The origin of the linoleic [18:2(n-6)], arachidonic [20:4(n-6)], and docosahexaenoic acid [22:6(n-3)] of the mucosal phospholipids in the gastrointestinal (GI) tract is not known. This study examines whether stomach, small intestine, and colon take up and desaturate-elongate unesterified polyunsaturated fatty acids (PUFA) from blood. Albumin-bound unesterified alpha-[14C]linolenate [18:3(n-3)] and [3H]eicosapentaenoate [20:5(n-3)] were injected intravenously. After 10 min, 1 h, and 18 h, radioactivity of tissue lipids and the degree of interconversion of the 3H-labeled and 14C-labeled fatty acids were determined. After 10 min, the lipids of the gastrointestinal tract contained 3.6% of the 14C (0.5% in stomach, 2.7% in small intestine, and 0.4% in colon) and 4.9% of the 3H (0.6% in stomach, 3.6% in small intestine, and 0.7% in colon). Both fatty acids were acylated mainly into phospholipids, both in liver and gastrointestinal tract. Although the proportions of radioactivity found in desaturation-elongation products increased with time, 15% of the 14C in the liver, 6% in the stomach, 8% in the small intestine, and 11% in the colon were in 20:5 already after 10 min. A rapid interconversion of 18:3 thus occurred both in the liver and in the gastrointestinal tract. In all tissues examined, interconversion of [3H]20:5 to [3H]22:5 also occurred. Interconversion of unesterified PUFA taken up from blood may be an important source of eicosanoid precursors in the gastrointestinal tract.