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Exposure Assessment of Catalonian Population to Mycotoxins

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Exposure Assessment of Catalonian Population to Mycotoxins Exposure assessment of Catalonian population to mycotoxins Avaluació de l'exposició de la població catalana a micotoxines German Cano Sancho The present work is included in the following research projects - Evaluación de la exposición de la población española a las toxinas de Fusarium . 2009. Ministerio de Ciencia e Innovación. AGL2008-05030-C02-01 - Aproximación integrada a la exposición humana simultánea a ocratoxina A y deoxinivalenol (MYCOEXPO). 2012. AGL2011-24862. - Exposure assessment of Catalonian population to mycotoxins. Agreement between Catalonian Food Safety Agency and University of Lleida between 2008 and 2010. Pre-doctoral fellowship - 2009 University of Lleida doctoral grant UdL-Competitive projects - 2010-2012 Catalonian Government fellowship. AGAUR FI-DGR 2010. Pre-doctoral training and collaborating residencies - 2010 Mathematics and Applied Informatics Unit- Centre de Jouy en Josas -Institut de National de Recherche Agronomique (INRA). Programming and execution of simulation models to assess the exposure of Catalonian population to the mycotoxin zearalenone and to analyse the sensitivity of the high percentiles - 2010 TOXALIM UMR 1331 INRA/INP/UPS. In vitro methods to assess the toxicity of Fusarium spp. Mycotoxins. - 2012 Departamento de Medio Ambiente, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA). Cytotoxic effects of the simultaneous exposure of Caco-2 cells to mycotoxins and antioxidants. University of Lleida, 2008-2013 Author German Cano Sancho Supervisors Vicente Sanchis Sònia Marín Major contributors Antonio J Ramos Jean P Gauchi Technical contributors Montse Prim Mª Bernarda Coronel Francisco M. Valle Misericordia Jiménez Pedro Burdaspal Teresa M. Legarda Joan Peris Carme Colom Beatriz López Exposure assessment of Catalonian population to mycotoxins Avaluació de l'exposició de la població catalana a micotoxines German Cano Sancho Doctoral Thesis Thesis supervisors Dr. Vicente Sanchis Almenar Dr. Sònia Marín Sillué Dep. Tecnologia d’Aliments Universitat de Lleida Lleida 2013 Acknowledgements / Agraïments/Agradecimientos Nora por tu infinita paciencia, tu fuerza y sensatos consejos. Als meus pares per ser com sou, per fer-me com sóc. Vicent, Sònia i Antònio per fer-me créixer científicament a la par que com a persona. Companys de laboratori aka “Peluqueria Tecal”, per fer d’aquests anys una gran acumulació de bons moments en un entorn a vegades difícil, sou els millors! Merci à J.P. Gauchi pour m’ouvrir les portes de votre laboratoire et le monde fascinant de la modélisation. Gràcies a tothom que ha col·laborat en els diferents treballs i que ha fet posible la realització d’aquesta tesis. Agraïr també a tothom amb qui he compartit els bons moments fora del laboratori durant aquest quatre anys, sense això res és possible. a Nora i als meus pares "No es más quién más alto llega, sino aquel que influido por la belleza que le envuelve, más intensamente siente." Maurice Herzog, alpinista Index Index Index 13 List of abbreviations 15 Chapter 1. Summary / Resum / Resumen 17 Chapter 2. Introduction 29 2.1. Exposure assessment in the framework of risk analysis of chemicals in foods 30 2.2. Exposure assessment highlights 33 2.2.1. Contamination data 34 2.2.2. Consumption data 40 2.2.3. Methods to combine consumption and contamination raw 43 datasets 2.2.4. Biomarkers 45 2.3. Mycotoxins in food 47 2.3.1. Aflatoxins B and G group 47 2.3.2. Aflatoxin M 1 49 2.3.3. Fumonisin B 1 and B 2 51 2.3.4. Patulin 54 2.3.5. Trichothecenes 55 2.3.6. Zearalenone 59 2.4. Legislation and governmental recommendations 60 Chapter 3. Objectives 72 Chapter 4. Global design and methodology 75 4.1 Global study design 76 4.2 Analytical study 77 4.3 Nutritional study 82 4.4 Exposure assessment 83 4.5 Risk characterization 88 13 Index Chapter 5. Occurrence and exposure assessment to aflatoxins B and G in Catalonia 91 Chapter 6. Occurrence and exposure assessment to aflatoxin M 1 in Catalonia 115 Chapter 7. Occurrence of fumonisins in Catalonia (Spain) and an exposure assessment of 130 specific population groups Chapter 8. Sphinganine and sphingosine levels and ratio in urine and blood samples from a 153 Catalonian population (Spain) Chapter 9. Presence of trichothecenes and co-occurrence in cereal-based food from Catalonia 177 (Spain) Chapter 10. Quantitative dietary exposure assessment of the Catalonian population (Spain) to 197 the mycotoxin deoxynivalenol Chapter 11. Effect of food processing on exposure assessment studies with mycotoxins 221 Chapter 12. Exposure assessment of T2 and HT2 toxins in Catalonia (Spain) 242 Chapter 13. Occurrence of zearalenone, an oestrogenic mycotoxin, in Catalonia (Spain) and 264 