Untargeted Liquid Chromatography-Mass Spectrometry-Based Metabolomics Analysis of Wheat Grain

Hayley Abbiss; Joel  P. A. Gummer; Michael Francki; Robert  D. Trengove

doi:10.3791/60851

Method Article

Untargeted Liquid Chromatography-Mass Spectrometry-Based Metabolomics Analysis of Wheat Grain

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DOI:

10.3791/60851

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March 13th, 2020

Hayley Abbiss¹^,²^,³ , Joel P. A. Gummer¹^,⁴ , Michael Francki⁵^,⁶ , Robert D. Trengove¹

¹Research and Innovation, Murdoch University, ²Centre for Digital Agriculture, Curtin University, ³Centre for Integrative Metabolomics and Computational Biology, School of Science, Edith Cowan University, ⁴Centre for Computational and System Medicine, Health Futures Institute, Murdoch University, ⁵Department of Primary Industries and Regional Development, ⁶State Agricultural Biotechnology Centre, Murdoch University

Summary

A method for the untargeted analysis of wheat grain metabolites and lipids is presented. The protocol includes an acetonitrile metabolite extraction method and reversed phase liquid chromatography-mass spectrometry methodology, with acquisition in positive and negative electrospray ionization modes.

Abstract

Understanding the interactions between genes, the environment and management in agricultural practice could allow more accurate prediction and management of product yield and quality. Metabolomics data provides a read-out of these interactions at a given moment in time and is informative of an organism's biochemical status. Further, individual metabolites or panels of metabolites can be used as precise biomarkers for yield and quality prediction and management. The plant metabolome is predicted to contain thousands of small molecules with varied physicochemical properties that provide an opportunity for a biochemical insight into physiological traits and biomarker discovery. To exploit this, a key aim for metabolomics researchers is to capture as much of the physicochemical diversity as possible within a single analysis. Here we present a liquid chromatography-mass spectrometry-based untargeted metabolomics method for the analysis of field-grown wheat grain. The method uses the liquid chromatograph quaternary solvent manager to introduce a third mobile phase and combines a traditional reversed-phase gradient with a lipid-amenable gradient. Grain preparation, metabolite extraction, instrumental analysis and data processing workflows are described in detail. Good mass accuracy and signal reproducibility were observed, and the method yielded approximately 500 biologically relevant features per ionization mode. Further, significantly different metabolite and lipid feature signals between wheat varieties were determined.

Introduction

Understanding the interactions between genes, environment and management practices in agriculture could allow more accurate prediction and management of product yield and quality. Plant metabolites are influenced by factors such as the genome, environment (climate, rainfall etc.), and in an agriculture setting, the way crops are managed (i.e., application of fertilizer, fungicide etc.). Unlike the genome, the metabolome is influenced by all of these factors and hence metabolomics data provides a biochemical fingerprint of these interactions at a particular time. There are usually one of two goals for a metabolomics-based study: firstly, to achieve a deeper understandi....