is 0.021μg/kg in muscle, 0.05μg/kg in liver.At the same time, the Comparison between the clean-up of IAC and purification of liquid-liquid distribution shows that the limited of determination(LOD) by IAC is less than that of liquid-liquid distribution and the number of impurity peak by IAC is less than that of the purification of liquid-liquid distribution. So, the clean-up of IAC shows its stronger specificity and selectivity than the method of liquid-liquid distribution in this study... |
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