In this paper, the components of amino acids in mixed starch wastewater (corn steep water/corn gluten water = 1/3, v/v) were analyzed by GC-MS. Effects of amino acids on lipid production by Rhodotorula glutinis and COD removal were studied. The results showed that mixed starch wastewater contained 9 kinds of amino acids and these amino acids significantly improved the biomass (13.63 g/L), lipid yield (2.48 g/L) and COD removal compared to the basic medium (6.23 g/L and 1.56 g/L). In a 5 L fermentor containing mixed starch wastewater as substrate to culture R. glutinis, the maximum biomass, lipid content and lipid yield reached 26.38 g/L, 28.90% and 7.62 g/L, with the associated removal rates of COD, TN and TP reaching 77.41%, 69.12% and 73.85%, respectively. The results revealed a promising approach for lipid production with using amino acids present in starch wastewater as an alternative nitrogen source. (C) 2016 Elsevier Ltd. All rights reserved.