Incorporation of a carbon-based nutrient enhancement strategy for drinking water biofiltration is an attractive option, especially for source waters which contain recalcitrant organics. This study compared biofilters that were operated in parallel and individually enhanced with amino acids (including alanine, phenylalanine, and tryptophan), inulin, and sucrose to increase biomass concentration and promote biodegradation of dissolved organic carbon (DOC) in the source water, including disinfection by-product (DBP) precursors. Biomass activity was characterized by measuring adenosine tri-phosphate (ATP), dissolved oxygen (DO) consumption, and through the use of laccase and esterase enzyme assays. Performance was evaluated in terms of headloss, turbidity, pH, DOC, UV254, and DBP formation potential (DBP FP). The introduction of carbon-based nutrients significantly increased biomass activity, where ATP values peaked at 976 ng/g of filter media, 853 ng/g, and 513 ng/g for amino acids, inulin, and sucrose-spiked biofilters, respectively, while a non-spiked control only reached 104 ng/g. DO utilization by the enhanced biofilters was significantly higher than the control, with a strong correlation between ATP and DO uptake observed for all filters (R2 > 0.74). Laccase and esterase enzyme activities of enhanced biofilters were also elevated (p > 0.05), suggesting greater biodegradation potential. Operational parameters such as headloss development and turbidity control were not impaired by carbon supplementation strategies or an increase in biomass concentration and activity. However, the enhancement strategy did not provide improvement in terms of source water carbon removal (DOC and UV254) or DBP FP when treated filters were compared to a control.