Development of a 32P-Postlabelling/HPLC Method for Detection of Dehydroretronecine-Derived DNA Adducts In Vitro and In Vivo

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Title:	Development of a 32P-Postlabelling/HPLC Method for Detection of Dehydroretronecine-Derived DNA Adducts In Vitro and In Vivo
Authors:	Ya-Chen Yang;Jian Yan;Mona Churchwell;Richard Beger;Po-Cheun Chan;Daniel R.Doerge;Peter P. Fu;Ming W. Chou
Contributors:	保健營養生技學系
Date:	2001
Issue Date:	2009-10-29 07:18:33 (UTC+0)
Abstract:	Pyrrolizidine alkaloids are naturally occurring genotoxic chemicals produced by a large number of plants. Metabolism of pyrrolizidine alkaloids in vivo and in vitro generates dehydroretronecine (DHR) as a common reactive metabolite. In this study, we report the development of a 32P-postlabeling/HPLC method for detection of (i) two DHR−3‘-dGMP and four DHR−3‘-dAMP adducts and (ii) a set of eight DHR-derived DNA adducts in vitro and in vivo. The approach involves (1) synthesis of DHR−3‘-dGMP, DHR−3‘-dAMP, and DHR−3‘,5‘-dG-bisphosphate standards and characterization of their structures by mass and 1H NMR spectral analyses, (2) development of optimal conditions for enzymatic DNA digestion, adduct enrichment, and 32P-postlabeling, and (3) development of optimal HPLC conditions. Using this methodology, we have detected eight DHR-derived DNA adducts, including the two epimeric DHR−3‘,5‘-dG-bisphosphate adducts both in vitro and in vivo.
Relation:	Chemical Research in Toxicology, 14(1):91–100.
Appears in Collections:	[Department of Food Nutrition and Healthy Biotechnology] Journal Article

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