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Strategy for identifying unknown hemoglobin adducts using adductome LC-MS/MS data: Identification of adducts corresponding to acrylic acid, glyoxal, methylglyoxal, and 1-octen-3-one
Stockholm University, Faculty of Science, Department of Environmental Science and Analytical Chemistry.
Stockholm University, Faculty of Science, Department of Environmental Science and Analytical Chemistry.
2016 (English)In: Food and Chemical Toxicology, ISSN 0278-6915, E-ISSN 1873-6351, Vol. 92, 94-103 p.Article in journal (Refereed) Published
Abstract [en]

Electrophilic compounds have the ability to form adducts with nucleophilic sites in proteins and DNA in tissues, and thereby constitute risks for toxic effects. Adductomic approaches are developed for systematic screening of adducts to DNA and blood proteins, with the aim to detect unknown internal exposures to electrophiles. In a previous adductomic screening of adducts to N-terminals in hemoglobin, using LC-MS/MS, 19 unknown adducts were detected in addition to seven previously identified adducts. The present paper describes the identification of four of these unknown adducts, as well as the strategy used to identify them. Using LC-MS data from the screening, hypotheses about adduct identities were formulated: probable precursor electrophiles with matching molecular weights were suggested based on the molecular weights of the modifications and the retention times of the analytes, in combination with comparisons of theoretical Log P calculations and databases. Reference adducts were generated by incubation of blood samples with the hypothesized precursor electrophiles. The four identified precursor electrophiles, corresponding to the observed unknown adducts, were glyoxal, methylglyoxal, acrylic acid and 1-octen-3-one. Possible origins/exposure sources and toxicological information concerning the electrophilic precursors are discussed. The identified adducts could be explored as possible biomarkers for exposure.

Place, publisher, year, edition, pages
2016. Vol. 92, 94-103 p.
Keyword [en]
Adductomics, Hemoglobin adducts, Glyoxal, Methylglyoxal, Acrylic acid, 1-Octen-3-one
National Category
Chemical Sciences
Research subject
Environmental Chemistry
Identifiers
URN: urn:nbn:se:su:diva-129040DOI: 10.1016/j.fct.2016.03.028ISI: 000376804400011OAI: oai:DiVA.org:su-129040DiVA: diva2:919226
Funder
Swedish Research CouncilCancer and Allergy Foundation
Available from: 2016-04-13 Created: 2016-04-13 Last updated: 2016-07-04Bibliographically approved
In thesis
1. Development of an adductomic approach to identify electrophiles in vivo through their hemoglobin adducts
Open this publication in new window or tab >>Development of an adductomic approach to identify electrophiles in vivo through their hemoglobin adducts
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Humans are exposed to electrophilically reactive compounds, both formed endogenously and from exogenous exposure. Such compounds could react and form stable reaction products, adducts, at nucleophilic sites in proteins and DNA. The formation of adducts constitutes a risk for effects, such as cancer and contact allergy, and plays a role in ageing processes. Adducts to proteins offer a possibility to measure electrophilic compounds in vivo.

Adductomic approaches aim to study the totality of adducts, to specific biomolecules, by mass spectrometric screening. This thesis describes the development and application of an adductomic approach for the screening of unknown adducts to N-terminal valine (Val) in hemoglobin (Hb) by liquid chromatography tandem mass spectrometry (LC/MS/MS).

The adductomic approach is based on the FIRE procedure, a modified Edman procedure for the analysis of adducts to N-terminal Val in Hb by LC/MS/MS. The adduct screening was performed by stepwise scanning of precursor ions in small mass increments and monitoring four fragments common for derivatives of detached Val adducts, in the multiple reaction monitoring mode. Samples from 12 smokers/nonsmokers were screened with the adductomic approach, and seven previously identified adducts and 19 unknown adducts were detected. A semiquantitative approach was applied for approximate quantification of adduct levels.

A strategy for identifying unknown Hb adducts using adductome LC/MS/MS data was formulated and applied for the identification of unknown adducts. Identifications were based on the observed m/z of precursor ions and retention times combined with databases and Log P calculations. Hypothesized adducts were generated in vitro for comparison and matching with the corresponding unknown adducts. Five identified adducts correspond to the precursor electrophiles ethyl vinyl ketone (EVK), glyoxal, methylglyoxal, acrylic acid, and 1-octen-3-one. These adducts, except the adducts corresponding to glyoxal and methylglyoxal, have not been observed as protein adducts before.  Probable exposure sources to these electrophiles are diet and/or endogenous formation. The observation of these adducts motivate further studies to evaluate possible contributions to health risks, as well as their potential as biomarkers of exposure.

The adduct from EVK was quantitatively assessed through different experiments to estimate the daily internal dose (area under the concentration-time-curve, AUC). EVK is about 2 × 103 more reactive than the reference compound acrylamide. The EVK adduct was shown to be unstable, with a relatively short half-life. The daily AUC in humans of EVK was estimated to be about 20 times lower than the corresponding AUC of acrylamide from intake via food.

To confirm the observation of the detected unknown adducts and obtain a statistical foundation, analysis of unknown adducts were performed in large sets of blood samples (n = 50–120) from human cohorts. The majority of the previously detected unknown adducts were found in all analyzed samples, and the levels of many adducts showed large variations between individuals. The cause and significance of these observed variations are not yet clarified, but are of importance for the directions of future studies.

In conclusion, a new approach for identification of unknown human exposure to electrophiles was developed and successfully applied. 

Place, publisher, year, edition, pages
Stockholm: Department of Environmental Science and Analytical Chemistry, 2016. 87 p.
Keyword
Adductomics, Hemoglobin adducts, LC/MS/MS, Ethyl vinyl ketone, Glyoxal, Methylglyoxal, Acrylic acid, 1-Octen-3-one
National Category
Other Chemistry Topics Analytical Chemistry
Research subject
Environmental Chemistry
Identifiers
urn:nbn:se:su:diva-129248 (URN)978-91-7649-348-9 (ISBN)
Public defence
2016-06-10, William-Olssonsalen, Geovetenskapens hus, Svante Arrhenius väg 14, Stockholm, 10:00 (English)
Opponent
Supervisors
Funder
Cancer and Allergy FoundationSwedish Research Council
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Submitted.

Available from: 2016-05-18 Created: 2016-04-18 Last updated: 2016-05-06Bibliographically approved

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Carlsson, HenrikTörnqvist, Margareta
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