Long-chain polyunsaturated fatty acids are important components of human milk that seem to influence infant development. After oral administration of U-13C-labeled linoleic acid to a lactating woman the recovery of tracer in milk linoleic acid was 6.4%, whereas tracer recovery in dihomo-gamma-linolenic acid (DGLA) was 0.3% and in arachidonic acid (AA), <0.01%. Some 14.9% of linoleic acid intake was converted to breath-CO2. In combination with data on dietary intake and quantitative milk output, we estimate that 23% of milk linoleic acid, 7% of milk dihomo-gamma-linolenic acids and 0.5% of milk arachidonic acid were contributed by direct endogenous conversion and transfer from dietary linoleic acid. These findings lead us to conclude that long-chain polyunsaturated fatty acids for human milk synthesis are produced in the body, but either a relatively low percentage is contributed under these dietary conditions, or there are large body pools for intermediate storage of these fatty acids that are utilized for secretion into milk.