This experiment was conducted to determine the variations in carotenoid, vitamins A and E concentrations, and color in the plasma and milk of dairy cows following a shift from a hay diet to diets containing increasing levels of carotenoids and vitamin E. This study was performed on 32 multiparous Montbéliarde dairy cows in midlactation. After a 6-wk preexperimental period on a diet based on hay and concentrates, the cows were allocated to 4 homogeneous groups, and thereafter fed for 6 wk on isoenergetic experimental diets where the hay was replaced by an experimental feed rich in carotenoids and vitamin E, consisting in 75% grass silage and 25% alfalfa protein concentrate (PX Agro Super Desialis, Châlons en Champagne, France). The hay-to-experimental feed ratios were 100/0 in group 1, 67/33 in group 2, 33/67 in group 3, and 0/100 in group 4, providing 1.6, 3.6, 5.4, and 7.4 g/d of total carotenoids, respectively. Variations in carotenoid, vitamins A and E concentrations as well as variations in color index (CI) were monitored from d -7 through to d 42 on the experimental diets. Zeaxanthin, lutein, 13-cis-beta-carotene, and all-trans-beta-carotene accounted for an average 3, 10, 9, and 78%, respectively, of total carotenoids in plasma and 0, 17, 12, and 71%, respectively, of total carotenoids in milk. The switch from preexperimental to experimental diets only slightly affected zeaxanthin, lutein, and vitamin A concentrations in plasma and milk. A rapid increase in vitamin E and beta-carotene (BC) was observed during the first week in both plasma and milk. For vitamin E, the time to reach a plateau was from 8 d (group 2) to 28 d (group 4) in plasma, and 5 d (groups 2-4) in milk. Plasma concentrations of BC had stabilized after 28 d in group 2 but were not stabilized after 42 d in groups 3 and 4, whereas milk concentrations of BC plateaued from d 21 in group 2 and d 28 in groups 3 and 4. At the end of the experimental period, BC and vitamin E concentrations in plasma and vitamin E concentrations in milk fat were linearly related to the proportion of experimental feed in the diet. In contrast, BC concentrations in milk fat did not differ between groups 2, 3, and 4, reflecting saturation at high levels of carotenoid intake (i.e., when plasma BC exceeded 5 mug/mL). These results suggested that under high-carotenoid diets, milk secretion of BC is not limited by the amount of plasma BC arriving to the mammary gland but by mechanisms involved in the BC transfer from plasma to milk. These mechanisms will need to be investigated. The BC concentrations were responsible for more than 80% of CI variations in plasma and 56% of CI variations in milk, where there was wide variability among individuals. Plasma CI appeared to be a more promising tool than milk CI as an indicator of the carotene content of the diets ingested by dairy cows.