Application of metabolomics for providing a new generation of selection tools for crop improvement

Quantitative trait loci mapping using metabolite data (metabolite quantitative trait loci mapping) has been successfully done in several crops, such as Arabidopsis thaliana, rice and tomato

Untargeted and targeted metabolomic profiling of Australian indigenous fruits

Selected Australian native fruits such as Davidson's plum, finger lime and native pepperberry have been reported to demonstrate potent antioxidant activity. However, comprehensive metabolite profiling of these fruits is limited, therefore the compounds responsible are unknown, and further, the compounds of nutritional value in these native fruits are yet to be described. In this study, untargeted and targeted metabolomics were conducted using the three fruits, together with assays to determine their antioxidant activities. The results demonstrate that targeted free and hydrolysed protein amino acids exhibited high amounts of essential amino acids. Similarly, important minerals like potassium were detected in the fruit samples. In antioxidant activity, Davidson's plum reported the highest activity in ferric reducing power (FRAP), finger lime in antioxidant capacity (ABTS), and native pepperberry in free radical scavenging (DPPH) and phosphomolybdenum assay. The compounds responsible for the antioxidant activity were tentatively identified using untargeted GC×GC-TOFMS and UHPLC-QqQ-TOF-MS/MS metabolomics. A clear discrimination into three clusters of fruits was observed using principal component analysis (PCA) and partial least squares (PLS) analysis. The correlation study identified a number of compounds that provide the antioxidant activities. GC×GC-TOFMS detected potent aroma compounds of limonene, furfural, and 1-R-α-pinene. Based on the untargeted and targeted metabolomics, and antioxidant assays, the nutritional potential of these Australian bush fruits is considerable and supports these indigenous fruits in the nutraceutical industry as well as functional ingredients for the food industry, with such outcomes benefiting Indigenous Australian communities

Using chain-length distributions to diagnose genetic diversity in starch biosynthesis

Amylopectin synthesis is controlled by the coordinated action of several types of enzymes, including starch synthases, branching and debranching enzymes. The contributions of some of these enzymes to the building of starch molecules have been previously established. Changes to the activity of an enzyme can affect amylopectin structure, which is associated with diversity in functional properties. One such property, gelatinisation temperature, has been studied at the genetic, biochemical and phenotypic level. A technique, the 'log(number distribution) approach', offers a means of collecting normalisation-free representations of the chain-length distributions of starch, with the potential to reveal new information about kinetics and processes of starch synthesis; this method of plotting the data can potentially reveal much more than that usually employed, viz., the simple number chain-length distribution. In this study, samples from genotypes with defined mutations in starch biosynthetic genes that specifically and differently alter the chain-length distribution of single-cluster chains, with a resultant effect on gelatinisation temperature, are used to show that log(number distribution) plots have sufficient discriminative capacity to diagnose the gene affected by mutations, provide new information, and determine those features of the plot which associate with genotype and phenotype. (C) 2010 Elsevier Ltd. All rights reserved

Understanding the Jasmine phenotype of rice through metabolite profiling and sensory evaluation

Introduction: Aromatic rices are culturally and economically important for many countries in Asia. Investigation of the volatile compounds emitted by rice during cooking is the key to understanding the flavour of elite aromatic rice varieties. Objectives: The objectives of this study were to compare Jasmine-type aromatic rices from the Greater Mekong Subregion and Australia in terms of their metabolomics and sensory profiles and to draw out associations between the volatile organic compounds and human sensory perception of rice aroma. Methods: A set of aromatic rice varieties from South East Asia and Australia, along with non-aromatic controls, was grown in tropical and temperate areas of Australia. Untargeted metabolite profiling of volatile compounds, from the heated rice flour, by static headspace extraction and separation by two dimensional gas chromatography time-of-flight mass spectrometry was performed. Volatile compounds were also assayed in the standard references used in the sensory evaluation and compared to the compounds detected in the headspace of rice. Results: While 2-acetyl-1-pyrroline (2-AP) was a discriminating compound, we identified several of its structural homologues, and a number of other metabolites that were consistently detected in fragrant Jasmine rice. 2-AP producing rice varieties have different sensory properties and these variations were defined by the discriminating compounds identified in each rice type. Conclusions: The results of this study are valuable in understanding the aspects of aromatic rice that are important to consumers, and in the identification of compounds that breeding programs can use to select for pleasant aromas, enabling breeding programs to target markets with greater accuracy

