Pure culture of Rhodococcus erythropolis NTU-1 was utilized to test the biodegradability and bioflocculate formation in treatment of petroleum pollutant in the soil. This study evaluated the cultivation in batch system, and discussed the irreversible adsorption of n-hexadecane and special equilibrium between solid phase and liquid phase. We also tried to find the optimum amount of n-hexadecane in our batch system was about 400-1500 μL/ 100mL MSM for NTU-1 consumption. The concentration was about 1400-6600 μL/ L in liquid phase. It means that NTU-1 would trap n-hexadecane in biofloccules. If the concentration in liquid phase was less than 1400 μL/L, the morphological structure may be flakes; if the concentration in liquid phase was more than 6600 μL/L, there might be too much n-hexadecane in the medium to form bioflocculate formation. In this study, we also evaluated different environment factors affecting the irreversible adsorption of n-hexadecane, including pH, temperature, salinity and aging. Different adsorption isotherm would be found by regression. By using the adsorption isotherm we got, we could estimate the n-hexadecane amount in soil by analyzing the concentration in the liquid phase. We could also estimate the biodegradability by NTU-1. Under the optimal condition, NTU-1 could remove more than 80% out of n-hexadecane after 108 hours incubation. To sum up, NTU-1 provided a feasible and potential method to treat contaminated soil in the batch system.