Fungal invasion is a main factor leading to the loss of postharvest herbs during storage. To prevent fungal invasion and clarify the favorable conditions for the stability of herbs during the storage period, uniform design and three-dimensional response surfaces were applied to investigate the relationship of the mildew degree of Radix Ophiopogonis induced by prevalent fungal isolates and the storage factors including air relative humidity (X(1)), temperature (X(2)), and moisture content (X(3)), in laboratory studies. Mildew degree was evaluated by ergosterol assay of mold isolates through a high-performance liquid chromatography-diode array detector (HPLC-DAD) technique. As a result, storage conditions, which satisfied the following correlation equations established by the relationships between the storage factors and the biomass of mildew (Y), could effectively prevent fungal spoilage: Y = -0.2484084028 + 0.00030711966572 x X(3) x X(3) + 0.00018881361186 x X(1) x X(2) - 0.00029473040679 x X(2) x X(3) <or= 0; Y = -0.2697457586 + 0.004127756022 x X(1) - 0.000015977780492 x X(1) x X(1) + 0.00021906984606 x X(2) x X(3) <or= 0; Y = -0.325655811 + 0.015464432582 x X(2) + 0.00004779394354 x X(1) x X(1) - 0.00021743815482 x X(2) x X(2) <or= 0. This compositive methodology might be useful to predict the occurrence of fungal invasion.