Biohydrogen production was evaluated in an integrated biosystem, using both dark and photo fermentation, using crude glycerol (CG) derived from the production of biodiesel from waste cooking oils (WCO). The dark fermentation assay was done with 20.0 g COD L-1 of CG by a mixed culture from Clostridiales order at 37 degrees C, pH 5.5 and generated 24.06 mmol H-2 g(-1) CODconsumed, with the removal of 45.74% of CG. The dark fermentation effluent (DFE) in concentrations of 1.0, 2.0 and 3.0 g COD L-1 was used in the photo fermentation by a phototrophic bacteria culture of Rhizobiales and Clostridiales order, at 37 degrees C, pH 7.0 at 18.50 W m(-2). The optimum condition was achieved with 1.0 g COD L-1 of DFE and generated 3.94 mmol H-2 g(-1) CODconsumed, with the removal of 76.10% of COD. The culture was able to consume ethanol (76.86%), acetic acid (95.73%), butyric acid (94.76%) and mainly methanol (99.18%), representing a sustainable alternative for the utilization of CG from WCO. (C) 2020 Elsevier Ltd. All rights reserved.