To investigate the safety of L-alanine, male and female Sprague-Dawley strain SPF rats [Crj:CD(SD)IGS] were administered L-alanine at 2,000 mg/kg/day by gavage for 4 weeks. After the end of the dosing period, reversibility was assessed following a 2-week recovery period. In the results, there were no toxicologically significant changes caused by L-alanine in general condition, body weight, food consumption, ophthalmology, hematology, blood chemistry, organ weight or at necropsy. In urinalysis, increased number of animals showing urine protein-positive or phosphate salt was observed in males and females. In addition, urine volume was significantly increased in males. In histopathological examination, squamous cell hyperplasia in the limiting ridge in the stomach was observed in males and females. These changes were reduced or no longer observed after the 2-week recovery period and thus were reversible changes. These results suggest that repeated oral administration of L-alanine at 2,000 mg/kg/day for 4 weeks is well tolerated in male and female rats.