Dissolved organic nitrogen (DON) is currently considered as one of the most important parameters in drinking water treatment due to its potential to form nitrogenous disinfection byproducts (N-DBPs). A comprehensive investigation was made in this study about the variation of DON concentrations in a full-scale drinking water treatment plant with the treatment train of coagulation/sedimentation, biofiltration and disinfection. The results showed that DON can be effectively removed by coagulation/sedimentation and disinfection, while the biofiltration significantly increases DON concentration. To reveal the mechanism of the increase, the DON concentration and other related parameters at different media depth of the biofilter were studied. The results showed that the DON concentration in biofilter presents a rapid decrease from 0.73 to 0.44 mg/l in the top (0-10cm) of the media, and a slow increase from 0.44 to 1.08 mg/l in the deeper media (10-100cm). SMPs (Soluble microbial products) released by bacterial metabolism might be the main source of the DON in the biofilter. These SMPs contained aromatic protein-like fractions, which were confirmed by EEM analysis.