Soil acts as sink and source of atmospheric carbon (C). Assessment of the long-term impacts of land-use change on soil C stocks is crucial for the sustainability of managed grasslands and for climate change mitigation. This study was conducted to assess the long-term (> 20 yr) impacts of grassland intensification on above- and below-ground C stocks in subtropical ecosystems. The experimental sites consisted of three replicated grassland ecosystems: native-rangeland (least-managed), slash-pine-bahiagrass silvopasture (moderately-managed), and improved bahiagrass pasture (intensively-managed). Soil cores (0-30cm depth) were collected within six adjacent experimental fields (3 ecological units × 2 replicates) to assess C stocks. Root and above-ground biomass were also quantified. Result shows that grassland intensification increased soil C stocks. Improved pasture and silvopasture contained greater soil C (62 and 69 Mg/ha, respectively) than the native rangeland (41 Mg/ha). Root biomass C in native rangeland and improved pasture was comparable (24 and 19 Mg/ha, respectively), but lower in silvopasture (15 Mg/ha). However, above-ground biomass C was greater in silvopasture compared to native rangeland (59 vs. 4.2 Mg/ha), but lower in improved pasture (2.1 Mg/ha). Particulate organic C was greater in improved pasture compared to native rangeland (34 vs. 21 Mg/ha, respectively) but did not differ from silvopasture (27 Mg/ha). Silvopastures exhibited greater amounts of stable mineral-associated C (42 Mg/ha) compared to native rangeland (20 Mg/ha) or improved pasture (28 Mg/ha). Our findings indicate that intensification of grasslands can increase soil C sequestration in the long-term, and can provide additional benefit in above-ground biomass C sequestration.