Polymerase chain reaction techniques for food allergen detection

Food allergies represent an important health problem in industrialized countries. Undeclared allergenic foods as contaminants in food products pose a major risk for sensitized persons. Reliable detection and quantification methods for food allergens are necessary to ensure compliance with food labeling and improve consumer protection. The methods currently used for the detection of potential allergens in foods are to target either the allergen itself or a marker that indicates the presence of the offending food. As markers for the presence of potentially allergenic foods or ingredients, specific proteins or DNA fragments are targeted. In routine food analysis, the enzyme-linked immunosorbent assay (ELISA) and the polymerase chain reaction (PCR) in the form of a real-time PCR or in combination with an ELISA have been used. The availability, the characteristics, and some future aspects of DNA-based methods in the rapid and sensitive detection of potentially allergenic food constituents or contaminations are discussed in this review.     

Die Reaktionen von Eisen(III)-chlorid mit Trithiazylchlorid. Die Kristallstruktur von [S4N4Cl]+[FeCl4]⊖

Reactions of Iron Trichloride with Trithyazyl Chloride. Crystal Structure of [S₄N₄Cl]⁺[FeCl₄]^? Iron trichloride reacts with (NSCl)₃ yielding S₄N₄[FeCl₄]₂, S₃N₃Cl₂[FeCl₄] or S₄N₄Cl[FeCl₄], depending on the reaction conditions. The i.r. spectra prove the presence of [FeCl₄]^? ions for all three compounds. The ⁵⁷Fe-Mössbauer spectra show a slight quadrupole splitting at 80 K for S₃N₃Cl₂[FeCl₄] (ΔE^Q = 0.42 mm · s^?1) and S₄N₄Cl[FeCl₄] (ΔE^Q = 0.23 mm · s^?1), which indicates a slight deformation of the FeCl₄^? tetrahedra. The crystal structure of S₄N₄Cl[FeCl₄] was determined and refined with X-ray diffraction data (2549 independent reflexions, R = 0.026). S₄N₄Cl[FeCl₄] crystallizes in the triclinic space group P1 with two formula units per unit cell. The lattice constants are a = 712, b = 911, c = 1006 pm, α = 76.5°, β = 83.8° and γ = 80.5°. The structure consists of the so far unknown [S₄N₄Cl]^⊕ cations and slightly deformed FeCl₄^? ions. The [S₄N₄Cl]^⊕ ion consists of a S₄N₄ ring built up of two nearly planar S₃N₂ fragments having a dihedral angle of 136°. The average SN bond length is 157 pm, the SCI bond length 214 pm.     

Validation studies and proficiency testing

Genetically modified organisms (GMOs) entered the European food market in 1996. Current legislation demands the labeling of food products if they contain <1% GMO, as assessed for each ingredient of the product. To create confidence in the testing methods and to complement enforcement requirements, there is an urgent need for internationally validated methods, which could serve as reference methods. To date, several methods have been submitted to validation trials at an international level; approaches now exist that can be used in different circumstances and for different food matrixes. Moreover, the requirement for the formal validation of methods is clearly accepted; several national and international bodies are active in organizing studies. Further validation studies, especially on the quantitative polymerase chain reaction methods, need to be performed to cover the rising demand for new extraction methods and other background matrixes, as well as for novel GMO constructs.     

Reaktionen von Uranpentabromid Die Kristallstrukturen von PPh4[UBr6], PPh4 [UBr6] · 2CCl4, (PPh4)2[UBr6] · 4CH3CN und (PPh4)2[UO2Br4] · 2CH2Cl2

Reactions of Uranium Pentabromide. Crystal Structures of PPh₄[UBr₆], PPh₄[UBr₆] · 2CCl₄, (PPh₄)₂[UBr₆] · 4CH₃CN, and (PPh₄)₂[UO₂Br₄] · 2CH₂Cl₂ PPh₄[UBr₆] and PPh₄[UBr₆] · 2CCl₄ were obtained from UBr₅ · CH₃CN and tetraphenylphosphonium bromide in dichloromethane, the latter being precipitated by CCl₄. Their crystal structures were determined by X-ray diffraction. PPh₄[UBr₆]: 2101 observed reflexions, R = 0.090, space group C2/c, Z = 4, a = 2315.5, b = 695.0, c = 1805.2 pm, β = 96.38°. PPh₄[UBr₆] · 2CCl₄: 2973 reflexions, R = 0.074, space group P2₁/c, Z = 4, a = 1111.5, b = 2114.2, c = 1718.7 pm, β = 95.42°. Hydrogen sulfide reduces uranium pentabromide to uranium tetrabromide. Upon evaporation, bromide is evolved from solutions of UBr₅ with 1 or more then 3 mol equivalents of acetonitrile in dichlormethane yielding UBr₄ · CH₃CN and UBr₄ · 3CH₃CN, respectively. These react with PPh₄Br in acetonitrile affording (PPh₄)₂[UBr₆] · 4CH₃CN, the crystal structure of which was determined: 2663 reflexions, R = 0.050, space group P2₁/c, Z = 2, a = 981.8, b = 2010.1, c = 1549.3 pm, β = 98.79°. By reduction of uranium pentabromide with tetraethylammonium hydrogen sulfide in dichloromethane (NEt₄)₂[U₂Br₁₀] was obtained; (PPh₄)₂[U₂Br₁₀] formed from UBr₄ and PPh₄Br in CH₂Cl₂. Both compounds are extremely sensitive towards moisture and oxygen. The crystal structure of the oxydation product of the latter compound, (PPh₄)₂[U0₂Br₄]· 2 CH₂Cl₂, was determined: 2163 reflexions, R = 0.083, space group C2/c, Z = 4, a = 2006.3, b = 1320.6, c = 2042,5 pm, β = 98.78°. Mean values for the UBr bond lengths in the octahedral anions are 266.2 pm for UBr₆-, 276.7 pm for UBr₆^2? and 282.5 pm for UO₂Br₄^2?     

