The adoption of conservationist systems can improve the protection of soil organic matter (SOM) and mitigate greenhouse gas emissions. Thus, we aimed to quantify the soil carbon (C) and nitrogen (N) stocks, C content in the SOM fractions and microbial C (Cmic) in the topsoil of an Acrisol under traditional and agrisilvicultural systems in the Southeast of Brazil. The evaluated treatments were native vegetation of Cerrado (NV), pasture of Urochloa brizantha (PAST), monoculture of pigeon pea (MCP), and agrisilvicultural system (Eucalyptus + pigeon pea; AGS). Soil samples were collected at depths of 0–5, 5–10, and 10–20 cm to determine the soil C and N contents and stocks, C associated with granulometric and humic fractions of SOM, and soil Cmic. The C contents in AGS and MCP systems were similar to NV at 5–10 and 10–20 cm, and soil C stocks in AGS were similar to NV and higher than in PAST system at 0–20 cm. We found a higher percentage of protected C in the silt + clay fraction, and the AGS system contributed to an increase in this organomineral fraction at 10–20 cm soil depth. The C contents in the free light fraction (0–5 cm) and humin (5–10 and 10–20 cm) were also similar between AGS and NV, giving greater protection of SOM in both systems. For Cmic and microbial quotient values we found no differences among the evaluated systems. The results indicated that after conversion of NV to different land uses, the AGS system more effectively increased the C stocks in the topsoil and protected the SOM.