Pigment contents and proximate compositions are important traits to evaluate the nutritional value of microalgae. The environmental condition and nutrient availability in the culture medium are hypothesised to regulate algal colour, nutritional composition and production. This study aimed to compare the pigment and proximate compositions in Tisochrysis lutea under phototrophic and mixotrophic conditions in an attempt to improve algal product quality. Algae were grown in the phototrophic and mixotrophic conditions at 50 μmol photons m−2 s−1 for 12 h daily, but 50 mM glycerol was added as a carbon source in the mixotrophic culture. The pigment contents in algae significantly increased in the mixotrophic condition, and the contents of chlorophylls a and c and carotenoids increased nearly 60 % compared with those in the phototrophic condition. The contents of saturated and polyunsaturated fatty acids in algae were significantly higher while the content of monounsaturated fatty acids was significantly lower in the mixotrophic condition than those in the phototrophic condition. Although the contents of protein, lipid and carbohydrate in T. lutea were not different, their overall production was significantly enhanced in the mixotrophic condition compared to the phototrophic condition. This study indicates that mixotrophic culture promotes pigment and proximate production and the change of fatty acid profile in algae depends on the addition of organic carbon to the culture medium.