Alfalfa saponins isolated by ethanol extraction and partial acid hydrolysis were intraruminally administered to sheep to investigate the biological activities and nutritional implications in ruminants. Mature wethers fitted with ruminal, duodenal and ileal cannulas were fed a concentrate or roughage diet formulated to be isonitrogenous and isoenergetic. Microbial fermentation and nutrient degradation in the rumen were reduced by saponins. Total protozoal count in the rumen was reduced 34 and 66% by saponins at levels of 2 and 4% dietary dry matter, respectively. Bacterial nitrogen flow to the duodenum was reduced 20 and 30% in the same sequence. Apparent digestion coefficients of organic matter, hemicellulose and cellulose in the total digestive tract were increased by saponins in sheep fed concentrate diets. Fractional digestion coefficients of organic matter, hemicellulose, cellulose and nitrogen were reduced in the stomach while they were increased in the small intestine by saponins in both diets. Saponins inhibited microbial fermentation and synthesis in the rumen and altered the sites of nutrient digestion in sheep.