Bärbel Vieth

Preparation and structure of high-chlorinated bornane derivatives for the quantification of toxaphene residues in environmental samples

SummaryThe HRGC-determination of Taxophene® residues in the presence of other chlorinated hydrocarbons with similar retention times is often difficult. This problem can satisfactorily be overcome by using purely isolated high-chlorinated bornane derivatives (7–17) as standard. The method is highly selective for measuring toxaphene in complex environmental matrices, such as fish, and can also be used for evaluating the changes in the relative distribution that may have resulted from bioaccumulation and environmental transformation.

CYP2E1 expression in bone marrow and its intra- and interspecies variability : approaches for a more reliable extrapolation from one species to another in the risk assessment of chemicals

Abstract When characterizing the health risks for man by exposure to chemicals, species-specific differences have to be taken into consideration, otherwise extrapolation from animal data to the human situation would be inadequate. The site-specific toxicity of chemicals may be explained by the following alternatives: (1) reactive metabolites are generated in the liver and subsequently transported to the target tissue(s); (2) metabolism of the parent compound occurs in the target tissue, a pathway by which the enzymes necessary for activation must be expressed in the target tissue. Cytochrome P450 2E1 (CYP2E1) is an important phase-I enzyme activating several chemicals. In the study described in this paper, myeloid intra- and interspecies variability in the expression of CYP2E1 has been investigated in rats, rabbits and man, because the bone marrow represents an important target organ for toxic effects of several chemicals, e.g. benzene. CYP2E1 at the protein level was detected by Western blotting and enzyme activities were determined by CYP2E1-dependent hydroxylation of chlorzoxazone (CLX). In the bone marrow of Wistar rats, the CLX hydroxylase activities were within the same order of magnitude (range: 0.1–0.4 pmol/mg protein per min) as previously described for mice (range 0.2–0.8 pmol/mg protein per min), whereas the CYP2E1 activities in two strains of rabbits were significantly higher (range: 1.7–4.7 pmol/mg protein per min) than in the rodents (P < 0.05). In human CD34+ bone marrow stem cells, CYP2E1 could also be detected on the protein level by Western blotting. The data demonstrate a presence of CYP2E1 in the bone marrow of all species investigated, thus supporting the hypothesis of CYP2E1-dependent local metabolism of several chemicals as a factor possibly contributing to their myelotoxicity and haematotoxicity. The data show that intraspecies/intrastrain variability of CYP2E1 activity in rodents is small. However, CYP2E1 activity between rodents and a non-rodent species was quite different indicating considerable interspecies variability.

CYP2E1-dependent benzene toxicity: the role of extrahepatic benzene metabolism

Abstract Benzene, a ubiquitous environmental pollutant, is haematotoxic and myelotoxic. As has been shown earlier, cytochrome P450 2E1 (CYP2E1)-dependent metabolism is a prerequisite for the cytotoxic and genotoxic effects of benzene, but which of the benzene metabolites produces toxicity is still unknown. The observed differences between the toxicity of benzene and that of phenol, a major metabolite of benzene, could be explained by alternative hypotheses. That is, whether (1) toxic benzene effects are caused by metabolites not derived from phenol (e.g. benzene epoxide, muconaldehyde), which are formed in the liver and are able to reach the target organ(s); or (2) benzene penetrates into the bone marrow, where local metabolism takes place, whereas phenol does not reach the target tissue because of its polarity. To further investigate hypothesis 2, we used various strains of mice (AKR, B6C3F1, CBA/Ca, CD-1 and C57Bl/6), for which different toxic responses have been reported in the haematopoietic system after chronic benzene exposure. In these strains, CYP2E1 expression in bone marrow was investigated and compared with CYP2E1 expression in liver by means of two independent methods. Quantification of CYP2E1-dependent hydroxylation of chlorzoxazone (CLX) by high-performance liquid chromatography (HPLC; functional analysis) was used to characterize specific enzymatic activities. Protein identification was performed by Western blotting using CYP2E1-specific antibodies. In liver microsomes of all strains investigated, considerable amounts of CYP2E1-specific protein and correspondingly high CYP2E1 hydroxylase activities could be detected. No significant differences in CYP2E1-dependent enzyme activities were found between the five strains (range of medians, 4.6–12.0 nmol 6-OH-CLX/[mg protein × min]) in hepatic tissue. In the bone marrow, CYP2E1 could also be detected in all strains investigated. However, chlorzoxazone hydroxylase activities were considerably lower (range of medians, 0.2–0.8 × 10−3 nmol 6-OH-CLX/[mg protein × min]) compared with those obtained from liver microsomes. No significant (P > 0.05) interstrain differences in CYP2E1 expression in liver and/or bone marrow could be observed in the mouse strains investigated. The data obtained thus far from our investigations suggest that strain-specific differences in the tumour response of the haematopoietic system of mice chronically exposed to benzene cannot be explained by differences in either hepatic or in myeloid CYP2E1-dependent metabolism of benzene.

