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Metals that are important for food safety control of bread product
Abstracts / Toxicology Letters 258S (2016) S62–S324
P09-013 In vitro study of mutagenicity of cylindrospermopsin and cytotoxic effects of the combination of microcystin and cylindrospermopsin A. Jos ∗ , D. Gutiérrez Praena, S. Maisanaba, M. Llana Ruíz Cabello, A. Cameán Area of Toxicology, Faculty of Pharmacy, University of Sevilla. C/Profesor García González 2, 41012 Sevilla, Spain Cylindrospermopsin (CYN) and microcystin-LR (MC-LR) are two cyanotoxins produced by several species of cyanobacteria and they are well distributed worldwide. It is known these toxins, separately, induce harmful effects in human, animals and plants, included those destined for human consumption. However, some of their toxic effects are still scarcely investigated, such as the mutagenicity induced by CYN. Moreover, CYN and MC-LR can appear in water reservoirs at the same time, so it could be assumed that both cyanotoxins would induce toxic effects, although the influence of their combination on their toxicity is still unknown. In the present study, the potential mutagenicity of CYN was studied using the Bacterial Reverse Mutation Test (OECD 471). In addition, the cytotoxic effects of the combination of different concentrations of CYN and MC-LR were studied in the hepatic cell line HepG2 through the MTS reduction assay. This combination was also evaluated using the CalcuSyn program in order to establish an additive or synergistic effect of the toxins. Preliminary results did not show a mutagenic response produced by CYN. With respect to the cytotoxicity, the mixture appeared to induce more damage than the cyanotoxins individually. Further works to clarify the mutagenic potential of CYN and how simultaneous exposure to cyanotoxins affects their toxicity are needed. Acknowledgements: The authors want to thank the Spanish Ministerio de Economía y Competitividad (AGL2015-64558-R) for the financial support. Moreover, the authors want to thank the Biology Service of the CITIUS of the University of Sevilla for their technical support. http://dx.doi.org/10.1016/j.toxlet.2016.06.1615 P09-014 Novel graphical tool to synthetise evidence from animal studies reporting on toxicological endpoints C. Croera 1,∗ , R. Pirow 2 , U. Gundert Remy 3 , T. Husøy 2 , C. Smeraldi 1 , A. Castoldi 1 1
European Food Safety Authority, Food Ingredients and Packaging Unit (FIP), Parma, Italy 2 Working Group on Bisphenol A Toxicology, European Food Safety Authority (EFSA), Parma, Italy 3 Working Group on Isoflavones, European Food Safety Authority (EFSA), Parma, Italy In 2015 the CEF and ANS EFSA Scientific Panels published the opinions on: (1) the health risks from bisphenol A (BPA) in foodstuffs, and on (2) the potential harmful effects of isoflavones in food supplements for peri- and post-menopausal women. For both assessments, a large amount of data was gathered from literature. The data were too heterogeneous for a formal meta-analysis. Hence, a novel graphical approach was developed to synthesise the data on specific endpoints and explore the dose–response relationship. The graphical figures were generated in the statistical computing envi-
S163
ronment R using an adapted code based on the R Package ‘lattice’. In BPA assessment, “Proliferative changes in mammary gland” were identified as toxicity endpoint. The related animal studies were grouped according to the histopathological changes observed. For isoflavones, “Increase of uterine weight” in ovariectomised (OVX) animals was identified as an endpoint. The studies were grouped according to the type of isoflavones administered and sorted for their duration. For BPA, the results were plotted on a common oral human equivalent dose scale. The graph included scores for study reliability and relevance, strengths and weaknesses, exposure and time of assessment. For isoflavones, statistical significant increase in uterine weight in treated animals versus OVX-control was indicated by the size of the dot, being proportionate to the relative effect compared to OVX-control. This graphical tool can be an effective way