Land use change is one of the anthropogenic interventions that may induce substantial modifications to both the quantity and quality of soil organic matter (SOM). Soils from three cultivated areas (two types of rainfed upland and one irrigated lowland) and two forests (subtropical managed and temperate unmanaged) in a midhill watershed of Nepal were studied to assess the effect of land use change on SOM quality. Bulk SOM was analyzed using solid-state 13C nuclear magnetic resonance (NMR) and fractionated by acid hydrolysis into pools with different chemical composition and, thus, different tendencies to decomposition. The NMR analysis showed a clear prevalence of O-alkyl C over alkyl C and aromatic C, with major differences between both soils and two depth intervals of the same soil (0–20 and 20–40 cm). Acid hydrolysis revealed that the labile C pool is larger in the 0- to 20- than the 20- to 40-cm soil depth of each land use. Labile C was larger in the temperate unmanaged forest and the irrigated cultivated land than in the other soils at both soil depths. Of the cultivated rainfed soils, the recently reclaimed soil contained less labile C than the historically cultivated soil, while the labile C in the irrigated soil exceeded that in both rainfed soils. The results suggest that changes in land use from forest to cultivation and the irrigation of the cultivated land may significantly affect not only the quantity but also the quality of the SOM and, more importantly, its chemical recalcitrance to degradation. As a consequence, any future decisions regarding land management in the fragile mountain areas of Nepal should be carefully considered with respect to the nature of the SOM.