Sustainable In Silico-Supported Ultrasonic-Assisted Extraction of Oligomeric Stilbenoids from Grapevine Roots Using Natural Deep Eutectic Solvents (NADES) and Stability Study of Potential Ready-to-Use Extracts.

Grapevine roots, as a side-stream of a vineyard, are a sustainable resource for the recovery of oligomeric stilbenoids, such as the bioactive r-viniferin. The aim of this study is to evaluate an in silico-supported method, based on the Conductor-like Screening Model for Real Solvents (COSMO-RS), for selection of environmentally friendly natural deep eutectic solvents (NADES) with regard to the extraction of grapevine roots. The most suitable NADES system for ultrasonic-assisted extraction of r-viniferin was choline chloride/1,2-propanediol. The optimal extraction parameters for r-viniferin were determined using single-factor experiments as follows: choline chloride/1,2-propanediol 1/2 mol/mol, 10 wt% H2O, biomass/NADES ratio 1/10 g/g, and 10 min extraction time. Under optimized conditions, the extraction yield of r-viniferin from grapevine roots reached 76% of the total r-viniferin content. Regarding stability, stilbenoids in choline chloride/1,2-propanediol remained stable during 128 days of storage at ambient temperature. However, fructose/lactic acid-based NADES were observed to degrade stilbenoids; therefore, the removal of the NADES will be of interest, with a suitable method implemented using Amberlite® XAD-16N resin. As green solvents, the NADES have been used as effective and environmentally friendly extractants of stilbenoid-containing extracts from grapevine roots for potential applications in the cosmetic and pharmaceutical industry or as nutraceuticals in the food industry.

In Silico-Assisted Isolation of trans-Resveratrol and trans-ε-Viniferin from Grapevine Canes and Their Sustainable Extraction Using Natural Deep Eutectic Solvents (NADES).

Grapevine canes are an important source of bioactive compounds, such as stilbenoids. This study aimed to evaluate an in silico method, based on the Conductor-like Screening Model for Real Solvents (COSMO-RS) to isolate stilbenoids from a grapevine cane extract by offline heart-cut high-performance countercurrent chromatography (HPCCC). For the following extraction of resveratrol and ε-viniferin from grapevine canes, natural deep eutectic solvents (NADES) were used as an environmentally friendly alternative to the traditionally used organic solvents. In order to evaluate a variety of combinations of hydrogen bond acceptors (HBAs) and hydrogen bond donors (HBDs) for the targeted extraction of stilbenoids, COSMO-RS was applied. In particular, ultrasonic-assisted extraction using a solvent mixture of choline chloride/1,2-propanediol leads to higher extraction yields of resveratrol and ε-viniferin. COSMO-RS calculations for NADES extraction combined with HPCCC biphasic solvent system calculations are a powerful combination for the sustainable extraction, recovery, and isolation of natural products. This in silico-supported workflow enables the reduction of preliminary experimental tests required for the extraction and isolation of natural compounds.

A Comparison between High-Performance Countercurrent Chromatography and Fast-Centrifugal Partition Chromatography for a One-Step Isolation of Flavonoids from Peanut Hulls Supported by a Conductor like Screening Model for Real Solvents.

Peanut hulls (Arachis hypogaea, Leguminosae), which are a side stream of global peanut processing, are rich in bioactive flavonoids such as luteolin, eriodictyol, and 5,7-dihydroxychromone. This study aimed to isolate these flavonoid derivatives by liquid-liquid chromatography with as few steps as possible. To this end, luteolin, eriodictyol and 5,7-dihydroxychromone were isolated from peanut hulls using two different techniques, high-performance countercurrent chromatography (HPCCC) and fast-centrifugal partition chromatography (FCPC). The suitability of the biphasic solvent system composed of n-hexane/ethyl acetate/methanol/water (1.0/1.0/1.0/1.5; v/v/v/v) was determined by the Conductor like Screening Model for Real Solvents (COSMO-RS), which allowed the partition ratio KD-values of the three main flavonoids to be calculated. After a one-step HPCCC separation of ~1000 mg of an ethanolic peanut hull extract, 15 mg of luteolin and 8 mg of eriodictyol were isolated with purities over 96%. Furthermore, 3 mg of 5,7-dihydroxychromone could be isolated after purification by semi-preparative reversed-phase liquid chromatography (semi-prep. HPLC) in purity of over 99%. The compounds were identified by electrospray ionization mass spectrometry (ESI-MS) and nuclear magnetic resonance spectroscopy (NMR).