for synthesising evidence from a large number of animal studies reporting on the same endpoint. http://dx.doi.org/10.1016/j.toxlet.2016.06.1616 P09-015 Assessment of mercury intake based on intervention research in Polish subpopulation R. Kuras ∗ , B. Janasik, M. Stanislawska, W. Wasowicz Department of Biological and Environmental Monitoring, Nofer Institute of Occupational Medicine, Lodz, Poland Fish and marine mammal consumptions are an important pathway for human exposure to mercury (Hg), and ingestion of fish contaminated with methylmercury can lead to adverse health outcomes. In this study, we analyzed mercury content in hair and blood samples collected from 67 volunteers during June to July 2015. An intervention study was based on 10-day fish consumption frequency. Hg concentrations in hair and blood samples derived from men were determined using TDA-AAS technique. Mean concentration of Hg in whole blood of volunteers in the first day of fish consumption was found to be: 0.62 ± 0.41 g/l (NS); after one week: 0.90 ± 0.46 g/l (p < 0.001) and in the end of the study 1.28 ± 0.49 g/l (p < 0.001) and one month after the end of the study (“wash–out” period) 0.78 ± 0.60 g/l (p < 0.001). Mean concentration of Hg in hair of volunteers in the first day of fish consumption was found to be: 0.24 ± 0.16 g/l (NS) and wash–out period: 0.29 ± 0.15 g/l (p < 0.05). The paper estimate weekly Hg intake from fish based on intervention study. The projected intake values of Hg through human consumption were calculated and were compared with the PTWI value established by JECFA. Acknowledgement: Financial support by Grant: 2013/11/B/NZ7/04934. http://dx.doi.org/10.1016/j.toxlet.2016.06.1617 P09-016 Metals that are important for food safety control of bread product Y. Feyzi 1 , A. Malekirad 1,∗ , M. Fazilati 1 , H. Salavati 1 , S. Habibollahi 1 , M. Rezaei 2 1
Department of Biology, Payame Noor University, Tehran, Iran Department of Food Safety and Hygiene, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran 2
Introduction: Breads are among the most important elements of our daily diet. Knowing about the heavy and essential elements
S164
Abstracts / Toxicology Letters 258S (2016) S62–S324
content in bread is required for different purposes such as estimating accumulation of major and trace elements in the bread that may influence water, soil, air, and environment quality as well as the role of bread in nutrition. Method: This study focuses on content of elements such as As, Hg, Cd, Pb, Ni, Fe, Cu and Zn in bread products available in Shahrekord, Iran. Totally 40 bread samples were collected in January 2014. Result: The mean concentrations of As, Hg, Cd, Pb, Ni, Fe, Cu and Zn in bread samples were 12.5 ± 0, 4.38 ± 0, 9.48 ± 4.75, 931.25 ± 722.5, 98.03 ± 26.89, 2324.69 ± 248, 14276.69 ± 10100.79 and 30316.88 ± 6802.19 g/kg, respectively. Conclusion: The concentration of the lead was higher than maximum allowable concentration that probably it is associated to activities of industrial factories around the farms and machine activities. http://dx.doi.org/10.1016/j.toxlet.2016.06.1618 P09-017 Multiresidue method for analysis of pesticides in tomatoes by liquid chromatography coupled with tandem mass spectrometry H. Kallah 1,∗ , R. Saadi 1 , S. Aouam 2 , S. Djelad 3 , Z. Gaouar 1 1
Environmental Health Research Laboratory, Oran University, Oran, Algeria 2 Department of Pharmacology Toxicology, CHU ORAN, Oran, Algeria 3 Department of Pharmacy, Faculty of Medicine, Oran University, Oran, Algeria Introduction: Pesticides are nowadays considered as toxic for human health. The maximal residues levels authorized in water and foodstuff are more and more strict. Therefore, sensitive and selective analytical techniques are necessary for their identification and their quantification. The aim of this study was to develop an analytical method for determining different classes of pesticides in tomatoes. Material and methods: A descriptive survey of farmers using a questionnaire has been carried. Then, a multiresidus method for the determination of 13 pesticides in tomatoes by HPLC/MS/MS after solid extraction in dispersive phase QuEChERS was validated, and applied