Cottage industry as a source of high exposure to lead: A biomonitoring study among people involved in manufacturing cookware from scrap metal.

In low-income countries, a widespread but poorly studied type of cottage industry consists of melting scrap metal for making cookware. We assessed the exposure to lead (Pb) among artisanal workers, and their families, involved in manufacturing cookware from scrap metal. In a cross-sectional survey, we compared artisanal cookware manufacturing foundries with carpentry workshops (negative controls) and car battery repair workshops (positive controls), all located in residential areas, in Lubumbashi (DR Congo). We collected surface dust in the workspaces, and blood and urine samples among workers, as well as residents living in the cookware workshops. Trace elements were quantified in the samples by Inductively Coupled Plasma Mass Spectrometry (ICP-MS). In surface dust, median Pb concentrations were higher in cookware foundries (347 mg/kg) than in carpentries (234 mg/kg) but lower than in battery repair workshops (22,000 mg/kg). In workers making the cookware (n = 24), geometric mean (GM) Pb blood cencentration was 118 µg/L [interquartile range (IQR) 78.4-204], i.e. nearly twice as high as among carpenters [60.2 µg/L (44.4-84.7), n = 33], and half the concentration of battery repair workers [255 µg/L (197-362), n = 23]. Resident children from the cookware foundries, had higher urinary Pb [6.2 µg/g creatinine (2.3-19.3), n = 6] than adults [2.3 (2.2-2.5), n = 3]. Our investigation confirms the high Pb hazard linked to car battery repair and reveals a high exposure to Pb among artisanal cookware manufacturers and their families, especially children, in residential areas of a city in a low-income country.

The use of nitrous oxide whippets as a recreational drug: Hidden health risks.

Whipped cream canisters, also known as nitrous oxide whippets, are traditionally used in the culinary arts to prepare food foams. In recent years, however, these gas canisters have been cracked open and inhaled to produce a "legal" high. Users of these whippets have reported the presence of an oily residue containing metallic particles. This contamination was investigated using liquid chromatography-, gas chromatography- and inductively coupled plasma-mass spectrometry (ICP-MS) and optical emission spectrometry (ICP-OES). The particulate matter was also analyzed by scanning transmission electron microscopy (STEM) combined with energy-dispersive X-ray spectroscopy (EDX). The presence of cyclohexyl isothiocyanate was confirmed at a maximum concentration of 67 µg per whippet. ICP-MS and ICP-OES analysis revealed the presence of mainly iron and zinc, but also, traces of aluminum, chromium, cobalt, nickel, and lead were found. STEM-EDX analysis confirmed the presence of nano-sized particles containing iron and zinc. When simulating inhalation, using the multiple path particle dosimetry model, it was confirmed that these nano-sized particles can reach the deeper parts of the lungs. Most users assume that inhaling a food-grade nitrous oxide whippet for a "legal" high poses no risks. However, this research shows that users are exposed to cyclohexyl isothiocyanate, a substance classified as a respiratory sensitizer. The presence of zinc in the particulate matter could potentially be linked to lung lesions.

Antidepressant-like effects of oxytocin in mice are dependent on the presence of insulin-regulated aminopeptidase.

Oxytocin is a neuromodulator with antidepressant-like effects. In vitro, oxytocin is rapidly cleaved by insulin-regulated aminopeptidase (IRAP). Oxytocin metabolites are known to exert strong central activities that are different from the effects of the parent molecule. Our goal is to investigate in vivo whether IRAP deletion modifies the antidepressant-like effects of oxytocin. Male and female C57Bl/6 mice, IRAP wild-type (IRAP(+/+)) and knock-out (IRAP(-/-)) mice were injected subcutaneously with saline, oxytocin or oxytocin combined with angiotensin IV. One hour after injection, immobility was timed during a 5 min forced swim that was preceded by an open field to study locomotor behaviour. Oxytocin induced antidepressant-like effects in male (0.25 mg/kg oxytocin) and female (0.15 mg/kg oxytocin) C57Bl/6 mice subjected to the forced swim test. Oxytocin did not influence locomotor behaviour in mice, as shown with the open field. These findings were reproduced in transgenic male (aged 3-6 months) and female (aged 12-18 months) IRAP(+/+) mice. However, the major findings of our study were that the antidepressant-like effect was reversed in angiotensin IV treated IRAP(+/+) mice and was completely absent in age- and gender-matched IRAP(-/-) mice. The lack of an antidepressant-like effect of oxytocin in young male and middle-aged female IRAP(-/-) mice attributes an important role to IRAP in mediating this effect.

