The effects of different wort gravity or ethanol concentration in initial wort on the fermentation performance of lager yeast and assimilation of free amino acids (FAAs) were studied. Results showed that compared with high wort gravity (24 degrees P), high ethanol concentration (10%, v/v) decreased yeast growth, cell viability, and wort fermentability significantly. The assimilation of FAAs was changed dramatically by high ethanol toxicity, and positive correlations between the assimilation amounts of 10 FAAs (Asp, Ser, Gly, Arg, Tyr, Val, Met, Lys, Ile, and Leu) and fermentation performance (cell viability, fermentability, and ethanol production) were identified, especially for Arg and Lys exhibiting extremely significant positive correlations. Furthermore, confirmatory testing was carried out by supplementing 24 degrees P worts with 10 FAAs of 0.5, 1, and 2 times of their standard concentrations, respectively. Results exhibited that 10 FAA supplementations improved physiological characteristics and fermentation performance of lager yeast significantly, especially for 1 times FAA supplementation increasing wort fermentability and ethanol yield by 6 and 17%, respectively, and upregulated the expression level of HSP12 and increased more intracellular trehalose accumulation in yeast cells, indicating that stronger protective function was stimulated in yeast cells. Therefore, it was suggested that these 10 FAAs could regulate yeast cells to adapt to high gravity environmental stresses.