Polyphenol-Rich Beverage Consumption Affecting Parameters of the Lipid Metabolism in Healthy Subjects.

Polyphenols are a diverse and widely distributed class of secondary metabolites, which possess numerous beneficial properties including a modulation of glucose and lipid metabolism. This placebo-controlled human intervention study was performed to explore effects of polyphenol-rich beverage (PRB) uptake on lipid metabolism, as well as DNA integrity. In this case, 36 healthy men were randomly divided to consume either 750 mL of a PRB (containing 51% chokeberry, cranberry, and pomegranate) or a placebo drink daily for eight weeks. Only PRB consumption was found to decrease fat and protein intakes significantly compared to the preceding one-week washout period. During the intervention with PRB an increased fat-free mass was shown after four weeks, whereas a significant elevation in body weight and leptin was observed in placebo group. Blood lipids were not significantly altered after PRB consumption, while triglyceride levels increased after placebo drink intake. In platelets, a significant inhibition of phosphodiesterase (PDE) activity was observed, more pronounced in test group. Consuming the PRB decreased total DNA strand breaks in whole blood as well as H2O2-induced breaks in isolated lymphocytes. Overall, our study suggested beneficial effects on lipid metabolism by reduced energy intake, modulation of biomarkers such as PDE activity and improved DNA integrity associated with PRB consumption.

Target-Guided Isolation of Progenitors of 1,1,6-Trimethyl-1,2-dihydronaphthalene (TDN) from Riesling Wine by High-Performance Countercurrent Chromatography.

High-performance countercurrent chromatography (HPCCC) was used for the target-guided isolation of precursors of 1,1,6-trimethyl-1,2-dihydronaphthalene (TDN) from Riesling wine. In separated HPCCC fractions of an Amberlite® XAD®-2 extract obtained from a German Riesling, TDN-generating fractions were identified by the acid-catalyzed hydrolysis of the progenitors at pH 3.0 and subsequent HS-GC-MS/MS analysis. The presence of multiple TDN-generating precursors in Riesling wine could be confirmed. From polar HPCCC fractions (11-13 and 14-16), 3,4-dihydroxy-7,8-dihydro-ß-ionone 3-O-rutinoside and 3,4-dihydroxy-7,8-dihydro-ß-ionone 3-O-ß-d-glucopyranoside were isolated as major TDN-precursors at a sufficient amount for structure elucidation by NMR spectroscopic studies. In the medium polar HPCCC factions (27-35), enzymatic hydrolysis liberated the aglycones 3-hydroxy-ß-ionone and 3-hydroxy-TDN in minor amounts. In further less polar TDN-generation fractions (36-44 and 45-50), glycosidic progenitors were absent; instead, a minor TDN formation most likely from non-conjugated constituents was observed.

Comprehensive Metabolite Profiling of Hydrangea macrophylla ssp. serrata Extracts Using Liquid Chromatography Coupled with Electrospray Ionization Ion Mobility Quadrupole Time-of-Flight Mass Spectrometry.

A wide range of secondary metabolites has been described for various Hydrangea species, including the sweet-tasting phenyldihydroisocoumarin phyllodulcin, which is found in the leaves of Hydrangea macrophylla ssp. serrata. This work aims at the development and validation of an analytical workflow for comprehensive semi-polar metabolite profiling using liquid chromatography coupled with electrospray ionization ion mobility quadrupole time-of-flight mass spectrometry (UPLC-ESI-IMS-QToF-MS) to complement existing analytical studies. The unsupervised analysis of this data set demonstrates the capability of this analytical workflow to distinguish different H. macrophylla ssp. serrata cultivars. In combination with supervised analysis, a list of metabolites responsible for the differentiation of the cultivars studied has been obtained. Suspect screening of phenyldihydroisocoumarins provides comprehensive information, which could help in the search for key enzymes related to the biosynthesis of phyllodulcin.

Separation of Dihydro-Isocoumarins and Dihydro-Stilbenoids from Hydrangea macrophylla ssp. serrata by Use of Counter-Current Chromatography.