to samples of tomatoes obtained from markets or directly from the open fields cultures. Results: The results of the descriptive study have guided the selection of a list of 13 pesticides belonging to different families (organophosphates, carbamates, triazoles, etc.) to quantifying. The multiresidus method validated for these pesticides in tomatoes according to SANCO guidelines shows recoveries range between 84 and 117% and coefficients of relative standard deviation (RSD) between 1.18 and 9.73%. The limits of detection and quantification are still below the MRLs of various analyzed pesticides. Thirty samples was analyzed, pesticides were detected in 47% of them at varying concentrations (2.29–64.27 g/kg), and 7% exceeded MRLs established by the directives of the European Union. Conclusion: It is will be interesting to complete this study by the development of other methods for determining of more pesticides in different matrices (water, fruits and vegetables) in order to evaluate the levels of pesticide residues to which the consumer is exposed. http://dx.doi.org/10.1016/j.toxlet.2016.06.1619
P09-018 Food safety evaluation of a soft drink and caramel color additive (e150d) P. Alves-Martínez 1 , T. Merinas-Amo 1 , F. Valenzuela-Gómez 1,∗ , R. Merinas-Amo 1 , L. Pérez-Raya 1 , A. Velasco-Ruiz 1 , Á. Alonso-Moraga 1 , Z. Fernández-Bedmar 1 , S. Demyda Peyrás 2 , M. Mateo-Fernández 1 1
Department of Genetics, Gregor Mendel Building, Faculty of Science, University of Córdoba, Córdoba, Spain 2 Institute of Veterinary Genetics (IGEVET), CCT La Plata, CONICET, Faculty of Veterinary Sciences, University of La Plata, Buenos Aires, Argentina Schools and local communities make the effort to reduce the consumption of sugary beverages since they are not part of a balanced diet. Despite this fact, Pepsi-Cola is one of the most consumed soda drinks. Therefore, studies are needed to evaluate the biological effects of lyophilised Pepsi-Cola (PEP) and a main additive like caramel color E-150d (CAR). We performed in vivo toxicity, antitoxicity and lifespan assays using Drosophila melanogaster; and in vitro cytotoxicity, DNA fragmentation, methylation status and comet assays using HL-60 promyelocytic cell line. Results showed that PEP is toxic at the highest assayed concentration (110 mg/ml) and protects the individuals against the toxicant hydrogen peroxide at the lowest one (0.86 mg/ml). CAR was toxic at all assayed concentrations. Regarding lifespan assays, PEP showed the same effect as negative control whereas CAR decreased the longevity of flies. According to in vitro assays, PEP and CAR were cytotoxic reaching IC50 at 25 mg/ml and 1 mg/ml respectively. However, none of the tested compounds were able to induce DNA internucleosomal fragmentation. PEP increased the repetitive elements methylation status and induced neither single nor double strand DNA breaks in HL-60 cells. In conclusion, we demonstrate the safety of PEP in the Drosophila model, and its antioxidant in vivo activity at the lowest assayed concentration. Although PEP and CAR show a chemopreventive property, it is not due to proapoptotic mechanisms. CAR could not be considered as a safe additive since it is toxic and decrease the lifespan expectancy in the Drosophila model. http://dx.doi.org/10.1016/j.toxlet.2016.06.1620 P09-019 The repeated-dose 90-day study oral toxicity study in rodents (OECD TG408) as a tool to investigate toxicological effects of the whole genetically modified (GM) food/feed: Differences from the 90-day study on chemicals and challenges A. Lanzoni 1,∗ , M. Ardizzone 1 , L. Martino 2 , S. Favilla 3 , C. Paoletti 1 1
European Food Safety Authority (EFSA), Genetically Modified Organism (GMO) Unit, Italy 2 European Food Safety Authority (EFSA), Assessment and Methodological Support (AMU) Unit, Italy 3 Functional Neuroimaging Group, Department of Biomedical, Metabolical and Neural Sciences University of Modena & R.E, Italy In food and feed risk assessment, the whole food/feed can be investigated for possible effects of toxicological relevance for consumers, and such a testing should be triggered by hazards identified by preceding analyses, or by specific properties of the food/feed under assessment [EFSA, 2011]. Relevant for GM foods/feeds and
