Biological Safety of LipoFullerene composed of Squalane and Fullerene-C60 upon Mutagenesis, Photocytotoxicity, and Permeability into the Human Skin Tissue

Fullerene-C60 (C60) is mainly applied in the aqueous phase by wrapping with water-soluble polymer or by water-solublizing chemical-modification, whereas C60 dissolved in oil is scarcely applied; still less explicable is its toxicity. We dissolved C60 in squalane at near-saturated or higher concentrations (220-500 ppm), named LipoFullerene (LF-SQ), and examined its biological safety. LF-SQ was administered at doses of 0.49-1000 mu g/ml to fibroblast cells Balb/3T3, and showed that cell viability was almost equal to that of the control regardless of the UVA- or sham-irradiation, indicating no phototoxicity. Reverse mutation by LF-SQ was examined on four histidine-demanding strains of Salmonella typhimurium and a tryptophan-demanding strain of Escherichia coli. As for the dosages of LF-SQ (313-5000 mu g/plate), the dose-dependency of the number of reverse mutation colonies of each strain did not show a marked difference when compared with the negative control, regardless of the metabolic activation, in contrast to twice or more differences for five positive controls (sodium azide, N-ethyl-N'-nitro-N-nitrosoguanidine, 2-nitrofluorene, 9-aminoacridine, and 2-aminoanthracene). In human skin biopsy built in a diffusion chamber, C60 permeated into the epidermis at 33.6 nmol/g tissue (24.2 ppm), on administration with LF-SQ containing 223 ppm of C60, but not detected in the dermis even after 24 hrs, as analysed by HPLC. It is presumed that LF-SQ can permeate into the epidermis via the corneum but can not penetrate the basement membrane, and so can not reach into the dermis, suggesting no necessity for considering a toxicity of C60 due to systemic circulation via dermal veins. Thus, C60 dissolved in squalane may not give any significant biological toxic effects such as photocytotoxicity, bacterial reverse mutagenicity, and permeability into the human skin.