Scallop shell extract inhibits squalene monohydroperoxide-induced skin erythema and wrinkle formation in rat

Approximately 300 000 t/annum of scallop shells are generated as industrial waste in the Hokkaido area of Japan alone. About 98% of scallop shell is composed of calcium carbonate, and the organic components constitute only 2% of weight of the scallop shell. Because calcium carbonate is a very cheap material, it is necessary to find a characteristic quality of scallop shells in order to effectively utilize this natural material. Calcium oxide after calcination of scallop shells have previously been used as an antibacterial agent, a food supplement, and an adsorbent of formaldehyde. The authors have previously shown that the organic components from scallop shell (scallop shell extract) enhance the turnover rate of the skin epidermal layer and increase the rate of recovery of UV-induced injury to rat dorsal skin. This property of scallop shell extract suggested that it may be suitable as a cosmetic material. The authors also reasoned that scallop shell extract may possess both growth-promoting activity for keratinocyte cells and antioxidative activity. Squalene monohydroperoxide is a primary oxidized lipid produced by UV radiation of skin. The generation of squalene monohydroperoxide has been reported in the pathogenesis of several skin conditions including acne and sunburn. The formation of squalene monohydroperoxide leads to peroxidation of cellular membranes, resulting in cell toxicity in vitro. Chiba et al. reported that topical application of squalene monohydroperoxide to hairless mouse skin induces wrinkle formation, roughness and water loss in skin. In addition, epidermal hyperplasia and dermal alterations such as collagen degradation and increased levels of glycosaminoglycan are induced in squalene monohydroperoxidetreated skin. These findings suggest that hairless mouse skin constitutes an effective model for studying wrinkle formation and skin aging in relation to oxidative stress in vivo. In this present study, the effect of scallop shell extract on squalene monohydroperoxide-induced changes in rat dorsal skin was investigated. Scallop shell extract was prepared as described previously. Male Wistar rats were purchased from Hokudo (Sapporo, Japan) and fed anormal diet and water. Rats were cared for according to the Guidelines of Animal Care issued by the Office of the PrimeMinister of Japan. Squalene monohydroperoxide was prepared as described by Chiba et al. Briefly, 5 mL of squalene solution (1 mg/mL) was UV-irradiated at 365 nm (0.24 J/cm) for 1 h, and the solution was extracted twice with 5 mL of methanol. The methanol extract, containing squalene monohydroperoxide, was separated by reverse phase C18 column chromatography using 100%methanol, and the peak containing squalene monohydroperoxide was pooled. The solvent was evaporated under a stream of nitrogen gas and the squalene monohydroperoxide dissolved in ethanol to a final concentration of 12.3 mg/mL (dry weight/vol.). The squalene monohydroperoxide solution (0.2 mL) or vehicle (0.2 mL of 100% ethanol) was *Corresponding author: Tel: 81-143-46-5745. Fax: 81-143-46-5801. Email: hasegawa@mmm.muroran-it.ac. jp Received 11 June 2007. Accepted 31 July 2007. FISHERIES SCIENCE 2008; 74: 217–219