The lipid production and biomass growth of microalgae Isochrysis galbana was investigated under the effects of different medium temperature and carbon dioxide levels. Algae production was studied in ten-liter, open-air, aerated bioreactors and sampling was done daily to monitor the cell growth. At the end of each fermentation, the dry mass, lipid amount and fatty acid profiles were determined. The algae were grown according to designed experiments (central composite design (CCD)) at 15, 22.5 and 30 °C medium and aerated with air containing 5, 10 and 15% v/v CO2. It was observed that in 15 °C experiments, the fatty acid profile was formed mainly by unsaturated fatty acid (both mono and poly unsaturated) and the amount of saturated fatty acids increased by increased CO2 levels. The saturated fatty acid content increased with increasing growth temperature. Dominantly observed fatty acids are Pentadecanoic Acid (C15:0), Pentadecanoic Acid (C15:0), Palmitic Acid (C16:0), Linolenic Acid (C18:3), Oleic Acid (C18:1). In the algal productions performed at 22.5 °C and 30 °C medium conditions, the fatty acid content was mainly formed by saturated fatty acids. However, the amount of lipid produced increased with increasing temperatures. The highest cell growth was seen at 15 °C experiments. The lowest growth rate was observed at 30° C experiments. It was observed that the medium temperature had the largest effect for the production of saturated and unsaturated fatty acids. The carbon dioxide was found effective linearly on the fatty acid production.