Previously, different Hydrangea macrophylla ssp. serrata cultivars were investigated by untargeted LC-MS analysis. From this, a list of tentatively identified and unknown compounds that differ significantly between these cultivars was obtained. Due to the lack of reference compounds, especially for dihydro-isocoumarins, we aimed to isolate and structurally characterise these compounds from the cultivar 'Yae-no-amacha' using NMR and LC-MS methods. For purification and isolation, counter-current chromatography was used in combination with reversed-phase preparative HPLC as an orthogonal and enhanced purification workflow. Thirteen dihydro-isocoumarins in combination with other metabolites could be isolated and structurally identified. Particularly interesting was the clarification of dihydrostilbenoid glycosides, which were described for the first time in H. macrophylla ssp. serrata. These results will help us in further studies on the biological interpretation of our data.

Variability of Constitutive Stilbenoid Levels and Profiles in Grape Cane (Vitis vinifera L.) Depending upon Variety and Clone, Location in the Vineyard, Pruning Time, and Vintage.

Stilbenoids in grape cane (Vitis vinifera L.) are bioactive compounds relevant for plant defense and the potential valorization of this byproduct. Our screening of grape cane from 102 varieties showed constitutive stilbenoid levels in a wide range (557-7748 mg/kg of dry weight). Analyses of genetically distinct clones of selected cultivars unraveled that intravarietal variability (e.g., cv. Riesling, 3236-6541 mg/kg) was higher than that across samples from a single clone but different vineyard positions (3017-3710 mg/kg). Furthermore, stilbenoid levels in samples obtained in October, December, and February (3 years, 2017-2019) showed pronounced quantitative and qualitative variability and the highest yields upon December pruning. For instance, vitisin B and ε-viniferin in cv. Pinot Noir and Accent were predominant in 2017 and 2019 (both >90% of total stilbenoids) but not in 2018 (both <55%) when temperatures were high and precipitation low. In brief, we report the variability of stilbenoid levels in grape cane depending upon genetic and environmental factors.

Bioactive Phenolic Compounds from Lingonberry (Vaccinium vitis-idaea L.): Extraction, Chemical Characterization, Fractionation and Cellular Antioxidant Activity.

Lingonberries contain high contents of bioactive compounds such as chlorogenic acids and anthocyanins. In addition to radical scavenging and antioxidant activities, these compounds can protect cells from DNA damage. For this reason, lingonberries might be well suited for nutraceuticals or natural biomedicines. To assess these applications, the present study characterized and identified the most effective extract, only consisting of anthocyanins, copigments or a mixture of both, obtained from a lingonberry juice concentrate. An extract was generated by using a XAD-7 column followed by fractionation into anthocyanins and copigments using adsorptive membrane chromatography. After identification of main polyphenols by HPLC-photodiode array-electrospray ionization-tandem mass spectrometry, free radical scavenging activity was analyzed by electron spin resonance spectroscopy using 2,2-diphenyl-1-picrylhydrazyl and galvinoxyl radicals. Furthermore, cyclic voltammetry analyses and the Trolox equivalent antioxidant capacity (TEAC) assay were applied. Finally, the reactive oxygen species (ROS) reducing effects of the lingonberry extract and its fractions were evaluated in HepG2 cells. While the combination of anthocyanins and copigments possessed the highest antioxidant activities, all samples (XAD-7 extract, anthocyanin and copigment fraction) protected cells from oxidative stress. Thus, synergistic effects between phenolic compounds may be responsible for the high antioxidant potential of lingonberries, enabling their use as nutraceuticals.

Fractionation of Extracts from Black Chokeberry, Cranberry, and Pomegranate to Identify Compounds That Influence Lipid Metabolism.

