Free AA in plasma and muscle were monitored in 36 dairy cows from 2 wk before until 15 wk after parturition. Cows were kept indoors and fed individually. The AA concentrations in plasma from 6 to 15 wk of lactation were compared with precalving concentrations. Plasma concentrations of Met, Phe, Glu, and Gln decreased by 16, 24, 25, and 25%, respectively. The ranking of the essential AA according to their decrease in plasma corresponded to the order in which they generally appear to be limiting for milk protein synthesis. The decrease of Glu and Gln in plasma exceeded that of the essential AA and contrasted strongly with all other nonessential AA. The change in the AA profile in muscle from pregnancy to lactation resembled that described for the catabolic state in mammals and suggests that muscle protein was degraded for supply of AA to the udder, despite excess protein and energy supply 15 wk into lactation. In muscle, the decrease in the pool of free Gln exceeds 25% and is higher than for any other AA. These observations support our hypothesis that Gln is potentially limiting for milk protein synthesis in the high yielding dairy cow.