DETECTION OF POLYCYCLIC AROMATIC HYDROCARBON DNA ADDUCTS IN HUMAN LUNG

Synchronous fluorescence spectroscopy has been combined with immunoaff inity chromatography (IAC) and HPLC to detect polycyclic aromatic hydr ocarbon (PAH)-DNA adducts and measure r-7,t-8-dihydroxy-t-9, 10-epoxy- 7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE)-DNA adducts in human tissues and cells. A monoclonal antibody (8E11) that recognizes a range of PAH -DNA adducts, but not chemically unrelated adducts, was used to prepar e IAC columns. Samples of DNA (25 from human lung and 8 positive and n egative controls) were hydrolyzed enzymically and subjected to IAC. Ad ducts captured by the antibodies and eluted in NaOH (50 mM) were analy zed for fluorescent properties. The spectral fluorescence excitation-e mission matrices suggested the presence of mixtures of PAH-DNA adducts in some of the eluates. The eluates were subsequently hydrolyzed with acid (HCI, 0.1 N, 3 hr) and re-analyzed by synchronous fluorescence s pectroscopy using a wavelength differential of 34 nm. In 6 of the 25 h uman lung DNA samples, materials with HPLC retention times identical t o benzo[a]pyrene-7,10/8,9-tetrahydrotetrol were found to have fluoresc ence characteristics indistinguishable from pyrene. Comparisons with a ppropriate standards indicated that BPDE-DNA adduct levels were betwee n 1 and 40 adducts in 10(8) unmodified nucleotides. No correlation was observed between lung DNA-adduct levels and measures of recent smokin g (serum cotinine), but tissue samples taken from different portions o f the same lungs showed variation in the DNA adduct levels detected. T his finding complicates interpretation of the data and has important i mplications for the design of future experiments.