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Characterization of alkyl-cobalamins formed in trapping of epoxide metabolites of 1,3-butadiene
Stockholm University, Faculty of Science, Department of Environmental Chemistry.
Stockholm University, Faculty of Science, Department of Materials and Environmental Chemistry (MMK), Environmental Chemistry.
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2004 (English)In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 27, no 7-8, 607-612 p.Article in journal (Refereed) Published
Abstract [en]

Analytical methods facilitating studies of electrophilically reactive and genotoxic compounds in vitro and in vivo are needed. The strong nucleophile, cob(I)alamin, formed by reduction of Vitamin B12 [cob(III)alamin], may be used for trapping and analysis of 1,2-epoxides and other electrophiles. In the present study, cob(I)alamin is evaluated as an analytical tool for 1,2-epoxide metabolites (oxiranes) of 1,3-butadiene. Products of reaction of cob(I)alamin with 1,2-epoxy-3-butene (EB), 1,2:3,4-diepoxybutane (DEB), and 1,2-epoxy-3,4-butanediol (EBdiol) have been analyzed by reversed phase high performance liquid chromatography (HPLC) coupled on-line to electrospray ionization mass spectrometry (ESI-MS) and ultraviolet diode array detection (UV-DAD). It was shown that a specific alkyl-Cbl complex is formed for each metabolite and that it was possible to discriminate between the products by HPLC-UV and by LC-MS. Quantification of DEB with the method by use of another 1,2-epoxide as an internal standard was successfully performed. The possibility of using cob(I)alamin for trapping and analysis of the three oxirane metabolites of 1,3-butadiene will facilitate quantitative comparisons of species in vitro with regard to metabolism of 1,3-butadiene.

Place, publisher, year, edition, pages
2004. Vol. 27, no 7-8, 607-612 p.
Keyword [en]
1, 3-Butadiene • Diepoxybutane • Cobalamin • Reactive metabolites • LC-MS
National Category
Environmental Sciences
URN: urn:nbn:se:su:diva-25174DOI: 10.1002/jssc.200301689OAI: diva2:199062
Part of urn:nbn:se:su:diva-78Available from: 2004-03-25 Created: 2004-03-25 Last updated: 2010-07-30Bibliographically approved
In thesis
1. Methods for measurement of reactive metabolites as a basis for cancer risk assessment: Application to 1,3-butadiene and isoprene
Open this publication in new window or tab >>Methods for measurement of reactive metabolites as a basis for cancer risk assessment: Application to 1,3-butadiene and isoprene
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

1,3-Butadiene is a general air pollutant associated with combustion of organic matter and is also an extensively used monomer in polymer production. The cancer risk estimation of 1,3-butadiene is encumbered with large uncertainties. Extrapolation from tumour frequencies in long-term animal tests has led to a relatively high figure for the risk associated with 1,3-butadiene exposure. This is mainly based on observations of very high tumour incidences in butadiene-exposed mice, which in this respect are about 100 times more sensitive than rats. It has been hypothesized that a high cancer risk from 1,3-butadiene could be associated with its metabolism to the bifunctional 1,2:3,4-diepoxybutane (DEB) which, in comparison with monofunctional epoxides, 1,2-epoxy-3-butene (EB) and 1,2-epoxy-3,4-butanediol (EBdiol), is a highly effective mutagen, i.e. cancer initiator. Measurement of in vivo doses of DEB is therefore essential for the risk assessment of 1,3-butadiene. Reaction products with hemoglobin offer a possibility of measuring reactive metabolites in vivo. Hemoglobin adducts from EBdiol have in this study been measured with available methods, which are, however, not applicable to the bifunctional DEB, and method development was therefore needed.

This work presents a procedure for measurement of a specific, ring-closed adduct, Pyr-Val, formed from the reaction of DEB with N-terminal valines in hemoglobin. It is based on LC-ESI-MS/MS analysis of the Pyr-modified N-terminal peptides enriched after trypsin digestion of globin. Mouse and rat could be compared regarding the metabolism of EB, DEB and EBdiol. From the data it was concluded that, in 1,3-butadiene exposure, about 60 times higher levels of DEB are formed in mice compared to rats. Estimates of in vivo doses in published cancer tests showed that carcinogenesis in mice is mainly due to DEB, whereas in rat, and possibly man, the monofunctional EBdiol is the predominant causative factor. Preliminarily, the cancer risk assessed from these data is compatible with the epidemiology-based risk estimate of US EPA.

Due to the structural similarity with 1,3-butadiene, certain parallel studies of isoprene (2-methyl-1,3-butadiene) metabolism were initiated. Isoprene is the major endogenously produced hydrocarbon in humans and mammals and shows a similar difference in sensitivity between species for tumour development as 1,3-butadiene. In mice treated with the isoprene monoepoxide, 1,2-epoxy-2-methyl-3-butene (IMO), an in vivo formation of the corresponding diepoxide, 1,2:3,4-diepoxy-2-methyl-butane (IDO), was demonstrated. The in vivo dose of IDO formed from IMO was about half of that of DEB formed from EB. In the analysis of bone marrow erythrocytes an increased frequency of micronuclei, induced by treatment with EB or IMO, showed correlation with the in vivo doses of the respective diepoxides.

With the ambition to reduce animal experiments a general procedure has been developed for trapping reactive metabolites in in vitro test systems, with the specific aim to study differences between species in metabolism of 1,3-butadiene. Vitamin B12 in its reduced form [Cbl(I)] has been used for instant trapping of 1,3-butadiene metabolites formed in S9-mixture. LC-ESI-MS/MS is then used for quantification of the formed alkyl-Cbls. The method has been applied to the epoxide metabolites of 1,3-butadiene, which all form specific alkyl-Cbls in the reaction with Cbl(I)

Place, publisher, year, edition, pages
Stockholm: Institutionen för miljökemi, 2004. 84 p.
National Category
Environmental Sciences
urn:nbn:se:su:diva-78 (URN)91-7265-840-1 (ISBN)
Public defence
2004-04-15, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 12 A, Stockholm, 13:00
Available from: 2004-03-25 Created: 2004-03-25Bibliographically approved

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Alsberg, ThomasRydberg, PerMinten, JohannaTörnqvist, Margareta
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