Microbial fuel cells (MFCs) need to be robust if they are to be applied in the field for bioremediation. This study investigated the effect of temperature (20–50 °C), salinity (0.5–2.5% (w/v) as sodium chloride), the use of redox mediators (riboflavin and anthraquinone-2-sulphonate, AQS) and prolonged fed-batch operation (60 days) on biodegradation of a petroleum hydrocarbon mix (i.e. phenanthrene and benzene) in MFCs. The performance criteria were degradation efficiency, % COD removal and electrochemical performance. Good electrochemical and degradation performance were maintained up to a salinity of 1.5% (w/v) but deteriorated by 35-fold and 4-fold respectively as salinity was raised to 2.5%w/v. Degradation rates and maximum power density were both improved by approximately 2-fold at 40 °C compared to MFC performance at 30 °C but decreased sharply by 4-fold when operating temperature was raised to 50 °C. The optimum reactor performance obtained at 40 °C was 1.15 mW/m2 maximum power density, 89.1% COD removal and a degradation efficiency of 97.10%; at moderately saline (1% w/v) conditions the maximum power density was 1.06 mW/m2, 79.1% COD removal and 91.6% degradation efficiency. This work suggests the possible application of MFC technology in the effective treatment of petroleum hydrocarbons contaminated site and refinery effluents.