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Doctoral thesis, 2017

New alkylresorcinol metabolites : identification, quantification and evaluation as dietary biomarkers

Wierzbicka, Roksana

Abstract

Alkylresorcinols (AR) are amphiphilic phenolic lipids that are extensively metabolized in the liver and form metabolites that can be detected in plasma and urine. Two AR metabolites, 3,5-dihydroxybenzoic acid (DHBA) and 3-(3,5-dihydroxyphenyl)-propanoic acid (DHPPA), have been evaluated as urinary biomarkers of whole grain wheat and rye intake. AR metabolite concentrations are currently measured by chromatographic techniques but for samples from large epidemiological studies, immunoassay techniques are preferred due to rapid and cheap analysis. The aim of this thesis was to develop high-throughput analytical methods for the analysis of alkylresorcinol metabolites and to evaluate recently discovered putative metabolites 3,5-dihydroxycinnamic acid (DHCA), 5-(3,5-dihydroxyphenyl) pentanoic acid (DHPPTA), 3,5-dihydroxycinnamic acid amide (DHCA-amide) and 2-(3,5-dihydroxybenzamido) acetic acid (DHBA-glycine) as biomarkers of whole grain wheat and rye intake. Polyclonal antibodies against DHBA and DHPPA were produced and characterized in terms of cross-reactivity, assay sensitivity, precision and accuracy. A developed ELISA method was used for analysis of urine samples and results were compared with GC-MS. A lack of agreement was found. Several putative AR metabolites were identified by LC-MS/MS, including DHPPTA, DHCA, DHBA-glycine and DHCA-amide. These compounds showed high cross reactivity in ELISA and likely explains the lack of agreement between the two methods. The medium-term reproducibility and relative validity of DHCA, DHPPTA and DHBA-glycine were found to be moderate to excellent in 24h urine collections. Moreover, the reproducibility was good for these metabolites also in spot-urine samples. Age and sex were found to be important determinants besides whole grain intake. The results suggest that determination of AR metabolites in 24h urine and single or duplicate spot urine samples appear to be suitable medium to long-term biomarkers of whole grain wheat and rye intake. These findings need to be confirmed in other populations before using these biomarkers in epidemiological studies.

Keywords

Alkylresorcinol; ELISA; antibodies; dietary biomarker; AR metabolites; whole grain

Published in

Acta Universitatis Agriculturae Sueciae
2017, number: 2017:20
ISBN: 978-91-576-8813-2, eISBN: 978-91-576-8814-9
Publisher: Department of Molecular Sciences, Swedish University of Agricultural Sciences

    UKÄ Subject classification

    Immunology

    Permanent link to this page (URI)

    https://res.slu.se/id/publ/79668