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Dissolved organic matter (DOM) in the multi-source watershed is of guiding significance for the comprehensive management and the assessment of sustainable safety of the watershed. We investigated the components and spectral characteristics of DOM in North Canal River as a typical multi-source watershed in northern China for the first time, using three-dimensional excitation-emission matrix and parallel factor analysis (PARAFAC). The relationship between DOM composition and water quality was also discussed. Results showed that the DOM in the North Canal River watershed was mainly composed of two similar humic acid-like components (230, 335/400nm and 260, 360/450nm) and a tryptophan-like component (280/290 ~ 350nm). The intensity of DOM shows obvious seasonal spatiotemporal variations. In terms of time, the relative concentration of DOM in winter is significantly higher than that in other seasons due to the influence of water volume, temperature, and photochemical degradation factors. As for the aspect of space, under the combined effect of land use and multiple sources of pollution, the relative concentration of tryptophan-like in the mainstream was significantly higher than tributaries, while the relative concentration of humic-like components in the tributaries was higher than that in the mainstream. In the North Canal River watershed, the primary sources of DOM are human-derived point sources and agricultural non-point sources in the mainstream, and terrestrial and microbial sources in the tributaries. Moreover, the composition of DOM is significantly related to water quality indicators, especially nitrogen and phosphorus, which shows that DOM can have an indicative impact on the trophic status in the North Canal River. The result of this study may provide an indicative effect and scientific basis for water quality monitoring and pollution controlling in the North Canal River watershed.