This study aimed to assess the effect of petrol and the Fyre Zyme reagent on selected chemical and biochemical properties of loamy sand. The experiment was conducted under laboratory conditions. First, petrol was introduced into the soil at doses of 0 and 50 g k−1dry matter (DM). Next, 6% Fyre-Zyme enzyme reagent solution was added to the samples contaminated and uncontaminated with petrol, in the following combinations: 0 (control), once at 40 cm3 kg−1 DM, twice at 20 cm3 kg−1 DM at two-week intervals, and four times at 10 cm3 kg−1 DM at weekly intervals. Contamination of loamy sand with petrol caused slight changes in the determined chemical parameters and stimulated dehydrogenase activities, but inhibited the activity of phosphatases. The introduction of the enzyme reagent into the soil increased the Corg and Ntot content. The greatest changes were observed in the activity of phosphatases. The receiver operating characteristic (ROC) curves revealed that the application of the enzyme reagent at the application of 4 × 10 cm3 kg−1 DM was the most beneficial. However, the results of the η2 analysis indicate that the greatest influence on the determined experimental parameters was found in the soil contaminated with petrol.