The fluorescent properties of dissolved organic matter (DOM) from groundwater in the coastal Dagu River watershed, North China were determined using excitation-emission matrix spectroscopy (EEMs) analysis. Surface water DOM samples were also investigated for comparison. Two humic-like components (C1: 250, 355/472 nm; C2: < 240, 325/400 nm) and one protein-like component (C3: <240, 280/340 nm) were identified using parallel factor analysis. Low intensities for all components were observed in groundwater DOM from the upper and lower reaches of the study area. However, higher abundances of these components occurred in the middle reaches, reflecting the combined effect of seepage of surface water with strong anthropogenic pollution and the alteration of groundwater circulation due to cutoff as a result of the construction of a cutoff wall since the late 1990s. The humic-like components were dominant in groundwater DOM, with the average percentage of the protein-like component being only 15%, which was less than half of the corresponding percentage in surface water DOM. The freshness index in groundwater DOM was lower than the surface water samples, while the fluorescence index and humification index were higher than in the latter. These indices demonstrated the much higher degree of humification for groundwater DOM, which may be related to the longer residence time of groundwater and greater contribution of microbial degradation in the aquifer environment. This study demonstrated that EEMs could distinguish between the effects of natural background and human activities on the quantity and characteristics of the groundwater DOM, and thus could be a useful tool for studing the carbon dynamics and the controlling factors in groundwater systems.