Isolation and identification of the toxic compounds of Tapura fischeri Engl

Abstract

Tapura fischeri is a member of the family Dichapetalaceae and the only other member of this family naturally occurring in South Africa is Dichapetalum cymosum. Poisoning by D. cymosum results in the deaths of many domestic livestock each year due to the presence of fluoroacetate. The aim of the study was to determine if monofluoroacetate or another fluorinated compound is present in T. fischeri, and the possible role endophytes might play in the production of these compounds. Through NMR and GCMS studies it was established that trifluoroacetate is present in T. fischeri. Bacterial endophytes were isolated from plant material and shown to produce a fluorinated compound other than mono and trifluoroacetate. Since trifluoroacetic acid is extremely volatile, and evaporate from the plant extract over time, column chromatography, together with NMR was employed to isolate and identify other compounds responsible for antibacterial activity against the bacterium Enterococcus faecalis previously observed on TLC plates. Two compounds were isolated, and identified with NMR as a fatty acid and a fatty acid attached to glycerol. The names of the compounds could not be established with GCMS due to insufficient derivatization of the compounds. The antibacterial activity of the compounds were also analyzed using 96 well microtitre plates in liquid media, where it was determined that the compounds do not have antibacterial activity against E. faecalis. This indicated that previous results on TLC plates were false positives due to the hydrophobic nature of the fatty acid compounds. Transmission electron microscopy was done on leaf material to determine the presence of bacterial endophytes in the intracellular spaces of plant material, but none was detected. These results suggest a possitive correlation between the plant, its endophytes and the production of the fluorinated compound.