The effect of dietary linoleic (C18:2n-6) and palmitic acids (C16:0) on rate of hepatic de novo fatty acid synthesis was assessed in normal subjects. The diet was formulated to provide combinations of high and low levels of C18:2n-6 and C16:0. After 21 days of diet treatment, plasma triacylglycerol level and incorporation of deuterium into the plasma very low density lipoprotein triacylglycerol (VLDL-TG) pool over 24 hours was measured. Plasma triacylglycerol levels were within the normal range. Increasing dietary intake of linoleic acid decreased plasma triacylglycerol level when subjects consumed a low level of dietary palmitic acid. The relative and net amount of de novo synthesized fatty acid in the plasma VLDL-TG pool was not influenced by the diet treatments. A relationship between plasma triacylglycerol level and rate of hepatic de novo fatty acid synthesis was observed.