ESR (electron spin resonance) spectra of a fatty acid spin probe (16-doxylstearic acid, 16-DS) incorporated into an aqueous surfactant system composed of oleic acid and oleate molecules were measured between 10 and 50 °C up to a total oleic acid + oleate concentration of 50 mM. Depending on the total concentration and the pH, different types of oleic acid/oleate aggregates formed. At the two ends of the pH range investigated (above pH 10.4 and below pH 6.4), the ESR spectra of 16-DS were highly symmetric, enabling calculation of the microviscosities in the surfactant aggregates to be 4 cP and 6 cP, respectively. In the high pH range, the observed aggregates are micelles. On the other hand, in the low pH range the microviscosity was considerably lower than that of neat oleic acid (measured to be 11 cP), indicating that the obtained emulsion system was not composed of pure oleic acid droplets. We postulate that the surfactant molecules at low pH form condensed aggregates of lamellar bilayers. Asymmetric high-field ESR lines were obtained at intermediate pH between pH 6.4 and pH 10.4. This indicates that the probe molecules were present in two physically different aggregation states. We assigned the two aggregation states to be vesicles and nonlamellar aggregates (most likely nonspherical micelles), based on the observation made by microscopy and light scattering techniques. The analysis of the ESR lines by spectral simulation using a modified Bloch equation supports the coexistence of vesicles and nonlamellar aggregates through the entire intermediate pH range; the relative amount of the two aggregation forms depends critically on pH, temperature, and concentration. Furthermore, the spectral simulation indicated that particularly stable oleic acid/oleate vesicles are formed around pH 8.5, where the protonated and ionized species exist in a stoichiometric ratio.

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DNA damage induced by solar ultraviolet (UV) radiation plays an important role in the induction of skin cancer. Although UVA constitutes the majority of solar UV radiation, it is less damaging to DNA than UVB. The DNA damage produced by UVA radiation, however, can be augmented in the presence of a photosensitizer. We previously used benzo[a]pyrene (BaP), an environmental carcinogenic polycyclic aromatic hydrocarbon, as an exogenous photosensitizer, and demonstrated that combined exposure to BaP and UVA resulted in DNA double-strand breaks (DSBs) in cultured Chinese hamster ovary (CHO-K1) cells. In this study, we investigated whether coexposure to BaP and UVA induces DSBs in a cell-free system and whether reactive oxygen species (ROS) were involved in the generation of the DSBs. DSBs were induced by the coexposure both in the cell-free system (in vitro) and in CHO-K1 cells (in vivo), but not by treatment with BaP or UVA alone. DSB induction in vitro required higher doses of UVA and BaP than were required in vivo, suggesting that the mechanism of DSB induction differed. A similar difference in efficiency also was observed in the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) by coexposure to BaP and UVA in vitro and in vivo. A singlet oxygen ((1)O2) scavenger (NaN3) effectively inhibited the production of DSBs and 8-oxodG, suggesting that (1)O2 is a principal ROS generated by BaP and UVA both in vitro and in vivo. Furthermore, repair-deficient xrs-5 cells were more sensitive to coexposure with BaP and UVA than were CHO-K1 cells, but the two cell lines were equally sensitive to the combined treatment in the presence of NaN3. This result suggested that the cell death produced by coexposure to BaP and UVA was at least partly due to the DSBs generated by (1)O2. Our findings indicate that coexposure to BaP and UVA effectively induced DNA damage, especially DSBs, which results in phototoxicity and possibly photocarcinogenesis.

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