Sulfur deficiency has been recognized as a limiting factor for crop production in many regions of the world. A 120-day incubation experiment was conducted to assess the effect of the applied waste elemental sulfur on sulfur bioavailability in soil. Four doses of sulfur were applied: 10, 20, 30 and 60 mg S kg−1 dry matter (d.m.) of soil. In order to assess the effect of soil pH adjustment on sulfur oxidation, the research was conducted on two sets of soil samples: one set of soil samples had natural pH, and the second one was limed before sulfur application. Application of waste sulfur slightly affected the soil pH, and increased the content of available sulfur in soil proportionally to sulfur dose. A beneficial effect of waste sulfur application on soil dehydrogenase and catalase activity was found. Liming reduced soil acidity, and significantly increased sulfate content and soil enzymatic activity. Waste elemental sulfur may be an alternative source of sulfur, supplementing the deficiencies of this element in soils. The described way of sulfur waste reuse corresponds with the increasingly common approach to create waste-free technologies in all economy.