Ash in lignocellulose is critical in interfering biomass pretreatment. Densifying Lignocellulosic biomass with Chemicals followed by Autoclave (DLCA) is a novel and efficient pretreatment method, whereas the specific effects of ash on DLCA pretreatment remain unclear. Herein, we quantitatively investigated the impacts of ash contents on the pretreatment effectiveness, enzymatic digestibility, and fermentability of DLCA pretreated corn stover (DLCA-CS). With the ash contents increasing from 3.36 % to 9.50 %, the DLCA-CS with addition of sulfuric acid (sa) showed decreased pretreatment efficiency and enzymatic digestibility, whereas these characteristics of DLCA-CS with addition of calcium hydroxide (ch) were improved with increasing ash contents. Further pH measurement during the biomass pretreatment revealed that the anions involved in ash neutralized the sulfuric acid, thus decreasing the pretreatment efficiency and enzymatic digestibility of DLCA(sa)-CS, while the anions can cooperated with the calcium hydroxide, thus increasing the pretreatment efficiency and enzymatic digestibility of DLCA(ch)-CS. The highest ethanol production of DLCA(sa)-CS (60.7 g/L) and DLCA(ch)-CS (66.5 g/L) at 25 % solid loading was achieved with a low ash content of 3.36 % without detoxification step. This study highlights the importance of controlling the ash contents for enhancing the efficiency of DLCA pretreatment.