Microalgae are a potential biodiesel feedstock, as well as an important source of many high value-added of biological products. In this work, lutein, polysaccharide and lipid of Chlorella protothecoides were studied to improve the comprehensive utilization value. The lutein in Chlorella protothecoides was mostly in the form of free lutein, rather than lutein esters. It could maximally improve the comprehensive utilization value of Chlorella protothecoides in the extraction sequence of lutein> polysaccharide> lipid with lutein extraction ratio 66.21%, polysaccharide content 6.45% and lipid content 37.26%. Under optimal extraction conditions (ultrasonic power 280 W, ultrasonic broken time 10 min, solid to liquid ratio of 1:40 (w/v), and extraction 3 times), the lutein extraction ratio and content were 94.59% and 0.012%, while polysaccharide content was 6.43% and lipid content was 33.56%. The polysaccharide of Chlorella protothecoides had the highest antimicrobial activity against Escherichia coli with inhibition halo diameter 15.84 mm, and weak antimicrobial activities against Bacillus subtilis and Pseudomonas aeruginosa. The fatty acid composition of Chlorella protothecoides had no significant change after the extraction of lutein.