Microorganism and enzyme activities in the soil are important biological indicators of soil quality. Farmland soil polluted by heavy metal in Ningyuanbu Town of the Jinchuan District in Gansu Province was the object of this study. To analyze the content of heavy metals (Zn, Pb, Cd, Cr, Cu, Ni, As, Hg), microorganisms (bacteria, fungi, actinomycetes), and enzyme activity (urease, alkaline phosphatase, catalase, dehydrogenase), 26 samples in soil layers of 0-20 cm and 20-40 cm were collected. The results showed that the over-standard rates of Ni, Cu, and As in the soil layer of 0-20 cm were 15.4%, 30.8%, and 38.5%, respectively. The over-standard rates of Ni, Cu, and As in the soil layer of 20-40 cm were 7.7%. There was a negative correlation between Pb, Hg, Ni, Cu, Cd, and As, along with bacteria, urease, alkaline phosphatase, catalase, and dehydrogenase. Cr showed a positive correlation with fungi and actinomycetes. Zn showed a positive correlation with bacteria. Pb, Zn, and Cr were the major factors that dominated soil biological traits, and the contribution rate was 72.4%, 16.2%, and 4.9%, respectively. Catalase activity was sensitive to nickel, copper, cadmium, and arsenic pollution, which can be used as an effective index for the evaluation of soil quality in these areas.