Application of the equivalency factor concept to the phototoxicity and -genotoxicity of furocoumarin mixtures.

Furocoumarins (FCs) are natural constituents widely occurring in plants used as food or in phytomedicines, cosmetics, etc. Some FCs exert dermal photo-toxicity and -genotoxicity when combined with UVA irradiation. For a few congeners, skin tumor formation has been described in humans and laboratory animals. Since almost no information is available on the photo-toxic properties of several congeners, we analyzed the photo-cytotoxic, photo-mutagenic, and photo-clastogenic properties in V79 cells for thirteen naturally occurring FCs, and for the coumarin limettin. Furthermore, nine FC mixtures including one mixture based on the FC pattern of an Angelica archangelica extract were tested in the same assays. We found that the concept of relative potency factors for photo-cytotoxic, -mutagenic, and -clastogenicpotencies of FCs, setting the value for 5-methoxypsoralen at 1.00, was applicable to all congeners tested. The concept was used successfully to describe the photo-toxic properties of binary mixtures of 5- and 8-methoxypsoralen. Furthermore, the photo-genotoxic (photo-mutagenic and -clastogenic) properties of complex FC mixtures comprising up to nine different congeners could be predicted. These data suggest that FCs can differ widely in their photo-toxic and photo-genotoxic properties but show relatively strict additivity with respect to their on target-effects when occurring as complex mixtures.

Application of the concept of relative photomutagenic potencies to selected furocoumarins in V79 cells.

Furocoumarins are phototoxic and photomutagenic natural plant constituents found in many medicinal plants and food items. Since plants contain mixtures of several furocoumarins, there is a need for a comparative risk assessment of a large number of furocoumarins. Previously, we have introduced the concept of relative Photomutagenicity Equivalency Factors (PMEFs) derived from the slope of the concentration-response curve of photomutagenicity of individual furocoumarins in V79 cells using the HPRT mutation assay in the presence of UVA irradiation at 125mJ/cm(2). Here we have applied this method to the furocoumarins bergamottin, isopimpinellin and psoralen using 5-methoxypsoralen (5-MOP) as a reference compound with a PMEF of 1.0. We found that neither bergamottin nor isopimpinellin, two furocoumarins abundant in plants, food etc., exerted any significant photomutagenicity while psoralen was clearly photomutagenic with a PMEF of 0.36. Similarly, isopimpinellin was not phototoxic in V79 cells, while bergamottin showed some cytotoxicity which, however, was completely independent of UVA irradiation. Only psoralen was photocytotoxic showing a similar concentration-response relationship for photomutagenicity, and for photocytotoxicity (at 72h after irradiation). Data from the micronucleus assay for DNA damage at 20h after irradiation were in complete agreement with the HPRT mutation data. Our findings indicate that individual furocoumarins differ enormously in their photomutagenic potency, and that a specific toxicological risk assessment is required for each furocoumarin instead of an assessment based on the sum of furocoumarins in a given sample.

Relative photomutagenicity of furocoumarins and limettin in the hypoxanthine phosphoribosyl transferase assay in V79 cells.

Furocoumarins are phototoxic and photomutagenic natural plant constituents found in many medicinal plants and food items. Because plants contain mixtures of several furocoumarins, there is a need for a comparative risk assessment of a large number of furocoumarins. Little is known about the photomutagenicity of the structurally related family of coumarins, which are also abundant in many plant species. In this study, we analyzed the photomutagenic potency of the linear furocoumarins 5-methoxypsoralen (5-MOP) and 8-methoxypsoralen (8-MOP), the angular furocoumarin angelicin, and the coumarin limettin. Above certain concentrations, all test compounds were more or less phototoxic in the presence of UVA doses between 50 and 200 mJ/cm(2), 5-MOP being the most phototoxic compound. At nonphototoxic concentrations, linear correlations were found between concentration and mutagenicity at a UVA dose of 125 mJ/cm(2) for all test compounds including limettin. For 5-MOP, strictly linear correlations were also found for the relationships of mutagenicity vs concentration at various UVA doses or vs UVA dose at given concentrations, respectively. These data indicate that the photomutagenicity of 5-MOP is proportional to the UVA dose x concentration product for noncytotoxic combinations of both factors. They also suggest that the slope of the concentration-photomutagenicity correlation at a given UVA dose may provide a basis for comparison between individual compounds. Applying this concept, in vitro photomutagenicity equivalency factors at 125 mJ/cm(2) were as follows: 1.0 (5-MOP, reference compound), 0.25 (8-MOP), and 0.02 (angelicin and limettin, respectively). These findings provide a new concept for the description of the relative photomutagenic potency of coumarins and furocoumarins and indicate that, in V79 cells, 8-MOP is less photomutagenic and limettin and angelicin are much less photomutagenic than 5-MOP.