Pheophorbide a, a major antitumor component purified from Scutellaria barbata, induces apoptosis in human hepatocellular carcinoma cells

Abstract SCUTELLARIA BARBATA has long been used as a Chinese medicine for the treatment of liver diseases such as hepatitis and hepatocellular carcinoma. In the present study, a bioassay-guided method was used to isolate the most active components from SCUTELLARIA BARBATA. The anti-proliferative effects on human hepatoma HepG2 and Hep3B cells of each fraction at every stage of the purification were monitored. An active component, which is 97 % pure by high performance liquid chromatographic analysis, was isolated. Based on nuclear magnetic resonance (NMR) and mass spectrophotometric (MS) analysis, this active component was identified to be pheophorbide a (C 35 H 36 N 4 O 5 ). Mechanistic studies showed that pheophorbide a induced apoptosis in Hep3B cells, a viral-induced hepatoma cell line. However, it was found to be non-toxic in normal human liver cells WRL-68. DNA fragmentation, sub-G 1 cell cycle arrest, as well as suppression of the anti-apoptotic protein Bcl-2, release of cytochrome c to the cytosol, and activation of pro-caspase 3 and pro-caspase 9 were observed when Hep3B cells were treated with 40 μg/mL (i. e., 67.5 μM) pheophorbide a for 48 hours. In conclusion, this is the first report describing the isolation of pheophorbide a from SCUTELLARIA BARBATA using a bioassay-guided isolation method. The anti-proliferative activity and possible mechanisms of action of pheophorbide a on hepatoma Hep3B cells are also discussed.