Conformational constraints in angiotensin IV to probe the role of Tyr², Pro5 and Phe6.

The aromatic amino acids Tyr and Phe in angiotensin IV (Ang IV) were conformationally constrained by the use of ß-Me substituted analogs, or cyclic constrained analogs. None of these modifications was allowed for Tyr¹, while only e-ß-MePhe6 substitution resulted in an AngIV analog with high IRAP potency and selectivity versus AP-N or the AT1 receptor. This indicates an important role of the orientation of the Phe6 for inducing selectivity. Pro5 replacement with 2-aminocyclopentanecarboxylic acid maintained IRAP potency and abolished AT1 affinity. These results confirm the importance of conformational constrained amino acids to generate selectivity in bioactive peptides.

Intake of food supplements based on algae or cyanobacteria may pose a health risk due to elevated concentrations of arsenic species.

Despite the health benefits of food supplements (FS) based on algae or cyanobacteria, the elevated arsenic (As) concentrations in these FS may raise a health concern. In the present study 33 FS containing algae or cyanobacteria were collected and As (species) were analysed to estimate consumer exposure. Based on hazard and exposure data, potential risks were evaluated using inorganic arsenic (Asi) and the potentially toxic As fraction (Astot minus arsenobetaine (AB)). Astot concentrations were in the range 0.053-57 mg/kg with highest concentrations in FS containing brown algae. Asi concentrations were in the range <0.02-4.7 mg kg-1. A large part of As in FS containing algae or cyanobacteria was identified as potentially toxic AsSugars species. Negligible amounts of AB were detected. According to a tentative risk evaluation, the intake of Asi related to all FS collected was of no health concern for the general population. In 8 out of 33 of the analysed FS, however, the Asi concentration was of concern for population groups with increased cancer risks. If all As species except the non-toxic AB were taken into consideration, only 26 out of 33 of the FS showed 'no concern' for the general population, while for the other 7 FS a potential health risk was identified. This study indicates the need to obtain more data on toxicity of AsSugars and to develop limits for As (species) in FS.

Does arsenic pose a health concern after consumption of clay products?

Clay products for oral use form a particular group of food supplements in relation to potential arsenic (As) toxicity, because - certainly in case of pure clay- all arsenic in these supplements is expected to be present in the most toxic inorganic form (Asi). In terms of risk, the most important questions to answer relate to the bioaccessibility and bioavailability of the inorganic arsenic present, rather than to the As species distribution, which often receives most attention in standard foodstuffs. In the present study, clay products for oral use were bought on the Belgian market and analysed for total arsenic (Astot), arsenic species (Asi, arsenobetaine, dimethylarsenate and monomethylarsenate)) and bioaccessible arsenic, in order to perform an exposure assessment and risk characterisation. Total As concentrations differed considerably between the samples and ranged from 0.20 to 6.4 mg Astot/kg. Bioaccessibility of Asi, determined via the Unified Barge Method (extraction making use of digestive enzymes) varied between 8% and 51%. The Asi concentration determined via HPLC-ICP-MS after extraction with diluted HNO3 + H2O2 (as in the CEN method for foodstuffs) was only a poor predictor of the bioaccessible Asi fraction, despite the significant relationship (R2 = 0.36; p < .05). The risk characterisation did not reveal acute risks related to Asi exposure. However, a potential concern with regard to chronic Asi intake was identified for the general population in 42% of the analysed food supplements, and for sensitive population groups in 67% of the samples, even after taking into account the bioaccessible fraction. The data presented illustrate that consumption of some of these clay products may contribute significantly to dietary Asi intake and that these should not be taken chronically.

