In the safety assessment of cosmetics and their ingredients, complete bans on animal testing are increasing globally. However, the complete replacement of many types of animal testing has yet to be accomplished. In the testing of phototoxicity, one alternative approach is the in vitro 3T3 Neutral Red Uptake Phototoxicity Test (3T3 NRU-PT), adopted by OECD as TG432. Even though this test is now widely used, it cannot accurately evaluate hydrophobic chemicals due to their limited water solubility. The objective of this study was to overcome the limitations of 3T3 NRU-PT and establish a new strategy to evaluate the phototoxic potential of a wide range of chemicals including those that are hydrophobic. Previously, some in vitro testing methods were developed that can be applied to hydrophobic chemicals, but each of them has advantages and disadvantages. In this study, we devised a tiered approach that utilizes four popular methods including 3T3 NRU-PT. Furthermore, we refined the experimental conditions of the method using reconstructed human epidermis and the modified the criteria for determining phototoxicity for hydrophobic chemicals. The results indicate that our tiered approach can be used to accurately and systematically evaluate the phototoxic potential of not only hydrophobic chemicals but also many others.

Transient receptor potential melastatin 8 (TRPM8) is the cold stimulus receptor found as a member of the temperature sensor transient receptor potentia (TRP) family in peripheral sensory nerves. Currently, some studies about TRPM8 functions besides the temperature sensor are in progress, but little is known about the role of TRPM8 in human skin melanocytes. Therefore, we examined the function and effect of Geranium robertianum L. on TRPM8 in human skin melano-cytes. As a result, mRNA expressions of melanin synthesis-related factors were decreased by this TRPM8 activator. On the other hand, a TRPM8 inhibitor exerted reverse effects. In UVB-irradi-ated melanocytes, TRPM8 activity and mRNA expression were decreased, and mRNA expressions of melanin synthesis-related factors were increased. Moreover, Geranium robertianum L. increased TRPM8 activity and mRNA expression, and decreased mRNA expressions of melanin synthesis-related factors in human skin melanocytes. On the basis of these results, it was concluded that TRPM8 had a suppressive function for melanin synthesis and the reduction of this function by UVB irradiation induced the acceleration of melanin synthesis in skin melanocytes. Since Geranium robertianum L. enhanced TRPM8 activity and mRNA level, it may prevent and improve pigmentation induced by the excessive synthesis of melanin.

The anionic surfactant, which is the main component of shampoo, interacts with the cationic polymer to form a water insoluble complex called coacervate in a certain concentration region upon shampoo dilution. It is known that coacervate adheres to hair and imparts a conditioning effect, and it has been reported that the properties of coacervate influence the feeling of use of shampoo. Fragrances are used in shampoo, and the aroma when using shampoo and the substantivity of fragrances on the hair are important. Since coacervate adheres to the hair, it is considered that the aroma when using shampoo and the substantivity of fragrances on the hair are affected by coacervate. In this study, we investigated that the effect of coacervate on the adhesion of fragrance on the hair. As a result of comparing the amount of fragrance in the supernatant part of shampoo solutions with/without coacervate, it is clear that the fragrance tended to be incorporated into the coacervate. The amount of fragrance adhering to the shampooed hair using a shampoo solution with/without coacervate was determined by gas chromatography; it is confirmed that the amount of fragrance on the hair shows more increase with coacervate than without it. It is considered that the fragrance is taken in together with coacervate and adheres to the hair.

There have been a lot of papers concerning the toxicity of inorganic nano-particles such as nano-sized titanium dioxide in recent years. However, no conclusion has been reached about this toxic matter. The situation has been resulted from a lack of long-term tests and epidemiological studies in humans. Natural-origin inorganic nano-particles were found in many volcanic ashes. The nano-particles were easy to suspend in water and higher-order aggregates were not found in the water suspension. We developed a method of separating nano-particles from volcanic ashes. If the contents of nano-particles are only 50 ppm, the absolute mass of the falling nano-particles will be large compared to the mass of artificial-origin nano-particles, because the mass unit of volcanic ashes per eruption may be over ten thousand tons. Sakurajima volcano, located in Kagoshima, Japan, is famous as one of the most active volcanoes in the world. In 1914, Sakurajima experience a large-scale eruption and since then long-term and frequent eruptions have continued. The Japanese government and medical associations have been conducting epidemiological studies for over 100 years and found the risk to human health was mainly acute disease such as eye and respiratory disorders, while chronic disease such as cancer, and reproductive toxicity and genotoxicity were not reported.