Choline metabolism in breast cancer cells and tumors has been investigated by multinuclear NMR in order to provide the biochemical basis for the presence of high phosphocholine in breast carcinoma relative to benign breast tumors and normal breast tissue. Choline was found to be transported into MCF7 human breast cancer cells and rapidly phosphorylated to phosphocholine which was then accumulated in the cells to high concentrations. The increased level of phosphocholine did not affect the rate of synthesis of phosphatidylcholine, indicating tight regulation of this pathway. The incorporation of [1,2-13 C]choline (100 vM) into phosphocholine and phosphatidylcholine after 24 h was 69.5 and 36% of the total respective pools. Incorporation of 2 H 9 -choline to tumors implanted in nude mice was achieved by infusing the deuterated choline to the blood circulation. The metabolism of deuterated choline was then monitored by 2 H localized MRS. The blood level of choline before the infusion was 58.6 9 10.3 vM (measured by 1 H-NMR of plasma samples) and increased 5-fold during the infusion (measured by 2 H-NMR). This increase in the blood level resulted in a gradual increase of a signal at 3.2 ppm due to deuterated choline metabolites. It appears that the increased availability of choline in the blood circulation leads to accumulation of phosphocholine in the tumors by the same mechanism as in the cells.