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Emmrich, Peter M.F., Rejzek, Martin, Hill, Lionel, Brett, Paul, Edwards, Anne, Sarkar, Abhimanyu, Field, Rob A. 
ORCID: https://orcid.org/0000-0001-8574-0275, Martin, Cathie and Wang, Trevor L.
(2019)
Linking a rapid throughput plate-assay with high-sensitivity stable-isotope label LCMS quantification permits the identification and characterisation of low β-L-ODAP grass pea lines.
BMC Plant Biology, 19 (1).
ISSN 1471-2229
Abstract
Background: Grass pea (Lathyrus sativus) is an underutilised crop with high tolerance to drought and flooding stress and potential for maintaining food and nutritional security in the face of climate change. The presence of the neurotoxin β-L-oxalyl-2,3-diaminopropionic acid (β-L-ODAP) in tissues of the plant has limited its adoption as a staple crop. To assist in the detection of material with very low neurotoxin toxin levels, we have developed two novel methods to assay ODAP. The first, a version of a widely used spectrophotometric assay, modified for increased throughput, permits rapid screening of large populations of germplasm for low toxin lines and the second is a novel, mass spectrometric procedure to detect very small quantities of ODAP for research purposes and characterisation of new varieties. Results: A plate assay, based on an established spectrophotometric method enabling high-throughput ODAP measurements, is described. In addition, we describe a novel liquid chromatography mass spectrometry (LCMS)-based method for β-L-ODAP-quantification. This method utilises an internal standard (di-13C-labelled β-L-ODAP) allowing accurate quantification of β-L-ODAP in grass pea tissue samples. The synthesis of this standard is also described. The two methods are compared; the spectrophotometric assay lacked sensitivity and detected ODAP-like absorbance in chickpea and pea whereas the LCMS method did not detect any β-L-ODAP in these species. The LCMS method was also used to quantify β-L-ODAP accurately in different tissues of grass pea. Conclusions: The plate-based spectrophotometric assay allows quantification of total ODAP in large numbers of samples, but its low sensitivity and inability to differentiate α- and β-L-ODAP limit its usefulness for accurate quantification in low-ODAP samples. Coupled to the use of a stable isotope internal standard with LCMS that allows accurate quantification of β-L-ODAP in grass pea samples with high sensitivity, these methods permit the identification and characterisation of grass pea lines with a very low ODAP content. The LCMS method is offered as a new 'gold standard' for β-L-ODAP quantification, especially for the validation of existing and novel low- and/or zero-β-L-ODAP genotypes.
| Item Type: | Article |
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| Additional Information: | Funding Information: This research was supported by the ‘Detoxed grass pea: sustainable sustenance for stressful environments’ grant provided under the BBSRC/DBT Joint Call in Crop Genomics. PMFE’s studentship was funded by the John Innes Foundation’s Student Rotation programme. Neither funding body was involved in the design of this study, nor the collection or analysis or interpretation of data, nor in writing the manuscript. Publisher Copyright: © 2019 The Author(s). |
| Uncontrolled Keywords: | c-internal standard,boaa,chickpea,grass pea,lathyrus sativus,lcms,pea,spectrophotometric assay,stable-isotope labelled,β-l-odap,plant science,sdg 13 - climate action ,/dk/atira/pure/subjectarea/asjc/1100/1110 |
| Faculty \ School: | University of East Anglia Research Groups/Centres > Theme - ClimateUEA Faculty of Social Sciences > School of Global Development (formerly School of International Development) Faculty of Science > School of Chemistry, Pharmacy and Pharmacology Faculty of Science > School of Biological Sciences |
| UEA Research Groups: | Faculty of Medicine and Health Sciences > Research Centres > Norwich Institute for Healthy Aging |
| Related URLs: | |
| Depositing User: | LivePure Connector |
| Date Deposited: | 02 Sep 2024 15:30 |
| Last Modified: | 25 Sep 2024 18:05 |
| URI: | https://ueaeprints.uea.ac.uk/id/eprint/96439 |
| DOI: | 10.1186/s12870-019-2091-5 |
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