We describe the characterization of the microalga Glossomastix chrysoplasta, an eicosapentaenoic acid (EPA) producer in the Pinguiophyceae class, Chromophyte division. Growth conditions were selected to optimize algal growth and EPA production. EPA represented up to 30% of the fatty acid content of Glossomastix chrysoplasta, at levels of 22 mg EPA per gram dry weight. Up to 72% of the EPA was produced as glycolipids, components of structural lipids. The optimal growth conditions in continuous culture were found to be greater than 500 mu mol photon s/m(2) (.) s light intensity, 0.33/day dilution rate, pH 7.20-7.45, and a temperature of 18-20 degrees C. Macronutrient studies indicated the limiting nutrient to be bicarbonate or dissolved carbon dioxide, and consequently decreasing pH increased EPA production. (c) 2005 Wiley Periodicals, Inc.