The influence of the oxygen and glucose supply on primary metabolism (fermentation, respiration, and anabolism) and astaxanthin production in the yeast Phaffia rhodozyma was investigated. When P. rhodozyma grew under fermentative conditions with limited oxygen or high concentrations of glucose, the astaxanthin production rate decreased remarkably. On the other hand, when the yeast grew under aerobic conditions, the astaxanthin production rate increased with increasing oxygen uptake. A kinetic analysis showed that the respiration rate correlated positively with the astaxanthin production rate, whereas there was a negative correlation with the ethanol production rate. The influence of glucose concentration at a fixed nitrogen concentration with a high level of oxygen was then investigated. The results showed that astaxanthin production was enhanced by an initial high carbon/nitrogen ratio (C/N ratio) present in the medium, but cell growth was inhibited by a high glucose concentration. A stoichiometric analysis suggested that astaxanthin production was enhanced by decreasing the amount of NADPH required for anabolism, which could be achieved by the repression of protein biosynthesis with a high C/N ratio. Based on these results, we performed a two-stage fed-batch culture, in which cell growth was enhanced by a low C/N ratio in the first stage and astaxanthin production was enhanced by a high C/N ratio in the second stage. In this culture system, the highest astaxanthin production, 16.0 mg per liter, was obtained.