Previously, overall protein assimilation after the ingestion of a pure protein meal was studied. In this study, the kinetics of protein assimilation in humans were investigated after the ingestion of a complex meal, which more closely represents a physiologically normal situation. Overall protein assimilation in humans after the ingestion of a pancake meal, containing 12 g of fat, 27 g of carbohydrate, and 19 g of protein, was evaluated in 26 normal volunteers. Both the egg white and yolk of L-[1-(13)C]-leucine-substituted eggs were used to make the batter. The labeled eggs were produced by feeding laying hens a standard chicken diet supplemented with 3 g/kg of L-[1-(13)C]-leucine (99%, mol:mol). High enrichment levels of protein with adequate labeling patterns were obtained in eggs from laying hens fed the L-[1-(13)C]-leucine-substituted diet. The isotopic enrichment of leucine at plateau was equal in egg white and yolk. The overall tracer recovery in egg proteins was 22.5%. The overall protein assimilation parameters in subjects that consumed the pancake meal did not differ from those obtained in subjects that consumed a single protein meal (mean cumulative (13)C recovery/6 h = 17.22 +/- 4.74%, with a maximal (13)C recovery/h of 5.65 +/- 1.48%, which was attained 145 +/- 25 min after ingestion of the meal). The pancake test meal prepared with eggs intrinsically labeled with L-[1-(13)C]-leucine is ideal for the study of protein assimilation. The incorporation of differently labeled substrates into a single test meal allows the assessment of different gastrointestinal processes in the overall assimilation of proteins.