Polyphenols show a spectrum of bioactive effects, including an influence on lipid metabolism. In this study, we performed activity-guided fractionations of black chokeberry (aronia), cranberry, and pomegranate extracts to identify the biologically active compounds. The extracts were prepared from fruit juice concentrates with the adsorbent resin Amberlite XAD-7 and were separated into a copigment and an anthocyanin fraction, followed by fractionation into a polymer and monomeric fraction by means of hexane precipitation. For further fractionation of the cranberry and pomegranate copigment fractions, high-performance countercurrent chromatography (HPCCC) was used. The compounds in each fraction were identified by high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS), and the quantification was performed by ultra high-performance liquid chromatography-diode array detector (UHPLC-DAD) analyses. Each of the (sub-)fractions was tested in three in vitro assays: phosphodiesterase 3B (PDE) activity, lipid accumulation, and lipolysis in 3T3-L1 cells. The results showed that various fractions and subfractions can inhibit lipid accumulation and PDE activity as well as increase lipolysis, particularly copigments. Overall, our results indicate an influence of polyphenol-rich (sub-)fractions on the lipid metabolism.

Shifts in biochemical and physiological responses by the inoculation of arbuscular mycorrhizal fungi in Triticum aestivum growing under drought conditions.

BACKGROUND: A decrease in rainfall is one of the main constraints on wheat production, although the association of wheat with arbuscular mycorrhizal fungi (AMF) may be an alternative for crop production under drought conditions. In the present study, we used three wheat cultivars (Purple, Ilustre and Maxi Baer) inoculated with two AMF strains, one obtained from the hyperarid Atacama Desert (northern Chile; Fm) and the other obtained from southern Chile (Cc). Plants were maintained under two irrigation conditions (normal irrigation and drought) and the physiological behaviour and enzymatic and nonenzymatic antioxidant activities in the shoots were determined. In addition, the phenolic compounds were identified by high-performance liquid chromatography-dioide array detection-electrospray ionization-tandem mass spectrometry and quantified. RESULTS: AMF colonization produced higher levels of efficiency of photosystem II and photosynthetic pigments. High values of catalase in Purple-Cc, ascorbate peroxidase in Purple-Cc, glutathione reductase in Maxi-Cc and superoxide dismutase in Purple-Cc, all under stress, were registered. Of the inoculated cultivars, Purple-Cc showed the highest flavonoid levels, whereas hydroxycinnamic acids were higher in Maxi-Fm without drought, with apigenin and luteolin being the most abundant. High levels of phenols were present in the Ilustre-Fm plants without drought. Under normal irrigation, high levels of antioxidant activity were registered in the AMF treatments, whereas, under stress conditions, in general, high values were observed under the Fm inoculation. CONCLUSION: Our results showed that the greatest antioxidant activity and phenolic content occurred in wheat plants inoculated with AMF, indicating their influence on coping with water stress, which is of importance in vast areas where global climate change is resulting in diminished rainfall. © 2021 Society of Chemical Industry.

Isolation of N-Ethyl-2-pyrrolidinone-Substituted Flavanols from White Tea Using Centrifugal Countercurrent Chromatography Off-Line ESI-MS Profiling and Semi-Preparative Liquid Chromatography.

N-Ethyl-2-pyrrolidinone-substituted flavanols (EPSF) are marker compounds for long-term stored white teas. However, due to their low contents and diasteromeric configuration, EPSF compounds are challenging to isolate. In this study, two representative epimeric EPSF compounds, 5'''R- and 5'''S-epigallocatechin gallate-8-C N-ethyl-2-pyrrolidinone (R-EGCG-cThea and S-EGCG-cThea), were isolated from white tea using centrifugal partition chromatography (CPC). Two different biphasic solvent systems composed of 1. N-hexane-ethyl acetate-methanol-water (1:5:1:5, v/v/v/v) and 2. N-hexane-ethyl acetate-acetonitrile-water (0.7:3.0:1.3:5.0, v/v/v/v) were used for independent pre-fractionation experiments; 500 mg in each separation of white tea ethyl acetate partition were fractionated. The suitability of the two solvent systems was pre-evaluated by electrospray mass-spectrometry (ESI-MS/MS) analysis for metabolite distribution and compared to the results of the CPC experimental data using specific metabolite partition ratio KD values, selectivity factors α, and resolution factors RS. After size-exclusion and semi-preparative reversed-phase liquid chromatography, 6.4 mg of R-EGCG-cThea and 2.9 mg of S-EGCG-cThea were recovered with purities over 95%. Further bioactivity evaluation showed that R- and S-EGCG-cThea possessed in vitro inhibition effects on α-glucosidase with IC50 of 70.3 and 161.7 µM, respectively.

