Strong interaction between natural dissolved organic matter (DOM) and Hg(II) may influence the transport, conversion, toxicity and bio-validity of mercury in the environment. In this paper ultraviolet (UV) absorbance titration was employed for the first time for the determination of the conditional stability constants of Hg(II) and (DOM). With increasing Hg(II) concentrations, the UV absorbance of fulvic acid, humic acid, and DOM in river increases progressively. By linear and non-linear model fitting, the conditional stability constants (lgK) of Hg(II) and DOM were worked out to be 3.54-4.93 and 3.64-4.85, respectively. The results are consistent with those acquired by the typical fluorescence quenching titration method, with the maximum relative error of lgK being 2.6% and the average relative error being 0.2%. The UV absorbance titration method has the advantages of rapid determination, simple performance, and it will probably become a new approach to studying interactions between DOM and trace metallic ions.