Enzyme activities play key roles in the biochemical functioning of soils, including soil organic matter formation and degradation, nutrient cycling, and decomposition of xenobiotics. Knowledge of enzyme activities can be used to describe changes in soil quality due to land use management and for understanding soil ecosystem functioning. In this study, we report the activities of the glycosidases (β-glucosidase, α-galactosidase, and β-glucosaminidase), acid phosphatase, and arylsulfatase, involved in C (C and N for β-glucosaminidase), P, and S cycling, respectively, as affected by soil order and land use within a watershed in north-central Puerto Rico (Caribbean). Representative surface soil (0–15 cm) samples were taken from 84.6% of the total land area (45,067 ha) of the watershed using a completely randomized design. The activity of α-galactosidase was greater in soils classified as Oxisols than in soils classified as Ultisols and Inceptisols, and it was not affected by land use. The activity of β-glucosidase was greater in Oxisols compared to the Inceptisols and Ultisols, and it showed this response according to land use: pasture > forest > agriculture. The activity of β-glucosaminidase was higher in Oxisols than the other soil orders