Qualitative Analysis of Polyphenols in Glycerol Plant Extracts Using Untargeted Metabolomics.

Glycerol is a reliable solvent for extracting polyphenols from food and waste products. There has been an increase in the application of glycerol over benchmark alcoholic solvents such as ethanol and methanol for natural product generation because of its non-toxic nature and high extraction efficiency. However, plant extracts containing a high glycerol concentration are unsuitable for mass spectrometry-based investigation utilising electrospray ionization, inhibiting the ability to analyse compounds of interest. In this investigation, a solid phase extraction protocol is outlined for removing glycerol from plant extracts containing a high concentration of glycerol and their subsequent analysis of polyphenols using ultra-performance liquid chromatography coupled with quadrupole time of flight tandem mass spectrometry. Using this method, glycerol-based extracts of Queen Garnet Plum (Prunus salicina) were investigated and compared to ethanolic extracts. Anthocyanins and flavonoids in high abundance were found in both glycerol and ethanol extracts. The polyphenol metabolome of Queen Garnet Plum was 53% polyphenol glycoside derivatives and 47% polyphenols in their aglycone forms. Furthermore, 56% of the flavonoid derivates were found to be flavonoid glycosides, and 44% were flavonoid aglycones. In addition, two flavonoid glycosides not previously found in Queen Garnet Plum were putatively identified: Quercetin-3-O-xyloside and Quercetin-3-O-rhamnoside.

Starch granule size: Does it matter?

Nature has developed starch granules varying in size from less than 1 µm to more than 100 µm. The granule size is an important factor affecting the functional properties and the applicability of starch for food and non-food applications. Within the same botanical species, the range of starch granule size can be up to sevenfold. This review critically evaluated the biological and environmental factors affecting the size of starch granules, the methods for the separation of starch granules and the measurement of size distribution. Further, the structure at different length scales and properties of starch-based on the granule size is elucidated by specifying the typical applications of granules with varying sizes. An amylopectin cluster model showing the arrangement of amylopectin from inside toward the granule surface is proposed with the hypothesis that the steric hindrance for the growth of lamellar structure may limit the size of starch granules.

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.

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.

Diversity of global rice markets and the science required for consumer-targeted rice breeding.

With the ever-increasing global demand for high quality rice in both local production regions and with Western consumers, we have a strong desire to understand better the importance of the different traits that make up the quality of the rice grain and obtain a full picture of rice quality demographics. Rice is by no means a 'one size fits all' crop. Regional preferences are not only striking, they drive the market and hence are of major economic importance in any rice breeding / improvement strategy. In this analysis, we have engaged local experts across the world to perform a full assessment of all the major rice quality trait characteristics and importantly, to determine how these are combined in the most preferred varieties for each of their regions. Physical as well as biochemical characteristics have been monitored and this has resulted in the identification of no less than 18 quality trait combinations. This complexity immediately reveals the extent of the specificity of consumer preference. Nevertheless, further assessment of these combinations at the variety level reveals that several groups still comprise varieties which consumers can readily identify as being different. This emphasises the shortcomings in the current tools we have available to assess rice quality and raises the issue of how we might correct for this in the future. Only with additional tools and research will we be able to define directed strategies for rice breeding which are able to combine important agronomic features with the demands of local consumers for specific quality attributes and hence, design new, improved crop varieties which will be awarded success in the global market.

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.