In this work, DOM was extracted from urban stormwater runoff samples collected at three typical regions (business, residential and campus regions) in Beijing, China. A comparison between the chemical characteristics of DOM extracted from these three regions was performed using UV-visible spectrometry (UV-vis), excitation–emission matrix (EEM) fluorescence, proton nuclear magnetic resonance spectroscopy (¹H NMR) and ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS). The UV-vis and EEM spectra revealed that the DOM in stormwater runoff samples mainly contained UV humic-like (peak A) compositions with higher molecular weights. ¹H NMR analysis indicated that the DOM of the three regions contained a similar distribution of functional groups, which mainly consisted of aliphatic chains and aromatic components with carbonyl, hydroxyl or alkyl terminal groups. And the UHPLC-Q-TOF-MS demonstrated the identification and characterization of possible compounds existing in the DOM. Further studies should focus on the interactive relationship between DOM and the co-existing contaminants (heavy metals and some typical organic pollutants) in stormwater runoff.