A 2 × 2 factorial experiment was conducted to investigate the interaction between cereal type (wheat v. barley) and an exogenous enzyme supplement (with or without) on nutrient digestibility, large intestinal microflora, volatile fatty acid profile and in vitro manure ammonia emissions from finisher pigs. The enzyme supplement used contained endo-1, 3-β-glucanase (EC 3·2·1·6) and endo-1, 4-β-xylanase (EC 3·2·1·8). The diets were formulated to contain similar concentrations of net energy (9·8 MJ/kg) and lysine (10·0 g/kg). Urine and faeces were collected over seven consecutive days from 16 boars (four boars per treatment, 80·0 kg live weight) that were housed in metabolism crates. After collections, the pigs were slaughtered and the contents of the intestinal tracts were removed for analysis. There was a significant interaction between cereal type and enzyme inclusion in the apparent total tract digestibility of dry matter (DMD), organic matter (OMD) and nitrogen. The inclusion of an enzyme supplement in barley-based diets increased (P < 0·05) DMD, OMD and nitrogen digestibility compared with unsupplemented diets, however there was no effect of enzyme supplementation in wheat-based diets. There was a significant interaction between cereal type and enzyme inclusion in selected components of the gut microflora. Pigs offered unsupplemented barley-based diets had higher populations of bifidobacteria (P < 0·05) in the caecum and colon than those on the enzyme supplemented barley diet, however, there was no effect of enzyme supplementation on bifidobacteria in wheat-based diets. There was a significant interaction between cereal type and enzyme inclusion in volatile fatty acid production and in in vitro ammonia emissions. In the absence of an enzyme supplement, barley-based diets reduced the proportion of isovaleric acid (P < 0·05) and isobutyric acid (P < 0·05) in the caecum and colon and also reduced manure ammonia emissions during storage from 0 to 240 h (P < 0·05) compared with the wheat-based diet, however there was no effect of cereal type in enzyme-supplemented diets. In conclusion, the inclusion of an enzyme in barley-based diets increased nutrient digestibility but also increased ammonia emissions.