Attempts have been made from comparative biochemical standpoint to isolate and determine free amino acids in the brain of the frog (Rana nigromaculata) in hibernation, the animal belonging to the amphybian family by the ion exchange cbromatography, and the following are the results: 1. Aspartic acid, glutamic acid, alanine, and serine have been found markedly decreased as in the case of the catfish brain as compared with those in the mammalian brains. 2. Taurine has been found more markedly decreased than either in the ease of the catfish brain or of the rat brain. 3. The amounts of unidentified substances X(1), X(2) and acidic amino-group show the values intermediate between those in the catfish brain and those in the rat brain. 4. The amount of γ-aminobutyric acid does not show any marked difference in the brains of all the animals tested. 5. Glycine, isoleucine, leucine, threonine, and lysine yield the values closer to those found in the mammalian brains than those in the catfish brain.