Stokes Extract and Its Flavonoids Protect PC-12 Cells against HO-Induced Cytotoxicity.

Stokes (RVS), an herbal medicine found in East Asia, was extracted and further fractionated to investigate its antioxidant capacity and neuroprotective effects. The RVS ethyl acetate (EtOAc) fraction had the highest level of total phenolics and antioxidant capacity among all solvent fractions tested. Pretreatment of PC-12 cells with the EtOAc fraction effectively attenuated HO-induced oxidative damage. Furthermore, the EtOAc fraction significantly attenuated caspase-3 activity, resulting in inhibition of HO-induced apoptosis. We identified and quantified fustin, sulfuretin, and butein in the EtOAc fraction using accurate mass quadrupole time-of-flight mass spectrometry and reversed-phase high-performance liquid chromatography. The intracellular antioxidant capacity and superoxide dismutase (SOD) activity were significantly increased in PC-12 cells treated with the EtOAc fraction and with individual flavonoids. When cells were pretreated with the EtOAc fraction or individual flavonoids and then co-incubated with diethyldithiocarbamic acid (an inhibitor of SOD activity), cell viability against HO-induced oxidative stress was attenuated. These results suggest that the RVS EtOAc fraction and its flavonoid constituents protect PC-12 cells against HO-induced neurotoxicity through their antioxidant properties.