Biosynthesis of anabasine in transformed root cultures of Nicotiana species

Abstract

The biosynthesis of 3-(2-piperidyl)pyridine (anabasine)(6) has been studied in transformed root cultures of Nicotiana rustica and N. tabacum using cadaverine precursors selectively labelled with deuterium. The labelling pattern in anabasine (6) derived biosynthetically from [2,2,4,4-²H₄]-cadaverine in N. tabacum could not be established by ²H NMR spectroscopy because of the closeness of the chemical shifts of the labelled ²H atoms. However, the complete labelling pattern for the tetradeuteriated anabasine (10) was established by ¹H NMR spectroscopy, because of the very high incorporation of ²H₄ species (dilution in anabasine ca. 1). Two AB systems were observed for the protons at C-4′ and C-6′, and a singlet for the C-2′ proton in anabasine (10). No deuterium was incorporated into the nicotinic acid portion of anabasine. The labelling patterns in anabasine derived biosynthetically from (S)- and (R)-[1-²H]cadaverine in N. rustica were determined by ²H NMR spectroscopy. These labelling patterns, due to mixtures of species (11) and (12) and (13)–(16), respectively, show that the pro-R hydrogen of cadaverine is retained and the pro-S hydrogen is lost at the position which becomes C-2′ of anabasine. In addition, the labelling patterns are consistent with formation of equal amounts of (R)- and (S)-anabasine.