The complexity of sound quantification of specialized metabolite biosynthesis: The stress related impact on the alkaloid content of Catharanthus roseus.

Medicinal plants grown under stress conditions reveal higher concentrations of relevant specialized metabolites than well-watered plants, putatively due to an enhanced biosynthesis. Yet, stress also reduced the biomass gain. Accordingly, the concentration increase in comparison to control plants could also be due to lesser biomass employed as the reference value, whereas the rate of biosynthesis may remain unchanged. For an unequivocal proof that stress indeed enhances the biosynthesis, the total amount of the substances per plant has to be determined. In this study, we investigated the stress-induced impact on the alkaloids accumulated in Catharanthus roseus and quantified both, the changes in concentration and in the entire amount of alkaloids. At any time, all Catharanthus roseus plants grown under drought stress exhibited a markedly higher alkaloid concentration compared to the well-watered controls. However, by calculating the entire alkaloid content per plant, a corresponding increment occurred only within the first two weeks of drought stress. Thereafter, no significant differences among drought treatments and control were detected. Finally, within the last week, the alkaloid content per plant decreased markedly, although there was a meaningfully higher concentration of alkaloids in the drought-stressed plants. In contrast, when plants had been exposed to high salt concentrations, the alkaloid concentrations were quite the same in stressed and control plants. The related total contents were significantly lower in plants exposed to salt stress. These results display that both phenomena, an increased rate of biosynthesis and lesser reference values, i.e., the biomass, contribute to the stress-related increase in the concentration of natural product. Moreover, it has to be considered that the enhancement of biosynthesis could be due to either an "active" up-regulation of biosynthetic capacity or a "passive" shift caused by the over-reduced status as a result of the stress-induced stomatal closure.

HPLC-DAD-MS and Antioxidant Profile of Fractions from Amontillado Sherry Wine Obtained Using High-Speed Counter-Current Chromatography.

In the present work, the polyphenolic profile of a complex matrix such as Amontillado sherry has been processed by means of high-speed counter-current chromatography (HSCCC) and characterized by HPLC-DAD-MS. An Amberlite XAD-7 column was used to obtain the wine extract, and three different biphasic solvent systems were applied for HSCCC separation: MTBE (methyl tert-butyl ether)/n-butanol/acetonitrile/water (1.1/3/1.1/5+0.1% trifluoroacetic acid), MTBE/n-butanol/acetonitrile/water (2/2/1/5), and hexane/ethyl acetate/ethanol/water (1/5/1/5). As a result, 42 phenolic compounds and furanic derivatives have been identified by means of HPLC-DAD-MS, with 11 of them being identified for the first time in Sherry wines: 3-feruloylquinic acid, isovanillin, ethyl vanillate, furoic acid, dihydro-p-coumaric acid, 6-O-feruloylglucose, ethyl gallate, hydroxytyrosol, methyl protocatechuate, homoveratric acid and veratraldehyde. In addition, the antioxidant capacity (ABTS) of the obtained fractions was determined, revealing higher values in those fractions in which compounds such as gallic acid, protocatechuic acid, protocatechualdehyde, trans-caftaric acid, syringic acid, isovanillin or tyrosol, among others, were present. This is the first time that HSCCC has been used to characterize the phenolic composition of Sherry wines.

Carbohydrate Hydrolase-Inhibitory Activity of Juice-Based Phenolic Extracts in Correlation to Their Anthocyanin/Copigment Profile.

Red fruits and their juices are rich sources of polyphenols, especially anthocyanins. Some studies have shown that such polyphenols can inhibit enzymes of the carbohydrate metabolism, such as α-amylase and α-glucosidase, that indirectly regulate blood sugar levels. The presented study examined the in vitro inhibitory activity against α-amylase and α-glucosidase of various phenolic extracts prepared from direct juices, concentrates, and purees of nine different berries which differ in their anthocyanin and copigment profile. Generally, the extracts with the highest phenolic content-aronia (67.7 ± 3.2 g GAE/100 g; cyanidin 3-galactoside; chlorogenic acid), pomegranate (65.7 ± 7.9 g GAE/100 g; cyanidin 3,5-diglucoside; punicalin), and red grape (59.6 ± 2.5 g GAE/100 g; malvidin 3-glucoside; quercetin 3-glucuronide)-showed also one of the highest inhibitory activities against α-amylase (326.9 ± 75.8 µg/mL; 789.7 ± 220.9 µg/mL; 646.1 ± 81.8 µg/mL) and α-glucosidase (115.6 ± 32.5 µg/mL; 127.8 ± 20.1 µg/mL; 160.6 ± 68.4 µg/mL) and, partially, were even more potent inhibitors than acarbose (441 ± 30 µg/mL; 1439 ± 85 µg/mL). Additionally, the investigation of single anthocyanins and glycosylated flavonoids demonstrated a structure- and size-dependent inhibitory activity. In the future in vivo studies are envisaged.

