Insights into the relative contribution of four precursors to 3-sulfanylhexan-1-ol and 3-sulfanylhexylacetate biogenesis during fermentation.

The desirable wine aroma compounds 3-sulfanylhexan-1-ol (3SH) and 3-sulfanylhexyl acetate (3SHA) are released during fermentation from non-volatile precursors present in the grapes. This work explores the relative contribution of four precursors (E-2-hexenal, 3-S-glutathionylhexan-1-ol, 3-S-glutathionylhexanal, and 3-S-cysteinylhexan-1-ol) to 3SH and 3SHA. Through the use of isotopically labelled analogues of these precursors in defined fermentation media, new insights into the role of each precursor have been identified. E-2-Hexenal was shown to contribute negligible amounts of thiols, while 3-S-glutathionylhexan-1-ol was the main precursor of both 3SH and 3SHA. The glutathionylated precursors were both converted to 3SHA more efficiently than 3-S-cysteinylhexan-1-ol. Interestingly, 3-S-glutathionylhexanal generated 3SHA without detectable concentrations of 3SH, suggesting possible differences in the way this precursor is metabolised compared to 3-S-glutathionylhexan-1-ol and 3-S-cysteinylhexan-1-ol. We also provide the first evidence for chemical conversion of 3-S-glutathionylhexan-1-ol to 3-S-(γ-glutamylcysteinyl)-hexan-1-ol in an oenological system.

Integration and holistic analysis of multiple multidimensional soil data sets.

Complex matrices such as soil have a range of measurable characteristics, and thus data to describe them can be considered multidimensional. These characteristics can be strongly influenced by factors that introduce confounding effects that hinder analyses. Traditional statistical approaches lack the flexibility and granularity required to adequately evaluate such matrices, particularly those with large dataset of varying data types (i.e. quantitative non-compositional, quantitative compositional). We present a statistical workflow designed to effectively analyse complex, multidimensional systems, even in the presence of confounding variables. The developed methodology involves exploratory analysis to identify the presence of confounding variables, followed by data decomposition (including strategies for both compositional and non-compositional quantitative data) to minimise the influence of these confounding factors such as sampling site/location. These data processing methods then allow for common patterns to be highlighted in the data, including the identification of biomarkers and determination of non-trivial associations between variables. We demonstrate the utility of this statistical workflow by jointly analysing the chemical composition and fungal biodiversity of New Zealand vineyard soils that have been managed with either organic low-input or conventional input approaches. By applying this pipeline, we were able to identify biomarkers that distinguish viticultural soil from both approaches and also unearth links and associations between the chemical and metagenomic profiles. While soil is an example of a system that can require this type of statistical methodology, there are a range of biological and ecological systems that are challenging to analyse due to the complex interplay of global and local effects. Utilising our developed pipeline will greatly enhance the way that these systems can be studied and the quality and impact of insight gained from their analysis.

Exploring the Equilibrium between Glut3SHal and Glut3SH-SO3: A Method for the Quantification of These Compounds in Wine.

3S-Gluthathionylhexanal (glut3SHal) is an early precursor to the important wine aroma compound 3-sulfanylhexan-1-ol (3SH), imparting tropical passion fruit aromas, even at trace concentrations. In wine, glut3SHal occurs in equilibrium with its bisulfite adduct (glut3SH-SO3), challenging its quantification. To circumvent the issues encountered when attempting to describe the equilibrium between these compounds, a method for their quantification in wine samples was developed. Separation of glut3SHal and glut3SH-SO3 using solid-phase extraction followed by oxime derivatization and analysis via liquid chromatography-mass spectrometry allowed for measurement of both compounds in wine samples. Analysis of commercial Sauvignon Blanc wines using the developed method confirmed that glut3SH-SO3 is the major species in the wine matrix. The method developed in this work will enable further exploration of the relationship between glut3SHal and glut3SH-SO3 and their contribution to production of 3SH in wines. There is potential to extrapolate this work to explore other aldehyde-sulfonic acid equilibria in foods and beverages.

A green liquid chromatography-tandem mass spectrometry method for the simultaneous analysis of volatile thiols and their precursors in oenological samples.

