Photochemical transformation and phototoxicity of 1-aminopyrene

Abstract

1-Aminopyrene (1-AP) is an environmental mutagen and a metabolite of 1-nitropyrene (1-NO2P). On light irradiation, 1-AP transforms into oxidation products with a half-life of 7.1 min in 10% methanolic buffer. The presence of DNA or free-radical/ singlet oxygen scavengers 1,4-dithiothreitol, histidine, or NaN3 slows down 1-AP photochemical reaction. The photoproducts identified include 1-hydroxyaminopyrene, 1-nitrosopyrene, 1-NO2P, 1-amino-x-hydroxypyrene, and three covalent dimers. Since it is known that 1-NO2P and 1-nitrosopyrene are genotoxic and 1-hydroxyaminopyrnene can react with DNA to form covalent adducts, we used the Mutatox test to assess the toxicity of 1-AP and its photoproducts. It was found that the lowest-observed-effect concentrations for 1-AP, 1-AP photoproducts, and 1-NO2P are 1.25 microM, 10 microM, and NA (no mutagenic response was seen at this concentration range) in direct medium (no S-9) and NA, 5 microM, and 0.625 microM in S-9 medium, respectively. Therefore, 1-AP photoproducts are more genotoxic than 1-AP itself in the S-9 medium and more mutagenic than 1-NO2P in the direct medium. Thus, 1-NO2P alone cannot account for all the mutagenicity of the photoproducts. Irradiation of 1-AP together with DNA leads to covalent DNA adduct formation possibly via the 1-hydroxyaminopyrene intermediate. In this study, ultraviolet-A (UVA) was used at approximately the same magnitude as the outdoor UVA irradiance. Considering the half-life of 1-AP in the test solutions in this study, the aquatic biota (including humans) near the surface layer of a static water body are most likely subjected to the photoinduced toxicity of the study compound. The biota at the lower depths will also be affected if turbulence becomes a significant factor in enhancing the exposure risk for aquatic organisms.

Figure 1

UV-Vis spectra for transformation of 1-aminopyrene in 10% methanolic buffer containing 40% acetonitrile monitored by UV-Vis absorption spectra. A solution of 50 μM 1-AP was irradiated with the UVA lamp and samples were taken at desired time intervals for recording of the absorption spectra.

Figure 2

HPLC chromatograms of 1-aminopyrene photo-transformation. A solution of 10 μM 1-aminopyrene in 30% acetonitrile/PBS buffer was irradiated with a UVA lamp. Samples were taken at 0, 5, 10, 20, 40, 80, 160 min and analyzed with HPLC.

Figure 3

Mass spectra of 1-aminopyrene and its photoproducts detected by LCQ^DUO LC-MS using the APCI ionization mode. Except for 1-aminopyrene, which was detected in the positive ion mode, all others were detected in the negative ion mode.

Figure 4

Absorption spectra of DNA covalent adduct of 1-AP. A solution of 30 μM 1-AP in 200 μM calf thymus DNA was irradiated with UVA light for 1 h. Its absorption spectrum (line # 2) is compared with the same solution before UVA irradiation (line #1), and the same solution after dialysis with semi-permeable membrane (MWCO 13,000) for 24×3 hours against the same buffer solution (line #3). Line #4 is the control.

Figure 5

Photochemical transformation pathways of 1-aminopyrene