To ascertain the effects of hair dyeing application on the DNA damage in human lymphocytes, a mixture of permanent black colored hair dye with the same amount of oxidant containing 6% hydrogen peroxide was used. A hair dyeing with contacting the scalp (conventional dyeing) and a hair dyeing with 3 to 4mm away from the scalp (alternative dyeing) were applied to each If young healthy women. Blood was taken from the brachial vein at two sampling times, just before and 6 hours after the hair dyeing, and tail extent moment(TEM) and tail length (TL) were measured by using a comet assay. After dyeing, TL was significantly increased in both conventional dyeing group and alternative dyeing group compared with before dyeing as an average of 47% and 28%, respectively, and TL for conventional dyeing group was higher than alternative dyeing group as an average of 1.2 fold. After dyeing, TEM was significantly increased in both conventional dyeing group and alternative dyeing group compared with before dyeing as an average of 192% and 76%, respectively, and TEM for conventional dyeing group was significantly higher than alternative dyeing group as an average of 1.7 fold. Therefore, alternative dyeing application was induced to lower lymphocyte DNA damage than conventional dyeing application, and TEM was appeared to be a more sensitive tool for the measurement of lymphocyte DNA damage than TL in this study.