Abstract: Using bamboo as raw material, the effect of formaldehyde/1, 4-dioxane pretreatment on the yield of enzymatic hydrolysis sugar from bamboo cellulose was studied. The producing process of lactic acid through fermentation with the enzymatic hydrolysate was also investigated. The results showed that the pretreatment could remove lignin and greatly increase the cellulose content of the pretreated bamboo substrate, with cellulose content reaching 75.6%-90.7%. The highest yield of cellulose-to-glucose by enzymatic hydrolysis after pretreatment was 92.8%. The enzymatic hydrolysate could be directly used for fermentation to produce lactic acid, and the yield of the glucose to lactic acid could reach 73.2%. The changes of bamboo structure before and after pretreatment were observed by scanning electron microscope(SEM), X-ray diffraction(XRD) and ion chromatograph. After pretreatment, the bamboo surface curled, the wax layer on the surface of bamboo was almost completely dissolved and removed, and the structure became loose.