Aim This study aimed to investigate the effects of vitamin C administration on skeletal muscle hypertrophy induced by mechanical overload in rats. Methods Male Wistar rats were randomly assigned to three groups: (i) sham-operated group (n=8), (ii) placebo-administered group (n=8) and (iii) vitamin C-administered group (n=8). In the placebo-administered and vitamin C-administered groups, the gastrocnemius and soleus muscles of the right hindlimb were surgically removed to overload the plantaris muscle. Vitamin C (500mgkg1) was orally administered to the vitamin C-administered group once a day for 14days. Results Synergist muscle ablation caused significant increases in wet weight and protein concentration of the plantaris muscle in both the placebo-administered (P<0.01) and vitamin C-administered groups (P<0.01) compared with the sham-operated group (SHA). However, the magnitude of plantaris muscle hypertrophy (expressed as a percentage of the contralateral plantaris muscle) was significantly smaller (P<0.01) in the vitamin C-administered group (141%) than in the placebo-administered group (PLA) (152%). Compared with the SHA, only the PLA showed higher expressions of phosphorylated p70s6k and Erk1/2 (positive regulators of muscle protein synthesis) and a lower expression of atrogin-1 (a muscle atrophy marker). Concentrations of vitamin C and oxidative stress markers in the overloaded muscle were similar between the placebo-administered and vitamin C-administered groups. Conclusion Oral vitamin C administration can attenuate overload-induced skeletal muscle hypertrophy, which may have implications for antioxidant supplementation during exercise training.