Assessment of hazards and risks associated with dietary exposure to mineral oil for the Belgian population.

Recently collected dietary exposure data on mineral oil saturated (MOSH) and aromatic (MOAH) hydrocarbons were used to evaluate the risks associated with exposure to mineral oil through food for the Belgian population. For MOSH, the no observed adverse effect level (NOAEL) value of 19 mg kg-1 bw day-1 based on the hepatic inflammation-associated granulomas found in a 90-day oral study in F-344 rats was used as point of departure (PoD). Due to existing toxicological uncertainties, the margin of exposure (MOE) approach was applied. In all investigated scenarios, the MOE values were well above 100, indicating that there is no direct health concern related to MOSH exposure for the Belgian population. Nevertheless, more appropriate risk assessment approaches for MOSH based on adequate PoD are needed. For dietary exposure to MOAH, which are potentially genotoxic and carcinogenic, no MOE values could be calculated due to the lack of adequate dose-response carcinogenicity data. In two investigated worst-case scenarios, a health concern related to MOAH exposure could not be excluded, highlighting that more data are needed to perform an adequate risk assessment. The possibility to use in vitro bioassays to collect such additional toxicological information for MOAH present in food samples was also investigated.

Involvement of the AT1 receptor subtype in the effects of angiotensin IV and LVV-haemorphin 7 on hippocampal neurotransmitter levels and spatial working memory.

Intracerebroventricular (i.c.v.) administration of angiotensin IV (Ang IV) or Leu-Val-Val-haemorphin 7 (LVV-H7) improves memory performance in normal rats and reverses memory deficits in rat models for cognitive impairment. These memory effects were believed to be mediated via the putative 'AT4 receptor'. However, this binding site was identified as insulin-regulated aminopeptidase (IRAP). Correspondingly, Ang IV and LVV-H7 were characterised as IRAP inhibitors. This study investigates whether and how IRAP may be involved in the central effects of Ang IV and LVV-H7. We determined the effects of i.c.v. administration of Ang IV or LVV-H7 on hippocampal neurotransmitter levels using microdialysis in rats. We observed that Ang IV modulates hippocampal acetylcholine levels, whereas LVV-H7 does not. This discrepancy was reflected in the observation that Ang IV binds with micromolar affinity to the AT1 receptor whereas no binding affinity was observed for LVV-H7. Correspondingly, we demonstrated that the AT1 receptor is involved in the effects of Ang IV on hippocampal neurotransmitter levels and on spatial working memory in a plus maze spontaneous alternation task. However, the AT1 receptor was not involved in the spatial memory facilitating effect of LVV-H7. Finally, we demonstrated that Ang IV did not diffuse to the hippocampus following i.c.v. injection, suggesting an extrahippocampal site of action. We propose that AT1 receptors are implicated in the neurochemical and cognitive effects of Ang IV, whereas LVV-H7 may mediate its effects via IRAP.

Brain and peripheral angiotensin II type 1 receptors mediate renal vasoconstrictor and blood pressure responses to angiotensin IV in the rat.

