To investigate the preventive effects of d-psicose, one of rare ketohexoses, on di-(2-ethylhexyl) phthalate (DEHP)-induced testicular injury, prepubertal male Sprague-Dawley rats were exposed to DEHP via their diet or orally, while under treatment with d-psicose. The rats given a diet-containing 1% DEHP alone for 7-14 days showed severe testicular atrophy accompanied by aspermatogenesis. On the other hand, those given the diet plus 2% but not 1% d-psicose-supplemented water for 14 days did not develop testicular atrophy, and exhibited an almost complete spermatogenesis. There was no significant difference in plasma mono-(2-ethylhexyl) phthalate (MEHP) levels between the d-psicose-free and d-psicose-treated groups. The testicular malondialdehyde (MDA) level after a single oral administration of 2g/kg of DEHP showed a similar pattern of increase to the plasma MEHP level and peaked in 24h suggesting a close and dose-dependent relation between plasma MEHP and testicular reactive oxygen species (ROS) levels. Pretreatment with d-psicose at a concentration of 2% and 4% resulted in an almost complete but not absolute suppression of testicular MDA production among rats administered 2g/kg of DEHP. The microarray analysis showed the induction of oxidative stress related genes including the thioredoxin, glutathione peroxidase 1 and 2, glutaredoixn 1 after 24h of the DEHP treatment in the testis. These results show that d-psicose prevents DEHP-induced testicular injury by suppressing the generation of ROS in the rat testis. This effect may be due to the direct scavenging by d-psicose of ROS generated in the testis.