. 1999. Effect of diet, digesta processing, freezing and extraction procedure on some polysaccharide-degrading activities of ruminal contents. Can. J. Anim. Sci. 79: 73-81. Three procedures for extracting polysaccharide-degrading enzymes (PDE) from ruminal contents were compared, and the effects of diet, inoculum preparation and freezing of samples on recovery of enzyme activities were evaluated. Four nonlactating cows and four heifers were fed either high-forage (83%, dry matter [DM] basis) or high-grain (91%, DM basis) diets. Three sample types, (1) enriched ruminal fluid; (2) blended enriched ruminal fluid; and (3) whole ruminal content, were analyzed fresh or after having been frozen (-40°C) for 48 h. Four cell-free preparations of each sample type were assayed: (1) control (no enzyme extraction); and enzymes extracted by (2) sonication; (3) lysozyme; and (4) lysozyme + EDTA. Animals fed a high-grain diet had lower carboxymethylcellulase and xylanase but higher amylase activities in the rumen. Blending ruminal solids with buffer and combining the filtrate with cheesecloth-filtered ruminal fluid produced higher polysaccharide-degrading activities per unit of DM compared with whole ruminal contents. In lysozyme treatments, freezing inactivated from 22 (amylase) to 52% (xylanase) of the enzyme activities measured in fresh ruminal fluid. Lysozyme treatment resulted in the highest enzyme recovery (from fresh ruminal fluid) of the extraction methods tested. With frozen samples, sonication and lysozyme gave similar results. If a large number of ruminal content samples are to be processed for PDE activities, freezing and subsequent sonication appear to be a practical alternative to extraction by lysozyme. Le passage au mixer des matières solides ruminales tamponnées et leur combinaison avec le filtrat ruminal donnaient lieu à des activités de dégradation des polysaccharides plus fortes par unité de m.s. que le contenu ruminal entier. Dans les traitements avec la lysozyme, la surgélation inactivait, de 22 % dans le cas de l'amylase, à 52 % dans celui de la xylanase, les activités enzymatiques dans le liquide ruminal frais. De toutes les méthodes d'extraction comparées, c'est le traitement avec la lysozyme qui produisait la récupération enzymatique la plus forte (à partir de liquide ruminal frais). Mais quand on utilisait des échantillons surgelés, la sonication et le traitement avec la lysozyme donnaient des résultats comparables. Si l'on doit préparer un grand nombre de prélèvements du contenu ruminal pour la mesure des activités EPD, la surgélation suivie de sonication apparaît comme une alternative pratique à l'extraction par la lysozyme.