Demands in research investigating small molecules by applying untargeted approaches have been a key motivator for the development of repositories for mass spectrometry spectra and automated tools to ...aid compound identification. Comparatively little attention has been afforded to using retention times (RTs) to distinguish compounds and for liquid chromatography there are currently no coordinated efforts to share and exploit RT information. We therefore present PredRet; the first tool that makes community sharing of RT information possible across laboratories and chromatographic systems (CSs). At http://predret.org, a database of RTs from different CSs is available and users can upload their own experimental RTs and download predicted RTs for compounds which they have not experimentally determined in their own experiments. For each possible pair of CSs in the database, the RTs are used to construct a projection model between the RTs in the two CSs. The number of compounds for which RTs can be predicted and the accuracy of the predictions are dependent upon the compound coverage overlap between the CSs used for construction of projection models. At the moment, it is possible to predict up to 400 RTs with a median error between 0.01 and 0.28 min depending on the CS and the median width of the prediction interval ranging from 0.08 to 1.86 min. By comparing experimental and predicted RTs, the user can thus prioritize which isomers to target for further characterization and potentially exclude some structures completely. As the database grows, the number and accuracy of predictions will increase.
Compelling evidence of the health benefits of phenolic compounds and their impact on food quality have stimulated the development of analytical methods for the identification and quantification of ...these compounds in different matrices in recent years. A targeted metabolomics method has been developed for the quantification of 135 phenolics, such as benzoates, phenylpropanoids, coumarins, stilbenes, dihydrochalcones, and flavonoids, in fruit and tea extracts and wine using UPLC/QqQ-MS/MS. Chromatography was optimized to achieve separation of the compounds over a period of 15 min, and MRM transitions were selected for accurate quantification. The method was validated by studying the detection and quantification limits, the linearity ranges, and the intraday and interday repeatability of the analysis. The validated method was applied to the analysis of apples, berries, green tea, and red wine, providing a valuable tool for food quality evaluation and breeding studies.
Flavonols are products of the flavonoid biosynthetic pathway, which also give rise to anthocyanins and condensed tannins in grapes. We investigated their presence in the berry skins of 91 grape ...varieties (Vitis vinifera L.), in order to produce a classification based on the flavonol profile. The presence of laricitrin 3-O-galactoside and syringetin 3-O-galactoside in red grapes is reported here for the first time. In red grapes, the main flavonol was quercetin (mean = 43.99%), followed by myricetin (36.81%), kaempferol (6.43%), laricitrin (5.65%), isorhamnetin (3.89%), and syringetin (3.22%). In white grapes, the main flavonol was quercetin (mean = 81.35%), followed by kaempferol (16.91%) and isorhamnetin (1.74%). The delphinidin-like flavonols myricetin, laricitrin, and syringetin were missing in all white varieties, indicating that the enzyme flavonoid 3‘,5‘-hydroxylase is not expressed in white grape varieties. The pattern of expression of flavonols and anthocyanins in red grapes was compared, in order to gain information on the substrate specificity of enzymes involved in flavonoid biosynthesis. Keywords: Vitis vinifera; grape; flavonols; anthocyanins; flavonoid 3‘-hydroxylase; flavonoid 3‘,5‘-hydroxylase; O-methyltransferase
The quantification of short-chain and medium-chain fatty acids is becoming more and more relevant in fecal and plasma samples due to their biological impact, which has been associated with colon ...rectal cancer and fiber consumption. For these reasons, a fast, cost-effective, and reproducible analytical method is highly required. In this research, a gas chromatography–mass spectrometry method based on full scan and multiple reaction monitoring (MRM) acquisition modes were optimized and validated for the analysis of short-chain and medium-chain fatty acids in three biological samples: human fecal water, fecal fermentation supernatants, and human plasma. Several extraction solvents (acidified water, diethyl ether, dichloromethane, ethyl acetate, and methyl
tert
-butyl ether (MTBE) were further evaluated, demonstrating that the latter was clearly the most suitable solvent with recoveries from 75.4 to 124.4% and coefficient of variations lower than 20%. The applicability of the GC–MS method was tested, for instance, acetic acid was quantified by using samples of plasma and feces from healthy donors at mean values of 66.9 μM and 24.5 mM, respectively. The optimized protocol could successfully find applications within multi-compartment human studies. In parallel, a second pilot experiment on fecal fermentation supernatants indicated that the proposed protocol is suitable to follow the formation of SCFAs during in vitro fermentation by the human gut microbiota. In summary, the present work provided an improved GC–MS method for precise and accurate quantification of SCFAs and MCFAs in human feces and plasma.
