A 13-week subchronic oral toxicity study of stevioside was carried out in F344 rats at dose levels of 0, 0.31, 0.62, 1.25, 2.5 and 5% in diet, to determine appropriate dose levels for a 2-year carcinogenicity study. The rats were randomly allocated to 6 groups, each consisting of 10 males and 10 females. No animals died during the administration period. Between the control and treated groups, there were no differences in body weight gain during the administration period and in food consumption in the later period of the study. LDH on biochemical investigation and single cell necrosis in the liver revealed by histopathological examination were increased in all male treated groups. These were not considered specific changes, because of the lack of any clear dose response, the relatively low severity and the limitation to males. Other parameters that were found to demonstrate significant differences on hematological and biochemical investigations were of minor toxicological significance. From these results, a concentration of 5% in diet was concluded to be a suitable maximum tolerable dose of stevioside for a 2-year carcinogenicity study in rats.