Congener patterns of polychlorinated dibenzo-p-dioxins, dibenzofurans and biphenyls as a useful aid to source identification during a contamination incident in the food chain

Polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs) and biphenyls (PCBs) are still considered among the most important groups of contaminants in the food chain. Self-control by food producers and official control by authorities are important activities that allow contaminant sources to be traced and promote further reduction in food and feed levels. Strict but feasible maximum levels were set by the EU Commission for food and feed to support this strategy, as well as action levels and thresholds. When products exceed these levels, it is important to trace the source of contamination and take measures to remove it. Congener patterns of PCDD/Fs and PCBs differ between sources and are important tools for source identification. Therefore, patterns associated with different sources and incidents relating to various feed matrices and certain agricultural chemicals were collated from published scientific papers, with additional ones available from some laboratories. The collection was evaluated for completeness by presentations at workshops and conferences. Primary sources appear to derive from 5 categories, i) by-products from production of organochlorine chemicals (e.g. PCBs, chlorophenols, chlorinated pesticides, polyvinyl chloride (PVC)), ii) the result of combustion of certain materials and accidental fires, iii) the use of inorganic chlorine, iv) recycling/production of certain minerals, and v) certain naturally occurring clays (ball clay, kaolinite). A decision tree was developed to assist in the identification of the source

Non-animal approaches for toxicokinetics in risk evaluations of food chemicals

The objective of the present work was to review the availability and predictive value of non-animal toxicokinetic approaches and to evaluate their current use in European risk evaluations of food contaminants, additives and food contact materials, as well as pesticides and medicines. Results revealed little use of quantitative animal or human kinetic data in risk evaluations of food chemicals, compared with pesticides and medicines. Risk evaluations of medicines provided sufficient in vivo kinetic data from different species to evaluate the predictive value of animal kinetic data for humans. These data showed a relatively poor correlation between the in vivo bioavailability in rats and dogs versus that in humans. In contrast, in vitro (human) kinetic data have been demonstrated to provide adequate predictions of the fate of compounds in humans, using appropriate in vitro-in vivo scalers and by integration of in vitro kinetic data with in silico kinetic modelling. Even though in vitro kinetic data were found to be occasionally included within risk evaluations of food chemicals, particularly results from Caco-2 absorption experiments and in vitro data on gut-microbial conversions, only minor use of in vitro methods for metabolism and quantitative in vitro-in vivo extrapolation methods was identified. Yet, such quantitative predictions are essential in the development of alternatives to animal testing as well as to increase human relevance of toxicological risk evaluations. Future research should aim at further improving and validating quantitative alternative methods for kinetics, thereby increasing regulatory acceptance of non-animal kinetic data

Toxicokinetic Modeling of the Transfer of Non-Dioxin-like Polychlorinated Biphenyls (ndl-PCBs) from Feed and Soil into Laying Hens and Their Eggs

Non-dioxin-like polychlorinated biphenyls (ndl-PCBs) are a subclass of persistent bioaccumulative pollutants able to enter the food chain. Toxicokinetic models for the transfer of the six ndl-PCB congeners (PCBs 28, 52, 101, 138, 153, and 180) from contaminated feed and soil into chicken eggs and meat are presented. Three independent controlled feeding study datasets were used to estimate the model parameters and four studies for evaluating the model performance. The yolk deposition of ndl-PCBs is modeled in a novel way that mimics the physiology of yolk growth and development, resulting in improved prediction of the experimental data without introducing an ad hoc time delay between ovulation and oviposition. Using the models, the highest level of 2.4 μg/kg dry matter (DM) was calculated for the sum of ndl-PCBs in laying hen feed to ensure that the current maximum levels in meat and eggs (40 ng/g fat) will not be exceeded. It is also shown how this highest level in feed should be adapted in case soil, in addition to feed, is also a source of ndl-PCBs for free-range chickens

PDE-5 inhibitors in selected herbal supplements from the Ghanaian market for better erectile function as tested by a bioassay

Herbal supplements sold as ‘all natural’ on various markets in Accra (Ghana) and advertised as highly efficacious in treating erectile dysfunction (ED) were bought and analysed by a PDE-5 enzyme inhibition assay. The claimed efficacy of these products could be the result of inherent plant constituents, but also of intentionally added pharmaceuticals. Medically, ED is treated with potent inhibitors of the phosphodiesterase-5 (PDE-5) enzyme, as in the case of sildenafil. To test the efficacy of the Ghanaian supplements, extracts were made and tested using a PDE-Glo phosphodiesterase assay, a luminescent high-throughput screening (HTS) method. Results revealed that about 90% of the selected samples were able to inhibit PDE-5 activity to a high extent. Estimated concentrations in sildenafil equivalents ranged from traces to very high, with 25 samples (62.5%) pointing at daily doses higher than 25 mg sildenafil equivalents and 9 (22.5%) of these at doses higher than the maximal recommended daily intake of 100 mg sildenafil equivalents. Further investigations are needed to confirm if the observed effects are due to inherent plant constituents or merely the result of added synthetic PDE-5 enzyme inhibitors, especially because doses above 100 mg sildenafil equivalents per day may result in severe health risks.</p

