To enhance the co-production of butanol and hydrogen by the acetone-butanol-ethanol (ABE) fermentation of Clostridium beijerinckii, a novel cathodic electro-fermentation (CEF) system was constructed with neutral red (NR) as electron mediator. With the mediation of NR, production of butanol and hydrogen from glucose in the CEF system achieved 5.49 ± 0.28 g/L and 3.74 ± 0.16 L/L, 569.5% and 325.0% higher than that in the open circuit (OC) system, respectively. The butanol and hydrogen yield of 0.30 ± 0.02 g/g and 206.53 ± 8.20 mL/g was 172.7% and 71.4% higher than that in the OC system, respectively. The effective co-production of butanol and hydrogen in the NR-mediated CEF system was attributed to the cooperation of the introduced polarized electrode and the additional NR. With the control of the polarized electrode, a feasible ORP was available for the effective hydrogen production. And the additional NR had induced more carbon source and electrons to the synthesis of butanol.