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
| Autor: | Paul Brett, Lionel Hill, Anne Edwards, Robert A. Field, Abhimanyu Sarkar, Martin Rejzek, Trevor L. Wang, Cathie Martin, Peter M. F. Emmrich |
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| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
Time Factors
Spectrophotometric assay Neurotoxins BOAA Plant Science Biology Grass pea Mass spectrometry Sensitivity and Specificity 01 natural sciences Mass Spectrometry Absorbance Lathyrus sativus 0404 agricultural biotechnology LCMS Chickpea Liquid chromatography–mass spectrometry lcsh:Botany Lathyrus Plate assay 2. Zero hunger Stable-isotope labelled Chromatography 13C-internal standard Stable isotope ratio Methodology Article 010401 analytical chemistry Pea Amino Acids Diamino Reproducibility of Results food and beverages 04 agricultural and veterinary sciences Reference Standards biology.organism_classification 040401 food science Mass spectrometric 0104 chemical sciences lcsh:QK1-989 Spectrophotometry 13. Climate action Isotope Labeling Costs and Cost Analysis β-L-ODAP Chromatography Liquid |
| Zdroj: | BMC Plant Biology, Vol 19, Iss 1, Pp 1-14 (2019) BMC Plant Biology |
| ISSN: | 1471-2229 |
| DOI: | 10.1186/s12870-019-2091-5 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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