1-Hydroxypyrene (HOP), a metabolite found in the urine of humans and laboratory animals exposed to polycyclic aromatic hydrocarbons (PAHs), is known to be both acutely toxic and genotoxic. It has been widely used as a biomarker for studying PAH exposure. In this research, we have found that, upon UVA irradiation, HOP causes DNA single-strand cleavages and forms HOP-DNA covalent adducts. The UVA-induced cleavage of supercoiled plasmid PhiX174 DNA is dependent upon both HOP concentration and UVA dosage. A longer irradiation time or higher HOP concentration induces more DNA cleavage. Results of the photocleavage experiments carried out in the presence of reactive oxygen species scavengers, histidine, sodium azide, mannitol, SOD, and desferal indicate that both the superoxide free radical and singlet oxygen are likely involved in causing DNA single-strand cleavage. The photocleavage is inhibited by the presence of an excited singlet-state quencher, KI, indicating that it is an excited-state reaction. Along with light-induced DNA cleavage, HOP also forms DNA covalent adducts while being degraded upon light irradiation. Light-induced degradation of 20 microM HOP follows first-order reaction kinetics in a 10% methanolic buffer (10 mM phosphate) solution in the absence or presence of 40 microM calf thymus DNA, with degradation half-lives of 20 or 15 min, respectively. The shorter degradation half-life in the presence of DNA is due to the formation of the HOP-DNA covalent adduct. The formation of the HOP-DNA covalent adduct is evidenced by comparing the UV-vis absorption and fluorescence emission spectra of the pure HOP with those of the HOP-DNA adduct. The covalent HOP-DNA adduct produced due to irradiation was purified by either extensive dialysis (3 x 500 mL buffer solutions), phenol and chloroform extraction followed by ethanol precipitation, or chloroform extraction alone. The isolated HOP-DNA adduct has an absorption peak at 353 nm, which is 8 nm red-shifted compared to that of free HOP. The fluorescence emission for HOP-DNA is at least 70 times weaker than that for free HOP in solution. In summary, the findings with HOP reveal that, in addition to metabolic activation that eventually leads to the formation of alkylated DNA adducts or other forms of DNA damage, HOP may be activated by light to produce DNA single-strand cleavage and covalent DNA adducts. These DNA lesions can be sources of toxicity.

No abstract

UVA light-induced DNA single strand cleavage by a set of 12 monomethyl substituted benz[a]anthracenes (MBAs) along with their parent compound, benz[a]anthracene (BA), and the potent carcinogen, 7,12-dimethylbenz[a]anthracene (DMBA), was studied. On the basis of the relative DNA single strand photocleavage efficiency of the fourteen compounds, they are divided into three groups: (1) strong DNA cleavers, 4-MBA, 5-MBA, 6-MBA, 8-MBA, 9-MBA, 10-MBA, and BA; (2) medium DNA cleavers, 1-MBA, 2-MBA, 3-MBA, and 11-MBA; and (3) weak DNA cleavers, 7-MBA, 12-MBA, and DMBA. The relative DNA photocleavage efficiency parallels very well with the energy gap between the highest-occupied-molecular-orbital (HOMO) and the lowest-unoccupied-molecular-orbital (LUMO) of each MBA, indicating that the DNA cleavage is related to their excited-state properties. The 7 and 12 positions of BA are two unique sites. Methyl substitution at either 7 or 12 (or both) positions lowers the HOMO-LUMO gap and greatly diminishes the DNA photocleavage efficiency. UVA light-induced photodegradation of selected MBAs reveals that methyl substitution at either 7 or 12 (or both) positions greatly enhances the degradation rate. Photodegradation of 7-MBA, 12-MBA, and DMBA yields products that are much less effective in mediating DNA cleavage. Photodegradation of other MBAs, exemplified by 5-MBA, yields a photooxidation product 5-MBA-7,12-quinone which is relatively stable under light and is a stronger DNA photocleaver than 5-MBA itself. The higher efficiency of DNA photocleavage for MBAs with methyl substitution at positions other than 7 or 12 is due, at least in part, to the formation of 7,12-quinone. Light-induced DNA single strand cleavage efficiency for several MBAs parallels the light-induced toxicity observed by other research groups, suggesting that light-induced DNA cleavage of MBAs are the source for phototoxicity. Since some PAHs such as coal tar are used commercially as creams, therapeutic agents, or ointments, or those roofers and asphalt workers that are subject to contamination with PAHs, the combination of PAHs and light (in the skin) may present a greater health risk to humans.

No abstract

No abstract