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© 1991 Oxford University Press

research-article

Fluorescence detection of benzo[a]pyrene-DNA adducts in human lung

Ainsley Weston and Elise D. Bowman

Laboratory of Human Carcinogenesis, National Cancer Institute, National Institutes of Health Bethesda, MD 20892, USA

Improved techniques are described for the specific identification of benzo[a]pyrene-diolepoxide (BPDE)-DNA adducts in human tissues. Immunoaffinity chromatography, synchronous fluorescence spectroscopy and second-derivative synchronous fluorescence spectroscopy have previously been used to detect BPDE-DNA adducts in human placenta. Here we report how these methods, together with HPLC and the generation of complete fluorescence excitation—emission matrices, have been used to identify unequivocally BPDE-DNA adducts in samples of human lung. BPDE nucleotide adducts were isolated with immunoaffinity chromatography columns bearing antibodies raised against the (±)anti-7,8-diol-9,10-epoxide-deoxyguanosine adduct of betizo[a]pyrene. These adducts were hydrolyzed to tetrahydrotetrols and the hydrolysis products subjected to HPLC. The major product isolated by HPLC, benzo[a]-pyrene-7,10/8,9-tetrahydrotetrol, was determined by fluorescence spectroscopy. Using this method, levels of BPDE-DNA adducts in the range of 1–40 in 108 nucleotides were measured in 6 out of 25 samples, with a lower detection limit of one adduct in 108 nucleotides. The data may also indicate that adduct levels show regional variation in different parts of the same lung.


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