In the present study, the removal performance of phenol and Chemical Oxygen Demand (COD) from a petroleum refinery wastewater (PRW) in a Rotating Biological Contactor (RBC) was investigated. The impact of temperature of wastewater (T) (35–45ºC), rotational speed (ω) (2–14 rpm) and disc submergence (Ω) (30–50%) on phenol and COD removal was examined. The response surface methodology (RSM) was applied to minimize the number of runs and investigate the optimum operating conditions. Seventeen runs were carried out and the optimum conditions for phenol and COD removals were statistically obtained at temperature of 35°C, rotational speed of 11 rpm and disc submergence of 46%. These predicted data were in good agreement with the observed ones, as well. The result indicated that, the removal efficiency of phenol, cyanide, ammonia nitrogen, hydrogen sulfide, COD, biological oxygen demand (BOD5), total dissolved solid (TDS), total suspended solid (TSS), total organic carbon (TOC) and turbidity respectively were under the 99%, 82%, 40%, 93%, 89%, 87%, 76%, 85%, 55% and 58%, in the optimum conditions that mentioned above. The major group of microbes (for the phenol microbial treatment) isolated from effluent of American Petroleum Institute (API) separator identified as Bacillus and Pseudomonas species. Furthermore, indigenous bacteria no need to acclimate. It also produced low sludge compared with the activated sludge in the RBC. Therefore, this type of bacteria was successfully applied in this research.