Several experimental tests were performed to study acidification during the anaerobic treatment of brewery wastewater. Batch test results suggested that a pH of 5-6 and a water temperature of 40°C were optimal conditions for acidification. Continuous-flow test results indicated a tendency for the carbohydrate reduction percentage to become higher with longer retention time in the acidification reactor. A high Specific Methanogenic Activity (SMA) of over 1.4 kg COD kg−1 MLVSS d−1 was observed, which the carbohydrate reduction percentage exceeded 75%. However, there was a tendency for the SMA to become lower due to residues of highly concentrated carbohydrates. The carbohydrate reduction percentage dropped to below 60% in the acidification treated water, when the retention time in the acidification reactor was extremely short. For achieving a carbohydrate reduction percentage of more than 75%, the retention time showed a tendency to become longer with higher organic concentration (e.g. COD 2,500-4,000 mg l−1: 3 h, COD 5,000-8,000 mg l−1: 6 h, and COD 10,000-16,000 mg l−1: 12 h).