Search

3-Amino-5-methylmorpholino-2-oxazolidinone (AMOZ) is the metabolite of furaltadone (FTD), one kind of nitrofuran antibiotics which have been banned as veterinary drugs due to the potential harmful effects on human health. AMOZ is stable in tissue matrices, and can be used as an indicator for illegal usage of FTD. In this study, a novel ultrasensitive competitive immunochromatographic assay (ICA) based on surface-enhanced Raman scattering (SERS) for direct detection of AMOZ in tissue and urine samples was developed. Monoclonal antibody (mAb) against AMOZ and Raman reporter 4-mercaptobenzoic acid (MBA) dual labelled colloidal GNPs (e.g. Ab-Au-MBA) was prepared, characterized and used as an immunoprobe. The specific Raman scattering intensity of MBA on the test line of the ICA strip was measured for quantitative determination of AMOZ. The assay was completed within 15 min. The values of IC50 and limit of detection (LOD) of the assay for AMOZ were 0.04 ng mL−1 and 0.28 pg mL−1, respectively. There was no cross-reactivity (CR) of the assay with other three nitrofurans and five compounds. The ICA strips could be used for 7 weeks without loss of activity. The tissue and urine samples were spiked with AMOZ and the recoveries of AMOZ were in range of 87.8–105.9% with the relative standard deviations (RSD) of 1.4–9.0% (n = 3). The results demonstrated that the proposed SERS-based ICA was able to directly detect AMOZ with high sensitivity, specificity, stability, precision and accuracy, and might be as a powerful method for the analysis of other analytes in different matrices.

BACKGROUND Furaltadone (FTD) is a type of nitrofuran and has been banned in many countries as a veterinary drug in food-producing animals owing to its potential carcinogenicity and mutagenicity. FTD is unstable in vivo, rapidly metabolizing to 3-amino-5-methylmorpholino-2-oxazolidinone (AMOZ); thus AMOZ can be used as an indicator for illegal usage of FTD. Usually, for the determination of nitrofurans, the analyte is often a derivative of the metabolite rather than the metabolite itself. In this study, based on the monoclonal antibody (mAb) against AMOZ, a competitive immunochromatographic assay (ICA) using a colloidal gold–mAb probe for rapid and direct detection of AMOZ without a derivatization step in meat and feed samples was developed. RESULTS The intensity of red color in the test line is inversely related to the analyte concentration and the visual detection limit was found to be 10 ng mL−1. The performance of this assay was simple and convenient because the tedious and time-consuming derivatization step was avoided. The ICA detection was completed within 10 min. The ICA strips could be used for 7 weeks at room temperature without significant loss of activity. The AMOZ spiked samples were detected by ICA and confirmed by enzyme-linked immunosorbent assay. The results of the two methods were in good agreement. CONCLUSION The proposed ICA provides a feasible tool for simple, sensitive, rapid, convenient and semi-quantitative detection of AMOZ in meat and feed samples on site. To our knowledge, this is the first report of the ICA for direct detection of AMOZ. © 2013 Society of Chemical Industry

Phenylethanolamine A (PA) is a new emerged β-adrenergic agonist illegally used as feed additives for growth promotion. In this study, a highly sensitive and specific indirect competitive enzyme-linked immunosorbent assay (ELISA) for the detection of PA in tissue and feed samples was developed and confirmed by liquid chromatography tandem mass spectrometry (LC–MS/MS). By reduction of nitryl group to amino group, the PA derivative was synthesized and coupled to carrier proteins with diazobenzidine method. The antisera obtained from four immunized rabbits were characterized in terms of sensitivity and specificity. All antisera displayed high sensitivity with IC50 values lower than 0.48 ng mL−1. The most sensitive ELISA was established with IC50 and limit of detection (LOD) values of 0.049 ng mL−1 and 0.003 ng mL−1, respectively. The cross-reactivity (CR) values of the antisera with three frequently used β-adrenergic agonists (clenbuterol, salbutamol and ractopamine) were lesser than 0.39%; there was no CR of the antisera with other six compounds including two structurally related substances (isoproterenol, phenylephrine). To investigate the accuracy and precision of the assay, swine kidney, liver, meat and feed samples were fortified with PA at different content and analyzed by ELISA. Acceptable recovery rates of 92.2–113.7% and intra-assay coefficients of variation of 3.8–10.9% (n=3) were achieved. Seven spiked samples were simultaneously analyzed by ELISA and LC–MS/MS. There was a high correlation coefficient of 0.9956 (n=7) between the two methods. The proposed ELISA proven to be a feasible quantitative/screening method for PA analysis in tissue and feed samples with the properties of high sensitivity and specificity, high sample throughput and low expensive.

Phenylethanolamine A (PA) is a new kind of β-agonist, which was illegally used as a feed additive for growth promotion in China. In this study, a novel immunochromatographic assay (ICA) based on surface-enhanced Raman scattering (SERS) for the ultrasensitive and quantitative detection of phenylethanolamine A is presented. The principle of this new ICA is similar to that based on colloidal gold particles, but using Au(MBA)@Ag-Ab [e.g., polyclonal antibody of PA labeled Au@Ag core-shell nanoparticles (NPs) sandwiched with a Raman reporter (4-mercaptobenzoic acid, MBA)] as a probe. After ICA procedures, the specific Raman scattering intensity of MBA on the test line was measured for quantitative detection of PA. This assay was completed within 15 min. The IC50 and limit of detection (LOD) values of the ICA for PA detection were 0.06 ng mL(-1) and 0.32 pg mL(-1), respectively, which were 1-3 orders of magnitude lower than those obtained by other immunoassays, indicating the ultrasensitivity of this ICA. There was no cross-reactivity (CR) of the assay with another three β-agonists (ractopamine, clenbuterol, and salbutamol), suggesting high specificity of the SERS-based ICA. A spiking experiment revealed that the recoveries of PA from pig urine samples were in range of 99.9- 101.2% with relative standard deviations (RSDs) of 3.6-5.8%. The results demonstrated that this SERS-based ICA was able to quantitatively detect PA in urine samples with high sensitivity, specificity, precision, and accuracy and might be a powerful method for the analysis of other target analytes in the food area.