Metabolic flux analysis was used to analyze the metabolic regulation mechanism of ethanol to promote the biosynthesis of astaxanthin. It was found that ethanol promoted the flux of pyruvate and acetyl-CoA metabolic nodules in the cell metabolism of Phaffia rhodozyma， and the flux of astaxanthin synthesis pathway was increased by 2.3 times of that of the control group， which promoted the synthesis of astaxanthin. According to the metabolic regulation mechanism， the metabolizing nodes of α-keto-glutaric acid and 5-phosphate ribulose were studied involved by the astaxanthin synthesis pathway of Phaffia rhodozyma JMU-MVP14. It was found that addition of 0.5 g/L α-keto-glutaric acid into culture broth could promote cell growth of Phaffia rhodozyma by 0.4 g/L. And astaxanthin yield reached 67.9 mg/L when 3 g/L glutamic acid was added into the broth， which was 1.7 times that of the control.