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Biological fate of styrene oxide adducts with globin: Identification of degradation products in rat urine
S60
Abstracts / Toxicology Letters 238S (2015) S56–S383
EC50 value than in rat cells. PCDDs and PCDFs (with a high dioxinlike activity) elicited similar activities in both human and rat cell models. In contrast, the dioxin-like PCBs did not induce the AhRmediated activity in AZ-AhR cells, with a sole exception of PCB126, which reached only 46% of maximal induction. The PAHs with a low AhR-mediated activity in rat cells did not induce any luciferase activity in human AZ-AhR cell line. Chrysene, benz[a]anthracene, benzo[a]pyrene, dibenzo[a,h]anthracene, and several other PAHs with a relatively high AhR-mediated activity in DR-CALUX assay, elicited only lower and partial induction responses AZ-AhR cells. Other evaluated PAHs (benzo[k]fluoranthene and indenopyrene) exhibited similar relative effective potencies (REPs) in both cell models, while higher relative potencies of benzo[b]fluoranthene and 5-methylchrysene in human cells were probably due to a different rate of metabolism in human and rat cells. In general, human cells were less sensitive to the AhR activation, which reflected well-known inter-species differences in the AhR affinity. The data also indicate that, for some PAHs and dioxin-like compounds, their REPs might differ between human and rodent cells. This study was supported by grant no. P503-12-G147 from the Czech Science Foundation. http://dx.doi.org/10.1016/j.toxlet.2015.08.215
P01-017 Biological fate of styrene oxide adducts with globin: Identification of degradation products in rat urine ˚ Duˇsková 1 , I. Hanzlíková 1 , L. J. Mraz 1,∗ , I. Linhart 2 , U. Dabrowská 1 , A. Moulisová 2 , K. Hejl 2 1
National Institute of Public Health, Centre of Occupational Health, Prague, Czech Republic 2 University of Chemistry and Technology, Department of Organic Chemistry, Prague, Czech Republic
Styrene oxide (SO), an electrophilic metabolic intermediate of the important industrial chemical styrene, forms covalent adducts in vivo and in vitro with blood protein globin via nucleophilic groups in Cys, His, Lys and N-terminal Val. Racemic SO can react with nucleophilic amino acid residues via both carbons of its epoxide ring to afford corresponding 2-hydroxy-1-phenylethyl-(2,1-HPE) and 2hydroxy-2-phenylethyl-(2,2-HPE) regioisomers, each in the form of two diastereomers. In His, the number of possible adduct forms is even doubled to 8 due to presence of two reactive nitrogen atoms in the imidazole ring. In our studies on the fate of globin adducts following physiological removal of erythrocytes from the circulation, the adducts with SO were employed as test compounds. Using a new experimental model, erythrocytes from donor rats were incubated with racemic SO (10 mM, 22 ◦ C, 20 h) and then intravenously administered to the acceptor rats. Because major part of the erythrocytes was removed from circulation within 24 h, degradation products of the globin adducts were massively excreted in the urine. HPLC-HRMS2 analysis of rat urine revealed an array of HPE regio- and stereoisomers resulting from all amino acid adducts investigated: (a) Val adducts mainly in the form of both 2,1- and 2,2-HPE-Val-Leu, produced by incomplete proteolysis of globin ␣chain, with only traces of 2,2- but not 2,1-HPE-Val; (b) Cys adducts in the form of N␣ -acetyl derivatives (2,1- and 2,2-HPE mercapturic acids); (c) His adducts in the form of six N␣ -acetyl HPE derivatives (regioisomerism not determined); (d) Lys adducts in the form of N␣ -acetyl 2,1- and 2,2-HPE species (single each).