P09-013 In vitro study of mutagenicity of cylindrospermopsin and cytotoxic effects of the combination of microcystin and cylindrospermopsin A. Jos ∗ , D. Gutiérrez Praena, S. Maisanaba, M. Llana Ruíz Cabello, A. Cameán Area of Toxicology, Faculty of Pharmacy, University of Sevilla. C/Profesor García González 2, 41012 Sevilla, Spain Cylindrospermopsin (CYN) and microcystin-LR (MC-LR) are two cyanotoxins produced by several species of cyanobacteria and they are well distributed worldwide. It is known these toxins, separately, induce harmful effects in human, animals and plants, included those destined for human consumption. However, some of their toxic effects are still scarcely investigated, such as the mutagenicity induced by CYN. Moreover, CYN and MC-LR can appear in water reservoirs at the same time, so it could be assumed that both cyanotoxins would induce toxic effects, although the influence of their combination on their toxicity is still unknown. In the present study, the potential mutagenicity of CYN was studied using the Bacterial Reverse Mutation Test (OECD 471). In addition, the cytotoxic effects of the combination of different concentrations of CYN and MC-LR were studied in the hepatic cell line HepG2 through the MTS reduction assay. This combination was also evaluated using the CalcuSyn program in order to establish an additive or synergistic effect of the toxins. Preliminary results did not show a mutagenic response produced by CYN. With respect to the cytotoxicity, the mixture appeared to induce more damage than the cyanotoxins individually. Further works to clarify the mutagenic potential of CYN and how simultaneous exposure to cyanotoxins affects their toxicity are needed. Acknowledgements: The authors want to thank the Spanish Ministerio de Economía y Competitividad (AGL2015-64558-R) for the financial support. Moreover, the authors want to thank the Biology Service of the CITIUS of the University of Sevilla for their technical support. http://dx.doi.org/10.1016/j.toxlet.2016.06.1615 P09-014 Novel graphical tool to synthetise evidence from animal studies reporting on toxicological endpoints C. Croera 1,∗ , R. Pirow 2 , U. Gundert Remy 3 , T. Husøy 2 , C. Smeraldi 1 , A. Castoldi 1 1
European Food Safety Authority, Food Ingredients and Packaging Unit (FIP), Parma, Italy 2 Working Group on Bisphenol A Toxicology, European Food Safety Authority (EFSA), Parma, Italy 3 Working Group on Isoflavones, European Food Safety Authority (EFSA), Parma, Italy In 2015 the CEF and ANS EFSA Scientific Panels published the opinions on: (1) the health risks from bisphenol A (BPA) in foodstuffs, and on (2) the potential harmful effects of isoflavones in food supplements for peri- and post-menopausal women. For both assessments, a large amount of data was gathered from literature. The data were too heterogeneous for a formal meta-analysis. Hence, a novel graphical approach was developed to synthesise the data on specific endpoints and explore the dose–response relationship. The graphical figures were generated in the statistical computing envi-
S163
ronment R using an adapted code based on the R Package ‘lattice’. In BPA assessment, “Proliferative changes in mammary gland” were identified as toxicity endpoint. The related animal studies were grouped according to the histopathological changes observed. For isoflavones, “Increase of uterine weight” in ovariectomised (OVX) animals was identified as an endpoint. The studies were grouped according to the type of isoflavones administered and sorted for their duration. For BPA, the results were plotted on a common oral human equivalent dose scale. The graph included scores for study reliability and relevance, strengths and weaknesses, exposure and time of assessment. For isoflavones, statistical significant increase in uterine weight in treated animals versus OVX-control was indicated by the size of the dot, being proportionate to the relative effect compared to OVX-control. This graphical tool can be an effective way for synthesising evidence from a large number of animal studies reporting on the same