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This research was conducted to investigate the comparative capabilities of microwave irradiation and chemical agents in the demulsification process of water-in-crude oil emulsion from Nigerian crude oil. Crude oil sample was obtained from a Niger Delta field in Isoko with API gravity of 23.52. The study was carried out in the laboratory of the Department of Chemical and Petroleum Engineering, Faculty of Engineering, Delta State University. The amine and the polyhydric alcohol group of demulsifiers were used as chemical agents. The results obtained in this study have exposed the capabilities of microwave irradiation and chemical methods in the destabilization of water in crude oil emulsions. All samples show separation of water and oil from water-in-oil emulsions as a function of time, respectively. All three replicates for the Microwave heating method were exposed to 40°C, 50°C and 60°C gave maximum water separation at an hour and 10 minutes, with replicate C exposed to highest temperature of 60°C giving highest separation rate of water from the emulsion at an early processing time of 5 minutes than replicates A and B showing that replicate C is less stable. Meanwhile, results from emulsion sample containing ethane 1, 2 diol is considered less stable than emulsions containing ethylene-di-amine-di-chloride, pentyl amine and ethane 1, 2, 3 triol, giving 67% water separation after 6 hours. Generally, results from the chemical processes averagely show that there is more stability of emulsion as against microwave irradiation method of separation. Separation of water-oil emulsion by microwave heating technology is suggested as alternative method because of its unique characteristic which is fast, very effective and enhancing gravity segregation to separate the emulsion into oil and water layers.