Lignocellulosic biomass, such as wood and agricultural residues, is an attractive material for fuel ethanol production since it contains large amounts of potentially fermentable sugars in the form of cellulose and hemicellulose. The interest of using hemicellulose as a resource for the efficient production of ethanol is currently increasing. Softwood is one of promising feedstock, since its principal hemicellulose component, O-acetyl galactoglucomannan, gives readily fermentable mannose as the major sugar after hydrolysis. This is advantageous over hemicellulose from hardwood or agricultural residues hydrolyzed to unfermentable xylose. In this study, we investigated an efficient enzymatic hydrolysis of both glucan and mannan components in a ball mill (BM)-treated softwood, Douglas-fir. A culture supernatant from Acremonium cellulolyticus CF-2612, which is a powerful cellulase-producing fungus, was used as the cellulase (ACase-CF) for the enzymatic hydrolysis. The glucan digestibility of BM-treated Douglas-fir was estimated to be 75% using 10 FPU/g-substrate ACase-CF, but mannose from the mannan component was hardly detectible in this hydrolysate. A supplemental commercial hemicellulase was essential to the mannan hydrolysis of BM-treated Douglas-fir. Two enzyme components, β-mannosidase and β-mannanase, were purified from the commercial hemicellulase and their effects on the mannan digestibility were evaluated. The results showed that a synergistic effect between ACase-CF and these mannan-degrading enzymes significantly improved the mannan hydrolysis. The glucan and mannan digestibility of BM-treated Douglas-fir reached 75% and 70%, respectively, using the enzyme cocktail consisting of 10 FPU/g-substrate ACase-CF, 1 U/g-substrate β-mannosidase, and 200 U/g-substrate β-mannanase.