The effect of carbon source and COD/NO3--N ratio on denitrification and methanogenesis in mixed methanogenic matrix was investigated in this study. Industrial wastewater, anaerobic treated cassava stillage (CS) and glucose synthetic wastewater were used as carbon sources respectively for comparison. Experimental results showed that denitrification was the main nitrate reduction pathway for all COD/NO3--N ratios tested in two substrates. Simultaneous denitrification and methanogenesis occurred at COD/NO3--N higher than 7 regardless of carbon sources. Incomplete denitrification was observed at COD/NO3--N ratio below 7 in both the anaerobic effluent of CS and glucose-fed cultures due to the insufficient available organic carbon. The nature of carbon sources was observed to play a key role in the nitrate and organic carbon utilization rates. COD/NO3--N ratio had a strong effect on the organic matter utilization pathways. Methanization consumed more organic matter than denitrification with further increase of COD/NO3--N ratio above 7 in two substrates. Results of VFA variation suggested that propionate and butyrate were preferably utilized by the denitrifiers than acetate. (C) 2012, The Society for Biotechnology, Japan. All rights reserved.