ARS | Publication request: Analysis of the toxic amino acid indospicine by liquid chromatography-tandem mass spectrometry

Submitted to: International Journal of Poisonous Plant Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: April 29, 2011
Publication Date: November 15, 2011
Repository URL: http://www.ars.usda.gov/is/np/PoisonousPlants/PoisonousPlantResearchJournalIntro.htm
Citation: Gardner, D.R., Riet-Correa, F. 2011. Analysis of the toxic amino acid indospicine by liquid chromatography-tandem mass spectrometry. International Journal of Poisonous Plant Research. 1(1):20-7.

Interpretive Summary: Some Indigofera species contain a toxic non-protein amino acid known as indospicine and are toxic to livestock. There are also cases of secondary poisonings in dogs that consume the meat of animals that previously grazed indospicine-containing forage. In order to analyze for this toxic amino acid in plants an analytical method based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed. The toxin, indospicine, was extracted from the plant with an acid alcohol solution and then derivatized with a phenylisothiocyanate solution and then analyzed by LC-MS/MS. The method was applied to the analysis of several samples of Indigofera that were collected in Brazil and associated the poisoning of horses and causing a nervous disease. Secondly, plants of a different species of Indigofera from Africa and associated with floppy trunk syndrome in elephants were also analyzed. Indospicine was detected in all samples collected in Brazil. No indospicine was detected in samples collected from Africa.

Technical Abstract: Some Indigofera species contain a toxic non-protein amino acid known as indospicine. Indospicine-containing plants are toxic to livestock and cases of secondary poisoning have been documented in dogs that consume the meat of animals that previously grazed indospicine-containing forage. For the analysis of indospicine in the plant material a method based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) of the phenylisothiocyanate derivative of indospicine was developed. Indospicine was extracted from the plant with ethanol/0.01N HCl (70:30). The sample was then derivatized with a phenylisothiocyanate solution and then analyzed by LC-MS/MS. The calibration was linear over the range of 1.0 – 25 µg/mL, recovery was 86% and the intra-day precision was 4.7% (RSD). Indospicine was confirmed in samples of I. lespedezioides collected in Brazil with a concentration range of 60 – 1400 ug/g (dry weight basis). No indospicine was detected in samples of I. praticola and I. tinctoria from Zimbabwe, Africa.