Microalgal oil is a potential energy source because it can be easily converted to fatty acid methyl ester or hydrocarbon type of diesel, and it is produced with relatively higher productivity compared with oil from plants and animals. Heterotrophic growth of microalgae is superior due to its high final product concentration; however, the cost of the raw materials is unacceptable if sugar is utilized as the carbon source. The aim of this study is to optimize the lipid accumulation of Chlorella protothecoides by using carbon sources other than glucose in heterotrophic and mixotrophic cultures. Different factors such as different carbon sources, carbon to nitrogen ratio, initial pH level, salinity, and rotational speed are studied in affecting the cell growth and the oil accumulation. Our experiments revealed that the heterotrophic and mixotrophic cultures of C. protothecoides grew better than autotrophic cultures. C. protothecoides can grow on glycerol or acetate, as well as on glucose. Several stress factors were confirmed or discovered to significantly increase the lipid content of microalgae cells. The replacement of glycerol and acetate as carbon sources for microalgae cultivations provides potential for waste utilization: glycerol from biodiesel industry and acetate from biohydrogen production.

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