In this study, the heterotrophic production potential of the secondary carotenoid lutein by the green microalgae Auxenochlorella protothecoides SAG 211-7a was investigated. A sequential statistical technique was applied to optimize modified bold’s basal media (MBB) to enhance the lutein production from microalgae Auxenochlorella protothecoides SAG 211-7a. Taguchi orthogonal array method was applied to select the various independent variables which affect the lutein production. It showed that sucrose, yeast extract, MgSO4.7H2O and EDTA were the significant factors affect the lutein production. Further, to increase the lutein yield and to study the interaction between these factors response surface methodology (RSM) was employed. The statistical model was validated with respect to lutein production under the conditions predicted by the model containing sucrose 14.0 g/l, yeast extract 3.0 g/l, MgSO4.7H2O 0.8 g/l and EDTA 0.76 g/l. The production of lutein obtained experimentally using the above medium was 1303 ± 25.32 μg/l, which is in correlation with the predicted value of 1337.21 g/l by the RSM regression study. Thus after sequential statistical media optimization strategy a 5-fold enhancement in lutein production was achieved.