Fruits are nowadays considered important suppliers of anti-oxidant molecules. Apples are particularly rich in phenolic compounds, non-nutritional phytochemicals that play active roles in controlling ...severe chronic diseases. In this work, 19 phenolic compounds were investigated in both skin and pulp tissues of seven apple accessions across the Malus genus collected at two stages: during fruit development and at harvest. The primary difference in phenolic concentration between wild and domesticated accessions, especially in the pulp, could be explained by the larger growth rate of the domesticated varieties. The proposed dilution effect was also confirmed through the observation of the increased content of procyanidin B2+B4 and phloridzin in russet-skinned apples, known to have higher concentrations of these compounds. The metabolite screening was also accompanied by the expression analysis of 16 polyphenolic genes showing, for nine elements, a higher expression at harvest than during fruit development. Finally, a polyphenolic comparison with red-fleshed apples was also carried out, underlying a larger amount of procyanidins and quercetin-3rhamnoside in the white-fleshed accessions. The results presented and discussed in this work suggest that specific white-fleshed apples, especially with russeted-skin, may play an important role in ameliorating the nutraceutical potential of apple fruit.
Accurate quantification and structural characterization of ellagitannins and ellagic acid conjugates in food, beverages, and food supplements are essential starting points for studying their effect ...on human health. However, accuracy is hindered both by the lack of pure standard compounds and by methods that maintain the compounds in their native form, avoiding any chemical modification of the structure. The objective of this work was to develop a new method for the purification, chromatographic separation, and accurate quantification of ellagitannins and ellagic acid conjugates to provide thorough characterization of the diversity in composition of 11 Rubus cultivars grown in Trentino, Italy. As such, two major steps were required: (i) the isolation and purification (with associated detailed structural characterization and determination of their molar extinction coefficients) of sanguiin H-6 and lambertianin C, providing essential data for their use, together with ellagic acid, as external standards, and (ii) the determination of the chemical structure of 20 novel minor ellagitannins and 4 ellagic acid conjugates on the basis of their Q-TOF-HDMS and DAD spectra. This survey of ellagitannins and ellagic acid conjugates provides evidence for the existence of significant differences in the pattern between and within blackberry and raspberry cultivars. To our knowledge, this is the first paper that has combined detailed metabolite profiling with accurate quantification of the main ellagitannins in Rubus using their respective standards.
The abundance of lipids in plants is influenced by genotype and phenotype. Despite being a very important class of plant metabolites, knowledge of grape lipids is still very limited to date, with the ...exception of those located in seeds. Few investigations of grape lipids have shown that their profile depends on grape maturity, the variety and their location in the berry. Recent advances in liquid chromatography coupled to mass spectrometry have paved the way for faster analysis of lipids with minimal sample preparation. Here we describe a validation method for the extraction, identification and quantification of different classes of grape lipids: fatty acids, sterols, glycerolipids, glycerophospholipids and sphingolipids using liquid chromatographic electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). The method was validated for 33 lipids, with linearity range (R2=0.95–1.00), LOQ (0.003–14.88ngmL−1) and intraday and interday repeatability being evaluated for each lipid. The lipid profiling method developed was successfully applied to the analysis of 18 grape samples (10 red grape and 8 white grape varieties) from 4 different genetic groups: Vitis vinifera, Vitis non-vinifera, Muscat and hybrid; 33 lipids were identified and quantified. This method, which can be easily expanded to include further compounds and other plant tissues, is the starting point for analysis of the lipid profile in different grape tissues, an essential goal for better understanding the role of lipids in grape physiology.
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•UHPLC–ESI-MS/MS method development for different classes of grape lipids.•Separation and quantification of the 33 grape lipids.•Application and validation of the method to 18 grape varieties.•The method can be easily expanded for studying plant tissues and physiology.
