This study evaluated the effects of Valeriana officinalis root extract on on survival and biochemical factors of rainbow trout, Oncorhynchus mykiss during transportation. For this purpose, fish were distributed in 24 plastic bags, of which 12 contained the V. officinalis root extract (50 mg/l) (group B) and 12 had no herbal extract as control (group A). Transport lasted 16 h and every 4 h, 3 bags of each treatment were sampled. Survival, water quality parameters, plasma  glucose and cortisol were studied. Results indicated that no significant differences in the pH, dissolved oxygen, ammonia levels and temperature were observed between treatments at all the assay periods (P > 0.05). Serum glucose and cortisol levels were significantly (P < 0.05) lower in group B over all the assay periods as compared to the control group. The survival rate (%) was significantly higher in group B compared to the control group (P < 0.05). The present work revealed that the application of V. officinalis root extract will be effective in suppressing stress responses in O. mykiss and it can be recommended for rainbow trout transport.