Acknowledgement: The study was supported by the grant NT13401-4/2012 from Internal Grant Agency of the Czech Ministry of Health. http://dx.doi.org/10.1016/j.toxlet.2015.08.216
P01-018 Safety evaluation of technically unavoidable traces of metals in cosmetics M. Marinovich 1,∗ , M.S. Boraso 1 , E. Testai 2 , C.L. Galli 1 1 Lab. of Toxicology and Risk Assessment, UNISAFE, Department of Pharmacological and Biomolecular Sciences, University of Milan, Milan, Italy 2 Instituto Superiore di Sanità, Department of Environment and Primary Prevention, Mechanisms of Toxicity Unit, Rome, Italy
According to EU Regulation no. 1223/2009/CE cosmetic products for daily use can contain “technically unavoidable traces” of metals. This definition is too vague. Authorities should set well defined limits, considering the risks associated with metal contamination of personal care products (PCPs). This presentation characterizes the risk arising from a number of metals (antimony, arsenic, cadmium, cobalt, chromium, mercury, nickel, lead) that may occur in raw materials and, consequently, in PCPs. A worst case scenario is adopted, based on the following assumptions: (i) the individual ingredients contained the maximum amount ever measured for each metal, (ii) the hypothetical PCP was produced exclusively with that single ingredient; (iii) when absorption through the skin was not known, data related to oral absorption were used. Risk characterization was performed calculating the Systemic Exposure Dosage (SED) and the Margin of Safety (MoS = NOAEL or BMDL10/SED). Exposure to the allegedly “technically unavoidable” maximum amounts of metals in cosmetic ingredients resulted in MoSs exceeding 100 (safety threshold) with one exception. This suggests that the availability of experimental dermal absorption rates could enable significant improvement in MoS, thus increasing safety levels. Although results are reassuring, the authors recommend minimization of contamination, according to the state of the art of manufacturing methods. http://dx.doi.org/10.1016/j.toxlet.2015.08.217
P01-019 Enniatin B mycotoxin is excreted as such to rat urine P. Koivisto 1,∗ , M. Jonsson 1 , M. Jestoi 1 , K. Peltonen 2 1 2
Evira, Chemistry and Toxicology, Helsinki, Finland Finnish Safety and Chemicals Agency, Helsinki, Finland
Introduction: Mycotoxins are natural toxins produced by moulds. Characteristic properties of some of the toxins are rather well known, while some still needs active research for better risk characterisation. Exposure to mycotoxins is a continuous process, particularly if cereal products are consumed regularly. Enniatins are cyclic toxins produced by Fusarium type fungi. In this study male Sprague Dawley rats were exposed to enniatins to get insight knowledge of ADME processes of enniatins. Materials and methods: Enniatins (commercial mixture of A, A1, B (major component), B1) were administered in carboxymethylcellulose emulsion resulting a exposure level of 50 mg/kg. A single by gavage administration was applied. Urine samples were collected
Abstracts / Toxicology Letters 238S (2015) S56–S383
EC50 value than in rat cells. PCDDs and PCDFs (with a high dioxinlike activity) elicited similar activities in both human and rat cell models. In contrast, the dioxin-like PCBs did not induce the AhRmediated activity in AZ-AhR cells, with a sole exception of PCB126, which reached only 46% of maximal induction. The PAHs with a low AhR-mediated activity in rat cells did not induce any luciferase activity in human AZ-AhR cell line. Chrysene, benz[a]anthracene, benzo[a]pyrene, dibenzo[a,h]anthracene, and several other PAHs with a relatively high AhR-mediated activity in DR-CALUX assay, elicited only lower and partial induction responses AZ-AhR cells. Other evaluated PAHs (benzo[k]fluoranthene and indenopyrene) exhibited similar relative effective potencies (REPs) in both cell models, while higher relative potencies of benzo[b]fluoranthene and 5-methylchrysene in human cells were probably due to a different rate of metabolism in human and rat cells. In general, human cells were less sensitive to the AhR activation, which reflected well-known inter-species differences in the AhR affinity. The data also indicate that, for some PAHs and dioxin-like compounds, their REPs might differ between human and rodent cells. This study was supported by grant no. P503-12-G147 from the Czech Science Foundation. http://dx.doi.org/10.1016/j.toxlet.2015.08.215
P01-017 Biological fate of styrene oxide adducts with globin: Identification of degradation products in rat urine ˚ Duˇsková 1 , I. Hanzlíková 1 , L. J. Mraz 1,∗ , I. Linhart 2 , U. Dabrowská 1 , A. Moulisová 2 , K. Hejl 2 1
National Institute of Public Health, Centre of Occupational Health, Prague, Czech Republic 2 University of Chemistry and Technology, Department of Organic Chemistry, Prague, Czech Republic
