Abstract: Cadmium (Cd) is the most severe heavy metal pollutant in China’s farmland soils. Soil dissolved organic matter (DOM) significantly influences Cd migration and transformation in soil environments, but whether the relationship between Cd mobility, bioavailability, and DOM spectral characteristics in paddy soils is affected by the rice-crayfish co-culture mode (RS) remains unclear. Therefore, this study used traditional rice cultivation (CK) as the control and RS as the research object. Through UV-Vis spectroscopy and three-dimensional fluorescence spectroscopy, we analyzed changes in DOM spectral characteristics and composition in RS-mode soils during rice growth stages pre-rice stage (T1), tillering stage (T2), booting stage (T3), and yellow maturity stage (T4). Correlation analysis and stepwise regression models were employed to establish relationships between soil DOM and exchangeable Cd (Cd_ex) content in RS and CK modes. The results are as follows: Compared with other growth stages, both modes exhibited the highest aromaticity and hydrophobic DOM at T4. The molecular weight of DOM in RS-mode soils showed a decreasing trend during rice growth, with DOM at T4 being more biogenic in origin. Soil DOM in both RS and CK modes primarily consisted of tryptophan-like components (C1), humic components (C2 and C3), and fulvic-like components (C4). However, the proportional distribution of these components showed no significant variation pattern across growth stages. The fluorescence index (FI) values of both CK and RS modes exceeded 1.9, while humification index (HIX) values were below 4. The biological index (BIX) of RS (0.68–0.80) was higher than that of CK (0.60–0.68), indicating that DOM in both modes possessed endogenous characteristics to some extent. Under both modes, soil exchangeable Cd (Cd_ex) content increased progressively during rice growth stages. However, RS mode exhibited lower Cd_ex content and smaller fluctuation ranges from T2 to T4 compared with CK. Cd_ex content in both modes showed significant or highly significant correlations (P < 0.05 or P < 0.01) with spectral slope ratio (S_R) and fulvic-like components (C4), demonstrating close associations between Cdex content and DOM humification degree, molecular weight, source, and composition. Stepwise regression analysis revealed that factors related to DOM molecular weight (E₂/E₃ and S_R) critically influenced Cd_ex content prediction.These findings provide theoretical references for understanding the potential impacts of rice-crayfish co-culture systems on soil Cd pollution.