Gold Immunochromatography Technology For The Rapid Detection Of The Enrofloxacin In Seafood

The veterinary drug residues in animal edible tissues present serious hazards topublic health, which, in turn, becomes the bottleneck in expanding export of theseafood in China. As Fluoroquinolones (FQs), a class of broad-spectrum antibiotics,are widely used in aquaculture. The FQs residues have aroused wide public concern.Recently, the colloidal gold-based immunochromatography is widely used indetection of the small molecules, especially the rapid detection of drug residues inanimal food with the introduction of colloidal gold. Currently, the commercializationof colloidal gold immunochromatography assay test strip（GICA）has been reported.But their applicability in rapid detection, such as the simplification for samplepretreatment to avoid the process of centrifuging and purification, has not been testedor reported.In this paper, an optimized sample pretreatment method with a high sensitivitywas constructed which could be simply performed only with some small equipent andtested for determination of FQs in turbot samples. And the effect of protein substrateswas eliminated by trichloroacetic acid (TCA). This established method wasappropriate for spot fast screening of food contaminants, solving the commonproblem that there have no effective pretreatment in the rapid detection by GICA teststrip.The protein was considered as the most important matrix effect by comparing theeffect of various matrix on the GICA, which was eliminated by trichloroacetic acid(TCA). The optimum concentration of TCA eliminating protein in the sample wasconfirmed to9%which could remain the high sensitivity of GICA. The application ofTCA could eliminate protein in samples significantly. Meanwhile, it could beconducted easily in a shorter time. Several experiments was carried out to determine the optimal pretreatmentcondition index. The sample pretreatment method and the LOD of detecting theEnrofloxacin (ENR) residues was preliminary confirmed as follows: samples weregrinded with mortar at first, then the extract liquid was filtering with pinhole filtermembrane and the pH of the liquid was adjusted by pH test strips in the end. Thewhole process could be completed within30min. The optimal extracting buffer forveterinary drug residues was the mixture of Methanol/PBS (1:1) and trichloroaceticacid (TCA) at the concentration to9%. The rate of recovery of this preliminarymethod was about40%-50%obtained by the application of HPLC. Limit of Detection(LOD) for Enrofloxacin (ENR) and Ciprofloxacin were10μg/kg and100μg/kg invarious samples, including turbot, Larimichthys crocea and Litopenaeus vannameisamples. This method proved to be a successful pretreatment method as the Limit ofDetection (LOD) for Enrofloxacin (ENR) meets the standards for veterinary drugresidues in animal edible tissues set by the government.In the preliminary application of the assay in real samples, the sensitivity ofGICA for Enrofloxacin (ENR) standard and ciprofloxacin (CIP) standard was1μg/kg~5μg/kg and20μg/kg~50μg/kg, respectively. Then3kinds ofFluoroquinolones and another2kinds of antibiotics (concentration all100μg/kg) wereused to confirm the specificity of GICA, showing a favourable specificity. The testresults of real samples from the two detection method, the National standard methodand the method established in this paper, were identical with each other which bothwere the negative results. It could be concluded that the optimized samplepretreatment method established in this paper could be applied in the real samples.