Lymphatic transport of eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids given as trieicosapentaenoyl glycerol (TriEPA) and tridocosahexaenoyl glycerol (TriDHA) was compared with that of ethyl ester and free acid in rats cannulated with thoracic duct. Trioleoylglycerol (TO) served as a control. EPA and DHA, compared with oleic acid, were slowly transported in lymph irrespective of fat types administered. Total 24-h recovery of DHA in all fat types and ethyl EPA was significantly lower compared to that of oleic acid. Lymphatic recovery of EPA and DHA in rats given TriEPA and TriDHA was significantly higher at the first 3 h after the administration compared to those given as free acid or ethyl ester. The recovery in rats given free acid at a later stage (9-24 h) was higher than that of the other fat types. As a result, the 24-h recovery was comparable between triacylglycerol (TAG) and free acid, while it was significantly lower in ethyl ester. Although TriEPA and TriDHA were slowly hydrolyzed by pancreatic lipase in vitro compared with TO and TAGs rich in EPA or DHA at the second position, the hydrolysis rate at 60 min incubation was comparable among the TAGs examined. The hydrolysis rate of ethyl esters was extremely low even in 6 h incubation with lipase. These observations show that presence of EPA and DHA at the 1- and 3-positions of TAGs does not result in their lower recovery in lymph. Processes after lipolysis may be responsible for their low recovery in lymph. In a separate study, slower lymphatic recovery of DHA given as free acid than TriDHA was improved by the simultaneous administration of TO, but not by free oleic acid. The observations suggest that the slow recovery of free acid is caused by delayed TAG synthesis in mucosal cells and/or low micellar solubility of fatty acids in the intestinal lumen due to a limited supply of 2-monoacylglycerol (MAG). A large portion of EPA and DHA were recovered in lymph chylomicrons and very low density lipoproteins (VLDL, > 95%) and incorporated into TAG (84-92%) fraction in all fat types examined. Lymphatic recovery rate of simultaneously administered cholesterol was influenced by the fat types given.