Understanding the quantity and chemical structure of dissolved organic matter (DOM) in subsoil and their response to land-use change is essential for evaluating the regional soil organic matter dynamics and soil carbon stability. In the present study, four adjacent land-use systems, including natural forest (control treatment), Chinese fir plantation, Chinese chestnut orchard, and sloping tillage were selected from a subtropical hilly landscape unit. The soil type was red soil derived from granite. Chemical analysis combined with spectrum scanning technology, including ultraviolet spectrum, two-dimensional fluorescence spectra, and near infrared spectroscopy, were used to analyze the quantity and chemical structure of DOM in the topsoil (0-0.2 m) and subsoil (0.2-1 m) in different land use areas. The results showed that DOM was mainly stored in the subsoil, accounting for 58-87% of all DOM in the soil profile. Among the four land-use systems, natural forest contained the most DOM and showed simpler chemical structures of DOM in the subsoil (mainly carbohydrate and proteoid) than in the topsoil. After the natural forest was changed to other land uses, the subsoil suffered more losses in DOM (26%-41%) than the topsoil (12%-49%), which was more obvious in winter than in summer. This indicates that the quantity of DOM in the subsoil was highly sensitive to human disturbance and vegetation changes. In addition, the chemical structure of DOM in the subsoil was generally more complex, and chemical recalcitrant components (aromatics, alkanes, and alkenes) accumulated at 80-100 cm depth following land use change. The spectrum curve, specific absorption peak, and characteristic value of soil DOM were sensitive to land use change and can be used as ecological indicators of vegetation and soil organic matter change after strong human disturbances. This study suggests that the change from natural forest to other land uses not only reduced the quantity of subsoil DOM, but also decreased the quality of soil organic matter, which may weaken carbon stability and the carbon sequestration capacity of subsoil in the long term.