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Microorganisms such as bacteria and microalgae are the most viable sources for the production of sustainable energy since they lack the major drawbacks of first and second generation biofuels. Microalgae have the capacity to store large quantities of lipids that can be used in the production of biodiesel. Due to the advantages microalgae present visa-vis other raw material sources in the production of biodiesel, there are great efforts to overcome technological and economical obstacles to develop this technology on an industrial scale. In this chapter the effect of nitrogen limitation and CO2 supply in the fatty acids profiles of microalgal oils from Dunaliella tertiolecta and Nannochloropsis oculata was determined. Nannochloropsis oculata and Dunaliella tertiolecta were cultured under different nutrient conditions and culture systems. In the photobioreactor culture, D. tertiolecta accumulated mainly C16: 0 (42%), C18: 0 (32%), and C18: 3 (10%), whereas N. oculata accumulated C16: 0 (41%), followed by C16: 1 (21%) and C18: 0 (10%). Combining the variation in the NO3 and CO2 concentrations did not modify importantly the fatty acids composition of D. tertiolecta in any of the analyzed levels. With sudden nitrogen limitation, the proportion of C18: 3 in D. tertiolecta increased from 10 to 30% of the fatty acids. In N. oculata no important change occurred in the fatty acids composition up to 72 h of limitation. In the semicontinuous raceway culture, N. oculata presented predominantly C16: 0 (37%), followed by C16: 1 and C18: 0. The C16: 1 and C18: 0 proportion was very different between the photobioreactor and the raceway cultures. C16: 1 proportion …