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Omzetting van polluenten in maag-darm systeem Tom Van de Wiele, PhD LabMET Laboratory of Microbial Ecology and Technology Ghent University Chemicals in People Studienamiddag TI-KVIV 15 mei 2006 1 Human exposure to pollutants Dermal contact Inhalation Isolation foam, pesticides... Paints, solvents Smoking, dust... NOx, ozone, VOC Ingestion Contaminated food / soil Dust particles PBDE, PCB, PAH, heavy metals... 2 Oral exposure to pollutants Food: Soy and hop isoflavones Heterocyclic aromatic amines from grilled meat Mycotoxins ... Environment: Soil ingestion Inhalation of dust and subsequent ingestion Flame retardants in house ... 3 Human health risk assessment Biological availability What fraction of the pollutant reaches the blood circulation? Biological activity What fraction of the pollutant causes toxicity in target organs? 4 What happens to ingested pollutants? 1 2 L I V E R 3 Release from matrix Complexation to organic matter BIOACCESSIBILITY Intestinal absorption 4 6 5 Biotransformation BIOAVAILABILITY 5 What happens to absorbed pollutants ? Liver and intestinal epithelium cells: Biotransformation reactions (phase I and II) Make compound more hydrophilic Removal from body in urine or bile DETOXIFICATION But: Biotransformation sometimes goes wrong Dead-end metabolite may be formed Higher toxicity than parent compound TOXIFICATION 6 What happens to non-absorbed pollutants ? Colon ascendens, colon transversum, colon descendens Non-absorbed pollutants, detoxified pollutants... enter the large intestine Vast microbial community 500 species, 1014 CFU/mL 7 Colon microbiota and health Further digestion Useful fatty acids Vitamin K, B12 Immunostimulation Health-promoting metabolic conversions Pathogens Formation of toxins Fat uptake and synthesis Production of (geno)toxic metabolites 8 How to study intestinal microbiota? In vivo studies: animals, humans (if possible) Most relevant Physiological factors taken into account But: Black box No mechanistic studies Ethical constraints Costly ! 9 How to study intestinal microbiota? In vitro studies: simulation of the gut Not physiologically accurate Validation in vivo needed But: Mechanistic studies Reproducible Microbial community from entire gut Metabolism of chemicals can be monitored 10 SHIME-Tec: gastrointestinal in vitro technology Simulator of the Human Intestinal Microbial Ecosystem 11 Twin SHIME : parallel treatment and control 12 Case study. Oral exposure to PAH Polycyclic Aromatic Hydrocarbons Ingestion of contaminated soil Industrial and urban areas Atmospheric deposition of PAH: 50 g.ha-1.yr-1 Oral uptake Adults: 50 mg.d-1 Children: 200 mg.d-1 Occasionally: 1-20 g.d-1 Recreation area Zelzate: 49.1 mg PAH/kg DW Human health risk assessment Focus on intestinal absorption and bioactivation by human enzymes Colon microbiota contribute to toxicity? If so: incorporate in risk assessment ! 13 Experimental set-up Incubate in SHIME: • pure PAH compounds • PAH contaminated soil Stomach Small Colon intestine • Check PAH release from soil matrix along the gut •If higher release > higher risk ? • Check biological activation of PAHs •Screening for hydroxylated PAH metabolites •Chemical analysis: LC-ESI-MS •Biological analysis: yeast estrogen bioassay 14 SHIME: colon microbiota activate PAHs Stomach Small intestine Colon Inactivated colon 3,00 nM EE2 equivalence 2,50 2,00 1,50 1,00 0,50 0,00 naphthalene phenanthrene pyrene benzo(a)pyrene PAH as such are not estrogenic !!! Hydroxylated PAH metabolites have estrogenic properties 15 Chemical analysis LC-ESI-MS: hydroxylation of PAHs 1-OH pyrene: 4.3 µg/L 7-OH B(a)P: 1.9 µg/L OH EE2 7-OH B(a)P Colon microbiota produce hydroxylated PAHs !!! 16 Urban playground soil: 49.1 ppm PAH PAH release estrogenicity % EE2 equivalence µg PAH/L released 25 20 15 10 5 0 stomach small intestine colon Lower release gives higher biological activity !!! 17 Biological activity assessment PAH exposure from contaminated soil ingestion Adult: 5 g PAH/d Child:50 g PAH/d Released PAHs lowest in colon, but highest bioactivity Colon microbiota convert PAH to pseudoestrogenic metabolites Relevant biological activity in vivo ? Contributes to general PAH toxicity? Van de Wiele et al. (2005) Environmental Health Perspectives 18 Other examples: Heterocyclic aromatic amines Intestinal bacteria convert procarcinogen PHIP in nonactive metabolite Detoxification mechanism Lower risk than expected 3' 4' 2' 1' 5' 6 7 CH3 8 N 6' 5 N 9 2 NH 2 N 3' Vanhaecke et al. (2006) Journal of Agricultural and Food Chemistry 4' 2' 5' 1' 6 7 CH 3 8 N 2 6' 5 N 9 N 10 NH 12 OH 11 19 Other examples: mycotoxins Conversion of zearalenone to zearalenol Increase in estrogenic properties Relationship with aetiology of cancer development 20 Other examples: phytoestrogens Gut bacteria convert isoxanthohumol to hoppein Most powerful phytoestrogen Food supplements Hormone substitution therapy Prevention of hormone related cancers (breast/prostate) Possemiers et al. (2005) Journal of Nutrition 21 Take home messages Metabolic potency from gut microbiota Identification of responsible bacteria and process conditions needed Interindividual variability ! Modulation of biological activation through dietary factors, microbial community composition... Higher than currently anticipated Consider this process for risk assessment 22 Contact information LabMET – Ghent University Coupure Links 653 B-9000 Gent [email protected] http://labMET.ugent.be/ www.shimetec.be www.food2know.be +32 9 264.59.76 23