.

“Exposure” VS “Exposome” Exposure assessment of most epi-studies is limited to a single or a small group of toxicants. Insufficient studies to observe developmental stages. Ref) Rappaport 2011

Can influence health here What happens here Prenatal programming of adult disease

Challenges to studying fetal chemical programming Overcome Limitations 1) Longitudinal birth cohort study : - time consuming and expense - measurements of maternal biomarkers (not fetal biomarkers) - cord blood is good matrix but largely detect long half life chemicals and provide only limited development period (over 3rd trimester) * New human matrix - retrospective - objective - capable of directly measuring fetal exposures - provide time-series exposure data

Tooth provide time series exposure

Key aspects of methodology Participants: 5 children (7 ~ 10 years old) Collected matrix: deciduous teeth Participants A B C D E 2nd, 3rd trimester dentine Prenatal (2nd, 3rd trimester), postnatal dentine

Key aspects of methodology Used equipment: QTOF-LC/MS (Agilent 6550 iFunnel QTOF with Jet Stream) Purpose: global screening of small molecules

Classification of qualified compounds 1. Known unknowns (Database OK, reference standard OK) 2. Suspected unknowns (Database OK, no reference standard) 3. Unknown unknowns (no database, no reference standard)

Multi-chemical exposure profiles during prenatal and early childhood periods Targeted analysis of organic compounds and metabolites accumulated in dentine formed during specific period

Bisphenol A (First report) Mono-methyl phthalate (First report) Mono-ethyl phthalate (First report) Mono-butyl phthalate (First report) Mono-benzyl phthalate (First report) Mono-ethylhexyl phthalate (First report) Hydroxycotinine (First report) Cotinine Nicotine

For suspected unknowns: Compound matching with literature for prenatal and children’s exposures to environmental factors (Bellinger 2013; Gonzalez-Alzaga and others 2014; Lyall and others 2014; Meeker 2012).

Multi-chemical exposure profiles during prenatal and early childhood periods Single measures of conventional biomarkers or use of whole teeth are vulnerable to assess exposome BPA

Multi-chemical exposure profiles during prenatal and early childhood periods Untargeted analysis of organic compounds and metabolites accumulated in dentine formed during prenatal period (Child C)

1) Molecular Formula Generation (MFG) algorithm to generate chemical formulas of different peaks

2) MassHunter Profinder was used to perform Batch Recursive Feature Extraction (BRFE)

3) Compounds selection Selected based on ion intensity in each tooth (n=100) Comparisons were made for inter/intra child variablilty in nontargeted analysis profiles. N = 309 N = 225

Metabolites pattern between pre- and post-natal samples for each child and between children

Conclusion and perspectives Similar methods can be applied to permanent/adult teeth for the study of adult disease outcomes.

The computational tool For targeted analysis - Personal Compound Database (PCD) - Find by Formula compiled by Agilent Tech. For untargeted screening - Molecular Feature Extraction (MFE) - Batch Recursive Feature Extraction (BRFE)