► A new HILIC–MS/MS method for the determination of 124 polar metabolites. ► ESI (positive/negative) ionisation of aminoacids, amines, organic acids, saccharides. ► Utilisation of new sub-3μm ...stationary phase material (HILIC Amide). ► Provides quantitative results that complement untargeted metabolomics studies. ► Applicability in different fields: biochemistry, food science, clinical chemistry.
A hydrophilic interaction liquid chromatography (HILIC)-MS/MS profiling method was developed for the efficient separation and quantification of small polar molecules, mostly primary metabolites. The method was based on an ultrahigh performance liquid chromatography (UHPLC) separation system coupled with ESI mass spectrometry on a triple quadrupole mass spectrometer, operating in both positive and negative ionisation mode using rapid polarity switching. With the developed method quantitation of 135 compounds belonging in four major classes of polar compounds (sugars, aminoacids, organic acids and amines) was achieved in a single run of 30min. The method was applied to grape extracts from different varieties and provided information on primary metabolite content. Multivariate statistical analysis was applied using the concentrations found, with the aim of investigating the differences in metabolite profiles. Classification of grapes according to their skin colour was carried out using principle component analysis based on the concentration variation of a number of the metabolites studied.
Glycosides are ubiquitous plant secondary metabolites consisting of a non-sugar component called an aglycone, attached to one or more sugars. One of the most interesting aglycones in grapes and wine ...is methyl salicylate (MeSA), an organic ester naturally produced by many plants, particularly wintergreens. To date, nine different MeSA glycosides from plants have been reported, mainly spread over the genera
and
From a sensorial point of view, MeSA has a balsamic-sweet odor, known as Wintergreen. MeSA was found in
grapes, in
. and in the Frontenac interspecific hybrid. We found that the MeSA glycosides content in Verdicchio wines and in some genetically related varieties (Trebbiano di Soave and Trebbiano di Lugana) was very high. In order to understand which glycosides were present in wine, the methanolic extract of Verdicchio wine was injected into a UPLC-Q-TOF-HDMS and compared to the extracts of different plants rich in such glycosides. Using pure standards, we confirmed the existence of two glycosides in wine: MeSA 2-
--d-glucoside and MeSA 2-
--d-xylopyranosyl (1-6) -d-glucopyranoside (gaultherin). For the first time, we also tentatively identified other diglycosides in wine: MeSA 2-
--l-arabinopyranosyl (1-6)--d-glucopyranoside (violutoside) and MeSA 2-
--d-apiofuranosyl (1-6)--d-glucopyranoside (canthoside A), MeSA 2-
--d-glucopyranosyl (1-6)-
--d-glucopyranoside (gentiobioside) and MeSA 2-
--l-rhamnopyranosyl (1-6)--d-glucopyranoside (rutinoside). Some of these glycosides have been isolated from
leaves by preparative liquid chromatography and structurally annotated by
H- and
C-NMR analysis. Two of the peaks isolated from
leaves, namely MeSA sambubioside and MeSA sophoroside, were herein observed for the first time. Six MeSA glycosides were quantified in 64 Italian white wines, highlighting the peculiar content and pattern in Verdicchio wines and related cultivars. The total concentration in bound and free MeSA in Verdicchio wines varied in the range of 456-9796 g/L and 5.5-143 g/L, respectively, while in the other wines the bound and free MeSA was below 363 g/L and 12 g/L, respectively. As this compound's olfactory threshold is between 50 and 100 g/L, our data support the hypothesis that methyl salicylate can contribute to the balsamic scent, especially in old Verdicchio wines.
All of the matrices entailed in olive oil processing were screened for the presence of known and new phenol constituents in a single study, combining an ultra high pressure liquid chromatography ...system with diode array and electrospray ionization quadrupole time-of-flight high resolution mass spectrometry (ESI-QTOF-HRMS) detection. Their trail was followed from the fruit (peel/pulp and stone) to the paste and final products, i.e. pomace, wastewater, and oil, providing important insight into the origin, disappearance, and evolution of each during the operational steps. Eighty different phenols, composed of fruit native representatives and their technologically formed and/or released derivatives, were detected in six olive matrices and fully characterized on the basis of HRMS and UV–vis spectroscopic data. In addition to phenols already known in olive matrices, four new molecular formulas were proposed and three new tentative identities assigned to newly discovered phenols, i.e., β-methyl-OH-verbascoside, methoxynüzhenide, and methoxynüzhenide 11-methyl oleoside.
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