Influence of the frequently used carbon sources in nitrate removal processes were evaluated in a lab-scale biofilm reactor. The NO3-N removal efficiency was in the order acetate > glucose > methanol > ethanol. Acetate-fed biofilm reduced nearly 100% NO3-N with negligible amount of NO2-N accumulation. Although 99% NO3-N was reduced in the glucose-fed biofilm, substantial NH3-N and NO2-N accumulated. Methanol-fed biofilm reduced 72% of NO3-N with accumulation of 2.2 mg L-1 of NO2-N, while biofilm formed in presence of ethanol showed 61% reduction in NO3-N although relatively higher ratio of denitrifiers were observed. Acetate and ethanol-fed biofilm displayed characteristic biofilm architecture with voids, but the former had relatively higher thickness and diffusion distance. In presence of glucose and methanol, a confluent biofilm without characteristic voids was formed. Pseudomonas sp. numerically dominated the acetate and ethanol-fed biofilm, while Enterobacter sp. and Methylobacillus sp., were abundant in glucose and methanol biofilms respectively. (C) 2012 Elsevier Ltd. All rights reserved.