exposure assessment Chapter 14. Quantitative exposure assessment of Catalonian (Spain) population and special 280 groups to the zearalenone mycotoxin Chapter 15. Presence and co-occurrence of aflatoxins, deoxynivalenol, fumonisins and 306 zearalenone in gluten-free and ethnic foods Chapter 16. Survey of patulin occurrence in apple juice and apple products in Catalonia, 324 Spain, and an estimate of dietary intake Chapter 17. Global discussion 339 Chapter 18. Concluding remarks and future research 357 Annex 1. Biomonitoring of Fusarium spp. mycotoxins: perspectives for an individual exposure 370 assessment tool Annex 2. Regulations of mycotoxins 395 14 Abbreviation List Abbreviation List 15 Abbreviation List Chemistry Organization IARC International Agency for Research on Cancer AFB 1 Aflatoxin B 1 JECFA Joint FAO/WHO Expert AFB Aflatoxin B 2 2 Committee on Food Additives AFG Aflatoxin G 1 1 SCF Scientific Committee on Food AFG Aflatoxin G 2 2 WHO World Health Organisation AFM Aflatoxin M 1 1 AFs Total aflatoxins (B +B +G +G ) 1 2 1 2 Other abbreviations DON Deoxinivalenol D3G Deoxinivalenol 3-glycoside ALARA As Low As Reasonably ELISA Enzyme Linked Achievable Immunosorbent Assay BMD Benmarch Dose FB 1 Fumonisin B 1 BMDL 10 BMD increasing cancer rates 10 FB 2 Fumonisin B 2 % over the control FBs Total fumonisins (B 1+B 2) CI b “Bootstrapping” Confidence HT2 HT-2 toxin Interval HPLC High performance liquid FFQ Food frequency questionnaire chromatography HBV Hepatitis B viruse IAC Immunoaffinity KM Kaplan-Meier chromatography (column) LOAEL Lowest observed adverse effect NIV Nivalenol level OPA o-phthaldialdehyde solution LOD Limit of detection OT ααα Ocratoxin alpha LOQ Limit of quantification PAT Patulin NOAEL No observed adverse effect level PBS Phosphate buffered saline MoE Margin of exposure Sa Sphinganine MLE Maximum likelihood estimation So Sphingosine ND Non detect T2 T-2 toxin Pdf Probability density function TRCs Trichothecenes PMTDI Provisional maximum tolerable daily intake ZEA Zearalenone R3 3 days dietary record R24 24 hours dietary recall Governmental Organisations TD50 Median toxic dose TDI Tolerable daily intake ACSA Agència Catalana de Seguretat Alimentària (Catalonian Safety Food Agency) TWI Tolerable weekly intake EC European Commission EFSA European Food Safety Agency FAO Food and Agriculture 16 Chapter 1. Summary Chapter 1. Summary 17 Chapter 1. Summary 1.1 Summary (English) Mycotoxins are toxic secondary metabolites produced by fungi that contaminate various agricultural commodities either before harvest or under post-harvest conditions. Their purpose is thought to be for fungal defence or competition. Although there are many species of toxigenic moulds, only a few mycotoxins, particularly those affecting cereals (maize, wheat, barley, oats and rice) and groundnuts are considered to be significant for humans. Several mycotoxins have been identified up to the present, but those of special interest in food and feed safety are: aflatoxins (B 1, B2, G 1, G 2, M 1), fumonisins (B 1, B 2), ochratoxin A, patulin, trichothecenes (deoxynivalenol, T-2 toxin, HT-2 toxin), and zearalenone. The general objective of the present thesis was to assess the exposure of Catalonian population to the major mycotoxins, aflatoxins group B and G, aflatoxin M 1, patulin and the Fusarium toxins deoxynivalenol, fumonisins, T-2 and HT-2 toxins, and zearalenone. To reach this general objective, several sub-objectives were developed: 1) To assess the dietary intake of Catalonian population in regard of those mycotoxin related foods, including the most vulnerable collectives like elderly, infants, immigrants or celiac sufferers. 2) To determine the mycotoxin levels in most relevant food categories, from a representative sample purchased around of the geography of Catalonia. 3) To combine accurately raw consumption and contamination datasets in order to obtain a thorough estimation of exposure levels of the population collectives, and a reliable characterization of the related health risk. 4) To assess the effect of any probable variable, as food processing or cooking, over the exposure assessment outputs. 5) To evaluate the novel biomarker ratio sphinganine (Sa) : sphingosine (So) as an indicator of exposure to fumonisins. The global design consisted in one hand, of a nutritional study and in the other, of an analytical study. Food samples were selected to be the most susceptible commodities to mycotoxin contamination and to be commonly consumed in Catalonia. Between 2008 and 2009 the samples were obtained in six hypermarkets and supermarkets from twelve main cities (Tortosa, Tarragona, Reus, Vilanova i la Geltrú, 18 Chapter 1. Summary l’Hospitalet de Llobregat, Barcelona, Terrassa, Sabadell, Mataró, Girona, Manresa and Lleida) of Catalonia, Spain, representative of 72 % of the population. Almost
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