FIRST PHASE INTER-LABORATORY VALIDATION OF THE IN VITRO EYE IRRITATION TESTS FOR COSMETIC INGREDIENTS: (4) EVALUATION OF HEMOGLOBIN DENATURATION (HD) TEST
HIROSHI ITAGAKI1,2, TOSHIKATSU HAYASHISUP>1,2, HIROSHI KAKISHIMASUP>1,3, TOMOYASU OGAWA1,3, MAYUMI KOTANI1,4, KIYOJI MATSUKAWA1,5, KUNIO MASUDA1,5, HAJIME KOJIMA1,6, KATSUYOSHI CHIBA1,7, IKUYO MAKINO1,7, KAZUTAMI SAKAMOTO1,8, YOSHINOBU TAKINO1,8, MINA KANARI1,8, TOYOZO KANEKO9, AKIHIKO HIROSE9, YASUO OHNO10 and AKIRA TAKANAKA10
1Japan Cosmetic Industry Assoc. (JCIA), 4th floor Hatsumei Bldg., 9-14, Tranomon, 2-chome, Minato-ku, Tokyo 105, Japan;
2Shiseido Safety & Analytical Research Center, 1050 Nippa-cho, Kohoku-ku, Yokohama 223, Japan;
3Kanebo Cosmetic Laboratory, 3-28, 5-chome, Kotobuki-cho, Odawara-shi, Kanagawa 256, Japan;
4SUNSTAR Inc., 3-1 Asahi-machi, Takatsuki-shi, Osaka 569, Japan;
5OPPEN Cosmetic Co. Ltd., 2-28-2 Shinaike, Setsu-shi, Osaka 569, Japan;
6Biochemical Research Institute, Nippon Menard Cosmetic Co Ltd., 4-66, Asakusa, Ohgaki-shi, Gifu 503, Japan;
7Safety Research Center, Yakult Central Institute for Microbiological Research, 1796 Yuo, Kunitachi-shi, Tokyo 186, Japan;
8Applied Research Laboratories, Central Research Laboratories, Ajinomoto Co. Inc., 1-1 Suzuki-cho, Kawasuki-ku, Kawasaki 210, Japan;
9Div. of Toxicology, Mitamura Building, 26-1 Kaitaicho, Shinjuku-ku, Tokyo 112-0012 Japan 1-18-1, Kamiyoga, Setagaya-ku, Tokyo 158, Japan;
10Div. of Phamacology, Mitamura Building, 26-1 Kaitaicho, Shinjuku-ku, Tokyo 112-0012 Japan 1-18-1, Kamiyoga, Setagaya-ku, Tokyo 158, Japan
Regular Article :AATEX 3(4):154-161
The hemoglobin denaturation (HD) test to detect eye irritation potential of cosmetic ingredients was primarily validated by eight laboratories using nine surfactants and physiological saline as test substances. The test procedures were controlled under the common standard operating procedure (SOP) in which the denaturation was measured spectrophotometrically using microplate reader and the hemoglobin denaturation ratio (HDR) for each concentration level of test chemicals was calculated. The rank order of test chemicals among laboratories was similar with respect to the concentration for 10% HDR However, HDR values themselves varied widely among laboratories. This seemed to be caused by the differences in the model of microplate reader and its filter. Multiple linear regression analysis proved that HDRs calculated in individual laboratories were highly correlated to in vivo maximal average Draize total scores (MAS). The mean correlation coefficient was 0.846. From these results, we concluded that the protein denaturation test using hemoglobin is a promising alternative method to the Draize rabbit eye irritation test (Draize test). Further validation of this method using a wider range of cosmetic ingredients is under way.
Key words: validation study, Draize eye irritation test, alternatives, in vitro, hemoglobin, hemoglobin denaturation, surfactant.