Styrene oxide (SO), an electrophilic metabolic intermediate of the important industrial chemical styrene, forms covalent adducts in vivo and in vitro with blood protein globin via nucleophilic groups in Cys, His, Lys and N-terminal Val. Racemic SO can react with nucleophilic amino acid residues via both carbons of its epoxide ring to afford corresponding 2-hydroxy-1-phenylethyl-(2,1-HPE) and 2hydroxy-2-phenylethyl-(2,2-HPE) regioisomers, each in the form of two diastereomers. In His, the number of possible adduct forms is even doubled to 8 due to presence of two reactive nitrogen atoms in the imidazole ring. In our studies on the fate of globin adducts following physiological removal of erythrocytes from the circulation, the adducts with SO were employed as test compounds. Using a new experimental model, erythrocytes from donor rats were incubated with racemic SO (10 mM, 22 ◦ C, 20 h) and then intravenously administered to the acceptor rats. Because major part of the erythrocytes was removed from circulation within 24 h, degradation products of the globin adducts were massively excreted in the urine. HPLC-HRMS2 analysis of rat urine revealed an array of HPE regio- and stereoisomers resulting from all amino acid adducts investigated: (a) Val adducts mainly in the form of both 2,1- and 2,2-HPE-Val-Leu, produced by incomplete proteolysis of globin ␣chain, with only traces of 2,2- but not 2,1-HPE-Val; (b) Cys adducts in the form of N␣ -acetyl derivatives (2,1- and 2,2-HPE mercapturic acids); (c) His adducts in the form of six N␣ -acetyl HPE derivatives (regioisomerism not determined); (d) Lys adducts in the form of N␣ -acetyl 2,1- and 2,2-HPE species (single each).
Acknowledgement: The study was supported by the grant NT13401-4/2012 from Internal Grant Agency of the Czech Ministry of Health. http://dx.doi.org/10.1016/j.toxlet.2015.08.216
P01-018 Safety evaluation of technically unavoidable traces of metals in cosmetics M. Marinovich 1,∗ , M.S. Boraso 1 , E. Testai 2 , C.L. Galli 1 1 Lab. of Toxicology and Risk Assessment, UNISAFE, Department of Pharmacological and Biomolecular Sciences, University of Milan, Milan, Italy 2 Instituto Superiore di Sanità, Department of Environment and Primary Prevention, Mechanisms of Toxicity Unit, Rome, Italy
According to EU Regulation no. 1223/2009/CE cosmetic products for daily use can contain “technically unavoidable traces” of metals. This definition is too vague. Authorities should set well defined limits, considering the risks associated with metal contamination of personal care products (PCPs). This presentation characterizes the risk arising from a number of metals (antimony, arsenic, cadmium, cobalt, chromium, mercury, nickel, lead) that may occur in raw materials and, consequently, in PCPs. A worst case scenario is adopted, based on the following assumptions: (i) the individual ingredients contained the maximum amount ever measured for each metal, (ii) the hypothetical PCP was produced exclusively with that single ingredient; (iii) when absorption through the skin was not known, data related to oral absorption were used. Risk characterization was performed calculating the Systemic Exposure Dosage (SED) and the Margin of Safety (MoS = NOAEL or BMDL10/SED). Exposure to the allegedly “technically unavoidable” maximum amounts of metals in cosmetic ingredients resulted in MoSs exceeding 100 (safety threshold) with one exception. This suggests that the availability of experimental dermal absorption rates could enable significant improvement in MoS, thus increasing safety levels. Although results are reassuring, the authors recommend minimization of contamination, according to the state of the art of manufacturing methods. http://dx.doi.org/10.1016/j.toxlet.2015.08.217
P01-019 Enniatin B mycotoxin is excreted as such to rat urine P. Koivisto 1,∗ , M. Jonsson 1 , M. Jestoi 1 , K. Peltonen 2 1 2
Evira, Chemistry and Toxicology, Helsinki, Finland Finnish Safety and Chemicals Agency, Helsinki, Finland
Introduction: Mycotoxins are natural toxins produced by moulds. Characteristic properties of some of the toxins are rather well known, while some still needs active research for better risk characterisation. Exposure to mycotoxins is a continuous process, particularly if cereal products are consumed regularly. Enniatins are cyclic toxins produced by Fusarium type fungi. In this study male Sprague Dawley rats were exposed to enniatins to get insight knowledge of ADME processes of enniatins. Materials and methods: Enniatins (commercial mixture of A, A1, B (major component), B1) were administered in carboxymethylcellulose emulsion resulting a exposure level of 50 mg/kg. A single by gavage administration was applied. Urine samples were collected