N-Hydroxyacylnornicotine, newly discovered from fresh plant tissue, was found entirely in the trichome exudate produced at the epidermis of the aerial part of Nicotiana stocktonii. Nicotine and nornicotine, but not N-hydroxyacylnornicotine, were present inside of the trichomes as well as other internal parts of the plant. Only nicotine was found in bleeding sap squeezed from cut roots or stems. Feeding of leaves with 2'-'4C-labeled nicotine primarily yielded labeled nicotine, nornicotine, and Nhydroxyacylnoraicotine. When similarly labeled nornicotine was fed to leaves as a precursor, a labeled N-hydroxyacylnornicotine was obtained, with a higher specific activity than with the 12'-4Cinicotine feeding.Based on these results, a synthesis route is suggested where nicotine is converted in the leaf to nornicotine, followed by trichome conversion of nornicotine to N-hydroxyacylnornicotine, and rapid secretion of this product.