Biosynthesis of Pyrrolizidine Alkaloids and Analogues

Denholm, Alastair Alexander (1990) Biosynthesis of Pyrrolizidine Alkaloids and Analogues. PhD thesis, University of Glasgow.

Abstract

The work presented in this thesis is divided into four sections : a) Synthesis of analogues of a known intermediate in pyrrolizidine alkaloid biosynthesis and the study of their incorporation into various pyrrolizidine alkaloid-producing plant species; b) Studies on the kinetics of enzymic oxidation of these analogues by pea seedling diamine oxidase; c) Further biosynthetic studies on the pyrrolizidine necine bases; d) Biosynthesis of the 3-hydroxy-3-methylglutarate portion of dicrotaline. a) Analogue Studies Radiolabelled analogues of N-(4-aminobutyl)-1,2-didehydropyrrolidinium (A) were synthesised and fed to various plant species which produce pyrrolizidine alkaloids. 14C-Labelled analogues (B) and (C) were synthesised by the extension of an established route to the natural precursor (A). These compounds were of low specific activity (2-30 Ci mmol-1). A new synthetic route was developed which furnished a series of analogues (D) and (E) labelled with tritium. These compounds were of higher specific activity (150-250 muCi mmol-1), and higher incorporations were observed in the feeding experiments (Senecio pleistocephalus, S. isatideus, and Cynoglossum australe spp.). All feeding experiments used radiolabelled putrescine as an internal standard. Iminium ion precursors (E) were utilised more efficiently than the corresponding saturated salts (D). The saturated salts (D) appeared to inhibit the incorporation of putrescine into pyrrolizidine alkaloids. The higher homologues were generally more poorly utilised than putrescine. b) Enzyme Kinetics The enzymic oxidation of non-labelled analogues of (B) and (C) by pea seedling diamine oxidase was studied using a spectrophotometric assay which allowed determination of and Vmax data for each substrate. Saturated analogues could be classified as putrescine or cadaverine analogues on the basis of binding affinities, as indicated by values. Iminium ions could similarly be divided into sub-groups. The strong binding affinities of the iminium ions are of particular note. This, along with the relatively low catalytic rate constants, makes the iminium ions potential candidates as inhibitors of the enzymic oxidation of putrescine. c) Biosynthesis of Necine Bases Platynecine and rosmarinecine were identified as intermediates on the biosynthetic pathway to rosmarinine by an intermediate trapping experiment. N - Acetylputrescine was incorporated more efficiently than putrescine into the alkaloids of Cynoglossum australe, while putrescine was the better precursor of rosmarinine in Senecio pleistocephalus. d) Biosynthesis of the 3-Hydroxy-3-methylglutarate (HMG) portion of Dicrotaline Acetate, mevalonate, and 3-hydroxy-3-methyl-glutarate were not specific precursors for the necic acid portion of dicrotaline (F) in Crotalaria lachnosema. The C-5 methyl group of isoleucine was specifically incorporated into the methyl group of HMG. This was shown by the specific incorporation of [4,5-H]isoleucine into the necic acid portion of dicrotaline and degradation to establish the site of labelling as the methyl group of dicrotaline.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Organic chemistry
• Date of Award: 1990
• Depositing User: Enlighten Team
• Unique ID: glathesis:1990-78248
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 28 Feb 2020 12:09
• Last Modified: 28 Feb 2020 12:09
• URI: https://theses.gla.ac.uk/id/eprint/78248