| In recent years, the food safety issues occurs frequently.The veterinary drug residues especially the drug residues in seafoods attracts wide attention at home and abroad. As Nitrofurans, a class of synthetic broad-spectrum antimicrobial drugs are widely used in animal breeding because of its good bactericidal effects, low price and other advantanges. but the metabolites of nitrofuran antibiotics can accumulate in animals, which causes serious hazards to public health, and as a result, most countries in the world including China have had the nitrofuran drugs as the banned drugs.Currently, the colloidal gold immunochromatography assay has seized its own unique advantages in the detection of veterinary drug residues.This detection technology of drug residues in different seafood becomes more simple and faster than large instrumental analysis methods, and as a result,it has been promoted in a wide range. After years of research, the colloidal gold immunochromatography assay test card for the corresponding drug residues has been launched in the market. But their applicability in rapid detection, such as the simplification for sample pretreatment to avoid the extraction of organic reagents and repeated centrifuging during the detection, has not been studied or reported.In this study, the experimental subject is Apostichopus japonicus. The optimized sample pretreatment method is the combination of the use of high temperature heating and hydrochloric acid treatment,which could effectively eliminate the matrix effects of protein and polysaccharide to omit the tedious step of the extraction of organic reagents after the derivatization and establish a new pretreatment method and achieve a desired direct detection. The entire testing process was limited to less than one hour, and performed without any large laboratory instruments,and what’s more the sensitivity of the detection corresponded with the relevant national standards. This established method solved the common problem that there was no effective pretreatment in the rapid detection by the existing commercially GICA test strip. The main contents were as follows:1. Exploring a variety of factors in early experiments, including the detection performance of the test card and the impact of Apostichopus japonicu matrixs on the test results of the direct detection,which was performed without extraction after the derivatization. It concluded that when C(AOZ)≥0.25μg/L, the test results was positive, and the result had good stability. It also concluded that the combined action of polysaccharide and protein produced the greatest interference on the detection.2. Exploring the best ways to eliminate matrix effects. It showed that the best pretreatment approach was heating the samples in a water bath at 80℃ for 5 min before the derivatization combining with the treatment of hydrochloric acid of which the final concentration was 0.1 mol/L, which can effectively eliminate matrix effects. Optimization result of derivatization was deriving for 45min in water bath at 60℃, which could further shorten the detection time and achieve the desired detection results. Limit of Detection (LOD) for furazolidone metabolite was 0.5μg/kg in the eventual established GICA, and the repeatability and specificity of detection method was also good.3. Verifying the eventual established pretreatment process in the real samples.The test results of the same real samples from the two detection method, the National standard method(LC-MS/MS) and the method established in this paper, were identical with each other which both were the strong positive results. It could be concluded that the optimized sample pretreatment method established in this paper could be applied in the rapid detection of AOZ in the real samples. |
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