Metabolomics and genomics combine to unravel the pathway for the presence of fragrance in rice

Since it was first characterised in 1983, 2-acetyl-1-pyrroline (2AP) has been considered to be the most important aroma compound in rice. In this study, we show four other amine heterocycles: 6-methyl, 5-oxo-2,3,4,5-tetrahydropyridine (6M5OTP), 2-acetylpyrrole, pyrrole and 1-pyrroline, that correlate strongly with the production of 2AP, and are present in consistent proportions in a set of elite aromatic rice varieties from South East Asia and Australia as well as in a collection of recombinant inbred lines (RILs) derived from indica Jasmine-type varieties, Australian long grain varieties (temperate japonica) and Basmati-type rice (Grp V). These compounds were detected through untargeted metabolite profiling by two-dimensional gas chromatography-time-of-flight mass spectrometry (GC × GC-TOF-MS), and their identity were confirmed by comparison with authentic standards analysed using gas chromatography mass spectrometry (GC-MS) and High Resolution GC × GC-TOF-MS (GC × GC HRT-4D). Genome-wide association analysis indicates that all compounds co-localised with a single quantitative trait locus (QTL) that harbours the FGR gene responsible for the production of GABA. Together, these data provide new insights into the production of 2AP, and evidence for understanding the pathway leading to the accumulation of aroma in fragrant rice

Biomolecular analyses of starch and starch granule proteins in the high-amylose rice mutant goami 2

Elevated proportions of amylose in cereals are commonly associated with either the loss of starch branching or starch synthase activity. Goami 2 is a high-amylose mutant of the temperate japonica rice variety Ilpumbyeo. Genotyping revealed that Goami 2 and Ilpumbyeo carry the same alleles for starch synthase IIa and granule-bound starch synthase I genes. Analyses of granule-bound proteins revealed that SSI and SSIIa accumulate inside the mature starch granules of Goami 2, which is similar to the amylose extender mutant IR36ae. However, unlike the amylose extender mutants, SBEIIb was still detectable inside the starch granules of Goami 2. Detection of SBEIIb after protein fractionation revealed that most of the SBEIIb in Goami 2 accumulates inside the starch granules, whereas most of it accumulates at the granule surface in Ilpumbyeo. Exhaustive mass spectrometric characterisations of granule-bound proteins failed to detect any peptide sequence mutation or major post-translational modifications in Goami 2. Moreover, the signal peptide was found to be cleaved normally from the precursor protein, and there is no apparent N-linked glycosylation. Finally, no difference was found in the SBEIIb structural gene sequence of Goami 2 compared with Ilpumbyeo. In contrast, a G-to-A mutation was detected in the SBEIIb gene of IR36ae located at the splice site between exon and intron 11, which could potentially introduce a premature stop codon and produce a truncated form of SBEIIb. It is suggested that the mutation responsible for producing high amylose in Goami 2 is not due to a defect in SBEIIb gene as was observed in IR36ae, even though it produces a phenotype analogous to the amylose extender mutation. Understanding the molecular genetic basis of this mutation will be important in identifying novel targets for increasing amylose and resistant starch contents in rice and other cereals

Relationship between 2-acetyl-1-pyrroline and aroma in Uganda rice populations with Oryza (barthi, glaberrima and sativa) backgrounds

