Establishment of pine (Pinus spp.) plantations on grasslands could increase carbon (C) sequestration to counteract increased atmospheric carbon dioxide concentrations. In the grasslands of the southern Brazilian highland (Campos), large areas have been converted to Pinus plantations over the last 30 years. In order to assess the impact of this land-use change on the amount and composition of soil organic matter (SOM), we investigated a grassland pasture site (G), and both an 8-year-old (P8) and a 30-year-old (P30) plantation with Pinus taeda. Soil samples down to 45 cm were analysed for texture, pH, soil organic carbon (SOC) and total nitrogen (Ntot) concentrations. Chemical composition of SOM was determined by using cross-polarization magic angle spinning (CPMAS) 13C NMR spectroscopy. We analysed for stable C isotope (δ13C) and assessed the lignin composition by CuO oxidation. Additionally, contents of pyrogenic organic material (PyOM) were determined because the Campos is regularly burnt. Both pine plantations revealed relatively small SOC concentrations in the mineral soil of 72.6 mg g−1 (P8) and 56.8 mg g−1 (P30) and Ntot concentrations of 4.0 mg g−1 (P8) and 2.9 mg g−1 (P30) for the A horizon, while grassland showed significantly (P < 0.01) larger contents of 100.2 mg g−1 for SOC and 5.9 mg g−1 for Ntot. Accumulation of litter layers suggests decreased input of organic material into the mineral soil under pine, which was confirmed by the δ13C values and lignin composition. Smaller contents of vanillyl- (V), syringyl- (S), and cinnamyl (C)-phenols, smaller ratios of S/V and C/V, and smaller ratios of acidic to aldehydic forms of V and S phenols indicated a high degree of decomposition of residual grass-derived SOM in the upper part of the mineral soil (0–10 cm) under pine plantations. This was confirmed by CPMAS 13C NMR spectroscopy, showing an increasing Alkyl C/O-Alkyl C ratio at the same depth. No significant changes in the contents of PyOM could be detected, but all sites tended to show the greatest concentrations at deeper soil depths > 15 cm, indicating a vertical relocation of PyOM. The results suggest that decomposition of residual SOM originating from grassland species contributes to the decrease of SOC and Ntot and to an acidification in the topsoil under pine plantations. We also suggest that slow litter decomposition and incorporation and the absence of fires at the plantations are additional reasons for the reduced amount of SOM. Depletion of SOM and the acidification of the topsoil may reduce the availability and supply of nutrients and diminish the C sequestration potential of the mineral soil.