Treatment of forage with enzyme mixtures can increase rate of degradation in vitro (Colombatto et al., 2000a). However, the complexity of natural forage makes it difficult to determine what fractions are most affected by enzyme treatment. The use of pure substrates (e.g. cellulose and xylan), provides a way of evaluating enzyme mode of action. Therefore, the present study examined the effects of pre-treating avicel and xylan with an enzyme mixture (EM) on a) the reducing sugars produced during treatment before rumen fermentation and b) the gas production profiles during in vitro rumen fermentation using the Reading Pressure Technique (RPT) (Mauricio et al., 1999).