•Two-stage and one-stage AD of vinasse were investigated and compared.•Two-stage AD of vinasse from Chinese liquor industry was first conducted.•Lag-phase and T80 of two-stage AD were shorter than that of one-stage.•The energy recovery and VS removal efficiency of two-stage AD was higher.•Two-stage fermentation was more suitable for the AD of vinasse. Vinasse is mainly produced during the brewing or liquor production, which is an potential substrate for biogas production. However, due to the rich content of easily degradation components in vinasse, the fermentation system during the initial AD stage of vinasse is easily acidifying, which leads to the fermentation system unstable, even failed. In this study, mesophilic hydrogen and methane production from vinasse through two-stage anaerobic digestion was investigated. In addition, one-stage anaerobic digestion of vinasse was also conducted to compare with two-stage anaerobic digestion. During two-stage anaerobic digestion, the hydrogen and methane yield were 14.8 and 274ml/g VSsubstrate, respectively. The methane yield from two-stage anaerobic digestion was 10.8% higher than that of one-stage. During the methane production process, the lag-phase of two-stage anaerobic digestion was 9.1days less than that of one-stage. In addition, the VS removal efficiency and energy recovery of two-stage anaerobic digestion were 10.4% and 12.9% higher than those of one-stage. Two-stage anaerobic digestion for hydrogen and methane production could efficiently improve the substrate utilization efficiency and energy recovery of vinasse.