Cyclo-alkannin and cycloshikonin were formerly considered to be derivatives of hydroanthraquinone 3. However, chemical and spectral investigation revealed that cycloalkannin possesses no secondary hydroxyl group. Thus, the structure of cyclo-alkannin leucoacetate (8) was determined by X-ray analysis, by the direct method ; the final R index with hydrogen atoms except for those of methyl groups was 0.065. The cyclization of alkannin and shikonin is a reaction between the hydroxyl group and double bond and does not involve the formation of a carbocyclic ring. In 1H-NMR both alkannin and cyclo-alkannin show two singlet signals arising from the protons of the aromatic and quinonic rings. The absence of coupling and lower chemical shift values suggests delocalization of the quinonic ring so that these compounds can be regarded as consisting of tautomeric structures (10, 11). The results of antitumor tests on alkannin and cycloalkannin derivatives (6, 7, 8, 10, 11) are also reported.