Efficient glutathione production by high-cell-density cultivation of Candida utilis was investigated. A series of batch glutathione fermentations were carried out and the optimal initial glucose concentration was found to be about 26 g/L. Then, fed-batch fermentation under diverse feeding strategies was used to enhance glutathione production with a total glucose concentration of 150 g/L. Constant glucose feeding strategy cannot meet the requirement of cells at the late period of feeding, while exponential glucose feeding strategy cannot satisfy the needs of cells at the beginning of feeding. Based on the results above, a polynomial glucose feeding strategy was developed to provide enough glucose for cells along with the cultivation, under which both the cell and glutathione productivity were satisfactorily improved. Furthermore, fed-batch fermentation under this strategy with a total glucose concentration of 200 g/L was successfully performed, the dry cell weight and glutathione concentration reached 91.2 g/L and 825 mg/L, respectively.