An applied synchronous fluorescence spectrophotometric assay to study benzo[a]pyrene-diolepoxide-DNA adducts

Synchronous scanning fluorescence with a fixed wavelength difference() of 34 nm between excitation and emission was used to quantitatebenzo[a]pyrene-diol epoxide (BPDE)-DNA adducts. Fluorescenceemission maxima occurred at 382 nm for BPDE-DNA and at 379 nmfor benzo[a]pyrene-tetrols and -triol, which are hydrolysisproducts of BPDE. Similarly, the peak for pyrene was at 372nm and for 1-nitropyrene at 386 nm. The minimum detectable amountof BPDE-moieties in in vitro modified BPDE-DNA, after hydrolysisin HCl, was 20 fmol in 100 µg of DNA, which is equivalentto 1 adduct per 1.4 x 10⁷ nucleotides. The correlation of fluorescenceintensity and the amount of BPDE-moieties was linear between20 fmol and 1 pmol. DNA isolated from human lymphoblasts incubatedwith BDPE had the same fluorescence peak and the correlationbetween the fluorescence intensity and the amount of BPDE inthe incubation mixture was linear. Among the DNA-samples fromperipheral blood lymphocytes of 30 aluminum plant workers, onlyone sample was found to contain a peak similar to BPDE-DNA.None of the DNA-samples from 10 persons not occupationally exposedwere positive. Measurement of BPDE-DNA adducts by synchronousfluorescence spectrophotometry should be useful in monitoringhuman exposure to benzo[a]pyrene.