Comparison of two post-column derivatization systems, ultraviolet irradiation and electrochemical determination, for the liquid chromatographic determination of aflatoxins in food

This study compared 2 post-column derivatization (PCD) techniques for the determination of aflatoxins B1, B2, G1, and G2 (AFB1, AFB2, AFG1, and AFG2) by fluorescence detection after liquid chromatographic separation: ultraviolet (UV) irradiation (PCD(UV)) and electrochemical bromination (PCD(EC)). Photochemical fluorescence enhancement was obtained with 2 different commercially available systems (PCD(UV1) and PCD(UV2)). An electrochemical bromination apparatus was used for bromination. Analyses of naturally contaminated or spiked samples of corn, pistachio paste, peanut butter, fig paste, and animal feed showed that neither of the techniques resulted in derivatization-specific matrix interferences for any of the matrixes under study, even when extracts were not completely purified. The response ratios PCD(UV)/PCD(EC) for AFB1, AFB2, AFG1, and AFG2 were 0.86, 0.96, 0.70, and 0.96, respectively, for PCD(UV1) and 0.82, 0.95, 0.60, and 0.90, respectively, for PCD(UV2). The long-term use of the UV lamps (300 h for PCD(UV1) and 343 h for PCD(UV2)) in the photochemical detectors showed that these ratios remained stable throughout the time frame investigated. The relative standard deviation obtained for each of the devices during the in-house validation study ranged from 0.3 to 1.8% for PCD(UV1), from 0.8 to 1.3% for PCD(UV2), and from 0.9 to 2.0% for PCD(EC).     

Subdivision Strategies for Boxes in Branch-and-Bound Nonlinear Solvers and Verification

Most verified solvers for nonlinear interval systems of equations comprise two strategies: a branch-and-bound-type “location” phase for excluding regions that cannot contain a solution, and a “verification” phase for proving that the remaining regions do indeed contain solutions. In the first phase, subdivision is crucial for the efficiency of the solvers. We discuss several ways for subdivision and present robust strategies that are suited for a variety of nonlinear systems. Particular focus is on the choice of subdivision directions, subdivision points and the handling of unbounded intervals. Furthermore we discuss a method to discard parts of a box within subdivision. Numerical evaluations are given based on the nonlinear interval solver SONIC. In the verification phase, further subdivision can increase the strength of the verification tests. In this paper, we address methods for the rigorous implementation of symmetrical subdivision which is needed, e.g., in existence tests based on Borsuk’s theorem.     

Development and validation of an LC–MS/MS method for the quantification of artificial sweeteners in human matrices

Artificial sweeteners are widely used as substitutes for sugar. The sweeteners are generally considered safe, however their whereabouts during pregnancy and lactation and the effect on child development are poorly explored. There is a need for new tools to measure these substances during pregnancy and lactation. Here, we describe the development and validation of a sensitive liquid chromatography–tandem mass spectrometry method for the simultaneous quantification of acesulfame, cyclamate, saccharin and sucralose in human plasma, umbilical cord blood, amniotic fluid and breast milk. The samples were prepared by protein precipitation and separated on a Luna Omega Polar C₁₈ column (2.1 × 50 mm, 1.6 μm). Electrospray ionization in negative mode and multiple reaction monitoring were used to monitor the ion transitions. The validated concentration ranges were from 1 to 500 ng/ml (10–500 ng/ml for sucralose). Interassay precisions were all ≤15% and the accuracies were within ±15%. Stability, linearity, dilution integrity, carryover and recovery were also examined and satisfied the validation criteria. Finally, this analytical method was successfully applied on spiked samples of plasma, umbilical cord blood, amniotic fluid and breast milk, proving its suitability for use in clinical studies on artificial sweeteners, including during pregnancy and lactation.