Acid condensation products of indole-3-carbinol and their in-vitro (anti)estrogenic, (anti)androgenic and aryl hydrocarbon receptor activities

The objective of the study was to investigate the (anti)estrogenic, (anti)androgenic and aryl hydrocarbon receptor (AhR) agonistic activities of a mixture of acid condensation products of indole-3-carbinol, termed RXM, and to identify the compounds most responsible for the observed effects, using in vitro receptor-reporter gene transcriptional activation bioassays. For this, HPLC-fractions of RXM were prepared and tested. LC-MS/MS analysis was carried out for the identification of some of the acid condensation products. The RXM displayed weak estrogenic and anti-androgenic, and strong AhR agonistic properties. The fraction containing 3,3-diindolylmethane (DIM) displayed a weak estrogenic and relatively strong anti-androgenic activity. DIM was confirmed to be an androgen receptor (hAR) antagonist and a partial estrogen receptor (hERα) agonist. Also the fraction containing the trimer [2-(indol-3-ylmethyl)indol-3-yl]indol-3-ylmethane (LTr1) showed anti-androgenic activities. It was shown for the first time that DIM is not only estrogenic and anti-androgenic, but also possesses anti-estrogenic properties. Though indolo[3,2-b]carbazole (ICZ) is a potent AhR activator and was detected in the RXM, it did not contribute to AhR-agonist activity. Instead, fractions containing the trimers LTr1 and 5,6,11,12,17,18-hexahydrocyclonona[1,2-b:4,5-b′:7,8-b″]tri-indole (CTr), as well as some unidentified compounds showed the highest AhR activation. The fraction, containing the linear trimer LTr1, showed a weak anti-androgenic activity which has not been reported before. The study demonstrates the importance of a bioassay directed approach for identifying compounds that contribute most to the effects of mixtures.</p

Fate of pyrrolizidine alkaloids during processing of milk of cows treated with ragwort

To investigate the fate of pyrrolizidine alkaloids (PAs) during milk processing, milk of cows treated via rumen fistula with a mixture of 84% (w/w) ragwort (Jacobaea vulgaris, syn. Senecio jacobaea) and 16% narrow-leaved ragwort (Senecio inaequidens) was processed using laboratory scale heating systems with industrial settings. Pasteurised and sterilised (UHT) milk were produced, as well as set-type yoghurt and cheese. Samples were analysed for 29 PAs using LC-MS/MS, of which 11 PAs were detected above LOQ in the samples (0.1 µg l−1). Alterations in the PA concentration and composition between the standardised milk and the corresponding end-product(s) were evaluated. The heat treatments applied for pasteurisation and UHT sterilisation to prepare semi-skimmed consumption milk did not affect the PA levels in the end-products. In yoghurt, after fermentation of standardised milk (6 h, pH 4.4), 73% of total PAs were recovered. The PA concentration, specifically dehydrojacoline, was decreased, although not quantifiable, during cheese production. A further decrease of 38% during 6 weeks of ripening was observed. The results show that the PA concentration of natural contaminated cow’s milk is not affected by heat treatment applied for pasteurised and sterilised milk, but that microbial fermentation of the milk leads to a lowered PA concentration in yoghurt and cheese. This is probably due to microbiological degradation, since PAs are fairly stable under acidic conditions

Food and feed safety : Cases and approaches to identify the responsible toxins and toxicants

There are food and feed safety monitoring programs to protect consumers. These programs however, are strongly focused on known and regulated substances. New or unexpected substances that might be of risk for consumers will thus escape routine controls. These risks are therefore mainly discovered by human or animal intoxications. All kind of analytical chemical methods, in-vitro bioassays, tracking, and chain analysis are then used to reveal the substance(s) responsible for the intoxication. Only in a few occasions (new) risks were revealed in time by analytical chemical methods or cell based in-vitro bioassays. This paper describes some relevant food and feed safety cases and how the causative substances were identified. This overview strongly indicates that more intense monitoring, including the use of cell based effect bioassays, can reduce the number of intoxications. Moreover, registration and follow-up actions should be arranged in a better way, for example by sharing information within the scientific communities or by establishing a national contact point. In addition, a strategy based on broad screening and bioassay directed identification with liquid chromatography high resolution mass spectrometry is proposed to prevent intoxications and identify toxin and toxicants relevant for food and feed safety

