In the present study, the soil column with radius of 30 cm and height of 200 cm was used to simulate a subsurface wastewater infiltration system. Under the hydraulic loading of 4 cm·d-1, composition and transformation of dissolved organic matter (DOM) from different depths were analyzed in a subsurface wastewater infiltration system for treatment of septic tank effluent using three-dimensional excitation emission matrix fluorescence spectroscopy (3D-EEM) with regional integration analysis (FRI). The results indicate that: (1) from different depth, the composition of DOM was also different; influent with the depth of 0.5 m was mainly composed of protein-like substances, and that at other depths was mainly composed of humic- and fulvic-like substances. (2) DOM stability gradually increased and part of the nonbiodegradable organic matter can be removed during organic pollutants degradation process. (3) Not only the organic pollutants concentration was reduced effectively, but also the stability of the DOM improved in subsurface wastewater infiltration system.