Pomegranate (Punica granatum L.) Extract and Its Anthocyanin and Copigment Fractions-Free Radical Scavenging Activity and Influence on Cellular Oxidative Stress.

Secondary plant metabolites, e.g., polyphenols, are widely known as health-improving compounds that occur in natural functional foods such as pomegranates. While extracts generated from these fruits inhibit oxidative stress, the allocation of these effects to the different subgroups of substances, e.g., anthocyanins, "copigments" (polyphenols without anthocyanins), or polymeric compounds, is still unknown. Therefore, in the present study, polyphenols from pomegranate juice were extracted and separated into an anthocyanin and copigment fraction using adsorptive membrane chromatography. Phenolic compounds were determined by high performance liquid chromatography with photodiode array (HPLC-PDA) detection and HPLC-PDA electrospray ionization tandem mass spectrometry (HPLC-PDA-ESI-MS/MS), while the free radical scavenging activity of the pomegranate XAD­7 extract and its fractions was evaluated by the Trolox equivalent antioxidant capacity (TEAC) assay and electron spin resonance (ESR) spectroscopy. Compared to juice, the total phenolic content and free radical scavenging potential was significantly higher in the pomegranate XAD-7 extract and its fractions. In comparison to the anthocyanin and copigment fraction, pomegranate XAD-7 extract showed the highest radical scavenging activity against galvinoxyl and DPPH radicals. Moreover, the enriched XAD-7 extract and its fractions were able to protect human hepatocellular HepG2 cells against oxidative stress induced by hydrogen peroxide. Overall, these results indicated that anthocyanins and copigments act together in reducing oxidative stress.

Rapid UV/Vis Spectroscopic Dye Authentication Assay for the Determination and Classification of Reactive Dyes, Monascus Pigments, and Natural Dyes in Coloring Foodstuff.

Food authenticity in the field of food dyes can be interpreted as the correctness of the coloring ingredients indicated. The Rapid UV/vis Spectroscopic Dye Authentication Assay (RaSDAY) presented in this work was used to verify the authenticity of water-soluble reddish colorings for food use. RaSDAY includes the processing of samples under different experimental conditions with pH variations and heat exposure. The absorbances measured are analyzed by principal component analysis and a k-nearest neighbors algorithm. As a result, classification of anthocyanins, betalains, and carmine and the detection of Monascus pigments, undeclared artificial food dyes, and reactive textile azo dyes can be performed by utilizing a rapid screening method. In 17 out of 20 samples of coloring food additives that were included in this work, reactive dyes, unpermitted Monascus pigments, and artificial food dyes were detected using the developed method. "Reactive Red 120", "Reactive Red 195", and "Reactive Red 198" were identified by subsequent 1H NMR spectroscopy in eight of those samples.

In Vitro Inhibition of Phosphodiesterase 3B (PDE 3B) by Anthocyanin-Rich Fruit Juice Extracts and Selected Anthocyanins.

