The aim of the study was to compare the activity and functional biodiversity in soil under two different cereals: common maize and winter wheat, both grown in the same pattern of cultivation techniques: conventional (to 25 cm depth) and reduced (to 10 cm depth). Soil samples for comparative analysis were collected at the same time (July 2016) at a long-term field experiment, which was carried out in 2013–2016. Soil biological activity was determined by measurement of dehydrogenases activity (DHa) with TTC (2,3,5-triphenyltetrazolium chloride) application, microbial biomass carbon (MBC) and nitrogen (MBN) content by fumigation-extraction method, and functional diversity of soil microorganisms using the Biolog EcoPlate System. The results demonstrated that the cultivation technique had a greater impact on the soil biological activity, compared to the type of cereal. Higher biological activity was found in the soil under reduced tillage in both cereals. Calculated correlations showed that DHa, MBC, MBN and acid phosphatases were positively correlated with each other. The negative correlation obtained between yield and biological parameters of activity in soil was not expected.