OBJECTIVES: Angiotensin (Ang) IV was reported to increase renal cortical blood flow (CBF) via putative angiotensin IV receptor (AT4) stimulation but reduce total renal blood flow (RBF) via angiotensin II type 1 (AT1) receptors. We investigated the effect of Ang IV on simultaneously measured mean arterial pressure (MAP), RBF, and CBF. The possible involvement of AT1 or AT4 receptors, the possible natriuretic effect, and responses to central administration were also explored. METHODS AND RESULTS: Intravenous injections of Ang IV dose dependently increased MAP and decreased CBF and RBF; these effects were abolished by AT1 receptor blockade. These reductions in CBF and RBF highly correlated as did renal vascular responses to Ang II and fenoldopam. Ang IV did not induce renal vasodilation even following AT1 receptor blockade. Intrarenal Ang IV infusion reduced CBF and RBF but had no natriuretic effect. Central Ang IV administration induced an AT1-mediated immediate increase in MAP and renal vascular resistance and a secondary increase in RBF. AT4 selective ligands, LVV-hemorphin-7 and AT4-16 (intravenous, intrarenal or intracerebroventricular), had no effects on MAP, RBF or urinary sodium excretion. Additional in-vitro experiments indicated that the majority of the Ang IV-sensitive aminopeptidase activity in kidney membranes is attributed to aminopeptidase-N. CONCLUSION: Insulin-regulated aminopeptidase (IRAP)/AT4 receptors are involved in neither the regulation of RBF or CBF nor in the handling of renal sodium. Ang IV increases MAP and induces renal vasoconstriction via stimulation of brain and peripheral AT1 receptors and may be involved in the regulation of renal blood flow and blood pressure.

Beta-homo-amino acid scan of angiotensin IV.

Angiotensin IV, a metabolite of angiotensin II, inhibits the enzyme insulin regulated aminopeptidase or IRAP and also, although with lower potency, aminopeptidase-N (AP-N). When both beta (2)-homo amino acid- and beta (3)-homo amino acid substitutions were used, allowed the identification of H-( R)beta (2)hVal-Tyr-Ile-His-Pro-beta (3)hPhe-OH as a potent and stable Ang IV analog with high selectivity for IRAP versus AP-N and the AT1 receptor.

Ligands to the (IRAP)/AT4 receptor encompassing a 4-hydroxydiphenylmethane scaffold replacing Tyr2.

Analogues of the hexapeptide angiotensin IV (Ang IV, Val(1)-Tyr(2)-Ile(3)-His(4)-Pro(5)-Phe(6)) encompassing a 4-hydroxydiphenylmethane scaffold replacing Tyr(2) and a phenylacetic or benzoic acid moiety replacing His(4)-Pro(5)-Phe(6) have been synthesized and evaluated in biological assays. The analogues inhibited the proteolytic activity of cystinyl aminopeptidase (CAP), frequently referred to as the insulin-regulated aminopeptidase (IRAP), and were found less efficient as inhibitors of aminopeptidase N (AP-N). The best Ang IV mimetics in the series were approximately 20 times less potent than Ang IV as IRAP inhibitors. Furthermore, it was found that the ligands at best exhibited a 140 times lower binding affinity to the membrane-bound IRAP/AT4 receptor than Ang IV. Although the best compounds still exert lower activities than Ang IV, it is notable that these compounds comprise only two amino acid residues and are considerably less peptidic in character than the majority of the Ang IV analogues previously reported as IRAP inhibitors in the literature.

Involvement of insulin-regulated aminopeptidase and/or aminopeptidase N in the angiotensin IV-induced effect on dopamine release in the striatum of the rat.

Locally administered angiotensin IV causes a dose-dependent increase of the dopamine levels in the striatum of the rat. The aminopeptidases insulin-regulated aminopeptidase (IRAP) and/or aminopeptidase N (AP-N) are proposed to be involved in this effect since both enzymes are inhibited by angiotensin IV. In agreement with this hypothesis we demonstrate that by using the AP-N selective inhibitor 7B, about 60% of the aminopeptidase activity in striatal membranes could be attributed to AP-N (pK(i)=9.20). Higher concentrations of 7B are capable of inhibiting IRAP as well (pK(i)=7.26). Interestingly, in vivo, inhibition of IRAP or AP-N activity does not appear to be involved in the angiotensin IV-mediated effect in the striatum since 7B itself is not capable to induce dopamine release such as observed with angiotensin IV. However, 7B at a concentration selective for inhibition of AP-N (100 nM) potentiates the angiotensin IV-mediated increase of dopamine, suggesting that inhibition of AP-N lengthens the half-life of angiotensin IV. On the other hand, inhibition of both AP-N and IRAP by perfusion of 500 nM 7B completely abolishes the effect of angiotensin IV. We therefore hypothesize that the effect of angiotensin IV on dopamine release in the striatum is mediated via activation of IRAP and/or AP-N, possibly acting as receptors for angiotensin IV.

