Rapid method for the determination of multiple mycotoxins in wines and beers by LC-MS/MS using a stable isotope dilution assay.

A "dilute and shoot" method for the liquid chromatography-tandem mass spectrometric (LC-MS/MS) determination of multiple mycotoxins (aflatoxins B1, B2, G1, G2, ochratoxin A (OTA), fumonisins (F) B1 and B2, zearalenone, deoxynivalenol, T-2 toxin, and HT-2 toxin) in wines and beers has been developed and validated. Separation was accomplished using ultrahigh-performance liquid chromatography (UHPLC) with <10 min analysis time. Mycotoxins were detected by dynamic multiple reaction monitoring (MRM) in positive electrospray ionization mode. Due to matrix effects, (13)C-uniformly labeled mycotoxins were added to the sample extracts prior to LC-MS/MS analysis. With external calibration, recoveries were 18-148% for white wines, 15-118% for red wines, and 20-125% for beers, at three spiking levels. The (13)C-labeled internal standards compensated for matrix effects effectively, with overall recoveries of 94-112% for white wines, 80-137% for red wines, and 61-131% for beers, with greater recoveries for FB1 and FB2, at three spiking levels. The relative standard deviation was <20% for all analytes in the wines and beers. This method was applied to a USDA-funded nationwide survey of domestic and imported wines and beers for the determination of OTA and extended to include other mycotoxins.

Validation of a rapid lateral flow test for the simultaneous determination of ß-lactam drugs and flunixin in raw milk.

ß-Lactam antibiotics are the most commonly used drugs on dairy farms. ß-Lactam residues in milk are kept out of the human milk supply with good agricultural practices and mandatory truck screening performed by the dairy industry under Appendix N of the Pasteurized Milk Ordinance. Flunixin, a nonsteroidal and anti-inflammatory drug, appears in dairy cattle tissue residues with a frequency similar to the occurrence of penicillin G. This creates concern that flunixin residues could be in milk and would go undetected under current milk screening programs. A single test that combines mandatory ß-lactam screening with voluntary flunixin screening is an economical approach for monitoring and controlling for potential flunixin or 5-hydroxyflunixin, the primary flunixin metabolite marker in milk. The objective of this study was to validate a ß-lactam and flunixin rapid lateral flow test (LFT) and compare the results obtained with a liquid chromatography-triple quadrupole tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of flunixin and 5-hydroxyflunixin in raw milk with a limit of detection of , 1 ppb, equivalent to 1 ng/ml. Using the LFT, three combined manufactured lots of test strips detected penicillin G at 2.0 ppb, ampicillin at 6.8 ppb, amoxicillin at 5.9 ppb, cephapirin at 13.4 ppb, ceftiofur (total metabolites) at 63 ppb, and 5-hydroxyflunixin at 1.9 ppb at least 90% of the time with 95% confidence. The LFT also detected incurred flunixin milk samples that were analyzed with the LC-MS/MS and diluted to tolerance in raw milk. The detection levels for the LFT are lower than the U.S. safe levels or tolerances and qualify the test to be used in compliance with U.S. milk screening programs.

Chemical inactivation of protein toxins on food contact surfaces.

We compared the kinetics and efficacies of sodium hypochlorite, peracetic acid, phosphoric acid-based detergent, chlorinated alkaline detergent, quaternary ammonium-based sanitizer, and peracetic acid-based sanitizer for inactivating the potential bioterrorism agents ricin and abrin in simple buffers, food slurries (infant formula, peanut butter, and pancake mix), and in dried food residues on stainless steel. The intrinsic fluorescence and cytotoxicity of purified ricin and abrin in buffers decreased rapidly in a pH- and temperature-dependent manner when treated with sodium hypochlorite but more slowly when treated with peracetic acid. Cytotoxicity assays showed rapid and complete inactivation of ricin and crude abrin in food slurries and dried food residues treated 0-5 min with sodium hypochlorite. Toxin epitopes recognized by ELISA decayed more gradually under these conditions. Higher concentrations of peracetic acid were required to achieve comparable results. Chlorinated alkaline detergent was the most effective industrial agent tested for inactivating ricin in dried food residues.

Effect of heat treatment on the quantitative detection of egg protein residues by commercial enzyme-linked immunosorbent assay test kits.

This study examined the changes in the solubility of egg proteins as affected by different heat treatments and compared the performances of three commercial test kits for the quantitation of protein residues in heat-treated samples. National Institute of Standards and Technology (NIST) whole egg standard reference material #8415 and Henningsen spray-dried whole egg powder were subjected to heating in the presence of water at 60 and 100 degrees C, autoclaving for 5 or 10 min, or dry heating at 60-400 degrees C for 10 min. The amount of protein in the heated samples was assayed using the bicinchoninic acid total protein assay as well as egg-specific commercial enzyme-linked immunosorbent assay (ELISA) kits. Elevated heat resulted in a lower level of proteins extracted. Neogen's Veratox kit, which is reactive to multiple proteins in egg, greatly underestimated the amount of residual proteins in the boiled or autoclaved samples. Tepnel BioSystems' Biokits assay, which employs antibodies specific to a heat-stable marker protein (ovomucoid), registered a higher level of protein in these samples. Both test kits substantially underestimated the amount of residual proteins in samples dry-heated at temperatures >176 degrees C. The Morinaga test, using an improved extraction buffer, registered the highest level of protein in the heat-treated NIST samples but not the Henningsen samples. The underestimation by the commercial test kits was attributed to changes in the immunoreactivity of residual proteins after heat treatments and not the differences in the amount of protein extracted. These results suggest that thermal processing may affect the quantitative analysis of allergens and needs to be taken into account in the validation of commercial ELISA test kits.