of Work: Photosensitization can result when light interacts with endogenous or exogenous chemical agents in the skin and eyes. This process can produce undesirable clinical consequences, such as phototoxicity (exaggerated sunburn), photoallergy, photocarcinogenicity and/or photomutagenicity ; or it can have beneficial effects as in tumor photodynamic therapy (PDT) and coal tar, anthralin or psoralen (PUVA) therapy for psoriasis. The objective of this research project is to elucidate the photochemical mechanisms whereby photosensitizers exert their toxic or therapeutic effects and to design appropriate in vitro or in vivo tests to detect photosensitizers. Fluoroquinolones (FLQ) are a relatively new class of antibacterials that are useful in the treatment of gram-negative and gram-positive bacterial infections. When used in humans FLQs often cause phototoxicity. Recent studies by others have shown that lomefloxacin (LOME) and fleroxacin (FLER) cause squamous cell carcinomas in HR/J mice injected with these drugs and irradiated with UV-A (315-400nm). However, we have been unable to demonstrate the photocarcinogenicity of UVA/LOME in the TgAC transgenic mouse after 12 weeks of exposure, even though the skin levels of LOME in these mice (~2ug/g) were the same or higher than those obtained in the HR/J animals. We have repeated this experiment with 8-methoxypsoralen/UVA and found that, while the TgAC mouse does develop papillomas, it does so more slowly than the HR/J mouse. Goldenseal is a herb, widely used for numerous medical applications including eyewashes and skin lotions, which is currently undergoing testing by the National Toxicology Program. We have studied the photochemistry and photobiology of berberine and hydrastine, the two most abundant Goldenseal alkaloids and minor alkaloids canadine, palmatine and hydrastinine. Berberine/UVA and hydrastine/UVA damage keratinocyte DNA as measured by the Comet assay. Canadine and palmatine were less effective. Neither berberine nor hydrastine photosensitizes singlet oxygen in aqueous solutions, but singlet oxygen is produced by berberine with a quantum yield 0.34 in methylene chloride. This suggests that a hydrophobic environment is required for photosensitization. Our data show that, although a weak singlet oxygen photosensitizer in water, berberine is able to produce reactive oxygen species in a hydrophobic environment. Thus DNA or membrane damage by berberine may involve a photodynamic mechanism. Juglone (5-hydroxy-1,4-naphthoquinone) is present in black walnuts and is a component of dietary supplements, hair dyes and walnut oil stains. Crushed black walnut hulls are sometimes applied to the skin to treat fungal, bacterial or viral infections. Juglone is currently undergoing study by the National Toxicology Program. We have found that juglone is toxic to keratinocytes which metabolize it to a semiquinone free radical. Reaction of the latter with oxygen generates superoxide and the hydroxyl radical. None of these reactions was enhaced by UV radiation. The sunscreen agent, 2-phenylbenzimidazole-5-sulfonic acid (PBSA), and its parent 2-phenylbenzimidazole (PBI) cause DNA photodamage via both Type I and Type II mechanisms when UVB irradiated. During UV irradiation of an aerobic solution of PBSA, superoxide and hydroxyl radicals were generated and trapped by 5,5-dimethyl-1-pyrroline N-oxide (DMPO). PBSA showed strong oxidizing properties when UV irradiated in neutral aqueous solution (pH 7.4) in the presence of cysteine, glutathione and azide, as evidenced by the detection of S-cysteinyl, glutathiyl and azidyl radicals with the aid of the spin trap DMPO. PBI and PBSA generated singlet oxygen with quantum yields 0.07 and 0.04 respectively in MeCN. Our studies demonstrate that UV irradiation of PBSA and PBI generates a variety of free radicals and active oxygen species that may be involved in the photodamage of DNA.
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