Andrée-Anne Chételat

Photomutagenesis test development: I. 8-methoxypsoralen, chlorpromazine and sunscreen compounds in bacterial and yeast assays

Two in vitro genotoxicity tests have been adapted to the evaluation of photomutagenic activity of test compounds. The study was initiated to obtain an experimental basis relating to newly proposed guidelines of the EC which request the screening of UV-absorbing compounds, for example, those employed in sunscreen preparations, for their photomutagenic potential. The well established photomutagens 8-methoxypsoralen and chlorpromazine were used to define relevant test protocols. The compounds were evaluated with the Ames test and the Saccharomyces cerevisiae D7 test for gene conversion. The influence of various parameters such as UV light sources, spectral composition, UV sensitivity of the test systems, absorbance by test materials and different exposure conditions is indicated. Two exemplary screening experiments with cosmetic ingredients are presented. Both test systems can be employed for the evaluation of compounds for photomutagenic activity although the standard excision-deficient strains of S. typhimurium pose problems because of their high UV sensitivity. The present experience in this complex field suggests that rigid test protocols and a restrictive test battery would be inadequate.

Photomutagenesis test development: II. 8-Methoxypsoralen, chlorpromazine and sunscreen compounds in chromosomal aberration assays using CHO cells.

Chromosomal changes were analysed in Chinese hamster ovary (CHO) cells treated with 8-methoxypsoralen (8-MOP) or chlorpromazine (CPZ) and irradiated with either a UVA fluorescent tube (emission spectrum ranging from 350 to 400 nm) or a xenon burner (continuous emission spectrum simulating ambient sunlight). In the dark neither 8-MOP nor CPZ was genotoxic by itself. If these compounds were used in combination with UV irradiation the rate of chromosome aberrations was significantly increased. The magnitude of the clastogenic response was dependent on compound concentration and UV dose. The spectral composition also played an important role. Care must be taken to account for spectral changes caused, e.g., by passage of the light through the plastic lid of the container. The possible clastogenicity of two sunscreens was tested with two protocols: (1) cells attached to the culture dish were treated in presence of the sunscreen in the medium or (2) cells were irradiated through a layer of sunscreen solution as a filter. With this a clear UVB-absorbing effect and a decreased frequency of UVAB-induced chromosome aberration was evident with the UVB-absorbing compound Parsol HS but was absent, as expected, with the UVA-absorbing compound Parsol 1789. The presence of the sunscreens in the irradiated cell sample did not cause a significant increase in UV-induced chromosome aberrations.

Phototoxicity and photogenotoxicity of nine pyridone derivatives.

Nine structurally related pyridone derivatives were assayed for photogenotoxicity and phototoxicity in the Ames test, the chromosomal aberration test in V79 cells and the neutral red uptake (NRU) test in 3T3 cells. All nine compounds absorb light to a comparable degree at wavelengths between 380 and 430 nm. Seven of the nine compounds were found to produce high quantities of singlet oxygen (1O(2)) upon irradiation in the presence of oxygen. These seven compounds were highly phototoxic in the NRU test, three were clearly and two were marginally photomutagenic in the Ames test, five were assessed as clearly and two as equivocally photoclastogenic in the chromosomal aberration test. Two compounds showed substantially lower 1O(2) yields. The pyridone ring of these two compounds is attached to a non-aromatic ring, while for the seven other compounds the chromophore system including the pyridone ring consists of two or three aromatic rings. One of the two compounds with low 1O(2) yields was distinctly less phototoxic and did not induce photogenotoxic effects. The other, structurally an indolo derivative and not the common thieno derivative, was, however, similarly phototoxic as the seven compounds with high 1O(2) quantum yield and was also clearly photogenotoxic indicating that different action pathways, not involving singlet oxygen, have to be considered at least for this compound.