A 12-week repeated-dose oral toxicity study of water-soluble Orostachys japonicus extract (WOJ) was performed in Sprague-Dawley (SD) rats of both genders. Each group of ten rats was orally administered in doses of either 0 or 250 mg/day over a 12-week period. As a result, no WOJ-related changes were observed in terms of survival rate, clinical signs, body weight, or food intake. In addition, no difference in organ weight between the control and treated groups was detected. Furthermore, serum biochemistry parameters revealed some changes within normal ranges although significant decreases in total-bilirubin in the females. In spite of some alterations in serum biochemistry, the clinical signs, body weight changes from food intake, and autoptical remarks indicated that WOJ was not toxic. This study suggests that repeated treatment of O. japonicus very low toxicity and the NOAEL (no observed adverse effect dose) of WOJ exceeds 250 mg/kg in the SD rats.