Toxicologically relevant phthalates in food.

Various phthalates have been detected in a wide range of food products such as milk, dietary products, fat-enriched food, meat, fish, sea food, beverages, grains, and vegetables as well as in breast milk. Here we present an overview on toxicologically considerable phthalate levels in food reported in the literature. The most common phthalates detected are di-(2-ethylhexyl) phthalate (DEHP), di-n-butyl phthalate (DnBP), and di-isobutyl phthalate (DiBP). Milk analyses demonstrate that background levels in unprocessed milk are usually low. However, during processing the phthalate contents may significantly increase due to migration from plastic materials in contact with food. Among dietary products fat-enriched food such as cheese and cream were identified with highest levels of DEHP. Plasticized PVC from tubes, conveyor belts, or disposable gloves used in food processing is an important source for contamination of food, especially of fatty food. Paper and cardboard packaging made from recycled fibers are another important source of contamination. In addition, gaskets used in metal lids for glass jars have been identified as possible source for the contamination of foodstuffs with phthalates. The highest concentrations of DEHP reported (>900 mg kg(-1)) were detected in food of high fat content stored in such glass jars. Beyond classical food, DEHP and DnBP were identified in human breast milk samples as the main phthalate contaminants. Phthalate monoesters and some oxidative metabolites were also quantified in breast milk.

Physiologically based toxicokinetic modelling as a tool to assess target organ toxicity in route-to-route extrapolation--the case of coumarin.

Coumarin (1,2-benzopyrone) is occurring in food, and is also used in cosmetics. In order to perform a risk assessment for both oral and dermal exposure, we applied a physiologically based approach to model kinetics in humans by simulating both routes of exposure. The concentration-time profile in liver revealed a higher peak concentration (C(max-hep)) for the oral when compared to the dermal route. The area under the concentration-time curve in the liver (AUC(hep)) was found the same for both routes if the same extent of absorption is assumed. Dose response information from published rat studies were used to identify the metric relevant for liver toxicity. Liver exposure levels resulting from doses and durations as outlined in the studies were simulated in a rat model. We obtained 31 data pairs of C(max-hep) and AUC(hep). Liver toxicity was observed at doses which resulted in simulated C(max-hep) values exceeding a certain liver concentration whereas we could not identify a clear cut off value of AUC(hep). Our findings support the notion that liver toxicity of coumarin in rats is related to C(max-hep) rather than to AUC(hep). If these findings can be transferred to the situation in humans, the result demonstrates that route specific differences in organ peak concentrations have to be considered when performing route-to-route extrapolation.

Skin permeation of polycyclic aromatic hydrocarbons: A solvent-based in vitro approach to assess dermal exposures against benzo[a]pyrene and dibenzopyrenes

ABSTRACT Consumer products with high contents of polycyclic aromatic hydrocarbons (PAHs) were repeatedly identified by market surveillance authorities. Since several of the individual compounds have been identified as genotoxic carcinogens, there might be health risks associated with the usage of these items. It therefore becomes reasonable to argue to reduce PAH contents in consumer products to a level as low as possible. This study presents data on the migration of PAHs from consumer products into aqueous sweat simulant or aqueous ethanol and on its combined migration and penetration into human skin. Product specimens were either submerged in simulant, or placed directly on test skins in Franz cell chambers to simulate dermal contacts. Migration of hexacyclic dibenzopyrenes became detectable by using ethanolic simulant, but not in aqueous sweat simulant. Similarly, migration of the pentacyclic model carcinogen benzo[a]pyrene (B[a]P) into aqueous sweat simulant was significantly lower when compared with human skin or skin models. The results point to a gross underestimation (about two orders of magnitude) when using aqueous sweat simulant instead of human skin for assessing PAH migration. On the other side, the usage of 20% ethanol as simulant revealed good agreement to the actual exposure of human skin against B[a]P migrating out of contaminated products. Our results underline that aqueous sweat simulant is not suitable to study dermal migration of highly lipophilic compounds.

