Rhizoma gastrodiae ethanol extracts were supplemented into the fermentation system of Grifola frondosa to study effects of R.gastrodiae ethanol extract on mycelia biomass, mycelia polysaccharides and β-glucan yield of Grifola frondosa by submerged fermentation. In addition, the key ingredients in Rhizoma gastrodiae ethanol extracts were identified. Meanwhile, the appropriate concentration of characteristic constituents of R.gastrodiae ethanol extracts were determined by high performance liquid chromatography. The results showed that R.gastrodiae ethanol extracts with the concentration of 7g/L could significantly promote cell growth, mycelia polysaccharides, and β-glucan yield. Under this concentration, the biomass, mycelia polysaccharides and β-glucan yield reached 1.708g/L, 5.974% and 2.055mg/g, and significantly (P<0.05) increased by 0.36,1.29, and 1.44 times, respectively, compared with the control (without additives). The characteristic ingredients of R.gastrodiae ethanol extracts were gastrodin (GA), p-hydroxybenzyl alcohol (HA), and p-hydroxylbenzaldehyde (HBA), and the contents were 5.8375,1.1072, and 0.6601mg/g, respectively. Meanwhile, HBA was the most responsible ingredients to promote the biomass, mycelia polysaccharides and β-glucan yield in submerged culture of Grifola frondosa and the optimum supplemental levels were 100,0, and 150mg/L, respectively.