Background: This study quantified the effect of four popular culture media in a biodiesel production study on the qualitative and quantitative lipid content, dry biomass, and lipid productivity of Nannochloropsis oculata. Methods: Culture of microalgae was done separately in Walne, F/2, Sato, and TMRL media. In the logarithmic and stationary growth phases, biomass production and lipid accumulation of microalgae were measured and the constituents were identified by gas chromatography. Results: N. oculata exhibited the highest rate of cell growth and biomass productivity of 0.2616 day-1 and 2.652 gl-1 in the Walne medium. The highest level of biomass conversion into lipids in TMRL medium revealed a cell dry weight of 37.22%. Walne medium proved to have the most efficient lipid productivity which was 0.1057 gl-1 day-1. The highest amount of triacylglycerol (TAG) was obtained in Sato medium in the stationary growth phase and was 75.25% of the fatty acids. Conclusion: The present study provides a practical benchmark, which allows the introduction of Walne as a suitable culture medium for N. oculata in biodiesel studies.