We have recently reported that hypelcin A is an α-aminoisobutyric acid (Aib)-containing peptide antibiotic (peptaibol) with uncoupling activity on rat liver mitochondria and growth-inhibitory activity against Lentinus edodes. Previously thought to be a single crystalline compound (uniform in thin layer chromatography), hypelcin A was found to be a mixture of more than four components by high performance liquid chromatography (HPLC). Here we report the isolation and structural elucidation of four new antibiotics, hypelcins A-I, A-II, A-III, and A-IV, from crude hypelcin A. Crude hypelcin A was purified by reversed-phase HPLC on Chemopak Nucreosil using methanol-water-2-propanol (45:27:20) to yield major components A-I (mp 265-266°C, [α]ᴰ²¹ -17.0°) and A-II (mp 254-256°C, [α]ᴰ²¹ -16.0°), and minor components A-III (mp 256-258°C) and A-IV (mp 259-261°C). Molecular weights of 1927, 1913, 1941, and 1927 were assigned to A-I to A-IV based on positive ion fast atom bombardment (FAB) mass spectra. Their structures (1-4) were established using FAB mass spectrometry (acyl cleavage ion series), amino acid analysis, ¹H-NMR (acetyl group), ¹³C-NMR (amide carbonyl groups), and gas chromatography mass spectrometry (GC-MS) confirmation of leucinol (Lol) from hydrolysates.