Sludge from a brewery was used to produce methane by feeding glycerol (propane-1,2,3-triol) at an organic loading rate (OLR) of Chemical Oxygen Demand (COD) at 2.5 g L-1 d(-1). Results from two different substrate-feeding approaches were compared: one was the shock loading of glycerol (Run A) and other was a gradual increase in the glycerol amount in a mixture with other carbon sources including glucose, sodium acetate, and lactate (Run B). Methane production rate was similar for both experiments (approximately 21 mmol L-1 d(-1)). Dominant bacteria in Run A were closely related to Mesotoga sp., Alkalibacter sp., and Garciella sp., while the dominant bacteria in Run B were closely related to Trichococcus sp. Dominant archaea were similar for both experiments and were closely related to Methanosaeta sp. and Methanobacterium sp. From these results, it was confirmed that the microbial consortium, especially the bacterial consortium, was strongly dependent on the feeding approach of the glycerol in the anaerobic digestion system. (C) 2016 Elsevier Ltd. All rights reserved.