CMR substances in consumer products: from food contact materials to toys

Good news first: toys must be safe! According to Directive 2009/48/EC on the safety of toys (Toy Safety Directive, TSD), substances classified as carcinogenic (Carc), mutagenic (Mut), or toxic for reproduction (Rep) category 1A, 1B, or 2 (so-called “CMR” substances), shall not be used in toys or components thereof. However, exemptions to this rule do exist. One of them stated that toy materials complying with the requirements laid down in Regulation (EC) No. 1935/2004 for Food Contact Materials (FCMs) are excluded from the prohibition, actually resulting in a higher level of protection (cf. below). However, this exemption expired on July 20, 2017. At present, only a few substances are listed in Appendix C to Annex II of the TSD and were thus specifically regulated. All remaining CMR substances, despite the ban, might be present in toys up to the maximum concentration to which they were classified as CMR according to CLP regulation (EC) No. 1272/2008 (EU 2008). For mutagenic or carcinogenic substances category 1A and 1B, this usually means a concentration of up to 0.1% (0.3% for those toxic to reproduction), while category 2 substances could be present at concentrations of up to 1% (Carc2 and Mut2) and 3% (Rep2), respectively. Noteworthy, these limit values were not derived based on toxicological risk assessments. Now, after July 20, 2017, these limits also apply to those materials for which the exemption based upon the requirements of the FCM Regulation was formerly valid. But what are the consequences? Did the exemption imply a higher safety level for the children in Europe in the recent past? Or was it, indeed, dispensable and thus okay not to extend it? Here, we try to address this issue by providing risk assessments of four exemplarily chosen substances.

Target Analysis of Polycyclic Aromatic Hydrocarbons (PAHs) in Consumer Products and Total Content of Polycyclic Aromatic Compounds (PACs)

ABSTRACT The fabrication of consumer products is tightly linked to the usage of polymeric materials. The application of mineral oil products as plasticizers or lubricants in this manufacture is very common and so is the addition of carbon black for coloring and abrasion resistance purposes. Whereas mineral oils often contain highly alkylated aromatic species, alkylation of polycyclic aromatic compounds (PACs) is rather low or even absent in the case of carbon black. Since they are broadly applied in various products, consumers might get exposed to PACs, for instance, via dermal contacts. Here, we characterized several commodities not only via the common way of targeted analysis for their contents on PAHs, but also with regard to its total amounts of unsubstituted and substituted aromatic hydrocarbons. The methods applied to analyze consumer products were as follows: (i) online coupling of liquid chromatography to gas chromatography with flame ionization detection (LC-GC-FID) for quantification of total PAH and mineral oil saturated hydrocarbon (MOSH) contents; (ii) coupling of tandem mass spectrometry to gas chromatography (GC-MS/MS) to quantify occasionally occurring PAHs with high specificity; and (iii) 2D gas chromatography coupled to time-of-flight mass spectrometry (GCxGC-MS) to characterize the complex mixtures concerning analytes’ ring systems and degrees of alkylation. We found PAH contents in the range of parts per million (ppm) in consumer products, while overall amounts of aromatics were in the percent range.

Safety limits for elements in toys: a comparison between the old and the new European toys safety directive

testing of toys for its compliance with legal provisions (EN 71-3:2002). Migration limits define the maximum release of a given element from a given toy material under defined testing conditions. As part of the revision of the TSD in the last decade, the safety limits for chemical elements in toys have been adapted (Directive 2009/48/EC). Based on the occurrence of adverse (health imparing) effects in animal experiments, the newly derived bioavailability limits were again translated into migration limits for the release of elements from toy materials, but this time assigned to three different kinds of materials separately. This revision of the legally binding safety limits has prompted an intense international discussion on whether the old bioavailability limits (and its subsequently derived migration limits) or the new migration limits, based on the onset of dose-dependent toxicity in animals, provide a higher level of health protection of children in terms of exposure against certain elements in toys. The present correspondence aims to contribute to the clarification of this controversial issue by evaluating the different approaches and calculation procedures, including the underlying assumptions and considerations by which the safety limits of the old and the new TSD were derived.