Improvements to the quantification of three white wine impact odorants 3-sulfanylhexan-1-ol, 3-sulfanylhexyl acetate, and 4-sulfanyl-4-methylpentan-2-one, and the non-volatile precursors from which they are released during fermentation, is of great interest to the wine science community. Recent reports of a "Quick, Easy, Cheap, Effective, Rugged and Safe" (QuEChERS) based method for the concurrent analysis of these thiols and their precursors via liquid chromatography tandem mass spectrometry (LC-MS/MS) has enabled the development of far simpler methods, as well as aligning these analyses with principles of green analytical chemistry. This current work reports the development and validation of a QuEChERS based LC-MS/MS method utilising a safer derivatising agent, 4,4'-dithiodipyridine, while greatly minimising the reagents involved and waste produced. We demonstrate that this new method compares favourably to the previously reported method with repeatability of 0.2-1.3%RSD and 0.4-5.2%RSD for precursors and free thiols. Further, the commercially available internal standard, 1-hexanethiol, used in previous analytical methods was compared to stable isotope labelled analogues of the analytes, with results suggesting that it may not be a reliable internal standard.

Hydrogen sulfide production during early yeast fermentation correlates with volatile sulfur compound biogenesis but not thiol release.

Yeasts undergo intensive metabolic changes during the early stages of fermentation. Previous reports suggest the early production of hydrogen sulfide (H2S) is associated with the release of a range of volatile sulfur compounds (VSCs), as well as the production of varietal thiol compounds 3-sulfanylhexan-1-ol (3SH) and 3-sulfanylhexyl acetate (3SHA) from six-carbon precursors, including (E)-hex-2-enal. In this study, we investigated the early H2S potential, VSCs/thiol output, and precursor metabolism of 11 commonly used laboratory and commercial Saccharomyces cerevisiae strains in chemically defined synthetic grape medium (SGM) within 12 h after inoculation. Considerable variability in early H2S potential was observed among the strains surveyed. Chemical profiling suggested that early H2S production correlates with the production of dimethyl disulfide, 2-mercaptoethanol, and diethyl sulfide, but not with 3SH or 3SHA. All strains were capable of metabolizing (E)-hex-2-enal, while the F15 strain showed significantly higher residue at 12 h. Early production of 3SH, but not 3SHA, can be detected in the presence of exogenous (E)-hex-2-enal and H2S. Therefore, the natural variability of early yeast H2S production contributes to the early output of selected VSCs, but the threshold of which is likely not high enough to contribute substantially to free varietal thiols in SGM.

Mixed yeast communities contribute to regionally distinct wine attributes.

There is evidence that vineyard yeast communities are regionally differentiated, but the extent to which this contributes to wine regional distinctiveness is not yet clear. This study represents the first experimental test of the hypothesis that mixed yeast communities-comprising multiple, region-specific, isolates, and species-contribute to regional wine attributes. Yeast isolates were sourced from uninoculated Pinot Noir fermentations from 17 vineyards across Martinborough, Marlborough, and Central Otago in New Zealand. New methodologies for preparing representative, mixed species inoculum from these significantly differentiated regional yeast communities in a controlled, replicable manner were developed and used to inoculate Pinot Noir ferments. A total of 28 yeast-derived aroma compounds were measured in the resulting wines via headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry. Yeast community region of origin had a significant impact on wine aroma, explaining ∼10% of the observed variation, which is in line with previous reports of the effects of region-specific Saccharomyces cerevisiae isolates on Sauvignon Blanc ferments. This study shows that regionally distinct, mixed yeast communities can modulate wine aroma compounds in a regionally distinct manner and are in line with the hypothesis that there is a microbial component to regional distinctiveness, or terroir, for New Zealand Pinot Noir.

Monitoring hydrogen sulfide de-novo formation from polysulfides present in wine using ion chromatography and ultra high-pressure liquid chromatography combined with fraction collection and high-resolution mass spectrometry.

The study of polysulfides has been a recent topic of interest for wine research due to the possibility of these compounds to release hydrogen sulfide (H2S) during storage. However, studying these compounds has been challenging for several reasons. Polysulfides are low in concentration in natural samples, they are chemically unstable and pure standards of the single compounds (RSnR with n > 2) are not commercially available. In the present study, a method was developed in order to collect a single polysulfide and study its degradation and the consequent formation of H2S. For this approach, ultra-high pressure liquid chromatography was used with an integrated fraction collector and subsequently coupled to high resolution mass spectrometry. After fractionation, the degradation of the di-cysteinyl pentasulfide (CS5C) was induced by exposure to 30 °C and the H2S formation was measured in parallel using ion-exchange chromatography. This method showed the evolutions of different polysulfides and the H2S release originating from the target compound, an observation that to the best of our knowledge has never been made before. The method in the present study demonstrated promising applications for polysulfide studies and brought us a step closer to the understanding of the chemistry of polysulfides in wine.