Angiotensin IV activates the nuclear transcription factor-kappaB and related proinflammatory genes in vascular smooth muscle cells.

Inflammation is a key event in the development of atherosclerosis. Nuclear factor-kappaB (NF-kappaB) is important in the inflammatory response regulation. The effector peptide of the renin angiotensin system Angiotensin II (Ang II) activates NF-kappaB and upregulates some related proinflammatory genes. Our aim was to investigate whether other angiotensin-related peptides, as the N-terminal degradation peptide Ang IV, could regulate proinflammatory factors (activation of NF-kappaB and related genes) in cultured vascular smooth muscle cells (VSMCs). In these cells, Ang IV increased NF-kappaB DNA binding activity, caused nuclear translocation of p50/p65 subunits, cytosolic IkappaB degradation and induced NF-kappaB-dependent gene transcription. Ang II activates NF-kappaB via AT1 and AT2 receptors, but AT1 or AT2 antagonists did not inhibit NF-kappaB activation caused by Ang IV. In VSMC from AT1a receptor knockout mice, Ang IV also activated NF-kappaB pathway. In those cells, the AT4 antagonist divalinal diminished dose-dependently Ang IV-induced NF-kappaB activation and prevented IkappaB degradation, but had no effect on the Ang II response, indicating that Ang IV activates the NF-kappaB pathway via AT4 receptors. Ang IV also increased the expression of proinflammatory factors under NF-kappaB control, such as MCP-1, IL-6, TNF-alpha, ICAM-1, and PAI-1, which were blocked by the AT4 antagonist. Our results reveal that Ang IV, via AT4 receptors, activates NF-kappaB pathway and increases proinflammatory genes. These data indicate that Ang IV possesses proinflammatory properties, suggesting that this Ang degradation peptide could participate in the pathogenesis of cardiovascular diseases.

Angiotensin AT4 receptor ligand interaction with cystinyl aminopeptidase and aminopeptidase N: [125I]Angiotensin IV only binds to the cystinyl aminopeptidase apo-enzyme.

Due to its high affinity for [(125)I]Angiotensin IV, cystinyl aminopeptidase (CAP) has recently been assigned as the 'angiotensin AT(4) receptor'. Since the aminopeptidase N (AP-N) activity is also susceptible to inhibition by Angiotensin IV, it might represent an additional target for this peptide. Based on [(125)I]Angiotensin IV binding and catalytic activity measurements, we compared the ligand interaction properties of recombinant human CAP and human AP-N. Both enzymes displayed distinct pharmacological profiles. Although their activity is inhibited by Angiotensin IV and LVV-hemorphin 7, both peptides are more potent CAP-inhibitors. On the other hand, substance P and l-methionine have a higher potency for AP-N. High affinity binding of [(125)I]Angiotensin IV to CAP occurs in the presence of chelators but not to AP-N in either the absence or presence of chelators. These differences were exploited to determine whether CAP and/or AP-N are present in different cell lines (CHO-K1, COS-7, HEK293, SK-N-MC and MDBK). We provide evidence that CAP predominates in these cell lines and that, comparatively, CHO-K1 cells display the highest level of this enzyme.

Synergistic inhibition of the enzymatic activity of aminopeptidase N by divalent metal ion chelators.

