High molecular weight (HMW) fractions are important components in root exudates. However, there is little available information concerning complexation of Cu(II) to the HMW fractions in root exudates. In the present study, complexation of root exudates from Salicornia europaea L. with Cu(II) was investigated using excitation emission matrix (EEM) fluorescence spectroscopy. Two protein-like fluorescence peaks were identified in the EEM spectra of root exudates. Fluorescence of both peaks was clearly quenched by Cu(II). The increase of conditional stability constant with increasing temperature indicates that the fluorescence quenching of the protein-like fluorescence by Cu(II)may be controlled by a dynamic process. The values of conditional stability constants (logKa) were in the range of 4.32–4.69, which were close to those of complexation of fulvic acid with Cu(II). This shows that the HMW fluorescent substances in root exudates from S. europaea L. were strong organic ligands for Cu(II). Our study suggests that the HMW fluorescent substances may affect chemical forms, mobility, and thus the fate of copper in wetland.