The sweet popcorn aroma conferred by 2-acetyl-1-pyrroline (2AP) is a highly economic trait of rice grain attracting premium price worldwide. This research study was conducted to determine the levels of 2AP in Ugandan rice lines with the aim of establishing a better understanding on the level and classes of 2AP and aroma phenotype. Concentration of 2AP was assayed using two-dimensional gas chromatography-time-of-flight mass spectrometry (GC × GC-TOF-MS) in tandem with sensory evaluation. Substantial variations in aroma intensity within and between the Uganda rice families were recorded. However, the levels of aroma variation were strongly influenced by the type of rice, and the breeding population it was derived from. Hence, three aroma based categories, namely, nonaromatic, moderately aromatic and highly aromatic were identified. GC with complementary sensory evaluation suggested a highly complex nature of rice aroma, as several rice lines were re-classified on the basis of this study. The 2AP contents and aroma intensity for genotypes with O. glaberrima were low compared to O. sativa and O. barthi. Genotypes of Supa 5, Supa 1052, Yasmin aromatic and MET 3 contained high 2AP levels whereas MET 16, MET 6, AGRA 78, AGRA 55, AGRA 41 and Sande TXD 306 exhibited moderate 2AP contents. Therefore, in developing an optimal breeding strategy aimed at improving the aroma in rice, quantitative information about 2AP and complementary sensory evaluation are a prerequisite

Biomolecular Analyses of Starch and Starch Granule Proteins in the High-Amylose Rice Mutant Goami 2

Elevated proportions of amylose in cereals are commonly associated with either the loss of starch branching or starch synthase activity. Goami 2 is a high-amylose mutant of the <i>temperate japonica</i> rice variety Ilpumbyeo. Genotyping revealed that Goami 2 and Ilpumbyeo carry the same alleles for <i>starch synthase IIa</i> and <i>granule-bound starch synthase I</i> genes. Analyses of granule-bound proteins revealed that SSI and SSIIa accumulate inside the mature starch granules of Goami 2, which is similar to the <i>amylose extender</i> mutant IR36ae. However, unlike the <i>amylose extender</i> mutants, SBEIIb was still detectable inside the starch granules of Goami 2. Detection of SBEIIb after protein fractionation revealed that most of the SBEIIb in Goami 2 accumulates inside the starch granules, whereas most of it accumulates at the granule surface in Ilpumbyeo. Exhaustive mass spectrometric characterisations of granule-bound proteins failed to detect any peptide sequence mutation or major post-translational modifications in Goami 2. Moreover, the signal peptide was found to be cleaved normally from the precursor protein, and there is no apparent N-linked glycosylation. Finally, no difference was found in the <i>SBEIIb</i> structural gene sequence of Goami 2 compared with Ilpumbyeo. In contrast, a G-to-A mutation was detected in the <i>SBEIIb</i> gene of IR36ae located at the splice site between exon and intron 11, which could potentially introduce a premature stop codon and produce a truncated form of SBEIIb. It is suggested that the mutation responsible for producing high amylose in Goami 2 is not due to a defect in <i>SBEIIb</i> gene as was observed in IR36ae, even though it produces a phenotype analogous to the <i>amylose extender</i> mutation. Understanding the molecular genetic basis of this mutation will be important in identifying novel targets for increasing amylose and resistant starch contents in rice and other cereals

A: Grains ordered from shortest to longest, showing different widths (mm) in each class except the extra-long class.

<p>B: Histogram showing the proportion of bold (black), medium (light grey) and slender (dark grey) grain shapes within each length class for all the popular rices discussed in the present paper.</p

Consumer preferences for aromatic rice for the countries, states, and provinces of Asia.

<p>In some regions, all popular rices are aromatic, in others, one or two is aromatic, and in some countries, aromatic rice is not popular. Additional information for other regions can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0085106#pone-0085106-t002" target="_blank">Table 2</a>. Data were obtained from INQR representatives from each region.</p