DNA Adducts Formedin vitrofrom Fractionated Extract of Urban Air Particles

Abstract

Abstract This study is a part of an ongoing interdisciplinary project on the health effects of air pollution (Teplice Program) in the highly polluted district of Teplice, Northern Bohemia. In our previous studies we found the relationship between DNA adduct levels detected in white blood cells of selected Teplice population and personal exposure to PAH associated with respirable particles. In this pilot study we used 32P-postlabeling assay for DNA adduct detection in an in vitro model system for evaluation of genotoxic activity of fractionated urban air extractable organic matter (EOM). To identify some of the specific DNA adducts formed we coupled TLC with HPLC analysis of labeled adducts. The urban air particles were collected by high-volume sampler during January-March 1992 in Teplice. EOM was extracted by dichlormethane (DCM) and crude extract was fractionated into five fractions to obtain a gross partition of different chemical classes. Fractions were incubated with calf thymus DNA (dose 100 μg/ml incubate) under oxidative and reductive conditions using two metabolic activation system: 1) an oxidative rat liver S9 system (S9) and 2) a reductive xanthine oxidase catalyzed system (XO). The different DNA adduct patterns and levels were determined using S9 and XO-mediated metabolism followed by postlabeling with both nuclease P1 and butanol extraction enrichment procedures for all fractions examined. The moderately polar fraction (DCM) contained over 50% of the total DNA adduct forming activity both without and with S9 activation. The highly polar fraction (methanol) contained about 60 % of the DNA adduct forming activity under reductive conditions. Some of the main distinct DNA adducts obtained with S9 and XO mediated metabolism were tentatively identified by HPLC comparing with standards of PAH- and nitro-PAH DNA adducts.