Thirty lactating Holstein cows were in a continuous trial from 21 to 120 days postpartum to evaluate diets containing whole, rolled sunflower seeds with or without additional limestone. Cows were fed individually total mixed rations of (dry matter) 47% corn silage, 9% alfalfa hay, and 44% concentrate. Concentrates were corn and soybean meal (control); corn, soybean meal, and 22% sunflower seeds; or corn, soybean meal, and sunflower seeds plus 3.5% additional limestone. Milk yield (32.2, 32.0, and 32.8 kg/day) was similar among rations. Yield of 4% fat-corrected milk was lower for cows fed sunflower seeds without additional limestone (30.2, 28.1, and 30.2 kg/day) because of lower milk fat percentages (3.57, 3.19, and 3.51). Milk protein percentage tended to be lower for cows fed sunflower seeds with additional limestone (3.01, 2.97, and 2.90). Milk, flavor score was acceptable but tended to be lower for milk from cows fed sunflower seeds with additional limestone (8.4, 8.5, and 7.9). Milk fat from cows fed sunflower seed rations contained less carbon-14:0, 16:0, and 16:1 fatty acids but more carbon-18:0. Dry matter intakes were 21.0, 18.4, and 20.0 kg/day. Dry matter digestibilities, body weight changes, and ruminal volatile fatty acid concentrations were similar among treatments. Total cholesterol in blood serum was elevated in cows fed sunflower seed rations. Insoluble salts of fatty acids were increased in ruminal fluid dry matter from cows fed sunflower seeds but were not increased further by additional limestone. Concentrations of nonesterified carbon-18:1 fatty acids in ruminal fluid dry matter were lower for cows fed sunflower seeds with additional limestone.