In recent years two novel types of bufotoxins in which the succinoyl and adipoyl groups are substituted for the suberoyl residue of the hitherto known bufotoxin have been separated from the skin of Japanese toad. We now wish to report the isolation and characterization of two new bufotoxins, one of which possesses the pimelate as a dicarboxylic acid moiety, from the skin of Bufo vulgaris formosus Boulenger. The ethanolic extract of the skin from 1,800 toads was repeatedly chromatographed to give a new bufotoxin (I), which showed a positive result with Sakaguchl's reagent and a negative test with ninhydrin, and exhibited nmr signals at δ 0.75 (3H, s, 18-CH3), 1.08 (3H, s, 19-CH3), 3.70 (1H, m, 11β-H), 5.05 (1H, m, 3α-H), 6.25 (1H, d, J=10Hz, 23-H), 7.45 (1H, d, J=2Hz, 21-H), and 7.95 ppm (1H, q, J=10,2Hz, 22-H). Hydrolytic cleavage with 6N hydrochloric acid furnished arginine. On enzymatic hydrolysis with a hog pancreas lipase preparation followed by methylation with diazomethane, compound I afforded the methyl ester of gamabufotalin 3-hemipimelate (II), and subsequent acetylation showed identity with an authentic specimen prepared from gamabufotalin, supporting the structure assignment of gamabufotalin 3-pimeloylarginine ester to this new type bufotoxin (the first recorded instance of the naturally occurring pimeloylarginine ester of bufogenin). Another new bufotoxin (IV) was similarly separated, and enzymatic hydrolysis and subsequent methylation with diazomethane provided the methyl ester of bufalin 3-hemisuccinate (V), leading to the assignment of bufalin 3-succinoylarginine ester. Further studies on the isolation of the new bufotoxin from the Japanese toad are being conducted.