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Topsoil layer contains various components of soil organic matter such as branches, leaves, bark, or metabolites and residues of soil biota. Soil organic matter (SOM) in forest ecosystems consists mostly of lignin, cellulose, chitin and other hydrocarbons. These compounds are decomposed mainly by soil fungi which produce extracellular enzymes to decompose wide range of organic residues. These enzymes may alter nutrient cycling and change soil properties such as water retention capacity, ion exchange capacity, formation of soil microstructure, soil microbial respiration, etc. In this study, we studied enzyme activity in the topsoil layer of postmining spoil heaps near Sokolov, Czech Republic. We investigated the effect of the following factors and their combinations on enzyme activity: i) dominant vegetation, ii) time of sampling, iii) reclamation, and iv) soil macrofauna.We measured enzyme activity in plastic mesocosms with autochthonous litter deposited on reclaimed and unreclaimed post-mining sites. We used mesocosms accessible for macrofauna (mesh size > 2 mm) and mesocosms inaccessible for macrofauna (mesh size < 2 mm). Under laboratory conditions, we measured enzyme activity (laccase, oxidase, peroxidase, Mn-peroxidase, alkaline phosphatase, acid phosphatase, endoglucanase, xylanase, cellobiohydrolase, glucosidase, xylosidase and chitinase) using buffer extraction method followed by spectrophotometric assay. We did not find any statistically significant difference between mesocosms accessible and inaccessible for fauna. However, we found significant effect of time of sampling on enzyme activity. Our results showed significant difference between reclaimed and unreclaimed post-mining sites. Reclaimed sites showed significantly higher enzyme activity than unreclaimed sites.