Using dioscorea root as substrate of Monascus species was found to stimulate monacolin K (cholesterol-lowering agent) formation in our previous study, but the mycotoxin−citrinin has never been studied. This study used dioscorea root as the liquid medium to culture Monascus purpureus NTU 568 using a 6.6 L jar fermentor. Culture pH value, dioscorea concentration, and ethanol concentration were used as the factors of response surface methodology (RSM) to investigate the optimal culture condition for high monacolin K production and low citrinin formation. Monacolin K and citrinin formation of M. purpureus NTU 568 under submerged dioscorea medium were respectively found to be significantly formed by 148% and 147%, as compared to that under submerged rice medium. The reason is due to the pH value (3.5) of dioscorea medium involved in the formation of Monascus cell amount and secondary metabolite. RSM results further indicated that lowering the pH value to 2.5 would result in high monacolin K and citrinin concentrations as well as high biomass in fixed dioscorea amount, implying that pH value may stimulate the formation of monacolin K and citrinin through increasing Monascus cell amount. Lowering dioscorea and ethanol concentration was able to increase the ratio of monacolin K level to citrinin level. The optimal culture condition (pH 5.7, 1% dioscorea concentration, and 0.5% ethanol concentration) would increase monacolin K levels to 27.9 mg/g (by 47%) and decrease citrinin level to 2.15 μg/g (by 54%), as compared to control conditions (pH 3.5, 5% dioscorea, and ethanol free).