Characterization of polysulfides in Saccharomyces cerevisiae cells and finished wine from a cysteine-supplemented model grape medium.

Polysulfide degradation in wine can result in hydrogen sulfide (H2S) release, imparting a rotten-egg smell that is detrimental to wine quality. Although the presence of wine polysulfides has been demonstrated, their biogenesis remains unclear. This study investigated the role of Saccharomyces cerevisiae in polysulfide formation during fermentation, with and without 5 mM cysteine supplementation as an H2S source. Using an established liquid chromatography-tandem mass spectrometry method, monobromobimane derivatives of hydropolysulfides, including CysSSSH, CysSSSSH and GSSSSH, and two oxidized polysulfides, GSSG and GSSSSG, were detected in yeast cells at the end of fermentation in a grape juice-like medium. Polysulfide production by four S. cerevisiae single deletion mutants (BY4743 Δcys3, Δcys4, Δmet17 and Δtum1) showed no significant differences compared to BY4743, suggesting that uncharacterized pathways maintain cellular polysulfide homeostasis. Five mM cysteine addition increased the formation of shorter sulfur chain species, including GSS-bimane and GSSG, but did not elevate levels of longer sulfur chain species. Additionally, polysulfides with even numbers of sulfur atoms tended to predominate in cellular lysates. Oxidized polysulfides and longer chain hydropolysulfides were not detected in finished wines. This evidence suggests that these polysulfides are unstable in wine-like environments or not transported extracellularly. Collectively, our data illustrate the complexity of yeast polysulfide metabolism under fermentation conditions.

Enantioselective Total Synthesis of (R,R)-Blumenol B and d9-(R,R)-Blumenol B.

C13-norisoprenoids are of particular importance to grapes and wines, as these molecules influence wine aroma and have been shown to significantly contribute to the distinct character of various wine varieties. Blumenol B is a putative precursor to a number of important wine aroma compounds, including the well-known compounds theaspirone and vitispirane. The enantioselective synthesis of (R,R)-blumenol B from commercially available 4-oxoisophorone was achieved using a short and easily scaleable route, which was then successfully applied to the synthesis of poly-deuterated d9-blumenol B.

Attempts to Create Products with Increased Health-Promoting Potential Starting with Pinot Noir Pomace: Investigations on the Process and Its Methods.

A process for using grape (Pinot noir) pomace to produce products with improved health-promoting effects was investigated. This process integrated a solid-liquid extraction (SLE) method and a method to acylate the polyphenolics in the extract. This report describes and discusses the methods used, including the rationale and considerations behind them, and the results obtained. The study begins with the work to optimize the SLE method for extracting higher quantities of (+)-catechin, (-)-epicatechin and quercetin by trialing 28 different solvent systems on small-scale samples of Pinot noir pomace. One of these systems was then selected and used for the extraction of the same flavonoids on a large-scale mass of pomace. It was found that significantly fewer quantities of flavonoids were observed. The resultant extract was then subject to a method of derivatization using three different fatty acylating agents. The antiproliferative activities of these products were measured; however, the resulting products did not display activity against the chosen cancer cells. Limitations and improvements to the methods in this process are also discussed.

Unraveling the Mystery of 3-Sulfanylhexan-1-ol: The Evolution of Methodology for the Analysis of Precursors to 3-Sulfanylhexan-1-ol in Wine.

Volatile polyfunctional thiol compounds, particularly 3-sulfanylhexan-1-ol (3SH) and 3-sulfanylhexyl acetate (3SHA), are key odorants contributing to the aroma profile of many wine styles, generally imparting tropical grapefruit and passionfruit aromas. 3SH and 3SHA are present in negligible concentrations in the grape berry, juice, and must, suggesting that they are released from non-volatile precursors present in the grape. The exploration of the nature and biogenesis of these precursors to 3SH and 3SHA has proven important for the elucidation of polyfunctional thiol biogenesis during alcoholic fermentation. The development and validation of appropriate analytical techniques for the analysis of 3SH precursors in enological matrices have been extensive, and this review explores the analysis and discovery of these precursor compounds. The development of analytical methods to analyze 3SH precursors, from the selection of the analytical instrument, sample preparation, and methods for standardization, will first be discussed, before highlighting how these techniques have been used in the elucidation of the biogenesis of 3SH and 3SHA in grape wines. Lastly, the future of thiol precursor analysis will be considered, with the development of new methods that greatly reduce the sample preparation time and enable multiple precursors, and the thiols themselves, to be quantitated using a single method.

