Fluorescence characterization of soil dissolved organic matter (DOM), which is one of the most important indexes concerning study on soil organic matter, can be effected by freezing and thawing cycles. In this paper, the fluorescence characterization of black soil DOM under the effect of freezing and thawing cycles was studied, using three-dimensional excitation-emission-matrix fluorescence spectroscopic (EEM). Based on the transformation of fluorescence characterization, the influences of humification degree and active humus in black soil were analysed. The result showed, compared with untreated by freezing and thawing cycles, (1) The phenomena red-shift of UV fulvic-like was found in soil DOM. It meant that the aromatization and humification degree increased. (2) Protein-like fluorescence peak was observed in some soil samples, which meant microbial activity was enhanced. (3) Active humus and humification degree are the indexs of soil fertility. The content of TOC in the active humus rose, just the same as the value of humification degree. It meant that soil fertility was improved. (4) Compared with the ratio of UV: visible humic-like fluorescence (r(a,c)), there were positive correlations with the active humus and humification degree. It meant that r(a, c) of soil DOM was one of the indexs on active humus, humification degree and soil fertility effected by freezing and thawing cycles.