Six mature Holstein cows in late lactation and fitted with permanent ruminal, duodenal and ideal cannulae were used in a double 3 X 3 Latin-square design to study the effect of roasting or extruding of cottonseed on protein degradation in the rumen and availability of amino acids in the small intestine. Raw whole cottonseed (CS), extruded whole cottonseed (ECS), and roasted whole cottonseed (RCS) were fed in diets containing 17% crude protein and composed of 42% whole cottonseed, 26% corn grain and 29% hay supplemented with dry molasses, vitamins and minerals. The diets were isocaloric and contained 1.86 Mcal NEL/kg. Acid detergent lignin was used as a digesta marker, and diaminopimelic acid as a bacterial marker. Ruminal ammonia concentration was higher (P less than .05) for CS than for ECS and RCS (15.7, 12.7 and 10.2 mM). Concentrations of isobutyrate (1.3 vs 0.8 and 0.8 mM) and isovalerate (1.8 vs 1.1 and 1.1 mM) were higher (P less than .05) for CS compared with ECS and RCS. Non-ammonia nitrogen flow to the duodenum (g/d) was 248, 300 and 350 for CS, ECS, and RCS, with RCS higher (P less than .05) that in CS. Apparent absorption of total amino acids in the small intestine was .80, 1.01, and 1.24 kg/d, for CS, ECS, and RCS, respectively, with RCS higher (P less than .05) than CS. Apparent absorption of essential and non-essential amino acids was 376, 425; 458, 548;610 and 628 g/d for CS, ECS and RCS respectively, with RCS higher (P less than .05) than CS.