Crude glycerol is a major byproduct of the biodiesel industry; previous research has proved the feasibility of producing docosahexaenoic acid (DHA, 22:6 n - 3) through fermentation of the algae Schizochytrium limacinum on crude glycerol. The objective of this work is to investigate the cell growth kinetics, substrate utilization efficiency, and DHA production of the algae through a continuous culture. Steady-state biomass yield, biomass productivity, growth yield on glycerol, specific glycerol consumption rate, and fatty acid composition were investigated within the range of dilution rate (D) from 0.2 to 0.6 day(-1), and the range of feed crude glycerol concentration (S(0)) from 15 to 120 g/L. The maximum specific growth rate was determined as 0.692 day(-1). The cells had a true growth yield of 0.283 g/g but with a relatively high maintenance coefficient (0.2216 day(-1)). The highest biomass productivity of 3.88 g/L-day was obtained at D = 0.3 day(-1) and S(0) = 60 g/L, while the highest DHA productivity (0.52 g/L-day) was obtained at D = 0.3 day(-1) and S(0) = 90 g/L due to the higher DHA content at S(0) = 90 g/L. The biomass and DHA productivity of the continuous culture was comparable to those of batch culture, while lower than the fed-batch culture, mainly because of the lower DHA content obtained by the continuous culture. Overall, the results show that continuous culture is a powerful tool to investigate the cell growth kinetics and physiological behaviors of the algae growing on biodiesel-derived crude glycerol. (C) 2010 Elsevier Ltd. All rights reserved.