Oil refinery waste activated sludge produced from oil wastewater biological treatment is a major industrial sludge. Two-phase anaerobic digestion of oil refinery waste activated sludge was studied for the first time. Thermal pretreatment under 170 °C is effective on sludge solubilization. At the optimum hydrolytic-acidogenic condition which was pH of 6.5, temperature of 55 °C and HRT of 2 days, 2754 mg/L volatile fatty acids (VFAs) were produced and acetic acid and butyric acid were the key components. Comparative studies of single-phase and two-phase anaerobic digestion in terms of organic removal, biogas production and methane concentration were conducted. The cumulative methane production and soluble COD (SCOD) removal efficiency in the two-phase system were 228 mL/g COD added and 77.8%, respectively, which were 1.6 and 2.1 times higher than those in single-phase anaerobic digestion. Such improved performance is attributed to intensification of dominant microbial population in separated reactors. Caloramator, Ureibacillus, Dechloromonas, Petrobacter, and T78 played important roles in hydrolytic-acidification and oil-organics degradation. Syntrophic bacteria in the family Porphyromonadaceae and the genus Anaerobranca provide acetate for methanogen. The results demonstrated the potential and operating condition of two-phase anaerobic digestion in treatment of oil refinery waste activated sludge.