The objective was to determine the effects of three exogenous polysaccharide-degrading enzyme preparations (EPDE) on ruminal fermentation and forestomach and total tract digestion of dietary nutrients in dairy cows. Four late-lactation, ruminally and duodenally cannulated, Holstein cows were allocated to dietary treatments in a 4×4 Latin square design experiment. The basal diet fed to the cows contained (DM basis): 400g/kg alfalfa and grass hays, 44g/kg corn and barley grains, 80g/kg whole cottonseed, and 80g/kg protein and mineral/vitamin supplements. The EPDE preparations, a blank, a predominantly amylase, a predominantly xylanase and an amylase/xylanase combination were dosed to the rumen through the cannula daily, during the morning feeding (06:00h) at 10g/cow/day. Treatments did not affect ruminal pH, ammonia concentration, protozoal counts, total and individual VFA concentration, acetate:propionate ratio, and solid and fluid ruminal digesta passage rates. Xylanase, carboxymethylcellulase and amylase activities of whole ruminal contents at 2, 4, and 6h following EPDE application were also not affected by treatments. Intake of nutrients and forestomach true and total tract apparent digestibility of neutral detergent fiber and starch also did not differ among treatments. Digestibility of DM, organic matter and N were reduced (P=0.045–0.050) by the amylase/xylanase combination versus the amylase or xylanase EPDE. The EPDE dosed intraruminally at 10g/cow/day did not affect ruminal fermentation, polysaccharide-degrading activities of ruminal contents or total tract apparent digestion of nutrients.