The cytotoxicity and antioxidant activity on human hepatoma cell line HepG2 of three flavonoids homogenous compounds from tartary buckwheat seeds and bran, namely quercetin, isoquercetin, and rutin, were investigated. The total antioxidant competency detection results indicated that the antioxidant capacity of quercetin was the strongest in a biological response system. A [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay showed that quercetin exhibited the strongest cytotoxic effects against the HepG2 cell line. Flow cytometric analysis indicated that quercetin significantly increased the production of reactive oxygen species, and led to the G2/M phase arrest accompanied by an increase of apoptotic cell death after 48 h of incubation. Quercetin-induced cell apoptosis was shown to involve p53 and p21 up-regulation, Cyclin D1, Cdk2, and Cdk7 down-regulation. These results suggested that the induction of G2/M arrest, apoptosis, and cell death by quercetin may associate with increased expression of p53 and p21, decrease of Cyclin D1, Cdk2, and Cdk7 levels, and generation of reactive oxygen species in cells. This study will help to better understand and fully utilize medicinal resources of plant flavonoids.