Food and feed safety : Cases and approaches to identify the responsible toxins and toxicants

There are food and feed safety monitoring programs to protect consumers. These programs however, are strongly focused on known and regulated substances. New or unexpected substances that might be of risk for consumers will thus escape routine controls. These risks are therefore mainly discovered by human or animal intoxications. All kind of analytical chemical methods, in-vitro bioassays, tracking, and chain analysis are then used to reveal the substance(s) responsible for the intoxication. Only in a few occasions (new) risks were revealed in time by analytical chemical methods or cell based in-vitro bioassays. This paper describes some relevant food and feed safety cases and how the causative substances were identified. This overview strongly indicates that more intense monitoring, including the use of cell based effect bioassays, can reduce the number of intoxications. Moreover, registration and follow-up actions should be arranged in a better way, for example by sharing information within the scientific communities or by establishing a national contact point. In addition, a strategy based on broad screening and bioassay directed identification with liquid chromatography high resolution mass spectrometry is proposed to prevent intoxications and identify toxin and toxicants relevant for food and feed safety.</p

Presence and risks of polycyclic aromatic hydrocarbons, dioxins and dioxin-like PCBs in dietary plant supplements as elucidated by a combined DR CALUX® bioassay and GC-HRMS based approach

Plant-based dietary supplements may contain undesirable contaminants such as polycyclic aromatic hydrocarbons, dioxins and dioxin-like polychlorinated biphenyls (dl-PCBs) due to the sources of raw materials or processing methods used. The presence of these contaminants in a series of herbal supplements sold on the Ghanaian market for improving sexual performance was examined using the DR CALUX® bioassay in combination with GC-HRMS analysis. Overall, cell responses at 4 and 48 h exposure to extracts prepared without an acid-silica clean-up were relatively higher than the responses obtained from extracts prepared with an acid-silica clean-up. This indicated that the 40 supplements contained only low levels of stable aryl hydrocarbon receptor (AhR) agonists like polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) and dl-PCBs, while some contained substantial amounts of less stable AhR-agonists. Ten supplements selected for confirmation with GC-HRMS analysis contained PCDD/Fs and dl-PCBs at levels ranging from 0.01 to 0.19 pg toxic equivalent (TEQ)/g only, while the level of the sum of 4 polycyclic aromatic hydrocarbons (Σ4PAHs) representing less stable AhR agonists, ranged from not detected (ND) to 25.5 ng/g. These concentrations were in line with the responses observed in the DR CALUX® bioassay. The concentration of PCDD/Fs and dl-PCBs corresponded to estimated daily intakes (EDIs) ranging from 0.01 to 1.20 pg TEQ/day, or 0.001 to 0.12 pg TEQ/kg bw/week for a 70 kg bw consumer, which was below the established tolerable weekly intake (TWI) of 2 pg TEQ/kg bw/week, thus indicating low concern for consumers’ health. Similarly, the EDIs based on the detected Σ4PAHs in supplements ranged from 7.2 to 111 ng/day, or 0.1 to 1.6 ng/kg bw/day, which corresponded to MOE values above 10,000, indicating a low health concern

High levels of dioxins and PCBs in meat, fat and livers of free ranging pigs, goats, sheep and cows from the island of Curaçao

Samples of adipose tissue, meat and livers from pigs, cows, sheep and goats from Curaçao were analysed for polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), and dioxin-like (dl-) and non-dioxin-like (ndl-) PCBs (polychlorinated biphenyls). Levels in many samples of adipose tissue were higher than the EU maximum levels (MLs) for PCDD/Fs and the sum of PCDD/Fs and dl-PCBs (sum-TEQ), indicating unusually high levels. Median sum-TEQ (Toxic Equivalents) levels for pigs, cows, sheep and goats were 0.9 (range 0.3–35), 3.0 (0.5–14), 5.7 (0.3–28) and 6.5 (0.5–134) pg TEQ g−1 fat. For most samples, the congener pattern pointed to the burning of waste as the major source, in line with the fact that most animals forage outside. MLs for ndl-PCBs were also exceeded in some of the samples, indicating that some areas are additionally contaminated with PCBs. Meat levels showed similar lipid based levels as adipose tissue, contrary to liver levels, which were much higher in most animals. Pigs showed liver sequestration at lower levels in adipose tissue than the ruminants. The relatively high levels observed in this study are likely to result in high exposure of consumers and measures were taken to reduce the contamination of areas where animals forage.</p