endpoint. http://dx.doi.org/10.1016/j.toxlet.2016.06.1616 P09-015 Assessment of mercury intake based on intervention research in Polish subpopulation R. Kuras ∗ , B. Janasik, M. Stanislawska, W. Wasowicz Department of Biological and Environmental Monitoring, Nofer Institute of Occupational Medicine, Lodz, Poland Fish and marine mammal consumptions are an important pathway for human exposure to mercury (Hg), and ingestion of fish contaminated with methylmercury can lead to adverse health outcomes. In this study, we analyzed mercury content in hair and blood samples collected from 67 volunteers during June to July 2015. An intervention study was based on 10-day fish consumption frequency. Hg concentrations in hair and blood samples derived from men were determined using TDA-AAS technique. Mean concentration of Hg in whole blood of volunteers in the first day of fish consumption was found to be: 0.62 ± 0.41 g/l (NS); after one week: 0.90 ± 0.46 g/l (p < 0.001) and in the end of the study 1.28 ± 0.49 g/l (p < 0.001) and one month after the end of the study (“wash–out” period) 0.78 ± 0.60 g/l (p < 0.001). Mean concentration of Hg in hair of volunteers in the first day of fish consumption was found to be: 0.24 ± 0.16 g/l (NS) and wash–out period: 0.29 ± 0.15 g/l (p < 0.05). The paper estimate weekly Hg intake from fish based on intervention study. The projected intake values of Hg through human consumption were calculated and were compared with the PTWI value established by JECFA. Acknowledgement: Financial support by Grant: 2013/11/B/NZ7/04934. http://dx.doi.org/10.1016/j.toxlet.2016.06.1617 P09-016 Metals that are important for food safety control of bread product Y. Feyzi 1 , A. Malekirad 1,∗ , M. Fazilati 1 , H. Salavati 1 , S. Habibollahi 1 , M. Rezaei 2 1
Department of Biology, Payame Noor University, Tehran, Iran Department of Food Safety and Hygiene, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran 2
Introduction: Breads are among the most important elements of our daily diet. Knowing about the heavy and essential elements
S164
Abstracts / Toxicology Letters 258S (2016) S62–S324
content in bread is required for different purposes such as estimating accumulation of major and trace elements in the bread that may influence water, soil, air, and environment quality as well as the role of bread in nutrition. Method: This study focuses on content of elements such as As, Hg, Cd, Pb, Ni, Fe, Cu and Zn in bread products available in Shahrekord, Iran. Totally 40 bread samples were collected in January 2014. Result: The mean concentrations of As, Hg, Cd, Pb, Ni, Fe, Cu and Zn in bread samples were 12.5 ± 0, 4.38 ± 0, 9.48 ± 4.75, 931.25 ± 722.5, 98.03 ± 26.89, 2324.69 ± 248, 14276.69 ± 10100.79 and 30316.88 ± 6802.19 g/kg, respectively. Conclusion: The concentration of the lead was higher than maximum allowable concentration that probably it is associated to activities of industrial factories around the farms and machine activities. http://dx.doi.org/10.1016/j.toxlet.2016.06.1618 P09-017 Multiresidue method for analysis of pesticides in tomatoes by liquid chromatography coupled with tandem mass spectrometry H. Kallah 1,∗ , R. Saadi 1 , S. Aouam 2 , S. Djelad 3 , Z. Gaouar 1 1
Environmental Health Research Laboratory, Oran University, Oran, Algeria 2 Department of Pharmacology Toxicology, CHU ORAN, Oran, Algeria 3 Department of Pharmacy, Faculty of Medicine, Oran University, Oran, Algeria Introduction: Pesticides are nowadays considered as toxic for human health. The maximal residues levels authorized in water and foodstuff are more and more strict. Therefore, sensitive and selective analytical techniques are necessary for their identification and their quantification. The aim of this study was to develop an analytical method for determining different classes of pesticides in tomatoes. Material and methods: A descriptive survey of farmers using a questionnaire has been carried. Then, a multiresidus method for the determination of 13 pesticides in tomatoes by HPLC/MS/MS after solid extraction in dispersive phase QuEChERS was validated, and applied to samples of tomatoes obtained from markets or directly from the open fields