Identification of herbivore-induced plant volatiles from selected Rubus species fed upon by raspberry bud moth (Heterocrossa rubophaga) larvae.

Heterocrossa rubophaga (raspberry bud moth) feed on a range of Rubus species, including commercial berryfruit crops where they are a pest. This study aimed to characterize the responses of native and non-native Rubus species to feeding by raspberry bud moth larvae. In a laboratory environment, in situ headspace volatiles of three Rubus species were collected from healthy plants and those fed upon by raspberry bud moth. Rubus cissoides (bush lawyer), the native host of raspberry bud moth, gave a limited response to larval feeding with green leaf volatiles (GLVs) representing the only new headspace constituents of the infested plants. The non-native hosts, Rubus ursinus var. loganobaccus cv Boysenberry (Boysenberry), and Rubus fruticosus (blackberry), gave strong responses to raspberry bud moth herbivory, releasing a number of unique nitrogenous compounds in conjunction with the GLVs. The nitrogenous compounds were identified as 2-methylbutanenitrile, (Z)- and (E)- 2-methylbutanal O-methyloxime, benzyl nitrile, (Z)- and (E)- phenylacetaldehyde O-methyloxime and indole. The four methyloximes and 2-methylbutanenitrile were confirmed by synthesis. Field collected phenology data showed that raspberry bud moth were active year round on both bush lawyer and blackberry.

Ultrasound assisted extraction and quantification of targeted bioactive compounds of Centella asiatica (Gotu Kola) by UHPLC-MS/MS MRM tandem mass spectroscopy.

Centella asiatica (Gotu Kola) is a green leafy vegetable rich in phytochemicals mainly including triterpenes and caffeoylquinic acids. Fresh leaves of this plant are consumed in salads and beverages in a variety of cuisines around the world. This is a well-known functional food for its neuroprotective and cognition enhancing properties in traditional societies. HPLC-DAD at lower wavelengths commonly used to identify and quantify major triterpenes of C. asiatica extracts, but associated with few drawbacks. This paper discusses a specific, sensitive and validated method developed based on UHPLC-ESI-MS-MS-MRM tandem mass spectroscopy for targeted quantification of C. asiatica bioactive compounds. The validated method enabled a precise estimation of major triterpenes and chlorogenic acid in C. asiatica in a shorter time. The findings of this study will contribute to the information on chemotype variation of C. asiatica plant grown under unique geographical, environmental and climatic conditions in New Zealand.

Metabolite characterization of fifteen by-products of the coffee production chain: From farm to factory.

Approximately 11.4 million tonnes of solid by-products and an increased amount of waste water will be generated during the 2020/21 coffee harvest. There are currently no truly value-adding uses for these potentially environmentally threatening species. This work presents the most wide-ranging chemical investigation of coffee by-products collected from farms to factories, including eight never previously investigated. Twenty compounds were found for the first time in coffee by-products including the bioactive neomangiferin, kaempferol-3-O-rutinoside, lup-20(29)-en-3-one and 3,4-dimethoxy cinnamic acid. Five by-products generated inside a factory showed caffeine (53.0-17.0 mg.g-1) and/or chlorogenic acid (72.9-10.1 mg.g-1) content comparable to coffee beans, while mature leaf from plant pruning presented not only high contents of both compounds (16.4 and 38.9 mg.g-1, respectively), but also of mangiferin (19.4 mg.g-1) besides a variety of flavonoids. Such by-products are a source of a range of bioactive compounds and could be explored with potential economic and certainly environmental benefits.

Time course accumulation of polysulfides in Chardonnay and model juice fermentations.

Over the last five years, polysulfides in wine have become a topic of interest. The finding that both yeast and the wine composition could contribute to the formation and evolution of these compounds has prompted several studies, further corroborated by the discovery of a link between polysulfides and H2S release during ageing. In the present study, the accumulation of cysteinylated and glutathionylated polysulfides was followed during fermentation for the first time. Synthetic grape media and Chardonnay juice treated with CuSO4, elemental sulfur or both were fermented, and subsequently analysed using an in-house UHPLC-HRMS method. Differences in polysulfide accumulation were observed between the two media, highlighting the role of the matrix composition. Elemental sulfur, especially when combined with CuSO4, resulted in significantly increased accumulations of polysulfides compared to controls in both media. Polysulfides with longer S-chains generally appeared later in the fermentation, an observation that prompted further questions on these metabolites' formation.

First use of grape waste-derived building blocks to yield antimicrobial materials.

