Tetrahydroxylated-benzo[a]pyrene isomer analysis after hydrolysis of DNA-adducts isolated from white blood cells

Abstract

On the assumption of associations between exposure to benzo[a]pyrene and certain health disorders, several recent research studies have been dedicated to the development of strategies aimed at assessing human exposure to this ubiquitous pollutant. As part of these research efforts, a method was designed for the analysis of four tetrahydroxylated-benzo[a]pyrene isomers resulting from the hydrolysis of their respective diol-epoxide precursors which are involved in DNA-adduct formation. The analytical sensitivity necessary to reach environmental levels of concentration was obtained by means of gas chromatography–tandem mass spectrometry. The suitability of the method was firstly evaluated by the analysis of DNA isolated from white blood cells of human volunteers. The results obtained confirm that this method was sufficiently sensitive to detect environmental exposure levels as all the samples measured were above the limit of quantification for benzo[a]pyrene-r-7,t-8,t-9,c-10-tetrahydrotetrol(±) and two of them tested positive for benzo[a]pyrene-r-7,t-8,c-9,c-10-tetrahydrotetrol(±). The method was subsequently applied to the analysis of DNA isolated from white blood cells of rats submitted to benzo[a]pyrene-controlled exposure. The study led to the detection of the four targeted tetra-OH-benzo[a]pyrenes as well as of two unknown isomers in all benzo[a]pyrene-treated animals. Interestingly, benzo[a]pyrene-r-7,t-8,c-9,c-10-tetrahydrotetrol(±) proved to be the most abundant isomer in both treated and control animals, followed by benzo[a]pyrene-r-7,t-8,t-9,c-10-tetrahydrotetrol(±); which points out to interspecies differences in the nature of the tetrahydroxylated-benzo[a]pyrene isomers formed. These results highlight the need to focus on all the tetrahydroxylated-benzo[a]pyrene isomers, which could be indicators of benzo[a]pyrene-associated toxicity related to an individual's own metabolism, rather than limit to a single form.