Irene Nehls

Determination of mycotoxins in foods: current state of analytical methods and limitations

Mycotoxins are natural contaminants produced by a range of fungal species. Their common occurrence in food and feed poses a threat to the health of humans and animals. This threat is caused either by the direct contamination of agricultural commodities or by a “carry-over” of mycotoxins and their metabolites into animal tissues, milk, and eggs after feeding of contaminated hay or corn. As a consequence of their diverse chemical structures and varying physical properties, mycotoxins exhibit a wide range of biological effects. Individual mycotoxins can be genotoxic, mutagenic, carcinogenic, teratogenic, and oestrogenic. To protect consumer health and to reduce economic losses, surveillance and control of mycotoxins in food and feed has become a major objective for producers, regulatory authorities and researchers worldwide. However, the variety of chemical structures makes it impossible to use one single technique for mycotoxin analysis. Hence, a vast number of analytical methods has been developed and validated. The heterogeneity of food matrices combined with the demand for a fast, simultaneous and accurate determination of multiple mycotoxins creates enormous challenges for routine analysis. The most crucial issues will be discussed in this review. These are (1) the collection of representative samples, (2) the performance of classical and emerging analytical methods based on chromatographic or immunochemical techniques, (3) the validation of official methods for enforcement, and (4) the limitations and future prospects of the current methods.

Determination of total sulfite in wine by ion chromatography after in-sample oxidation.

Sulfur dioxide (SO2) or sulfites are the most common preservatives used in winemaking. The level of total SO2 is subject to regulation. Currently, the regulatory determination of total SO2 (including sulfites) is done by the optimized Monier-Williams (OMW) method, which includes time-consuming distillation and titration steps. This paper describes the development and application of an alternative, rapid, straightforward, and reliable method for the determination of total sulfite in wine. In this method, a simple oxidation step using alkaline hydrogen peroxide (H2O2) solution is followed by ion chromatographic (IC) analysis of sulfate coupled with conductometric detection. Thirteen wines were analyzed in order to compare the in-sample oxidation method with the OMW-procedure. A t-test revealed satisfying compliance regarding sample preparation, i.e., alkaline H2O2 treatment and acidic distillation (OMW method). Comparable results were also obtained between IC analysis and acid/base titration. Our results indicate that the novel method (limit of quantification: 4 mg SO2 L(-1)) is well suited for the cost-efficient monitoring of regulatory limits.

Hexabromocyclododecane Enantiomers: Microsomal Degradation and Patterns of Hydroxylated Metabolites

The degradation of the enantiomers of α-, β-, and γ-hexabromocyclododecane (HBCD) by phase I metabolism was investigated using induced rat liver microsomes. HBCD isomers were quantified using HPLC-MS/MS (ESI(-)) after separation on a combination of a reversed phase and a chiral analytical column. The degradation of all six isomers followed first-order kinetics and the estimated half-lives ranged from 6.3 min for both β-HBCD enantiomers to 32.3 min in case of (+)-γ-HBCD. (+)-α- and (-)-γ-HBCD displayed significantly shorter half-lives than their corresponding antipodes. It could be shown that this degradation led to a significant enrichment of the first eluting enantiomers (-)-α- and (+)-γ-HBCD. Individual patterns of mono- and dihydroxylated derivatives obtained from each α- and γ-HBCD enantiomer were seen to be distinctly characteristic. The patterns of monohydroxylated HBCD derivatives detected in liver and muscle tissues of pollack, mackerel and in herring gull eggs were largely similar to those observed in the in vitro experiments with rat liver microsomes. This enabled individual hydroxy-HBCDs to be assigned to their respective parent HBCD enantiomers.

Analysis and Migration of Phthalates in Infant Food Packed in Recycled Paperboard

The contamination of infant food with substances from its packaging due to migration processes is still a problem. Most recently, great attention was paid to the migration of epoxidized soybean oil (ESBO) and phthalates from twist-off closures into baby food packed in glass jars. Besides, packaging made of recycled fiber materials such as paper and paperboard were found to be the source of contaminants in dry and powdery foodstuffs such as sugar, rice, and maize flour. In this study 20 infant food samples packed in recycled paperboard containers were tested for phthalates and diisopropyl naphthalenes (DIPN), known incorporated substances in recycled paper. Furthermore, the barrier function of different secondary packaging materials (paper and aluminum-coated foil) was investigated. The highest contents of phthalates (mainly diisobutyl phthalate, DiBP) and DIPN in infant food samples were found for those foods packed in inner bags made of paper. Migration experiments were performed under authentic conditions to evaluate possible transfer mechanism (gas phase, direct contact) of phthalate esters into foodstuff. It is shown that paper does not provide an appropriate barrier against migration of semipolar compounds such as phthalates. The air space itself otherwise effectively prevents migration of the less volatile phthalates under the applied conditions.

Determination of the Alternaria mycotoxin tenuazonic acid in cereals by high-performance liquid chromatography–electrospray ionization ion-trap multistage mass spectrometry after derivatization with 2,4-dinitrophenylhydrazine

Tenuazonic acid (TA) is a major Alternaria mycotoxin. In the present work a novel approach for the detection of TA in cereals by liquid chromatography-ion-trap multistage mass spectrometry after derivatization with 2,4-dinitrophenylhydrazine is described. The product of the derivatization reaction and its major MS(2) fragments were characterised by Fourier transform-ion cyclotron resonance tandem mass spectrometry. Without preconcentration, the established method features a limit of detection of 10 microg/kg using 2g of sample in a rapid workup procedure. Accuracy, precision and linearity were evaluated in the working range of 50-5000 microg/kg. TA was detected in 13 and quantified in 3 out of 27 cereal samples obtained from a local supermarket, the average content being 49 microg/kg (highest incidence: 851+/-41 microg/kg).

