A computational DFT B3LYP method with 6-311G (d,p) basis set, the in vitro chemical-based and cellular antioxidant activity (CAA) assays were applied in this study to explain the structure-antioxidant activity relationships of delphinidin and petunidin. The compound molecular structures, spectral properties, frontier orbital energy, and transition state of delphinidin and petunidin were compared. In transition state, the result of the active site (O21-H32 and O22-H33) was consistent with the result of bond length. The frontier orbital theory results indicated that the probable antioxidant activity order was petunidin (0.09126 a.u.) > delphinidin (0.09175 a.u.), which agreed well with the cell-based antioxidant activity determined by CAA. However, the order of ABTS•+ and DPPH radical scavenging activity was delphinidin > petunidin. Our study could help to provide a rational approach for the investigation of antioxidant activity of phytochemicals. PRACTICAL APPLICATIONS: As anthocyanins, delphinidin and petunidin with great antioxidant activity are widely found in various fruits and vegetables. However, there are many kinds of methods used to measure their antioxidant activity and the antioxidant mechanism which are not concrete and clear. Therefore, it is crucial to study the antioxidant actvity of anthocyanins utilizing the DFT method combined with in vitro chemical-based and cell-based assays. Our study could contribute not only to the elucidation of chemical mechanism of antioxidants and exploration the structural features in essence, but also to promote the further development of phytochemicals in the field of food chemistry and pharmacy.