In this study, the effects of different pretreatment methods on electricity generation in microbial fuel cell (MFC) were investigated. Organic fraction of municipal solid waste (MSWOF) was used as substrate and the samples were subjected to ultrasonication and heat/alkali pretreatment methods before being fed into the reactor. MFC was operated in fed-batch mode with seven days of retention time at 100 Ω external resistor. Diluted raw MSWOF sample, ultrasonically pretreated samples for 5 and 20 minutes and samples pretreated by heat/alkali using NaOH and KOH generated maximum power densities of 53.3 mW/m , 59.0 mW/m , 72.3mW/m , 130.7 mW/m and 114.4 mW/m and also soluble chemical oxygen demand (SCOD) removal efficiencies of 23,3%, 27,8%, 31,6%, 52,4% and 46,8% respectively. Power generation performance of MFC enhanced, as the SCOD value of substrate increased after pretreatment. Substrate subjected to heat/alkali pretreatment gave better results than ultrasonication in terms of power generation and organic matter removal. It is important to apply the proper pretreatment method regarding the type of substrate fed into the MFC in order to maximize overall efficiency of the system.