Enantiomer-specific analysis of hexabromocyclododecane in fish from Etnefjorden (Norway).

High-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) was applied to sterospecifically quantify the content of alpha-, beta-, and gamma-hexabromocyclododecane (HBCD) in six fish species from the Norwegian Etnefjorden. A combination of a beta-PM cyclodextrin and an achiral column enabled the paired chromatographic separation of the stereoisomers in the order (-)-alpha-, (+)-alpha-, (-)-beta-, (+)-beta-, (+)-gamma- and, (-)-gamma-HBCD. The limits of detection were in the range of 6-21 pg g(-1) depending on the stereoisomer and the concentrations of alpha-, beta-, and gamma-HBCD in fillets ranged from <5.4 ng g(-1) to 11.1 microg g(-1) lipid weight. alpha-HBCD enantiomers were throughout dominating, and in most cases the accumulation of the respective first eluted enantiomers ((-)-alpha-, (-)-beta- and (+)-gamma-HBCD) was observed. Deviations from the racemic EF-value were considered to be significant if it was outside of the expanded uncertainty range for each of the racemic HBCD-ratios. The composition of HBCD isomers varied between the investigated fish species and the relative high values for the gamma-HBCD concentrations for the bottom-dwellers flounder and thorny skate seems to echo the HBCD pattern of ocean sediments.

Hydrothermally carbonized plant materials: Patterns of volatile organic compounds detected by gas chromatography

The nature and concentrations of volatile organic compounds (VOCs) in chars generated by hydrothermal carbonization (HTC) is of concern considering their application as soil amendment. Therefore, the presence of VOCs in solid HTC products obtained from wheat straw, biogas digestate and four woody materials was investigated using headspace gas chromatography. A variety of potentially harmful benzenic, phenolic and furanic volatiles along with various aldehydes and ketones were identified in feedstock- and temperature-specific patterns. The total amount of VOCs observed after equilibration between headspace and char samples produced at 270°C ranged between 2000 and 16,000μg/g (0.2-1.6wt.%). Depending on feedstock 50-9000μg/g of benzenes and 300-1800μg/g of phenols were observed. Substances potentially harmful to soil ecology such as benzofurans (200-800μg/g) and p-cymene (up to 6000μg/g in pine wood char) exhibited concentrations that suggest restrained application of fresh hydrochar as soil amendment or for water purification.

13C-N.M.R. spectroscopic study of the homogeneous sulphation of cellulose and xylan in the N2O4-DMF system

Abstract By means of 13 C-n.m.r. spectroscopy the substituent distribution within the anhydroglucose and anhydroxylose units, respectively, in the systems N 2 O 4 -DMF-cellulose, and N 2 O 4 -DMF-SO 3 -xylan has been investigated. In the systems containing cellulose, a derivatization exclusively at C-6 is observed on dissolution in N 2 O 4 -DMF and on subsequent reaction with SO 2 , while with SO 3 as the acylating agent some substitution also takes place on the secondary OH groups at higher d.s. The OH groups of xylan are substituted to a rather low d.s. by nitrite ester groups in the system N 2 O 2 -DMF and by sulphate half-ester groups on subsequent reaction with SO 2 or SO 3 . The results are discussed on the basis of a transesterification from the nitrite to the sulphate ester.

Temporal trend (1988-2008) of hexabromocyclododecane enantiomers in herring gull eggs from the german coastal region

Levels of α-, β-, and γ-hexabromocyclododecane (HBCD) were determined in pooled eggs from herring gulls (Larus argentatus) sampled on three bird sanctuaries near the German North Sea coast between 1988 and 2008 (Mellum and Trischen) and the German Baltic Sea coast between 1998 and 2008 (Heuwiese) and archived by the German Environmental Specimen Bank. Pressurized fluid extraction, gel permeation chromatography, and LC-MS/MS using (13)C(12)-labelled isotope standards and a chiral column were applied. α-HBCD was the dominating diastereomer and ranged between 3.7 and 107 ng g(-1)lw while β- and γ-HBCD were throughout close to LOQ. The highest α-HBCD concentration was found in eggs from Mellum sampled in the year 2000. Interestingly, HBCD in eggs from the three islands displayed similar time courses with levels increasing to a peak contamination around 2000 and decreasing levels ever since. Chiral signatures of α-HBCD in eggs differed among the islands but indicated a preferential enrichment of the first eluting enantiomer (-)-α-HBCD.

Quantification of Ethyl Glucuronide in Hair: Effect of Milling on Extraction Efficiency

AIM The objective of the study was to provide conclusive evidence for the effect of particle size reduction as by milling on the extractable content of ethyl glucuronide (EtG) of hair samples. METHODS A number of real case hair samples and two pooled hair materials with EtG contents in the range of 10-30 pg/mg were systematically compared with regard to the extraction yield of EtG after cutting to 2-3 mm length and pulverization with a ball mill. After the respective treatment the samples were submitted to aqueous extraction followed by quantification of EtG using HPLC-MS/MS. RESULTS It was unequivocally demonstrated that milling of hair samples prior to aqueous extraction significantly increases the extractable EtG content compared with cut hair. The effect ranged between 137 and 230% and was seen to occur regardless of the extent of pulverization. Cooling of samples was not necessary to prevent partial degradation of EtG during the grinding procedure. CONCLUSION The options currently employed at choice in analytical practice (cutting or milling) were seen to significantly affect the extractable amount of EtG in hair. This is suspected to influence the degree of equivalence of quantification results obtained in different laboratories as well as their respective classification of a test subjects drinking behaviour on the basis of currently recommended cut-off values.