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Yellow poplar was selected for alcohol-based biofuel production through alkaline hydrolysis, enzymatic saccharification and fermentation. In alkaline hydrolysis, 50.0% of biomass remained as residual, which chemical composition was 82.0% of cellulose, 17.0% xylan and 1.0% of lignin. Enzymatic saccharification led to almost complete conversion of cellulose and xylan in alkaline hydrolysis residue to monosaccharide. After enzymatic saccharification, the composition of hydrolysate was analyzed by HPLC. The sugar composition of enzymatic hydrolysate was 95.1 g/L of glucose and 21.4 g/L of xylose. The enzymatic hydrolysate also contained 0.5 g/L of acetic acid and 0.48 g/L of total phenolics. The furfural and 5-hydroxymethylfurfural (5-HMF) was not detected in this liquor. It means that the fermentation inhibitors such as phenolics and furans are not existed or under control. The hydrolysate from enzymatic saccharification was used for the production of ethanol and butanol by Saccharomyces cerevisiae and Clostridium beijerinckii, respectively. The ethanol production yield for sugar was about 58% in the fermentation of hydrolysate, which shows the same level of the control. In ethanol fermentation, glucose was completely utilized, where xylose was not consumed. The butanol was produced to the level of the control and xylose was consumed a little bit.