Determination of Sudan I by high performance liquid chromatography with immuno-affinity column clean-up

摘要

To prepare an immuno-affinity column (IAC) for Sudan I, Sepharose 4FF was activated by epichlorohydrin, and the density of epoxy is 80 μmol/g. Then it was coupled with 6-aminocaproic acid. The Sudan I immunoaffinity column was prepared by carbodiimide method in which modified Sepharose 4FF was used as carrier to couple with monoclonal antibody against Sudan I. A reversed-phase high performance liquid chromatographic method was used to determine the samples elution after the IAC cleaning up. Sudan I was determined by HPLC with a retention time of 5.82 min (flow rate of 1 ml/min). The column capacity is 1582.8 ng Sudan I when using 0.3 g Sepharose 4FF. Average recoveries of Sudan I from red chili powder spiked at levels of 0.25-3 mg/kg range from 44.52% to 77.40%. The coefficient of variation is 4.6% to 8.3%. The IAC method was found to bo highly effective, sensitive and convenient for isolating the target analyte from sample.