Membranes of HEK293 cells that were transfected with human aminopeptidase N (AP-N, CD13, EC 3.4.11.2) and purified soluble porcine kidney AP-N were used to study inhibition of its enzyme activity by divalent cation chelators. Whereas pre-incubation for 10 min with ethylenediaminetetraacetic acid (EDTA), did not or only weakly affected the enzyme activity, the bidentate chelator 1,10-phenanthroline produced a complete and concentration-dependent inhibition of AP-N. The corresponding curves had Hill slopes of 2.50 +/- 0.23 and 2.73 +/- 0.01 for soluble and recombinant AP-N respectively. EDTA increased the potency of 1,10-phenanthroline till a limit, at which Hill slopes became close to unity. In the absence of EDTA, the inhibition by 1,10-phenanthroline was only weakly affected by the substrate concentration. On the other hand, competition between 1,10-phenanthroline and the substrate took place in the presence of EDTA. Similar findings were reported for the related metallopeptidase cystinyl aminopeptidase and point towards a model in which 1,10-phenanthroline inhibit enzyme activity by decreasing the free Zn2+ concentration. Moreover, EDTA is capable of removing a modulatory ion from an allosteric site at the enzyme, facilitating the direct interaction between 1,10-phenanthroline and the catalytic Zn2+. Compatible with this model, Ca2+ may bind to this allosteric site resulting in the potentiation of Zn2+-mediated re-activation of the enzyme activity in the presence of EDTA and 1,10-phenanthroline.

Metal ion modulation of cystinyl aminopeptidase.

Cystinyl aminopeptidase has one Zn2+-binding motif and is a member of the M1 aminopeptidase family. Ion modulation of its catalytic activity was studied in membranes of CHO-K1 cells (Chinese-hamster ovary K1 cells) using L-leucine-p-nitroanilide as substrate. The planar bidentate chelators 1,10-phenanthroline and 2,2'-bipyridine inhibited the activity in a concentration-dependent manner with Hill slopes of 3.32+/-1.78 and 2.10+/-0.26 respectively. The acetic acid-containing chelators EDTA, EGTA and DTPA (diethylenetriamine-N,N,N',N'',N''-penta-acetic acid) weakly affected the activity, but they increased the potency of the planar chelators up to a limit, at which Hill slopes became close to unity. Moreover, competition between 1,10-phenanthroline and the substrate only took place in the presence of EDTA. These findings are compatible with a model in which the bidentate chelators inhibit enzyme activity by decreasing the free Zn2+ concentration. By removing a modulatory ion from an allosteric site at the enzyme, the acetic acid-containing chelators facilitate the direct interaction between the bidentate chelators and the catalytic Zn2+. The inhibitory effect of EDTA plus 1,10-phenanthroline could be completely reversed by Zn2+. Ca2+ and Mg2+ increased the potency of Zn2+ for this process. This is expected if they interact with the modulatory site to decrease the sensitivity of the enzyme towards 1,10-phenanthroline. Conversely, the bidendate chelators increased the high-affinity [125I]angiotensin IV binding to the membranes and this was potentiated by the acetic acid-containing chelators. These findings support the concept that high-affinity [125I]angiotensin IV binding, previously referred to as 'AT4 receptor binding', only occurs for the cystinyl aminopeptidase apoenzyme.

Endocrine activity of mycotoxins and mycotoxin mixtures.

Reporter gene assays incorporating nuclear receptors (estrogen, androgen, thyroid ß and PPARγ2) have been implemented to assess the endocrine activity of 13 mycotoxins and their mixtures. As expected, zearalenone and its metabolites α-zearalenol and ß- zearalenol turned out to have the strongest estrogenic potency (EC50 8,7 10-10 ± 0,8; 3,1 10-11 ± 0,5 and 1,3 10-8 ± 0,3 M respectively). The metabolite of deoxynivalenol, 3-acetyl-deoxynivalenol also had estrogenic activity (EC50 3,8 10-7 ± 1,1 M). Furthermore, most of the mycotoxins (and their mixtures) showed anti-androgenic effects (15-acetyldeoxynivalenol, 3-acetyl-deoxynivalenol and α-zearalenol with potencies within one order of magnitude of that of the reference compound flutamide). In particular, deoxynivalenol and 15-acetyl-deoxynivalenol acted as antagonists for the PPARy2 receptor. When testing mixtures of mycotoxins on the same cell systems, we showed that most of the mixtures reacted as predicted by the concentration addition (CA) theory. Generally, the CA was within the 95% confidence interval of the observed ones, only minor deviations were detected. Although these reporter gene tests cannot be directly extrapolated in vivo, they can be the basis for further research. Especially the additive effects of ZEN and its metabolites are of importance and could have repercussions in vivo.

