Mycotoxin contamination of cereals and related products used for feed can cause intoxication, especially in farm animals. Therefore, efficient analytical tools for the qualitative and quantitative analysis of toxic fungal metabolites in feed are required. Current methods usually include an extraction step, a clean-up step to reduce or eliminate unwanted co-extracted matrix components and a separation step with suitably specific detection ability. Quantitative methods of analysis for most mycotoxins use immunoaffinity clean-up with high-performance liquid chromatography (HPLC) separation in combination with UV and/or fluorescence detection. Screening of samples contaminated with mycotoxins is frequently performed by thin layer chromatography (TLC), which yields qualitative or semi-quantitative results. Nowadays, enzyme-linked immunosorbent assays (ELISA) are often used for rapid screening. A number of promising methods, such as fluorescence polarization immunoassays, dipsticks, and even newer methods such as biosensors and non-invasive techniques based on infrared spectroscopy, have shown great potential for mycotoxin analysis. Currently, there is a strong trend towards the use of multi-mycotoxin methods for the simultaneous analysis of several of the important Fusarium mycotoxins, which is best achieved by LC-MS/MS (liquid chromatography with tandem mass spectrometry). This review focuses on recent developments in the determination of mycotoxins with a special emphasis on LC-MS/MS and emerging rapid methods.
Food allergies have become an important health issue especially in industrialized countries. Undeclared allergenic ingredients or the presence of "hidden" allergens because of contamination during the food production process pose great health risks to sensitised individuals. The EU directive for food labelling lists allergenic foods that have to be declared on food products by the manufacturers. The list includes gluten-containing cereals, crustaceans, eggs, fish, peanuts, soybeans, milk, various nuts (e.g. almond, hazelnut, and walnut, etc.), celery, mustard, sesame seeds, lupin, and molluscs. Reliable methods for detection and quantification of food allergens are needed that can be applied in a fast and easy-to-use manner, are portable, and need only limited technical equipment. This review focuses on the latest developments in food allergen analysis with special emphasis on fast immunoanalytical methods such as rapid enzyme-linked immunosorbent assays (ELISA), lateral-flow immunochromatographic assays (LFA) and dipstick tests. Emerging technologies such as immunochemical microarrays and biosensors are also discussed and their application to food allergen analysis is reviewed. Finally, a comprehensive overview of rapid immunochemical test kits that are currently available commercially is given in tabular form.
Solid phase extraction (SPE) based on molecularly imprinted polymers (MIPs) is a novel approach for sample preparation and preconcentration, gaining increased interest in the fields of environmental, clinical, and food analysis. The first application combining MIPs with SPE for advanced beverage analysis is reported. MIPs for the flavonoid quercetin have been generated, using quercetin as a template molecule in a self-assembly approach and yielding imprinting of 1% of the used template. The MIP achieved a capacity of 0.4 g quercetin per gram polymer and a recovery rate of 98.2%. The application of these synthetic receptors as SPE material for the selective extraction and preconcentration of quercetin from synthetic and red wine samples was investigated. Red wine samples from a French Merlot were directly applied onto the SPE cartridge. The collected fractions were analyzed by high-pressure liquid chromatography. For verification of the obtained results, a similarly prepared nonimprinted polymer and a classical octadecyl silane reversed-phase cartridge were applied as the SPE matrix during control experiments. The MIP enabled the selective extraction of quercetin from a complex matrix, such as red wine, spiked with 8.8 mg per liter quercetin, demonstrating the potential of molecularly imprinted solid phase extraction for rapid, selective, and cost-effective sample pretreatment.
In this paper, the results obtained by five independent methods for the quantification of fumonisins B(1), B(2), and B(3) in raw maize are reported. Five naturally contaminated maize samples and a reference material were analyzed in three different laboratories. Although each method was validated and common calibrants were used, a poor agreement about fumonisin contamination levels was obtained. In order to investigate the interactions among analyte and matrix leading to this lack of consistency, the occurrence of fumonisin derivatives was checked. Significant amounts of hidden fumonisins were detected for all the considered samples. Furthermore, the application of an in vitro digestion protocol to raw maize allowed for a higher recovery of native fumonisins, suggesting that the interaction occurring among analytes and matrix macromolecules is associative rather than covalent. Depending on the analytical method as well as the maize sample, only 37-68% of the total fumonisin concentrations were found to be extractable from the samples. These results are particularly impressive and significant in the case of the certified reference material, underlying the actual difficulties in ascertaining the trueness of a method for fumonisin determination, opening thus an important issue for risk assessment.
Novel qualitative as well as semiquantitative rapid strip tests for screening of T-2 mycotoxin in agricultural commodities were developed. Colloidal gold particles were coated with monoclonal anti-T-2 antibodies and used as detector reagent, indicating the strip test results by formation of up to two colored lines in a competitive assay format. The test line comprises a protein conjugate of the T-2 mycotoxin and the control line an antispecies-specific antibody to confirm the correct test development. To perform the test, 5 g of sample was extracted in a ratio of 1:5 with methanol/water (70:30) by shaking for 3 min and the extract directly used without further cleanup steps. The T-2 toxin lateral flow device (LFD) presented has a cutoff level around 100 microg/kg for naturally contaminated wheat and oat. The semiquantitative test may be used in the lower micrograms per kilogram range and allows for rapid semiquantitative photometric classification of the level of sample contamination. For both tests, results were obtained within 4 min. The developed LFDs therefore allow for the first time fast and on-site screening for the determination of T-2 toxin in cereals.
An overview is given on recent trends and applications of rapid immunodiagnostic tests for screening of food and feed for mycotoxins. Different test formats are discussed, and challenges in the development of lateral-flow devices for on-site determination of mycotoxins, with requirements such as being robust, fast, and cost-effective, are briefly elucidated.
This article covers the latest activities in mycotoxin analysis and the advances of its respective quality assurance. The majority of mycotoxin analyses carried out in the laboratories is still based on physicochemical methods, which are continually improved. For example, immunoaffinity columns and multifunctional clean-up columns have become of increasing importance and in some areas of mycotoxin analysis they have more or less displaced conventional liquid-liquid partitioning or column chromatography during clean-up. The need for rapid yes/no decisions on the other hand has led to a number of new screening methods. In particular, rapid and easy-to-use test kits based on immunoanalytical principles or the generation of artificial macromolecular receptors employed in molecularly imprinted polymers (MIPs) have made good progress. Further research in mycotoxin analysis is pursued in the field of biosensors and also the potential of infrared spectroscopic techniques as screening method has been demonstrated. In the area of multi mycotoxin analysis the most promising development was observed in mass spectrometry. At the same time, several interlaboratory studies in the field of mycotoxin analysis revealed problems proven by high between laboratory standard deviation and non-traceable results. This not only shows the necessity of reliable methods and well defined performance characteristics but also the need for appropriate calibrants of defined concentration and stated purity. A certified zearalenone (ZON) calibrant is already available and a certified calibrant containing various trichothecenes is currently under development. (Certified) reference materials are available for aflatoxins in a number of commodities, ochratoxin A (OTA) in wheat, deoxynivalenol (DON) in maize and wheat, and ZON in maize. With these measures important steps towards traceability of results in mycotoxin analysis have been achieved.
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