Reaction of β-Alkannin (Shikonin) with Reactive Oxygen Species: Detection of β-Alkannin Free Radicals Original Research Article

β-Alkannin (shikonin), a compound isolated from the root of Lithospermum erythrorhizon Siebold Zucc., has been used as a purple dye in ancient Japan and is known to exert an anti-inflammatory activity. This study aimed to understand the biological activity in terms of physico-chemical characteristics of β-alkannin. Several physico-chemical properties including proton dissociation constants, half-wave potentials and molecular orbital energy of β-alkannin were elucidated. This compound shows highly efficient antioxidative activities against several types of reactive oxygen species (ROS), such as singlet oxygen (1O2), superoxide anion radical (radical dotO2), hydroxyl radical (radical dotOH) and tert-butyl peroxyl radical (BuOOradical dot) as well as iron-dependent microsomal lipid peroxidation. During the reactions of β-alkannin with 1O2, radical dotO2 and BuOOradical dot, intermediate organic radicals due to β-alkannin were detectable by ESR spectrometry. Compared with the radicals due to naphthazarin, the structural skeleton of β-alkannin, the β-alkannin radical observed as an intermediate in the reactions with 1O2 and radical dotO2 was concluded to be a semiquinone radical. On the other hand, during the reactions of β-alkannin and naphthazarin with BuOOradical dot, ESR spectra different from the semiquinone radical were observed, and proposed to result from the abstraction of hydrogen atoms from phenolic hydroxyl groups of β-alkannin by BuOOradical dot. Based on the ROS-scavenging abilities of β-alkannin, the compound was concluded to react directly with ROS and exhibits antioxidative activity, which in turn exerts anti-inflammatory activity.