Investigation of the genotoxicity of substances migrating from polycarbonate replacement baby bottles to identify chemicals of high concern.

Due to the worldwide concern that bisphenol A might act as an endocrine disruptor, alternative materials for polycarbonate (PC) have been introduced on the European market. However, PC-replacement products might also release substances of which the toxicological profile--including their genotoxic effects--has not yet been characterized. Because a thorough characterization of the genotoxic profile of all these substances is impossible in the short term, a strategy was developed in order to prioritize those substances for which additional data are urgently needed. The strategy consisted of a decision tree using hazard information related to genotoxicity. The relevant information was obtained from the database of the European Chemicals Agency (ECHA), in silico prediction tools (ToxTree and Derek Nexus(TM)) and the in vitro Vitotox(®) test for detecting DNA damage. By applying the decision tree, substances could be classified into different groups, each characterized by a different probability to induce genotoxic effects. Although none of the investigated substances could be unequivocally identified as genotoxic, the presence of genotoxic effects could neither be excluded for any of them. Consequently, all substances require more data to investigate the genotoxic potential. However, the type and the urge for these data differs among the substances.

Evaluation of the potential health risks of substances migrating from polycarbonate replacement baby bottles.

Since the European Commission prohibited the use of bisphenol A in the production of polycarbonate (PC) baby bottles, many other materials have replaced PC for the manufacture of this type of food contact materials. In the present study, the potential migration risks associated with these alternative materials were investigated. First, all substances were evaluated for endocrine disruptive (ED) activity by using different existing lists of (suspected) ED chemicals. Next, the potential non-ED risks were assessed. A distinction was made between migrants listed in Annex I of European Regulation 10/2011 and the unlisted substances (e.g. non-intentionally added substances). For the listed substances, concentrations in the migration solutions were compared to their respective specific migration limits (SML) (when applicable). Migration of all substances was shown to be below their SML. The unlisted substances were evaluated using toxicological information from previous evaluations, or if not available, by applying the Threshold of Toxicological Concern (TTC) approach. In case the estimated exposure to the unlisted substance exceeded the human exposure TTC value, a more indepth risk assessment was performed. Based on the results of both parts of the study, four baby bottles were considered of high concern because of the potential toxicity of migrating compounds.

Screening of endocrine activity of compounds migrating from plastic baby bottles using a multi-receptor panel of in vitro bioassays.

Endocrine activity of 65 compounds migrating from polycarbonate replacement plastic baby bottles was assessed using in vitro cell based assays (reporter gene assays) involving 7 nuclear receptors, i.e. human steroid hormones receptors (oestrogen, androgen, progesterone and glucocorticoid receptors), human thyroid beta and peroxisome proliferator-activated gamma receptors, and the mouse aryl hydrocarbon receptor. The chemicals were tested at 4 concentrations ranging from 0.001mM to 1mM. Only twelve chemicals did not show any activity towards any of the nuclear receptors, while fifty three compounds showed a possible endocrine activity. Most of the agonistic activities were observed towards the oestrogen receptor while the PPARγ was the target for most of the recorded antagonistic activities. Agonistic activities were recorded for several phthalates, benzophenones, aromatic hydrocarbons and phenols, while compounds such as benzaldehydes, ketones and esters of fatty acid showed antagonistic activities. Thirty five chemicals were able of agonistic activities on 1 to 4 receptors and antagonistic activities were recorded for 35 compounds as well, towards 1 to 7 receptors. Sixteen compounds were able of both agonistic and antagonistic activities, but not on the same receptors, except in 2 cases for the oestrogen receptor and 4 cases for the PPARγ.