Isolation and partial characterization of dissolved copper-complexing ligands in streamwaters.

We have separated two groups of copper-complexing ligands (the weak and strong ligands) from streamwaters in the Lake Biwa watershed by modified immobilized metal ion affinity chromatography (IMAC). The weak ligands were about 0.54-1.21% of the total dissolved organic matter (DOM), as determined by UV absorbance, and the strong ligands were about 0.06-0.21%. The results show that the stronger ligands were retained longer on the IMAC column, eluted later, and were accompanied by shorter wavelength UV absorbers, fluorescence maxima patterns with shorter wavelength excitation, and relatively "fresher" organic matter. The weak ligands with logK'(CuL) values of 6.6-7.7 had predominant humic-like fluorescence and may have been considerably degraded, while stronger ligands with logK'(CuL) values of 8.9-9.3 had only protein-like fluorescence and were relatively newly produced, labile material, as indicated from their amino acid composition. The protein-like fluorescence was mainly due to aromatic tryptophan probably bound to proteins or peptides. The results presented here have significant implications regarding the possible sources and biogeochemical role of organic ligands in aquatic environments.