Grape marc is an underutilised waste material that poses significant environmental issues. This study offers the first proof-of-concept investigation into the polymerisation of both crude and purified Sauvignon blanc grape marc extracts using the diacyl chlorides terephthaloyl chloride, succinyl chloride, adipoyl chloride, sebacoyl chloride, and the tartaric acid derivative (4R,5R)-2,2-dimethyl-1,3-dioxolane-4,5-dicarbonyl dichloride to obtain new materials, in what to the best of our knowledge is the first reported example of a direct polymerisation of an agricultural waste extract. A total of 26 novel materials were prepared. It has also shown that quercetin, a phenolic monomer found in grape marc extracts, can be polymerised with (4R,5R)-2,2-dimethyl-1,3-dioxolane-4,5-dicarbonyl dichloride to give a polymer that shows activity towards S. aureus.

A single run liquid chromatography-tandem mass spectrometry method for the analysis of varietal thiols and their precursors in wine.

This research details the development of a novel LC-MS/MS method involving a QuEChERS-based extraction that, for the first time, achieves the fast and concurrent analysis of both free varietal thiols (3-sulfanylhexan-1-ol, 3-sulfanylhexyl acetate, and 4-methyl-4-sulfanylpentan-2-one) and their S-glutathionylated/S-cysteinylated precursors in wine samples. Limits of detections for the free thiols were comparable to or lower than those obtained with previously reported more laborious methods. Repeatability for the free thiols ranged between 3-19% and from 0.1-7.7% for the thiol precursors. The method was subsequently applied to the analysis of a selection of 22 wines from a range of different grape varieties. The rapidity of this method combined with its sensitivity makes it suitable for both research and industry applications.

Elucidation of Endogenous Aroma Compounds in Tamarillo (Solanum betaceum) Using a Molecular Sensory Approach.

Glycosidically bound volatiles (GBVs) are flavorless compounds in fruits and may undergo hydrolysis during fruit maturation, storage, and processing, releasing free aglycones that are odor active. However, the contribution of glycosidic aglycones to the sensory attributes of fruits remains unclear. Herein, the key odor-active aglycones in tamarillo fruits were elucidated through the molecular sensory approach. We extracted GBVs from three cultivars of tamarillo fruits using solid-phase extraction and subsequently prepared aglycone isolates by enzymatic hydrolysis of GBVs. Gas chromatography-mass spectrometry-olfactometry (GC-MS-O) coupled with odor activity value (OAV) calculation, comparative aroma extract dilution analysis (cAEDA), and omission tests were used to identify key aromatic aglycones. A total of 42 odorants were determined by GC-MS-O analysis. Among them, trans-2,cis-6-nonadienal, 2,5-dimethyl-4-hydroxy-3(2H)-furanone (DMHF), linalool, 4-vinylguaiacol, geraniol, and α-terpineol showed high OAVs. The cultivar Amber had more aglycones with flavor dilution (FD) factors >16 than the Mulligan cultivar (27 vs 21, respectively), and the Laird's Large fruit showed the highest FD of 1024 for glycosidic DMHF. Omission tests indicated 14 aglycones as essential odorants related to GBVs in tamarillo fruits. Moreover, the enzymatic liberation of aglycones affected the sensory attributes of the tamarillo juice, resulting in an intensified odor profile with noticeable fruity and sweet notes. This study gives insights into the role of endogenous aroma during tamarillo-flavor perception, which lays the groundwork for developing tamarillo-based products with improved sensory properties.

Syntheses of mono-acylated luteolin derivatives, evaluation of their antiproliferative and radical scavenging activities and implications on their oral bioavailability.

Luteolin is a flavonoid found in a wide range of plant materials, including commonly eaten fruits and vegetables. It displays a wide range of biological activities but is known to have poor bioavailability. In this study, ten different mono-acyl (nine 5-O-acyl and one 7-O-acyl) derivatives of luteolin were synthesised for the purpose of improving bioactivity and bioavailability, and therefore enhance their therapeutic potential. The antiproliferative activity of these derivatives was assessed against the HCT116 colon cancer and MDA-MB-231 breast cancer cell lines using a 3[H] thymidine incorporation assay. The radical scavenging activity of these derivatives against 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical cation and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical using Trolox as a standard, was also assessed. Some of these derivatives were found to have improved antiproliferative activity with comparable radical scavenging activity compared to luteolin. Increased lipophilicity has been shown to increase the bioavailability of flavonoids implying these analogues will also have increased bioavailability.