cultures. Results: The results of the descriptive study have guided the selection of a list of 13 pesticides belonging to different families (organophosphates, carbamates, triazoles, etc.) to quantifying. The multiresidus method validated for these pesticides in tomatoes according to SANCO guidelines shows recoveries range between 84 and 117% and coefficients of relative standard deviation (RSD) between 1.18 and 9.73%. The limits of detection and quantification are still below the MRLs of various analyzed pesticides. Thirty samples was analyzed, pesticides were detected in 47% of them at varying concentrations (2.29–64.27 g/kg), and 7% exceeded MRLs established by the directives of the European Union. Conclusion: It is will be interesting to complete this study by the development of other methods for determining of more pesticides in different matrices (water, fruits and vegetables) in order to evaluate the levels of pesticide residues to which the consumer is exposed. http://dx.doi.org/10.1016/j.toxlet.2016.06.1619
P09-018 Food safety evaluation of a soft drink and caramel color additive (e150d) P. Alves-Martínez 1 , T. Merinas-Amo 1 , F. Valenzuela-Gómez 1,∗ , R. Merinas-Amo 1 , L. Pérez-Raya 1 , A. Velasco-Ruiz 1 , Á. Alonso-Moraga 1 , Z. Fernández-Bedmar 1 , S. Demyda Peyrás 2 , M. Mateo-Fernández 1 1
Department of Genetics, Gregor Mendel Building, Faculty of Science, University of Córdoba, Córdoba, Spain 2 Institute of Veterinary Genetics (IGEVET), CCT La Plata, CONICET, Faculty of Veterinary Sciences, University of La Plata, Buenos Aires, Argentina Schools and local communities make the effort to reduce the consumption of sugary beverages since they are not part of a balanced diet. Despite this fact, Pepsi-Cola is one of the most consumed soda drinks. Therefore, studies are needed to evaluate the biological effects of lyophilised Pepsi-Cola (PEP) and a main additive like caramel color E-150d (CAR). We performed in vivo toxicity, antitoxicity and lifespan assays using Drosophila melanogaster; and in vitro cytotoxicity, DNA fragmentation, methylation status and comet assays using HL-60 promyelocytic cell line. Results showed that PEP is toxic at the highest assayed concentration (110 mg/ml) and protects the individuals against the toxicant hydrogen peroxide at the lowest one (0.86 mg/ml). CAR was toxic at all assayed concentrations. Regarding lifespan assays, PEP showed the same effect as negative control whereas CAR decreased the longevity of flies. According to in vitro assays, PEP and CAR were cytotoxic reaching IC50 at 25 mg/ml and 1 mg/ml respectively. However, none of the tested compounds were able to induce DNA internucleosomal fragmentation. PEP increased the repetitive elements methylation status and induced neither single nor double strand DNA breaks in HL-60 cells. In conclusion, we demonstrate the safety of PEP in the Drosophila model, and its antioxidant in vivo activity at the lowest assayed concentration. Although PEP and CAR show a chemopreventive property, it is not due to proapoptotic mechanisms. CAR could not be considered as a safe additive since it is toxic and decrease the lifespan expectancy in the Drosophila model. http://dx.doi.org/10.1016/j.toxlet.2016.06.1620 P09-019 The repeated-dose 90-day study oral toxicity study in rodents (OECD TG408) as a tool to investigate toxicological effects of the whole genetically modified (GM) food/feed: Differences from the 90-day study on chemicals and challenges A. Lanzoni 1,∗ , M. Ardizzone 1 , L. Martino 2 , S. Favilla 3 , C. Paoletti 1 1
European Food Safety Authority (EFSA), Genetically Modified Organism (GMO) Unit, Italy 2 European Food Safety Authority (EFSA), Assessment and Methodological Support (AMU) Unit, Italy 3 Functional Neuroimaging Group, Department of Biomedical, Metabolical and Neural Sciences University of Modena & R.E, Italy In food and feed risk assessment, the whole food/feed can be investigated for possible effects of toxicological relevance for consumers, and such a testing should be triggered by hazards identified by preceding analyses, or by specific properties of the food/feed under assessment [EFSA, 2011]. Relevant for GM foods/feeds and