Abstract Two-stage thermophilic hydrogen fermentation and mesophilic methanogenic process for methane production with hydrogenic effluent from co-digestion of decanter cake (DC) and crude glycerol (GLC) was investigated. For the single stage, hydrogen reactor with 2% wv−1 total solid (TS) of DC and crude glycerol was operated at 4 days hydraulic retention time (HRT) with varying glycerol loading of 0.75–1.5% GLC while methane reactor with 2% wv−1 total solid (TS) of DC was operated at 13 days HRT. The maximum hydrogen production rate and hydrogen yield in the single stage obtained from the 1.5% GLC co-digestion condition were 461 mL H2 L−1 d−1 and 23 L H2 kg−1 TSadded, respectively. Crude glycerol was found to be an alternative pH-adjust substance in hydrogen reactor. At the condition of 0.75% GLC co-digestion, the pH was maintained in the range higher than 5.3 throughout the fermentation process. Semi-continuous methane production with 0.75% GLC hydrogenic effluent achieved the maximum methane production rate and methane yield about 736 mL CH4 L−1 d−1 and 44 L CH4 kg−1 TSadded compared with the single stage with 2% TSDC and the second stage with 1% GLC hydrogenic effluent. Total energy recovery from two-stage fermentation about 0.056 kWh kg−1 TSadded was achieved from the condition of 0.75% GLC hydrogenic effluent condition. It was improved by 6.2 and 1.6 times compared to the single stage in both hydrogen and methane reactor, respectively. This work demonstrated the practical enhancement of energy recovery and the efficiency of waste utilization by two-stage fermentation for hydrogen and methane production from decanter cake and crude glycerol.