The concentration of NaNO3, MgSO4 center dot 7H(2)O and proteose, in Chlorella protothecoides medium were optimized for algal biomass and lipid production by using response surface methodology with Box-Behnken design. The optimal concentrations were 0.45 g/L of NaNO3, 6 mg/L of MgSO4 center dot 7H(2)O, and 0.25 g/L of proteose for maximum biomass production and 2 mg/L of MgSO4 center dot 7H(2)O and no addition of NaNO3 and proteose for lipid accumulation. In optimized biomass production medium, a final biomass concentration of 1.19 g/L was obtained, which was 1.8 times higher than that in the original medium. For lipid accumulation, a 12.9% lipid content was obtained from the biomass in the lipid production medium, which was three times higher than that from the original medium. The fatty acid profile of algae grown in the optimized medium demonstrated a higher unsaturated fatty acid content (i.e. methyl linoleate (C18:2) and methyl linolenate (C18:3)) than that of the algae grown in the original medium. The results provide a strategy for limiting the amount of nutrients required in large scale outdoor cultivation systems of C. protothecoides to make the production of algal biomass more economically attractive. (C) 2012 Elsevier Ltd. All rights reserved.