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HPLC-FLD and LC-MS/MS determination of Aflatoxin M1 from milk after biofixators usage

The toxicity of aflatoxin M1 (AFM1) has a high importance for the food quality and possible entrance in human body. Mycotoxin-binding agents (MBA) could be used for fixation of AFM1 to minimise exposure through dairy products. The purpose of this study was to determine unbounded amounts of AFM1 after adding of lactic acid bacteria (LAB) and β-glucan (cellular yeast components) added firstly for binding then removing aflatoxin AFM1 from milk. LAB were isolated from fresh cow's milk and traditional dairy products such as: fresh cow cheese and cream. All milk samples used in the experiment were first analysed and verified for AFM1 contamination free, than spiked with known amount of AFM1. Membrane filtration method and Centricon Plus-70, a MWCO 100 kDa were selected for removal of resulting biofixator-AFM1 complex from milk.The concentration of AFM1 residues in the samples were determined by high performance liquid chromatography using a fluorescent detector (HPLC-FLD). Previously obtained values were confirmed by high performance liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Before HPLC or LC-MS/MS analysis the samples were purified on immunoaffinity columns. Both methods were fully validated by determination of following validation parameters: selectivity, linearity, precision, accuracy, recovery, limits of detection and quantification. Validation data proved that both methods are suitable for determination of AFM1 residues in milk. Obtained results of AFM1 after removed biofixator-AFM1 complex from milk shown very good correlation between both methods.

Aflatoxin M1, Food safety, Mycotoxin-binding agents, HPLC-FLD, LC-MS/MS

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