In the course of screening for reactive oxygen species scavengers from natural products, an antioxidant was isolated from the mycelial culture broth of Phellinus linteus and identified as hispidin. The hispidin content was reached its maximum level at 12 days after onset of inoculation. About 2.5 mg/mL of hispidin was produced by P. linteus in a yeast-malt medium (pH 5.8, 25 degrees C). Hispidin inhibited 22.6 and 56.8% of the super oxide anion radical, 79.4 and 95.3% of the hydroxyl radical, and 28.1 and 85.5% of the DPPH radical at 0.1 and 1.0 mM, respectively. The positive control alpha-tocopherol scavenged 25.6 and 60.3%, 74.6 and 96.3%, and 32.7 and 77.5% of each radical, respectively, at the same concentrations. However, hispidin showed no significant activity on the hydrogen peroxide radical.