Phosphodiesterases (PDEs) are essential enzymes for the regulation of pathways mediated by cyclic adenosine monophosphate (cAMP). Secondary plant compounds like anthocyanins (ACs) can inhibit PDE activity and, consequently, may be beneficial for lipid metabolism. This study investigated 18 AC-rich juice extracts and pure reference compounds from red fruits for potential inhibitory effects on PDE 3B activity. Extracts were obtained through adsorption on Amberlite® XAD 7 resin. Based on this screening, the chokeberry, blueberry, pomegranate, and cranberry extracts were active, with half maximal inhibitory concentrations (IC50) ranging from 163 ± 3 µg/mL to 180 ± 3 µg/mL. The ACs in these extracts, peonidin-3-glucoside and cyanidin-3-arabinoside, were the most active single compounds (IC50 = 56 ± 20 µg/mL, 108 ± 6 µg/mL). All extracts comprised high amounts of phenolic compounds, as determined by the Folin-Ciocalteu assay, ranging from 39.8 ± 1.5 to 73.5 ± 4.8 g gallic acid equivalents (GAE)/100 g extract. Pomegranate and chokeberry extracts exhibited the largest amounts of polyphenols (72.3 ± 0.7 g GAE/100 g, 70.6 ± 4.1 g GAE/100 g, respectively). Overall, our results showed that fruit juice extracts and their ACs can inhibit PDE activity. Any potential health benefits in vivo will be investigated in the future.

Pyrrolizidine alkaloid profiling of four Boraginaceae species from Northern Germany and implications for the analytical scope proposed for monitoring of maximum levels.

Pyrrolizidine alkaloids (PAs) and their corresponding N-oxides (PANOs) have been determined in food and feed at levels relevant for consumer health. More than 660 different PAs have been detected, but few are available as reference substances for analytical demands. In the context of the European legislation on maximum levels of PAs in food products, a defined analytical scope of 21 PAs for determination has been suggested. An expansion of the scope from 21 to 35 PAs, including 14 structural isomers, is currently under discussion. In the present study, a target screening method was established for a comprehensive characterisation of PA profiles of the species Echium vulgare, Heliotropium europaeum, Cynoglossum officinale and Symphytum spp. to assess whether an expansion of the analytical scope is required to quantitatively cover the total PA contents of Boraginaceae species. The scope of the method comprised known and unknown PAs previously screened and confirmed in the respective plant extracts. A total of 176 PAs and PANOs were detected. The toxic 1,2-unsaturated PAs represent the predominant PA type with about 98% of the mean total content. This PA profiling demonstrates that an expansion of the scope from 21 to 35 PAs is required to adequately cover the mean total PA contents of Cynoglossum officinale and H. europaeum, whereas in the case of Symphytum spp. and Echium vulgare an expansion would not be necessary. ABBREVIATIONS: Pyrrolizidine alkaloid: PA, Pyrrolizidine alkaloid N-oxide: PANO, European Food Safety Authority: EFSA, German Federal Institute for Risk Assessment: BfR, ultra-high performance liquid chromatography: UHPLC, high-resolution: HR, tandem mass spectrometer: MS/MS, multiple reaction monitoring: MRM, data-independent MS2: ddms2, electrospray ionisation: ESI, limit of detection: LOD, limit of quantification: LOQ, gas chromatography coupled with mass spectrometry: GC-MS, fragmentation (data): MS2 (data), full scan: MS1, variable data-independent acquisition: vDIA, monoester esterified with a necic acid at position C9 of the necine base: O9-monoester, relative proportion: rel. prop., intraperitoneal: i.p., intravenous: i.v., higher-energy C-trap dissociation: HCD, all-ion fragmentation: AIF, parallel reaction monitoring: PRM.

Impact of Rootstock, Clonal Selection, and Berry Size of Vitis vinifera sp. Riesling on the Formation of TDN, Vitispiranes, and Other Volatile Compounds.

The C13-norisoprenoid aroma compounds 1,1,6-trimethyl-1,2-dihydronaphthalene (TDN) and biosynthetically related vitispirane (VS) are important contributors to the varietal aroma of Riesling wines and are released from glycosidically bound carotenoid breakdown products during bottle aging. TDN is appreciated by numerous winemakers, particularly in aged Riesling wines. Higher levels of TDN, however, are perceived as a "petrol" off-flavor, which is expected to increase due to ongoing climate change. Wines produced from the same Riesling clone, which was grafted on six different rootstocks, varied significantly throughout two vintages in respect to their concentrations of free and bound TDN and VS as well as other volatiles. Over three vintages, the same compounds differed significantly among wines made from eight Riesling clones grafted on the same rootstock. Genetically determined loose grape clusters favored the formation of TDN and yielded wines of stronger sensory petrol intensity. Berry size, however, had no relevant impact on TDN and VS formation.