We examined the circadian variation of holo-transcobalamin (holo-TC), total transcobalamin (total TC) and plasma cobalamins, and the association between food intake and fluctuations in the biochemical markers. A total of 17 healthy women (mean age 33 years; range 24-40) participated. The subjects were admitted at 08:00 h after an overnight fast, and blood samples were obtained every 20 min for 24 h from 12:00 h. Holo-TC and total TC were measured by an ELISA, and plasma cobalamins were measured by a routine method (Centaur, Bayer). All subjects, except one, had 12:00 h levels within the reference interval for all variables studied, and all subjects had normal renal function as judged from plasma creatinine. We found a small intra-individual variation (12%, 10%, 10%) and a considerable inter-individual variation (56%, 26%, 43%) in the level of holo-TC, total TC and plasma cobalamins, respectively. During the night, the absolute values of all components decreased, as did plasma albumin. There was no systematic association between intake of food and fluctuation of the biochemical markers. In conclusion, we found no significant circadian variation in serum holo-TC in healthy vitamin-replete subjects on a standard diet. This supports the view that holo-TC is a marker of long-term vitamin B(12) status, and that samples may also be obtained from non-fasting subjects.